Distribution of Vitamin D-Dependent Calcium-Binding Protein Messenger Ribonucleic Acid in Rat Placenta and Duodenum*

Abstract

The vitamin D-dependent calcium-binding protein (CaBP), cholecalcin or calbindin, is one of the best documented molecular expressions of 1,25-dihydroxyvitamin D, the hormonal metabolite of vitamin D. In this report, DNA/RNA hybridization assays have been used to examine cholecalcin (CaBP) mRNA production in the placenta and duodenum of 21-day pregnant rats. A cloned CaBP cDNA which codes for the rat intestinal 9000 mol wt cholecalcin (9KCaBP) was radiolabeled and used in hybridization assays to explore 1) the size and relative quantities of CaBP mRNA extractable from placenta and duodenum by molecular hybridization, and 2) the localization and quantification, by in situ hybridization histochemistry, of CaBP mRNA in specific cells in rat placenta and duodenum. Northern hybridization studies show that the [32P]cDNA sequence hybridizes to a single 500- to 600-nucleotide species in the placenta as in the duodenum and, therefore, demonstrate identical 9KCaBP mRNA processing in both tissues. Dot blot hybridization studies show that the concentration of 9KCaBP mRNA was greatest in the duodenum, while that of the inner (fetal) placenta was about 50% the duodenal level. Considerably less CaBP mRNA was found in the outer (maternal) placenta. The observed differences in 9KCaBP mRNA levels correlate well with the in vivo variations in 9KCaBP concentrations. In situ hybridization histochemistry using [3H]cDNA reveals that 9KCaBP mRNA visualized by silver grains was concentrated in the inner placenta over the cytoplasm of syncytial cells in the trophoblastic epithelium of the labyrinth and much less frequently in the cells of the outer placenta. In the duodenum, 9KCaBP mRNA was found only in the absorptive epithelial cells from the crypt region to the upper part of the villi. The silver grains were distributed throughout the cytoplasm of the columnar cells; they were densest in the perinuclear region and rarest in the nuclear region. The concentration was greater in the cells at the villous tips than in those of the crypts. This difference in 9KCaBP mRNA levels correlates well with the distribution of the protein itself along the villi. CaBP mRNA quantities detected by hybridization histochemistry showed greater labeling in the syncytial cells of the trophoblastic epithelium of the labyrinth than in the absorptive epithelial cells of the upper part of the villi (200% less), indicating accumulation of CaBP mRNA at a greater rate in the trophoblastic epithelium than in the absorptive epithelial cells. These results indicate that in the rat, 9KCaBP is synthesized in both the absorptive cells of the duodenum and the cells of the trophoblastic epithelium of the placenta.