【作者】 张莹；

【导师】 宋光华；

【作者基本信息】 大连医科大学 ， 内科学， 2002， 硕士

【摘要】 目的：糖尿病肾病(Diabetic Nephropathy，DN)是导致终末期肾功能衰竭的主要原因之一，造成DN发生、发展的因素较为复杂，除遗传因素外，尚与血流动力学因素、糖代谢及各种生长因子如血管紧张素Ⅱ(AngⅡ)、内皮素(ET)、转化生长因子-β1(TGF-β1)等因素有关。近年来，二脂酰甘油(DAG)-蛋白激酶C(PKC)通路的活化在DN中的作用已成为国内外众多学者关注的焦点，已证实，DAG-PKC通路激活后，可引起肾小球的高灌注、高滤过，细胞外基质过度生成，从而促进DN的发生、发展，故PKC抑制剂的应用有可能成为阻止和延缓DN发生发展的关键环节。目前国内外关于PKC抑制剂在DN中的作用机制研究甚少，本研究采用对PKC具有较强抑制作用的黄酮类化合物野黄芩甙元(EB)预防性治疗DM大鼠，首次观察其对AngⅡ、ET、TGF-β1水平的影响，并分析其相互关系，探讨PKC抑制剂EB对DN的保护作用及作用机制，同时与血管紧张素转换酶抑制剂(ACEI)卡托普利(captopril)的作用进行比较。 方法：动物随机分为正常对照组(NC组)、糖尿病对照组(DM组)、糖尿病野黄芩甙元(40mg/kg.d-1)治疗组(DM+EB组)及糖尿病卡托普利(100mg/kg.d-1)治疗组(DM+C组)。8周后，采用放射免疫法测定肾组织Angn及血、尿、肾ET水平;应用免疫组织化学法、光镜及电镜技术观察肾组织TGF一pl的表达及病理学改变，并用计算机图象分析系统进行定量分析。 结果:(1)8周时，三组搪尿病大鼠血糖水平相近伊>0.05)，明显高于正常对照组大鼠伊<0.001).(2)与NC组比较，DM组大鼠相对肾重、Cer、UAIb、肾小球基质面积百分比及GBMT(1 3 .5妊1 .24 vs5 .57士0.44，5.93士0.81 vs 3.3肚0.48，34.77士5.98 vs 4.38土1.47，36.5处3.80vs 1 5.2狂3.25，204.25士19.19 vs 143.79土8.53，P值均<0.01)明显增高，表明此时已存在糖尿病肾脏改变;而DM+EB组和DM+C组的上述指标均较nM组明显降低(P值均<0.05)。(3)oM组尿、肾ET、肾^ngll较NC组明显升高(4.3牡1 .20 vs 1 .04土0.13， 8.7矢0.95 vs 4.11士0.74，15.86士1.40 vs 6.3肚0.69，P值均<0.01);肾小球、肾小管TGF一pl灰度值较NC组明显降低(107.26士12.54 vs 164.74士10.01，92.26士12.71 vs132，26士8.43，P<0.0一);而DM+EB组和DM+C组的上述指标较nM组明显改善(P<0.05)。各组大鼠血内皮素水平无明显差异(P>0.05)。(4)相关分析:肾组织的ET、Angll与E了Bw、BS、UAlb、肾小球基质面积百分比、GBMT呈显著正相关(P值均<.0 .01);TGF一pl灰度值与上述指标呈显著负相关(P值均<0.01);肾ET与肾Angll含量呈显著正相关(r=0.74，P<0.0一);肾ET、Angll与肾TGF一pl灰度值呈显著负相关(P<0.01)。 结论:1.DN时，肾内局部Angn、ET及TGF一pl的生成增多，共同促进DN的发生、发展。 2.EB可能通过对PKC的抑制作用抑制肾内局部Angn、ET及TGF一pl的生成，从而起到改善肾血流量、降低尿蛋白、抑制ECM的过度生成的作用，故对DN具有一定的保护作用。更多还原

【Abstract】 Objection: Diabetic nephropathy (DN) is one of the leading cause of end -stage renal failure. A variety of mechanisms have been implicated as contributing to the development of DN such as haemodynamic factors , glucose metabolism and the role of several growth factors. It has been elucidated that activation of protein kinase C(PKC) can cause glumerular hyperfiltration and synthesis of excess extracellular matrix via mediating the production of angiotensin II(AngII), endotheLin-1(ET-1), transforming growth factor- β1(TGF- β1). In the current study, the diabetic rats were prevently treated with PKC inhibitor erigeron breviscapus (EB), observing its effects on the level of serium ET, urne ET , renal ET, renal Angll and renal TGF-β1 .We will shed light on the renoprotective effects and mechanism of EB on DN, at the same time comparing with the role of angiotensin-converting enzyme inhibitor(ACEI) captopril.Method: The rats were randomly devided into normal control rats(group NC), streptozotocin diabetic rats(group DM), diabetic rats treated with EB(groupDM+EB) and diabetic rats treated with captoril(groupDM+C). 8 weeks later, ET in serum, urine and kidney as well as Angll in kidney weremeasured by radioimmunnoassay. Intrarenal TGF-β1 content and the extent of renal pathologic changes were observed by immunohistochemisty, light microscopy, electron microscopy, and then quantitated by using computer image analysis system.Results: (1)8 weeks later, the levels of blood glucose were similar in the three groups of diabetic rats (P>0.05),but much higher than the group NC(P<0.01). (2)Kidney weight/body weight, urinary albumin , creatinine clearance rate (Ccr), percentage of matrix area/glomorular area and glomorular basement membrane thickness(GBMT)in group DM were much higher than those in group NC(13.59± 1.24 vs 5.57±0.44, 5.93±0.81 vs 3.30±0.48, 34.77±5.98 vs 4.38±1.47, 36.59±3.80 vs 18.27±3.25, 204.25±19.19 vs 143.79±8.53, P<0.01 all), which indicate the development of DN. At the same time, the level of ET, Angll and TGF- P 1 in kidney in group DM were significantly increased compared with group NC(P<0.001all). While all above parameters were improved in the diabetic rats treated with EB and Captopril (P<0.05 all). (3) The levels of ET, Angll and TGF- β1 in kidney positively correlated with kidney weight/body weight, blood glucose, Ccr, UAlb, the percentage of matrix area/Glomorular area, and GBMT (P<0.01 all). The renal ET concentration positively correlated with the renal Angll concentration (r=0.74, P<0.01).The levels of renal ET and Angll positively correlated with those of renal TGF- P 1(P<0.01).Conclusions: (1) In DN, the intrarenal Angll, ET and TGF- β1 generation may be increased, which promote the development and progression of DN together. (2) EB can modify glomerular hyperfiltration, reduce urinary albumin protein excretion, decrease matrix synthesis by inhibiting the activity of PKC which can further decrease the production of intrarenal Angll, ETand TGF-β1.更多还原