骨形态发生蛋白9激活PI3K/Akt信号通路抑制退变髓核细胞的炎症反应和凋亡  

BMP9 inhibits inflammatory response and apoptosis of degenerative nucleus pulposus cells through activating PI3K/Akt signaling pathway

作　　者：鲁花[1] 于露[1] 甄欢欢[1] 刘汝银[1] 岳宗进[1] 

机构地区：[1]河南省中医院脊柱科,郑州450002

出　　处：《第三军医大学学报》2016年第18期2047-2052,共6页Journal of Third Military Medical University

摘　　要：目的研究骨形态发生蛋白9(bone morphogenetic protein 9,BMP9)对退变髓核细胞炎症反应和凋亡的影响,并探究其与PI3K/Akt信号通路的关系。方法通过氧糖剥夺(oxygen glucose deprivation,OGD)培养建立退变髓核细胞模型;重组腺相关病毒(adeno-associated virus,AAV)将BMP9转染入人髓核细胞内(human nucleus pulposus cells,HNPCs),实验共分5组:Control组(正常培养的HNPCs)、OGD组(氧糖剥夺模型)、AAV组(氧糖剥夺模型,转染AAV空载体)、AAV-BMP9组(氧糖剥夺模型,转染AAV-BMP9),AAV-BMP9+LY294002组(AAV-BMP9组基础上添加PI3K抑制剂LY294002)。免疫荧光方法检测蛋白聚糖(Aggrecan)和Ⅱ型胶原(typeⅡcollagen,Col2a1)的表达;Western blot检测p-Akt和p-m TOR的蛋白表达;酶联免疫吸附实验检测细胞培养上清液中TNF-α、IL-1β、IL-6和IL-8的表达;流式细胞术检测细胞的凋亡。结果在氧糖剥夺培养模型中Aggrecan和Col2a1表达明显降低;AAV-BMP9组中p-Akt和p-m TOR的表达明显高于OGD组和AAV组(P<0.05);此外,AAV-BMP9组HNPCs分泌的TNF-α、IL-1β、IL-6和IL-8明显减少,细胞凋亡被显著抑制(P<0.05);LY294002的加入能够显著逆转BMP9的上述效果。结论 BMP9能够抑制退变HNPCs的炎症反应和凋亡,并且这种作用是通过激活PI3K/Akt信号通路实现的。Objective To investigate the effects of bone morphogenetic protein 9 （BMP9） on inflammatory response and apoptosis of degenerative nucleus pulposus ceils and to mechanistically explore its association with PI3K/Akt signaling pathway. Methods The degenerative nucleus pulposus ceils were generated by oxygen glucose deprivation （OGD） method. BMP9 gene was transinduced into human nucleus pulposus cells （HNPCs） by recombinant adeno-associated virus （AAV） mediated gene transfer. According to the transfection and treatment, HNPCs were divided into 5 groups： Control group （cultured HNPCs without treatment）, OGD group （ OGD model）, AAV group （ OGD ＋ AAV empty vector）, AAV-BMP9 group （ OGD ＋ AAV-BMP9 ）, and AAV-BMP9 ＋ LY294002 group （ OGD ＋ AAV-BMP9 ＋ PI3K inhibitor LY294002 ）. Immunofluorescence staining was used to detect Aggrecan and typeⅡ collagen （Co12a1）. Western blot were performed to detect the protein expression of p-Akt and p-mTOR. Enzyme linked immunosorbent assay （ELISA） was carried out to measure the levels of TNF-a, IL-1β, IL-6 and IL-8 in the supematant of medium. Flow cytometry was performed to detect cell apoptosis. Results The expression of Aggrecan and Col2al was significantly decreased in OGD model. The protein expression of p-Akt and p-mTOR was higher in AAV-BMP9 group than those in OGD and AAV groups （P 〈0. 05）. In addition, the levels of TNF-a, IL-1β, IL-6 and IL-8 were significantly decreased （P 〈0.05 ） and cell apoptosis was markedly reduced in HNPCs of AAV-BMP9 group. But, LY294002 treatment significantly reversed the above-mentioned effects induced by BMP9 overexpression in degenerative HNPCs. Conclusion BMP9 inhibits the inflammatory response and apoptosis of degenerative HNPCs through activating PI3K/Akt signaling pathway.

关 键 词：骨形态发生蛋白9 炎症反应 细胞凋亡 髓核细胞 

分 类 号：R341[医药卫生—基础医学] R363.22