Khathi, Andile.Ngubane, Philani Smangaliso.Dimba, Nosipho Rosebud.2023-09-222023-09-2220232023https://researchspace.ukzn.ac.za/handle/10413/22304Masters Degree. University of KwaZulu-Natal, Durban.Introduction Type 2 diabetes (T2D) is the most common type of diabetes mellitus, which is reported to be associated with life-threatening co-morbidities. This condition is characterized by hyperglycaemia due to a defect of the insulin receptor, and its often preceded by prediabetes. Chronic consumption of a calorie diets is the primary cause of T2D, and this diet has been associated with altered intestinal permeability in diabetic patient. However, it remains unknown whether increased intestinal permeability complications begin in the prediabetic state. Previous studies done in our laboratory developed a high-fat high carbohydrate (HFHC) diet-induced prediabetic animal model, using male Sprague Dawley rats. This model was found to mimic the human condition of prediabetes. In this model, the animals develop prediabetes after 20 weeks of ingesting a HFHCdiet. Using this HFHC diet-induced animal model of prediabetes, this study sought to investigate the changes on gut microbiota and the association between prediabetes and markers associated with intestinal permeability. Furthermore, this study sought to investigate changes in concentration level of markers associated with a leaky gut and glucose homeostasis, following change to a low carbohydrate, high unsaturated fat (LCHUF) diet. Method and Materials 12 male Sprague Dawley rats (3 weeks old) were randomly assigned into the non-prediabetic group and diet-induced prediabetic group (n=6). Group A animals were exposed to a standard diet with normal drinking water for 20 weeks, and group B animals were exposed to a HFHC diet supplemented with 15% fructose for a period of 20 weeks. After 20 weeks, the American Diabetes Association criteria for diagnosis of prediabetes was used to diagnose prediabetes in all animals. The fecal samples were analyzed to measure the gut microbiota composition of Firmicutes, Bacteroidetes, and Proteobacteria in both animal groups. Furthermore, blood glucose, plasma insulin, serum zonulin, plasma lipopolysaccharide (LPS), soluble CD14 (sCD14), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), C-reactive protein (CRP), and intestinal fatty-acid binding protein (IFABP) concentrations were measured. The first manuscript measured all these parameters at 20 weeks. In the second manuscript, 12 male Sprague Dawley rats were used and fed a HFHC diet for 20 weeks. The prediabetic animals were subdivided into two groups. Group A animals remained on the HFHC diet (the prediabetic control group), while the other 6 animals in group B were switched to a low carbohydrate, high unsaturated fat (LCHUF) diet. Group B was then categorized as the prediabetic group that had dietary intervention (PD+DI). All animals were then maintained on their respective diets and monitored for further 12 weeks. The fecal samples were analyzed to measure the gut microbiota composition of Firmicutes, Bacteroidetes, and Proteobacteria in both animal groups. Furthermore, blood glucose, glycated haemoglobin (HbA1c), serum zonulin, plasma LPS, sCD14, TNFxvii α, IL-6, CRP, and IFABP concentrations were measured. The second manuscript measured all these parameters at 32 weeks. Results and Discussion Prolonged consumption of a HFHC diet results in the development of prediabetes. This was evidenced by a significant increase in fasting blood glucose and plasma insulin in the prediabetic animals compared to the non-prediabetic animals. The HFHC diet also showed to dysregulate the gut microbiota causing gut dysbiosis which enhances translocation of endotoxins from the gut lumen into the bloodstream that elicits an inflammatory response. In the prediabetic group (PD), there was a reduction in the Firmicutes levels and an increase in Bacteroidetes and Proteobacteria compared to the nonprediabetic group (NPD). Serum zonulin, plasma sCD14, TNF-α, IL-6, CRP, and IFABP concentrations in the PD group were increased compared to the NPD group, while plasma LPS concentrations were similar. The low-grade inflammation that is observed in the prediabetic state is suggested to further progresses onset of prediabetes. However, to reverse prediabetes and to combat a leaky gut problem, the second manuscript illustrated that switching to a LCHUF diet can effectively improve glucose homeostasis thus reverse prediabetes. This was evidenced by a significant decrease in fasting blood glucose and HbA1c concentration. These results were accompanied by a decrease of Firmicutes and an increase of Bacteroidetes and Proteobacteria suggesting that a LCHUF diet effectively improved gut microbiome composition. This caused the release of serum zonulin and its effect on disassembling the tight junctions to decrease. This was evidenced by a decrease in plasma LPS and sCD14 concentration. In addition, we also observed a decrease in plasma TNF-α, IL-6, CRP, and IFABP indicating another beneficial effect of this diet on reducing intestinal inflammation, and risks of insulin resistance. Conclusion Taken together, these results suggest that chronic consumption of the HFHC diet may be associated with the dysregulation of gut microbiota, leading to increased intestinal permeability. This could be associated with chronic subclinical inflammation that have been reported to result in the development of insulin resistance in the pre-diabetic stage. In addition, a LCHSF diet markedly improved intestinal permeability as well as the glucose regulationenPre-diabetes.High -Fat High Carbohydrates.Intestinal Permeability.Zonulin.Gut Microbiota.Thesis