Role of CCN2/CTGF in the proliferation of Mastomys enterochromaffin-like cells and gastric carcinoid development
Role of CCN2/CTGF in the proliferation of Mastomys enterochromaffin-like cells and gastric carcinoid development
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DOI：
10.1152/ajpgi.00131.2006
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发表时间：
2007-01-01
影响因子：
4.5
通讯作者：
Mane, S. M.
中科院分区：
文献类型：
--
作者：
Kidd, M.;Modlin, I. M.;Mane, S. M.
乳鼠肠嗜铬样（ECL）细胞增殖最初是胃泌素驱动的，但一旦肿瘤形成，细胞成为胃泌素自主。我们推测CCN 2（CTGF），一种促有丝分裂生长因子，可能调节ECL细胞增殖。检查乳鼠基因芯片数据库（dCHIP）以鉴定CCN 2表达水平。通过实时PCR检测使用FACS（100 nM吖啶橙子）获得的正常和肿瘤ECL细胞制备物中的CCN 2。通过免疫组织化学在粘膜和ECL细胞制备物中鉴定CCN 2蛋白。用CCN 2或CCN 2 + EGF刺激短期培养的细胞，并测量增殖（MTT测定）。根据CCN 2（1 ng/ml）介导的增殖和ERK 1/2磷酸化评估ERK 1/2抑制剂PD-98059（0.1-100 μ M）。然后检测临床胃类癌中CCN 2转录物和蛋白。与正常粘膜相比，肿瘤样品中ccn 2转录物上调（+2.36倍，P < 0.01）。PCR证实ccn 2在FACS制备的（> 98%纯）正常ECL细胞中不表达，但在肿瘤ECL细胞级分中升高（41.3 +/-10.7倍）。Mastomys胃粘膜和FACS制剂的免疫染色证实，CCN 2蛋白存在于ECL肿瘤中，但不存在于正常ECL细胞中。CCN 2和CCN 2 + EGF均不能刺激正常ECL细胞增殖。CCN 2刺激肿瘤细胞增殖（EC 50 = 0.01 ng/ml），EGF显著增强（P < 0.01）CCN 2诱导的肿瘤细胞增殖（EC 50 = 20 pg/ml）。PD-98059抑制肿瘤细胞中CCN 2诱导的增殖（-1/2 +/-3%，P < 0.05）和ERK 1/2磷酸化（-34 +/-5%，P < 0.05）。在临床标本中，胃泌素自主性类癌中CCN 2的转录和蛋白表达均高于正常粘膜（P < 0.02）。总之，CCN 2可能是乳鼠ECL肿瘤增殖的增殖调节剂，一旦这些细胞成为自主的胃泌素调节。胃类癌组织中CCN 2的鉴定可能既可用作ECL细胞转化的指标，又可定义胃泌素自主性，胃类癌恶性程度的标准。
Mastomys enterochromaffin-like (ECL) cell proliferation is initially gastrin driven, but once neoplasia develops, cells become gastrin autonomous. We hypothesized that CCN2 (CTGF), a mitogenic growth factor, may regulate ECL cell proliferation. A Mastomys GeneChip database was examined (dCHIP) to identify CCN2 expression levels. CCN2 in normal and tumor ECL cell preparations obtained using FACS (100 nM acridine orange) was examined by real-time PCR. CCN2 protein was identified in mucosal and ECL cell preparations by immunohistochemistry. Short-term cultured cells were stimulated with either CCN2 or CCN2 + EGF, and proliferation was measured (MTT assay). The ERK1/2 inhibitor PD-98059 (0.1-100 mu M) was assessed in terms of CCN2 ( 1 ng/ml)-mediated proliferation and ERK1/2 phosphorylation. CCN2 transcript and protein was then examined in clinical gastric carcinoids. The ccn2 transcript was upregulated in tumor samples compared with the normal mucosa (+2.36-fold, P < 0.01). PCR demonstrated that ccn2 was not expressed in FACS-prepared (> 98% pure) normal ECL cells but was elevated in tumor ECL cell fractions (41.3 +/- 10.7-fold). Immunostaining of the Mastomys gastric mucosa and FACS preparations confirmed that CCN2 protein was present in ECL tumors but not in normal ECL cells. Neither CCN2 nor CCN2 + EGF stimulated normal ECL cell proliferation. CCN2 stimulated tumor proliferation (EC50 similar to 0.01 ng/ml); EGF significantly augmented (P < 0.01) CCN2-induced tumor cell proliferation (EC50 = 20 pg/ml). PD-98059 inhibited CCN2-induced proliferation (-12 +/- 3%, P < 0.05) and ERK1/2 phosphorylation (-34 +/- 5%, P < 0.05) in tumor cells. In clinical samples, both CCN2 transcript and protein were elevated in gastrin-autonomous carcinoids (P < 0.02) compared with the normal mucosa. In conclusion, CCN2 may be a proliferative regulator of Mastomys ECL neoplastic proliferation once these cells become autonomous of gastrin regulation. Identification of CCN2 in gastric carcinoid tissue may be useful both as an indicator of ECL cell transformation and may define gastrin autonomy, a criteria of gastric carcinoid malignancy.