The human gut microbiome is a complex microbial ecosystem residing in the lumen of our gastrointestinal tract. The type and amounts of microbes present in this ecosystem varies based on numerous factors, including host genetics, diet, and environmental factors. The human gut microbiome plays an important role in normal host physiological functions, including providing energy to colonocytes in the form of short-chain fatty acids. However, gut microbial metabolites have also been associated with numerous disease states. Current tools for analyzing the gut microbiome, such as high-throughput sequencing techniques, are limited in their predictive ability. Additionally, “-omic” approaches of studying the complex array of molecules, such as transcriptomics (RNA), proteomics (proteins), and metabolomics (previously identified physiologically active molecules), give important insight as to the levels of these molecules but do not provide adequate explanations for their production in a complex environment. With a better physiological understanding of why specific metabolites are produced by the gut microbiome, more directed therapies could be developed to target their production. Therefore, it is immensely important to study the specific bacteria that reside within the gut microbiome to gain a better understanding of how their metabolic actions might impact the host. Within this framework, this study aimed to better understand the production of secondary bile acid metabolites by bacterial in the gut microbiome. High levels of secondary bile acids are associated with numerous pathophysiological disorders including colon cancer, liver cancer, and cholesterol gallstone disease. In the current study, three bile acid metabolizing strains of bacteria that are known members of the gut microbiome were studied. A novel strain of Eggerthella lenta was identified and characterized, along with the type strain, for its ability to modulate bile acid and steroid metabolism based on the atmospheric gas composition. Additionally, it was shown that the oxidation of hydroxyl groups on primary bile acids by E. lenta C592 inhibited subsequent 7α-dehydroxylation by Clostridium scindens. The gene involved in the production of a Δ4,6-reductase enzyme, responsible for catalyzing two of the final reductive steps in the 7α-dehydroxylation pathway, was putatively identified and characterized in Clostridium scindens ATCC 35704. Lastly, the transcriptomic profile of Clostridium scindens VPI 12708 in the presence of numerous bile acids and steroid molecules was studied. These studies contribute significantly to the understanding of why specific bile acid metabolites are made by members of the gut microbiome and suggest ways of modulating their production.

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 5 uppsatser.

A dissertation submitted to the Faculty of Science, University of' the Witwatersrand, in partial fulfilment of the requirements of the Degree of Master of Science in the field ·of Biotechnology. February 1991. / Nocardioform bacteria were studied for their ability to interconvert bile acids. From the studies of utilisation and resistance curves, the most suitable donor and recipient strains for complementary gene cloning, were Arthrobacter oxydans strain C1 and Rhodococcus erythropolis strain ATCC 4217-1 respectively. [Abbreviated Abstract. Open document to view full version] / MT2016

Bile acids (BAs) comprise a diverse group of cholesterol metabolites with multiple modes of action. Much of the role of BAs and their receptors in energy homeostasis has been discerned from studies on genetic and/or pharmacologic manipulations. Additionally, changes in BA metabolism and transport have been reported in settings of insulin resistance, obesity, and liver dysfunction. Thus, BA-based interventions have been proposed for treatment of metabolic diseases. However, the heterogeneity of endogenous BAs lends to different affinities for and potencies in activating the various BA receptors, and the effects of altering BA composition per se are incompletely understood. In this dissertation, we aimed to characterize the effects of altering BA composition by stimulating intestinal organoids with distinct BA pools modeled after those in humans and mice. Unexpectedly, we found that BA composition regulated expression of the manganese transporter encoded by Slc30a10 and manganese efflux from cells, suggesting a role for BAs in metal homeostasis. We also identified genes that were similarly and differentially regulated by the distinct mouse and human BA pools. Overall, our studies reveal a pathway by which BAs could modulate micronutrient metabolism, which might also mediate known effects of BAs on macronutrient metabolism.

Alcoholic liver disease (ALD) is one of the most common liver diseases worldwide characterized by the accumulation of lipids within the liver, inflammation and the possibility of progressing to cirrhosis and liver failure. More importantly, there are currently no effective treatments for ALD and liver transplantation remains the only therapeutic option for end-stage liver disease. Previous studies have shown that ALD is a result of a combination of endoplasmic reticulum (ER) stress, lipid metabolism dysregulation and inflammation. It has been previously reported that alcohol disrupts gut microbiota homeostasis and causes increased endotoxins that contribute to the pathology of ALD. However, the detailed mechanism(s) underlying ALD and disease progression is poorly understood. We have discovered that sphingosine kinase 2 (SphK2) deficient (SphK2-/-) mice on an alcohol diet exhibit increased steatosis and inflammation compared to wild type mice. Sphingosine 1-phosphate receptor 2 (S1PR2) and SphK2 have been previously shown to play a key role in nutrient metabolism and signaling. However, their roles in alcohol-induced liver injury have not been characterized. The overall objective of this study is to determine the molecular mechanism(s) by which disruption of S1PR2-mediated SphK2 signaling contributes to ALD. The effects of alcohol on mouse primary hepatocytes and cultured RAW264.7 macrophages were examined. The acute on chronic alcohol mouse model from NIAAA that recapitulates the drinking pattern of human ALD patients was used to study the effects of SphK2 deficiency in ALD. In addition, 60-day chronic alcohol mouse model was used to determine whether a more severe form of ALD was present in SphK2-/- mice. The results indicated that SphK2-/- mice on an alcohol diet exhibited an increased amount of hepatic steatosis compared to wild type mice. Genes regulating lipid metabolism were also dysregulated in SphK2-/- mice. SphK2-/- mice also had increased inflammation and liver injury as shown by an upregulation of inflammatory markers and increased levels of liver enzymes. Moreover, SphK2 protein expression levels were downregulated in the human livers of alcoholic cirrhotic and hepatocellular carcinoma (HCC) patients. These findings contribute to a greater understanding of the pathophysiology of ALD and could provide information on the development of novel therapeutics against ALD.

No description available.

Colon cancer is the second most common type of cancer in the United States. Its incidence is linked epidemiologically to high levels of bile acids in the feces. Bile acids have been implicated as promoters and cocarcinogens in the etiology of colon cancer and as comutagens and mutagens in bacteria. These observations suggest the hypothesis that bile acids may damage DNA. By using the DNA-damage inducible SOS system in Escherichia coli, this study shows that when bacteria are exposed to bile acids there is induction of the SOS repair system and preferential survival of cells undergoing repair. Additionally, differential killing assays using repair defective bacteria show strains defective in recombinational repair or excision repair have lower survival when treated with bile acids than their parental wild-type counterparts. Human fibroblasts were treated with bile acids and unscheduled DNA synthesis (UDS) was measured. UDS is considered to represent the DNA synthesis step in excision repair. UDS, measured by autoradiography, was found to significantly increase in human fibroblasts upon treatment with bile acids. In addition, differential cytotoxicity assays with Chinese Hamster Ovary cells showed that different DNA-repair pathway defective cells were sensitive to different bile acids. Introduction of DNA damage and induction of DNA-repair by bile acids implicates them as possible direct carcinogens in the etiology of colon cancer.

Research has identified a linear relationship between saturated fat intake and colon cancer, and has demonstrated that high fat diets enhance tumorigenesis through elevation of secondary bile acids such as deoxycholic acid (DCA). We and others have shown that DCA can manipulate cell adhesion by decreasing expression of E-cadherin and increasing expression of beta-catenin. We have also shown that DCA significantly reduces EphB2 expression, which regulates cell positioning and segregation. Importantly, vitamin D can reinstate membranous E-cadherin/beta-catenin interactions and increase E-cadherin expression. In the present study, we sought to analyze the effects of DCA and vitamin D (cholecalciferol) treatment on EphB2 in colorectal cancer cells. Pre-treatment with cholecalciferol restored EphB2 expression in a dose-dependent manner, even with combined DCA treatment. This observation may be EGFR-dependent, suggesting that cholecalciferol may antagonize the effects of DCA. Taken together, these results suggest that cholecalciferol may represent an adjuvant therapy for colorectal cancer patients.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2007. / Title from first page of PDF file (viewed Feb. 7, 2008). Includes bibliographical references (p. 114-133).

Thesis (Ph.D.)--Kent State University, 2006. / Title from PDF t.p. (viewed May 25, 2007). Advisor: John Chiang. Keywords: biology, molecular. Includes bibliographical references (p. 208-228).