乳杆菌A-2代谢产物LSPM1通过JAK/STAT3通路调节人舌鳞状细胞癌细胞增殖和凋亡  

Lactobacillus A-2 Metabolite LSPM1 Regulates Proliferation and Apoptosis of Human Tongue Squamous Cell Carcinoma Cells through JAK/STAT3 Pathway

作　　者：孙智辉 徐仲秋 董丽 毕闯 王惠琳 韩新文 李志帅 李丽 SUN Zhi-hui;XU Zhong-qiu;DONG Li;BI Chuang;WANG Hui-lin;HAN Xin-wen;LI Zhi-shuai;LI Li(Department of Stomatology,the Affiliated Hospital of Qingdao Binhai University(Qingdao Military Civilian Integration Hospital),Qingdao 266400,China;Department of Pathology,the Affiliated Hospital of Qingdao Binhai University(Qingdao Military Civilian Integration Hospital),Qingdao 266400,China;Department of Neurosurgery,the Affiliated Hospital of Qingdao Binhai University(Qingdao Military Civilian Integration Hospital),Qingdao 266400,China;Department of Rehabilitation,the Affiliated Hospital of Qingdao Binhai University(Qingdao Military Civilian Integration Hospital),Qingdao 266400,China)

机构地区：[1]青岛滨海学院附属医院(青岛军民融合医院)口腔科,青岛266400 [2]青岛滨海学院附属医院(青岛军民融合医院)病理科,青岛266400 [3]青岛滨海学院附属医院(青岛军民融合医院)神经外科,青岛266400 [4]青岛滨海学院附属医院(青岛军民融合医院)康复科,青岛266400

出　　处：《南昌大学学报（医学版）》2022年第5期14-18,37,共6页Journal of Nanchang University：Medical Sciences

基　　金：山东省医药卫生科技发展计划项目(202008021206);山东省高等学校青创人才引育计划康复治疗学研究创新团队项目。

摘　　要：目的探讨乳杆菌A-2代谢产物LSPM1对人舌鳞状细胞癌(鳞癌)细胞Cal-27的生物学功能影响和机制。方法制备LSPM1和培养人舌鳞癌细胞Cal-27,分组实验:Blank组(空白对照)、LSPM1组(加入LSPM1冻干粉稀释液,终浓度30 mg·mL^(-1))、AG-490组(加入JAK/STAT3信号通路抑制剂AG-490)和LSPM1+AG-490组(同时加入LSPM1和AG-490)。采用MTT法、细胞划痕实验和流式细胞术测定细胞增殖、迁移和凋亡,采用JAK/STAT3信号通路抑制剂AG-490观察细胞生物学功能,Western Blot方法检测JAK/STAT3信号通路关键蛋白JAK2和STAT3的表达和磷酸化水平。结果72 h内LSPM1组细胞增殖率(0.81±0.03)明显低于Blank组(0.93±0.03)(P<0.01)。加入AG-490后,LSPM1对细胞增殖的抑制作用减弱;72 h内AG-490组和LSPM1+AG-490组细胞增殖率与Blank组相比差异无统计学意义(P>0.05),即30 mg·mL^(-1) LSPM1对Cal-27细胞的增殖具有明显抑制作用;LSPM1组72 h内细胞划痕面积缩小2.3×10^(4)个单位,Blank组划痕面积缩小4.2×10^(4)个单位,2组比较差异有统计学意义(P<0.01),LSPM1抑制人舌鳞癌细胞Cal-27迁移;Blank组细胞凋亡率为0.11±0.01,LSPM1组细胞凋亡率为0.21±0.01,2组比较差异有统计学意义(P<0.05),LSPM1促进人舌鳞癌细胞Cal-27细胞凋亡。Western blot实验中Blank组、LSPM1组、AG-490组和LSPM1+AG-490组JAK蛋白相对表达量分别为1.00、1.62、0.20、0.60;p-JAK蛋白相对表达量分别为1.00、1.66、0.30、0.60;STAT3蛋白相对表达量分别为1.00、1.22、0.09、0.12;p-STAT3蛋白相对表达量分别为1.00、1.17、0.07、0.10;加入AG-490抑制剂后,LSPM1+AG-490组STAT3、JAK2蛋白表达及磷酸化水平较LSPM1组降低,差异有统计学意义(P<0.05)。结论乳杆菌A-2代谢产物LSPM1通过JAK2/STAT3信号抑制人舌鳞癌细胞系Cal-27增殖和迁移,促进细胞凋亡。Objective To investigate the effect and mechanism of lactobacillus A-2 metabolite LSPM1 on biological function of human tongue squamous cell carcinoma cell line Cal-27.Methods LSPM1 was prepared and human tongue squamous cell carcinoma Cal-27 cells were cultured.The cells were divided into blank group(blank control),LSPM1 group(treatment with lyophilized LSPM1 powder diluent at the final concentration of 30 mg·mL^(-1)),AG-490 group(treatment with JAK/STAT3 signal pathway inhibitor AG-490),and LSPM1+AG-490 group(treatment with combined LSPM1 and AG-490).MTT assay,scratch test and flow cytometry were performed to determine cell proliferation,migration and apoptosis,respectively.JAK/STAT3 signal pathway inhibitor AG-490 was used to observe the biological function of cells.The expression and phosphorylation levels of JAK2 and STAT3 were detected by Western blot.Results Treatment with 30 mg·mL^(-1) LSPM1 significantly inhibited proliferation and migration and promoted apoptosis in CAL-27 cells.After 72 hours of treatment,the cell proliferation rate in LSPM1 group(0.81±0.03)was lower than that in blank group(0.93±0.03)(P<0.01).However,the inhibitory effect of LSPM1 on cell proliferation was weakened by supplementation with AG-490.There was no difference in the proliferation rate between AG-490 group and blank group(P>0.05).The cell scratch area in LSPM1 and blank groups respectively decreased by(2.3×10^(4))and(4.2×10^(4))units with in 72 hours,and the differencewas significant between the two groups(P<0.01).The 36 hours apoptosis rate in blank group(0.11±0.01)was lower than that in LSPM1 group(0.21±0.01)(P<0.05).In blank group,LSPM1 group,AG-490 group and LSPM1+AG-490 group,respectively,the relative expression levels were 1.00,1.62,0.20 and 0.60 for JAK protein,1.00,1.66,0.30 and 0.60 for p-JAK protein,1.00,1.22,0.09 and 0.12 for STAT3 protein,and 1.00,1.17,0.07 and 0.10 for p-STAT3.Furthermore,the expression and phosphorylation levels of STAT3 and JAK2 in LSPM1+AG-490 group were lower than those in LSPM1 group

关 键 词：人舌鳞状细胞癌 Cal-27 乳杆菌A-2 LSPM1 JAK2/STAT3 

分 类 号：R739.85[医药卫生—肿瘤]