替米沙坦联合冬虫夏草对糖尿病肾病大鼠肾小管上皮细胞转分化的调节作用
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摘要
近年来，随着糖尿病患者族群的不断扩大，糖尿病目前已经成为了一个全球关注的健康问题。作为糖尿病比较常见的微血管并发症，糖尿病肾病（diabetic nephropathy, DN）的基本病理改变为细胞外基质（extra-cellular matrix, ECM）重构，引起肾小球硬化和肾间质纤维化。在肾间质纤维化的过程中，肾小管上皮细胞间充质转分化（Epithelial-mesenchymal-transition, EMT）起到关键的作用。α平滑肌肌动蛋白（α-smoothmuscle actin, α-SMA）的生成是EMT过程中关键的一步，提示着EMT发生的程度。转化生长因子-β（transforming growth factor-β, TGF-β）在体外是最有效的引起EMT的因子，骨成形蛋白-7(bone morphogenic protein-7, BMP-7)属于TGF-β超家族，一般来说主要表达于肾小管的上皮细胞已经以及部分足细胞。有研究显示，当肾小管的上皮细胞受到损伤时，肾组织中的BMP-7表达明显减少。而当对组织进行外源性的补充BMP-7后，肾功能明显得到改善，提示BMP-7可能为逆转肾间质纤维化的重要的肾脏保护因子。血管紧张素Ⅱ受体拮抗剂（Angiotensin Ⅱ receptor angtagonist, ARB）目前已应用于DN的临床治疗，主要是通过其血流动力学保护肾脏，在抑制肾间质纤维化方面的机制并不明了，冬虫夏草作为一种中药制剂对于肾脏的保护作用机制更是扑朔迷离。本课题通过研究糖尿病肾病大鼠模型肾组织中α-SMA、 BMP-7和TGF-β的表达情况，以及替米沙坦联合冬虫夏草干预下三者的表达情况，同时观察24小时尿蛋白定量和肾功能的变化，以此探讨其对于糖尿病肾病肾小管上皮间充质转分化的调节作用，并寻找替米沙坦联合冬虫夏草对于糖尿病肾病的保护机制，为糖尿病肾病的治疗提供更多的途径及理论依据。
目的：通过研究糖尿病肾病大鼠模型肾组织中α-SMA、 BMP-7和TGF-β的表达情况，以及替米沙坦联合冬虫夏草干预下三者的表达情况，同时观察24小时尿蛋白定量和肾功能的变化，以此探讨其对于糖尿病肾病肾小管上皮间充质转分化的调节作用。
方法：选用雄性Wistar大鼠作为实验对象，动物模型建立使用链脲佐菌素对大鼠一次性腹腔内注射，造模成功后，将大鼠随机分为4组：DN模型组（n＝9）、替米沙坦组（n＝9），冬虫夏草组（n＝9），替米沙坦联合冬虫夏草组（n＝10），另设正常对照组（n＝10）。替米沙坦组每日给予8mg/kg体重的替米沙坦灌胃一次；冬虫夏草组按药量2.5g/Kg虫草水提取液灌胃给药；联合治疗组两种药物均等量灌胃，对照组与模型组以等量饮用水灌胃，每日一次。所有大鼠不限食水。治疗8周后，对大鼠收集24小时尿量测定尿蛋白定量；心包取血测定血糖、血尿素氮、肌酐水平；处死大鼠后采集肾脏标本，测量大鼠肾重及肾重体重比；肾组织HE染色，测定各组大鼠肾小管-间质病理损伤评分，利用组织免疫化学法检测各组大鼠肾组织中BMP-7、α-SMA和TGF-β的表达情况。
结果：（1）对血中BUN、SCr的影响：替米沙坦组、冬虫夏草组及联合用药组能明显降低BUN、SCr水平，与模型组比较有显著性差异（P<0.01），但仍高于正常对照组；联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较差异无显著性意义（P>0.05）。（2）对24小时尿蛋白的影响：替米沙坦组、冬虫夏草组及联合用药组能明显降低24小时尿蛋白水平，与模型组比较有显著性差异（P<0.01），联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较有显著性意义（P<0.01）。（3）肾组织病理：替米沙坦组、冬虫夏草组及联合用药组大鼠与正常对照组大鼠相比，出现了不同程度肾小球硬化、肾间质炎症细胞浸润、肾小管上皮细胞空泡变性及间质纤维化，肾小管-间质损伤指数明显升高（P<0.01），与DN组对比则明显减轻（P<0.01）。（4）肾组织免疫组化：替米沙坦组、冬虫夏草组及联合用药组大鼠肾间质α-SMA及TGF-β阳性表达与模型组相比均显著下调（P<0.01），BMP-7的阳性表达则显著上调（P<0.01），联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较差异无显著性差异（P>0.01）。（5）α-SMA及TGF-β与24小时尿蛋白定量和肾功能成明显正相关（P＜0.01）；BMP-7与24小时尿蛋白定量和肾功能明显负相关（P＜0.01）；α-SMA与TGF-β呈明显正相关（P＜0.01）；BMP-7与α-SMA及TGF-β呈负相关，结果具有统计学意义（P＜0.01）。
结论：
1.糖尿病肾病模型组α-SMA表达明显升高，提示糖尿病肾病过程存在肾小管上皮细胞间充质转分化；
2. α-SMA表达上调与TGF-β明显相关，提示TGF-β可诱导α-SMA的表达，促进EMT的发生；
3. BMP-7与TGF-β、α-SMA表达明显负相关，提示BMP-7可能通过抑制TGF-β的表达，从而抑制EMT的进展；
4.替米沙坦、冬虫夏草可以减少肾组织α-SMA，抑制EMT的发生发展，延缓和改善肾间质纤维化，且联合使用效果更佳。
As the incidence of diabetes (DM) increased in recent years, it has become a global health burden.Diabetic nephropathy (DN) is one of the most common and severe microvascular complications of thediabetes, and its basic pathological change is extracellular matrix (ECM) reconstruction, which lead toglomerulosclerosis and renal interstitial fibrosis. EMT is now recognized as a key contributor to the loss ofrenal function throughout the nephron in diabetic nephropathy. The formation of α-smooth muscle actin(α-SMA) is a critical step in the EMT process, suggesting the extent of EMT. Studies have shown thattransforming growth factor-β (TGF-β) is the most effective factor in vitro caused the EMT. Bonemorphogenetic protein-7(BMP-7) belongs to the TGF-β superfamily, mainly expressed in tubular epithelialand the podocytes tubule cell, when the cells are injured, the expression of BMP-7is decreased. Throughexogenous BMP-7to protect kidney function now is considered as one of the most important factor toreversal renal fibrosis. Angiotensin II receptor antagonists (ARB) has been used clinically, primarilythrough its hemodynamic protection, but the mechanism of inhibition of renal interstitial fibrosis is notclear. Cordyceps sinensis as an important agents for renal protection mechanism is more complicated andconfusing. We will observe the expression of α-SMA、BMP-7and TGF-β in renal tissue of diabeticnephropathy rat model, as well as the change of the three indexes after the treatment of telmisartan andCordyceps, in order to find the effect to renal tubular epithelial to mesenchymal transdifferentiation and themechanism of telmisartan Cordyceps sinensis for protecting kidneys.
Objective: To observe the expression of α-SMA、BMP-7and TGF-β in renal tissue of diabeticnephropathy rat model, as well as the change of the three indexes after the treatment of telmisartan andCordyceps, in order to find the effct of EMT in the development of DN and the mesenchymaltransdifferentiation and the mechanism of telmisartan Cordyceps sinensis for protecting kidneys.
Methods: We established the diabetic nephropathy rat modle by single intraperitoneal injection ofstreptozotocin (STZ). Then randomly divided the DN model rats into four groups: non-treatment DN modelgroup (n=9), telmisartan-treated group (n=9), Cordyceps sinensis treated group (n=9),combination-treated group (n=10). In addition, we selected10normal rats served as control group. Thetelmisartan-treated group rats were gavaged once a day with telmisartan (8mg/kg of body weight). The Cordyceps sinensis-treated group rats were gavaged with Cordyceps sinensis(2.5g/kg of body weight) oncea day. The combination-treated group rats were gavaged with the two drugs once a day. The other twogroups were gavaged with drinking water everyday. The test time lasted for eight weeks, then determinatedthe urine of24hours, the volume of urinary protein and renal function markers. Then observed theexpreesion ofα-SMA、BMP-7and TGF-β by immunohistochemistry ABC method.
Results:(1) The treatment groups rats, including one drug separately and two drugs combination,compare with DN model group rats, the kidney function (both BUN and Cr) is significantly decreased (P<0.01); but increased to normal group.(2) Compared with normal control group rats, the24hours urinaryprotein quality of the DN model group rats were significantly increased (P <0.01), after the treatment oftelmisartan and Cordyceps sinensis separately and both two drugs combination, the24hours urinaryprotein quality significantly decreased (P <0.01).(3) Inflammatory cells infiltration, tubular vacuolizationand tubulointerstitial fibrosis could be seen in DN group, and its index of renal tubular-interstitial injuryelevated significantly （P<0.01）when it was compared with normal group. Renal tubular-interstitial injuryin other three treatment groups significantly released（P<0.01）.(4) The expression of both α-SMA andTGF-β were significantly suppressed among subjects in DN group（P<0.01）, but the expression of BMP-7were contrast decreased (P <0.01). In other three treatment groups, the expression of both α-SMA andTGF-β were significantly reduced and the expression of BMP-7was increased compare with DN group (P<0.01).(5) α-SMA and TGF-β were positively correlated with urinary protein quality and renal functionmarkers significantly (P <0.01); but BMP-7was negatively correlated with them (P <0.01); α-SMA waspositively correlated with TGF-β(P <0.01), BMP-7was negatively correlated with both α-SMA and TGF-βsignificantly (P <0.01).
Conclusions:
1. The increased of expression of α-SMA in DN rats suggestted that there is EMT in DN rats.
2. The expression of α-SMA is concerned with TGF-β significantly, suggesting that TGF-β caninduce the expression of α-SMA, and promote the occurrence of EMT
3. The expression of BMP-7was significantly negatively correlated with α-SMA and TGF-β,suggesting that BMP-7maybe can through inhibition of TGF-β expression to inhibite the progressof the EMT
4. Telmisartan and Cordyceps sinensis can slow down the process of renal fibrosis decreasing ECMand impeding EMT, and the effect is better if they joint application.
目的：通过研究糖尿病肾病大鼠模型肾组织中α-SMA、 BMP-7和TGF-β的表达情况，以及替米沙坦联合冬虫夏草干预下三者的表达情况，同时观察24小时尿蛋白定量和肾功能的变化，以此探讨其对于糖尿病肾病肾小管上皮间充质转分化的调节作用。
方法：选用雄性Wistar大鼠作为实验对象，动物模型建立使用链脲佐菌素对大鼠一次性腹腔内注射，造模成功后，将大鼠随机分为4组：DN模型组（n＝9）、替米沙坦组（n＝9），冬虫夏草组（n＝9），替米沙坦联合冬虫夏草组（n＝10），另设正常对照组（n＝10）。替米沙坦组每日给予8mg/kg体重的替米沙坦灌胃一次；冬虫夏草组按药量2.5g/Kg虫草水提取液灌胃给药；联合治疗组两种药物均等量灌胃，对照组与模型组以等量饮用水灌胃，每日一次。所有大鼠不限食水。治疗8周后，对大鼠收集24小时尿量测定尿蛋白定量；心包取血测定血糖、血尿素氮、肌酐水平；处死大鼠后采集肾脏标本，测量大鼠肾重及肾重体重比；肾组织HE染色，测定各组大鼠肾小管-间质病理损伤评分，利用组织免疫化学法检测各组大鼠肾组织中BMP-7、α-SMA和TGF-β的表达情况。
结果：（1）对血中BUN、SCr的影响：替米沙坦组、冬虫夏草组及联合用药组能明显降低BUN、SCr水平，与模型组比较有显著性差异（P<0.01），但仍高于正常对照组；联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较差异无显著性意义（P>0.05）。（2）对24小时尿蛋白的影响：替米沙坦组、冬虫夏草组及联合用药组能明显降低24小时尿蛋白水平，与模型组比较有显著性差异（P<0.01），联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较有显著性意义（P<0.01）。（3）肾组织病理：替米沙坦组、冬虫夏草组及联合用药组大鼠与正常对照组大鼠相比，出现了不同程度肾小球硬化、肾间质炎症细胞浸润、肾小管上皮细胞空泡变性及间质纤维化，肾小管-间质损伤指数明显升高（P<0.01），与DN组对比则明显减轻（P<0.01）。（4）肾组织免疫组化：替米沙坦组、冬虫夏草组及联合用药组大鼠肾间质α-SMA及TGF-β阳性表达与模型组相比均显著下调（P<0.01），BMP-7的阳性表达则显著上调（P<0.01），联合用药组与替米沙坦组、冬虫夏草组比较有显著性差异（P<0.01）；替米沙坦组与冬虫夏草组比较差异无显著性差异（P>0.01）。（5）α-SMA及TGF-β与24小时尿蛋白定量和肾功能成明显正相关（P＜0.01）；BMP-7与24小时尿蛋白定量和肾功能明显负相关（P＜0.01）；α-SMA与TGF-β呈明显正相关（P＜0.01）；BMP-7与α-SMA及TGF-β呈负相关，结果具有统计学意义（P＜0.01）。
结论：
1.糖尿病肾病模型组α-SMA表达明显升高，提示糖尿病肾病过程存在肾小管上皮细胞间充质转分化；
2. α-SMA表达上调与TGF-β明显相关，提示TGF-β可诱导α-SMA的表达，促进EMT的发生；
3. BMP-7与TGF-β、α-SMA表达明显负相关，提示BMP-7可能通过抑制TGF-β的表达，从而抑制EMT的进展；
4.替米沙坦、冬虫夏草可以减少肾组织α-SMA，抑制EMT的发生发展，延缓和改善肾间质纤维化，且联合使用效果更佳。
As the incidence of diabetes (DM) increased in recent years, it has become a global health burden.Diabetic nephropathy (DN) is one of the most common and severe microvascular complications of thediabetes, and its basic pathological change is extracellular matrix (ECM) reconstruction, which lead toglomerulosclerosis and renal interstitial fibrosis. EMT is now recognized as a key contributor to the loss ofrenal function throughout the nephron in diabetic nephropathy. The formation of α-smooth muscle actin(α-SMA) is a critical step in the EMT process, suggesting the extent of EMT. Studies have shown thattransforming growth factor-β (TGF-β) is the most effective factor in vitro caused the EMT. Bonemorphogenetic protein-7(BMP-7) belongs to the TGF-β superfamily, mainly expressed in tubular epithelialand the podocytes tubule cell, when the cells are injured, the expression of BMP-7is decreased. Throughexogenous BMP-7to protect kidney function now is considered as one of the most important factor toreversal renal fibrosis. Angiotensin II receptor antagonists (ARB) has been used clinically, primarilythrough its hemodynamic protection, but the mechanism of inhibition of renal interstitial fibrosis is notclear. Cordyceps sinensis as an important agents for renal protection mechanism is more complicated andconfusing. We will observe the expression of α-SMA、BMP-7and TGF-β in renal tissue of diabeticnephropathy rat model, as well as the change of the three indexes after the treatment of telmisartan andCordyceps, in order to find the effect to renal tubular epithelial to mesenchymal transdifferentiation and themechanism of telmisartan Cordyceps sinensis for protecting kidneys.
Objective: To observe the expression of α-SMA、BMP-7and TGF-β in renal tissue of diabeticnephropathy rat model, as well as the change of the three indexes after the treatment of telmisartan andCordyceps, in order to find the effct of EMT in the development of DN and the mesenchymaltransdifferentiation and the mechanism of telmisartan Cordyceps sinensis for protecting kidneys.
Methods: We established the diabetic nephropathy rat modle by single intraperitoneal injection ofstreptozotocin (STZ). Then randomly divided the DN model rats into four groups: non-treatment DN modelgroup (n=9), telmisartan-treated group (n=9), Cordyceps sinensis treated group (n=9),combination-treated group (n=10). In addition, we selected10normal rats served as control group. Thetelmisartan-treated group rats were gavaged once a day with telmisartan (8mg/kg of body weight). The Cordyceps sinensis-treated group rats were gavaged with Cordyceps sinensis(2.5g/kg of body weight) oncea day. The combination-treated group rats were gavaged with the two drugs once a day. The other twogroups were gavaged with drinking water everyday. The test time lasted for eight weeks, then determinatedthe urine of24hours, the volume of urinary protein and renal function markers. Then observed theexpreesion ofα-SMA、BMP-7and TGF-β by immunohistochemistry ABC method.
Results:(1) The treatment groups rats, including one drug separately and two drugs combination,compare with DN model group rats, the kidney function (both BUN and Cr) is significantly decreased (P<0.01); but increased to normal group.(2) Compared with normal control group rats, the24hours urinaryprotein quality of the DN model group rats were significantly increased (P <0.01), after the treatment oftelmisartan and Cordyceps sinensis separately and both two drugs combination, the24hours urinaryprotein quality significantly decreased (P <0.01).(3) Inflammatory cells infiltration, tubular vacuolizationand tubulointerstitial fibrosis could be seen in DN group, and its index of renal tubular-interstitial injuryelevated significantly （P<0.01）when it was compared with normal group. Renal tubular-interstitial injuryin other three treatment groups significantly released（P<0.01）.(4) The expression of both α-SMA andTGF-β were significantly suppressed among subjects in DN group（P<0.01）, but the expression of BMP-7were contrast decreased (P <0.01). In other three treatment groups, the expression of both α-SMA andTGF-β were significantly reduced and the expression of BMP-7was increased compare with DN group (P<0.01).(5) α-SMA and TGF-β were positively correlated with urinary protein quality and renal functionmarkers significantly (P <0.01); but BMP-7was negatively correlated with them (P <0.01); α-SMA waspositively correlated with TGF-β(P <0.01), BMP-7was negatively correlated with both α-SMA and TGF-βsignificantly (P <0.01).
Conclusions:
1. The increased of expression of α-SMA in DN rats suggestted that there is EMT in DN rats.
2. The expression of α-SMA is concerned with TGF-β significantly, suggesting that TGF-β caninduce the expression of α-SMA, and promote the occurrence of EMT
3. The expression of BMP-7was significantly negatively correlated with α-SMA and TGF-β,suggesting that BMP-7maybe can through inhibition of TGF-β expression to inhibite the progressof the EMT
4. Telmisartan and Cordyceps sinensis can slow down the process of renal fibrosis decreasing ECMand impeding EMT, and the effect is better if they joint application.
引文
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[16]杨念生,武庆庆,姜宗培,等.维甲酸抑制大鼠单侧输尿管梗阻模型肾间质纤维化[J].中国肾脏病杂志,2004,20(3):194-197.
[17] Eddy AA.Molecular basis of renal fibrosis [J]. Pediatr Nephrol,2010,15:290-301.
[18] Either F, Floege J. Novel insights into renal fibrosis [J]. Curr Opin Nephrol Hypertens.2010,12(3):227-232.
[19] Zeisberg M, Kalluri R. The role of epithelial-to-mesenchymal transition in renal fibrosis[J]. MolMed2009,82:175-181.
[20] Kalluri R, Neilson EG. Epithelial-mesenchymal transition and its implication for fibrosis[J]. ClinInvest,2010,112(12):1776-1784.
[21] Junwei Y, Youhus L. Dissection of key events in tubular epithelial to myofibroblast transition and itsimplication in renal interstitial fibrosis [J]. Am J Pathol,2011,159(4):1465-1475.
[22] Patel S, Takagi K I, Suzuki J, et al. RhoGTPase acti-vation is a key step in renal epithelialmesenchymal transdifferentiation [J]. J Am Soc Nephrol,2010,16:1977-1984.
[23] Rhyu D Y, Yang G Y, Ha H, et al. Role of reactive oxygen species in TGF-beta1-inducedmitogen-activated protein kinase activation and epithelial-mesenchymal transition in renal tubularepithelial cells [J]. J Am Soc Nephrol,2008,16:667-675.
[24] Ozkaynak E, Rueger DC, Drier EA, et al. OP-1cDNA encodes an osteogeneic protein in the TGF-βfamily [J]. EMBO J,2009,9(7):2085-2093.
[25]徐少伟，郑法雷，苏震等.马兜磷酸和TGF-β对肾小管上皮细胞凋亡和转分化的初步探讨[D]．北京：北京协和医院，2007．
[26] Hogan BL. Bone morphogenetic proteins in development [J] Curr Opin Genet Dev.2009,6:432-438．
[27] Vukicevic S, Latin V, Chen P, el a1. Localization of osteogenic protein-1and bone morphogeneticprotein-7[J]. Biol Chem1994;269:16985-16988.
[28] Lu o J. Renin-angiotensin system in diabetic nephropathy [J]. Nefrologia,2010,25(2):73-81.
[29] Yokoi H, Suga Wara A, Mukoyama M, et al. Role of connective tissue growth factor in profibroticaction of transforming growth factor-beta: apotential target for preventing renal fibrosis[J]. Am JKidney Dis,2011,38:134-138.
[30] Goumenos DS, Tsamandas AC, Enahas AM, et al. Apoptos is and myofibrob last expression inglomerular disease: apossible link with transforming growth factor-beta1[J]. Nephron,2010,92(2):287.
[31]秦林燕,刘章锁.替米沙坦对糖尿病肾病大鼠蛋白尿的影响[J].河南职工医学院学报,2009,21(1):1-3.
[32]胡杨青,周巧玲,刘抗寒,等.冬虫夏草对糖尿病肾病鼠肾组织转化生长因子β/c-myc表达的影响[J].肾脏病与透析肾移植杂志,2005,14(5):413-417.
[33]刘丽秋,岳少女亘,赵秀珍,等.冬虫夏草对肾小球硬化大鼠组织抑制因子-1,2mRNA表达的影响[J].肾脏病与透析肾移植杂志,2004,13(5):457-458.
[34]郑丰,杨俊伟,黎磊石.冬虫夏草治疗肾毒性急性肾功能衰竭[J].中国病理生理杂志,1994,10(3):314-316.
[35]戴春笋,刘志红,李恒,等.冬虫夏草防治环孢素A肾毒性的体内与体外研究[J].肾脏病与透析肾移植杂志,2000,9(4):322-325.
[1] Ozkaynak E, Rueger DC, Drier EA, et al. OP-1cDNA encodes an osteogeneic protein in the TGF-βfamily [J]. EMBO J,1990,9(7):2085-2093.
[2]徐少伟，郑法雷，苏震等.马兜磷酸和TGF-β对肾小管上皮细胞凋亡和转分化的初步探讨[D]．北京：北京协和医院，2007．
[3] Hogan B L. Bone morphogenetic proteins in development [J].Curr Opin Genet Dev,2010,6:432-438.
[4] Vukicevic S, Chen P, Latin V, et al. Localization of OP-1and BMP-7[J]. Biol Chem,2009;269:16895-16988.
[5] Waite K A, Eng C. From development disorder to heritable cancer: it’s all in the BMP and TGFfamily[J].Nat Rev Genet,2008,10:73-76.
[6]郭顺华，谌赌璞．BMP-7与肾脏保护作用[J]．国外联学泌尿系统分册，2003,23(4)：411-413．
[7] Luo G, Hofmann C, Bronckers A L, et al. BMP-7is an inducer of nephrogenesis and also required foreye development and skeletal patterning [J].Genes Dev,2005;9:2808-2820.
[8] Boskonda D, Shi M S, Sampath K T, et al. Chcracterization of receptors for OP-1and BMP-7in ratkidneys [J]. Kidjnt,2010,58:1902-1911.
[9] Archdeacon P, Deteiler R K. BMP-7: a critial role in kidney development and a putative madulator ofkidney injury [J]. Adv Chronic Kidney Dis,2008,15(3):314-320.
[10] Goiuld S E, Day M, Simon S, et al. BMP-7regulates chemokine, cytokine and hemodynamic geneexpression in proximal tubule cells [J]. Kid Int,2011,61:51-60.
[11] Simon M, John G M, Stephen E H, et al. Expression of BMP-7jin ischemic rat kidney [J].Am JPhysiol Renal Physiol,1999,276: F382-F389.
[12] Klahr S. The bone morphogenelic proteins (BMPs), Their role in renal fibrils and renal function [J].Nephrol,2009,16(2):179-185.
[13] Mitu G M, Wang S, Hirschberg R. BMP-7is a rodocyte survival factor of diabetic injury [J]. Am JPhysiol Renal Physiol,2007,293(5):1641-1648.
[14] Dudley A T, Robertson E, et al. Overlapping expression of BMPs potentially account for limited tissuedefects in BMP-7[J].Dev Dynamics,2009,208:349-362.
[15] Piscione T D, Yager, Gupta I R, et al. BMP-2and OP-1direct and opposite effect on renal branchingmorphogenesis [J]. Am J Physiol Renal Physiol,1997,273: F961-F975.
[16] Andrew T D, Rbert E G, Elizabeth J R. Interaction between FGF and BMP signaling pathway regulatesdevelopment [J].Genes and Development,2010,13:1601-1613.
[17] Oxburgh L, Dudley AT, Godin RE, et al. BMP4substitutes for loss of BMP7during kidneydevelopment [J]. Devel Biol,2008,286(2):637-646.
[18] Iwano M. Curropin Opin Nephml Hypertens.2004,13(3):279-284.
[19] Zeisberg M, Hanai J, Sugimoto H, et al. BMP-7counteracts TGF-β induced EMT and chronic renalinjury [J]. Nat Med,2009,9(7):964-968.
[20] Wang S N, Lapage J. Loss of tubular BMP-7in diabetic nephropathy [J]. Am Soc Nephrol,2011,12(11):2392-2399.
[21] Michos O, Goncalves A, Lopez-Rios J, et al. Reduction of BMP4activity by gremlin1enablesureteric bud outgrowth and GDNF/WNT11feedback signaling during kidney branchingmorphogenesis [J]. Development,2007,134(13):2397-2405.
[22] Yeh CH, Chang CK, Cheng MF, et al. Decrease of bone morphogenetic protein-7(BMP-7) and its typeⅡ receptor (BMP-RⅡ) in kidney of type1-like diabetic rats [J]. Horm Metab Res,2009,41(8):605-611
[23] De Petris L, Hruska KA, Chiehcio S, et al. Bone morphogenetic protein-7delays podocyte injury dueto high glucose [J].Nephrol Dial Transplant,2007,22(12):3442-3450.
[24] Mitu GM, Wang S, Hischberg R. BMP-7is a podocyte survival factor and rescues podocytes fromdiabetic injury [J].Am J Physiol Renal Physiol,2009,293(5):1641-1648.
[25] Otani H, Otsuka F, Inagaki K, et al. Antagonistic effects of bone morphogenetic protein-4and-7onrenal mesangial cell proliferation induced by aldosterone through MAPK activation [J].Am J PhysiolRenal Physiol,2010,292(5):1513-1525.
[26] Vukicevic S, Basic V, Rogic D, et al. BMP-7reduces seventy of injury after ischemic acute renalfailure in rats [J].Clin Invest,1998,102:201-214.
[27] Marumo T, Hishikawa K, Yoshikawa M, et al. Epigenetic regulation of BMP-7in the regenerativeresponse to ischemia [J].J Am Soe Nephrol,2008,19(7):1311-1320.
[28] Zwsborg M, Bottiglio C, Kumar N, et al. BMP-7inhibits progression of chronic renal fibrosisassociated with two genetic mouse modles [J].Am J Physiol Renal Physiol,2003,285:1060-1067.
[29] Chan WL, Leung JC, Chan LY, et a1.BMP-7protects mesangial cells from injury by polymeric IgA[J].Kidney Int,2008,74(8):1026-1039.
[30] Tyler JR, Robertson H, Booth TA,et a1.Chronic allograft nephropathy: intraepithelial signalsgenerated by transforming growth factor-beta and bone morphogenetic protein-7[J].Am J Transplant,2010,6(6):1367-1376.
[31] Hruska K A, Guo G, Wozniak M, et al. OP-1prevent renal fibrosis associated with ureteral obstruction.[J].Am J Physiol Renal Physiol,2010,279(1):130-143.
[32] Klahr S, Morrissry J. Obstructive nephropathy and renal fibrosis: the role of bone morphogeneticprotein-7and hepatocyte growth factor [J]. Kidney Int Suppl,2009,10(2):105-112.
[33]郑法雷,袁群生.肾-骨对话和肾性骨病:机制与治疗进展[J].中国中西医结合肾病杂志,2008,9(2):95-97.
[34] Davies MR, Lund RJ, Mathew St et a1. Low turnover osteodystrophy and vascular calcification areamenable to skeletal anabolism in an animal model of chronic kidney disease and the metabolicsyndrome [J].J Am Soc Nephrol,2010,16(4):917-928.
[35] Hnmka K A, Mathew S, Davies M R, et al. Connection between vascular calcifiwetion andprogression of chronic kidney disease [J]. Kidney Int Suppl,2011,11(99):142-151.
[36] Freedman BI, Bowden DW, Ziegler JT, et al. Bone morphogenetic protein-7(BMP-7) genepolymorphisms are associated with inverse relationships between vascular calcification and BMD: thediabetes heart study [J].J Bone Miner Res,2009,24(10):1719-1727.
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[4] Bottinger E P, Bitzer M. TGF-β signaling in renal disease [J]. J Am Soc Nephrol,2009,13:2600-2610.
[5] Phanish M K, Wahab N A, Colville-Nash P, et al.The differential role of Smad2and Smad3in theregulation of pro-fibrotic TGF beta1responses in human proximal-tubule epithelial cells [J]. BiochemJ,2006,393:601-607.
[6] Tyler J R, Robertson H, Booth T A, et al. Chronic allograft nephropathy: intraepithelial signalsgenerated by transforming growth factor-beta and bone morphogenetic protein-7[J]. Am J Transplant,2007,6:1367-1376.
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[9] Barnett AH. Preventing renal complications in diabetic patients the diabetics exposed to telmisartan andenalapril(DETAIL) study [J]. Acta Diabetol,2005,42(2):42-48.
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[13]王伯沄,李玉松,黄高昇,等.病理学技术.[M].北京:人民卫生出版社2000:336.
[14] Budwit-Novotny DA, McCarty KS, Cox EB, et al. Immunohistochemical analysis of estrogen receptorin endometrial adenocarcinoma using a monoclonal antibody[J]. Cancer Res,2008,46(10):5419-5425.
[15]刘华锋,陈孝文,吴瑗,等.原发性肾小球肾炎患者肾组织白介素-18的表达[J].中华肾脏病杂志,2003,19(2):127-128.
[16]杨念生,武庆庆,姜宗培,等.维甲酸抑制大鼠单侧输尿管梗阻模型肾间质纤维化[J].中国肾脏病杂志,2004,20(3):194-197.
[17] Eddy AA.Molecular basis of renal fibrosis [J]. Pediatr Nephrol,2010,15:290-301.
[18] Either F, Floege J. Novel insights into renal fibrosis [J]. Curr Opin Nephrol Hypertens.2010,12(3):227-232.
[19] Zeisberg M, Kalluri R. The role of epithelial-to-mesenchymal transition in renal fibrosis[J]. MolMed2009,82:175-181.
[20] Kalluri R, Neilson EG. Epithelial-mesenchymal transition and its implication for fibrosis[J]. ClinInvest,2010,112(12):1776-1784.
[21] Junwei Y, Youhus L. Dissection of key events in tubular epithelial to myofibroblast transition and itsimplication in renal interstitial fibrosis [J]. Am J Pathol,2011,159(4):1465-1475.
[22] Patel S, Takagi K I, Suzuki J, et al. RhoGTPase acti-vation is a key step in renal epithelialmesenchymal transdifferentiation [J]. J Am Soc Nephrol,2010,16:1977-1984.
[23] Rhyu D Y, Yang G Y, Ha H, et al. Role of reactive oxygen species in TGF-beta1-inducedmitogen-activated protein kinase activation and epithelial-mesenchymal transition in renal tubularepithelial cells [J]. J Am Soc Nephrol,2008,16:667-675.
[24] Ozkaynak E, Rueger DC, Drier EA, et al. OP-1cDNA encodes an osteogeneic protein in the TGF-βfamily [J]. EMBO J,2009,9(7):2085-2093.
[25]徐少伟，郑法雷，苏震等.马兜磷酸和TGF-β对肾小管上皮细胞凋亡和转分化的初步探讨[D]．北京：北京协和医院，2007．
[26] Hogan BL. Bone morphogenetic proteins in development [J] Curr Opin Genet Dev.2009,6:432-438．
[27] Vukicevic S, Latin V, Chen P, el a1. Localization of osteogenic protein-1and bone morphogeneticprotein-7[J]. Biol Chem1994;269:16985-16988.
[28] Lu o J. Renin-angiotensin system in diabetic nephropathy [J]. Nefrologia,2010,25(2):73-81.
[29] Yokoi H, Suga Wara A, Mukoyama M, et al. Role of connective tissue growth factor in profibroticaction of transforming growth factor-beta: apotential target for preventing renal fibrosis[J]. Am JKidney Dis,2011,38:134-138.
[30] Goumenos DS, Tsamandas AC, Enahas AM, et al. Apoptos is and myofibrob last expression inglomerular disease: apossible link with transforming growth factor-beta1[J]. Nephron,2010,92(2):287.
[31]秦林燕,刘章锁.替米沙坦对糖尿病肾病大鼠蛋白尿的影响[J].河南职工医学院学报,2009,21(1):1-3.
[32]胡杨青,周巧玲,刘抗寒,等.冬虫夏草对糖尿病肾病鼠肾组织转化生长因子β/c-myc表达的影响[J].肾脏病与透析肾移植杂志,2005,14(5):413-417.
[33]刘丽秋,岳少女亘,赵秀珍,等.冬虫夏草对肾小球硬化大鼠组织抑制因子-1,2mRNA表达的影响[J].肾脏病与透析肾移植杂志,2004,13(5):457-458.
[34]郑丰,杨俊伟,黎磊石.冬虫夏草治疗肾毒性急性肾功能衰竭[J].中国病理生理杂志,1994,10(3):314-316.
[35]戴春笋,刘志红,李恒,等.冬虫夏草防治环孢素A肾毒性的体内与体外研究[J].肾脏病与透析肾移植杂志,2000,9(4):322-325.
[1] Ozkaynak E, Rueger DC, Drier EA, et al. OP-1cDNA encodes an osteogeneic protein in the TGF-βfamily [J]. EMBO J,1990,9(7):2085-2093.
[2]徐少伟，郑法雷，苏震等.马兜磷酸和TGF-β对肾小管上皮细胞凋亡和转分化的初步探讨[D]．北京：北京协和医院，2007．
[3] Hogan B L. Bone morphogenetic proteins in development [J].Curr Opin Genet Dev,2010,6:432-438.
[4] Vukicevic S, Chen P, Latin V, et al. Localization of OP-1and BMP-7[J]. Biol Chem,2009;269:16895-16988.
[5] Waite K A, Eng C. From development disorder to heritable cancer: it’s all in the BMP and TGFfamily[J].Nat Rev Genet,2008,10:73-76.
[6]郭顺华，谌赌璞．BMP-7与肾脏保护作用[J]．国外联学泌尿系统分册，2003,23(4)：411-413．
[7] Luo G, Hofmann C, Bronckers A L, et al. BMP-7is an inducer of nephrogenesis and also required foreye development and skeletal patterning [J].Genes Dev,2005;9:2808-2820.
[8] Boskonda D, Shi M S, Sampath K T, et al. Chcracterization of receptors for OP-1and BMP-7in ratkidneys [J]. Kidjnt,2010,58:1902-1911.
[9] Archdeacon P, Deteiler R K. BMP-7: a critial role in kidney development and a putative madulator ofkidney injury [J]. Adv Chronic Kidney Dis,2008,15(3):314-320.
[10] Goiuld S E, Day M, Simon S, et al. BMP-7regulates chemokine, cytokine and hemodynamic geneexpression in proximal tubule cells [J]. Kid Int,2011,61:51-60.
[11] Simon M, John G M, Stephen E H, et al. Expression of BMP-7jin ischemic rat kidney [J].Am JPhysiol Renal Physiol,1999,276: F382-F389.
[12] Klahr S. The bone morphogenelic proteins (BMPs), Their role in renal fibrils and renal function [J].Nephrol,2009,16(2):179-185.
[13] Mitu G M, Wang S, Hirschberg R. BMP-7is a rodocyte survival factor of diabetic injury [J]. Am JPhysiol Renal Physiol,2007,293(5):1641-1648.
[14] Dudley A T, Robertson E, et al. Overlapping expression of BMPs potentially account for limited tissuedefects in BMP-7[J].Dev Dynamics,2009,208:349-362.
[15] Piscione T D, Yager, Gupta I R, et al. BMP-2and OP-1direct and opposite effect on renal branchingmorphogenesis [J]. Am J Physiol Renal Physiol,1997,273: F961-F975.
[16] Andrew T D, Rbert E G, Elizabeth J R. Interaction between FGF and BMP signaling pathway regulatesdevelopment [J].Genes and Development,2010,13:1601-1613.
[17] Oxburgh L, Dudley AT, Godin RE, et al. BMP4substitutes for loss of BMP7during kidneydevelopment [J]. Devel Biol,2008,286(2):637-646.
[18] Iwano M. Curropin Opin Nephml Hypertens.2004,13(3):279-284.
[19] Zeisberg M, Hanai J, Sugimoto H, et al. BMP-7counteracts TGF-β induced EMT and chronic renalinjury [J]. Nat Med,2009,9(7):964-968.
[20] Wang S N, Lapage J. Loss of tubular BMP-7in diabetic nephropathy [J]. Am Soc Nephrol,2011,12(11):2392-2399.
[21] Michos O, Goncalves A, Lopez-Rios J, et al. Reduction of BMP4activity by gremlin1enablesureteric bud outgrowth and GDNF/WNT11feedback signaling during kidney branchingmorphogenesis [J]. Development,2007,134(13):2397-2405.
[22] Yeh CH, Chang CK, Cheng MF, et al. Decrease of bone morphogenetic protein-7(BMP-7) and its typeⅡ receptor (BMP-RⅡ) in kidney of type1-like diabetic rats [J]. Horm Metab Res,2009,41(8):605-611
[23] De Petris L, Hruska KA, Chiehcio S, et al. Bone morphogenetic protein-7delays podocyte injury dueto high glucose [J].Nephrol Dial Transplant,2007,22(12):3442-3450.
[24] Mitu GM, Wang S, Hischberg R. BMP-7is a podocyte survival factor and rescues podocytes fromdiabetic injury [J].Am J Physiol Renal Physiol,2009,293(5):1641-1648.
[25] Otani H, Otsuka F, Inagaki K, et al. Antagonistic effects of bone morphogenetic protein-4and-7onrenal mesangial cell proliferation induced by aldosterone through MAPK activation [J].Am J PhysiolRenal Physiol,2010,292(5):1513-1525.
[26] Vukicevic S, Basic V, Rogic D, et al. BMP-7reduces seventy of injury after ischemic acute renalfailure in rats [J].Clin Invest,1998,102:201-214.
[27] Marumo T, Hishikawa K, Yoshikawa M, et al. Epigenetic regulation of BMP-7in the regenerativeresponse to ischemia [J].J Am Soe Nephrol,2008,19(7):1311-1320.
[28] Zwsborg M, Bottiglio C, Kumar N, et al. BMP-7inhibits progression of chronic renal fibrosisassociated with two genetic mouse modles [J].Am J Physiol Renal Physiol,2003,285:1060-1067.
[29] Chan WL, Leung JC, Chan LY, et a1.BMP-7protects mesangial cells from injury by polymeric IgA[J].Kidney Int,2008,74(8):1026-1039.
[30] Tyler JR, Robertson H, Booth TA,et a1.Chronic allograft nephropathy: intraepithelial signalsgenerated by transforming growth factor-beta and bone morphogenetic protein-7[J].Am J Transplant,2010,6(6):1367-1376.
[31] Hruska K A, Guo G, Wozniak M, et al. OP-1prevent renal fibrosis associated with ureteral obstruction.[J].Am J Physiol Renal Physiol,2010,279(1):130-143.
[32] Klahr S, Morrissry J. Obstructive nephropathy and renal fibrosis: the role of bone morphogeneticprotein-7and hepatocyte growth factor [J]. Kidney Int Suppl,2009,10(2):105-112.
[33]郑法雷,袁群生.肾-骨对话和肾性骨病:机制与治疗进展[J].中国中西医结合肾病杂志,2008,9(2):95-97.
[34] Davies MR, Lund RJ, Mathew St et a1. Low turnover osteodystrophy and vascular calcification areamenable to skeletal anabolism in an animal model of chronic kidney disease and the metabolicsyndrome [J].J Am Soc Nephrol,2010,16(4):917-928.
[35] Hnmka K A, Mathew S, Davies M R, et al. Connection between vascular calcifiwetion andprogression of chronic kidney disease [J]. Kidney Int Suppl,2011,11(99):142-151.
[36] Freedman BI, Bowden DW, Ziegler JT, et al. Bone morphogenetic protein-7(BMP-7) genepolymorphisms are associated with inverse relationships between vascular calcification and BMD: thediabetes heart study [J].J Bone Miner Res,2009,24(10):1719-1727.