| 王晓辉,徐 倩,杨志华,李 兵,邢 媛,张媛媛,高保东,哈小琴.KGF联合HIF-1α对BECN1敲除IEC-6细胞缺氧的保护作用及机制[J].,2023,(10):1849-1856 |
| KGF联合HIF-1α对BECN1敲除IEC-6细胞缺氧的保护作用及机制 |
| Protective Effect and Mechanism of KGF Combined with HIF-1α on Hypoxia in BECN1 Knockout IEC-6 Cells |
| 投稿时间：2022-09-30 修订日期：2022-10-28 |
| DOI：10.13241/j.cnki.pmb.2023.10.008 |
| 中文关键词: 角质细胞生长因子 缺氧诱导因子1 肠影窝上皮细胞 缺氧 自噬 |
| 英文关键词: Keratinocyte Growth Factor Hypoxia Inducible Factor 1 Intestinal Epithelial Cells Hypoxia Autophagy |
| 基金项目:兰州市创新人才项目（2016-RC-61）；军队后勤科研项目（CWH17C008） |
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| 中文摘要: |
| 摘要 目的：探讨KGF联合HIF-1α对BECN1敲除后的IEC-6细胞在缺氧应激状态下的保护作用及相关机制。方法：构建敲除BECN1基因的稳定细胞系IEC-6B-细胞，分为空白对照组（BC）、阴性对照组（NC）、HIF-1α组、KGF联合HIF-1α组（KH）。观察细胞形态学改变。检测细胞存活率、ATP含量、细胞周期和细胞凋亡率。检测自噬相关基因表达水平和凋亡、自噬相关蛋白的表达。结果：低氧处理24 h后，各组细胞可见梭形、星形及其它异常形态变化；NC组细胞胞质内可见大量大小不一的吞噬溶酶体泡；其他各组细胞呈现细胞早期凋亡形态变化，BC组和KH组细胞胞质中可见自噬泡。与BC组相比较，各敲除组细胞存活率均显著降低（P<0.01），其中KH组细胞存活率高于NC组、KGF组及HIF-1α组（P<0.05）。BC组细胞内ATP含量均显著高于其他各组（P<0.05）。与BC组和KH组比较，其他三组细胞G0/G1期百分比显著增加，细胞凋亡率均显著增加（P<0.05）。与BC组相比较，其余各组细胞自噬基因BECN1、SQSTM1及LC3基因mRNA表达均显著降低（P<0.05）；Beclin 1蛋白及LC3 II/LC3 I的比值均显著降低（P<0.05），且p62蛋白表达显著增加（P<0.05）。KH组的Bax/Bcl-2的比值、Caspase3蛋白表达低于NC组（P<0.05）。结论：KGF联合HIF-1α能促进细胞增殖、增强细胞能量代谢、减少G0/G1期阻滞细胞率、抑制细胞凋亡作用，对低氧应激的IEC-6B-细胞具有保护作用。 |
| 英文摘要: |
| ABSTRACT Objective: To investigate the protective effect and mechanism of KGF combined with HIF-1α on IEC-6 cells knocked out by BECN1 under hypoxia stress. Methods: The stable BECN1 knockout cell line IEC-6B-cells were constructed and divided into blank control group (BC), negative control group (NC) and HIF-1 α Group, KGF joint HIF-1 α Group (KH). Observed the changes of cell morphology in each group. Cell survival rate, ATP content, cell cycle and apoptosis rate, autophagy related gene expression, apoptosis and autophagy related protein expression were detected in each group. Results: After 24 hours of hypoxia treatment, spindle, star and other abnormal morphological changes were observed in cells of each group. Electron microscopy showed that a large number of lysosomal vesicles with different sizes were found in the cytoplasm of NC cells. Other groups of cells showed morphological changes of early apoptosis, and autophagic vesicles were found in the cytoplasm of cells in BC group and KH group. Compared with BC group, the cell survival rate of each knockout group was significantly lower(P<0.01), and the cell survival rate of KH group was higher than NC group, KGF group and HIF-1α group(P<0.05). The content of ATP in BC group was significantly higher than that in other groups (P<0.05). Compared with BC group and KH group, the percentage of G0/G1 phase and apoptosis rate of other three groups were significantly increased (P<0.05). There was no significant difference between KH group and BC group (P>0.05). Compared with BC group, the expression of BECN1, SQSTM1 and LC3 genes in NC group, KGF group, HIF-1α group and KH group decreased significantly (P<0.05). Compared with BC group, the expression of Beclin1 protein and LC3 II/LC3 I in NC group, KGF group, HIF-1 α group and KH group decreased significantly(P<0.05), and the expression of p62 protein increased significantly(P<0.05). The ratio of Bax/Bcl-2 and the expression of Caspase3 protein in KH group were lower than those in NC group(P<0.05). Conclusion: KGF combined with HIF-1α can protect IEC-6B- cells from hypoxia stress by promoting cell proliferation, enhancing cell energy metabolism, reducing cell arrest rate in G0/G1 phase and inhibiting apoptosis. |
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