胃癌腹腔转移、小肠Cajal间质细胞凋亡致胃肠动力障碍机制的研究

Patients who suffered gastric cancer peritoneal metastasis (GCPM) often appear some symptoms such as abdominal pain and distension, the above phenomenon cannot be explained with simple tumor adhesion or oppression. In the preliminary experiments on a mouse model for GCPM, we found that the number of ICC (interstitial cells of Cajal) decreased and the slow wave frequency of ICC slowed down. The expression of ICC increased and peristalsis enhanced after after SCF or ouabain adminstration. ICC occurred pyknosis after co-cultured with primitive cells and gastric cancer cell, the expression of ICC and gap junction increased after SCF administration. Our study firstly put forward the damage of ICC is the pathological foundation of gastrointestinal motility disorder. But, the mechanism of ICC damage is unclear now. Our study put forward the hypothesis: Tumor infiltrating lymphocytes maybe promote apoptosis of tumor cells through Fas/FasL pathway, and induce apoptosis of ICC. The apoptosis of ICC will cause the communication between motor nerve and muscle obstacles, spontaneous rhythmic contraction in gastrointestinal smooth muscle becomes weak or occurs stagnation. To verify this hypothesis, our study will explore the pathophysiological mechanisms of ICC damaged by tumor infiltrating lymphocytes through peritoneal fluid from human gastric cancer, non cancerous tissues at the edge of colon cancer, the rat model of B6 and ICC primary cells with pathology and molecular biology techniques, whereby looking for therapeutic targets.

胃癌术后腹腔转移，CT尚未发现转移病灶前，患者常出现腹痛、胀气等肠梗阻症状，单纯用肿瘤粘连或压迫不能解释。前期胃癌腹腔转移模型结果发现，小肠Cajal间质细胞(ICC)减少、固缩，慢波频率减慢。干细胞因子/哇巴因干预，ICC表达增多，蠕动增强。ICC原代细胞与胃癌细胞共培养，ICC固缩，加干细胞因子后，ICC表达和细胞连接增多。首次提出小肠ICC损伤是胃肠蠕动障碍的病变基础。但引起ICC损伤机制不清楚。本研究提出假说：肿瘤浸润性淋巴细胞通过Fas/FasL凋亡途径促进肿瘤细胞凋亡，也诱导ICC凋亡。ICC凋亡使肠运动神经与肌肉间信息传递障碍，胃肠平滑肌自发节律性收缩减弱或停滞。为验证这一假说，本研究通过胃癌术后腹腔转移的腹腔液体、肠癌边缘非癌组织、B6鼠胃癌腹腔转移模型，ICC原代细胞，采用病理学和分子生物学技术，探讨肿瘤浸润性淋巴细胞诱导ICC凋亡致胃肠动力障碍的分子机制，寻找治疗靶点。

恶性肿瘤腹腔转移或伴腹水的形成，对腹腔晚期恶性肿瘤患者的生存时间和生存质量均造成严重影响，是致命并发症。本研究通过模拟胃癌腹腔转移模型，采用胃癌或卵巢癌晚期伴有腹水沉淀的肿瘤细胞、癌组织和体外卵巢癌细胞系，通过病理学、分子病理学及分子生物学技术，得出如下结果及结论。第一：导致胃肠蠕动功能障碍的病变基础是小肠Cajal间质细胞（ICC）减少、固缩凋亡，体外给予SCF干细胞因子/哇巴因干预，可改善小肠ICC结构及功能。第二：小肠ICC损伤、凋亡，c-kit表达减少，ICC细胞膜受损，HCN离子通道蛋白丢失，SCF/c-kit信息传递障碍是导致胃肠蠕动减慢的病理生理基础。第三：鼠模型在体实验和小肠ICC原代细胞体外实验，采用SCF、抗炎药、胃癌细胞、巨噬细胞单独或联合与ICC共培养，并收集高表达的HCN注射到模型鼠小肠壁的体内实验，寻找治疗的新思路、新靶点。第四：从临床获得大样本卵巢癌腹水的自噬/凋亡相关指标检测，如LC-3, Beclin-1, CA125, CASP -9,CASP-8, c-CASP -3和Survivin等，特别是对化疗后再腹水、或反复腹水的病例检测发现，腹水中的肿瘤细胞自噬高表达，抑制了凋亡，与耐药有关。第五：应用miRNA预测软件搜索靶基因LC3B的特异性miRNA并确定miR-204为候选miRNA。研究提出了卵巢癌或卵巢癌腹水耐药与自噬的增高，凋亡受抑制，miR-204的功能缺失致肿瘤细胞的周期发生了改变，同时提高了耐药蛋白的高表达，是导致卵巢癌或卵巢癌腹水后续治疗的困难所在。所以，miR-204可以作为潜在的分子标记物用于肿瘤的治疗。第六：通过人类基因芯片筛选未化疗与化疗后腹水细胞沉淀的基因表达谱，微阵列数据将腹水未化疗组数据与腹水化疗后组数据进行比较筛选出826个差异基因，其中165个基因上调和662个基因下调。其中，PIK3CB基因相互作用网络中处于一个关键节点。PIK3CB在化疗后卵巢癌腹水细胞沉淀中表达降低，反复化疗中升高。PIK3CB/Akt/mTOR通路与卵巢癌细胞顺铂耐药可能相关。反复化疗复发的卵巢癌腹水可激活卵巢癌细胞PIK3CB/Akt/mTOR通路。上述研究结果进一步验证了，对卵巢癌腹水最佳的治疗首选仍是化疗，靶向抑制自噬可促进凋亡，达到治疗的疗效。