lncRNA H19和IGF2基因在乳腺癌组织中的表达水平及印记状态

doi:10.13481/j.1671-587X.20240425

摘要/Abstract

摘要：

目的研究长链非编码RNA（lncRNA） H19和胰岛素样生长因子2（IGF2）基因在乳腺癌组织中的表达水平，分析其印记状态。方法采用实时荧光定量PCR（RT-qPCR）法检测乳腺癌组织及癌旁组织中H19和IGF2 mRNA表达水平，分析H19和IGF2 mRNA在乳腺癌组织及癌旁组织中的表达差异，利用单核苷酸多态性（SNP）区分等位基因表达情况（纯合或杂合），基因组DNA中IGF2（ApaⅠ位点）或H19（AluⅠ位点）为杂合则进行印记分析，确定H19和IGF2在乳腺癌组织中的印记状态，即印记保持（MOI）或印记丢失（LOI）。分析乳腺癌组织中H19和IGF2表达与分子分型的关系。结果 RT-qPCR法检测，乳腺癌组织中H19和IGF2 mRNA表达水平高于癌旁组织（P<0.01），H19 mRNA表达水平与IGF2 mRNA表达水平呈正相关关系（r=0.567，P<0.01）。不同分子分型乳腺癌患者癌组织中H19 mRNA表达水平均高于癌旁组织（P<0.05或P<0.01）。H19和IGF2在乳腺癌组织中均存在LOI，IGF2 的LOI发生率为36.7%，高于H19的LOI发生率（4.3%）。RT-qPCR法检测，IGF2 LOI组乳腺癌组织中IGF2 mRNA表达水平明显高于IGF2 MOI组（P<0.01）。结论乳腺癌组织中H19和IGF2 mRNA表达水平明显高于癌旁组织，IGF2的LOI发生率高于H19的LOI发生率，IGF2的LOI可能是乳腺癌发病的关键因素之一。

关键词: 乳腺肿瘤, 长链非编码RNA, H19, 胰岛素样生长因子2, 印记丢失, 单核苷酸多态性

Abstract:

Objective To study the expression levels of long non-coding RNA （lncRNA） H19 and insulin-like growth factor 2 （IGF2） genes in breast cancer tissue， and to analyze their imprinting status. Methods Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of H19 and IGF2 mRNA in breast cancer tissue and adjacent tissue， and the differences in the expressions of H19 and IGF2 mRNA in breast cancer tissue and adjacent tissue were analyzed； single nucleotide polymorphism （SNP） was used to distinguish the allele expression status （homozygous or heterozygous）. For heterozygous IGF2 （ApaⅠ site） or H19 （AluⅠ site） in genomic DNA， imprinting analysis was used to detect the imprinting status of H19 and IGF2 in breast cancer tissue， that were maintenance of imprinting （MOI） or loss of imprinting （LOI）； the relationship between the expressions of H19 and IGF2 and molecular subtypes in breast cancer tissue were also analyzed. Results The RT-qPCR results showed that the expression levels of H19 and IGF2 mRNA in breast cancer tissue were higher than those in adjacent tissue （P<0.01）. There was a positive correlation between the expression levels of H19 mRNA and IGF2 mRNA （r=0.567， P<0.01）. Compared with adjacent tissue， the expression levels of H19 mRNA in cancer tissue of the breast cancer patients with various molecular subtypes were increased （P<0.05 or P<0.01）. LOI was observed in both H19 and IGF2 in breast cancer tissue， and the incidence of IGF2 LOI was 36.7%， which was higher than that of H19 LOI（4.3%）. The RT-qPCR results showed that the expression level of IGF2 mRNA in breast cancer tissue in IGF2 LOI group was significantly higher than that in IGF2 MOI group （P<0.01）. Conclusion The expression levels of H19 and IGF2 mRNA in breast cancer tissue are significantly higher than those in adjacent tissue. The incidence of IGF2 LOI is higher than that of H19 LOI， and IGF2 LOI may be one of the key factors in the pathogenesis of breast cancer.

Key words: Breast neoplasm, Long chain non-coding RNA H19, Insulin-like growth factor 2, Loss of imprinting, Single nucleotide polymorphism

中图分类号:

R737.9

魏雪,文雪,谢潇,王月媛,黄丹,杨明. lncRNA H19和IGF2基因在乳腺癌组织中的表达水平及印记状态[J]. 吉林大学学报(医学版), 2024, 50(4): 1109-1115.

Xue WEI,Xue WEN,Xiao XIE,Yueyuan WANG,Dan HUANG,Ming YANG. Expression levels and imprinting status of lncRNA H19 and IGF2 genes in breast cancer tissue[J]. Journal of Jilin University(Medicine Edition), 2024, 50(4): 1109-1115.

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Primer	Primer sequence（3'-5'）
IGF2 Forward：	CTTGGACTTTGAGTCAAATTGGCCT
Reverse：	GAGGAGCCAGTCTGGGTTGTTGCTA
H19 Forward：	CTTTACAACCACTGCACTACCTG
Reverse：	GCCATGAAGATGGAGTCGCCG

Primer	Primer sequence（3'-5'）
IGF2 Forward：	CTTGGACTTTGAGTCAAATTGGCCT
Reverse：	GAGGAGCCAGTCTGGGTTGTTGCTA
H19 Forward：	GATCGGTGCCTCAGCGTTCG
Reverse：	GTCCTGCTTGTCACGTCCAC
β-actin Forward：	CAGGTCATCACCATTGGCAATGAGC
Reverse：	CGGATGTCCACGTCACACTTCATGA

Group	n	H19/IGF2 homozygous	H19/IGF2 heterozygous	H19 homozygous /IGF2 heterozygous	H19 heterozygous /IGF2 homozygous
Luminal	86	33 (38.4)	19 (22.1)	15 (17.4）	19 (22.1)
HER2+	14	6 (42.9)	6 (42.9)	1 (7.1)	1 (7.1)
Triple-negative	17	6 (35.3)	7 (41.2)	3 (17.6)	1 (5.9)

Group	n
Group	n	H19 genotype	Expressed	IGF2 genotype	Expressed
LOI of H19 and IGF2	1	A/C	a/c	T/C	t/c
LOI of IGF2	8	A/C	a	T/C	t/c
	6	A/C	c	T/C	t/c
	8	A	a	T/C	t/c

Group	n	H19/IGF2 MOI	H19/IGF2 LOI	H19 MOI/IGF2 LOI
Luminal	19	4 (21.1)	2 (10.5)	13 (68.4)
HER2+	6	1 (16.7)	0 (0.0)	5 (83.3)
TNBC	7	1 (14.3)	0 (0.0)	6 (85.7)