木香内酯对结肠癌细胞增殖和凋亡的影响

Effect of Micheliolide on Proliferation and Apoptosis of Colon Cancer Cells

关键词:  木香内酯  结肠癌  PTEN  细胞增殖  细胞凋亡

Key words:  micheliolide  colon cancer  PTEN  proliferation  apoptosis

摘要:

目的 探讨木香内酯（micheliolide，MCL）对结肠癌细胞增殖和凋亡的作用及潜在机制。方法 通过腹腔注射致癌剂氧化偶氮甲烷（azoxymethane，AOM）和饮用致炎剂葡聚糖硫酸钠（dextran sodium sulfate，DSS）构建AOM/DSS小鼠结肠癌模型，分别给予尾静脉注射2.5 mg·kg^-1顺铂、MCL 0，10，20，50 mg·kg^-1，连续30 d记录小鼠存活率。取肿瘤组织，称肿瘤质量。免疫组化检验小鼠肿瘤组织Ki67和胱天蛋白酶3（caspase-3）蛋白表达水平，Western blotting检测小鼠肿瘤组织PTEN蛋白表达量；结肠癌细胞系SW620分别用含10 μmol·L^-1顺铂、MCL 0，2.5，5，10 μmol·L^-1培养基培养，或转染siPTEN后在MCL 0和10 μmol·L^-1培养基培养。BrdU细胞增殖试验检测细胞增殖，流式细胞术检测细胞凋亡，Western blotting检测细胞增殖抗原（Ki67、PCNA）、细胞凋亡相关蛋白（cleaved caspase-3、Bcl-2、Bax）及PTEN蛋白表达量。结果 与模型组相比，MCL增加小鼠生存率，减轻肿瘤质量，增加抑瘤率，上调PTEN蛋白表达，减少Ki67阳性细胞，增加caspase-3阳性细胞（P<0.05或P<0.01）；与对照组相比，MCL降低结肠癌细胞BrdU阳性细胞百分比，下调Ki67和PCNA的表达量，增加细胞凋亡率，下调Bcl-2的表达量，上调Bax、cleaved caspase-3和PTEN的表达（P<0.05或P<0.01）；抑制PTEN表达能逆转MCL对SW620细胞增殖和凋亡的作用（P<0.01）。结论 MCL抑制结直肠癌细胞增殖并诱导细胞凋亡，这与上调PTEN表达有关。

Abstract:

OBJECTIVE To explore the effects of micheliolide(MCL) on the proliferation and apoptosis of colon cancer cells and its potential mechanism. METHODS Azoxymethane(AOM)/dextran sodium sulfate(DSS) mouse colon cancer model was established by intraperitoneal injection of AOM and DSS. The mice were injected with cisplatin 2.5 mg·kg^-1, 0, 10, 20 and 50 mg·kg^-1 of MCL into the tail vein, respectively, and the survival rate was recorded for 30 d. Tumor tissues were taken and the tumors were weighed. Ki67 and caspase-3 protein expression levels in mouse tumor tissues were examined by immunohistochemistry, and PTEN protein expression in mouse tumor tissues was detected by Western blotting. Colon cancer cell line SW620 was cultured in medium containing cisplatin 10 μmol·L^-1, MCL 0, 2.5, 5, 10 μmol·L^-1, or transfected with siPTEN in MCL 0 and 10 μmol·L^-1, respectively. Cell proliferation was detected by BrdU cell proliferation assay, apoptosis was detected by flow cytometry, and cell proliferation antigen(Ki67, PCNA), apoptosis-related protein(cleaved caspase-3, Bcl-2, Bax) and PTEN protein expression were detected by Western blotting. RESULTS Compared with the model group, MCL increased the survival rate of mice, reduced tumor weight, increased tumor inhibition rate, up-regulated PTEN protein expression, decreased Ki67 positive cells, and increased caspase-3 positive cells(P<0.05 or P<0.01). Compared with the control group, MCL decreased the percentage of BrdU-positive cells, down-regulated the expression of Ki67 and PCNA, increased the apoptosis rate, down-regulated the expression of Bcl-2, and up-regulated the expression of Bax, cleaved caspase-3 and PTEN in colon cancer cells(P<0.05 or P<0.01). Inhibition of PTEN expression could reverse the effects of costunolide on SW620 cell proliferation and apoptosis(P<0.01). CONCLUSION MCL inhibits the proliferation and induces apoptosis of colorectal cancer cells, which is related to the up-regulation of PTEN expression.