Abstract 231: Upregulation of Tenascin-C Expression in Rheumatic Heart Valve Disease by Synergistic TNF-a and Shear Stress via RhoA/ROCK Signaling Pathway
Background: Group A streptococcal infection has traditionally been viewed as the pathologic agent of rheumatic heart valvular disease (RHVD), but can not account for differential morbidity of the four heart valves. Higher aortic and mitral valve morbidity indicates that mechanical forces may play a defining pathologic role. Tenascin-C (TN-C) expression, promiment in rheumatic diseases, has been found to be induced during times of mechanical duress. However, its expression, regulation, and relation to RHVD is unidentified.
Methods: Valvular specimens, interstitial cells, and sera were obtained from valve replacement operations of RHVD patients (n=30) and congenital valvular malformation control (n=20). Sera TN-C and TNF-α were detected by ELISA, Western blot, and immunohistochemistry. Valvular interstitial cells were cultured in corresponding sera (10%) under shear stress (5 Dyne, 60min) provided by an original cyclic fluid culture system. Expression of Tenascin-C, RhoA, and ROCK activity were determined by Western blot, and the effects of TNF-a neutralizing antibody and ROCK-I/II inhibitor Y27632 were investigated.
Results: TN-C and TNF-α levels in RHVD patients were significantly higher than control (93.4±32.6 ng/ml vs. 29.6±7.3 ng/ml, 19.3±5.2 pg/ml vs. 3.8±1.7 pg/ml respectively, P<0.01), and were confirmed via immunohistochemical methods. Co-culture under shear stress and TNF-α stimulation significantly enhanced TN-C expression in synergistic fashion (TN-C/actin, 0.98±0. 25 vs. 3.62±0.34, P<0.01), and increased both RhoA (membrane-bound RhoA/total RhoA protein, 32.7%±4.6% vs. 61.9%±5.5%, P<0.01) and ROCK activity (p-MYPT1/total MYPT1, 21.8%±3.7% vs. 49.2%±4.1%, P<0.01) in valvular interstitial cells. This effect was inhibited by neutralization antibody against TNF-α or ROCK-I/II inhibitor Y27632.
Conclusion: Our results suggest TN-C, a potential biomarker of RHVD, is upregulated in synergistic fashion by TNF-α and shear stress. Activation of the RhoA/ROCK signaling pathway plays a critical role in valvular pathogenesis, and its blockade may provide a novel therapeutic approach for RHVD treatment or prevention.