The Association Of Adiponectin With Cardiovascular Diseases And Its Mechanism

Part1:Meta-Analysis of the Association between ADIPOQ Polymorphisms and Cardiovascular DiseasesBackgrounds:Adiponectin is encoded by the ADIPOQ gene, which is located on the chromosomal region3q27, spanning17kb and comprising three exons. Whole genome linkage studies have identified that this genomic region is a susceptible locus for type2diabetes, the metabolic syndrome, and cardiovascular diseaseas. In the past decade, several single nucleotide polymorphisms(SNPs) of the ADIPOQ gene have been identified. Many molecular epidemiological studies have been conducted focusing on the association between variants in the ADIPOQ and the risk of CVDs. However, the evidence is inadequate to draw robust conclusions because of the small effect of these SNPs on the susceptibility to CVDs and the relatively small sample size in each study. To summarize the published data, we conducted a meta-analysis from all eligible studies to assess the association between the SNPs of ADIPOQ and the risk of CVDs.Methods:A comprehensive search was performed within these electronic databases of PubMed, EMBASE and Science Citation Index Expanded up to April,2011to find all studies about the association between the SNPs of ADIPOQ and the risk of CVDs. All studies was reviewed and selected according to the inclusion criteria and exclusion criteria, the quality of studies assessed by assigning a quality score using a standardized extraction form, data were collected independently by using a standardized data extraction form. All statistical analyses were performed using Stata statistical software. Q statistic and I2statistic was used to estimate heterogeneity across all eligible studies, possible causes of heterogeneity were explored and stratification analysis was performed. A logistic regression approach proposed for molecular association studies was used to estimate a biological effect of SNPs and evaluated genetic model. Data were pooled according to the best genetic model. The pooled data was calculated by the inverse variance method and the significance of pooled data was tested by Z statistic. Sensitivity analysis was performed to assess the stability of the results by excluding a single study in the meta-analysis each time to reflect the influence of the individual data on the pooled data. Publication bias was assessed qualitatively by performance of funnel plots and quantitatively by means of Begg’s and Egger’s tests.Results:26eligible articles concerned about17SNPs of the ADIPOQ association with CVDs were identified based on our criteria. The enough number of eligible studies for meta-analysis was focused on four SNPs of rs1501299,rs2241766,rs266729and rs822396. Each SNP association with CVDs was analyzed respectively.18eligible studies including5,365cases and10,241controls determined the association of the SNP rs1501299(+276G>T) with CVDs. There is no statistic significance(p=0.36) about the overall effect of SNP rs1501299(+276G>T) on CVDs risk by Logistic regression analysis. Stratification analysis was performed according to ethnicity, quality score of study and sample population, to explore potential sources of heterogeneity and estimate the association of the SNP rs1501299(+276G>T) with CVDs risk in different subgroups. Although there was no significant heterogeneity in subgroup of general population, Logistic regression analysis showed it is no statistic significance(p=0.85) about the overall effect of SNP rs1501299(+276G>T) on CVDs risk in this subgroup, even excluding the low quality studies(p=0.78). There was nearly no heterogeneity in subgroup of T2DM population, but the heterogeneity remained even excluded the low quality score studies in other groups. Only data of T2DM subgroup was pooled. The gene effect of SNP rs1501299(+276G>T) is significant in the subgroup of T2DM population including1433cases and2621 controls(p=0.04).The genetic model was most likely to be recessive genetic model (TT versus TG plus GG) for T2DM population. Data pooled according to this genetic model yielded an odds ratio of0.74(95%CI:0.58,0.94; z=2.47,p=0.013). Sensitivity analysis showed every individual study had no significant influence on the overall pooled estimates.Publication bias showed the dots were nearly symmetrically distributed in the funnel plot, and the P values from Begg’s test (p=1.00) and Egger’s test (p=0.96) were greater than0.05.15eligible studies including5,151cases and8,704controls determined the association between the SNP rs2241766(+45T>G) and CVDs. There was significant heterogeneity across the studies. Although the heterogeneity was acceptable in subgroup of Caucasian by stratification, Logistic regression analysis showed it is no statistic significance about the overall effect of SNP rs2241766(+45T>G) on CVDs(p=0.84). The heterogeneity remained even excluded the low quality score studies in other groups,it is no significance to pool data.20eligible studies including6,401cases and11,897controls determined the association between the SNP rs266729(-11377C>G) and CVDs. There was significant heterogeneity across the studies, and the heterogeneity remained in each subgroup even excluded the low quality score studies by stratification.5eligible studies including2,582cases and3,611controls determined the association between the SNP rs822396(-3971A>G) and CVDs. Although there was no significant heterogeneity across the studies, Logistic regression analysis showed it is no statistic significance about the overall effect of SNP rs822396(-3971A>G) on CVDs(p=0.44).Conclusions:There is a recessive protective effect of SNP rs1501299(+276G>T) on CVDs for T2DM population, the TT homozygote had a lower risk of developing CVDs than carriers of G allele, but the association between SNP rs1501299(+276G>T) and CVDs in other population remains an unresolved issue. As for SNP rs2241766(+45T>G) and SNP rs266729(-11377C>G), there is no sufficient evidence to evaluate the association with CVDs. The present evidence indacates that the association between SNP rs822396(-3971A>G) and CVDs is not significant.lt suggested that the SNP rs1501299was one of the reasons that T2DM population was susceptibility to CVDs. Some programs could be specifically targeted at T2DM population early to prevent from CVDs events. Part2:The Effect of Adiponectin on Insulin Resistance and Oxidative Stress in CardiomyocyteBackgrounds:Insulin resistance is the key pathological feature of obesity, type2diabetes(T2DM) or metabolic syndrome(MS), and also characteristic of many clinical and experimental settings. Insulin resistance also plays an important role in cardiovascular diseases, diabetic cardiomyopathy or insulin-resistant cardiomyopathy, retinopathy, and neuropathy etc. A growing body of evidence indicates that oxidative stress, a significant imbalance between the production and elimination of reactive oxygen species(ROS), is always accompanied with insulin resistance and plays a major role in insulin resistance. Adiponectin is abundantly secreted by mature adipocytes and also by other cells. Studies indicated that adiponectin can ameliorate insulin sensitivity and regulate biological processes like apoptosis, proliferation, migration and inflammation.Both in vitro studies and animal experiments demonstrate that adiponectin acted directly on cardiomyocytes to protect the heart from ischaemic injury, hypertrophy, cardiomyopathy and systolic dysfunction. The effect of adiponectin on oxidative stress and related mechanisms are not understanded. We used high concentration of insulin (high-insulin) to induce the insulin resistance and oxidative stress in Cardiomyoblast H9c2cells, and to investigate the potential mechanisms of insulin resistance and oxidative stress as well as the effects of full-length adiponectin on insulin resistance and oxidative stress. Methods:Insulin resistant cells were induced by treatment with high-insulin. The insulin resistance was detected by glucose consumption and uptake. Oxidative stress was detected with2’,7’-dichlorodihydrofluorescein diacetate, a cell-permeable non-fluorescent probe can be deesterified by intracellular esterases and be oxidized to highly fluorescent2’,7’-dichlorodihydrofluorescein(DCF) by reactive oxygen species. The relevant gene and protein expression such as GLUT4, the member of NADPH oxidases (Nox) family, antioxidant enzymes and related singal transduction molecules were measured by reverse transcription polymerase chain reaction(TR-PCR) and western blots to investigate the mechanisms underlying insulin resistance and oxidative stress, as well as the effects of adiponectin.Results:Both basal and insulin-stimulated glucose uptake as well as glucose consumption were decreased after treating with200nM insulin for24h(p<0.05for basal and p<0.01for insulin-stimulated). GLUT-4expression also decreased markedly when treating with high-insulin for24h(p<0.05).High-insulin markedly augmented ROS production(p<0.01). Both DPI and Rotenone could inhibit ROS generation in response to high-insulin(p<0.05for Rot and p<0.01for DPI).RT-PCR showed expression of Noxl, Nox2, Nox4mRNA, but not Nox3and Nox5in H9c2cells. Only Nox4increased significantly in high-insulin-treated H9c2cells by RT-PCR and Western boltting(p<0.05).The mRNA and protein of SOD1and Gpxl decreased markedly when treated for24h with high-insulin(p<0.05). p-Akt(Ser473) decreased and p-Erkl/2(Thr202and Tyr204) increased significantly in high-insulin-treated H9c2cells(p<0.05for p-Akt and p<0.01for Erkl/2).Insulin-stimulated glucose uptake inhibited by high-insulin was increased significantly when pretreated with5.0μg/ml adiponectin in H9c2cells(p<0.05). The decreased expression of GLUT4also recovered significantly(p<0.05). High-insulin-induced ROS was decreased markedly by pretreating with adiponectin(p<0.05).Both alteration of Nox4and antioxidant enzymes were partly recovered in high-insulin-treated H9c2cells by pretreating with adiponectin(p<0.05). SOD1and Gpxl expression also increased significantly(p<0.05), but the alteration of Nox4was not significant when treated with adiponectin alone.Decreased p-Akt(Ser473) in high-insulin-treated H9c2cells could restore by pretreated with full-length adiponectin. The level of p-Erk(Thr202and Tyr204) increased in high-insulin treated H9c2cells could down-regulate by pretreated with full-length adiponectin or Erkl/2inhibitor U0126(p<0.05). The alteration of GLUT4, Nox4, SOD1and Gpxl induced by high-insulin were all recovered partly by pretreating with Erkl/2inhibitor U0126(p<0.05).Conclusions:High-insulin could induce insulin resistance alone in H9c2cells.High-insulin-induced insulin resistance was accompanied with oxidative stress. The elevated ROS mainly generated from Nox4and also involved in anti-oxidant enzymes impairment. Both insulin resistance and oxidative stress induced by high-insulin were alleviated by pretreatment with full-length adiponectin.The Erkl/2 and Akt pathways were involved in insulin resistance, oxidative stress and the effects of adiponectin on high-insulin-treated H9c2cells.