Inflammation predicts risk of cardiovascular disease (CVD) events, but the relation of drugs that directly target inflammation with CVD risk is not established. Methotrexate is a disease-modifying anti-rheumatic drug broadly used for treatment of chronic inflammatory disorders. We performed a systematic review and meta-analysis of evidence for relationships of methotrexate with CVD occurrence. Cohorts, case-control studies or randomized trials were included if they reported an association between methotrexate and CVD risk. Inclusions/exclusions were independently adjudicated, and all data were extracted in duplicate. Pooled effects were calculated using inverse-variance-weighted meta-analysis. Of 694 identified publications, 10 observational studies, in which methotrexate was administered among patients with rheumatoid arthritis, psoriasis, or polyarthritis met inclusion criteria. Methotrexate was associated with 21% lower risk of total CVD (n=10 studies, 95% CI=0.73-0.87, p<0.001), and 18% lower risk of myocardial infarction (n=5, 95% CI=0.71-0.96, p=0.01), without evidence for statistical between-study heterogeneity (p=0.30, p=0.33, respectively). Among prespecified sources of heterogeneity explored, stronger associations were observed in studies that adjusted for underlying disease severity (RR=0.64, 95% CI=0.43-0.96, p<0.01), and for other concomitant medication (RR=0.73, 95% CI=0.63-0.84, p<0.001). Publication bias was potentially evident (funnel plot, Begg’s test, p=0.06); excluding studies with extreme risk estimates did not, however, alter results (RR=0.81, 95% CI=0.74-0.89). In conclusion, methotrexate use is associated with lower risk of CVD among patients with chronic inflammation. These findings suggest that a direct treatment of inflammation may reduce CVD risk.

BackgroundCurrent evidence suggests that endothelial progenitor cells (EPC) contribute to ischemic tissue repair by both secretion of paracrine factors and incorporation into developing vessels. We tested the hypothesis that cell-free administration of paracrine factors secreted by cultured EPC may achieve an angiogenic effect equivalent to cell therapy.Methodology/Principal FindingsEPC-derived conditioned medium (EPC-CM) was obtained from culture expanded EPC subjected to 72 hours of hypoxia. In vitro, EPC-CM significantly inhibited apoptosis of mature endothelial cells and promoted angiogenesis in a rat aortic ring assay. The therapeutic potential of EPC-CM as compared to EPC transplantation was evaluated in a rat model of chronic hindlimb ischemia. Serial intramuscular injections of EPC-CM and EPC both significantly increased hindlimb blood flow assessed by laser Doppler (81.2±2.9% and 83.7±3.0% vs. 53.5±2.4% of normal, P<0.01) and improved muscle performance. A significantly increased capillary density (1.62±0.03 and 1.68±0.05/muscle fiber, P<0.05), enhanced vascular maturation (8.6±0.3 and 8.1±0.4/HPF, P<0.05) and muscle viability corroborated the findings of improved hindlimb perfusion and muscle function. Furthermore, EPC-CM transplantation stimulated the mobilization of bone marrow (BM)-derived EPC compared to control (678.7±44.1 vs. 340.0±29.1 CD34+/CD45− cells/1×105 mononuclear cells, P<0.05) and their recruitment to the ischemic muscles (5.9±0.7 vs. 2.6±0.4 CD34+ cells/HPF, P<0.001) 3 days after the last injection.Conclusions/SignificanceIntramuscular injection of EPC-CM is as effective as cell transplantation for promoting tissue revascularization and functional recovery. Owing to the technical and practical limitations of cell therapy, cell free conditioned media may represent a potent alternative for therapeutic angiogenesis in ischemic cardiovascular diseases.

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