基于转录组学探讨LF-UTMD对PTC细胞的影响  

Exploring the impact of LF-UTMD on PTC cells based on transcriptome analysis

作　　者：周祖邦[1] 牛彦强 张佳琰 仇菊梅 慈霞 谢金会[1] ZHOU Zu-bang;NIU Yan-qiang;ZHANG Jia-yan;QIU Ju-mei;CI Xia;XIE Jin-hui(Department of Ultrasound Medicine,Gansu Provincial Hospital,Lanzhou 730000,China;First School of Clinical Medical,Gansu University of Chinese Medicine,Lanzhou 730000,China;Department of Geriatrics,Gansu Provincial Hospital,Lanzhou 730000,China)

机构地区：[1]甘肃省人民医院超声医学科,兰州730000 [2]甘肃中医药大学第一临床医学院,兰州730000 [3]甘肃省人民医院老年医学科,兰州730000

出　　处：《兰州大学学报(自然科学版)》2025年第2期246-256,264,共12页Journal of Lanzhou University(Natural Sciences)

基　　金：甘肃省科技厅重点研发计划项目(22YF7FA098);甘肃省自然科学基金项目(24JRRA606);甘肃省人民医院国家自然科学基金培育项目(24GSSYA-3);甘肃省人民医院院内基金(2024KYQDJ-A-37)。

摘　　要：为了探讨低频超声靶向微泡破坏技术(LF-UTMD)对甲状腺乳头状癌(PTC)细胞的影响,揭示其潜在的治疗机制,并寻找可能的治疗靶点分子,使用人类乳头状甲状腺癌细胞系BCPAP细胞进行实验,正交试验筛选最佳超声参数和微泡体积分数φ(微泡)联合干预条件,六氟化硫微泡和超声对细胞分别单独或联合干预;细胞计数试剂盒检测细胞增殖;通过Oligo(dT)磁珠富集mRNA,构建cDNA文库,进行高通量测序;对原始测序数据进行质量控制、过滤、比对、表达量计算和差异基因筛选;进行基因本体(GO)、京都基因与基因组百科全书(KEGG)和基因集富集分析(GSEA),以及核心基因的蛋白质互作网络(PPI)分析;对各类凋亡相关差异基因进行分析,通过实时荧光定量聚合酶链式反应验证各类凋亡差异基因的表达.结果表明,抑制PTC细胞增殖的最佳组合为:机械指数1.2、辐照时间90 s、φ(微泡)=10%、占空比为20%、脉冲重复频率为1 kHz;主成分分析显示样本重复性好,且对照组和超声联合微泡(U-M)组之间存在显著性差异.与对照组相比,U-M组中显著变化的基因有1 552个,其中上调基因937个,下调基因615个.GO和KEGG分析揭示与免疫反应、代谢过程、细胞增殖和分化、信号传导等相关的差异基因;GSEA分析表明U-M组治疗可能通过增强免疫反应、调节代谢过程、影响细胞增殖和分化、调节信号传导、基因沉默和RNA代谢、干扰细胞周期和DNA修复以及促进细胞死亡影响PTC细胞.PPI互作分析发现STAT4、 IRF7、 CXCL10、 FOS、 TNF等为核心基因.U-M组中,双硫死亡标志物SLC7A11显著升高,坏死性凋亡标志物Ripk1降低而Ripk4升高,TLR1、 TLR2、 TLR5升高,Rnf31降低.铜死亡相关基因未显著变化,铁死亡相关基因CPEB1、Hmox1、 Atf3、 Aloxe3、 PTGS2升高.焦亡相关基因Caspase1、 Gbp5、 Aim2降低,GSDMA升高,其他焦亡相关基因无变化.LF-UTMD能够显著影响PTC细胞的基因Aimed to explore the effects of low-frequency ultrasound-targeted microbubble destruction(LFUTMD) on papillary thyroid carcinoma(PTC) cells through transcriptome analysis,to reveal potential therapeutic mechanisms and to identify possible therapeutic target molecules.BCPAP cells were used for the experiments;orthogonal experiments were conducted to select the optimal conditions for the combined intervention of ultrasound and microbubbles,with sulfur hexafluoride microbubbles and ultrasound applied to cells either individually or in combination;cell counting kit-8 was used to detect cell proliferation;mRNA was enriched using Oligo(dT) magnetic beads,cDNA libraries were constructed and highthroughput sequencing performed;raw sequencing data were subjected to quality control,filtering,alignment,expression quantification,and differential gene screening;gene ontology(GO),Kyoto encyclopedia of genes and genomes(KEGG) and gene set enrichment analysis(GSEA) enrichment analyses were conducted,along with protein-protein interaction networks(PPI) interaction analysis of core genes;further analysis was performed on various apoptosis-related differential genes,and the expression of these genes was validated by quantitative real-time polymerase chain reaction.The results were that the optimal combination for inhibiting PTC cell proliferation was a 1.2 mechanical index,90 s irradiation time,volume fraction 10% microbubble,20% duty cycle and 1 kHz pulse repetition frequency;further transcriptome analysis revealed that principal component analysis showed good sample repeatability and significant differences between the control and the ultrasound-targeted microbubble(U-M) group.Compared to the control,there were 1 552 significantly changed genes in the U-M group,with 937 upregulated and 615 downregulated genes.GO and KEGG analyses revealed differential genes related to immune response,metabolic processes,cell proliferation and differentiation,and signal transduction;GSEA analysis indicated that U-M group treatment might affect PTC

关 键 词：甲状腺乳头状癌 低频超声靶向微泡破坏技术 转录组 死亡方式 富集分析 

分 类 号：R736.1[医药卫生—肿瘤]