Correlated Expression Studies Of CHI3L1 And MMP-9 In Human Carotid Atherosclerotic Plaque

BackgroundNow China, with the rapid development of economy and society, the improvement of people's living standards, the increase of diet diversity, greater intake of high-fat and high-cholesterol food, the increasing pressure of life, the amount of exercise reduced and the aging of society advances, lead the incidence of atherosclerotic disease increased year by year. Atherosclerosis(atherosclerosis AS) is the foundation of most vascular disease and potential risk factors. With Chinese society gradually entered the aging progress, atherosclerosis has tremendous threat to people's health, poses a great burden to family and society. The incidence of ischemic cerebral apoplexy higher year by year, with poor prognosis and high mortality and disability rate. Its etiology and pathogenesis is still not completely clear. Carotid atherosclerosis is the most important risk factor for ischemic stroke, and it is closely related to the location, progression and recurrence of cerebral infarction. Foreign studies have reported the carotid atherosclerosis and the degree of stenosis has a close relationship with the occurrence of ischemic cerebral apoplexy.In recent years a large number of domestic and foreign research shows that, the unstable plaque formation is one of the main causes of acute cardiovascular events, therefore, unstable plaque has become the popular direction of research at home and abroad, the main pathological features include thinning fibrous cap, lipid core is larger and the large amount of inflammatory cell infiltrate. The risk factors that lead to the occurrence and development of AS can lead to vascular endothelial dysfunction, and then the expression of adhesion molecules increased significantly, and chemotaxis monocytes and T lymphocyte adhesion. Monocytes and lymphocytes into the intima start inflammatory reaction, leading to secretion of large amounts of cytokines, such as tumor necrosis factor(tumor necrosis factor, TNF) and interrukin(IL), monocyte chemoattractant protein-1(MCP-1) etc.They can further activate the EC and vascular smooth muscle cells(VSMCs), macrophages, and promote further development of the AS. Chitinase 3-like protein 1(CHI3L1), also known as YKL-40 and cartilage glycoprotein 39, breast regression protein. The protein exists chitin binding activity, but no chitinase activity. Studies have shown that it can be secreted by a series of cells, and it plays a very important role in the process of acute and chronic inflammation, and extracellular matrix(ECM) reconstruction and other diseases, such as atherosclerosis, diabetes, asthma, hypertension and other, also in the unstable plaque plays a very important role. MMP-9 is one important member of the matrix metalloproteinase family, MMP- 9 role in the pathological process of atherosclerosis is mainly manifested in the following aspects: 1) Promote the migration of smooth muscle cells under the lining formate atheromatous plaque, then lead to the progress of atherosclerosis plaques, 2) Degradate of extracellular matrix and collagen, lead to unstable plaque and plaque rupture by regulating metabolism and degradation of extracellular matrix, participation in lipid retention, calcinosis, endothelial cells and smooth muscle cells of biological behaviour change, etc. It's on the degradation of extracellular matrix(ECM) in vascular smooth muscle cells migrate into the vascular intima-media resulting in IMT thickening and in the process of plaque formation and plaque instability plays an important role.However at present there is no reports said in carotid atheromatous plaque tissues the expression of CHI3L1 and the expression of MMP- 9 whether there is any change, and whether there is a correlation between the two. ObjectDiscuse the expression level of CHI3L1 and MMP-9 in human carotid atherosclerotic plaque, and the relationship between them, and hope to further explore the pathogenesis of carotid atherosclerosis, and find new therapeutic targets. Materials and methods1.From April 2012 to April 2014 we collected 42 cases of carotid atherosclerotic plaque specimens in our department by carotid endarterectomy, 25 cases of them were male, the other 17 cases were female. Before operation all cases through carotid artery ultrasonography and carotid CTA and angiography make definite diagnosis. In addition, as control group wecollected 20 cases of splenic artery from whom suffered spleen resection due to trauma,exclude patients with portal hypertension,16 cases of them were male, the other 4 cases were female. Before operation, all patients signed "The informed consent of the study of carotid atherosclerotic plaque specimens ", the specimens were divided into two parts, one part quickly were placed in 80 DEG cryogenic refrigerator for mRNA detection, the other part were fixed in 10% formaldehyde solution to be embedded in paraffin, which were used for the detection of immunohistochemistry.2.All specimens were stained with HE, the carotid atherosclerosis plaque tissue classified into stable plaque group(20 cases) and unstable plaque group(22 cases) according the preoperative ultrasonography and carotid CT angiography, splenic artery tissue as control group(20 cases). 3.The expression levels of the protein of CHI3L1 and MMP-9 in three groups were determined by immunohistochemistry SP method, and analyze the significance of the protein of CHI3L1 and MMP- 9 as well as the relationship between them.4. Reverse transcription polymerase chain reaction(RT-PCR) was used to detect the expression of CHI3L1 mRNA and MMP-9 mRNA in the three groups, and analyze the significance of CHI3L1 mRNA and MMP-9 mRNA as well as the relationship between them.5. Statistical method: use SPSS17.0 statistical software analyze the experimental data, results were expressed by sx±, use single factor analysis of variance compare the three groups, compare any two means by Bonferroni, the expression of the protein CHI3L1 and MMP-9,and their mRNA expression in carotid atherosclerotic plaques, and the relationship between was analyzed by Pearson correlation analysis. With P< 0.05 for the difference was statistically significant Results1.Immunohistochemical results:1.1CHI3L1 staining was mainly located in inflammatory cells, EC, VSMCS and foam cells.The protein CHI3L1 strongly expressed in the patch tissues, and the expression was weakly in the control group. There was significant difference in the expression level of CHI3L1 protein between the three groups(F=482.407,P<0.001).In the stable plaque group(2.087±0.027) CHI3L1 expressed higher than the control group(1.946±0.019),in the unstable plaque tissue(2.168±0.022) CHI3L1 expressed higher than the control group, and the expression of CHI3L1 in unstable plaque tissue(2.168±0.022) was higher than in stable plaque tissue.1.2MMP-9 was expressed in inflammatory cells, EC and VSMCS, and foam cells.The protein MMP-9 strongly expressed in the patch tissues, and the expression was weakly in the control group. There was significant difference in the expression level of MMP-9 between the three groups(F=488.210,P<0.001). In the stable plaque group(2.098± 0.025) MMP-9 expressed higher than the control group(196.7 + 0.026), in the unstable plaque tissue(2.233± 0.031) MMP-9 expressed higher than the control group, and the expression of MMP-9 in the unstable plaque tissue(2.233 ± 0.031) was higher than in the stable plaque tissue.1.3The expression of CHI3L1 and MMP-9 in the plaque tissue was analyzed by Pearson correlation analysis. Results showed that both expressed by the positive correlation(rstable =0.555,P<0.001;runstable =0.618,P<0.001).2.The results of Reverse transcription-polymerase chain reaction(RT-PCR):2.1The expression difference of CHI3L1 mRNA was statistically significant(F=675.837,P<0.001).The expression of CHI3L1 mRNA in the unstable plaque tissue(1.592± 0.036) was the highest, the expression of CHI3L1 mRNA in the stable plaque group(133.8 ±+ 0.046) was higher than the control group(1.157± 0.033).2.2The expression difference of MMP-9 mRNA was statistically significant( F=518.019,P<0.001), The expression of MMP-9 mRNA in the unstable plaque tissue(1.402± 0.024) was the highest, the expression of MMP-9 mRNA in the stable plaque group(1.268 ± 0.019) was higher than the control group(1.183±0.023).2.3The expression of CHI3L1 mRNA and MMP-9 mRNA in the plaque tissue was analyzed by Pearson correlation analysis. Results showed that both expressed by the positive correlation(rstable mRNA=0.536,P<0.001;r unstable mRNA=0.582,P<0.001). Conclusion1.CHI3L1 and MMP-9 expressed highly in human carotid atherosclerotic plaque.2.The high expression of CHI3L1 and MMP-9 may be related to the instability of carotid atherosclerotic plaque.3.CHI3L1 and MMP-9 may have synergistic effect in promoting the instability of carotid atherosclerotic plaque.