中国医药
中國醫藥
중국의약
CHINA MEDICINE
2015年
1期
130-134
,共5页
肾小球系膜细胞%脂联素%转化生长因子β1
腎小毬繫膜細胞%脂聯素%轉化生長因子β1
신소구계막세포%지련소%전화생장인자β1
Mesangial cells%Adiponectin%Transforming growth factor beta1
目的 探讨脂联素对脂肪细胞条件培养基促系膜细胞(MsMC)合成转化生长因子β1(TGFβ1)的作用.方法 将MsMC分为7组:A组加入无血清DMEM培养基;B组加入脂肪细胞条件培养基;C组加入pSuper质粒转染后条件培养基;D组加入pcDNA3.1(-)质粒转染后条件培养基;E组加入脂联素siRNA-pSuper表达质粒转染后条件培养基;F组加入脂联素pcDNA3.1(-)表达质粒转染后条件培养基;C组加入添加3 mg/L脂联素的脂肪细胞条件培养基.用脂联素pcDNA3.1(-)表达质粒、脂联素siRNA-pSuper表达质粒、pcDNA3.1(-)质粒及pSuper质粒分别转染3T3-L1细胞(脂质体2000转染法),诱导细胞分化为成熟脂肪细胞后收集细胞,以蛋白质印迹法检测细胞裂解液中脂联素蛋白含量.孵育MsMC 9 h后,收取细胞,用实时定量聚合酶链反应检测细胞裂解液中TGFβ1 mRNA水平;孵育24 h后收取上清液,用酶联免疫吸附试验检测上清液中TGFβ1蛋白质水平.结果 与未转染的成熟脂肪细胞比较,用脂联素pcDNA3.1(-)表达质粒转染3T3-L1细胞得到的成熟脂肪细胞的细胞裂解液中脂联素含量增加[(0.59±0.06)比(0.21±0.02)],差异有统计学意义(P＜0.05);用脂联素siRNA-pSuper表达质粒转染3T3-L1细胞得到的成熟脂肪细胞的细胞裂解液中脂联素含量减少[(0.12±0.01)比(0.39±0.02)],差异有统计学意义(P＜0.05).B组TGFβ1 mRNA水平及蛋白质表达含量高于A、F、G组[TGFβ1 mRNA:(3.09 ±0.15)比(1.50±0.35)、(2.1±0.53)、(1.8±0.42),TGFβ1蛋白质水平:(708±10)比(224±16)、(382±11)、(300 ±25)],差异有统计学意义(P＜0.05).结论 脂肪细胞条件培养基可刺激系膜细胞合成TGFβ1,而脂联素对这一效应具有明显抑制作用.
目的 探討脂聯素對脂肪細胞條件培養基促繫膜細胞(MsMC)閤成轉化生長因子β1(TGFβ1)的作用.方法 將MsMC分為7組:A組加入無血清DMEM培養基;B組加入脂肪細胞條件培養基;C組加入pSuper質粒轉染後條件培養基;D組加入pcDNA3.1(-)質粒轉染後條件培養基;E組加入脂聯素siRNA-pSuper錶達質粒轉染後條件培養基;F組加入脂聯素pcDNA3.1(-)錶達質粒轉染後條件培養基;C組加入添加3 mg/L脂聯素的脂肪細胞條件培養基.用脂聯素pcDNA3.1(-)錶達質粒、脂聯素siRNA-pSuper錶達質粒、pcDNA3.1(-)質粒及pSuper質粒分彆轉染3T3-L1細胞(脂質體2000轉染法),誘導細胞分化為成熟脂肪細胞後收集細胞,以蛋白質印跡法檢測細胞裂解液中脂聯素蛋白含量.孵育MsMC 9 h後,收取細胞,用實時定量聚閤酶鏈反應檢測細胞裂解液中TGFβ1 mRNA水平;孵育24 h後收取上清液,用酶聯免疫吸附試驗檢測上清液中TGFβ1蛋白質水平.結果 與未轉染的成熟脂肪細胞比較,用脂聯素pcDNA3.1(-)錶達質粒轉染3T3-L1細胞得到的成熟脂肪細胞的細胞裂解液中脂聯素含量增加[(0.59±0.06)比(0.21±0.02)],差異有統計學意義(P＜0.05);用脂聯素siRNA-pSuper錶達質粒轉染3T3-L1細胞得到的成熟脂肪細胞的細胞裂解液中脂聯素含量減少[(0.12±0.01)比(0.39±0.02)],差異有統計學意義(P＜0.05).B組TGFβ1 mRNA水平及蛋白質錶達含量高于A、F、G組[TGFβ1 mRNA:(3.09 ±0.15)比(1.50±0.35)、(2.1±0.53)、(1.8±0.42),TGFβ1蛋白質水平:(708±10)比(224±16)、(382±11)、(300 ±25)],差異有統計學意義(P＜0.05).結論 脂肪細胞條件培養基可刺激繫膜細胞閤成TGFβ1,而脂聯素對這一效應具有明顯抑製作用.
목적 탐토지련소대지방세포조건배양기촉계막세포(MsMC)합성전화생장인자β1(TGFβ1)적작용.방법 장MsMC분위7조:A조가입무혈청DMEM배양기;B조가입지방세포조건배양기;C조가입pSuper질립전염후조건배양기;D조가입pcDNA3.1(-)질립전염후조건배양기;E조가입지련소siRNA-pSuper표체질립전염후조건배양기;F조가입지련소pcDNA3.1(-)표체질립전염후조건배양기;C조가입첨가3 mg/L지련소적지방세포조건배양기.용지련소pcDNA3.1(-)표체질립、지련소siRNA-pSuper표체질립、pcDNA3.1(-)질립급pSuper질립분별전염3T3-L1세포(지질체2000전염법),유도세포분화위성숙지방세포후수집세포,이단백질인적법검측세포렬해액중지련소단백함량.부육MsMC 9 h후,수취세포,용실시정량취합매련반응검측세포렬해액중TGFβ1 mRNA수평;부육24 h후수취상청액,용매련면역흡부시험검측상청액중TGFβ1단백질수평.결과 여미전염적성숙지방세포비교,용지련소pcDNA3.1(-)표체질립전염3T3-L1세포득도적성숙지방세포적세포렬해액중지련소함량증가[(0.59±0.06)비(0.21±0.02)],차이유통계학의의(P＜0.05);용지련소siRNA-pSuper표체질립전염3T3-L1세포득도적성숙지방세포적세포렬해액중지련소함량감소[(0.12±0.01)비(0.39±0.02)],차이유통계학의의(P＜0.05).B조TGFβ1 mRNA수평급단백질표체함량고우A、F、G조[TGFβ1 mRNA:(3.09 ±0.15)비(1.50±0.35)、(2.1±0.53)、(1.8±0.42),TGFβ1단백질수평:(708±10)비(224±16)、(382±11)、(300 ±25)],차이유통계학의의(P＜0.05).결론 지방세포조건배양기가자격계막세포합성TGFβ1,이지련소대저일효응구유명현억제작용.
Objective To investigate if adiponectin has an inhibitive effect on transforming growth factor β1 (TGFβ1) up-regulation in mouse mesangial cells induced by adipocyte conditioned medium.Methods Mouse mesangial cells were divided into the following groups:group A had serumfree DMEM medium; group B had fat cell conditioned medium; group C had fat cell transfected with pSuper gene conditioned medium; group D had fat cell transfected with pcDNA3.1 plasmid conditioned medium; group E had fat cell transfected with adiponectin siRNA-pSuper expression plasmid conditioned medium; group F had fat cell transfected with adiponectin pcDNA3.1 expression plasmid conditioned medium;group G had fat cell conditioned medium including 3 mg/L adiponectin.After the mesangial cells were incubated for 9 h,TGFβ1 mRNA in cell lysate was detected by real time PCR; after incubation for 24 h,TGFβ1 protein in supernatant was measured by ELISA.Results Compared with non transfected mature fat cells,the content of adiponectin in cell lysates from mature fat cells which were transfected by adiponectin pcDNA3.1 (-) expression plasmid was significantly increased[(0.59 ± 0.06) vs (0.21 ± 0.02)] ; the content of adiponectin in cell lysates from mature fat cells which were transfected by adiponectin siRNA-pSuper expression plasmid was significantly decreased [(0.12 ± 0.01) vs (0.39 ± 0.02)] (P ＜ 0.05).Compared with group B,the mRNA and protein expression of TGFβ1 was significantly down-regulated in the group A,F,G [TGFβ1 mRNA:(3.09 ±0.15) vs (1.50±0.35),(2.1 ±0.53),(1.8 ±0.42) ;TGFβ1 protein:(708 ± 10) vs (224 ± 16),(382±11),(300 ±25),P＜0.05].Conclusion Adipocyte CM can significantly up-regulate TGFβ1 expression in mouse mesangial cells and adiponectin can signific antly inhibit this effect.
