Su MiaoShang, He Yifan, Xue Sichen, Yu Jueke, Ren Xikai, Huang Nan, Abdullahi Rukkaiya, Xu Manhuan
The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Zhejiang 325000, China.
Laboratory Medical and Life Science College, Wenzhou Medical University, Zhejiang 325035, China.
Biosci Rep. 2021 Jun 25;41(6). doi: 10.1042/BSR20203639.
To investigate whether butyric acid could alleviate chronic intermittent hypoxia (CIH)-induced lipid formation in human preadipocytes-subcutaneous (HPA-s) through accumulation of human antigen R (HuR) and inactivation of AMP-activated protein kinase (AMPK) pathway, HPA-s were obtained and divided into three groups: Control group: cells were cultured under normal conditions; CIH group: cells were cultured in a three-gas incubator (10% O2); Butyric acid group: 10 mmol/l butyric acid added into cell culture medium. HuR-siRNA was futher transfected into CIH group for verification the function of HuR. Oil Red O was implemented for observation of lipid droplets within cells. Cell Counting Kit-8 (CCK8) assay was used for detecting cell viability. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-nick end labeling (TUNEL) assay as well as flow cytometry analysis was employed for determining cell apoptosis. Western blotting was used for measurement of protein expression levels. RT-qPCR analysis was used for detecting mRNA expression. CIH treatment increased adipocytes proliferation, while butyric acid inhibited cell proliferation and promoted cell apoptosis. The treatment of butyric acid in CIH group down-regulated expression of inflammatory factors and increased cell apoptotic rate. Butyric acid treatment increased HuR expression in both cytoplasm and nucleus and decreased the level of p-AMPK and p-ACC, while transfection of AMPK activator or HuR-siRNA would down-regulate HuR expression. Moreover, butyric acid alleviated CIH-induced cell proliferation, lipid formation and inflammatory status and promoted cell apoptosis through regulating related genes including p21, PPARγ, C/EBPa, IL-1β, IL-6, TLR4, caspase-8 and caspase-3. In conclusion, butyric acid could alleviate CIH-induced inflammation, cell proliferation and lipid formation through accumulation of HuR and inactivation of AMPK pathway.
为了研究丁酸是否能通过人抗原R（HuR）的积累和AMP激活的蛋白激酶（AMPK）途径的失活来减轻慢性间歇性缺氧（CIH）诱导的人皮下前脂肪细胞（HPA-s）中的脂质形成，获取HPA-s并将其分为三组：对照组：细胞在正常条件下培养；CIH组：细胞在三气培养箱（10% O2）中培养；丁酸组：在细胞培养基中加入10 mmol/l丁酸。将HuR-siRNA进一步转染到CIH组以验证HuR的功能。采用油红O观察细胞内脂滴。使用细胞计数试剂盒-8（CCK8）检测细胞活力。采用末端脱氧核苷酸转移酶介导的脱氧尿苷三磷酸缺口末端标记（TUNEL）检测以及流式细胞术分析来确定细胞凋亡。使用蛋白质免疫印迹法测量蛋白质表达水平。采用RT-qPCR分析检测mRNA表达。CIH处理增加了脂肪细胞增殖，而丁酸抑制细胞增殖并促进细胞凋亡。在CIH组中进行丁酸处理下调了炎症因子的表达并提高了细胞凋亡率。丁酸处理增加了细胞质和细胞核中HuR的表达，并降低了p-AMPK和p-ACC的水平，而转染AMPK激活剂或HuR-siRNA会下调HuR表达。此外，丁酸通过调节包括p21、PPARγ、C/EBPa、IL-1β、IL-6、TLR4、caspase-8和caspase-3在内的相关基因，减轻了CIH诱导的细胞增殖、脂质形成和炎症状态，并促进了细胞凋亡。总之，丁酸可通过HuR的积累和AMPK途径的失活来减轻CIH诱导的炎症、细胞增殖和脂质形成。
