Mitogenic response of canine fundic epithelial cells in short-term culture to transforming growth factor alpha and insulinlike growth factor I.
Mitogenic response of canine fundic epithelial cells in short-term culture to transforming growth factor alpha and insulinlike growth factor I.
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DOI：
10.1172/jci115189
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发表时间：
1991-05
期刊：
影响因子：
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通讯作者：
Monica C. Chen;Alexander T. Lee;A. H. Soll
中科院分区：
文献类型：
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作者：
Monica C. Chen;Alexander T. Lee;A. H. Soll
我们报告的方法，允许快速分裂的胃上皮细胞培养，以调查粘膜细胞复制的调节。将犬胃底粘膜细胞经酶处理分散，过滤和淘洗富集，并在DMEM/F12培养基中的胶原凝胶上培养。48小时后，超过95%的细胞显示免疫反应性与抗体的细胞角蛋白，上皮标记。细胞在72 h时形成融合单层，当安装在Ussing室中时，跨膜电阻为1，600 ohm.cm2，表明保留了上皮细胞特征。小牛血清（0.1-2%）产生了剂量依赖性促有丝分裂效应，这一点通过[3 H]-胸苷掺入酸沉淀物质和细胞数量的增加而得到证实。用[3 H]-胸苷孵育18-24小时后，在2%血清中培养的约55%的细胞通过放射自显影显示DNA合成的证据，并且所有复制细胞均为细胞角蛋白阳性。使用可比的培养条件，与溴脱氧尿苷孵育18-24小时的细胞比例相似，显示出核抗溴脱氧尿苷免疫反应性，从而表明这些培养物中超过一半的细胞在此期间合成DNA。与血清一样，表皮生长因子和转化生长因子α（TGF α）（10 pM至1 nM）、胰岛素（10 nM至1 microM）和胰岛素样生长因子-I（IGF-I，1-100 nM）增加[3 H]-胸苷摄取。与胰岛素相比，IGF-I的效力更大，表明存在IGF-I受体。我们的结论是，这种文化准备是由胃底粘膜上皮细胞，并含有一个分裂上皮细胞的优势。EGF/TGF α和IGF-I是直接调节胃底粘膜细胞增殖的潜在因子。
We report methods allowing the culture of rapidly dividing gastric epithelial cells to investigate the regulation of mucosal cell replication. Cells from canine fundic mucosa were dispersed by enzyme treatment, enriched by filtration and elutriation, and cultured on collagen gel in DMEM/F12 medium. After 48 h, greater than 95% of the cells displayed immunoreactivity with antibody to cytokeratin, an epithelial marker. The cells formed confluent monolayers by 72 h with a transmembrane resistance of 1,600 ohm.cm2 when mounted in a Ussing chamber indicating retention of epithelial cell characteristics. Calf serum (0.1-2%) produced a dose-dependent mitogenic effect evident by increases in [3H]-thymidine incorporation into acid-precipitated material and in cell number. After an 18-24-h incubation with [3H]-thymidine, approximately 55% of the cells cultured in 2% serum showed evidence of DNA synthesis by autoradiography and all of the replicating cells were cytokeratin positive. Using comparable culture conditions, a similar proportion of cells incubated for 18-24 h with bromodeoxyuridine displayed nuclear anti-bromodeoxyuridine immunoreactivity, thus indicating that over half of the cells in these cultures synthesized DNA during this period. As with serum, epidermal growth factor and transforming growth factor alpha (TGF alpha) (10 pM to 1 nM), insulin (10 nM to 1 microM) and insulinlike growth factor-I (IGF-I, 1-100 nM) increased [3H]-thymidine uptake. The greater potency of IGF-I, compared to insulin, suggests the presence of IGF-I receptors. We conclude that this culture preparation is composed of fundic mucosal epithelial cells and contains a predominance of dividing epithelial cells. EGF/TGF alpha and IGF-I are potential factors directly regulating proliferation of fundic mucosal cells.