CAJ | 학술논문

目的探讨慢性肾功能衰竭大鼠主动脉泛素(Ub)-NF-κB通路的表达变化规律.方法将大鼠随机分为假手术对照组(CON)、肾功能衰竭组(CRF)、肾功能衰竭加MG-132干预组(CRF+M),观察6个月.采用部分肾动脉结扎加对侧肾切除法复制慢性肾功能衰竭大鼠模型.ELISA法测定血液中炎性因子白细胞介素1β(IL-1β)和肿瘤坏死因子α(TNF-α)的含量.RT-PCR半定量测定主动脉中NF-κB及Ub mRNA表达.免疫组织化学方法检测主动脉中NF-κB及Ub蛋白表达.Western印迹法测定主动脉泛素化蛋白的含量.凝胶电泳迁移率(EMSA)法测定动脉中NF-κB的活化情况.结果与CON组比较,CRF组大鼠术后4～6个月时血清中IL-1β[(9.02±1.29)比(2.74±0.96)mg/L]和TNF-α[(50.02±9.52)比(14.04±1.29)mg/L]水平显著升高(P＜0.01);主动脉NF-κB、Ub的mRNA表达升高1.38倍和1.29倍(P＜0.01);N-κB、Ub的蛋白质表达升高3.75倍和20.5倍(P＜0.01);NF-κB活性增强1.82倍(P＜0.01),术后6个月各指标均有进一步上升趋势.与CRF组比较,CRF+M组大鼠干预治疗4～6个月后,大鼠血清中IL-1β[(2.94±0.33)mg/L]和TNF-α[(2.80±2.12)mg/L]含量显著下降(P＜0.01);NF-κB、Ub的mRNA及蛋白质表达显著减少(P＜0.01);NF-κB的活性显著降低(P＜0.01),与CON组比较,差异已无统计学意义,而主动脉中泛素化蛋白含量显著升高(P＜0.01).结论慢性肾功能衰竭大鼠主动脉泛素.NF-κB炎性反应信号明显活化,抑制蛋白酶体活性可能是降低主动脉NF-κB活化的重要药物靶点.
목적 탐토만성신공능쇠갈대서주동맥범소(Ub)-NF-κB통로적표체변화규률.방법 장대서수궤분위가수술대조조(CON)、신공능쇠갈조(CRF)、신공능쇠갈가MG-132간예조(CRF+M),관찰6개월.채용부분신동맥결찰가대측신절제법복제만성신공능쇠갈대서모형.ELISA법측정혈액중염성인자백세포개소1β(IL-1β)화종류배사인자α(TNF-α)적함량.RT-PCR반정량측정주동맥중NF-κB급Ub mRNA표체.면역조직화학방법검측주동맥중NF-κB급Ub단백표체.Western인적법측정주동맥범소화단백적함량.응효전영천이솔(EMSA)법측정동맥중NF-κB적활화정황.결과 여CON조비교,CRF조대서술후4～6개월시혈청중IL-1β[(9.02±1.29)비(2.74±0.96)mg/L]화TNF-α[(50.02±9.52)비(14.04±1.29)mg/L]수평현저승고(P＜0.01);주동맥NF-κB、Ub적mRNA표체승고1.38배화1.29배(P＜0.01);N-κB、Ub적단백질표체승고3.75배화20.5배(P＜0.01);NF-κB활성증강1.82배(P＜0.01),술후6개월각지표균유진일보상승추세.여CRF조비교,CRF+M조대서간예치료4～6개월후,대서혈청중IL-1β[(2.94±0.33)mg/L]화TNF-α[(2.80±2.12)mg/L]함량현저하강(P＜0.01);NF-κB、Ub적mRNA급단백질표체현저감소(P＜0.01);NF-κB적활성현저강저(P＜0.01),여CON조비교,차이이무통계학의의,이주동맥중범소화단백함량현저승고(P＜0.01).결론 만성신공능쇠갈대서주동맥범소.NF-κB염성반응신호명현활화,억제단백매체활성가능시강저주동맥NF-κB활화적중요약물파점.
Objective To investigate the activation of NF-κB and regulation by ubiquitin (Ub)-proteasome pathway in the aorta of rats with chronic renal failure(CRF).Methods The CRF rat model was established by right nephreetomy and left branch renal artery ligation.The CRF rats were were randomly divided into simple CRF group(n=20)and CRF+M used as control group(CON).The NF-κB and the Ub mRNA expression were detected by RT-PCR,and its protein expression was analyzed by immunohistochemistry method.The activity of NF-κBwas mesured by EMSA method.The concentration of IL-1 and TNF-α was detected by ELISA.Results Compared with the CON group,the concentration of serum IL-1 and TNF-α was increased significantly in CRF group [IL-1:(9.02±1.29) vs (2.74±0.96)mg/L,P＜0.01;TNF-α:(50.02±9.52) vs (14.04±1.29)mg/L,P＜0.01]at month 4 after operation.The mRNA expression of NF-κB and Ub in the aorta of CRF group was 1.38 and 1.29 times as that of CON group(P＜0.01).and the protein expression of NF-κB and Ub was 3.75 and 20.5 times as that of CON group(P＜0.01).Compared with the CON group,the activity of NF-κB in the aorta of rats of CRF group was elevated markedly at month 4 after operation(P＜0.01).All the indices were further increased at month 6 after operation.Compared with CRF group,the concentrations of serum IL-1and TNF-α were decreased significantly in CRF+M group[IL-1:(2.94±0.33)mg/L,P＜0.01;TNF-α:(12.80±2.12)mg/L,P＜0.01].The mRNA and protein expression of NF-κB and Ub were also decreased markedly(P＜0.01),and the activity of NF-κB was decreased significantly at month 4 to 6 after operation(P＜0.01).But the amount of ubiquitnative protein was increased significantly in the aorta of CRF+M group as compared to CRF group(P＜0.01). Conclusion The inflammatory signal pathway of ubquitin-proteasome-NF-κB pathway was activated in the aorta of CRF rats,and the proteasome was probablely an important pharmacological intervention target to regulate the activation of NF-κB.