结缔组织生长因子通过上调肾间质成纤维细胞组织型转谷氨酰胺酶和Ⅲ型胶原蛋白促进肾间质纤维化  

Increase of tissue transglutaminase and collagen type Ⅲ induced by connect tissue growth factor in renal interstitial fibroblast contributes to renal interstitial fibrosis

作　　者：刘森炎[1] 钱一欣[1] 卞蓉蓉[1] 郁胜强[1] 

机构地区：[1]上海长征医院肾内科,上海200003

出　　处：《上海医学》2017年第3期173-178,共6页Shanghai Medical Journal

摘　　要：目的观察结缔组织生长因子(CTGF)对肾间质成纤维细胞上组织型转谷氨酰胺酶(tTG)和Ⅲ型胶原蛋白的影响,并探讨其相关的调控机制。方法以大鼠肾间质成纤维细胞(NRK-49F细胞)为研究对象,给予CTGF刺激(不同剂量或不同时间),检测tTG的蛋白相对表达量和mRNA水平。观察CTGF对丝裂原活化蛋白激酶(MAPK)、细胞外信号调节激酶(ERK)1/2和磷酸肌醇3-激酶(PI3K)/蛋白激酶B(Akt)信号转导通路的激活作用和不同信号通路阻断对CTGF调节tTG和Ⅲ型胶原蛋白作用的影响。结果以25、50、100、200 ng/mL的CTGF刺激NRK-49F细胞24 h后的tTG蛋白相对表达量逐渐递增(P<0.05),且均显著高于对照组(P值均<0.05)。以100 ng/mL的CTGF刺激NRK-49F细胞6、12、24 h的tTG蛋白相对表达量逐渐递增(P<0.05),刺激12、24、48 h时的tTG蛋白相对表达量均显著高于对照组(P值均<0.05)。以100 ng/mL的CTGF刺激NRK-49F细胞6、12、24 h后tTG mRNA水平逐渐递增(P<0.05),且均显著高于对照组(P值均<0.05)。以100 ng/mL的CTGF刺激NRK-49F细胞5 rmin时ERK和Akt通路迅速磷酸化,刺激30 rmin后又恢复至基础水平。p38和c-Jun氨基末端激酶信号通路未检测到激活。单独以100.g/mL的CTGF刺激NRK-49F细胞24 h后的tTG蛋白相对表达量显著高于对照组(P<0.05);以100ng/mL的CTGF和20μmol/L的LY294002,以及单独以20μmol/L的LY294002刺激NRK-49F细胞24 h后的tTG蛋白相对表达量与对照组的差异均无统计学意义(P值均>0.05)。使用LY294002阻断P13K/Akt通路后,CTGF对tTG蛋白表达的诱导作用被抑制。单独以100 ng/mL的CTGF,以及100 ng/mL的CTGF和20μmol/L的PD98059刺激NRK-49F细胞24 h后的tTG蛋白相对表达量均显著高于对照组(P值均<0.05);而单独以20μmol/L的PD98059刺激NRK-49F细胞24 h后的tTG蛋白相对表达量与对照组的差异无统计学意义(P>0.05)。使用PD98059阻断ERK1/2 MAPK通路后,CTGF对tTG蛋白表达的诱导作用并未受到影响。单独以100 ng/mL的CTObjective To investigate the effect of connective tissue growth factor （CTGE） on tissue transglutaminase （tTG） and collagen type Ⅲ in renal interstitial fibroblast and the underlying mechanisms. Methods NRK-49F 〈renal interstitial fibroblast） was treated with OTGF at different time points and doses. Then tTG levels were determined by Western blot and reverse transcription-polymerase chain （RT-PCR）. The activation of CTGF on mitogen-activated protein kinases （MAPKs） and phosphoinositide 3-kinase （PI3K）/ protein kinase B （Akt） signal pathway were measured. The effects of blockade of different signal pathways on induction of CTGF on tTG and collagen type m were determined. Results After treatment with OTGF for 24 hours at doses of 25, 50, 100, and 200 ng/mL, tTG protein level of NRK-49F cells was increased in a dose-dependent manner （P〈0.05）. After treatment with 100 ng/mL CTGF for 6, 12, and 24 hours, tTG protein and mRNA levels of NRK-49F cells were increased in a time-dependent manner （both P〈0.05）. After treatment with 100 ng/mL CTGF for 12, 24, and 48 hours, tTG protein level were significantly higher than those in control group （all P〈0.05）. Treatment with 100 ng/mL CTGF induced phosphorylation of extracellular regulated protein kinase （ERK） and Akt pathway at 5 minutes and the pathway recovered at 30 minutes in NRK-49F cells. Activation of p38 and c-Jun was not detected. Treatment with 100 ng/mL CTGF for 24 hours significantly increased tTG protein level in NRK-49F cells （P〈0.05）. Not blockade of PD98059 （20 pmol/L） on ERK pathway but blockade of LY294002 （20 pmol/L） on PI3K/Akt pathway could abolish this upregulation by CTGF （P〈0. 05）. LY294002 （20 μmol/L） or PD98059 （20 μmol/L） alone had no effect on tTG protein level in NRK-49F cells （both P〉0.05）. Treatment with 100 ng/mL CTGF for 24 hours significantly increased cellular and secretory collagen type Ⅲ protein level in NRK-49F cells （both P〈 0.05）. Blockade of L

关 键 词：慢性肾脏病 结缔组织生长因子 组织型转谷氨酰胺酶 Ⅲ型胶原蛋白 

分 类 号：R692[医药卫生—泌尿科学]