Plasma Beta-2 microglobulin (β2M) level is inversely associated with glomerular filtration rate (GFR) and ejection fraction (EF) in patients with chronic kidney disease. However, few studies have evaluated roles of β2M in cardiac remodeling induced by hypertension. The present data showed that serum β2M level was increased in patients with hypertension as compared to control group, and it was significantly higher in patients with chronic heart failure than in those with hypertension. Serum β2M level was related to diastolic dysfunction in patients with hypertension and heart failure. During 4 weeks after transverse aortic constriction (TAC) in mice, β2M level in serum or heart tissue increased progressively in time-dependent manner. Cardiomyocyte but not fibroblast secretory β2M was rapidly increased after mechanical stretch (MS) in vitro. Exogenous β2M showed pro-fibrotic effects and enhanced migratory capability in cardiac fibroblasts. Conditional medium (CM) from mechanical stretched cardiomyocytes showed the similar effects in fibroblasts, but these effects were partly abolished by CM from β2M-downregulated-cardiomyocytes subjected to MS. In vivo, AAV9 mediated-knockdown of cardiac β2M greatly improved the cardiac fibrosis and dysfunction induced by TAC. Further analysis indicated that β2M promoted the phosphorylation of p38, smad2/3 and epidermal growth factor receptor (EGFR) in fibroblasts, and inhibition of EGFR attenuated p-p38 and p-smad2/3 levels, and significantly inhibited pro-fibrotic effects in cardiac fibroblasts treated with β2M. The present data demonstrated the importance of β2M as a paracrine pro-fibrotic mediator associated with hypertension or heart failure. It suggests that inhibition of β2M may have the therapeutic potential for the prevention of cardiac fibrosis or the progression of heart failure associated with hypertension.