Date

2021-8

Author

Ulutaş, Mehmet Sefa

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Inflammatory bowel diseases (IBDs) are chronic inflammatory disorders that cause prolonged inflammation in the gastrointestinal tract. In western countries high fat and protein consumption is shown as the main cause of inflammatory bowel diseases. However, traditional Turkish diet heavily relies on carbohydrate-based foods, the incidence of IBD in Turkey is very similar to western countries. We hypothesize that this may be due to high carbohydrate consumption. In the first part of the study, the interaction between intestinal microbiota and intestinal epithelial cells were investigated in mice fed with a high carbohydrate diet to mimic high carbohydrate consumption in Turkey. To analyze the changes in the microbiota composition and population dynamics, the stool samples of mice with fed different diets were studied by denaturing gradient gel electrophoresis (DGGE) method. To investigate the effect of different diets on inflammatory markers; pro-inflammatory and anti-inflammatory cytokine expression levels were measured by qRT-PCR in small intestine and colon tissues of mice. Moreover, in order to investigate how tight junction (TJ) proteins changed with different diets, TJ mRNA and protein levels were measured by qRT-PCR and western blotting method respectively. In the second part of the study, microbiota composition of mice fed with either ketogenic diet or fasted for 24-hour were stidied by next generation gequencing (NGS). These dietary interventions are known to increase the rate limiting step of ketogenesis, Hmgcs2 expression in intestinal stem cells. We have also studied the effect of microbial derived short chain fatty acid (SCFA), butyrate, on intestinal epithelial stem cells function and metabolism, using in vivo and in vitro methods. In order to investigate the effect of microbiota on intestinal stem cells function and Hmgcs2 expression in the proximal and distal colon, antibiotic treatments were used. After four mounts of feeding, DGGE profiles of stool samples from high carbohydrate with zero fiber (HC-0F), high carbohydrate (HC) and western (WD) groups showed significant reduction of bacterial diversity compared with standard chow diet group (S). Immunoblotting and qRT-PCR results of cytokines and tight junction showed that the inflammation is higher in HC-0F and HC diet groups than S or LC-0F diet groups. There is also a negative correlation between bacterial richness and colonic inflammation in our results. While bacterial diversity was less in HC-0F and HC mice groups than in other groups after diet feeding, it was observed that colonic inflammation is increased. Next generation sequencing of 24 hours fasting mice showed a significant change in microbiota of colon and small intestine lumen. Microbial derived SCFA, boosted the organoid formation capacity of proximal and distal colon at an optimum 0.5 mM concentration, increased the expression of Hmgcs2, Cpt1a and PPAR-γ. While in antibiotic treated mice, intestinal stem cells proriferation decreased and Hmgcs2 expression decreased.

Subject Keywords

High Carbohydrate Diet, Microbiota, Inflammation, Butyrate, Hmgcs2

URI

https://hdl.handle.net/11511/92885

Collections

Graduate School of Natural and Applied Sciences, Thesis