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目的持续暴露脱氧胆酸(deoxycholic acid,DOC)可诱导肠腺瘤癌变,机制尚不清楚。本研究旨在探讨肠黏膜屏障损伤在DOC诱导肠腺瘤癌变过程中的作用。方法将20只4周龄Apcmin/+小鼠均分为对照组和DOC组,对照组常规饮水,DOC组给予含质量分数0.2%DOC饮用水,给药12周后处死,观察两组小鼠肠道腺瘤大小、数目及癌变率。采用HE染色评价肠道腺瘤病理类型,采用异硫氰酸荧光素标记的葡聚糖(fluorescein isothiocyanate-conjuagted dextran,FITC-D)检测小鼠肠道通透性,采用实时荧光定量PCR及免疫组织化学染色法评价两组小鼠肠道组织各种紧密连接蛋白、杯状细胞及潘氏细胞相关分子的表达水平。结果 DOC组Apcmin/+小鼠肠道息肉数量为57.00±3.07,较对照组21.50±4.69明显增加,t=20.03,P<0.000 1。DOC组小鼠血清中FITC-D浓度为(1.17±0.12)μg/mL,明显高于对照组(0.51±0.07)μg/mL,t=4.30,P=0.004;DOC组小鼠肠道ZO-1(0.31±0.04)和Claudin 3 mRNA表达量(0.14±0.02)明显低于对照组的0.78±0.07(t=5.53,P=0.000 3)和0.92±0.08(t=9.80,P=0.000 6),而DOC组增加肠道通透性的Claudin 7mRNA表达量为5.79±0.53,明显高于对照组的1.61±0.30,t=6.83,P=0.002 4;DOC组小鼠肠道染色阳性细胞数PAS(20.88±0.79)和MUC2(13.10±1.16)均较对照组35.44±1.68(t=7.84,P<0.000 1)及28.50±0.96(t=10.24,P<0.000 1)明显减少;且DOC组小鼠肠道MUC2mRNA表达量为0.15±0.04,低于对照组的0.91±0.05,t=11.34,P<0.000 1;DOC组胃型黏蛋白MUC5mRNA表达量为2.40±0.26,高于对照组的0.96±0.02,t=4.21,P=0.005 6;DOC组小鼠肠道潘氏细胞溶菌酶染色阳性细胞数为5.19±0.26,较对照组7.49±0.33显著下降,t=5.49,P=0.000 6。同时DOC组潘氏细胞分泌防御素(0.12±0.27)及隐凹素(0.20±0.35)基因表达量均低于对照组的0.87±0.05(t=13.75,P<0.000 1)和0.87±0.05(t=10.95,P<0.000 1)。结论 DOC可通过增加肠道黏膜通透性、破坏肠道上皮间紧密连接蛋白、减少杯状细胞及潘氏细胞数量损伤肠黏膜屏障,从而促进Apcmin/+小鼠腺瘤生长及癌变。
Purpose Continuous exposure to deoxycholic acid (DOC) induces carcinogenesis of enteric adenomas, but the mechanism is not clear. This study aimed to investigate the role of intestinal mucosal barrier injury in the carcinogenesis of intestinal adenoma induced by DOC. Methods Twenty 20-week-old Apcmin / + mice were randomly divided into control group and DOC group. The control group received routine drinking water. The DOC group received 0.2% DOC drinking water. After 12 weeks of administration, Intestinal adenoma size, number and rate of cancer. The pathological types of gut adenomas were evaluated by HE staining and the intestinal permeability of mice was detected by fluorescein isothiocyanate-conjuagged dextran (FITC-D). Real-time fluorescence quantitative PCR and immunofluorescence Histochemical staining was used to evaluate the expression of various tight junction proteins, goblet cells and Paneth cells in intestinal tissue of the two groups. Results The number of intestinal polyps in Apcmin / + mice was 57.00 ± 3.07, which was significantly higher than that in control group (21.50 ± 4.69, t = 20.03, P <0.0001). The concentration of FITC-D in DOC group was (1.17 ± 0.12) μg / mL, significantly higher than that in control group (0.51 ± 0.07) μg / mL, t = 4.30, P = 0.004. 1 (0.31 ± 0.04) and Claudin 3 mRNA (0.14 ± 0.02) were significantly lower than the control group (0.78 ± 0.07, t = 5.53, P = 0.0003 and 0.92 ± 0.08, , While the expression of Claudin 7 in DOC group was 5.79 ± 0.53, which was significantly higher than that in control group (1.61 ± 0.30, t = 6.83, P = 0.002 4). The number of intestinal staining positive cells in DOC group was PAS (20.88 ± 0.79) and MUC2 (13.10 ± 1.16) were significantly decreased compared with those in control group 35.44 ± 1.68 (t = 7.84, P <0.000 1) and 28.50 ± 0.96 MUC2mRNA expression in murine intestine was 0.15 ± 0.04, lower than that in control group (0.91 ± 0.05, t = 11.34, P <0.0001). The expression of gastric mucin MUC5mRNA in DOC group was 2.40 ± 0.26, which was higher than that in control group 0.02, t = 4.21, P = 0.005 6; The number of lysozyme positive cells in intestinal Pawan’s cells in DOC group was 5.19 ± 0.26, which was significantly lower than that in control group (7.49 ± 0.33, t = 5.49, P = 0.0006). At the same time, the expression of secreted defensins (0.12 ± 0.27) and cryptogenin (0.20 ± 0.35) in Pancreatic cells of DOC group were all lower than that of the control group (0.87 ± 0.05, t = 13.75, t = 10.95, P <0.000 1). Conclusion DOC can promote adenocarcinoma growth and carcinogenesis in Apcmin / + mice by increasing the permeability of intestinal mucosa, destroying the intestinal epithelial tight junction protein, reducing the number of goblet cells and intestinal mucosal barrier.