Intervention effects of Compound Houttuyniae Herba to diabetic renal damage based on SOCS-JAK/STAT negative feedback regulation
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摘要:
目的：糖尿病腎?。╠iabetic nephropathy, DN）是糖尿病（Diabetic mellitus，DM）臨床常見的慢性并發(fā)癥之一，本實驗旨在研究復(fù)方魚腥草三種提取部位及其完整成分對db/db小鼠腎損害的保護(hù)作用，并通過探究各提取物對JAK2/STAT3通路及其下游蛋白的影響來探討復(fù)方魚腥草防治糖尿病腎病的可能機(jī)制.方法：1、提取復(fù)方魚腥草水提物、醇提物、揮發(fā)油成分,并將三種成分按照所得質(zhì)量比配制得到復(fù)方魚腥草完整成分。2、選取8只db/m小鼠為正常組；56只db/db小鼠分為模型組、鹽酸二甲雙胍組、AG490組、復(fù)方魚腥草水提物組、復(fù)方魚腥草醇提物組、復(fù)方魚腥草揮發(fā)油組以及復(fù)方魚腥草完整成分組，每組8只，給藥8周，記錄小鼠體重以及血糖，8周后檢測小鼠血糖、腎功能指標(biāo)及腎臟指數(shù)，酶聯(lián)免疫試劑盒法檢測血漿胰島素水平（FINS）、轉(zhuǎn)化生長因子-β1（TGF-β1）、纖連蛋白（FN）、HE染色觀察小鼠腎臟病理變化。3、RT-PCR法檢測各組小鼠腎組織中的JAK2mRNA、STAT3mRNA，SOCS-1mRNA表達(dá)的變化情況； Western blot法檢測腎組織中的JAK2、p-JAK2、STAT3、p-STAT3、SOCS-1、c-fos及c-jun蛋白表達(dá)變化，運(yùn)用免疫熒光法檢測腎組織c-fos、c-jun蛋白的表達(dá)。結(jié)果：1、給藥八周后，與正常組比較，模型組db/db小鼠血漿轉(zhuǎn)化生長因子-β1（TGF-β1）、纖連蛋白（FN）、治療組小鼠血清胰島素水平、胰島素敏感指數(shù)較模型組升高，轉(zhuǎn)化生長因子-β1（TGF-β1），纖連蛋白（FN），水平降低（P<0.05），各治療組小鼠腎臟組織病理改善明顯。2、模型組小鼠腎組織p-JAK2，p-STAT3及JAK2 mRNA，STAT3mRNA的表達(dá)均對比正常組小鼠腎組織的表達(dá)明顯增高(P<0.05)；與模型組相比，各給藥組小鼠腎組織p-JAK2，p-STAT3表達(dá)均有降低，揮發(fā)油組及完整成分組小鼠腎組織SOCS-1基因及蛋白明顯升高（p<0.05）；各給藥組的JAK2、STAT3基因表達(dá)及蛋白變化不顯著。各治療組的c-fos、c-jun在腎小管及腎間質(zhì)的表達(dá)均有不同程度下降。結(jié)論：復(fù)方魚腥草對糖尿病腎損傷具有改善作用，其保護(hù)腎臟的作用可能與降低尿蛋白排泄，調(diào)節(jié)血糖，維持腎臟結(jié)構(gòu)、功能的完整性有關(guān)。其改善糖尿病腎損傷的分子機(jī)制可能通過SOCS 負(fù)反饋調(diào)節(jié)JAK/STAT信號轉(zhuǎn)導(dǎo)通路相關(guān)基因及蛋白表達(dá)，從而抑制降低下游原癌基因c-fos、c-jun的表達(dá)，減少炎癥因子的活化，降低FN的分泌，減少ECM聚積有關(guān)。
Abstract:
Objective To research the protective effects of different extracts from Compound Houttuyniae Herba (CHH) and its mechanism through JAK/STAT-SOCS-1 signaling pathway. Methods The normal group comprised db/m mice (n = 8). db/db mice were randomly divided into seven groups with eight mice in each group according to the applied treatment method: model group, metformin hydrochloride (MH) group, AG490 group, water extract (WE) group, ethanol extract (EE) group, volatile oil (VO) group, and mixture (MG) group (mixture of above three extract) of CHH. After 8 weeks, the general status, biochemical indicators, and renal histological changes in the mice were evaluated, the total RNA and protein were collected and RT-PCR method was used to examine the effect on mRNA expression of JAK2, STAT3, SOCS1, and Western blot method was used to detect the protein expression of JAK2, P-JAK2, STAT3, P-STAT3, SOCS1, c-fos, and c-jun. Immunofluorescence was used to observe the protein expression of c-fos and c-jun in kidney tissue. Results Compared with normal group, the serum level of TGF-β1, FN of EE, VO, and MG groups were decreased (P < 0.05). Renal function was also improved but not significantly. Also, the renal histology was improved especially in the mixture group. The protein expression of JAK2, STAT3, P-JAK2, P-STAT3, and the genetic expression of JAK2 and STAT3 in kidney tissue were significantly increased in the model group (P < 0.05). Compared with the model group, the expression of P-JAK2 and P-STAT3 was down-regulated in treatment groups; The expression of SOCS-1 of VO and mixture groups were elevated (P < 0.05). Conclusion CHH has beneficial effects on diabetic renal injury, which may protect and improve the kidney function and reduce urinary protein, maintain the integrity of the kidney structure and function.
關(guān)鍵詞:
復(fù)方魚腥草;糖尿病腎病;JAK/STAT-SOCS;c-fos;c-jun
Keywords:
Project Supported:
National Natural Science Foundation of China (No. 81403345) and Innovation Program of Shanghai Municipal Education Commission (No. 14YZ059)
