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There is strong evidence that thyroid hormones through triiodothyronine (T3) regulate Sertoli cell proliferation and differentiation in the neonatal testis. However, the mechanism(s) by which they are able to control Sertoli cell proliferation is unclear. In the present study in vivo approaches (PTU-induced neonatal hypothyroidism known to affect Sertoli cell proliferation) associated with in vitro experiments on a Sertoli cell line were developed to investigate this question. We demonstrated that the inhibitory effect of T3 on Sertoli cell growth, analyzed by evaluating DNA-incorporated [3H] thymidine, was associated with a time and dose-dependent increase in the levels of Cx43, a constitutive protein of gap junctions, known to participate in the control of cell proliferation and the most predominant Cx in the testis. These Cx43 changes were associated with increased gap junction communication measured by gap FRAP. Consistent with these results two specific inhibitors of gap junction coupling, AGA and oleamide, were able to significantly reverse the T3 inhibitory effect on Sertoli cell proliferation. The present data also revealed a nongenomic effect of T3 on Cx43 Sertoli cells that was evidenced by a rapid up-regulation of gap junction plaque number as identified in Cx43-GFP transfected cells exposed to the hormone. This process appears mediated through actin cytoskeleton since incubation of the cells with cytochalasin D totally reversed the T3 stimulatory effect on Cx43-GFP gap junction plaques. Based on these data, we propose a working hypothesis in which Cx43 could be an intermediate target for T3 inhibition of neonatal Sertoli cell proliferation.

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SUMMARYWe localized REG protein in Paneth cells and nonmature columnar cells of the human small intestinal crypts and speculated that this protein was associated with growth and/or differentiation. The aim of this study was to determine whether REG protein is present in two human colon cancer cell lines that exhibit enterocytic differentiation after confluence and to investigate changes in the level of its expression during growth and differentiation. Results were compared to those obtained on cells that remain undifferentiated. Western immunoblotting and immunofluorescence demonstrated the presence of REG protein in the three cell lines. With the antisera against human REG protein, the staining was diffusely spread throughout the cytoplasm at Day 2, and after Days 3-4 it appeared to have migrated to cell boundaries. After confluence, we observed only a punctate staining array along cell boundaries, which disappeared at Day 15. REG mRNA expression was demonstrated by RTPCR and REG mRNA hybridization until Day 13, but not after, in the three cell types. REG protein may be involved in cellular junctions. Its presence appears to be associated with the cell growth period and the protein must be downregulated when growth is achieved and differentiation is induced. (J Histochem Cytochem 47:863-870, 1999)

We have previously isolated from human pancreatic juice a secretory glycoprotein of 19 KD (P19), devoid ofknown enzymatic activity. P19 gave by proteolysis a protein of 141W (P14), at first named protein X and also called pancreatic thread protein or pancreatic stone protein. Specific rabbit immunosera prepared against P19 and P14 were applied to localize these proteins in human small intestine. By comparison, antibodies directed against some human pancreatic enzymes (amylase, lipase, chymotrypsin, trypsinogen 1, trypsinogen 2, and trypsin 1) were also tested.

Using the peroxidase-anti-peroxidase (PAP) technique with a specific rabbit anti-swine intestinal-phospholipase-A2 serum, the immunoreactivity of this phospholipase A2 was localized in rat-intestinal Paneth cells. The specific rabbit anti-swine intestinal-phospholipase-A2 serum did not stain the rat-pancreatic acinar cells which were stained by a specific rabbit anti-swine pancreatic-phospholipase-A2 serum. Specific rabbit anti-swine pancreatic-phospholipase-A2 serum did not stain rat-intestinal Paneth cells. Therefore, there is no cross-immunoreactivity between pancreatic and intestinal phospholipases.

The nutritional benefits of lactic acid bacteria in fermented dairy products have been well documented, especially in terms of weight gain and feed efficiency, but not in terms of small intestine adaptation. The effects of a diet supplemented (30% wt/wt) with milk fermented either by Lactobacillus casei DN-114 001 or yoghurt for 3 or 15 days were investigated in the small intestine of mice by morphometry, kinetic analysis and determination of brush-border enzyme activities. Results were compared with those obtained with standard or milk isocaloric diets. Cell proliferation and villous area were significantly increased in the proximal intestine of mice fed the fermented-milk-supplemented diets for 3 days and were associated with hypertrophy and hyperplasia of Paneth and goblet cells. Lactase-specific activity was increased by fermented-milk diets at days 3 and 15, whereas there was no variation in maltase-specific activity. Alkaline phosphatase-specific activity was increased after 3 days of the three tested diets in the whole intestine, and after 15 days in the proximal intestine. Aminopeptidase activity was increased in the distal part of the intestine after 3 days of the 3 diets. Our findings suggest that diets supplemented with fermented milks have a positive effect on the trophicity of the mucosa in the small intestine of mice.

Variations of the intestinal mucosa were studied in male adult Wistar rats on day 8 and day 15 after hypophysectomy. All results were compared with those obtained in pair-fed control rats. Morphological variations were observed at two intestinal levels (high part [hp] and low part [lp] of the jejunoileum) both with morphometrical analysis (villous area, microvillous area, enterocytes, goblet and Paneth cell number, mitotic index...), in light or electron microscopy and with histochemical techniques (proteins, polysaccharides, nucleic acids). Levels of the main intestinal hydrolysis were also measured by means of biochemical assays. Atrophy of the intestinal mucosa was confirmed, but atrophy produced not only a reduction of the villous area (hp = –30%p < 0.01, lp = –22%p < 0.05 on day 8, hp = –25 %p < 0.05 lp = –36% p < 0.01 on day 15) but also a significant decrease of the microvillous area (hp ≈ –39% p < 0.001, lp = –14% p < 0.05 on day 8, hp = –24% p < 0.02, lp = –30% p < 0.05 on day 15) in accordance with a significant decrease of brush border enzyme levels. The atrophy was generalized but concerned mainly the high part of the small intestine and could be attributed mostly to a significant decrease of glandular mitosis (hp = –57% p < 0.01, lp = –77% p < 0.001 on day 8, hp = –74% p < 0.01, lp = -75% p < 0.001 on day 15) without apparent modification of cell extrusion at the top of intestinal villi. Hypophysectomy affected the different cell types of the villous intestinal epithelium in different ways. Absorbing cells were the more reduced in number. Defects of maturation of goblet cells were noted. On the other hand, hypertrophy (p < 0.001) and hyperplasia (p < 0.001) of Paneth cells were observed with a significant decrease of the lytic activity. These apparent discrepancies can be attributed to important cytological variations and partial mucoid transformation of the secretory component of Paneth cells (observed both by histochemical and electron microscopic observation). Cytological variations after hypophysectomy seem to be induced early, since no significant differences could be observed between days 8 and 15 for most of the parameters tested. Direct or indirect (local relay) effect of pituitary function is discussed.