Proliferation and differentiation of Gastric Mucosal Cells in Culture

胃黏膜细胞在培养过程中的增殖和分化。分离胃粘膜细胞，按细胞大小用离心式淋洗器进行分级。用流式细胞仪对这些分离的细胞进行细胞周期分析，发现15-35%的细胞处于S期，因此胃干细胞广泛分布于不同的分离部位。本研究检测了多种生长因子对胃粘膜细胞的影响，包括TGFa、TGFb、ET-1。用人胃活检标本进行了TGFa和表皮生长因子受体的免疫组织化学定位。TGFa和EGFR共同定位于胃粘膜上皮细胞、壁细胞和胃干细胞(增殖细胞核抗原阳性细胞)。在消化性溃疡周围的再生胃粘膜中，TGFa免疫反应阳性细胞与增殖细胞核抗原阳性细胞共同定位于再生腺的底部。因此，TGFa可能以自分泌或旁分泌的方式支持胃粘膜细胞的增殖。采用RT-PCR方法检测人胃活检组织中TGFa mRNA的表达。研究表明，随着胃溃疡的愈合，再生胃粘膜中TGFa mRNA的表达增加，c-myc基因在胃肠道多种肿瘤中均有过表达。C-myc反义寡核苷酸(18mer和27mer)在0.1~10uM的浓度范围内对MKN45胃癌细胞的生长抑制率为40~75%，而正义寡核苷酸对MKN45胃癌细胞的生长抑制作用不明显。TUNEL(缺口末端标记)法显示细胞凋亡率与肿瘤细胞生长受抑制程度有关。这些结果提示c-myc反义寡核苷酸有可能用于胃癌的治疗。

PROLIFERATION AND DEFFERENTIATION OF GASTRC MUCOSAL CELL IN CULTURERabbit gastric mucosal cells were isolated and fractionated by a centrifugal elutriator according to cell-size. Cell cycli analysis of these fractionated cells by flowcytometry demonstrated that 15-35% of cells are in S-phase and therefore gastric stem cells are widely distributed in various fractions. Effects of a variety of growth factors including TGFa, TGFb, ET-1 on these gastric mucosal cells were examined in this study.Immunohistochemical localization of TGFa and epidermal growth factor receptor was performed using biopsy samples of human stomach. Both TGFa and EGFR immunoreactivity were co-localized in surface mucous epithelias cells, parietal cells, and gastric stem cells (PCNA-positive cells). In the regenerating gastric mucosa arround peptic uleer, TGFa immunoreactivity was co-localized with PCNA-positive cells at the basal portion of the regenerative glands. Thus, TGFa may support gastric mucosal cell proliferation in a autocrine or paracrine fashion. TGFa mRNA expression was also examined in biopsy samples of human stomach using RT-PCR method. It was demonstrated that TGFa mRNA expression in regenerating gastric mucosa is increased with gastric ulcer healing.Overexpression of c-myc gene has been detected in a variety of tumors in the gastrointestinal tract. Antisense oligonneleotides to c-myc (18 mer and 27 mer) inhibited growth of MKN45 gastric cancer cells by 40-75% at concentrations of 0.1-10uM, while sense oligonucleotide did not. TUNEL (Nick-end labeling) method revealed that frequency of apoptotic cells is correlated with the degree of supression of tumor cell growth. These results suggests that antisense oligonucleotide to c-myc may be promising for the treatment of gastric cancer.