Doctoral Degrees (Anatomy and Histology)
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- ItemThe anatomy of seismic signalling: morphological adaptations of the hind limb in drumming and non-drumming African mole-rats (Bathyergidae)(Stellenbosch : Stellenbosch University, 2021., 2021-03-31) Sahd, Lauren; Kotze, Sanet H.; Bennett, Nigel C.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Anatomy and Histology.ENGLISH ABSTRACT: Hind foot drumming is one of the most common forms of seismic signalling. The family Bathyergidae (African mole-rats) is well known for exhibiting hind foot drumming during courtship and territorial behaviour. Hind foot drumming in bathyergid species arises as a result of the rapid flexion and extension of the hip and knee joints by either a single or alternating hind limbs. While the ecological context of drumming in these species has been well described, the possible morphological adaptations to enable the production of these seismic signals, have yet to be unravelled. Therefore, the primary aim of the present study was to investigate if morphologically discernible adaptations to the hind limb osteology and muscles could be determined in two drumming (Georychus capensis and Bathyergus suillus) and one non-drumming species (Cryptomys hottentotus natalensis) of African mole-rats using a variety of techniques. The gross anatomy of the hind limb was investigated by undertaking detailed dissections to determine the origin and insertion points as well as the innervation of the muscles of the hind limb. Thereafter, 32 muscles were removed from a single limb (with joint angles closest to 90°) for muscle architecture measurements which included belly length, muscle fascicle length, muscle mass and physiological cross-sectional area. The remaining soft tissue was removed from the specimens by maceration to enable the description of the osteology of the hind limb. Additionally, morpho-functional indices were used to morphometrically compare the bones between species. Twenty-one hind limb muscles were selected to undergo muscle fibre typing using myosin heavy chain slow antibody immunohistochemical staining. The amount of positive fibres was quantified to determine the total percentage of slow muscle fibres in each muscle section. Micro computed tomography (CT) scans of contrast enhanced stained specimens were used to determine accurate volumetric measurements of 26 muscles per sample in all three species as well as to assemble three dimensional reconstructions of the musculature of the limbs. Musculus gracilis anticus may play a key role in hind foot drumming as it was the only muscle that was morphologically different between the drummers and non-drummer. In the two drumming species, m. gracilis was a single muscle, whereas it was double in C. h. natalensis. Additionally, it was the only muscle to be significantly different in G. capensis and C. h. natalensis for all muscle architecture parameters analysed. Furthermore, m. gracilis anticus was the only muscle that had a significant difference in the volume as determined by the micro CT scans between the two drumming species and C. h. natalensis. However, the number of slow fibres of m. gracilis anticus was not significantly different between G. capensis and C. h. natalensis. Furthermore, the robust tibias in the drumming species, as indicated by the tibial robustness index, possibly counter the additional biomechanical load caused by the muscles (specifically m. gracilis anticus) involved with hind foot drumming. Thus, it can be concluded that there are distinct morphological adaptations to the osteology and musculature of the two hind foot drumming species.
- ItemHyperglycaemia and its implication on the Pancreatic islet microvasculature in diabetic rat models(Stellenbosch : Stellenbosch University, 2020-12) Ngounou, Eleonore; Alblas, Amanda; Baatjes, Karin J.; Greyling, Linda Magdalena; Page, Benedict; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Anatomy and Histology.SUMMARY BACKGROUND: Despite the considerable progress made in the treatment of diabetes mellitus, vascular damage remains the leading cause of patient death. The mechanisms underlying vascular abnormalities in obesity and diabetes mellitus remain to be elucidated and may be the main cause of β-cell death. In addition, the detailed description of islet microvasculature in the pancreas is lacking in the literature; therefore, a better understanding of the characteristics of the blood vessel and the factors that maintain β-cell function is needed in clinical practice. OBJECTIVE: To describe the spatial distribution and histomorphology of islet microvasculature under the effect of hyperglycaemia in two experimental diabetic models. METHODS: Eight week old male Wistar rats (n=50) were divided into two groups that received either a standard diet (RAC) (n=20) or a high-fat diet (HFD) (n=30) for two weeks. By the end of the two weeks, altered glucose uptake was confirmed in the HFD group by an oral glucose tolerance test (OGTT). A subgroup (RAC / STZ) of the RAC group (n=10) and another (HFD / STZ) of the HFD group (n=10) then received 50 and 35mg/kg of body weight (BW) of streptozotocin (STZ) to induce type I diabetes mellitus and type II diabetes mellitus, respectively. They were kept diabetic for an additional eight weeks. The body weight and blood glucose (BGL) of the animals were recorded throughout the experimental period (88 days). Blood was collected for flow cytometry and Luminex assay before half the number of animals were sacrificed for pancreatic tissue collection for histological procedure. The remaining half was used to replicate (cast) the pancreatic vasculature by perfusion with polyurethane-based casting resin (PU4ii). Haematoxylin and Eosin (H&E) stained sections were used to assess the general morphology of pancreatic tissue. Methenamine silver and immunostaining using CD34 antibody, delineated the basement membrane and endothelial cells, respectively, of islet microvasculature. A digital camera and a nano-computed tomography (nano-CT) scanner made it possible to generate digital and 3D images. Quantitative evaluation of topographic morphometric parameters of the pancreatic vascular network in the duodenal and splenic regions of the pancreas in each experimental condition was performed using the imageJ and Volume Graphics VGStudioMax 3.0®. Reconstruction of the pancreatic vascular network was attempted using the vascular tree scale laws. RESULTS: A significant increase in the mean body weight was accompanied by a slight increase in mean BGL within 2 weeks in HFD. Streptozotocin caused the development of two diabetic models with all clinical symptoms (polyuria, polyphagia, high BGL (> 28mmol/L) and a significant decrease in body mass in both diabetic groups (26.68% and 15.54% in RAC / STZ and HFD / STZ respectively). The results of the flow cytometry and the Luminex assay validated the presence of islet vascular lesions in animals, which also justified the significant necrosis of endothelial cells, a decrease (p<0.05) in the mean percentage of the stained area of CD34 pixels in islets, and thickening of the basement membrane. The scaling law was used to obtain the relationships between 1) the length and volume of the pancreatic vascular tree up to capillary level (R2=0.693±0.053), 2) the diameter of the lumen and the blood flow in each pancreatic vascular branch (R2=0.988±0.055), and 3) the diameter and length of the branches of the vessels (R2=0.838±0.0123). CONCLUSION: This investigation has established detailed morphological features of the vasculature of the pancreas in the duodenal and splenic regions in normal and diabetic rat models. There were large differences in the structure of the pancreatic vasculature between the two regions appearing to be dictated by metabolic demand. However, there are still challenges in 3D visualisation of the capillary networks of the pancreatic vascular tree, which was the main limitation of this study.
- ItemMulti-element analysis of human brain regions(Stellenbosch : Stellenbosch University, 2020-04) Cilliers, Karen; Muller, Christo J. F.; Page, Benedict; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Anatomy and Histology.ENGLISH ABSTRACT: Trace elements are vital for normal cellular function. An imbalance of these elements can result in oxidative stress and cellular damage, which can contribute to ageing, neurodegenerative diseases and brain tumours. While some neurodegenerative diseases have been thoroughly investigated with regards to trace element imbalance, human immunodeficiency virus (HIV)- associated neurocognitive disorder (HAND) has not been investigated. Chelation or supplementation of trace elements could assist with treating HAND, as has been attempted with Alzheimer’s disease and brain tumours. Additionally, there may exist differences between population groups, sex and brain regional distribution. A trace element imbalance may predispose individuals to neurodegenerative diseases. Therefore, the aim of this study was to determine if HIV, population group (geographical location), sex and anatomical brain regions have an effect on trace element concentrations in the brain. A thorough literature review was conducted, consisting of two literature review chapters and two published systematic reviews. Trace element changes due to ageing and Alzheimer’s disease, as well as brain tumours were reviewed. Furthermore, the effect of HIV on the brain, such as atrophy, diffusion changes, and brain matter hyperintensities, were reviewed to ascertain which brain regions are commonly affected. Since the HIV status of the cadavers is unknown, a published systematic review was included to ascertain the most reliable test, tissue or bodily fluid, as well as the duration that HIV remains reliably detectable after death. Prior to formalin embalming, blood was drawn and tested in triplicate with Determine HIV1/2 rapid tests and confirmed with a SD HIV Device 1/2 3.0 rapid HIV Kit. After embalming, tissue was sampled from the caudate nucleus from HIV-positive and HIV-negative individuals. To determine population group, sex and brain regional differences, samples from the caudate nucleus, putamen, globus pallidus and hippocampus were taken. Trace element concentrations were determined using inductively coupled plasma mass spectrometry. A Kruskal-Wallis test was used to determine statistically significant differences between HIV-positive and HIV negative groups. A multiple median regression model was used to determine significant differences in trace element concentrations between sex and regions. To determine the effect of HIV, 15 HIV-positive and 14 HIV-negative male cadavers were included in the study (mean age 44, range 22 to 61). To determine differences between the population, sex and brain region differences, 29 male and 13 female cadavers from a South African population within the Western Cape region were also included in the study (mean age 35, range 19 to 45). In the HIV-infected group, cadmium was marginally decreased, and nickel was marginally increased. Trace element levels were comparable to other population groups, such as India, the United Kingdom and Canada, although magnesium was considerably lower compared to the literature. While there were no significant sex differences, significant anatomical regional differences existed. Regional distribution of iron, selenium and barium in the brain were consistent with the literature, while zinc, manganese, copper, calcium, magnesium and strontium were inconsistent. In conclusion, the present study provides information for the first time on the alterations of trace element levels in the brains of HIV-infected individuals. More research should be conducted to ascertain the different ways that HIV alters the brain. A larger cohort, including individuals with and without HAND would provide valuable information on trace element content. Additionally, it should be investigated whether chelation or supplementation of trace elements could improve the symptoms of HAND. This is also the first study to report the trace element concentrations of different brain regions from a South African population within the Western Cape region. Currently no studies have been reported in other regions of Africa, and the reasons for the differences between population groups should be explored. Additionally, supplementation could possibly improve the low magnesium levels observed in the brains of individuals from a Western Cape population.
- ItemAssessment of health status in a 20th century skeletal collection from the Western Cape(2019-04) Alblas, Amanda; Friedling, Louise Jacqui; Greyling, Linda Magdalena; Stellenbosch University. Faculty of Medicine and Health Science. Dept. of Biomedical Sciences: Anatomy and Histology.ENGLISH ABSTRACT: Studies on the health status of skeletal remains give insight into the standard of living and survival pattern of historic populations. Analysis of trauma and pathological conditions in human skeletal remains are important in biological anthropology, explaining patterns of malnutrition, stress, disease and trauma in a population. However, the difficulty to overcome is the fact that the majority of skeletal pathological conditions are limited in their interpretive significance, since they are nonspecific and a range of stressors can cause the lesions. By analysing multiple conditions within a known population, pathological responses for specific insults can be outlined and in return help in interpretation of the frequencies and distributions within and between populations. The Kirsten Skeletal Collection, housed at Stellenbosch University, Division of Clinical Anatomy broadly represents individuals from mainly low socio-economic communities of different population groups in the Western Cape, dating throughout the 20th century. The aim of this study was to macroscopically and radiologically examine adult individuals (n=624, nmales=438, nfemales=186) in this collection for skeletal markings that included malnutrition (diet and metabolic deficiencies), osteoarthritic lesions, neoplasms, infective diseases and antemortem trauma lesions to be used as baseline information in further anthropological studies on the people of the Western Cape region. Statistically, the prevalence of specific diseases or trauma were correlated between the sexes, three different age-at-death and population groups as well as three different time periods throughout the 20th century using two-way frequency-tests and correspondence analyses. During the 20th century, many factors resulted in poverty, and "non white” people, namely the South African Black (SAB) and South African Coloured (SAC) population groups, was especially disadvantaged by the laws introduced by the ruling political party. The influx of people from rural areas during World War II to work in the manufacturing industry resulted in the already overcrowded, unsanitary informal settlements around the Cape Peninsula to be flooded, influencing the disease patterns in the communities. The results demonstrated that the lowest prevalence of metabolic deficiencies, iron deficiency anaemia (porotic hyperostosis), growth arrest signs (Harris’ lines), infections such as tuberculosis, osteomyelitis and non-specific periosteal reactions were observed in the South African White (SAW) population group. This confirms that higher socio economic societies, that escaped the unsanitary conditions associated with poor housing and overcrowding environments, were more succesfully buffering themselves from malnutrition and exposure to pathogens. Better dental health as well as dental fillings were also more associated with the SAW population group that had unrestrained access to dentists and health care facilities. In contrast, the ‘non-white’ population groups, that were supressed during the Apartheids regime, demonstrated a high prevalence of malnutrition, metabolic deficiencies, tuberculosis and trauma lesions. The difference between the higher and lower social categories was especially recognised during the late time period when the Apartheid laws of population group segregation among others, started to show results in the 1960 and 1970. Later, during the 1980s and 1990s, political unrest caused by the supressed majority, and the world due to the Apartheid laws, resulted in sanctions and lower economic opportunities for South Africa. A notable higher frequency of infective markings on bone was observed during the late time period in the study, an indication of the successful use of antibiotics during the last decades of the 20th century, which provided more time for lesions to manifest on bones due to the increased life-span of people. Studies on skeletal collections rely on the assumption that the remains represent a past community, population group or populations from a specific region and can be used as a valid comparative reference for reconstructing different aspects of skeletal biology of past people that lived in that population. The Kirsten Skeletal Collection represents adult age groups between 18 and 100, three population groups, both sexes, various time periods over the 20th century as well as known cause of death and last known residence. However, this skeletal collection relies on body donations or retention of adult unclaimed or family donated bodies under the statues of the Inspector of Anatomy, and therefore, resulted in a biased sample. This bias is perceived in the fact that the Kirsten Skeletal Collection have an overrepresentation of males, aged individuals and people with lower socio-economic status. Although three major South African population groups are represented, suggesting depiction in population variation, it is highly unequal, especially representing the overpresented heterogenous mixed population group (SAC) that lived in and around the northern townships of Cape Town. Despite limitations, in general this study of the Kirsten Skeletal Collection may represent many of the traits in the population at that time and may be useful in future studies on honours, masters and PhD level to refine region- and population specific reference data and play a supportive role in research and training of specialists. These data to be collected and interpreted in future studies, include estimation of demographic parameters (age, sex, ancestry origin) as well as human variation, trauma biomechanics and pathological conditions. If the existing predispositions of the collection are acknowledged and accounted for by the use of suitable methodology, the Kirsten Skeletal Collection holds much potential to become a valuable resource for future research projects in osteology and related fields.
- ItemAn in vitro study of mesenchyme–islet cell interactions in islet neogenesis: A model for tissue replacement therapy in diabetes mellitus(Stellenbosch : Stellenbosch University, 2017-12) Manda, Juziel Kampando; Tchokonte-Nana, Venant; Page, Benedict; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Division Anatomy and Histology.ENGLISH ABSTRACT : Shortages of donor islets, immune rejection, and the need for life-long immuno-suppressors remain the clinical challenges of islet transplantation in the treatment of diabetes mellitus. An alternative to these challenges is the in vivo generation of beta cells within the patient’s pancreas. The animal model of pancreatic injury has been reported to be a potential source of islet cells for tissue replacement therapy in type 1 diabetes mellitus. However, the in vitro regenerative capacity of endogenous beta cells in this model needs more investigation. This study investigated, in vitro, the effect of pancreatic duct ligation (PDL)-induced islet/duct-mesenchymal stromal cells (MSCs) interactions on islet and duct cells development and assessed the long-term transplantation outcome of islet-mesenchymal cells isografts. Islets, duct fragments, and MSCs were isolated from post PDL tissues harvested from eighty adult male Wistar rats (250 - 300g) 24- and 120 h following duct ligation. Islets or duct fragments were cultured with or without MSCs ([Islet/MSC+ or Islet/MSC-] or [PEDC/MSC+ or PEDC/MSC-]). Development of islets and duct fragments in culture were evaluated morphologically and by immunocytochemistry using antibodies against Pdx1, Ngn3, CK7 and insulin. Islets were also transplanted with or without MSCs (Islet/MSC+ or islet/MSC-) in diabetic animals (n = 40). Isografts survival and function were evaluated by monitoring blood glucose levels, and immunohistochemistry of graft tissues were studied. Results showed activation of Pdx1+ islet cells in both cultures with or without MSCs, however, expansion of Pdx1+ cells were promoted in the presence of MSCs and this was followed by activation of Ngn3 expression and expansion of Ngn3+ cells, which was maintained in islet cells up to 4 weeks. This resulted into low levels of insulin expression in islet-like aggregates formed between the third and the fourth week. Co-culturing of duct fragments with MSC similarly resulted into maintenance of endocrine precursors that expressed Ngn3, which later formed islet-like aggregates. In cultures with MSCs, duct epithelial cells developed growth areas with cells that co-expressed CK7 and Ngn3 in periductal cells. When periductal cells formed islet-like aggregates, Ngn3 co-expressed with insulin in islet-like cell clusters closer to ducts. Transplantation of early harvested (24 h PPDL) islets showed better curative capacity than late (84 h PPDL) islets. The average glucose levels were lower throughout the 5 weeks monitoring period in 24 h PPDL transplanted rats. The average time to reverse hyperglycemia in 80% of the 24 h PPDL transplant group was 32 ± 2 days (~4.5 weeks), while only 20% in the 84 h PPDL transplant group attained normoglycemia at 61 ± 2 days (~9 weeks) (p = 0.0011) post transplantation. Graft survival rate was higher in islets co-transplanted with MSC (Islet/MSC+) compared to grafts transplanted with islets alone (Islet/MSC-). Islet morphology and distribution of beta cells was normal in Islet/MSC+ similar to the endogenous islets in the pancreas. In conclusion, MSCs promote the expansion of Pdx1+ cells and maintain the expression of Ngn3 in islet cells and duct–derived neogenetic cells. MSCs prolong graft survival and improve the capacity of early harvested post PDL islets to reverse hyperglycemia; this novel observation may be applicable to clinical transplantation.