Association of Inflammatory Gene Polymorphisms With Ischemic Stroke in a Chinese Han Population

Nan Zhao; Xin Liu; Yongqin Wang; Xiaoqiu Liu; Jiana Li; Litian Yu; Liyuan Ma; Shuyu Wang; Hongye Zhang; Lisheng Liu; Jingbo Zhao; Xingyu Wang

Disclosures

J Neuroinflammation. 2012;9(162) 

In This Article

Discussion

In this case–control study, we assessed the relationship of 51 SNPs in 35 genes related to inflammatory response with the risk of ischemic stroke in 1,124 cases and 1,163 controls. In the control group, there were more individuals with hypertension and BMI was slightly higher compared to the case group. This was due to our effort to recruit controls to match the cases with blood pressure in the search for genes predisposing to ischemic stroke with and without the influence of hypertension.

Initial analysis showed that CCL11 rs4795895 and CCL11 rs3744508 were significantly associated with ischemic stroke by adjusting for age, sex, BMI and hypertension status, and after adjusting for multiple testing by FDR, CCL11 rs4795895 remained significantly associated with ischemic stroke. The CCL11 rs4795895 genetic polymorphism was the only SNP that retained significant association with ischemic stroke in the whole group. After stratification by hypertension, SNP rs4795895 was still significant in both the non-hypertensive and hypertensive groups, though after adjusting for the multiple hypothesis test it was not significantly associated with ischemic stroke. Polymorphisms rs4795895 and rs3744508 were located in the 5′ flanking region and the coding region of the eotaxin 1 gene, respectively.[14] Previous reports have noted that the presence of the CCL11 rs3744508 A allele is associated with lower CCL11 production[15] and an increased risk of myocardial infarction.[16] Ours is the first study to report any association with SNPs of CCL11 and ischemic stroke. There are two possible mechanisms by which eotaxin might be related to stroke risk. First, it may be the direct consequence of atherosclerosis development. The eotaxin gene and protein are over expressed in human atherosclerosis.[17] Vascular smooth muscle cells (VSMCs)[18] and endothelium[19] in human atheroma prominently express eotaxin, suggesting that eotaxin contributes to the progression of atherosclerosis. Second, increased eotaxin levels can mirror the development of inflammatory responses in heart and brain. Farahi et al. reported that circulating eotaxin levels were increased in patients with coronary artery disease.[15]

To explore the influence of inflammatory genes on ischemic stroke in the hypertension and non-hypertension groups, we stratified study subjects by their hypertension status and found that CCR2 rs1799864 and CCR5 rs1799987 polymorphisms were associated with ischemic stroke in the hypertensive group, and CCL11 rs3744508, LTC4S rs730012, FCER1B rs569108, TGFB1 rs1800469 and LTA rs909253 were associated with ischemic stroke in the non-hypertensive group.

CCR2 rs1799864, a SNP causing an isoleucine-for-valine substitution at position 64 in the CCR2 receptor on chromosome 3, was found to be associated with ischemic cardiomyopathy and myocardial infarction in Czech populations.[20,21] And in an African-American population from the Johns Hopkins Sibling and Family Heart Study, it was reported that the combination of homozygotic or heterozygote CCR2 rs1799864 genotype and monocyte chemoattractant protein 1 (MCP-1) rs2857656 homozygous genotype was associated with a particularly high prevalence of carotid artery plaque.[22] Chemokine receptor 2 (CCR2) is the receptor for MCP-1. Recent studies have indicated that MCP-1 and CCR2 could have an important role in regulating blood–brain barrier permeability. An animal study had indicated that a lack of CCR2 greatly reduced brain edema formation and BBB disruption, as well as decreasing leukocyte infiltration, and CCR2−/− mice had decreased expression of a wide range of proinflammatory cytokines during reperfusion.[23] Polymorphism CCR5 rs1799987 is located at position 59,029 in the promoter region of CCR5 gene. The A allele of the SNP is associated with susceptibility to diabetic nephropathy.[24] Chemokine receptor 5 (CCR5) is the receptor for expressed on regulated on activation, normal T cell expressed and secreted (RANTES). RANTES was found as a mediator of cerebral ischemia-reperfusion (I/R) induced blood–brain barrier disruption, tissue injury, and the inflammatory and prothrombogenic phenotype assumed by the cerebral microvasculature after focal I/R.[25] In our study, CCR2 rs1799864 and CCR5 rs1799987 were associated with ischemic stroke only in the hypertensive group. We tested the interaction between the polymorphisms and hypertension status, and observed no relationship between them. A possible explanation is that in conditions of high blood pressure, patients carrying risk allele(s) suffer from cerebrovascular accident more than others due to direct or indirect effects of CCR2/CCR5 on blood–brain barrier and blood vessel of brain.

CCL11CCR2 and CCR5 belong to the CC family of chemokines. CC chemokines and their receptor polymorphisms have been shown to be associated with a number of infection or inflammation-related disease states, such as atherosclerosis.[26] To the best of our knowledge, our study is the first to demonstrate the association of CCL11 CCR2 and CCR5 genetic polymorphisms with the risk of ischemic stroke in a Chinese population. CC chemokines may play an important role in both the etiology and pathophysiology of ischemic stroke; the functional relevance of these SNPs to ischemic stroke remains to be further elucidated and CC chemokines could represent promising therapeutic targets in primary and secondary prevention of ischemic stroke.

In our study TGFB1 rs1800469, LTC4S rs730012 and LTA rs909253 were associated with ischemic stroke in the non-hypertensive group, which is in line with previous reports,[9,27–30] although these findings drew from different ethnic populations and different stroke subtypes. Transforming growth factor β (TGFB) is a pleiotropic cytokine with a diversity of effects, such as inducing or facilitating vascular stenosis and thrombogenesis.[31,32]TGFB1 rs1800469 polymorphism is located in the promoter region and considered possible modulator of expression of TGF-β1 gene and levels. A case–control study had found an increased risk of stroke associated with the risk allele of the rs1800469 in a population from The Netherlands.[27] The rs730012 variant of leukotrieneC4 Synthase (LTC4S) upregulates LTC4S mRNA expression, increasing the synthesis of proinflammatory leukotrienes.[28] One study found that the LTC4S rs730012 variant contributed to large vessel stroke risk in children with sickle cell anemia.[29] Our previous study, a meta-analysis of association of candidate polymorphisms and ischemic stroke in six populations, indicated LTA rs909253 was associated with increased risk for ischemic stroke in normotensive subjects but not in hypertensive ones.[9] This is in concordance with the present report.

The β subunit of high affinity IgE receptor (FCER1B) is a critical component of the IgE receptor, serving as a chaperone to promote assembly and expression and a strong amplifier of IgE-mediated signals.[33] The FCER1B rs569108 G allele was found to have an association with the development of allergic asthma in a Chinese population.[30] In our study, the G allele of rs569108 increased the risk of ischemic stroke in non-hypertensive subjects. To explore the role of FCER1B polymorphism in pathogenesis of ischemic stroke, more genetic and functional studies will be needed.

There were five SNPs identified in the non-hypertension group and two SNPs in the hypertension group, however there were many more patients in the hypertension group. One possible explanation is that in the non-hypertensive group, the effect of genetic variation in inflammation gene may be easier to discern. Clinical and experimental studies have reported that hypertension itself is a chronic, low-grade inflammatory process.[34] High blood pressure levels are associated with an increase in circulating levels of inflammation markers that can reflect a vascular inflammatory process.[35] Hypertension could be a stronger factor causing inflammatory process. Therefore, it is possible that in the hypertensive group the genetic influence of inflammatory polymorphisms is concealed. Another possible explanation is that different ischemic stroke subtypes may occur in the different groups stratified by hypertensive status, and stratification could result in a more homogeneous population. According to Arboix et al.'s research,[13] hypertension was the main cardiovascular risk factor only for lacunar and atherothrombotic infarction. In the present study, different significant SNPs were observed in the hypertensive and non-hypertensive groups, indicating that these polymorphisms may be associated with different subtypes of ischemic stroke, although we could not further define the subtypes. To the best of our knowledge, there is no study with the same design as ours, although hundreds of genetic studies of ischemic stroke have been performed in different geographic and ethnic populations. The ambiguous definition and stratification of ischemic stroke subtypes may be a major reason why no confirmed inflammatory genetic factor was found despite so many research projects on the genetics of ischemic stroke.

There are limitations to our study. First, the current study was a retrospective study, and the included stroke patients were stroke survivors. It could not exclude possible selection bias. Secondly, the cases were not further diagnosed with stroke subtypes, which may confound the results. Thirdly, due to the limitations of the study scope we did not collect information on subjects' diabetes status, hypercholesterolemia, and coronary artery disease, as well as plasma concentrations of inflammatory markers, which play an important role in the development of ischemic stroke. Also, smoking status was not included in the multivariable analyses. Finally, the results from this study have not been replicated, and should be further validated by a carefully designed study.

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