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Background and Aims: microRNA223 (miR-223) plays an important role in the development of atherosclerosis and ischemic stroke. It is involved in regulation of multiple physiological and pathophysiological processes such as cholesterol metabolism, endothelial cell (EC) function, and thrombosis. Here we investigated the role of methylation regulation of MIR-223 promoter region in atherosclerotic cerebral infarction (ACI) patients., Methods: A total of 23 patients with ACI and 32 healthy individuals were recruited. We performed bisulfite sequencing PCR and real-time PCR to detect methylation levels of MIR-223 promoter region and miR-223, respectively, in genomic DNA isolated from peripheral blood leukocytes., Results: Mean methylation levels of a total of nine CpGs of MIR-223 promoter were significantly lower in ACI patients than in healthy individuals (p < 0.01), and were also significantly lower in individuals with carotid atherosclerosis than those without carotid atherosclerosis (p < 0.05). Meanwhile, miR-223 expression in leukocytes was significantly higher in ACI patients than in healthy individuals (p < 0.05). miR-223 level was negatively correlated with mean methylation levels of MIR-223 promoter (r = -0.4451, p < 0.01). The methylation level of MIR-223 promoter revealed a positive correlation with the circulating total cholesterol level (r = 0.318, p = 0.019)., Conclusions: Hypomethylation of MIR-223 promoter is associated with atherosclerotic cerebral infarction., (Copyright © 2017. Published by Elsevier B.V.)

The aim of the study was to investigate the expression of miR-223 and FAM5C in the peripheral blood mononuclear cells (PBMCs) of cerebral infarction patients with or without diabetes. Sixteen cases with diabetes mellitus (DM), 14 cases with cerebral infarction (CI), 12 cases with cerebral infarction and diabetes mellitus (CIDM), and 18 healthy subjects were included in this study. Real-time PCR was used to quantify mRNA expression. Western blot was used to detect FAM5C protein level. Recombinant plasmids expressing miR-223-3p and 3' UTR of FAM5C were constructed. Dual-luciferase reporter system was used to analyze the binding of miR-223-3p to FAM5C 3' UTR. FAM5C mRNA and protein level were significantly higher in the PBMCs of CIDM patients compared with healthy controls (P < 0.05). miR-223-3p expression in PBMCs was significantly lower in DM patients than in healthy controls (P < 0.05). The expression of miR-223-3p was negatively correlated with FAM5C mRNA in all patients and healthy controls. Co-transfection of miR-223-3p plasmid with FAM5C 3'UTR dual-luciferase plasmid significantly inhibited the luciferase activity (P < 0.01). FAM5C, but not miR-223, is a risk factor for CI in type 2 DM patients.

The Basigin (BSG, also known as CD147/extracellular matrix metalloproteinase inducer) belongs to the immunoglobulin superfamily (IgSF). It is a cellular receptor for cyclophilin A (CypA), and is originally known as tumor cell collagenase stimulatory factor (TCSF), which could abundantly expressed on the surface of tumor cells, haematopoietic, monocytes, epithelial endothelial cells and smooth muscle cells. Accumulating evidence showed that BSG played an important role in stimulating the secretion of matrix metalloproteinases (MMPs), which has been reported to be involved in the development of atherosclerosis. Since atherosclerosis is an important risk factor for atherosclerotic cerebral infarction (ACI), we speculate that BSG genetic polymorphisms may influence formation of atherosclerosis and then development of ACI. This study aimed to detect the potential association of the single nucleotide polymorphisms (SNP, -631 G > T, -318 G > C, 10141 G > A and 10826 G > A) of BSG gene in Hunan Han Chinese population with ACI. We genotyped 199 ACI patients and 188 matched healthy controls for the four BSG SNP by method of matrix-assisted laser desorption/ionization-time-offlight mass spectrometry (MALDI-TOF MS). Our results suggested that all the polymorphisms were observed in the subjects from Changsha area of Hunan Province. However, no significant difference was observed between the distribution of these SNP in cases and controls. Therefore, we speculate that BSG genetic polymorphisms might not be an important factor in the development of ACI in our Chinese Han population.

Background: and purpose: Matrix metalloproteinases 9 (MMP-9) has been reported to play a critical role in the pathophysiology of atherosclerotic cerebral infarction (ACI). Here we assessed association of MMP-9 polymorphisms with ACI susceptibility and the function of SNPs through microRNA mediated regulation., Methods: Genotyping was performed using MALDI-TOF mass spectrometry. Reporter gene plasmids with the MMP-9 3'UTR carrying either the mutant or the wild-type MMP-9 allele were constructed. Also, we constructed pcDNA-3.1-miR-491-5p recombinant plasmid, which transiently co-transfected human umbilical vein endothelial cells (HUVEC) with the reporter plasmids. Reporter plasmids, miR-491-5p mimics and inhibitor were transfected into HUVE cells line by lipofectamine. MMP-9 mRNA expression in HUVEC was detected by RT-PCR and protein level by ELISA., Results: The rs1802908 and rs2664517 polymorphisms were not observed in all subjects from Hunan Han Chinese. No significant difference in genotype distribution of rs20544 and rs9509 between cases and controls were observed (p＞0.05). The rs1056628CC genotype had a significantly increased risk for ACI as compared with carries of the rs1056628 A allele (total χ(2) = 12.041, P = 0.002). Reporter gene assay revealed that the rs1056628 A allele showed lower reporter activity than the rs1056628C allele. Hsa-miR-491-5p had effect on modulation of MMP-9 gene in vitro. The rs1056628 A→C variant in the 3'-UTR of the MMP-9 increased MMP-9 protein expression in cultured HUVECs., Conclusions: Our data suggested that the rs1056629A→C variation contributes to an increased risk of ACI by increasing MMP-9 expression through affecting binding of miR-491 to the polymorphic site in the 3'-UTR of MMP-9., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

The phosphatase and tensin homologue (PTEN) gene is critical to the pathological development of atherosclerosis, a major risk factor for atherosclerotic cerebral infarction (ACI). However, it remains unclear whether genetic polymorphism in the PTEN gene can affect the risk of developing ACI. In the present haplotype-based case-control study, we investigated whether polymorphisms of the PTEN gene were associated with ACI in a Han Chinese population. Using matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS), we genotyped 300 patients with ACI and 204 matched healthy controls for five single nuclear polymorphisms (SNP) of the PTEN gene (rs2299939, rs17431184, rs555895, rs12357281 and rs2673836). Linkage disequilibrium and haplotype construction were analyzed using SHEsis software. We found that the rs2299939 PTEN intronic polymorphism was significantly associated with a risk of ACI in our Han Chinese population. The rs555895 and rs17431184C/T variants were not associated with risk of ACI. Two other polymorphisms were not detected in any subject in this study. The rs2299939C/rs17431184T/rs555895T PTEN haplotype was also significantly associated with risk of ACI. Our study provides the first evidence of an association between a PTEN intronic variant and the risk of ACI. It also provides new a new line of evidence for the critical role of PTEN in the development of stroke., (Copyright © 2012 Elsevier Ltd. All rights reserved.)