Figure 8.

Effect of diet on renal enzymes involved in the synthesis and degradation of 1,25-(OH)₂D₃ in lactating and virgin rats. Kidney homogenates were obtained from dams (D) and virgin rats fed a glucose (G), fructose (F), or starch (S) diet for 6 wk. A) mRNA and protein levels of 1α-hydroxylase CYP27B1. mRNA levels were normalized to EF1α mRNA levels, while β-actin was used as loading and transfer control in Western blots. B) mRNA and protein levels of 24-hydroxylase CYP24A1. mRNA expression data were normalized relative to levels in virgins fed S. C) Immunolocalization of CYP27B1 in the kidney of dams. Yellow solid arrows indicate 1α-hydroxylase occupying the basolateral cytosolic pole of cells lining the proximal tubules (p) in panels DG and DS. White arrows indicate more modest quantities of 1α-hydroxylase from distal tubules (d), while blue arrows indicate its virtual absence from glomeruli (g) of panels DG and DS. In dams fed fructose (panel DF), distal tubular and glomerular cells also have low abundance of CYP27B1, and there is reduced expression (dotted yellow arrows) of CYP27B1 in the proximal tubule. CYP27B1 immunolocalization for virgin rats is not shown since no effect of dietary fructose was observed (see panel A). D) Relative 1α-hydroxylase activity expressed as ratio of 1,25-(OH)₂D₃ /25-(OH)D₃. Bars, analyses, and normalization as in Fig. 1.