CD147通过AIM2炎症小体介导宫颈癌细胞焦亡和增殖

王玲; 秦祥川; 李金秋; 阿仙姑·哈斯木

doi:10.12259/j.issn.2095-610X.S20240103

CD147通过AIM2炎症小体介导宫颈癌细胞焦亡和增殖

doi: 10.12259/j.issn.2095-610X.S20240103

1.
新疆医科大学基础医学院/新疆地方病分子生物学重点实验室，新疆乌鲁木齐　836001

基金项目: 国家自然科学基金资助项目（82260516）

详细信息

作者简介:
王玲（1995～），女，山西运城人，在读硕士研究生，主要从事宫颈癌发病机制的研究工作

通讯作者:
阿仙姑·哈斯木，E-mail：axiangu75@126.com

中图分类号: R737.33
计量
- 文章访问数: 711
- HTML全文浏览量: 462
- PDF下载量: 12
- 被引次数: 0
出版历程
- 收稿日期: 2023-10-30
- 网络出版日期: 2024-01-09
- 刊出日期: 2024-01-25

CD147 Mediates Cervical Cancer Cell Pyroptosis and Proliferation through AIM2 Inflammasome

1.
Basic Medicine College of Xinjiang Medical University/Xinjiang Key Laboratory of Molecular Biology of Endemic Diseases，Urumqi Xinjiang 836001，China

摘要

摘要: 目的探讨跨膜蛋白CD147表达量变化对AIM2炎症小体介导的宫颈癌细胞焦亡及增殖的影响。方法采用Western blot 实验检测 CD147 在宫颈癌细胞 SiHa（HPV+）、C33a（HPV-）和正常宫颈上皮细胞 H8（HPV+）、HCer Epic（HPV-）中的表达水平；通过慢病毒转染 SiHa细胞下调CD147的表达，根据不同处理分为SiHa组、阴性对照组（shCD147-NON）、敲低组1（shCD147-1）和敲低组2（shCD147-2），通过Western blot、RT-qPCR和观察细胞绿色荧光表达验证转染效果；通过Western blot 和 RT-qPCR 检测CD147和AIM2炎症小体相关因子 AIM2、Caspase-1、IL-18、GSDMD蛋白和 mRNA 的表达；检测细胞培养上清中乳酸脱氢酶（LDH）释放度，荧光倒置显微镜下观察细胞的形态；CCK-8 实验检测细胞的增殖能力；细胞克隆实验检测细胞集落形成能力。结果 Western blot 结果显示，与 HCerEpic细胞比较，SiHa 细胞中 CD147 蛋白表达最高（P < 0.05）； CD147低表达慢病毒有效下调了SiHa细胞 CD147表达水平（P < 0.05）；Western blot 及 RT-qPCR实验结果表明，与SiHa组相比， shCD147-1 组和 shCD147-2 组AIM2、Caspase-1、IL-18、GSDMD蛋白和 mRNA 表达明显升高（P < 0.05）；乳酸脱氢酶（LDH）释放实验显示，与SiHa组相比，shCD147组LDH释放度明显升高（P < 0.05）；荧光倒置显微镜下显示，shCD147组出现肿胀和空泡化，表现出典型的细胞焦亡现象；与SiHa组相比，shCD147-1组和shCD147-2组细胞增殖能力和集落形成能力明显降低（P < 0.05）。结论 CD147低表达有效上调宫颈癌 SiHa 细胞AIM2炎症相关因子的表达，诱发细胞焦亡，抑制细胞的增殖和克隆。
- 宫颈癌 /
- CD147 /
- AIM2炎症小体 /
- 细胞焦亡 /
- 增殖
Abstract: Objective To investigate the effect of transmembrane protein CD147 expression on AIM2 inflammasome-mediated pyroptosis and proliferation of cervical cancer cells. Methods Western Blot was used to detect the expression of CD147 in cervical cancer cell lines SiHa（HPV+） and C33a（HPV-） and normal cervical epithelial cells H8（HPV+） and HCer Epic（HPV-）. SiHa cells were transfected with lentivirus to down-regulate the expression of CD147. According to the different treatments, SiHa cells were divided into SiHa group, negative control group（shCD147-NON）, knockdown group 1（shCD147-1） and knockdown group 2（shCD147-2）. The transfection effect was verified by Western Blot, RT-qPCR and green fluorescence expression. The protein and mRNA expressions of AIM2, Caspase-1, IL-18 and GSDMD were detected by Western Blot and RT-qPCR. The lactate dehydrogenase（LDH） release was measured in the cell culture supernatant, and the cell morphology was observed under the fluorescence inverted microscope; the proliferation ability of cells was measured by CCK-8 and the colony formation ability was measured by cell cloning experiments. Results Western Blot results showed that CD147 protein expression in SiHa cells was the highest compared with that in HCerEpic cells. CD147 low expression lentivirus effectively down-regulated the expression of CD147 in SiHa cells. The results of Western Blot and RT-qPCR experiments showed that the expression of AIM 2, Caspase-1, IL-18, GSDMD protein and mRNA increased in shCD147-1 and shCD147-2 group（P < 0.05）. Lactate dehydrogenase（LDH） release assay showed that compared with the SiHa group, the shCD147 group had a significant increase in LDH release（P < 0.05）. Fluorescence inverted microscope showed that the shCD147 group had swelling and vacuolization, showing typical pyroptosis. Compared with the SiHa group, the shCD147-1 and shCD147-2 groups had significantly reduced the cell proliferation and colony formation ability（P < 0.05）. Conclusion Low expression of CD147 effectively up-regulates the expression of AIM2 inflammation-related factors in cervical cancer SiHa cells, induces the pyroptosis, and inhibits the cell proliferation and cloning.
- Cervical cancer /
- CD147 /
- AIM2inflammasome /
- Pyroptosis /
- Proliferation

HTML全文

图 1 检测 CD147在宫颈癌细胞中的表达水平及慢病毒转染效率

A：CD147蛋白在宫颈癌及正常宫颈上皮细胞中的相对表达量；B：转染慢病毒后CD147的 mRNA 的表达情况；C：转染慢病毒后CD147蛋白的相对表达量；D：慢病毒转染CD147细胞后绿色荧光表达情况 ( 标尺为100 μm)；ns：差异无统计学意义；^****P < 0.0001。

Figure 1. The expression level of CD147 in cervical cancer cells and the transfection efficiency of lentivirus were detected

下载: 全尺寸图片幻灯片

图 2 下调CD147 对 SiHa 细胞AIM2炎症小体信号通路的影响

A：下调 CD147后 AIM2、Caspase-1、IL-18、GSDMD 的 mRNA 的表达情况；B：下调 CD147后 AIM2、Caspase-1、IL-18、GSDMD 蛋白的相对表达量；ns：无统计学差异；^***P < 0.001；^****P < 0.0001。

Figure 2. Effect of down-regulating CD147 on AIM2 inflammasome signaling pathway in SiHa cells

下载: 全尺寸图片幻灯片

图 3 下调CD147对宫颈癌SiHa细胞焦亡的影响

A：荧光倒置显微镜观测下调 CD147对SiHa 细胞形态影响（标尺为100 μm）；B：下调 CD147对SiHa 细胞释放乳酸脱氢酶水平的影响；^***P < 0.001。

Figure 3. Effect of down-regulation of CD147 on pyroptosis in cervical cancer SiHa cells

下载: 全尺寸图片幻灯片

图 4 下调 CD147 后宫颈癌 SiHa 细胞增殖克隆能力情况

A：下调 CD147对SiHa 细胞增殖影响；B：下调 CD147对SiHa 细胞集落形成情况；^***P < 0.001；^****P < 0.0001。

Figure 4. The proliferation and cloning ability of cervical cancer SiHa cells after down-regulation of CD147

下载: 全尺寸图片幻灯片

表 1 RT-qPCR检测基因的引物序列

Table 1. Primer sequences of genes detected by RT-qPCR

基因	引物序列
CD147	F-TGTTCGTGCTGCTGGGATTCG R-GAGCCAAGGTCTTCTACGGTAGTG
AIM2	F-TATCGGCACAGTGGTTTCTTAGAGG R-GGGCTGAGTTTGAAGCGTGTTG
Caspase-1	F-CCCACATCCTCAGGCTCAGAAG R-TGCGGCTTGACTTGTCCATTATTG
IL-18	F-TGGCTGCTGAACCAGTAGAAGAC R-GAGGCCGATTTCCTTGGTCAATG
GSDMD	F-CCAGAAGAAGACGGTCACCATCC R-TGGAACGCTTGTGGCCTGTC
GAPDH	F-TGCACCACCAACTGCTTAGC R-GGCATGGACTGTGGTCATGAG

下载: 导出CSV

参考文献(21)

[1]	Sung H,Ferlay J,Siegel R L,et al. Global cancer statistics 2020: Globocan estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin,2021,71(3):209-249. doi: 10.3322/caac.21660
[2]	Kendrick A A,Schafer J,Dzieciatkowska M,et al. Cd147: A small molecule transporter ancillary protein at the crossroad of multiple hallmarks of cancer and metabolic reprogramming[J]. Oncotarget,2017,8(4):6742-6762. doi: 10.18632/oncotarget.14272
[3]	Dana P,Kariya R,Vaeteewoottacharn K,et al. Upregulation of cd147 promotes metastasis of cholangiocarcinoma by modulating the epithelial-to-mesenchymal transitional process[J]. Oncol Res,2017,25(7):1047-1059. doi: 10.3727/096504016X14813899000565
[4]	Schroder K,Tschopp J. The inflammasomes[J]. Cell,2010,140(6):821-832. doi: 10.1016/j.cell.2010.01.040
[5]	Shi J,Gao W,Shao F. Pyroptosis: Gasdermin-mediated programmed necrotic cell death[J]. Trends Biochem Sci,2017,42(4):245-254. doi: 10.1016/j.tibs.2016.10.004
[6]	Rao Z,Zhu Y,Yang P,et al. Pyroptosis in inflammatory diseases and cancer[J]. Theranostics,2022,12(9):4310-4329. doi: 10.7150/thno.71086
[7]	Kumari P,Russo A J,Shivcharan S,et al. Aim2 in health and disease: Inflammasome and beyond[J]. Immunol Rev,2020,297(1):83-95. doi: 10.1111/imr.12903
[8]	Fidler T P,Xue C,Yalcinkaya M,et al. The aim2 inflammasome exacerbates atherosclerosis in clonal haematopoiesis[J]. Nature,2021,592(7853):296-301. doi: 10.1038/s41586-021-03341-5
[9]	郭文涛. Cd147调控srebps影响宫颈癌细胞脂肪酸合成及侵袭迁移能力的实验研究[D]. 乌鲁木齐: 新疆医科大学硕士学位论文, 2020.
[10]	Lee C L,Lam M P,Lam K K,et al. Identification of cd147 (basigin) as a mediator of trophoblast functions[J]. Hum Reprod,2013,28(11):2920-2929. doi: 10.1093/humrep/det355
[11]	Yang H,Chen B. Cd147 in ovarian and other cancers[J]. Int J Gynecol Cancer,2013,23(1):2-8. doi: 10.1097/IGC.0b013e3182749139
[12]	Poli G,Fabi C,Bellet M M,et al. Epigenetic mechanisms of inflammasome regulation[J]. Int J Mol Sci,2020,21(16):5758. doi: 10.3390/ijms21165758
[13]	Wang L,Sun L,Byrd K M,et al. Aim2 inflammasome's first decade of discovery: Focus on oral diseases[J]. Front Immunol,2020,11(8):1487.
[14]	Lee S,Karki R,Wang Y,et al. Aim2 forms a complex with pyrin and zbp1 to drive panoptosis and host defence[J]. Nature,2021,597(7876):415-419. doi: 10.1038/s41586-021-03875-8
[15]	Du T,Gao J,Li P,et al. Pyroptosis,metabolism,and tumor immune microenvironment[J]. Clin Transl Med,2021,11(8):e492. doi: 10.1002/ctm2.492
[16]	Yu P,Zhang X,Liu N,et al. Pyroptosis: Mechanisms and diseases[J]. Signal Transduct Target Ther,2021,6(1):128. doi: 10.1038/s41392-021-00507-5
[17]	Pushkarsky T,Zybarth G,Dubrovsky L,et al. Cd147 facilitates hiv-1 infection by interacting with virus-associated cyclophilin A[J]. Proc Natl Acad Sci USA,2001,98(11):6360-6365. doi: 10.1073/pnas.111583198
[18]	Majno G,Joris I. Apoptosis,oncosis,and necrosis. An overview of cell death[J]. Am J Pathol,1995,146(1):3-15.
[19]	Butera A,Quaranta M T,Crippa L,et al. Cd147 targeting by ac-73 induces autophagy and reduces intestinal fibrosis associated with tnbs chronic colitis[J]. J Crohns Colitis,2022,16(11):1751-1761. doi: 10.1093/ecco-jcc/jjac084
[20]	Kulyar M F,Yao W,Ding Y,et al. Cluster of differentiation 147 (cd147) expression is linked with thiram induced chondrocyte's apoptosis via bcl-2/bax/caspase-3 signalling in tibial growth plate under chlorogenic acid repercussion[J]. Ecotoxicol Environ Saf,2021,213(4):112059.
[21]	曹勋荣,吕桂雪,魏雯雯. Rna干扰抑制cd147表达对宫颈癌细胞增殖、转移和侵袭能力的影响[J]. 中国免疫学杂志,2023,39(2):349-353,358. doi: 10.3969/j.issn.1000-484X.2023.02.023

相关文章(20)

[1]	张晔琳, 马丽娅, 彭旭晖, 杨禾丰, 佘睿. hsa-let-7a-5p调控牙周膜干细胞增殖及凋亡, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20231028
[2]	王静, 米弘瑛, 张熠, 李丽, 余建华, 刘丽巧, 刘庆瑜, 王立伟. 细胞焦亡参与早期子鼠坏死性小肠结肠炎的发病, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20230124
[3]	张梁, 王保全, 雷喜锋, 王旭, 柯阳, 张玮. miR-29c-3p/IGF1分子轴对肝星状细胞活化，增殖和凋亡的作用机制, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20230926
[4]	海燕, 美力班·吐尔逊, 阿仙姑·哈斯木. Pin1通过调控脂质代谢关键酶影响宫颈癌细胞的增殖及凋亡, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20230107
[5]	王保全, 张伟, 田园, 雷喜锋, 王旭. miR-142-5p通过CCND1调控胆囊癌细胞的增殖和转移, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20220223
[6]	廖周俊, 杨少华, 刘立鑫, 胡晟, 陈轶晖, 康强, 张小文. AK4对肝内胆管癌细胞HUCCT1增殖、迁移的影响, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20220611
[7]	赵斌, 段元鹏, 张国颖, 毕城伟, 杨李波, 施致裕, 杨勇, 张建朋, 高婷. CircRNA EZH2通过调控miR-30c-5p促进前列腺癌细胞增殖和迁移, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20220731
[8]	张玮, 王保全, 雷喜锋, 王旭, 张梁. miR-125b-5p调控HK2抑制胆囊癌细胞增殖和糖酵解, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20221206
[9]	龙熙翠, 刘贝贝, 卢绍波, 李志红, 金文娇, 陆金芝, 韩雪松. 细胞焦亡因子Caspase-1、IL-1β与IL-18在子宫内膜息肉组织中的表达和意义, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20210917
[10]	刘洋, 廖婧, 卢又汇, 孙春意, 孟昱时. miR-21负向调控宫颈癌HeLa细胞株中hTERT的表达, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20210307
[11]	杨兰, 贾霄, 姜奕彤, 崔琪, 刘光赐, 何颖红. UBE2C基因沉默表达对人胃癌AGS细胞增殖和迁移的影响, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20210504
[12]	王小莹, 刘作金, 申丽娟. 缺血再灌注损伤与细胞焦亡的相关性研究进展, 昆明医科大学学报. doi: 10.12259/j.issn.2095-610X.S20201240
[13]	贾楠楠. Gnaq对SH-SY5Y细胞增殖的作用及机制, 昆明医科大学学报.
[14]	魏东. 脆性位点基因WWOX调控人胆囊癌细胞的体外增殖效应, 昆明医科大学学报.
[15]	朱胜章. 宫颈液基细胞学检查在贵州黔南地区宫颈癌筛查中的临床运用, 昆明医科大学学报.
[16]	杨正浩. 细胞DNA定量分析法和液基薄层细胞学检测技术在宫颈癌早期诊断中的应用对比, 昆明医科大学学报.
[17]	王海峰. 上调microRNA-101沉默EZH2基因表达对人膀胱癌T24细胞系增殖和凋亡的影响, 昆明医科大学学报.
[18]	武斌. 大豆苷元对人乳牙牙髓干细胞增殖和成骨分化的影响, 昆明医科大学学报.
[19]	夏英杰. SD大鼠髁状突颈部骨折对大鼠髁状突软骨细胞增殖与凋亡的影响, 昆明医科大学学报.
[20]	李雄. 氧自由基在血管紧张素Ⅱ诱导ECV304细胞增殖中的作用, 昆明医科大学学报.