Acra S A, Bulus N, Bogatcheva G, Coffey R J, Barnard J A
Department of Pediatrics, Vanderbilt University School of Medicine, Nashville, TN 37232 2576, USA.
Regul Pept. 1998 Jun 30;74(2-3):105-12. doi: 10.1016/s0167-0115(98)00029-9.
The epidermal growth factor (EGF) peptide family includes six closely-related proteins, all of which bind to the EGF receptor. In the intestinal epithelium, transforming growth factor alpha (TGFalpha) appears to be the most physiological ligand for the EGF receptor. The present studies were designed to examine the effect of TGFalpha overexpression on duodenal epithelial proliferation using a metallothioneine-inducible promoter/enhancer transgenic mouse (MT-TGFalpha). The MT-TGFalpha mouse model was further studied to determine the in vivo effect of unregulated TGFalpha production on the physiological proliferative responses to fasting and refeeding.
MT-TGFalpha mice were given 25 mM oral ZnSO4 to induce transgene expression and were studied 1 to 2 months later. Duodenal histology was analyzed morphometrically in well-oriented transverse sections. The vincristine metaphase-arrest technique was used to assess proliferation in duodenal crypts. Immunohistochemical staining and in situ hybridization were used to assess transgenic TGFalpha protein and mRNA expression, respectively.
Normally fed MT-TGFalpha mice had deeper crypts (0.12 vs. 0.08 mm), longer villi (0.66 vs. 0.54 mm), and greater luminal diameters (2.65 vs. 2.19 mm) compared to controls (P<0.05 for all three dimensions). The crypt cell mitotic index in normally fed transgenic mice was 1.5 fold greater than the index in normally fed controls (20+/-2 vs. 35+/-4 mitoses per crypt; P <0.05). Fasting and refeeding MT-TGFalpha mice resulted in no significant change in their high baseline rate of crypt proliferation, while proliferation in control mice rose from a lower baseline during fasting to a level with refeeding that approximated rates in MT-TGFalpha mice. Transgenic TGFalpha protein and mRNA were localized to the villus epithelial compartment with little or no evidence of mRNA or protein expression in the crypt epithelium.
Overproduction of TGFalpha in the mouse duodenal epithelium results in a pronounced increase in crypt epithelial cell proliferation and a resulting increase in the dimension of the crypt/villus unit. This appears to be mediated through a paracrine and/or juxtacrine effect on crypt cells by TGFalpha produced in the villus epithelium. Fasting and refeeding experiments suggest that TGFalpha may also play a role in the proliferative response to refeeding or that the full potential for proliferation is realized by TGFalpha overexpression alone. Collectively, these studies suggest that TGFalpha is a physiological autocrine and paracrine regulator of small intestinal epithelial proliferation.
表皮生长因子（EGF）肽家族包含六种密切相关的蛋白质，它们均能与EGF受体结合。在肠道上皮中，转化生长因子α（TGFα）似乎是EGF受体最具生理性的配体。本研究旨在利用金属硫蛋白诱导型启动子/增强子转基因小鼠（MT-TGFα）来检测TGFα过表达对十二指肠上皮增殖的影响。对MT-TGFα小鼠模型进行进一步研究，以确定不受调控的TGFα产生对禁食和再喂养生理增殖反应的体内影响。
给MT-TGFα小鼠口服25 mM硫酸锌以诱导转基因表达，并在1至2个月后进行研究。在取向良好的横切面上对十二指肠组织学进行形态计量分析。采用长春新碱中期阻断技术评估十二指肠隐窝中的增殖情况。分别使用免疫组织化学染色和原位杂交来评估转基因TGFα蛋白和mRNA表达。
与对照组相比，正常喂养的MT-TGFα小鼠隐窝更深（0.12对0.08 mm）、绒毛更长（0.66对0.54 mm）且管腔直径更大（2.65对2.19 mm）（所有三个维度P<0.05）。正常喂养的转基因小鼠隐窝细胞有丝分裂指数比正常喂养的对照组高1.5倍（每个隐窝有丝分裂数分别为20±2对35±4；P<0.05）。禁食和再喂养MT-TGFα小鼠后，其隐窝增殖的高基线率无显著变化，而对照组小鼠的增殖从禁食期间较低的基线上升至再喂养时接近MT-TGFα小鼠的水平。转基因TGFα蛋白和mRNA定位于绒毛上皮区室，在隐窝上皮中几乎没有或没有mRNA或蛋白表达的证据。
小鼠十二指肠上皮中TGFα的过量产生导致隐窝上皮细胞增殖显著增加，进而导致隐窝/绒毛单元尺寸增大。这似乎是通过绒毛上皮产生的TGFα对隐窝细胞的旁分泌和/或邻分泌作用介导的。禁食和再喂养实验表明，TGFα可能也在对再喂养的增殖反应中起作用，或者单独的TGFα过表达实现了全部增殖潜能。总体而言，这些研究表明TGFα是小肠上皮增殖的生理性自分泌和旁分泌调节因子。
