An assessment of bait availability, utilization and management guidelines in Eastern Cape estuaries
- Authors: Jooste, Jakobus Gerrit
- Date: 2003
- Subjects: Upogebia african -- Effects of sediments on -- South Africa -- Eastern Cape , Estuarine ecology -- South Africa -- Eastern Cape , Fishing baits -- Conservation -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11073 , http://hdl.handle.net/10948/334 , Upogebia african -- Effects of sediments on -- South Africa -- Eastern Cape , Estuarine ecology -- South Africa -- Eastern Cape , Fishing baits -- Conservation -- South Africa -- Eastern Cape
- Description: Aspects around the management of benthic soft sediment bait organisms, with special focus on the mud prawn (Upogebia africana) of eastern Cape estuaries was investigated. The recreational linefishery of the Gamtoos estuary was described, and compared to previous studies. Target fish species was identified, and a large dependency on bait sourced from estuaries needed for the capture of these species was noted, especially for spotted grunter (Pomadasys commersonnii). A comparison between bait use, success and the natural diet of target fish species was also made. The structure and distribution of sediments in the old channel mud banks was described and compared with historical data. The influence of sediments on mud prawn distribution was investigated, but no significant interactions were found at the study site. The impacts of once-off pumping and digging events, and monthly trampling on the sediments and mud prawn population was studied over a 7-month period. Initial removal rates as well as recovery time varied significantly between the two collection methods, while the largest decline in prawn numbers (to zero) with no recovery visible after seven months was caused by trampling. All disturbances caused some alteration in sediment composition, but not to such an extent that the sediments became unfavorable for mud prawns. Trampling did, however, result in the compaction of sediments to such a degree that prawns could not construct burrows. Issues around current removal quotas of bait species as well as the creation of a small-scale commercial (SSC) bait selling operation at Swartkops estuary were critically evaluated, and suggestions for the future removal rates of mud prawns based on production export calculations were made. The distribution, size, sex ratios and number of gravid females occurring along a tidal gradient as well as along horizontal gradient of the mud bank during growth (January – March) and reproductive (September – October) periods was investigated. Changes in the distribution of females between the two study periods were significant, while the distribution of reproductively active females were closely linked to the low water mark (Lower tidal levels). Females occurring in this zone were also significantly larger than females occurring towards the back of the study site. Some minor changes along the horizontal gradient were also observed. The sediment compos ition of the mud bank was found not to play a role in this distribution, leading to the suggestion that exposure to ebb and flood tide currents could influence female prawn distributions. Management recommendations based on these observations were made.
- Full Text:
- Date Issued: 2003
- Authors: Jooste, Jakobus Gerrit
- Date: 2003
- Subjects: Upogebia african -- Effects of sediments on -- South Africa -- Eastern Cape , Estuarine ecology -- South Africa -- Eastern Cape , Fishing baits -- Conservation -- South Africa -- Eastern Cape
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11073 , http://hdl.handle.net/10948/334 , Upogebia african -- Effects of sediments on -- South Africa -- Eastern Cape , Estuarine ecology -- South Africa -- Eastern Cape , Fishing baits -- Conservation -- South Africa -- Eastern Cape
- Description: Aspects around the management of benthic soft sediment bait organisms, with special focus on the mud prawn (Upogebia africana) of eastern Cape estuaries was investigated. The recreational linefishery of the Gamtoos estuary was described, and compared to previous studies. Target fish species was identified, and a large dependency on bait sourced from estuaries needed for the capture of these species was noted, especially for spotted grunter (Pomadasys commersonnii). A comparison between bait use, success and the natural diet of target fish species was also made. The structure and distribution of sediments in the old channel mud banks was described and compared with historical data. The influence of sediments on mud prawn distribution was investigated, but no significant interactions were found at the study site. The impacts of once-off pumping and digging events, and monthly trampling on the sediments and mud prawn population was studied over a 7-month period. Initial removal rates as well as recovery time varied significantly between the two collection methods, while the largest decline in prawn numbers (to zero) with no recovery visible after seven months was caused by trampling. All disturbances caused some alteration in sediment composition, but not to such an extent that the sediments became unfavorable for mud prawns. Trampling did, however, result in the compaction of sediments to such a degree that prawns could not construct burrows. Issues around current removal quotas of bait species as well as the creation of a small-scale commercial (SSC) bait selling operation at Swartkops estuary were critically evaluated, and suggestions for the future removal rates of mud prawns based on production export calculations were made. The distribution, size, sex ratios and number of gravid females occurring along a tidal gradient as well as along horizontal gradient of the mud bank during growth (January – March) and reproductive (September – October) periods was investigated. Changes in the distribution of females between the two study periods were significant, while the distribution of reproductively active females were closely linked to the low water mark (Lower tidal levels). Females occurring in this zone were also significantly larger than females occurring towards the back of the study site. Some minor changes along the horizontal gradient were also observed. The sediment compos ition of the mud bank was found not to play a role in this distribution, leading to the suggestion that exposure to ebb and flood tide currents could influence female prawn distributions. Management recommendations based on these observations were made.
- Full Text:
- Date Issued: 2003
An evaluation of the Xenopus laevis liver slice model to study the toxic effects of microcystin
- Authors: Coates, Nadya
- Date: 2003
- Subjects: Zenopus laevis , Microcystis aeruginosa -- Toxicology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11069 , http://hdl.handle.net/10948/307 , Zenopus laevis , Microcystis aeruginosa -- Toxicology
- Description: Blooms of cyanobacteria have increased in occurrence in the past three decades and have been reported to cause severe problems for animals and humans, leading to death in extreme instances. The majority of poisonings that have taken place have been attributed to a hepatotoxin produced by the species Microcystis aeruginosa, namely microcystin. The appearance of a cyanobacterial bloom does not give any indication as to its toxicity and therefore, it is imperative that simple, yet sensitive, bioassays are developed to overcome this problem. This study was undertaken to evaluate the effects of microcystin-LR on the liver of Xenopus laevis both in vitro and in vivo. This animal provides an opportunity to study the long-term hepatotoxic effects of the toxin compared to in vitro studies performed with mice and rats. The use of the liver slice model system as a potential bioassay to study the effects of microcystin-LR on Xenopus laevis liver was evaluated. Liver slices were cultured in RPMI- 1640 culture medium for periods ranging from 30 hours to 10 days and the liver slices were exposed to toxin concentrations ranging from 1nM to 500nM. The use of frog liver slices to study the longer-term effects of low-dose exposure to microcystin-LR was evaluated by observing the ultrastructural changes within hepatocytes using transmission electron microscopy, the release of the enzymes alanine aminotransferase and lactate dehydrogenase into the surrounding culture medium, as well as using a 3-[4,5-dimethylthiazol-2yl]-2,5- diphenyl tetrazolium bromide assay to determine the viability of the liver slices in culture. The amount of lipid peroxidation in the liver slices after exposure to microcystin-LR was assessed using the Thiobarbituric Acid Test. Results showed the frog liver slice culture system to be an inadequate method to evaluate the hepatotoxic effects of microcystin-LR. An in vivo assessment of the effects of microcystin-LR on Xenopus laevis was carried out using a total of 9 frogs (3 groups of 3 frogs). Frogs received a single intraperitoneal dose of 120mg/kg of microcystin-LR and were sacrificed at 8 and 24 hours post exposure. Microcystin-LR caused no significant change in serum lactate dehydrogenase levels, hepatosomatic index (liver weight as a percentage body weight), glutathione peroxidase activity, glycogen or lipid peroxidation. There was, however, an increase in glutathione sii transferase activity in the liver. The presence of the toxin in the liver was confirmed by immunohistochemistry. This study suggests that Xenopus laevis has, in some way, adapted to detoxifying aquatic toxins in the environment.
- Full Text:
- Date Issued: 2003
- Authors: Coates, Nadya
- Date: 2003
- Subjects: Zenopus laevis , Microcystis aeruginosa -- Toxicology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11069 , http://hdl.handle.net/10948/307 , Zenopus laevis , Microcystis aeruginosa -- Toxicology
- Description: Blooms of cyanobacteria have increased in occurrence in the past three decades and have been reported to cause severe problems for animals and humans, leading to death in extreme instances. The majority of poisonings that have taken place have been attributed to a hepatotoxin produced by the species Microcystis aeruginosa, namely microcystin. The appearance of a cyanobacterial bloom does not give any indication as to its toxicity and therefore, it is imperative that simple, yet sensitive, bioassays are developed to overcome this problem. This study was undertaken to evaluate the effects of microcystin-LR on the liver of Xenopus laevis both in vitro and in vivo. This animal provides an opportunity to study the long-term hepatotoxic effects of the toxin compared to in vitro studies performed with mice and rats. The use of the liver slice model system as a potential bioassay to study the effects of microcystin-LR on Xenopus laevis liver was evaluated. Liver slices were cultured in RPMI- 1640 culture medium for periods ranging from 30 hours to 10 days and the liver slices were exposed to toxin concentrations ranging from 1nM to 500nM. The use of frog liver slices to study the longer-term effects of low-dose exposure to microcystin-LR was evaluated by observing the ultrastructural changes within hepatocytes using transmission electron microscopy, the release of the enzymes alanine aminotransferase and lactate dehydrogenase into the surrounding culture medium, as well as using a 3-[4,5-dimethylthiazol-2yl]-2,5- diphenyl tetrazolium bromide assay to determine the viability of the liver slices in culture. The amount of lipid peroxidation in the liver slices after exposure to microcystin-LR was assessed using the Thiobarbituric Acid Test. Results showed the frog liver slice culture system to be an inadequate method to evaluate the hepatotoxic effects of microcystin-LR. An in vivo assessment of the effects of microcystin-LR on Xenopus laevis was carried out using a total of 9 frogs (3 groups of 3 frogs). Frogs received a single intraperitoneal dose of 120mg/kg of microcystin-LR and were sacrificed at 8 and 24 hours post exposure. Microcystin-LR caused no significant change in serum lactate dehydrogenase levels, hepatosomatic index (liver weight as a percentage body weight), glutathione peroxidase activity, glycogen or lipid peroxidation. There was, however, an increase in glutathione sii transferase activity in the liver. The presence of the toxin in the liver was confirmed by immunohistochemistry. This study suggests that Xenopus laevis has, in some way, adapted to detoxifying aquatic toxins in the environment.
- Full Text:
- Date Issued: 2003
An investigation into the induction of oxidative stress and apoptosis by microcystin-LR in the CaCo2 cell line and intestinal tract of Balb/c mice
- Authors: Botha, Nicolette
- Date: 2003
- Subjects: Microcystis aeruginosa -- Toxicology , Apoptosis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11066 , http://hdl.handle.net/10948/349 , Microcystis aeruginosa -- Toxicology , Apoptosis
- Description: This study reports the findings on the effect of Microcystin-LR (MCLR) on the gastrointestinal tract cells of mice and on two different cell lines, Caco2 and MCF-7. The cyanobacterium Microcystis aeruginosa produces the potent toxin, MCLR. This toxin has been implicated in a number of cases of ill-health. It was decided to investigate whether microcystin-LR induced apoptosis in the gastrointestinal tract of mice and also which possible mechanisms were involved in the induction in vitro. Balb/c mice were given a 75% LD50 intraperitoneal dose of pure microcystin -LR and sacrificed at 8, 16, 24 and 32 hours post-exposure. The small intestinal sections were stained with haematoxylin and eosin and examined for apoptotic cells. There was a time-dependent increase in the number of apoptotic cells with most in the duodenum and the jejunum. No change in glycogen content was evident at 24 hours post exposure when PAS-stained sections were examined. To determine that microcystin was the agent responsible for the changes, fluoroscein isothiocyanate (FITC) immunostaining for the toxin was done on the sections. Apoptosis in vitro was investigated in Caco2, a cell line that behaves like normal enterocytes when the cells are differentiated at confluency, and MCF-7, a breast cancer cell line deficient in pro-caspase-3, cells by 3-[dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays and by staining with DAPI and Rhodamine 123. MCLR exposure induced apoptosis, as seen in decreased cell viability and increased leakage of LDH, as well as mitochondrial damage shown by Rhodamine staining. The MCF-7 cells, deficient in pro-caspase-3, and Caco2 cells did not show cleavage of poly(ADP)ribose polymerase (PARP) after exposure to 50μM MCLR after 72 hours exposure. Both micro- and milli-calpain activity was however significantly increased in both cell lines exposed to the toxin. There was a significant increase in H2O2, one of the key reactive oxygen species, production during the first 30 minutes that the cells were exposed to 50 mM MCLR.
- Full Text:
- Date Issued: 2003
- Authors: Botha, Nicolette
- Date: 2003
- Subjects: Microcystis aeruginosa -- Toxicology , Apoptosis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11066 , http://hdl.handle.net/10948/349 , Microcystis aeruginosa -- Toxicology , Apoptosis
- Description: This study reports the findings on the effect of Microcystin-LR (MCLR) on the gastrointestinal tract cells of mice and on two different cell lines, Caco2 and MCF-7. The cyanobacterium Microcystis aeruginosa produces the potent toxin, MCLR. This toxin has been implicated in a number of cases of ill-health. It was decided to investigate whether microcystin-LR induced apoptosis in the gastrointestinal tract of mice and also which possible mechanisms were involved in the induction in vitro. Balb/c mice were given a 75% LD50 intraperitoneal dose of pure microcystin -LR and sacrificed at 8, 16, 24 and 32 hours post-exposure. The small intestinal sections were stained with haematoxylin and eosin and examined for apoptotic cells. There was a time-dependent increase in the number of apoptotic cells with most in the duodenum and the jejunum. No change in glycogen content was evident at 24 hours post exposure when PAS-stained sections were examined. To determine that microcystin was the agent responsible for the changes, fluoroscein isothiocyanate (FITC) immunostaining for the toxin was done on the sections. Apoptosis in vitro was investigated in Caco2, a cell line that behaves like normal enterocytes when the cells are differentiated at confluency, and MCF-7, a breast cancer cell line deficient in pro-caspase-3, cells by 3-[dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays and by staining with DAPI and Rhodamine 123. MCLR exposure induced apoptosis, as seen in decreased cell viability and increased leakage of LDH, as well as mitochondrial damage shown by Rhodamine staining. The MCF-7 cells, deficient in pro-caspase-3, and Caco2 cells did not show cleavage of poly(ADP)ribose polymerase (PARP) after exposure to 50μM MCLR after 72 hours exposure. Both micro- and milli-calpain activity was however significantly increased in both cell lines exposed to the toxin. There was a significant increase in H2O2, one of the key reactive oxygen species, production during the first 30 minutes that the cells were exposed to 50 mM MCLR.
- Full Text:
- Date Issued: 2003
Investigations into the asymmetric reduction of ketones
- Authors: Bena, Luvuyo Clifford
- Date: 2003
- Subjects: Ketones , Asymmetric synthesis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11065 , http://hdl.handle.net/10948/323 , Ketones , Asymmetric synthesis
- Description: A six-step synthesis of salbutamol from methyl salicylate with an overall yield of 17% has been completed, although the yield was not optimised. In the process, Zn(BH4)2 was found to selectively reduce a ketone carbonyl group in the presence of an ester unit. In contrast, borane was found to reduce both the ketone and ester carbonyl groups. Reduction of phenacyl bromide with borane in the presence of chiral catalysts based on (R)-alaninol and (R,S)-ephidrine resulted a measure of enantioselectivity in the product. However, the configuration of the alcohol obtained in the case of (R)-alaninol was contrary to expectations based both on experimental trends observed elsewhere as well as our own theoretical predictions. The asymmetric reduction of methyl 5-bromoacetyl-2-benzyloxybenzoate was accomplished with both borane and Zn(BH4)2 in the presence of a range of chiral catalysts. Optically active products were obtained in all cases, although the optical rotations were significantly smaller in the case of Zn(BH4)2. Unfortunately, we were not successful in determining the enantiomeric excesses of these reactions. The use of a NMR lanthanide shift reagent resulted in a complex spectrum that was impossible to interpret unambiguously. This presumably arises from the presence of several Lewis base sites in the product at which complexation with the shift reagent can take place. It was also not possible to determine the optical rotation of salbutamol itself owing to the relatively small amount of material obtained. A conformational analysis of salbutamol, where NMR data was correlated with molecular modelling results, was successfully carried out and revealed a strong preference for that conformer family characterised by O–C–C–N and Ar–C–C–N torsion angles of ca. 60º and 180º, respectively. Interestingly, these conformers are found to be stabilised by OH…N rather than NH…O hydrogen bonding. This study has also confirmed the effectiveness of the MMFF94 force field for conformational analysis studies in compounds of this kind. Lastly, a relatively simple method for modelling the BH3/oxazaborolidine reduction of ketones at the PM3 semiempirical MO level of approximation was devised. This approach has provided insights into the mechanism of the reaction and has furthermore enabled us to predict the enantioselectivities likely to result from various catalysts and ketones. In comparing our theoretical and experimental findings, an anomalous result was observed in the case of (R)-alaninol; this will have to be investigated further, particularly at the experimental level. However, we believe that our approach provides a sound basis for aiding the design and screening of new, potentially better catalysts.
- Full Text:
- Date Issued: 2003
- Authors: Bena, Luvuyo Clifford
- Date: 2003
- Subjects: Ketones , Asymmetric synthesis
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11065 , http://hdl.handle.net/10948/323 , Ketones , Asymmetric synthesis
- Description: A six-step synthesis of salbutamol from methyl salicylate with an overall yield of 17% has been completed, although the yield was not optimised. In the process, Zn(BH4)2 was found to selectively reduce a ketone carbonyl group in the presence of an ester unit. In contrast, borane was found to reduce both the ketone and ester carbonyl groups. Reduction of phenacyl bromide with borane in the presence of chiral catalysts based on (R)-alaninol and (R,S)-ephidrine resulted a measure of enantioselectivity in the product. However, the configuration of the alcohol obtained in the case of (R)-alaninol was contrary to expectations based both on experimental trends observed elsewhere as well as our own theoretical predictions. The asymmetric reduction of methyl 5-bromoacetyl-2-benzyloxybenzoate was accomplished with both borane and Zn(BH4)2 in the presence of a range of chiral catalysts. Optically active products were obtained in all cases, although the optical rotations were significantly smaller in the case of Zn(BH4)2. Unfortunately, we were not successful in determining the enantiomeric excesses of these reactions. The use of a NMR lanthanide shift reagent resulted in a complex spectrum that was impossible to interpret unambiguously. This presumably arises from the presence of several Lewis base sites in the product at which complexation with the shift reagent can take place. It was also not possible to determine the optical rotation of salbutamol itself owing to the relatively small amount of material obtained. A conformational analysis of salbutamol, where NMR data was correlated with molecular modelling results, was successfully carried out and revealed a strong preference for that conformer family characterised by O–C–C–N and Ar–C–C–N torsion angles of ca. 60º and 180º, respectively. Interestingly, these conformers are found to be stabilised by OH…N rather than NH…O hydrogen bonding. This study has also confirmed the effectiveness of the MMFF94 force field for conformational analysis studies in compounds of this kind. Lastly, a relatively simple method for modelling the BH3/oxazaborolidine reduction of ketones at the PM3 semiempirical MO level of approximation was devised. This approach has provided insights into the mechanism of the reaction and has furthermore enabled us to predict the enantioselectivities likely to result from various catalysts and ketones. In comparing our theoretical and experimental findings, an anomalous result was observed in the case of (R)-alaninol; this will have to be investigated further, particularly at the experimental level. However, we believe that our approach provides a sound basis for aiding the design and screening of new, potentially better catalysts.
- Full Text:
- Date Issued: 2003
Observed metabolic changes in male Wistar rats after treatment with an antidepressant implied in undesirable weight gain, or Sutherlandia frutescens for Type II diabetes
- Authors: Chadwick, Wayne
- Date: 2003
- Subjects: Rats -- Metabolism , Non-insulin-dependent diabetes -- Research , Rats as laboratory animals
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11068 , http://hdl.handle.net/10948/313 , Rats -- Metabolism , Non-insulin-dependent diabetes -- Research , Rats as laboratory animals
- Description: Type II diabetes is fast becoming a growing problem in developed countries worldwide. Traditionally the median age for diagnosis was around sixty, but recent surveys have shown that the entire age distribution curve has shifted to the left. Western countries boast the worst statistics in which type II diabetes is being reported in children under the age of ten. At such a young age the disease often goes undiagnosed for long periods of time allowing considerable damage to occur. The incidence of type II diabetes is thought to be parallel with the growing rate of obesity associated with a characteristically unhealthy western diet. Type II diabetes is an extremely expensive disease to manage, and with the rapid growth of this pandemic our country will soon feel the economic burden of this disease. It is for this reason that cheaper medication needs to be investigated in the form of traditional plants, such as Sutherlandia frutescens. Prescription medication, such as tricyclic antidepressants, may also increase body weight or appetite thereby playing a role in obesity. The cause of weight gain in such cases may go unrecognized or lead to cessation of the medication with or without the practitioner’s knowledge or approval. It is therefore necessary to investigate the causative agents responsible for the excessive weight gain. Drinking water containing extracts of the S. frutescens, metformin (a well known type II diabetes medication) and amitriptyline (a common tricyclic antidepressant) was administered to three groups of ten male Wistar rats. The control group received water without any medication. The rat’s weight and food consumption was monitored throughout the trial and their oxygen consumption was also determined. Rats were sacrificed after four months of medicinal compliance and glucose uptake, in the presence and absence of insulin, was tested in epididymal fat, liver and muscle. Fasting plasma glucose levels, lipoprotein, cholesterol and triglyceride concentrations were also determined.
- Full Text:
- Date Issued: 2003
- Authors: Chadwick, Wayne
- Date: 2003
- Subjects: Rats -- Metabolism , Non-insulin-dependent diabetes -- Research , Rats as laboratory animals
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11068 , http://hdl.handle.net/10948/313 , Rats -- Metabolism , Non-insulin-dependent diabetes -- Research , Rats as laboratory animals
- Description: Type II diabetes is fast becoming a growing problem in developed countries worldwide. Traditionally the median age for diagnosis was around sixty, but recent surveys have shown that the entire age distribution curve has shifted to the left. Western countries boast the worst statistics in which type II diabetes is being reported in children under the age of ten. At such a young age the disease often goes undiagnosed for long periods of time allowing considerable damage to occur. The incidence of type II diabetes is thought to be parallel with the growing rate of obesity associated with a characteristically unhealthy western diet. Type II diabetes is an extremely expensive disease to manage, and with the rapid growth of this pandemic our country will soon feel the economic burden of this disease. It is for this reason that cheaper medication needs to be investigated in the form of traditional plants, such as Sutherlandia frutescens. Prescription medication, such as tricyclic antidepressants, may also increase body weight or appetite thereby playing a role in obesity. The cause of weight gain in such cases may go unrecognized or lead to cessation of the medication with or without the practitioner’s knowledge or approval. It is therefore necessary to investigate the causative agents responsible for the excessive weight gain. Drinking water containing extracts of the S. frutescens, metformin (a well known type II diabetes medication) and amitriptyline (a common tricyclic antidepressant) was administered to three groups of ten male Wistar rats. The control group received water without any medication. The rat’s weight and food consumption was monitored throughout the trial and their oxygen consumption was also determined. Rats were sacrificed after four months of medicinal compliance and glucose uptake, in the presence and absence of insulin, was tested in epididymal fat, liver and muscle. Fasting plasma glucose levels, lipoprotein, cholesterol and triglyceride concentrations were also determined.
- Full Text:
- Date Issued: 2003
The effect of human soluble FceRII on the RPMI 8866 B-Lymphoblastoid and the U937 Monocyte cell lines
- Authors: Daniels, Brodie Belinda
- Date: 2003
- Subjects: Developmental inmmunology , Cellular control mechanisms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11084 , http://hdl.handle.net/10948/322 , Developmental inmmunology , Cellular control mechanisms
- Description: Due to the diverse functions of Fc eRII, such as its roles in cellular adhesion, growth and differentiation of B and T lymphocytes, rescue of B cells from apoptosis and release of cytotoxic mediators, it is clear why it is believed to be a central molecule in allergic response. Because of its important role in the regulation of IgE production, FceRII may be the primary cause of certain allergic conditions. This study attempted to express and purify a recombinant human soluble FceRII to test its effect on a B-lymphoblastoid (RPMI 8866) and a monocytic (U937) cell line. The protein was expressed in Escherichia coli inclusion bodies, before being refolded and purified in a single gel chromatography step. This pure protein was then tested for biological activity by testing its IgE binding func tion. Once proven functional, it was used to test its effect on the cell lines at three concentrations for its apoptotic rescue properties and its cytokine effects. The recombinant protein did not seem to have any significant effect on the apoptotic rescue of either cell line. While the recombinant sFceRII appeared to have a slight effect on the stimulation of IL-1ß and TNFa in the RPMI 8866 cells, there was no apparent effect on the production of NF?B. In U937 cells, the protein did not seem to have any effect on the stimulation of IL-1ß, TNFa or NF?B. However, the cytokine effects of the recombinant protein were tested on isolated PBMCs from a healthy individual and a hyper-IgE syndrome patient. The recombinant protein was able to stimulate the production of cytokines in both individuals’ PBMCs, proving that it has the same effect as the natural protein. The upregulation of these cytokines indicates that the recombinant protein is able to stimulate the immune system. Therefore, this recombinant soluble FceRII protein could possibly be used for immune therapy.
- Full Text:
- Date Issued: 2003
- Authors: Daniels, Brodie Belinda
- Date: 2003
- Subjects: Developmental inmmunology , Cellular control mechanisms
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11084 , http://hdl.handle.net/10948/322 , Developmental inmmunology , Cellular control mechanisms
- Description: Due to the diverse functions of Fc eRII, such as its roles in cellular adhesion, growth and differentiation of B and T lymphocytes, rescue of B cells from apoptosis and release of cytotoxic mediators, it is clear why it is believed to be a central molecule in allergic response. Because of its important role in the regulation of IgE production, FceRII may be the primary cause of certain allergic conditions. This study attempted to express and purify a recombinant human soluble FceRII to test its effect on a B-lymphoblastoid (RPMI 8866) and a monocytic (U937) cell line. The protein was expressed in Escherichia coli inclusion bodies, before being refolded and purified in a single gel chromatography step. This pure protein was then tested for biological activity by testing its IgE binding func tion. Once proven functional, it was used to test its effect on the cell lines at three concentrations for its apoptotic rescue properties and its cytokine effects. The recombinant protein did not seem to have any significant effect on the apoptotic rescue of either cell line. While the recombinant sFceRII appeared to have a slight effect on the stimulation of IL-1ß and TNFa in the RPMI 8866 cells, there was no apparent effect on the production of NF?B. In U937 cells, the protein did not seem to have any effect on the stimulation of IL-1ß, TNFa or NF?B. However, the cytokine effects of the recombinant protein were tested on isolated PBMCs from a healthy individual and a hyper-IgE syndrome patient. The recombinant protein was able to stimulate the production of cytokines in both individuals’ PBMCs, proving that it has the same effect as the natural protein. The upregulation of these cytokines indicates that the recombinant protein is able to stimulate the immune system. Therefore, this recombinant soluble FceRII protein could possibly be used for immune therapy.
- Full Text:
- Date Issued: 2003
The specification and design of a prototype 2-D MPEG-4 authoring tool
- Authors: Viljoen, Deon Walter
- Date: 2003
- Subjects: MPEG (Video coding standard) , Digital video -- Standards , Multimedia systems
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11092 , http://hdl.handle.net/10948/d1015741 , MPEG (Video coding standard) , Digital video -- Standards , Multimedia systems
- Description: The purpose of this project was the specification, design and implementation of a prototype 2-D MPEG-4 authoring tool. A literature study was conducted of the MPEG-4 standard and multimedia authoring tools to determine the specification and design of a prototype 2- D MPEG-4 authoring tool. The specification and design was used as a basis for the implementation of a prototype 2-D MPEG-4 authoring tool that complies with the Complete 2-D Scene Graph Profile. The need for research into MPEG-4 authoring tools arose from the reported lack of knowledge of the MPEG-4 standard and the limited implementations of MPEG-4 authoring tools available to content authors. In order for MPEG-4 to reach its full potential, it will require authoring tools and content players that satisfy the needs of its users. The theoretical component of this dissertation included a literature study of the MPEG-4 standard and an investigation of relevant multimedia authoring systems. MPEG-4 was introduced as a standard that allows for the creation and streaming of interactive multimedia content at variable bit rates over high and low bandwidth connections. The requirements for the prototype 2-D MPEG-4 authoring system were documented and a prototype system satisfying the requirements was designed, implemented and evaluated. The evaluation of the prototype system showed that the system successfully satisfied all its requirements and that it provides the user with an easy to use and intuitive authoring tool. MPEG-4 has the potential to satisfy the increasing demand for innovative multimedia content on low bandwidth networks, including the Internet and mobile networks, as well as the need expressed by users to interact with multimedia content. This dissertation makes an important contribution to the understanding of the MPEG-4 standard, its functionality and the design of a 2-D MPEG-4 Authoring tool. Keywords: MPEG-4; MPEG-4 authoring; Binary Format for Scenes.
- Full Text:
- Date Issued: 2003
- Authors: Viljoen, Deon Walter
- Date: 2003
- Subjects: MPEG (Video coding standard) , Digital video -- Standards , Multimedia systems
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11092 , http://hdl.handle.net/10948/d1015741 , MPEG (Video coding standard) , Digital video -- Standards , Multimedia systems
- Description: The purpose of this project was the specification, design and implementation of a prototype 2-D MPEG-4 authoring tool. A literature study was conducted of the MPEG-4 standard and multimedia authoring tools to determine the specification and design of a prototype 2- D MPEG-4 authoring tool. The specification and design was used as a basis for the implementation of a prototype 2-D MPEG-4 authoring tool that complies with the Complete 2-D Scene Graph Profile. The need for research into MPEG-4 authoring tools arose from the reported lack of knowledge of the MPEG-4 standard and the limited implementations of MPEG-4 authoring tools available to content authors. In order for MPEG-4 to reach its full potential, it will require authoring tools and content players that satisfy the needs of its users. The theoretical component of this dissertation included a literature study of the MPEG-4 standard and an investigation of relevant multimedia authoring systems. MPEG-4 was introduced as a standard that allows for the creation and streaming of interactive multimedia content at variable bit rates over high and low bandwidth connections. The requirements for the prototype 2-D MPEG-4 authoring system were documented and a prototype system satisfying the requirements was designed, implemented and evaluated. The evaluation of the prototype system showed that the system successfully satisfied all its requirements and that it provides the user with an easy to use and intuitive authoring tool. MPEG-4 has the potential to satisfy the increasing demand for innovative multimedia content on low bandwidth networks, including the Internet and mobile networks, as well as the need expressed by users to interact with multimedia content. This dissertation makes an important contribution to the understanding of the MPEG-4 standard, its functionality and the design of a 2-D MPEG-4 Authoring tool. Keywords: MPEG-4; MPEG-4 authoring; Binary Format for Scenes.
- Full Text:
- Date Issued: 2003
Using E-learning to support IT education in a university environment a case study approach
- Authors: Taljaard, Marinda
- Date: 2003
- Subjects: Education, Higher -- South Africa -- Port Elizabeth -- Computer-assisted instruction , Internet in education , Information technology -- Study and teaching (Higher) -- South Africa -- Port Elizabeth , College teaching -- South Africa -- Port Elizabeth -- Computer network resources , University of Port Elizabeth
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11091 , http://hdl.handle.net/10948/d1015740 , Education, Higher -- South Africa -- Port Elizabeth -- Computer-assisted instruction , Internet in education , Information technology -- Study and teaching (Higher) -- South Africa -- Port Elizabeth , College teaching -- South Africa -- Port Elizabeth -- Computer network resources , University of Port Elizabeth
- Description: At the University of Port Elizabeth (UPE), the End User Computing course (EUC) acts as a service course for many departments. This implies that many students are forced by their curricula to register for this course. The ever-increasing numbers in EUC place a considerable load on existing human and physical resources. In lecture groups of 120 –160, students rarely get the attention they need, and the pace at which the content is delivered (too slow or too fast) may also inhibit the learning process. During an initial investigation into E-learning at UPE in 1999, a prototype virtual classroom was developed. There were, however, a number of problems with this prototype. Firstly, it was implemented using a number of different technologies, which made it difficult to extend and maintain. Secondly, it only addressed some aspects of an E-learning environment, which proved insufficient for the EUC course. In the existing EUC course at UPE, the students are already exposed to some E-learning concepts, as a section of their skills training component is handled by using multimedia software in a simulated environment. The objective of this project was to extend the E-learning component further to determine the advantages and disadvantages of using E-learning to support information technology (IT) education in a contact-university environment. This project included a literature search and survey of existing E-learning environments at other universities. This research was used to develop a draft framework for an E-learning environment. The framework was used to select a tool to create an E-learning environment at UPE. An experiment was designed using this E-learning environment to support two IT courses at different year levels. The results of the experiment were analysed using qualitative and quantitative methods to determine the impact of using E-learning to support IT education at UPE. The results of this research show that E-learning can be used to support IT education at UPE. More success, however, was achieved at postgraduate level than at first-year level. Making use of Elearning increased student satisfaction and promoted active learning, while providing benefits like convenience, communication, flexibility and scaffolding. We conclude, therefore, that E-learning can provide a flexible approach to IT education in a university environment in the future.
- Full Text:
- Date Issued: 2003
- Authors: Taljaard, Marinda
- Date: 2003
- Subjects: Education, Higher -- South Africa -- Port Elizabeth -- Computer-assisted instruction , Internet in education , Information technology -- Study and teaching (Higher) -- South Africa -- Port Elizabeth , College teaching -- South Africa -- Port Elizabeth -- Computer network resources , University of Port Elizabeth
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11091 , http://hdl.handle.net/10948/d1015740 , Education, Higher -- South Africa -- Port Elizabeth -- Computer-assisted instruction , Internet in education , Information technology -- Study and teaching (Higher) -- South Africa -- Port Elizabeth , College teaching -- South Africa -- Port Elizabeth -- Computer network resources , University of Port Elizabeth
- Description: At the University of Port Elizabeth (UPE), the End User Computing course (EUC) acts as a service course for many departments. This implies that many students are forced by their curricula to register for this course. The ever-increasing numbers in EUC place a considerable load on existing human and physical resources. In lecture groups of 120 –160, students rarely get the attention they need, and the pace at which the content is delivered (too slow or too fast) may also inhibit the learning process. During an initial investigation into E-learning at UPE in 1999, a prototype virtual classroom was developed. There were, however, a number of problems with this prototype. Firstly, it was implemented using a number of different technologies, which made it difficult to extend and maintain. Secondly, it only addressed some aspects of an E-learning environment, which proved insufficient for the EUC course. In the existing EUC course at UPE, the students are already exposed to some E-learning concepts, as a section of their skills training component is handled by using multimedia software in a simulated environment. The objective of this project was to extend the E-learning component further to determine the advantages and disadvantages of using E-learning to support information technology (IT) education in a contact-university environment. This project included a literature search and survey of existing E-learning environments at other universities. This research was used to develop a draft framework for an E-learning environment. The framework was used to select a tool to create an E-learning environment at UPE. An experiment was designed using this E-learning environment to support two IT courses at different year levels. The results of the experiment were analysed using qualitative and quantitative methods to determine the impact of using E-learning to support IT education at UPE. The results of this research show that E-learning can be used to support IT education at UPE. More success, however, was achieved at postgraduate level than at first-year level. Making use of Elearning increased student satisfaction and promoted active learning, while providing benefits like convenience, communication, flexibility and scaffolding. We conclude, therefore, that E-learning can provide a flexible approach to IT education in a university environment in the future.
- Full Text:
- Date Issued: 2003
The effect of nutrient levels and ratios on the growth of Microcystis aeruginosa and microcystin production
- Authors: Sember, Craig Stewart
- Date: 2002
- Subjects: Microcystis aeruginosa -- Toxicology , Nitrates , Microcystins
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11076 , http://hdl.handle.net/10948/287 , Microcystis aeruginosa -- Toxicology , Nitrates , Microcystins
- Description: This study reports the findings on the effect of nitrates and phosphates on the biomass and toxin production of various strains of the unicellular non-nitrogen fixing cyanobacterium, Microcystis aeruginosa. The occurrence of blooms of Microcystis aeruginosa and microcystin in freshwater impoundments across the globe has been on the increase lately due to increased levels of eutrophication, resulting in human and animal deaths and illness, as well as drinking and recreational water foulment. A range of environmental factors have been shown to effect growth and microcystin production. Existing literature however is somewhat contradictory as to the effects of these physical and chemical factors on toxin production. Therefore Microcystis aeruginosa strains were cultured under batch and continuous conditions to determine the effect of nitrate and phosphate concentrations and ratios on biomass and toxin production. Cultures were analysed with regards to internal nutrient stores, biomass production, nutrient depletion, photosynthetic efficiency and microcystin production. Results showed that microcystin production correlated to growth rate, photosynthetic efficiency and internal nitrogen stores and that an optimal N:P ratio was associated with microcystin levels, growth rate and photosynthetic efficiency. Results therefore led to the conclusion that the nitrogen, carbon, and phosphate balance within the cell is closely associated with microcystin production. Whether or not microcystin is produced to maintain this balance or produced as a function of this balance remains to be determined.
- Full Text:
- Date Issued: 2002
- Authors: Sember, Craig Stewart
- Date: 2002
- Subjects: Microcystis aeruginosa -- Toxicology , Nitrates , Microcystins
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11076 , http://hdl.handle.net/10948/287 , Microcystis aeruginosa -- Toxicology , Nitrates , Microcystins
- Description: This study reports the findings on the effect of nitrates and phosphates on the biomass and toxin production of various strains of the unicellular non-nitrogen fixing cyanobacterium, Microcystis aeruginosa. The occurrence of blooms of Microcystis aeruginosa and microcystin in freshwater impoundments across the globe has been on the increase lately due to increased levels of eutrophication, resulting in human and animal deaths and illness, as well as drinking and recreational water foulment. A range of environmental factors have been shown to effect growth and microcystin production. Existing literature however is somewhat contradictory as to the effects of these physical and chemical factors on toxin production. Therefore Microcystis aeruginosa strains were cultured under batch and continuous conditions to determine the effect of nitrate and phosphate concentrations and ratios on biomass and toxin production. Cultures were analysed with regards to internal nutrient stores, biomass production, nutrient depletion, photosynthetic efficiency and microcystin production. Results showed that microcystin production correlated to growth rate, photosynthetic efficiency and internal nitrogen stores and that an optimal N:P ratio was associated with microcystin levels, growth rate and photosynthetic efficiency. Results therefore led to the conclusion that the nitrogen, carbon, and phosphate balance within the cell is closely associated with microcystin production. Whether or not microcystin is produced to maintain this balance or produced as a function of this balance remains to be determined.
- Full Text:
- Date Issued: 2002
The effect of selenium in the detoxification of the microcystin hepatotoxins
- Authors: Downs, Kerry
- Date: 2002
- Subjects: Cynaobacterial toxins , Microcystins , Selenium
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11070 , http://hdl.handle.net/10948/284 , Cynaobacterial toxins , Microcystins , Selenium
- Description: Blooms of cyanobacteria have been known to cause illness in humans and death in wild and domestic animals. One of the toxins produced by cyanobacteria is microcystin, which is a potent hepatotoxin. Microcystin is taken up by bile acid transporters in the intestine and transported into the liver. After exposure to acute doses of microcystin, severe haemorrhage has been observed along with apoptotic and necrotic hepatocytes. The cytoskeletal structure of the hepatocytes is disrupted and oxidative stress is induced. Selenium, a known anti-oxidant, has been shown to induce increased activity of glutathione peroxidase. Glutathione peroxidase removes peroxides from cells protecting them from oxidative stress. This study set out to determine if selenium could play a role in preventing the damage to mice livers due to microcystin toxin. The protective role of selenium was explored in three main studies: in the first study, the ability of selenium to increase the survival time of mice exposed to a lethal dose of toxin was determined. In the second study the mice were exposed to sublethal chronic doses of toxin over 30 days. The ability of selenium to minimise liver damage under these conditions was determined. The final study investigated the mechanism of the protective effect of selenium. The results of the first study suggested that selenium could extend survival time. In the second study the selenium supplemented mice showed a reduction in the extent of the increase in liver weight and a decrease in the amount of lipid peroxidation induced compared to the mice that received only toxin. The histology of the selenium supplemented mice also showed a decrease in the severity and amount of morphological changes in the liver. The third study indicated that the protection shown by selenium might be mediated by an increase in the glutathione peroxidase (GPX) activity in selenium supplemented mice. This increase in GPX activity would increase the removal of the lipid hydroperoxides and prevent the damage they would cause in the cell. A further result indicated an increase in glutathione S-transferase in only the toxin control mice when compared to the selenium supplemented and control mice. ii In conclusion selenium offers protection against microcystin but further studies need to be done to provide statistically valid results to clarify the level of protection.
- Full Text:
- Date Issued: 2002
- Authors: Downs, Kerry
- Date: 2002
- Subjects: Cynaobacterial toxins , Microcystins , Selenium
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11070 , http://hdl.handle.net/10948/284 , Cynaobacterial toxins , Microcystins , Selenium
- Description: Blooms of cyanobacteria have been known to cause illness in humans and death in wild and domestic animals. One of the toxins produced by cyanobacteria is microcystin, which is a potent hepatotoxin. Microcystin is taken up by bile acid transporters in the intestine and transported into the liver. After exposure to acute doses of microcystin, severe haemorrhage has been observed along with apoptotic and necrotic hepatocytes. The cytoskeletal structure of the hepatocytes is disrupted and oxidative stress is induced. Selenium, a known anti-oxidant, has been shown to induce increased activity of glutathione peroxidase. Glutathione peroxidase removes peroxides from cells protecting them from oxidative stress. This study set out to determine if selenium could play a role in preventing the damage to mice livers due to microcystin toxin. The protective role of selenium was explored in three main studies: in the first study, the ability of selenium to increase the survival time of mice exposed to a lethal dose of toxin was determined. In the second study the mice were exposed to sublethal chronic doses of toxin over 30 days. The ability of selenium to minimise liver damage under these conditions was determined. The final study investigated the mechanism of the protective effect of selenium. The results of the first study suggested that selenium could extend survival time. In the second study the selenium supplemented mice showed a reduction in the extent of the increase in liver weight and a decrease in the amount of lipid peroxidation induced compared to the mice that received only toxin. The histology of the selenium supplemented mice also showed a decrease in the severity and amount of morphological changes in the liver. The third study indicated that the protection shown by selenium might be mediated by an increase in the glutathione peroxidase (GPX) activity in selenium supplemented mice. This increase in GPX activity would increase the removal of the lipid hydroperoxides and prevent the damage they would cause in the cell. A further result indicated an increase in glutathione S-transferase in only the toxin control mice when compared to the selenium supplemented and control mice. ii In conclusion selenium offers protection against microcystin but further studies need to be done to provide statistically valid results to clarify the level of protection.
- Full Text:
- Date Issued: 2002
The immobilization of Microcystis aeruginosa PCC7806 on a membrane nutrient-gradostat bioreacator for the production of the secondary metobolites
- Authors: Strong, Peter James
- Date: 2002
- Subjects: Microcystis aeruginosa , Myrocystins , Bioreactors
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11083 , http://hdl.handle.net/10948/283 , Microcystis aeruginosa , Myrocystins , Bioreactors
- Description: A module and an inoculation technique were developed that would allow for the efficient immobilization of Microcystis aeruginosa PCC7806 on a synthetic membrane. A variety of module types, membranes (ceramic, tubular polyethersulfone and externally skinless polyethersulfone capillary membrane), and methods of immobilization (adsorption, pressure filtration and a developed technique that involved drying a cell slurry on a membrane) were assessed. The morphological properties that affected the immobilization of Microcystis aeruginosa PCC7806, as well as the effects of immobilization upon cell morphology were assessed. Cells in the stationary growth phase, which had a well-developed extra-cellular polysaccharide layer and no gas vesicles, were optimal for immobilization. Microcystin production under immobilized conditions was assessed under different nitrate concentrations, light intensities, biofilm thickness and immobilization times. Additional work included assaying for Microcystin production of two airlift-grown cultures under a high light intensity and complete nutrient deprivation and the inoculation of a ceramic membrane. An immunological technique was used to elucidate where toxin production was greatest within a biofilm immobilized upon an externally skinless polyethersulfone capillary membrane. The externally skinless polyethersulfone capillary membrane was evaluated to assess homogeneity and the physical differences between membrane batches that led to the erratic, incomplete biofilm formation, as a biofilm of a constant thickness could not be immobilized. Microcystis aeruginosa PCC7806 was exposed to a variety of solvents in order to permeabilize the cyanobacteria, as that would have enabled a truly continuous extraction process for the metabolite. FDA hydrolysis had to be optimized in order to use it as an indicator of cell viability. In addition a single-step extraction of Microcystin was attempted using live bacteria. A capillary membrane module, containing the externally skinless polyethersulfone capillary membrane, inoculated using pressure filtration, was the most efficient combination to establish a biofilm. Cells that were no longer actively dividing and that lacked buoyancy displayed superior immobilization to cells that were actively dividing and buoyant. The immobilized cells did produce Microcystin but in much lower concentrations to cells grown in an airlift culture. Biofilms grown with a higher nitrate concentration, a lower biofilm thickness and a lower light intensity had a higher specific microcystin content, while biofilms with a higher nitrate concentration a lower light intensity and a longer growth period displayed the a greater toxin production per mm2 of membrane. Microcystin occurred at its highest concentration in cells just above the pore opening. The diffusion of nutrients occurred relatively quickly to the outside layers of the biofilm, with a true gradient being established laterally from these nutrient veins that were above the pores. Permeabilization of the cells proved unsuccessful, as cells that remained viable did not release the intracellular compound into the surrounding medium.
- Full Text:
- Date Issued: 2002
- Authors: Strong, Peter James
- Date: 2002
- Subjects: Microcystis aeruginosa , Myrocystins , Bioreactors
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11083 , http://hdl.handle.net/10948/283 , Microcystis aeruginosa , Myrocystins , Bioreactors
- Description: A module and an inoculation technique were developed that would allow for the efficient immobilization of Microcystis aeruginosa PCC7806 on a synthetic membrane. A variety of module types, membranes (ceramic, tubular polyethersulfone and externally skinless polyethersulfone capillary membrane), and methods of immobilization (adsorption, pressure filtration and a developed technique that involved drying a cell slurry on a membrane) were assessed. The morphological properties that affected the immobilization of Microcystis aeruginosa PCC7806, as well as the effects of immobilization upon cell morphology were assessed. Cells in the stationary growth phase, which had a well-developed extra-cellular polysaccharide layer and no gas vesicles, were optimal for immobilization. Microcystin production under immobilized conditions was assessed under different nitrate concentrations, light intensities, biofilm thickness and immobilization times. Additional work included assaying for Microcystin production of two airlift-grown cultures under a high light intensity and complete nutrient deprivation and the inoculation of a ceramic membrane. An immunological technique was used to elucidate where toxin production was greatest within a biofilm immobilized upon an externally skinless polyethersulfone capillary membrane. The externally skinless polyethersulfone capillary membrane was evaluated to assess homogeneity and the physical differences between membrane batches that led to the erratic, incomplete biofilm formation, as a biofilm of a constant thickness could not be immobilized. Microcystis aeruginosa PCC7806 was exposed to a variety of solvents in order to permeabilize the cyanobacteria, as that would have enabled a truly continuous extraction process for the metabolite. FDA hydrolysis had to be optimized in order to use it as an indicator of cell viability. In addition a single-step extraction of Microcystin was attempted using live bacteria. A capillary membrane module, containing the externally skinless polyethersulfone capillary membrane, inoculated using pressure filtration, was the most efficient combination to establish a biofilm. Cells that were no longer actively dividing and that lacked buoyancy displayed superior immobilization to cells that were actively dividing and buoyant. The immobilized cells did produce Microcystin but in much lower concentrations to cells grown in an airlift culture. Biofilms grown with a higher nitrate concentration, a lower biofilm thickness and a lower light intensity had a higher specific microcystin content, while biofilms with a higher nitrate concentration a lower light intensity and a longer growth period displayed the a greater toxin production per mm2 of membrane. Microcystin occurred at its highest concentration in cells just above the pore opening. The diffusion of nutrients occurred relatively quickly to the outside layers of the biofilm, with a true gradient being established laterally from these nutrient veins that were above the pores. Permeabilization of the cells proved unsuccessful, as cells that remained viable did not release the intracellular compound into the surrounding medium.
- Full Text:
- Date Issued: 2002
Studies on the kallikrein-kininogen system of the ostrich (Struthio camelus)
- Authors: Bothma, Leonard Frederick
- Date: 2001
- Subjects: Kallikrein , Kinins , Ostriches
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11067 , http://hdl.handle.net/10948/275 , Kallikrein , Kinins , Ostriches
- Description: Ostrich organs/tissue/fluids were screened for plasma kallikrein-like, tissue kallikrein-like and tonin-like activity in a continuous-fluorogenic-assay system using Pro-Phe-Arg-7-amino-4-methylcoumarine, Phe- Arg-7-amino-4-methylcoumarine and Val-Leu-Arg--7-amino-4-trifluoro-methylcoumarine as substrates. Ostrich liver and kidney showed the highest specific plasma kallikrein-like activity. Ostrich adrenal glands and kidney showed the highest specific tissue kallikrein-like and tonin-like activity. Ostrich high molecular weight kininogen was purified from plasma and low molecular weight kininogen was partially purified. The N-terminal amino acid sequences of both high- and low molecular weight kininogens from ostrich plasma were determined. Ostrich plasma high molecular weight kininogen was purified as a 118 kD protein. The purified high molecular weight kininogen inhibits the cysteine proteinase papain at a ratio of one molecule HKG to two molecules of papain. Ornitho kinin-like molecules were detected in ostrich urine using reverse phase HPLC.
- Full Text:
- Date Issued: 2001
- Authors: Bothma, Leonard Frederick
- Date: 2001
- Subjects: Kallikrein , Kinins , Ostriches
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11067 , http://hdl.handle.net/10948/275 , Kallikrein , Kinins , Ostriches
- Description: Ostrich organs/tissue/fluids were screened for plasma kallikrein-like, tissue kallikrein-like and tonin-like activity in a continuous-fluorogenic-assay system using Pro-Phe-Arg-7-amino-4-methylcoumarine, Phe- Arg-7-amino-4-methylcoumarine and Val-Leu-Arg--7-amino-4-trifluoro-methylcoumarine as substrates. Ostrich liver and kidney showed the highest specific plasma kallikrein-like activity. Ostrich adrenal glands and kidney showed the highest specific tissue kallikrein-like and tonin-like activity. Ostrich high molecular weight kininogen was purified from plasma and low molecular weight kininogen was partially purified. The N-terminal amino acid sequences of both high- and low molecular weight kininogens from ostrich plasma were determined. Ostrich plasma high molecular weight kininogen was purified as a 118 kD protein. The purified high molecular weight kininogen inhibits the cysteine proteinase papain at a ratio of one molecule HKG to two molecules of papain. Ornitho kinin-like molecules were detected in ostrich urine using reverse phase HPLC.
- Full Text:
- Date Issued: 2001
On using AMOS, EQS, LISREL, Mx, RAMONA and SEPATH for structural equation modeling
- Authors: Peprah, Syvester
- Date: 2000
- Subjects: Latent variables , Multivariate analysis , Mathematical statistics -- computer programs
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11082 , http://hdl.handle.net/10948/279 , Latent variables , Multivariate analysis , Mathematical statistics -- computer programs
- Description: Structural Equation Modeling is a common name for the statistical analysis of Structural Equation Models. Structural Equation Models are models that specify relationships between a set of variables and can be specified by means of path diagrams. A number of Structural Equation Modeling programs have been developed. These include, amongst others, AMOS, EQS, LISREL, Mx, RAMONA and SEPATH. A number of studies have been published on the use of some of the applications mentioned above. They include, amongst others, Brown (1986), Waller (1993) and Kano (1997). Structural Equation Models are increasingly being used in the social, economic and behavioral sciences. More and more people are therefore making use of one or more of the Structural Equation Modeling applications on the market. This study is performed with the aim of using each of the Structural Equation Modeling applications AMOS, EQS, LISREL, Mx, RAMONA and SEPATH for the first time and document the experience, joy and the difficulties encountered while using them. This treatise is different from the comparisons already published in that it is based on the use of AMOS, EQS, LISREL, Mx, RAMONA and SEPATH to fit a Structural Equation Model for peer influences on ambition, which is specified for data obtained by Duncan, Haller and Portes (1971), by myself as a first time user of each of the programs mentioned. The impressive features as well as the difficulties encountered are listed for each application. Recommendations for possible improvements to the various applications are also proposed. Finally, recommendations for future studies on the use of Structural Equation Modeling programs are made.
- Full Text:
- Date Issued: 2000
- Authors: Peprah, Syvester
- Date: 2000
- Subjects: Latent variables , Multivariate analysis , Mathematical statistics -- computer programs
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11082 , http://hdl.handle.net/10948/279 , Latent variables , Multivariate analysis , Mathematical statistics -- computer programs
- Description: Structural Equation Modeling is a common name for the statistical analysis of Structural Equation Models. Structural Equation Models are models that specify relationships between a set of variables and can be specified by means of path diagrams. A number of Structural Equation Modeling programs have been developed. These include, amongst others, AMOS, EQS, LISREL, Mx, RAMONA and SEPATH. A number of studies have been published on the use of some of the applications mentioned above. They include, amongst others, Brown (1986), Waller (1993) and Kano (1997). Structural Equation Models are increasingly being used in the social, economic and behavioral sciences. More and more people are therefore making use of one or more of the Structural Equation Modeling applications on the market. This study is performed with the aim of using each of the Structural Equation Modeling applications AMOS, EQS, LISREL, Mx, RAMONA and SEPATH for the first time and document the experience, joy and the difficulties encountered while using them. This treatise is different from the comparisons already published in that it is based on the use of AMOS, EQS, LISREL, Mx, RAMONA and SEPATH to fit a Structural Equation Model for peer influences on ambition, which is specified for data obtained by Duncan, Haller and Portes (1971), by myself as a first time user of each of the programs mentioned. The impressive features as well as the difficulties encountered are listed for each application. Recommendations for possible improvements to the various applications are also proposed. Finally, recommendations for future studies on the use of Structural Equation Modeling programs are made.
- Full Text:
- Date Issued: 2000
Ostrich calpastatin purification and partial characterization of the liver inhibitor
- Authors: Roman, Henry James
- Date: 2000
- Subjects: Calpastatin , Protease inhibitors , Ion exchange chromatography , Ostriches
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11090 , http://hdl.handle.net/10948/d1015522
- Description: The isolation and purification of calpastatin from ostrich liver is presented, along with its physicochemical and kinetic properties. By using extraction from liver, ion-exchange chromatography on DEAE-Toyopearl, heating to 90 °C for 10 min and rechromatography on Toyopearl Super-Q 650 S, ostrich calpastatin was isolated and purified from ostrich liver. The purified intact calpastatin showed homogeneity on SDS-PAGE (Mr of 105.6 K). Amino acid analysis showed that ostrich calpastatin resembled that of rabbit liver and human erythrocyte calpastatin. An N-terminal sequence could not be obtained because the N-terminus was found to be blocked by an as yet unknown amino acid residue. The Mr values of degradative forms of ostrich liver calpastatin were determined to be 56 K and 90 K. By using PAG-IEF the pI of the intact form was determined to be 5.1. Ostrich liver calpastatin behaved characteristically like other calpastatins during kinetic analysis. Calpastatin inhibited calpain from pH 6 to 9 and was found to be unaffected by temperatures as high as 100 °C. Calpastatin also inhibited calpain activity at Ca2+ concentrations ranging from 1 to 10 mM. The inhibitor was shown to be phosphorylated because after incubation with alkaline phosphatase there was a decrease in inhibitory activity. No inhibitory effects were detected against other proteases such as chymotrypsin and trypsin, with both proteases inactivating calpastatin completely. Ostrich liver calpain was shown to have a pH optimum of 7.5 and a temperature optimum of 30 °C. In terms of its thermodynamic properties it resembled that of other ostrich proteases; DH, DS and DG being 47.07 kJ/mol, -91.1 J/mol/K and 74.237 kJ/mol, respectively. Ostrich liver calpain showed a Km of 0.14 % (w/v). The enzyme was active at both milli- and micro-molar concentrations of Ca2+. Ostrich liver calpastatin showed many physical, chemical and kinetic properties similar to those of other known calpastatins.
- Full Text: false
- Date Issued: 2000
- Authors: Roman, Henry James
- Date: 2000
- Subjects: Calpastatin , Protease inhibitors , Ion exchange chromatography , Ostriches
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11090 , http://hdl.handle.net/10948/d1015522
- Description: The isolation and purification of calpastatin from ostrich liver is presented, along with its physicochemical and kinetic properties. By using extraction from liver, ion-exchange chromatography on DEAE-Toyopearl, heating to 90 °C for 10 min and rechromatography on Toyopearl Super-Q 650 S, ostrich calpastatin was isolated and purified from ostrich liver. The purified intact calpastatin showed homogeneity on SDS-PAGE (Mr of 105.6 K). Amino acid analysis showed that ostrich calpastatin resembled that of rabbit liver and human erythrocyte calpastatin. An N-terminal sequence could not be obtained because the N-terminus was found to be blocked by an as yet unknown amino acid residue. The Mr values of degradative forms of ostrich liver calpastatin were determined to be 56 K and 90 K. By using PAG-IEF the pI of the intact form was determined to be 5.1. Ostrich liver calpastatin behaved characteristically like other calpastatins during kinetic analysis. Calpastatin inhibited calpain from pH 6 to 9 and was found to be unaffected by temperatures as high as 100 °C. Calpastatin also inhibited calpain activity at Ca2+ concentrations ranging from 1 to 10 mM. The inhibitor was shown to be phosphorylated because after incubation with alkaline phosphatase there was a decrease in inhibitory activity. No inhibitory effects were detected against other proteases such as chymotrypsin and trypsin, with both proteases inactivating calpastatin completely. Ostrich liver calpain was shown to have a pH optimum of 7.5 and a temperature optimum of 30 °C. In terms of its thermodynamic properties it resembled that of other ostrich proteases; DH, DS and DG being 47.07 kJ/mol, -91.1 J/mol/K and 74.237 kJ/mol, respectively. Ostrich liver calpain showed a Km of 0.14 % (w/v). The enzyme was active at both milli- and micro-molar concentrations of Ca2+. Ostrich liver calpastatin showed many physical, chemical and kinetic properties similar to those of other known calpastatins.
- Full Text: false
- Date Issued: 2000
Possible crosstalk between signal transduction pathways in the induction of differentiation in HT-29 cells
- Authors: Jamie, Hajierah
- Date: 2000
- Subjects: Cellular signal transduction , Cell differentiation
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11096 , http://hdl.handle.net/10948/d1019684
- Description: The investigation into the mechanisms by which compounds such as butyrate induce differentiation in HT-29 cells, is lacking. The colonic carcinoma cell line, HT-29, undergoes differentiation induction in the presence of butyrate and acetoacetate. The Caco-2 cell line spontaneously differentiates on contact inhibition. In this study, a signal transduction pathway involving ATP, cAMP, Ca2+ and the transcriptional factor CREB was investigated following suggestions that the energy state of the cell and diffferentiation are linked. The activity of the MAP kinase cascade, including possible crosstalk that may exist between these pathways was determined. The HT-29 cells were exposed to 5 mM acetoacetate, butyrate, DMSO and propionate. The results of this differentiation induction were compared to Caco-2 and HeLa cells, which are cervical carcinoma cells. It was found that ATP levels are decreased on differentiation induction in HT-29 cells, which, in turn affected the cAMP concentrations. Theoretically, the inducers do not have any effect on PDE 4 activity, and may facilitate the interaction between cAMP and PKA. Influx of Ca2+ into the cells was inhibited to a degree by the inducers, which was possibly overcome by crosstalk between the cAMP and Ca2+ pathways. CREB activation, lineage-specific gene expression, ERK activity and c-myc expression were all dependent on both the inducers used and the cell-type. PKA played a major role in CREB activation in acetoacetate- and butyrate -induced HT-29, Caco-2 and HeLa cells, while a2+/Calmodulin-dependent kinases I/IV may have a secondary role. Alkaline phosphatase expression in HeLa cells was independent of CREB. Evidence that crosstalk between the MAP kinase cascade and the REBactivation pathways exist, was illustrated by increased CREB activation on ERK inhibition in acetoacetate- and butyrate-induced HT-29 and HeLa cells. Also, the role that ERK played in the cells differed with inducer and cell-type. The dependence of cmyc expression on c-jun and c-fos, appeared to be differentiation induction- and celltype specific. Results from this study indicate the potential use of acetoacetate and butyrate as anti-cancer compounds.
- Full Text:
- Date Issued: 2000
- Authors: Jamie, Hajierah
- Date: 2000
- Subjects: Cellular signal transduction , Cell differentiation
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11096 , http://hdl.handle.net/10948/d1019684
- Description: The investigation into the mechanisms by which compounds such as butyrate induce differentiation in HT-29 cells, is lacking. The colonic carcinoma cell line, HT-29, undergoes differentiation induction in the presence of butyrate and acetoacetate. The Caco-2 cell line spontaneously differentiates on contact inhibition. In this study, a signal transduction pathway involving ATP, cAMP, Ca2+ and the transcriptional factor CREB was investigated following suggestions that the energy state of the cell and diffferentiation are linked. The activity of the MAP kinase cascade, including possible crosstalk that may exist between these pathways was determined. The HT-29 cells were exposed to 5 mM acetoacetate, butyrate, DMSO and propionate. The results of this differentiation induction were compared to Caco-2 and HeLa cells, which are cervical carcinoma cells. It was found that ATP levels are decreased on differentiation induction in HT-29 cells, which, in turn affected the cAMP concentrations. Theoretically, the inducers do not have any effect on PDE 4 activity, and may facilitate the interaction between cAMP and PKA. Influx of Ca2+ into the cells was inhibited to a degree by the inducers, which was possibly overcome by crosstalk between the cAMP and Ca2+ pathways. CREB activation, lineage-specific gene expression, ERK activity and c-myc expression were all dependent on both the inducers used and the cell-type. PKA played a major role in CREB activation in acetoacetate- and butyrate -induced HT-29, Caco-2 and HeLa cells, while a2+/Calmodulin-dependent kinases I/IV may have a secondary role. Alkaline phosphatase expression in HeLa cells was independent of CREB. Evidence that crosstalk between the MAP kinase cascade and the REBactivation pathways exist, was illustrated by increased CREB activation on ERK inhibition in acetoacetate- and butyrate-induced HT-29 and HeLa cells. Also, the role that ERK played in the cells differed with inducer and cell-type. The dependence of cmyc expression on c-jun and c-fos, appeared to be differentiation induction- and celltype specific. Results from this study indicate the potential use of acetoacetate and butyrate as anti-cancer compounds.
- Full Text:
- Date Issued: 2000
The isolation and partial characterization of a2-antiplasmin and plasminogen from ostrich plasma
- Authors: Thomas, Adele René
- Date: 2000
- Subjects: Serpins , Ostriches , Antifibrinolytic agents , Plasminogen , Plasmin
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11080 , http://hdl.handle.net/10948/274 , http://hdl.handle.net/10948/d1005751 , Serpins , Ostriches , Antifibrinolytic agents , Plasminogen , Plasmin
- Description: This study reports the isolation, purification and partial characterisation of the ostrich serpin, a2AP, as well as its target enzyme, ostrich plasmin, in its active and inactive proenzyme, viz. plasminogen, forms. Three different procedures were undertaken to isolate and purify ostrich a2AP. The first one involved L-lysine-Sepharose chromatography, ammonium sulfate fractionation, ion-exchange chromatography on Toyopearl Super-Q 650S, and ostrich plasminogen-Sepharose affinity chromatography. The second procedure replaced the latter chromatographic step with gel filtration on Sephadex G-200 and hydroxylapatite chromatography, while the third one employed instead the theoretically more efficient LBSI-Sepharose chromatographic step. The third procedure yielded purified ostrich a2AP, but the degree of purity and yield were relatively low. Ostrich plasminogen was highly purified after L-lysine-Sepharose chromatography and ostrich plasmin was obtained by the urokinase-activation of the purified ostrich plasminogen Ostrich a2AP revealed an Mr of 77-84 K and two isoelectric forms of pI 3.85 and 6.18. Nterminal sequence analysis showed ostrich a2AP to have only 2 out of 11 residues in common with both those of human and bovine a2AP. Ostrich a2AP showed the largest inhibitory effects on ostrich plasmin, followed by comm. bovine chymotrypsin, trypsin and plasmin, in that order, and it appeared to be a much less potent plasmin inhibitor than bovine aprotinin, but a much more potent one than the synthetic inhibitors, DFP and EACA. Ostrich plasminogen showed an Mr of 92 K and multiple isoelectric forms (~7) in the pI range 6.01-9.18, with a major one of pI 6.01. It showed a total of 775 amino acid residues and its N-terminal sequence showed ~53 percent identity with those of human, rabbit, cat, and ox plasminogens. Ostrich plasmin revealed an Mr of 78 K, two isoelectric forms of pI 4.07 and 6.01, and a total of 638 amino acid residues. N-terminal sequence analysis showed that 2-4 residues are identical to the 5 of human, cat, dog, rabbit, and ox plasmins. The pH and temperature optima of ostrich plasmin were determined as 8.0 and 40 oC, respectively. The thermodynamic and kinetic parameters of ostrich plasmin were computed, and plasmin was shown to prefer Lys to Arg residues in the S1 position. In conclusion, ostrich a2AP, plasminogen and plasmin showed definite similarities to their mammalian counterparts, but there were also significant differences.
- Full Text:
- Date Issued: 2000
- Authors: Thomas, Adele René
- Date: 2000
- Subjects: Serpins , Ostriches , Antifibrinolytic agents , Plasminogen , Plasmin
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11080 , http://hdl.handle.net/10948/274 , http://hdl.handle.net/10948/d1005751 , Serpins , Ostriches , Antifibrinolytic agents , Plasminogen , Plasmin
- Description: This study reports the isolation, purification and partial characterisation of the ostrich serpin, a2AP, as well as its target enzyme, ostrich plasmin, in its active and inactive proenzyme, viz. plasminogen, forms. Three different procedures were undertaken to isolate and purify ostrich a2AP. The first one involved L-lysine-Sepharose chromatography, ammonium sulfate fractionation, ion-exchange chromatography on Toyopearl Super-Q 650S, and ostrich plasminogen-Sepharose affinity chromatography. The second procedure replaced the latter chromatographic step with gel filtration on Sephadex G-200 and hydroxylapatite chromatography, while the third one employed instead the theoretically more efficient LBSI-Sepharose chromatographic step. The third procedure yielded purified ostrich a2AP, but the degree of purity and yield were relatively low. Ostrich plasminogen was highly purified after L-lysine-Sepharose chromatography and ostrich plasmin was obtained by the urokinase-activation of the purified ostrich plasminogen Ostrich a2AP revealed an Mr of 77-84 K and two isoelectric forms of pI 3.85 and 6.18. Nterminal sequence analysis showed ostrich a2AP to have only 2 out of 11 residues in common with both those of human and bovine a2AP. Ostrich a2AP showed the largest inhibitory effects on ostrich plasmin, followed by comm. bovine chymotrypsin, trypsin and plasmin, in that order, and it appeared to be a much less potent plasmin inhibitor than bovine aprotinin, but a much more potent one than the synthetic inhibitors, DFP and EACA. Ostrich plasminogen showed an Mr of 92 K and multiple isoelectric forms (~7) in the pI range 6.01-9.18, with a major one of pI 6.01. It showed a total of 775 amino acid residues and its N-terminal sequence showed ~53 percent identity with those of human, rabbit, cat, and ox plasminogens. Ostrich plasmin revealed an Mr of 78 K, two isoelectric forms of pI 4.07 and 6.01, and a total of 638 amino acid residues. N-terminal sequence analysis showed that 2-4 residues are identical to the 5 of human, cat, dog, rabbit, and ox plasmins. The pH and temperature optima of ostrich plasmin were determined as 8.0 and 40 oC, respectively. The thermodynamic and kinetic parameters of ostrich plasmin were computed, and plasmin was shown to prefer Lys to Arg residues in the S1 position. In conclusion, ostrich a2AP, plasminogen and plasmin showed definite similarities to their mammalian counterparts, but there were also significant differences.
- Full Text:
- Date Issued: 2000
Physiological and non-physiological induction of gastrointestinal differentiation
- Authors: Brauns, Seth Clint Aron
- Date: 1999
- Subjects: Gastrointestinal system -- Differentiation , Gastrointestinal system -- Physiology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11089 , http://hdl.handle.net/10948/d1015521
- Description: The human colonic carcinoma cell lines HT-29 and Caco-2 both exhibit structural and functional differentiation under appropriate culture conditions. HT-29 can be induced to differentiate by treatment with short-chain fatty acids or acetoacetate. Caco-2 cells differentiate spontaneously upon contact inhibition. In this study HT-29 cells were treated with 5 mM acetate, propionate, butyrate and acetoacetate (physiological inducers) to assess their effects on the expression of carbonic anhydrase 1, sucrase-isomaltase and alkaline phosphatase which are reported to be markers of gastrointestinal differentiation. The maturation induction observed was compared to that of the spontaneous differentiation observed in Caco-2 cells. Assays were performed over an 18 day period. Results showed a close correlation (p < 0.05) between HT-29 and Caco-2 cell on days 4 and 12. These results indicate that differentiation reported in both cell lines is comparable and can be used as a basis for further comparative studies. In addition, parallel experiments to the above were conducted using a selection of nine rationally designed cyclic dipeptides (CDPs) potential drug entities which were chosen as non-physiological inducers. The results showed that the cyclic dipeptides were able to induce the gastrointestinal phenotype as observed in HT-29 cells treated with physiological inducers. Studies on the effects of energy-related metabolism in HT-29 and Caco-2 cells as induced by physiological and non-physiological inducers indicated that energy metabolism is a significant role player in gastrointestinal differentiation. The results reported show a decrease in ATP concentrations indicating that the cyclic dipeptides, like physiological inducers, affect the energy state of the HT-29 cells and thus may effect the differentiation of these cells. A positive correlation was found between histone phsophorylation and differentiation confirming that histone phsophorylation was partly responsible for the decrease in ATP concentrations. It is suggested that the induction of differentiation in HT- 29 cells could be either due to non-specific transcription of genes by activation of a chromatin switch or specific by the activation of signal transduction pathways based on the flux of ATP through the cells. Differential display RT-PCR is probably the most sensitive method that could be used to validate the suggestion of either a nonspecific transcription of genes or a specific differentiation reported for HT-29 cells.
- Full Text:
- Date Issued: 1999
- Authors: Brauns, Seth Clint Aron
- Date: 1999
- Subjects: Gastrointestinal system -- Differentiation , Gastrointestinal system -- Physiology
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:11089 , http://hdl.handle.net/10948/d1015521
- Description: The human colonic carcinoma cell lines HT-29 and Caco-2 both exhibit structural and functional differentiation under appropriate culture conditions. HT-29 can be induced to differentiate by treatment with short-chain fatty acids or acetoacetate. Caco-2 cells differentiate spontaneously upon contact inhibition. In this study HT-29 cells were treated with 5 mM acetate, propionate, butyrate and acetoacetate (physiological inducers) to assess their effects on the expression of carbonic anhydrase 1, sucrase-isomaltase and alkaline phosphatase which are reported to be markers of gastrointestinal differentiation. The maturation induction observed was compared to that of the spontaneous differentiation observed in Caco-2 cells. Assays were performed over an 18 day period. Results showed a close correlation (p < 0.05) between HT-29 and Caco-2 cell on days 4 and 12. These results indicate that differentiation reported in both cell lines is comparable and can be used as a basis for further comparative studies. In addition, parallel experiments to the above were conducted using a selection of nine rationally designed cyclic dipeptides (CDPs) potential drug entities which were chosen as non-physiological inducers. The results showed that the cyclic dipeptides were able to induce the gastrointestinal phenotype as observed in HT-29 cells treated with physiological inducers. Studies on the effects of energy-related metabolism in HT-29 and Caco-2 cells as induced by physiological and non-physiological inducers indicated that energy metabolism is a significant role player in gastrointestinal differentiation. The results reported show a decrease in ATP concentrations indicating that the cyclic dipeptides, like physiological inducers, affect the energy state of the HT-29 cells and thus may effect the differentiation of these cells. A positive correlation was found between histone phsophorylation and differentiation confirming that histone phsophorylation was partly responsible for the decrease in ATP concentrations. It is suggested that the induction of differentiation in HT- 29 cells could be either due to non-specific transcription of genes by activation of a chromatin switch or specific by the activation of signal transduction pathways based on the flux of ATP through the cells. Differential display RT-PCR is probably the most sensitive method that could be used to validate the suggestion of either a nonspecific transcription of genes or a specific differentiation reported for HT-29 cells.
- Full Text:
- Date Issued: 1999
A linear model for valuating preferences of freshwater inflows into forty selected estuaries along the South African coastline
- Authors: Smith, Melnick Jurgen
- Subjects: Estuaries -- South Africa -- Eastern Cape , Mathematical statistics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10581 , http://hdl.handle.net/10948/d1020916
- Description: According to the National Water Act of 1998, an estuary is an enclosed body of water that is either periodically or permanently open to the ocean. Within an estuary, the seawater is diluted to a measurable degree, creating a unique aquatic environment for animals and plants. Estuaries are environmental and economic assets to the population. The health status of our local estuaries, however, is being compromized due to a steady decrease in the freshwater inflow and supply. Tides and climatic conditions do have an impact upon the dynamics of an estuary, but these factors remain relatively constant throughout each year. The freshwater inflow and supply, however, are highly variable and are directly influenced by human involvement. Upstream abstraction for industrial and domestic use, for example, could lead to mouth closure where the ocean meets the river. The National Water Act of 1998 was established to address the lack of research and predominant mismanagement of freshwater inflow into South Africa’s estuaries (Allanson and Baird, 1999). To ensure proper water resource management, different water allocation costs and benefits need to be compared and analyzed to secure an optimum solution (Mlangeni, 2007). Like many environmental services yielded to man, estuary services are not traded in any markets. Alternative markets are thus sought to allow the estimation of the values of such services. Among the available valuation techniques are the Contingent Valuation Method (CVM), Travel Cost Method (TCM) and Hedonic Pricing Method (HPM). The involved benefits of water allocations are predicted in this study by use of the CVM which elicits respondents’ willingness to pay (WTP) towards predetermined changes in freshwater inflow into estuaries. The CVM was applied throughout the Water Research Commission’s (WRC) Project K5/1413 from 2000 to 2008 (Hosking, 2010). Each individual study employed specialized surveys which ideally created a close correspondence between the answers provided by respondents to the supposed scenarios and their voluntary exchanges in markets should money actually have been handled (Mlangeni, 2007). Much criticism has been directed towards the CVM, but careful use and application of the method has been shown to produce significant and satisfactory results (Hosking, 2010). The primary aim of this study was to collectively analyze the collated data provided by the WRC and compare the results with the findings of previous studies. Each variable was analyzed separately in order to reveal any discrepancies between the respective findings. A supplementary objective of this study was to add to the body of knowledge pertaining to South Africa’s estuaries and guide management in the distribution of freshwater towards proficient levels (Du Preez and Hosking, 2010). The associated change in the cumulative consumer surplus with an increased freshwater supply into forty selected estuaries was therefore investigated. The subsequent benefits due to a superior freshwater supply are therefore reflected (Du Preez and Hosking, 2010). The data gathered by each of the individual researchers throughout their studies (supported by the WRC) were combined to form a single dataset including all recorded information supplied by the corresponding respondents. As the investigation progressed, improvements were made upon the questionnaires posed to the considered estuary populations. Consequently, some of the data in the combined dataset were “missing”, since previous studies did not include certain questions, while later studies omitted others. Data imputation was employed to create an imputed dataset, enabling the modeling of the public’s WTP through regression techniques. A linear model was utilized in this study, also incorporating interaction between the predictor variables. The double-log functional form was implemented to estimate the public’s WTP. The population’s total willingness to pay (TWTP) was further estimated by aggregation. A summary of the respective results is displayed in in Table 1.
- Full Text:
- Authors: Smith, Melnick Jurgen
- Subjects: Estuaries -- South Africa -- Eastern Cape , Mathematical statistics
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10581 , http://hdl.handle.net/10948/d1020916
- Description: According to the National Water Act of 1998, an estuary is an enclosed body of water that is either periodically or permanently open to the ocean. Within an estuary, the seawater is diluted to a measurable degree, creating a unique aquatic environment for animals and plants. Estuaries are environmental and economic assets to the population. The health status of our local estuaries, however, is being compromized due to a steady decrease in the freshwater inflow and supply. Tides and climatic conditions do have an impact upon the dynamics of an estuary, but these factors remain relatively constant throughout each year. The freshwater inflow and supply, however, are highly variable and are directly influenced by human involvement. Upstream abstraction for industrial and domestic use, for example, could lead to mouth closure where the ocean meets the river. The National Water Act of 1998 was established to address the lack of research and predominant mismanagement of freshwater inflow into South Africa’s estuaries (Allanson and Baird, 1999). To ensure proper water resource management, different water allocation costs and benefits need to be compared and analyzed to secure an optimum solution (Mlangeni, 2007). Like many environmental services yielded to man, estuary services are not traded in any markets. Alternative markets are thus sought to allow the estimation of the values of such services. Among the available valuation techniques are the Contingent Valuation Method (CVM), Travel Cost Method (TCM) and Hedonic Pricing Method (HPM). The involved benefits of water allocations are predicted in this study by use of the CVM which elicits respondents’ willingness to pay (WTP) towards predetermined changes in freshwater inflow into estuaries. The CVM was applied throughout the Water Research Commission’s (WRC) Project K5/1413 from 2000 to 2008 (Hosking, 2010). Each individual study employed specialized surveys which ideally created a close correspondence between the answers provided by respondents to the supposed scenarios and their voluntary exchanges in markets should money actually have been handled (Mlangeni, 2007). Much criticism has been directed towards the CVM, but careful use and application of the method has been shown to produce significant and satisfactory results (Hosking, 2010). The primary aim of this study was to collectively analyze the collated data provided by the WRC and compare the results with the findings of previous studies. Each variable was analyzed separately in order to reveal any discrepancies between the respective findings. A supplementary objective of this study was to add to the body of knowledge pertaining to South Africa’s estuaries and guide management in the distribution of freshwater towards proficient levels (Du Preez and Hosking, 2010). The associated change in the cumulative consumer surplus with an increased freshwater supply into forty selected estuaries was therefore investigated. The subsequent benefits due to a superior freshwater supply are therefore reflected (Du Preez and Hosking, 2010). The data gathered by each of the individual researchers throughout their studies (supported by the WRC) were combined to form a single dataset including all recorded information supplied by the corresponding respondents. As the investigation progressed, improvements were made upon the questionnaires posed to the considered estuary populations. Consequently, some of the data in the combined dataset were “missing”, since previous studies did not include certain questions, while later studies omitted others. Data imputation was employed to create an imputed dataset, enabling the modeling of the public’s WTP through regression techniques. A linear model was utilized in this study, also incorporating interaction between the predictor variables. The double-log functional form was implemented to estimate the public’s WTP. The population’s total willingness to pay (TWTP) was further estimated by aggregation. A summary of the respective results is displayed in in Table 1.
- Full Text:
An investigation into the localization of peptide-gold nanoparticles in an in vitro and in vivo colorectal cancer model
- Authors: Cairncross, Lynn
- Subjects: Colon (Anatomy) -- Cancer -- Treatment , Cancer -- Early detection
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10351 , http://hdl.handle.net/10948/d1020921
- Description: Background: Colorectal cancer is the third most common cancer and cause of related deaths worldwide. Early colorectal cancer diagnosis is vital in reducing incidence and mortality. There is a need for the development of non-invasive screening tools for enhancing the detection of the disease. Cancer specific peptides are useful cancer targeting agents that can be used to specifically improve early detection strategies. Several cancer targeting peptides have been identified. Previous work investigated the specific binding of three of these peptides (p.C, p.L and p.14) conjugated to quantum dots and were found to bind to colorectal cancer cell lines (HT-29 and Caco-2). However, their uptake, localization and biodistribution in an in vitro and in vivo colorectal cancer model have not been determined. This is essential in gaining an understanding for future diagnostic or therapeutic based applications. Primary Aim: The aim of this study was investigate the localization of three selected peptides p.C, p.L and p.14 conjugated to gold nanoparticles in an in vitro and in vivo colorectal cancer model using HRTEM. Methodology: The AuNP/peptide conjugates were characterized by HRTEM and DLS. For in vitro studies; HT-29, Caco-2 and C3A cells were exposed to the AuNP-p.C, AuNP-p.L and AuNP-p.14, collected and processed for HRTEM to assess targeting and localization. For in vivo studies; the establishment of a colorectal cancer model using the AOM/DSS model 1 and 2 was conducted. Wistar rats were assigned to 6 groups, five experimental and 1 control group. Group 1 received AOM/DSS method 1 and was treated with AuNP-p.L. Group 2 and 3 received AOM/DSS method 2 and were treated with AuNP-p.C and AuNP-p.14. Group 4 and 5 remained healthy and treated with AuNP-p.C and AuNP-p.14. Group 6 remained healthy receiving no nanoparticle treatment. After treatment, rats were sacrificed and tissue was processed for HRTEM. Tissue chosen for HRTEM analysis included: Group 1 (inflamed colon, rectum, pancreatic and kidney), Group 4 (kidney) and Group 5 (liver). Results: results obtained from nanoparticle characterization suggested that nanoparticles were conjugated to their respective peptides and were stable in dispersion. For in vitro studies, results suggested no AuNP targeting and localization in HT-29 cell lines. For in vivo studies, no colorectal cancer tumours were induced. TEM micrographs did not indicate the presence of nanoparticles in colon, rectum, pancreatic, kidney and liver tissue. However, AuNPs were found in the kidney tissue (group 4). Conclusion: Although the overall objectives were not met, this study provided insight into TEM cell preparation and optimization for future nanoparticle cell interaction research. This study also demonstrated the absence of AuNPs in healthy tissue and the presence of AuNPs in healthy kidney tissue through renal clearance, a favourable quality for diagnostic or therapeutic applications.
- Full Text:
- Authors: Cairncross, Lynn
- Subjects: Colon (Anatomy) -- Cancer -- Treatment , Cancer -- Early detection
- Language: English
- Type: Thesis , Masters , MSc
- Identifier: vital:10351 , http://hdl.handle.net/10948/d1020921
- Description: Background: Colorectal cancer is the third most common cancer and cause of related deaths worldwide. Early colorectal cancer diagnosis is vital in reducing incidence and mortality. There is a need for the development of non-invasive screening tools for enhancing the detection of the disease. Cancer specific peptides are useful cancer targeting agents that can be used to specifically improve early detection strategies. Several cancer targeting peptides have been identified. Previous work investigated the specific binding of three of these peptides (p.C, p.L and p.14) conjugated to quantum dots and were found to bind to colorectal cancer cell lines (HT-29 and Caco-2). However, their uptake, localization and biodistribution in an in vitro and in vivo colorectal cancer model have not been determined. This is essential in gaining an understanding for future diagnostic or therapeutic based applications. Primary Aim: The aim of this study was investigate the localization of three selected peptides p.C, p.L and p.14 conjugated to gold nanoparticles in an in vitro and in vivo colorectal cancer model using HRTEM. Methodology: The AuNP/peptide conjugates were characterized by HRTEM and DLS. For in vitro studies; HT-29, Caco-2 and C3A cells were exposed to the AuNP-p.C, AuNP-p.L and AuNP-p.14, collected and processed for HRTEM to assess targeting and localization. For in vivo studies; the establishment of a colorectal cancer model using the AOM/DSS model 1 and 2 was conducted. Wistar rats were assigned to 6 groups, five experimental and 1 control group. Group 1 received AOM/DSS method 1 and was treated with AuNP-p.L. Group 2 and 3 received AOM/DSS method 2 and were treated with AuNP-p.C and AuNP-p.14. Group 4 and 5 remained healthy and treated with AuNP-p.C and AuNP-p.14. Group 6 remained healthy receiving no nanoparticle treatment. After treatment, rats were sacrificed and tissue was processed for HRTEM. Tissue chosen for HRTEM analysis included: Group 1 (inflamed colon, rectum, pancreatic and kidney), Group 4 (kidney) and Group 5 (liver). Results: results obtained from nanoparticle characterization suggested that nanoparticles were conjugated to their respective peptides and were stable in dispersion. For in vitro studies, results suggested no AuNP targeting and localization in HT-29 cell lines. For in vivo studies, no colorectal cancer tumours were induced. TEM micrographs did not indicate the presence of nanoparticles in colon, rectum, pancreatic, kidney and liver tissue. However, AuNPs were found in the kidney tissue (group 4). Conclusion: Although the overall objectives were not met, this study provided insight into TEM cell preparation and optimization for future nanoparticle cell interaction research. This study also demonstrated the absence of AuNPs in healthy tissue and the presence of AuNPs in healthy kidney tissue through renal clearance, a favourable quality for diagnostic or therapeutic applications.
- Full Text: