近端肾小管上皮细胞5羟色胺降解系统激活与肾脏缺血再灌注损伤的关系  

The activation of 5-hydroxytryptamine degradation system in proximal renal tubular epithelial cells induced by renal ischemia-reperfusion injury

作　　者：徐凡 金佳琦 张怡 关晶 梁秀睿 张誉馨 袁安生 刘润坤 傅继华 XU Fan;JIN Jiaqi;ZHANG Yi;GUAN Jing;LIANG Xiurui;ZHANG Yuxin;YUAN Ansheng;LIU Runkun;FU Jihua(School of Basic Medicine and Clinical Pharmacy,China Pharmaceutical University,Nanjing 210009,China;School of Life Science and Technology,China Pharmaceutical University,Nanjing 211198,China;School of Science,China Pharmaceutical University,Nanjing 211198,China)

机构地区：[1]中国药科大学基础医学与临床药学学院,南京210009 [2]中国药科大学生命科学与技术学院 [3]中国药科大学理学院

出　　处：《肾脏病与透析肾移植杂志》2021年第4期337-344,共8页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：国家自然科学基金(81570720);中国药科大学“双一流”学科创新团队建设项目(CPU2018GY23);江苏省大学生创新人才培养计划基金(202010316072S)。

摘　　要：目的:探讨肾脏缺血再灌注损伤(RIRI)与5羟色胺(5-HT)降解的关系,评估抑制5-HT合成及5-HT2A受体(5-HT2AR)对RIRI的治疗效果。方法:雄性Wistar大鼠,通过阻断左肾动脉及静脉血流45 min,切除右肾,然后恢复左肾血液供应诱导RIRI。动物随机分为假手术组,RIRI模型组,5-HT2AR拮抗剂盐酸沙格雷酯(SH)、芳香族L-氨基酸脱羧酶(AADC)抑制剂卡比多巴(CDP)预防性给药组,及SH和CDP联合(SH∶CDP=2∶1)预防性给药组(p-SC)及治疗性给药组(t-SC)。各给药组以相同剂量给药,在再灌注24 h后收集血和肾脏。检测血清肌酐(SCr)、血尿素氮(BUN)浓度及肾组织丙二醛(MDA)、总超氧化物歧化酶(T-SOD)、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)、5-HT和活性氧(ROS)水平。HE染色评估肾损伤。Western印迹和免疫组化检测肾组织色氨酸羟化酶1(Tph1)、AADC、5-HT2AR、单胺氧化酶A(MAO-A)的表达和定位,Western印迹检测Bcl-2相关x蛋白(Bax)、B淋巴细胞瘤2(Bcl-2)、半胱氨酸天冬氨酶蛋白酶3(Caspase3)和Caspase9蛋白表达。TUNEL染色检测肾组织细胞凋亡。结果:与假手术组相比,RIRI模型组肾组织中Tph1、AADC、5-HT2AR、单胺氧化酶A(MAO-A)蛋白表达量显著升高(P<0.05),其高表达部位为近端肾小管上皮细胞,与肾组织病变部位重叠。与模型组比较,SH组、p-SC组及t-SC组的肾组织Tph1、AADC、MAO-A表达量升高被明显抑制(P<0.05),但所有给药组均不能抑制5-HT2AR的表达量升高(P>0.05),且CDP组也明显抑制MAO-A的表达量升高(P<0.05);各给药组明显改善肾小管病变、肾功能指标SCr、BUN及氧化应激指标MDA、T-SOD(P<0.05);降低肾组织5-HT及ROS含量(P<0.05);降低TNF-α、IL-6含量;改善凋亡相关蛋白Bax、Bcl-2、Caspase3、Caspase9表达量(P<0.05)及TUNEL染色显示的细胞凋亡。且p-SC组疗效明显好于SH组及CDP组,显示协同效应,但p-SC组与t-SC组疗效相同。结论:RIRI可能起因于近端肾小管上皮细胞的5-HT合成Objective:To investigate the relationship between renal ischemia-reperfusion injury(RIRI)and 5-hydroxytryptamine(5-HT)degradation and evaluate effect of inhibiting 5-HT synthesis and 5-HT2 A receptor(5-HT2 AR)on RIRI.Methodology:In male Wistar rats,the right kidney was removed and the left kidney was ischemia for 45 min,and then the blood supply was restored for 24 h to induce RIRI.The rats were randomly divided into the sham group;the RIRI model group;pretreated groups,which were treated with 5-HT2 AR antagonist sarpogrelate hydrochloride(SH),aromatic amino acid decarboxylase(AADC)inhibitor carbidopa(CDP),and SH and CDP co-treated preventive administration(p-SC);and therapeutic administration(t-SC)groups(SH∶CDP=2∶1).Each drug-treated group was administered the same dose by gavage.After 24 h of reperfusion,the blood and kidneys were collected.Serum creatinine(SCr)and blood urea nitrogen(BUN),and levels of malondialdehyde(MDA),total superoxide dismutas(T-SOD),tumor necrosis factor-α(TNF-α),interleukin-6(IL-6),5-HT and reactive oxygen species(ROS)in renal tissue were measured.Renal injuries were evaluated by HE staining.The expression and localization of Tryptophan hydroxylase(Tph1),AADC,5-HT2 AR and monoamin oxidase-A(MAO-A)in renal tissue were detected by Western blotting and immunohistochemistry,and the expression of Bax,Bcl-2,Caspase3 and Caspase9 were detected by Western blotting.Apoptosis was detected by TUNEL staining.Results:Compared with the sham group,the expression of Tph1,AADC,5-HT2 AR and MAO-A were significantly increased in renal tissue from the RIRI model group(P<0.05).Furthermore,the high expression regions were in the proximal renal tubular epithelial cells,overlapping with the regions of kidney lesions.Compared with the model group,the expression of Tph1,AADC and MAO-A were significantly prevented in SH,p-SC and t-SC groups(P<0.05);however,the expression of 5-HT2 AR was not suppressed in all drug-treated groups(P>0.05).In the CDP group,the upregulation of MAO-A was also significantly suppre

关 键 词：肾缺血再灌注损伤 5羟色胺 5羟色胺2A受体 活性氧 

分 类 号：R692[医药卫生—泌尿科学]