摘要：目的研究血小板源生长因子(PDGF)对肾小球系膜细胞(MC)表型转化的影响，并探讨丝裂素活化蛋白激酶(MAPK)信号转导途径在其作用中的调控机制。方法以体外培养的大鼠MC为对象，观察PDCF对细胞增殖、α-平滑肌肌动蛋白(SMA)表达和细胞内。C-Jun氨基末端激酶(JNK)活性的作用；观察细胞外信号调节激酶(ERK)途径上游信号MEK-1阻断剂PD98059对PDGF所致上述作用的影响；应用raf-1基因转染并稳定高表达的MC观察其对α-SMA表达、细胞形态及其超微结构的影响。结果PDCF可刺激MC持续增殖；刺激6～48h可使α-SMA表达增加，但持续刺激72h以上则可导致其表达被抑制近50％。PDCF对JNK活性无明显影响，阻断ERK活化不影响JNK活性及PDGF对α-SMA表达的短时刺激作用。Raf-1蛋白激酶稳定高表达的MC细胞内α-SMA表达缺失，同时细胞内微丝、微管结构减少，细胞形态改变。结论PDGF对系膜细胞表型转化的影响表现为促进细胞持续增殖、短时作用刺激α-SMA表达而持续作用则抑制α-SMA表达，提示其为具有调控MC增殖和分化双重作用的细胞因子。PDGF的作用可能与MAPK不同亚类的调控有关，其中Raf-1和JNK活性可能对细胞表型具有关键调控作用。

Regulatory effects of signal transduction through MAPK pathways on PDGF-induced phenotypic change in rat glomerular mesangial cells

LI Xiaomei ,TANG Jiawei ,LI Biao,et al.

(Renal Division,Department of Medicine,First Hospital and Institute of Nephrology,Peking University,Beijing 100034,China)

Abstract：Objective To investigate effects of PDGF on phenotypic change of glomerular mesangial cells(MC),and study intracellular mechanisms through mitogen-activated-protein kinase(MAPK)pathways.Methods After stimulation with PDGF,cell proliferation and expression of α-smooth muscle actin(α-SMA) were detected in cultured rat MC.Expression of α-SMA and activity of JNK,which may be a intracellular signaling event in cell differentiation,was measured with or without antagonist of MEK-1.Then MC was stably transfected with constitutively active Raf-1 (BxB Raf).Phenotypic differences including expression of α-SMA and cell morphology between Raf-1 transfectants and normal cells were assessed.Results PDGF stimulated MC proliferation from 24 to 120 hours.Expression of α-SMA was enhanced frmn 6 to 48 hours stimulation,but was inhibited about 50% after 72 hours stimulation.PDGF did uot influence JNK activity.PDGF-induced level of JNK activity and enhanced expression of α-SMA were not influenced by treatment with MEK-1 antagonist PD98059.In the BxB-Raf expressing MC with a constitutively elevated ERK activity,JNK activity decreased by 50% and expression of α-SMA was completely absent.Less detectable microfilaments and morphologic change were found in these cells.Conclusions PDGF stimulates cell proliferation,promotes expression of α-SMA at a short period but inhibits it at persistent stimulation.PDGF regulates not only MC proliferation but also their differentiation process.These effects appear to be regulated by MAPK signaling pathways,in which activation of Raf-1 and JNK may be the critical events.

Keywords：Platelet derived growth factor;Cell differentiation;Cytoskeleton;Signal transduction

基金项目：国家自然科学基金资助项目(39770346)；教育部跨世纪人才基金资助项目(39910210474-231-C04)

参考文献：

［1］Wang HY,Chen YP.Mesangioproliferative disease.In:Neilson EG,Couse WG.eds.Immunologic Renal Diseases.Philadelphia:Lippiocott-Raven Publishers.1997.993-1002.

［2］Johnson RJ.Nephrology forum:the glomerular response to inury.Mechanisms of progression and resolution.Kidney Int.1994,45:1769-1782.

［3］王玉,邹万忠,李晓玫.肾小球肾炎中系膜细胞α-平滑肌肌动蛋白表过与细胞表型变化的关系.中华肾脏病杂志,1998.14:206-209.

［4］Li Xiaomei.Zarinetehi F.Schrier RW.et al.Inhibition of MAP kinase by prostaglandin E2 and forskolin in rat renal mesangial cells.Am J Physiol.1995,269:986-991.

［5］李彪,李晓玫.raf-1基因高表达对肾小球系膜细胞增殖表型的影响.中华医学杂志,1999,79:529-532.

［6］蔡琪,李晓玫,王海燕.c-Jun氨基末端激酶活性测定.北京医科大学学报,1999,31:283-284.

［7］Wills JW.Cytoskeletal regulation of membrane transport events.FASEB J.1994.8:1161-1165.

［8］Abboub HE.Platelet-derived growth factor and mesangial cells.Kidney Int.1992.41:581-583.

［9］Johnson RJ,Floege J.Couser WG,et al.Role of platelet-derived growth factor in glomerular disease.J Am Soc Nephrol.1993.4:119-128.

［10］Providence KM,Kutz SM,Higgins PJ,et al.Perturbation of the actin cvtoskeleton induces PAI-1 gene expression in euhured epithelial cells independent of substrate anchorage.Cell Motil Cytoskeleton.1999.42:218-229.

［11］Utsunomiya Y.Kawamura T,Abe A,et al.Significance of mesan gial expression of alpha smooth muscle actin in the progression of IgA nephropathy.Am J Kidney Dis.1999,34:902-910.

［12］Bornfeldt KE,Raines EW,Graves LM,et al.Platelel-derived growth factor:distract signal transduction pathways associated with migration versus proliferation.Ann New York Acad Sci.1995,416-430.

［13］Choudhury GG,Marra F,Kiyomoto H,el al.PDGF stimulates tyrosine phosphrylation of JAK1 protein tyrosine kinase in human mesangial cells.Kidney Int.1996.49:19-25.

［14］Garrington TP,Johnson GL,Organization and regulation of mito gen-activated protein kinase signaling pathways.Curr Opin Cell Biol.1999.11:211-218.

［15］李晓玫,唐嘉薇,王海燕.IL-1对肾系膜细胞的分裂原活化蛋白激酶不同亚类的作用.中国免疫学杂志,1998,14:300-303.

收稿日期：2000-10-12