摘要目的 观察肝再生模型中上皮细胞黏附分子(EpCAM)和转化生长因子-β(TGF-β)的动态表达.方法 将35只雄性F344大鼠(Fisher344大鼠)随机分为假手术组、肝切除组(PH组)和2-乙酰氨基芴组(2-AAH/PH组),采用免疫组织化学染色法观察不同时间点EpCAM阳性细胞在肝组织中的增殖及分布;应用Western blot和反转录-聚合酶链反应(RT-PCR)检测不同时间点TGF-β和激活素受体样激酶-5(ALK-5)的蛋白表达和基因表达.结果 EpCAM阳性细胞在假手术组很难见到;PH组在术后第7天可见少量EpCAM阳性细胞,术后14 d可见阳性细胞向肝内迁移,术后21 d仅见很少量的阳性细胞;2-AAF/PH组术后7、14 d可见与PH组相似结果,但EpCAM阳性细胞明显增多且术后21 d依然可见多量的EpCAM阳性细胞.RT-PCR的结果显示:假手术组TGF-β和ALK-5三时间点的表达差异无统计学意义(TGF-β 2-ΔΔCt值:7 d:1.00±0.00、14 d:1.09±0.16、21 d:1.08±0.13,FTGF-β=0.57,PTGF-β=0.595;ALK-5 2-ΔΔCt值:7 d:1.00 ±0.00、14 d:1.10±0.14、21 d:1.07±0.16,FALK-5=0.54,P=0.610);PH组和2-AAF/PH组TGF-β和ALK-5三时间点表达差异有统计学意义(TGF-β PH组和2-AAF/PH组2-ΔΔCt值:7 d:8.66±0.72、7.79±0.58,14 d:5.36 ±0.53、4.04±0.42,21 d:7.64±0.64、6.88±0.68,FTGF-βpH=21.52,PTGF-β PH=0.002,FTGF-β2.AAF/PH=35.10,PTGF-β2-AAF/PH =0.001;ALK-5 PH组和2-AAF/PH组:7d:4.32±0.52、5.91 ±0.54,14 d:7.79 ±0.74、9.73 ±1.04,21 d:4.85 ±0.46、5.90±0.89,FALK-5PH=30.43,PALK-5 PH=0.001,FALK-52-AAF/PH=20.33,P =0.002).Western blot的结果显示:假手术组TGF-β和ALK-5三时间点的表达差异无统计学意义(TGF-β灰度值:7d:456.4±46.8、14d:454.8±47.1、21 d:458.8±55.6,FTGF-β=0.14,P=0.805;ALK6灰度值:7d:20.9±4.5、14 d:21.3±7.9、21 d:22.4 ±6.5,FALK-5=0.42,P=0.960);PH组和2-AAWPH组TGF-β和ALK-5三时间点表达差异有统计学意义(TGF-β PH组和2-AAF/PH组灰度值:7 d:1 175.2±106.5、882.6 ±83.7,14 d:780.4 ±63.9、639.6±61.3,21 d:1 067.0±91.6、816.2±83.1,FTGF-βpH=6.61,P=0.009,FTGF-β2-AAF/PH=8.06,P=0.012;ALK-5 PH组和2-AAF/PH组:7 d:72.9±7.0、110.1±13.1,14 d:90.1 ±9.4、136.9±12.5,21 d:57.8 ±5.6,91.8±9.7,FALK-5pH=14.05,P=0.005,FALK-52-AAF/PH=10.96,P=0.011).结论 在肝再生过程中,TGF-β表达呈现动态趋势并与EpCAM的表达趋势不同;TGF-β受体ALK-5的表达并不和TGF-β同步.

abstractsObjective Studies of epithelial cell adhesion molecule (EpCAM) and transforming growth factor beta (TGF-β) dynamic expression on liver regeneration model.Provide theoretical basis for further discussion of TGF-β regulatory mechanism in liver regeneration.Methods 35 male F344 (Fisher344) rats were randomly divided into the sham-operated group,partial-hepatectomy (PH) group and the 2-acetaminofluorence (2-AAF/PH) group.Using immunohistochemistry,Western blotting and reverse transcription-polymerase chain reaction (RT-PCR),EpCAM positivecells proliferation,distributionand dynamic expression of TGF-β and active receptor-like kinase 5 (ALK-5) of two model sets were observed at different time points.Results EpCAM positive cell sarescarce in sham-operated group.In PH and 2-AAF/PH groups,a few EpCAM positive cells are observed at 7thand migrated to liver parenchyma at 14th but the positive cells number of later is more.At 21th,EpCAM positive cells are scare in PH group but many inthe 2-AAF/PH.The RT-PCR showed that the expression of TGF-β and ALK-5 was no statistically significant difference on three time points in sham-operated group (2-ΔΔCi value of TGF-β:7 d:1.00±0.00,14 d:1.09 ±0.16,21 d:1.08±0.13,FTGF-β=0.57,P=0.595;ALK-5:7 d:1.00 ±0.00,14 d:1.10 ±0.14,21 d:1.07 ±0.16,FALK-5 =0.54,P =0.610).In PH group and 2-AAF/PH group the difference was statistically significant (TGF-β 2-ΔΔCtvalue of PH group and 2-AAF/PH group:7 d:8.66 ±0.72,7.79 ±0.58;14 d:5.36 ±0.53,4.04 ±0.42;21 d:7.64 ±0.64,6.88 ±0.68;FTGF-βpH =21.52,P =0.002,FTGF-β2-AAF/PH =35.10,P =0.001;ALK-5:7 d:4.32±0.52,5.91 ±0.54;14 d:7.79 ±0.74,9.73 ± 1.04;21 d:4.85 ±0.46,5.90 ±0.89;FALK-5PH =30.43,P =0.001,FALK-52-AAF/PH =20.33,P =0.002).The results of Western blotting indicated that the expression of TGF-5 and ALK-5 was no statistically significant difference on three time points in sham-operated group (gray value of TGF-β:7 d:456.4 ± 46.8;14 d:454.8 ± 47.1;21 d:458.8 ± 55.6;FTGF-β =0.14,P =0.805,FALK-5 =0.42,P =0.960).But the difference was statistically significantin PH group and 2-AAF/PH group (TGF-β gray value of PH group and 2-AAF/PH group:7 d:1 175.2± 106.5,882.6 ±83.7;14 d:780.4 ±63.9,639.6 ±61.3;21 d:1 067.0±91.6,816.2±83.1;FTGF-βpH =6.61,P=0.009,FTGF-β2-AAF/PH =8.06,P=0.012;ALK-5:7 d:72.9 ±7.0,110.1 ±13.1;14 d:90.1 ±9.4,136.9 ±12.5;21 d:57.8 ±5.6,91.8±9.7;FALK-5pH =14.05,P=0.005,FALK-52-AAF/pH =10.96,P=0.011).Conclusion Expression tendency ofTGF-βis different with EpCAM and the expression of ALK-5 and TGF-β are not synchronized in liver regeneration.But the specific mechanism of two phenomena is unknown.