Vitamin D metabolites and their intestinal responses
Jesse P. Goff
Vitamin D metabolites regulate calcium absorption in the intestines through activation of the vitamin D receptor (VDR) and resulting in increased expression of calcium-associated proteins. Presently, it is thought that vitamin D metabolizes into a hormonal form, 1,25-dihydroxyvitamin D (1,25D), and the hormone then circulates via blood to the duodenum to stimulate calcium absorption. Recently, Koszewski et al. proposed that delivery of the hormone occurs via digestive passage rather than by the blood (Koszewski et al., 2012). Growing evidence has suggested, instead, that the hormonal precursor, 25-hydroxyvitamin D (25D), may have functional activity by either providing substrate for local hormonal production or by agonistically stimulating a response. In addition, the liver is recognized to produce and excrete both 25D and the glucuronide form, 25D-Gluc, in bile. The hypothesis guiding the present dissertation is that both 25D and 25D-Gluc may have hormonal effects in the intestine when delivered by way of digesta.
The following dissertation investigates the delivery and mechanism driving intestinal responses by these metabolites. Acute dosing of vitamin D metabolites, 25D and 1,25D, as well as their respective glucuronide conjugated derivatives, in vitamin D-normal mice was used to assess intestinal responses in vivo. Assays performed in cell culture were used to examine mechanisms of action. Responses to the metabolites both in vivo and in vitro were evaluated by measuring Cyp24 mRNA, an early and robust indicator of the VDR-mediated response. These assessments of Cyp24 were analyzed quantitatively by PCR and qualitatively by RNAscope, a recently developed RNA in situ hybridization technique. Results from these studies suggest that the vitamin D-mediated response is limited to intestinal epithelial cells nearest the luminal interior in both the duodenum and colon. Orally administered 25D was shown to elicit a response in the duodenum through agonistic binding and activity. Lastly, both oral and subcutaneous 25D-Gluc exhibited hormonal actions in the colon, but not the duodenum. The overall conclusion drawn from this dissertation is that 25D and its glucuronide conjugate are meaningful participants in regulating the VDR-mediated response in the intestines.