浅表血栓性静脉炎管壁基质金属蛋白酶及其抑制剂表达研究

doi:10.3877/cma.j.issn.1674-0793.2024.05.005

中华普通外科学文献(电子版) ›› 2024, Vol. 18 ›› Issue (05) : 338 -343. doi: 10.3877/cma.j.issn.1674-0793.2024.05.005

论著

浅表血栓性静脉炎管壁基质金属蛋白酶及其抑制剂表达研究

李星月¹, 董伟², 徐永波³, 张文法¹, 胡晓璇⁴, 钟玉绪⁴, 褚海波¹^,^†(

)

^1. 261035　潍坊，潍坊医学院附属医院普通外科
^2. 266300　青岛，同济大学附属东方医院胶州医院普通外科
^3. 261200　潍坊，中国人民解放军陆军第80集团军医院普通外科
^4. 100850　北京，中国人民解放军军事科学院军事医学研究院毒物药物研究所

收稿日期:2024-04-12 出版日期:2024-10-01
通信作者: 褚海波
基金资助:
中国人民解放军后勤重大项目(AWS17J008); 潍坊市青年医学人才托举工程基金

Expression of matrix metalloproteinases and their tissue inhibitors in the vein wall following superficial venous thrombosis

Xingyue Li¹, Wei Dong², Yongbo Xu³, Wenfa Zhang¹, Xiaoxuan Hu⁴, Yuxu Zhong⁴, Haibo Chu¹^,^†()

^1. Department of General Surgery, Affiliated Hospital of Weifang Medical College, Weifang 261035, China
^2. Department of General Surgery, Jiaozhou Branch of Shanghai East Hospital, Tongji University, Qingdao 266300, China
^3. Department of General Surgery, the 80th Group Army Hospital of People’s Liberation Army, Weifang 261200, China
^4. State Key Laboratory of Toxicology and Medical Countermeasures, Institute of Pharmacology and Toxicology, Beijing 100850, China

Received:2024-04-12 Published:2024-10-01
Corresponding author: Haibo Chu

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引用本文：

李星月, 董伟, 徐永波, 张文法, 胡晓璇, 钟玉绪, 褚海波. 浅表血栓性静脉炎管壁基质金属蛋白酶及其抑制剂表达研究[J]. 中华普通外科学文献(电子版), 2024, 18(05): 338-343.

Xingyue Li, Wei Dong, Yongbo Xu, Wenfa Zhang, Xiaoxuan Hu, Yuxu Zhong, Haibo Chu. Expression of matrix metalloproteinases and their tissue inhibitors in the vein wall following superficial venous thrombosis[J]. Chinese Archives of General Surgery(Electronic Edition), 2024, 18(05): 338-343.

目的

检测浅表血栓性静脉炎（SVT）、大隐静脉曲张（VGSVs）和正常大隐静脉（GSVs）管壁基质金属蛋白酶（MMPs）和其组织抑制剂（TIMPs）表达变化。

方法

应用免疫组织化学（IHC）、聚合酶链反应（PCR）、Western blotting方法，检测三组静脉管壁中MMP-2、MMP-9、TIMP-1、TIMP-2蛋白阳性表达率、mRNA和蛋白表达。

结果

SVT组MMP-2、MMP-9、TIMP-1、TIMP-2蛋白阳性表达率、mRNA和蛋白表达均显著高于VGSVs组和GSVs组，差异有统计学意义（P<0.05）。MMP-2/TIMP-2比值SVT组和VGSVs组均低于GSVs组，差异有统计学意义（P<0.05）；MMP-9/TIMP-1比值SVT组均低于VGSVs组和GSVs组，差异有统计学意义（P<0.05）。

结论

在高流体静力压和炎症条件下，SVT管壁MMPs和TIMPs蛋白表达明显失调，提示MMPs和TIMPs可能参与静脉管壁重塑过程。

关键词: 浅表血栓性静脉炎, 隐静脉, 基质金属蛋白酶, 金属蛋白酶组织抑制剂

Objective

To investigate the changes in expression of matrix metalloproteinases (MMPs) and the tissue inhibitor of metalloproteinases (TIMPs) from superficial venous thrombosis (SVT), varicose great saphenous veins (VGSVs), and great saphenous veins (GSVs).

Methods

In the venous walls of the three groups, the expression of MMP-2, MMP-9, TIMP-1, and TIMP-2 proteins, protein-positive expression ratios, mRNA expression, and protein expression were determined by immunohistochemistry, polymerase chain reaction, and Western blotting.

Results

The MMP-2, MMP-9, TIMP-1, and TIMP-2 protein-positive expression ratios, mRNA and protein expression in the SVT group were significantly higher than those in the VGSVs and the GSVs groups (P<0.05). The MMP-2/TIMP-2 ratios in the SVT and the VGSVs groups were significantly lower than those in the GSVs group (P<0.05), respectively. The MMP-2/TIMP-2 and MMP-9/TIMP-1 ratios in the SVT group were significantly lower than those in the VGSVs and the GSVs groups (P<0.05), respectively.

Conclusions

Disequilibrium of MMPs and TIMPs in SVT wall occurs due to underlying high hydrostatic pressure and inflammation. These results suggested that MMPs and TIMPs participate in the process of venous wall remodeling.

Key words: Superficial venous thrombosis, Saphenous vein, Matrix metalloproteinases, Tissue inhibitor of metalloproteinases

表1 MMP-2、MMP-9、TIMP-1、TIMP-2相关基因的引物序列及扩增产物

基因名称	基因编码	引物序列	产物大小(bp)
MMP-2	NM_001127891.1	上游引物：5'-ATCTTTGCTGGAGACAAATTCTGGA-3'	175
		下游引物：5'-GCTTCAGGTAATAGGCACCCTTGA-3'
MMP-9	NM_004994.2	上游引物：5'- ACGCACGACGTCTTCCAGTA -3'	94
		下游引物：5'- CCACCTGGTTCAACTCACTCC -3'
TIMP-1	NM_003254.2	上游引物：5'-CCTTATACCAGCGTTATGAGATCAA-3'	194
		下游引物：5'-AGTGATGTGCAAGAGTCCATCC-3'
TIMP-2	NM_003255.4	上游引物：5'-CTGGACGTTGGAGGAAAGAAGG-3'	148
		下游引物：5'-CCATCTGGTACCTGTGGTTCAGG-3'
GAPDH	NM_002046.3	上游引物：5'-GCACCGTCAAGGCTGAGAAC-3'	138
		下游引物：5'-TGGTGAAGACGCCAGTGGA-3'

表1 MMP-2、MMP-9、TIMP-1、TIMP-2相关基因的引物序列及扩增产物

基因名称	基因编码	引物序列	产物大小(bp)
MMP-2	NM_001127891.1	上游引物：5'-ATCTTTGCTGGAGACAAATTCTGGA-3'	175
		下游引物：5'-GCTTCAGGTAATAGGCACCCTTGA-3'
MMP-9	NM_004994.2	上游引物：5'- ACGCACGACGTCTTCCAGTA -3'	94
		下游引物：5'- CCACCTGGTTCAACTCACTCC -3'
TIMP-1	NM_003254.2	上游引物：5'-CCTTATACCAGCGTTATGAGATCAA-3'	194
		下游引物：5'-AGTGATGTGCAAGAGTCCATCC-3'
TIMP-2	NM_003255.4	上游引物：5'-CTGGACGTTGGAGGAAAGAAGG-3'	148
		下游引物：5'-CCATCTGGTACCTGTGGTTCAGG-3'
GAPDH	NM_002046.3	上游引物：5'-GCACCGTCAAGGCTGAGAAC-3'	138
		下游引物：5'-TGGTGAAGACGCCAGTGGA-3'

图1 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-2蛋白表达　A~C分别为SVT组、VGSVs组和GSVs组MMP-2蛋白表达；D~F为SVT组、VGSVs组和GSVs组MMP-9蛋白表达；G~I为SVT组、VGSVs组和GSVs组TIMP-1蛋白表达；J~L为SVT组、VGSVs组和GSVs组TIMP-2蛋白表达（箭头所示，IHC, ×200, Bars=50 μm）

表2 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-2阳性表达率比较（%,

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	70.040±4.257^a	52.000±3.189^b	5.933±2.375	335.864	<0.001
MMP-9	70.400±3.175^a	52.960±2.475^b	5.000±1.927	2888.871	<0.001
TIMP-1	71.920±3.752^a	52.880±2.744^b	5.000±2.299	1169.311	<0.001
TIMP-2	72.600±4.052^a	53.600±2.769^b	4.200±2.007	637.307	<0.001

表2 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-2阳性表达率比较（%,

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	70.040±4.257^a	52.000±3.189^b	5.933±2.375	335.864	<0.001
MMP-9	70.400±3.175^a	52.960±2.475^b	5.000±1.927	2888.871	<0.001
TIMP-1	71.920±3.752^a	52.880±2.744^b	5.000±2.299	1169.311	<0.001
TIMP-2	72.600±4.052^a	53.600±2.769^b	4.200±2.007	637.307	<0.001

表3 3组静脉管壁阳性表达率MMP-2/ TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2/TIMP-2	0.968±0.087^a	0.972±0.071^a	1.651±0.965	-40.137	<0.001
MMP-9/TIMP-1	0.982±0.080	1.005±0.077	1.176±0.741	-141.682	0.226

表3 3组静脉管壁阳性表达率MMP-2/ TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2/TIMP-2	0.968±0.087^a	0.972±0.071^a	1.651±0.965	-40.137	<0.001
MMP-9/TIMP-1	0.982±0.080	1.005±0.077	1.176±0.741	-141.682	0.226

表4 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-2 mRNA表达比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	2.135±0.301^a	1.752±0.172	1	129.971	<0.001
MMP-9	1.810±0.228^a	1.501±0.177	1	95.427	<0.001
TIMP-1	1.809±0.128^a	1.451±0.099	1	304.501	<0.001
TIMP-2	1.840±0.261^a	1.364±0.158	1	97.464	<0.001

表4 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-2 mRNA表达比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	2.135±0.301^a	1.752±0.172	1	129.971	<0.001
MMP-9	1.810±0.228^a	1.501±0.177	1	95.427	<0.001
TIMP-1	1.809±0.128^a	1.451±0.099	1	304.501	<0.001
TIMP-2	1.840±0.261^a	1.364±0.158	1	97.464	<0.001

表5 3组静脉管壁基因表达MMP-2/ TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2/TIMP-2	1.186±0.251^a	1.302±0.206^a	1	-10.475	<0.001
MMP-9/TIMP-1	1.003±0.129	1.038±0.139	1	-0.751	0.476

表5 3组静脉管壁基因表达MMP-2/ TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2/TIMP-2	1.186±0.251^a	1.302±0.206^a	1	-10.475	<0.001
MMP-9/TIMP-1	1.003±0.129	1.038±0.139	1	-0.751	0.476

表6 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-1蛋白表达比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	1.105±0.128^a	0.636±0.079^b	0.346±0.016	1598.563	<0.001
MMP-9	1.085±0.039^a	0.704±0.024^b	0.349±0.021	2873.253	<0.001
TIMP-1	2.234±0.177^a	1.002±0.084^b	0.428±0.035	2226.214	<0.001
TIMP-2	1.235±0.131^a	0.530±0.065^b	0.192±0.065	2176.540	<0.001

表6 3组静脉管壁MMP-2、MMP-9、TIMP-1、TIMP-1蛋白表达比较（

±s）

检测指标	SVT组	VGSVs组	GSVs组	F值	P值
MMP-2	1.105±0.128^a	0.636±0.079^b	0.346±0.016	1598.563	<0.001
MMP-9	1.085±0.039^a	0.704±0.024^b	0.349±0.021	2873.253	<0.001
TIMP-1	2.234±0.177^a	1.002±0.084^b	0.428±0.035	2226.214	<0.001
TIMP-2	1.235±0.131^a	0.530±0.065^b	0.192±0.065	2176.540	<0.001

表7 3组静脉管壁蛋白表达MMP-2/TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSV组	GSVs组	F值	P值
MMP-2/TIMP-2	0.906±0.154^a	1.207±0.141^b	2.043±0.779	-12.405	<0.001
MMP-9/TIMP-1	0.489±0.045	0.708±0.056	0.822±0.098	-1.523	0.226

表7 3组静脉管壁蛋白表达MMP-2/TIMP-2、MMP-9/ TIMP-1比值比较（

±s）

检测指标	SVT组	VGSV组	GSVs组	F值	P值
MMP-2/TIMP-2	0.906±0.154^a	1.207±0.141^b	2.043±0.779	-12.405	<0.001
MMP-9/TIMP-1	0.489±0.045	0.708±0.056	0.822±0.098	-1.523	0.226

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Expression of matrix metalloproteinases and their tissue inhibitors in the vein wall following superficial venous thrombosis

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Expression of matrix metalloproteinases and their tissue inhibitors in the vein wall following superficial venous thrombosis