No abstract

The activity of the type 2 isozyme of steroid 5 alpha-reductase is crucial for normal development of the external genitalia and prostate in human males. We used immunohistochemistry to localize type 2 isozyme expression in the human male fetal reproductive tract and adult prostate. In fetal tissue, the stroma of the seminal vesicles, corpus cavernosum, corpus spongiosum, dorsal vein complex, scrotal skin, and prostate expressed the enzyme. In addition, the epithelial cells of the fetal urethra and proximal prostatic ducts stained positively. The type 2 isozyme could not be detected in epithelial cells of the fetal prostatic acini, seminal vesicles, prostatic utricle, ejaculatory ducts, epididymides, and Cowper's glands. Adult prostate specimens were derived from transurethral prostatectomies performed for benign prostatic hyperplasia. Enzyme expression in these benign prostatic hyperplasia samples localized to the stroma and epithelial cells of the urethra and proximal ducts. No staining was detected in the acinar (luminal and basal) epithelial cells. Double staining with an antismooth muscle actin antibody localized type 2 isozyme expression to the stromal fibroblast cells of the prostate. Double staining with an androgen receptor antibody localized AR expression to the acinar epithelial cells and stromal fibroblasts. These data indicate that 5 alpha-reductase type 2 is expressed throughout the developing male genitourinary tract and functions as both an autocrine and a paracrine mediator of growth and differentiation.

Chronic pancreatitis (CP) is associated with impaired glucose tolerance and with reduced hepatic sensitivity to insulin. We have previously shown that in normal and sham-operated rats, insulin suppresses hepatic glucose production, and this suppression is associated with a decrease in the hepatocyte plasma membrane-bound quantity of the facilitative glucose transport protein GLUT2. The insulin-mediated reduction in membrane-bound GLUT2 is impaired in CP, and may play a role in the glucose intolerance associated with CP. To determine whether GLUT2 is actively internalized and whether this mechanism is disordered in CP, livers from fed and fasting rats in whom CP had been induced 2-3 months earlier by pancreatic duct oleic acid infusion, and in sham-operated (sham) rats, were fractionated to yield endosome (E)- and plasma membrane (PM)-enriched fractions. Forty-five minutes after duodenal intubation alone (fasting) or intubation plus duodenal feeding, livers were removed, homogenized and ultracentrifuged, and microsomal pellets were separated by sucrose density gradient ultracentrifugation. GLUT2 content of fractions was determined by Western blotting and scanning densitometry. The E:PM ratio of GLUT2 increased from 0.68 +/- 0.11 (mean +/- SEM) in fasting sham livers (n = 8) to 1.04 +/- 0.09 in fed sham livers (n = 8; p < 0.05). However, there was no change in the E:PM ratio of GLUT2 in CP livers after duodenal feeding (0.90 +/- 0.12 vs. 0.86 +/- 0.10; n = 8,8; p = NS). To test our findings using confocal laser scanning microscopy, liver specimens from fed and fasting CP and sham rats were minced, fixed in 4% paraformaldehyde, sectioned, and stained with rabbit antirat GLUT2 antibody followed by rhodamine-labeled secondary antibody. GLUT2 was quantified by mean pixel intensity in an 8 x 16-pixel area of PM and a 16 x 16-pixel area of cytosol (CYT) in each of 30 random cells/field (400x) in each of three rats per group. As in the fractionation study, duodenal feeding increased the CYT:PM ratio of GLUT2 from 0.75 +/- 0.01 in fasting sham liver to 0.86 +/- 0.01 in fed sham liver (p < 0.0001), while the CYT:PM ratio in CP remained unchanged. We conclude that feeding induces a shift in GLUT2 from the plasma membrane to the endosomal pool. The feeding-induced internalization of GLUT2 is absent in livers from rats with CP and may play a role in the glucose intolerance associated with CP.

No abstract