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）

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

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