雌激素对十二指肠黏膜碳酸氢盐分泌的影响

Effects of estrogen on bicarbonate secretion of duodenal mucosa

目的探讨雌二醇对十二指肠黏膜碳酸氢盐分泌的影响,观察雌二醇作用的雌激素受体(ER)亚型。方法取4周龄C57雄性小鼠16只,分成对照组和雌二醇组,每组8只。采用化学发光法检测小鼠血清雌二醇水平;实时荧光定量PCR检测十二指肠组织中囊性纤维化穿膜传导调节蛋白(CFTR)、溶质载体家族26(SLC26)A3和SLC26A6 mRNA的表达。采用实时荧光定量PCR分别检测雌二醇加ERα或ERβ阻断剂,以及雌二醇作用于沉默ERα或ERβ24、48 h后SCBN细胞CFTR、SLC26A3和SLC26A6 mRNA的表达。采用高速离子成像系统观察雌二醇对SCBN细胞分泌碳酸氢盐的影响,以及雌二醇对沉默ERα和ERβ后SCBN细胞分泌碳酸氢盐的影响。采用t检验、秩和检验进行统计学分析。结果雌二醇组小鼠血清雌二醇水平高于对照组[(4874±942)pmol/L比(657±187)pmol/L],差异有统计学意义(t=-11.579,P<0.01)。雌二醇组小鼠十二指肠CFTR、SLC26A3和SLC26A6 mRNA水平均高于对照组(0.856±0.302比0.452±0.246,2.910±1.680比1.120±0.540,1.272±0.667比0.319±0.114),差异均有统计学意义(t=-2.317、-2.483、-3.721,P均<0.05)。雌二醇加ERβ阻断剂作用24、48 h后CFTR mRNA和SLC26A6 mRNA水平均低于雌二醇作用24、48 h(CFTR mRNA:0.171±0.059比0.522±0.260,0.111±0.014比0.578±0.297;SLC26A6 mRNA:0.486±0.289比1.118±0.571,0.492±0.231比1.551±0.605),差异均有统计学意义(tCFTRmRNA=2.974、2.655,tSLC26A6 mRNA=2.393、3.272;P均<0.05)。沉默ERα加入雌二醇作用24、48 h后CFTR、SLC26A3和SLC26A6 mRNA水平均高于沉默ERα组(作用24 h:5.073±2.270比1.185±0.494,1.796±1.168比0.468±0.108,3.085±1.357比0.706±0.347;作用48 h:5.025±1.998比1.185±0.494,1.557±0.653比0.468±0.108,3.290±1.750比0.706±0.347),差异均有统计学意义(t24 h=-4.122、-2.773、-4.162,t48 h=-4.604、-4.034、-3.250;P均<0.05)。与沉默ERα组相比,沉默ERα组加入雌二醇作用24、48 h后碳酸氢盐分泌均增加,pH值均升高(0.35±0.17比0.72±0.17、1.15±0.25),差异均有统计学意义(t=-6.516、-12.387,P均<0.01)。结论雌二醇主要通过ERβ上调十二指肠黏膜上皮细胞碳酸氢盐转运蛋白的表达,促进十二指肠黏膜碳酸氢盐的分泌。

Objective To study the effects of estrogen on bicarbonate secretion of duodenal mucosal,and to observe estrogen receptor(ER)subtypes of estrogen.Methods Sixteen 4-week-old male C57 mice were divided into control group and estrogen group,with eight mice in each group.The mice serum level of estrogen was detected by chemiluminescence.The expression of cystic fibrosis transmembrane conductance regulator(CFTR),solute carrier family 26(SLC26)A3 and SLC26A6 in the duodenum tissues were determined by real-time polymerase chain reaction(RT-PCR).After SCBN cells treated with estrogen,ERαand ERβblocking agent,and transfected with silenced ERαand ERβfor 24 and 48 hours,the expression levels of CFTR,SLC26A3 and SLC26A6 mRNA in cells were detected by RT-PCR.The effects of estrogen before and after silenced ERαand ERβon bicarbonate secretion of SCBN cells were observed by high-speed ion imaging system.T test and rank sum test were used for statistical analysis.Results Compared with that of control group,the serum estrogen level of estrogen group was significantly high((4874±942)pmol/L vs.(657±187)pmol/L,t=-11.579,P<0.01).The expression levels of CFTR,SLC26A3 and SLC26A6 mRNAs in duodenum tissues of estrogen group were higher than those of control group(0.856±0.302 vs.0.452±0.246,2.910±1.680 vs.1.120±0.540,1.272±0.667 vs.0.319±0.114),and the differences were statistically significant(t=-2.317,-2.483 and-3.721,all P<0.05).Compared with those treated with estrogen for 24 and 48 hours,the levels of CFTR mRNA and SLC26A6 mRNA were lower after the ERβblocking agent were added into estrogen for 24 and 48 hours(CFTR mRNA:0.171±0.059 vs.0.522±0.260 and 0.111±0.014 vs.0.578±0.297;SLC26A6 mRNA:0.486±0.289 vs.1.118±0.571 and 0.492±0.231 vs.1.551±0.605),and the differences were statistically significant(tCFTRmRNA=2.974 and 2.655,tSLC26A6 mRNA=2.393 and 3.272;all P<0.05).Compared with those of silenced ERαgroup,the levels of CFTR mRNA,SLC26A3 mRNA and SLC26A6 mRNA were higher after ERαsilenced and then added estrogen for 24 and 48 hours(24 h:5.073±2.270 vs.1.185±0.494,1.796±1.168 vs.0.468±0.108 and 3.085±1.357 vs.0.706±0.347;48 h:5.025±1.998 vs.1.185±0.494,1.557±0.653 vs.0.468±0.108 and 3.290±1.750 vs.0.706±0.347),and the differences were statistically significant(t24 h=-4.122,-2.773 and-4.162,t48 h=-4.604,-4.034 and-3.250;all P<0.05).Compared with that of silenced ERαgroup,the bicarbonate secretion increased after ERαsilenced and then added estrogen for 24 and 48 hours(0.72±0.17 and 1.15±0.25 vs.0.35±0.17),and pH also elevated,and the differences were statistically significant(t=-6.516 and-12.387,both P<0.01).Conclusion Estrogen mainly up-regulates the expression of bicarbonate transporter protein in duodenal mucosal epithelial cells through ERβ,and promotes the bicarbonate secretion of duodenal mucosa.