Browsing by Author "Ngounou, Eleonore"

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    Hyperglycaemia 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.
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    Immunohistomorphology of pancreatic islet microvasculature and the immunophenotypic analysis of CEPC in adult diabetic rats
    (Sociedad Chilena de Anatomia, 2017) Tchokonte-Nana, Venant; Le Roux, Danie Jacobus; Kotze, Patricia Clara; Ngounou, Eleonore
    ENGLISH ABSTRACT: Hyperglycaemia is one of the main causes for the endothelial cell (EC) damage in diabetic patients. Even though circulating endothelial progenitor cells (CEPC) could be used as a prognosis for microvascular complications, there is very little information on the islet microvasculature. We analysed by immunohistochemistry and by flow cytometric immunophenotyping, the expression of CD34 on EC and the expressions of CD31, CD34, CD45 and CD133 on CEPC in Streptozotocin (STZ)-induced diabetic rats. Peripheral blood and tissue specimens were obtained from rats of different treatment regimens: STZ treatment, control saline (NS) and sodium citrate (CB) treatments. Blood cells were exposed to flow cytometric immunophenotyping for CD133, CD31, CD34, CD45 and CD133. While tissues from the pancreas, liver and kidney were routinely processed and stained immunohistochemically for CD34. There was a tendency of an increased in CD45-/CD133+/CD31+/CD34+ cells (0.04 ± 0.11 %) in diabetic rats compared to the controls (CB: 0.03 ± 0.04 %; Saline: 0.01 ± 0.03 %). But there was no significant statistical difference between them. The expression pattern of CD34 on the EC in the organs’ vascular beds including arterioles, venules, capillaries and sinusoids was extremely heterogeneous across and within treatment regimens. The ECs in the sinusoids of the liver presented similar CD34 expression patterns across different treatment regimens, while the expression of CD34 on the ECs of sinusoidal capillaries in the pancreas vary with the treatment regimen. We conclude that the degree of endothelial cell damage is not uniform across organs’ vascular beds in the rat, contrary to mice and humans. Furthermore, the sinusoids in the pancreas and the kidney may have the same degree of endothelial damage when exposed to the same deleterious causes.