国际口腔医学杂志

国际口腔医学杂志 ›› 2024, Vol. 51 ›› Issue (1): 52-59.doi: 10.7518/gjkq.2024013

• 论著 • 上一篇    下一篇

长链非编码RNA小核仁RNA宿主基因22调控微小RNA-27b-3p对口腔鳞状细胞癌细胞增殖、侵袭和迁移的影响

周金阔1(),张晋弘1,史晓晶2,刘广顺2,姜磊3,刘倩峰1()   

  1. 1.河北医科大学第一医院口腔二科 石家庄 050000
    2.河北医科大学第一医院口腔一科 石家庄 050000
    3.河北医科大学第一医院中心实验室 石家庄 050000
  • 收稿日期:2023-04-27 修回日期:2023-08-21 出版日期:2024-01-01 发布日期:2024-01-10
  • 通讯作者: 刘倩峰
  • 作者简介:周金阔，主治医师，硕士，Email：jcwrni@163.com
  • 基金资助:
    河北省2022年度医科科学研究课题(20221387)

Influences of long noncoding RNA small nucleolar RNA host gene 22 on the cell proliferation, invasion and migration of oral squamous carcinoma cells by regulating microRNA-27b-3p

Zhou Jinkuo1(),Zhang Jinhong1,Shi Xiaojing2,Liu Guangshun2,Jiang Lei3,Liu Qianfeng1()   

  1. 1.Dept. of Stomatology Ⅱ, the First Hospital of Hebei Medical University, Shijiazhuang 050000, China
    2.Dept. of Stomatology Ⅰ, the First Hospital of Hebei Medical University, Shijiazhuang 050000, China
    3.Central Laboratory, the First Hospital of Hebei Medical University, Shijiazhuang 050000, China
  • Received:2023-04-27 Revised:2023-08-21 Online:2024-01-01 Published:2024-01-10
  • Contact: Qianfeng Liu
  • Supported by:
    Hebei Province 2022 Annual Medical Scien-ce Research Project(20221387)

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摘要：

目的 探究长链非编码RNA（LncRNA）小核仁RNA宿主基因（SNHG）22调控微小RNA（miR）-27b-3p对口腔鳞状细胞癌（OSCC）细胞增殖、侵袭和迁移的影响。 方法 收集52例OSCC患者的癌组织及癌旁组织标本，体外培养人正常口腔角质细胞HOK和3种人OSCC细胞（CAL-27、SCC-25和HSC-3），实时荧光定量聚合酶链反应（qRT-PCR）检测癌组织、癌旁组织、HOK细胞和3种OSCC细胞中SNHG22、miR-27b-3p表达情况。对SCC-25细胞进行转染并将其分为Ctrl组（未进行转染）、si-SNHG22组、si-NC组、miR-27b-3p inhibitor组、inhibitor-NC组、si-SNHG22+inhibitor-NC组和si-SNHG22+miR-27b-3p inhibitor组，检测各组SCC-25细胞增殖情况［细胞计数试剂盒8（CCK-8）法检测增殖率、流式细胞术检测增殖指数（PI）］；Transwell实验检测各组SCC-25细胞侵袭情况；划痕愈合实验检测各组SCC-25细胞迁移情况；双荧光素酶实验验证SNHG22与miR-27b-3p的靶向作用关系。 结果 与癌旁组织比较，OSCC癌组织中SNHG22表达显著升高，miR-27b-3p表达显著降低（P<0.05）；与HOK细胞比较，CAL-27、SCC-25和HSC-3细胞中SNHG22表达显著升高，miR-27b-3p表达显著降低，且SCC-25细胞中SNHG22与miR-27b-3p的表达与HOK细胞差异最大（P<0.05）。与Ctrl组比较，si-SNHG22组SCC-25细胞增殖率、PI、侵袭数和划痕面积愈合率均显著减少（P<0.05），miR-27b-3p inhibitor组SCC-25细胞增殖率、PI、侵袭数和划痕面积愈合率均显著增加（P<0.05）；与si-SNHG22组比较，si-SNHG22+miR-27b-3p inhibi-tor组SCC-25细胞增殖率、PI、侵袭数和划痕面积愈合率均显著增加（P<0.05）。双荧光素酶实验显示SNHG22与miR-27b-3p存在靶向作用关系。 结论 SNHG22在OSSC中高表达，miR-27b-3p在OSSC中低表达，SNHG22可能通过海绵化miR-27b-3p促进SCC-25细胞增殖、侵袭和迁移，SNHG22/miR-27b-3p轴可能是OSCC一个新的诊断和治疗靶点。

关键词: 长链非编码RNA小核仁RNA宿主基因22, 微小RNA-27b-3p, 口腔鳞状细胞癌, 增殖, 侵袭, 迁移

Abstract:

Objective This study aimed to investigate the influences of long noncoding RNA small nucleolar RNA host gene (SNHG) 22 on the proliferation, invasion, and migration of oral squamous cell carcinoma (OSCC) cells by regulating microRNA (miR)-27b-3p. Methods Cancer tissue and paracancerous tissue specimens of 52 OSCC patients were collected. Human normal oral keratinocytes (HOK) and three kinds of human OSCC cells (CAL-27, SCC-25, and HSC-3) were cultured in vitro. Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of SNHG22 and miR-27b-3p in cancer tissues, adjacent tissues, HOK cells, and three kinds of OSCC cells. SCC-25 cells were transfected and separated into the following groups: control (no transfection), si-SNHG22, si-NC, miR-27b-3p inhibitor, inhibitor-NC, si-SNHG22+inhibitor-NC, and si-SNHG22+miR-27b-3p inhibitor. The proliferation of SCC-25 cells in each group was detected by cell-counting kit 8 method, and proliferation index (PI) was detected by flow cytometry. Transwell assay was applied to detect the invasion of SCC-25 cells in each group. Scratch-area healing experiment was applied to detect the migration of SCC-25 cells in each group. Dual-luciferase experiment was applied to verify the targeting relationship between SNHG22 and miR-27b-3p. Results Compared with adjacent tissues, SNHG22 expression in OSCC cancer tissues significantly increased, and the expression of miR-27b-3p significantly decreased (P<0.05). Compared with HOK cells, SNHG22 expression significantly increased in CAL-27 cells, SCC-25 cells, and HSC-3 cells, and the expression of miR-27b-3p significantly decreased. The expression in SCC-25 cells differed the most from that in HOK cells (P<0.05). Compared with the control group, the SCC-25 cell-proliferation rate, PI, invasion number, and scratch-area healing rate in the si-SNHG22 group decreased significantly (P<0.05). The SCC-25 cell-proliferation rate, PI, invasion number, and scratch-area healing rate in the miR-27b-3p inhibitor group increased significantly (P<0.05); compared with the si-SNHG22 group. The SCC-25 cell-proliferation rate, PI, invasion number, and scratch-area healing rate in the si-SNHG22+miR-27b-3p inhibitor group increased significantly (P<0.05). Dual-luciferase experiments showed that SNHG22 had a targeting relationship with miR-27b-3p. Conclusion SNHG22 was highly expressed in OSSC, whereas miR-27b-3p was lowly expressed. SNHG22 may promote the proliferation, invasion, and migration of SCC-25 cells through sponge miR-27b-3p. The SNHG22/miR-27b-3p axis may be a new diagnostic and therapeutic target for OSCC.

Key words: long noncoding RNA small nucleolar RNA host gene 22, microRNA-27b-3p, oral squamous cell carcinoma, proliferation, invasion, migration

中图分类号: 

  • R782

表 1

qRT-PCR引物序列"

基因引物序列
SNHG22Forward 5’-GACTCAAGCCCTGAGCATGT-3’
Reverse 5’-CTGGTCACCTCCAACTGCAA-3’
miR-27b-3pForward 5’-CGCCTTGAATCGGTG-3’
Reverse 5’-GTGCAGGGTCCGAGGT-3’
GAPDHForward 5’-CTTTGGTATCGTGGAAGGACTC-3’
Reverse 5’-GTAGAGGCAGGGATGATGTTCT-3’
U6Forward 5’-GTGCTCGCTTCGGCAGCACAT-3’
Reverse 5’-TACCTTGCGAAGTGCTTAAAC-3’

图1

SNHG22与miR-27b-3p的结合位点"

图2

SNHG22和miR-27b-3p在组织中的表达情况A：OSCC组织中SNHG22表达水平；B：OSCC组织中miR-27b-3p表达水平；*与癌旁组织比较，P<0.05。"

图3

SNHG22、miR-27b-3p在细胞中的表达情况A：OSCC细胞系中SNHG22表达水平；B：OSCC细胞系中miR-27b-3p表达水平；*与HOK细胞比较，P<0.05。"

图4

成功抑制SCC-25细胞中SNHG22或miR-27b-3p表达A：各组SCC-25细胞中SNHG22表达水平；B：各组SCC-25细胞中miR-27b-3p表达水平。a：与Ctrl组比较，P<0.05；b：与si-NC组比较，P<0.05；c：与inhibitor-NC组比较，P<0.05；d：与si-SNHG22组比较，P<0.05；e：与si-SNHG22+inhibitor-NC组比较，P<0.05。"

图5

敲低SNHG22或miR-27b-3p表达对SCC-25细胞增殖、侵袭和迁移的影响A：各组SCC-25细胞增殖率比较；B：各组SCC-25细胞PI比较；C：各组SCC-25细胞侵袭数比较；D：各组SCC-25细胞划痕面积愈合率比较。a：与Ctrl组比较，P<0.05；b：与si-NC组比较，P<0.05；c：与inhibitor-NC组比较，P<0.05；d：与si-SNHG22组比较，P<0.05；e：与si-SNHG22+inhibitor-NC组比较，P<0.05。"

图 6

敲低SNHG22或miR-27b-3p表达对SCC-25细胞PI的影响"

图 7

敲低SNHG22或miR-27b-3p表达对SCC-25细胞侵袭的影响"

图 8

敲低SNHG22或miR-27b-3p表达对SCC-25细胞迁移的影响"

图9

SNHG22与miR-27b-3p靶向关系验证*表示与SNHG22-WT+mimic-NC组比较，P<0.05。"

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