不同型別幽门螺杆菌感染血清微RNA表达水平及表达特征研究

doi:10.3877/cma.j.issn.2095-5820.2023.01.006

目的

探讨不同型别的幽门螺杆菌（Hp）感染后是否导致特定微RNA（miRNA）表达的改变。

方法

从135例研究对象中依据Hp感染情况分为无Hp感染组、Ⅰ型（毒力型）Hp感染组和Ⅱ型（非毒力型）Hp感染组各40例（其余15例未明确分组，剔除数据）。采集血清，利用荧光定量PCR（Rt-PCR）方法检测miR-133a和miR-130a的表达水平，研究不同型别Hp感染者血清中miRNA的表达水平和特征。

结果

不同型别的Hp感染者血清miR-133a和miR-130a表达水平不同，与无Hp感染组和Ⅱ型Hp感染组相比，Ⅰ型Hp感染组血清中的miR-133a和miR-130a表达水平发生了特征性的改变，miR-133a表达水平低至原来的25%，miR-130a表达水平高达9.51倍。

结论

不同型别Hp感染可能同血清特定miRNA miR-133a和miR-130a的表达水平相关，Ⅰ型Hp感染与致细胞恶性变相关，提示不同型别Hp感染可能具有不同的致细胞恶性变作用。

关键词: 幽门螺杆菌, 分型, 微RNA, 荧光定量PCR

Objective

Helicobacter pylori (Hp) is an important pathogen of gastrointestinal diseases. Depending on whether Hp produces cytotoxin-associated protein A (CagA) and vacuolating toxin (VacA), Hp can be divided into type Ⅰ (virulence type) and Type Ⅱ (non-virulent type).

Methods

According to the Hp infection situation, 135 subjects were divided into non-Hp infection group, type Ⅰ Hp infection group and type Ⅱ Hp infection group with 40 cases each. Serum was collected, and the levels of miR-133a and miR-130a were analyzed by fluorescent quantitative PCR, so as to study the expression level and characteristics of miRNA in the serum of patients with different types of Hp infection.

Results

The expression levels of miR-133a and miR-130a in the serum of patients with different types of Hp infection were different. Compared with non-Hp infection group and type Ⅱ infection group, the expression levels of miR-133a and miR-130a in the serum of typeⅠ Hp infection group were characteristively changed. The expression level of miR-133a was as low as 0.25 times and the expression level of miR-130a was as high as 9.51 times.

Conclusions

Different types of Hp infection may be related to the expression levels of specific miRNAs miR-133a and miR-130a in serum, reflecting that type I Hp is related to cell malignancy, which suggesting that different types of Hp may have different effects on cell malignancy.

Key words: helicobacter pylori, typing, miRNA, real-time PCR

表1 Hp检测结果/例

C¹³呼气试验阳性分型	C¹³呼气试验
C¹³呼气试验阳性分型	阳性	阴性	合计
Ⅰ型	40	2	42
Ⅱ型	40	2	42
其他	11	40	51
合计	91	44	135

表1 Hp检测结果/例

C¹³呼气试验阳性分型	C¹³呼气试验
C¹³呼气试验阳性分型	阳性	阴性	合计
Ⅰ型	40	2	42
Ⅱ型	40	2	42
其他	11	40	51
合计	91	44	135

表2 miRNA列表及序列和扩增试剂盒

miRNA	序列	引物及试剂盒
has-miR-133a-3p	UUUGGUCCCCUUCAACCAGCUG	has-miR-133a-3p miRCURY LNA PCR Assay
has-miR-130a-3p	CAGUGCAAUGUUAAAAGGGCAU	has-miR-130a-3p miRCURY LNA PCR Assay

表2 miRNA列表及序列和扩增试剂盒

miRNA	序列	引物及试剂盒
has-miR-133a-3p	UUUGGUCCCCUUCAACCAGCUG	has-miR-133a-3p miRCURY LNA PCR Assay
has-miR-130a-3p	CAGUGCAAUGUUAAAAGGGCAU	has-miR-130a-3p miRCURY LNA PCR Assay

表3 miR-133a在GES-1细胞中的表达水平

项目	样本量	miR-133a C_t	GAPDH C_t	ΔC_t	ΔΔC_t	以Hp阴性感染血清为参考，其它血清样本中miR-133a表达量比值
Ⅰ型Hp表达均值	40	30.65±0.20	24.48±0.21	6.17±0.33	1.98±0.33	0.25（0.20~0.35）
Ⅱ型Hp表达均值	40	28.37±0.15	24.11±0.12	4.26±0.16	0.07±0.16	0.95（0.85~1.05）
正常对照表达均值	40	27.14±0.08	22.95±0.07	4.19±0.05	0.00±0.04	1.00（0.90~1.10）

表3 miR-133a在GES-1细胞中的表达水平

项目	样本量	miR-133a C_t	GAPDH C_t	ΔC_t	ΔΔC_t	以Hp阴性感染血清为参考，其它血清样本中miR-133a表达量比值
Ⅰ型Hp表达均值	40	30.65±0.20	24.48±0.21	6.17±0.33	1.98±0.33	0.25（0.20~0.35）
Ⅱ型Hp表达均值	40	28.37±0.15	24.11±0.12	4.26±0.16	0.07±0.16	0.95（0.85~1.05）
正常对照表达均值	40	27.14±0.08	22.95±0.07	4.19±0.05	0.00±0.04	1.00（0.90~1.10）

表4 miR-130a在不同血清中的表达水平

项目	样本量	miR-130a C_t	GAPDH C_t	ΔC_t	ΔΔC_t	以Hp阴性感染血清为参考，其它血清样本中miR-130a表达量比值
I型Hp表达均值	40	31.20±0.09	25.26±0.10	6.02±0.06	-3.25±0.06	9.51（9.00~9.90）
II型Hp表达均值	40	33.70±0.10	24.00±0.08	9.30±0.06	0.03±0.06	0.98（0.94~1.02）
正常对照表达均值	40	34.20±0.13	25.16±0.10	9.20±0.10	0.00±0.10	1.00（0.90~1.10）

表4 miR-130a在不同血清中的表达水平

项目	样本量	miR-130a C_t	GAPDH C_t	ΔC_t	ΔΔC_t	以Hp阴性感染血清为参考，其它血清样本中miR-130a表达量比值
I型Hp表达均值	40	31.20±0.09	25.26±0.10	6.02±0.06	-3.25±0.06	9.51（9.00~9.90）
II型Hp表达均值	40	33.70±0.10	24.00±0.08	9.30±0.06	0.03±0.06	0.98（0.94~1.02）
正常对照表达均值	40	34.20±0.13	25.16±0.10	9.20±0.10	0.00±0.10	1.00（0.90~1.10）

1	CAMILO V, SUGIYAMA T, TOUATI E. Pathogenesis of Helicobacter pylori infection[J]. Helicobacter, 2017, 22(S1).
2	ANSARI S, YAMAOKA Y. Role of vacuolating cytotoxin A in Helicobacter pylori infection and its impact on gastric pathogenesis[J]. Expert Review of Anti-infective Therapy, 2020, 18(10): 987-996.
3	WANG Y C. Medicinal plant activity on Helicobacter pylori related diseases[J]. World Journal of Gastroenterology, 2014, 20(30):10368-10382.
4	MUZAHEED. Helicobacter pylori Oncogenicity: Mechanism, Prevention, and Risk Factors[J]. The Scientific World Journal, 2020, 2020: 3018326.
5	KROL J, LOEDIGE I, FILIPOWICZ W. The widespread regulation of microRNA biogenesis, function and decay[J]. Nature Reviews Genetics, 2010, 11: 597-610.
6	RACHAGANI S, KUMAR S, BATRA S K. MicroRNA in pancreatic cancer: pathological, diagnostic and therapeutic implications[J]. Cancer Letters, 2010, 292: 8-16.
7	TIE Y, LIU B, FU H, et al. Circulating miRNA and cancer diagnosis[J]. Science in China. Series C, Life Sciences, 2009, 52:1117-1122.
8	CHEN X, BA Y, MA L, et al. Characterization of microRNAs in serum: a novel class of biomarkers for diagnosis of cancer and other diseases[J]. Cell Research, 2008, 18: 997-1006.
9	MITCHELL P S, PARKIN R K, KROH E M, et al. Circulating microRNAs as stable blood-based markers for cancer detection[J]. Proceedings of the National Academy of Sciences of the United States of America, 2008, 105: 10513-10518.
10	ZHANG Z, LI Z, GAO C, et al. miR-21 plays a pivotal role in gastric cancer pathogenesis and progression[J]. Laboratory Investigation; A Journal of Technical Methods and Pathology, 2008, 88: 1358-1366.
11	MOTOYAMA K, INOUE H, MIMORI K, et al. Clinicopathological and prognostic significance of PDCD4 and microRNA-21 in human gastric cancer[J]. International Journal of Oncology, 2010, 36: 1089-1095.
12	LAI K W, KOH K X, LOH M, et al. MicroRNA-130b regulates the tumour suppressor RUNX3 in gastric cancer[J]. European Journal of Cancer (Oxford, England: 1990), 2010, 46:1456-1463.
13	WU Q, JIN H, YANG Z, et al. MiR-150 promotes gastric cancer proliferation by negatively regulating the pro-apoptotic gene EGR2[J]. Biochemical and Biophysical Research Communications, 2010, 392:340-345.
14	JIANG Z, GUO J, XIAO B, et al. Increased expression of miR-421 in human gastric carcinoma and its clinical association[J]. Journal of Gastroenterology, 2010, 45: 17-23.
15	LIVAK K J, SCHMITTGEN T D. Analysis of relative gene expression data using real-time quantitative PCR and the 2^-ΔΔCt method[J]. Methods, 2001, 25(4): 402-408.
16	TOMOMITSU T, SAYUMI T, TETSUYA T, et al. Telomere length in the gastric mucosa after Helicobacter pylori eradication and its potential role in the gastric carcinogenesis[J]. Clinical and Experimental Medicine, 2018, 18(1): 21-26.
17	LI S, ZENG J P, LI S Y, et al. Helicobacter pylori Induces a Novel NF-kB/LIN28A/let-7a/hTERT Axis to Promote Gastric Carcinogenesis[J]. Molecular Cancer Research: MCR, 2021, 19 (1): 74-85.
18	KANG S S, JANG H J, KIM J H, et al. The relationship between telomerase activation and helicobacter pylori Infection in Gastric Cancer[J]. The Korean Journal of Gastroenterology, 2002, 63(6) : 634-642.
19	CHEN C, RIDZON D A, BROOMER A J, et al. Real-time quantification of microRNAs by stem-loop RT-PCR[J]. Nucleic Acids Research, 2005, 33: e179.
20	SAITO Y, SUZUKI H, TSUGAWA H, et al. Dysfunctional gastric emptying withdown-regulation of muscle-specific microRNAs in Helicobacter pylori-infected mice[J]. Gastroenterology, 2011, 140(1): 189-198.
21	HE M Q, WAN J F, ZENG H F, et al. MiR-133a-5p suppresses gastric cancer through TCF4 down-regulation[J]. Journal of Gastrointestinal Oncology, 2021, 12(3): 1007-1019.
22	CHENG Z, LIU F, WANG G, et al. MiR-133 is a key negative regulator of CDC42-PAK pathway in gastric cancer[J]. Cell Signal, 2014, 26(12): 2667-2673.
23	DUAN J, ZHANG H, QU Y et al. Onco-miR-130 promotes cell proliferation and migration by targeting TGFβR2 in gastric cancer[J]. Oncotarget, 2016, 7(28): 44522-44533.

[1]	刘伟, 牛云峰, 安杰. LINC01232 通过miR-516a-5p/BCL9 轴促进三阴性乳腺癌的恶性进展[J/OL]. 中华乳腺病杂志(电子版), 2024, 18(06): 330-338.
[2]	于溟璇, 杜华, 张彩虹, 师迎旭. miRNA-192家族在乳腺癌中的作用机制及诊断价值[J/OL]. 中华乳腺病杂志(电子版), 2024, 18(04): 235-240.
[3]	魏艺, 周羽西, 杨烨, 凌秀凤, 赵纯. 微小RNA对子宫内膜容受性影响的研究现状[J/OL]. 中华妇幼临床医学杂志(电子版), 2024, 20(03): 266-270.
[4]	崔玉峰, 林毅军, 王志民. 幽门螺杆菌分子检测技术研究进展[J/OL]. 中华实验和临床感染病杂志(电子版), 2024, 18(05): 257-262.
[5]	郭伟仪, 林沛玲. 不同抗体型幽门螺杆菌感染与溃疡性结肠炎患者疾病活动及组织学评分的关系[J/OL]. 中华实验和临床感染病杂志(电子版), 2024, 18(04): 237-244.
[6]	张升荣, 李晓东, 王伟. B族链球菌感染分型对孕产妇未足月胎膜早破的预测价值及其与绒毛膜羊膜炎的关系[J/OL]. 中华实验和临床感染病杂志(电子版), 2024, 18(03): 170-175.
[7]	王欢欢, 郑少祥, 郝金锦, 陈文亮. 胃癌分子分型的研究进展及相关联系[J/OL]. 中华普通外科学文献(电子版), 2024, 18(03): 229-234.
[8]	赵国栋, 陆锦俊, 许永强. 不同肠系膜下动脉分型的腹腔镜低位直肠前切除术临床效果及经验总结[J/OL]. 中华普外科手术学杂志(电子版), 2024, 18(03): 334-337.
[9]	杨莎莎, 张毛为, 孙宜田, 刘亚南, 位娟, 魏建, 陈碧. 结缔组织疾病相关间质性肺病并发小气道功能障碍临床分析[J/OL]. 中华肺部疾病杂志(电子版), 2024, 17(05): 738-743.
[10]	蒋永东, 耿启砥, 吴宇峰, 高世华. 肩锁关节脱位的文献计量学分析[J/OL]. 中华肩肘外科电子杂志, 2024, 12(03): 246-256.
[11]	张峻, 赵建民, 姚晓克, 吉浩宇, 越瑞祥. 增加CT对桡骨远端骨折分型的可靠性及可重复性评价[J/OL]. 中华老年骨科与康复电子杂志, 2024, 10(05): 264-272.
[12]	周锐, 罗飞. 骨质疏松椎体骨折的分型进展[J/OL]. 中华老年骨科与康复电子杂志, 2024, 10(05): 315-320.
[13]	杨卫东, 周威, 向洪涛. 慢性萎缩性胃炎患者幽门螺杆菌感染与炎性细胞因子及病理特征的关系[J/OL]. 中华消化病与影像杂志(电子版), 2024, 14(05): 459-464.
[14]	李金星, 陈志宏, 孙铭均, 孙岩岩, 党树伟, 李广恩, 彭书奇, 付大伟, 李宏伟, 李国东. 基于多层螺旋CT 的腹腔干解剖变异与新分型研究[J/OL]. 中华临床医师杂志(电子版), 2024, 18(08): 756-763.
[15]	何妍, 檀丽冰, 周雯, 魏敏, 连军松. 老年男性人群幽门螺杆菌感染与血脂谱的关联研究[J/OL]. 中华胃肠内镜电子杂志, 2024, 11(03): 171-175.

阅读次数

全文

摘要

选择文件类型/文献管理软件名称

选择包含的内容

Study on the expression levels and characteristics of miRNA in different types of Helicobacter pylori infection

模态框（Modal）标题

选择文件类型/文献管理软件名称

选择包含的内容

Study on the expression levels and characteristics of miRNA in different types of Helicobacter pylori infection