The use of antiinflammatory agents has been suggested as a

The use of antiinflammatory agents has been suggested as a potential treatment in HF. One strategy targeted tumor necrosis factor (TNF) in HF,, but the negative results of the clinical trials have been discouraging., Other clinical trials have attempted to suppress proinflammatory cytokine production in patients with HF with the use of transcriptional or translational approaches, and the benefits were doubtful., Glutathione S-transferase P1-1 (GSTP1) inhibits apoptosis and detoxifies reactive oxygen species to maintain the cellular redox state., , Importantly, GSTP1 has antiinflammatory effects that are mediated through inhibition of mitogen-activated protein kinase (MAPK) activity, primarily c-Jun N-terminal kinase 1 (JNK1) and the p38 signaling pathway., GSTP1 also participates in regulation of stress signaling and protects Azaserine receptor against apoptosis via its noncatalytic ligand-binding activity. In the setting of MI-induced HF, prosurvival pathways such as MAPK, and mediators of apoptosis such as caspases have been identified as potential treatment targets, allowing rescue of damaged myocardial cells. GSTP1 interacts with TNF-α receptor–associated factor 2 (TRAF2), JNK1, and p38, resulting in inhibition of TRAF2-induced MAPK activation., We have recently shown that GSTP1 is overexpressed in screening protein microarrays obtained from HF patients and that serum GSTP1 is a specific and sensitive marker in HF. However, whether GSTP1 could be beneficial in HF treatment is unknown. The present study addressed this issue by examining cardiac tissues of patients with ischemic HF followed by evaluation of GSTP1 efficacy in an experimental rat model of MI-induced HF. Methods
Results Both GSTP1 and TRAF2 mRNA (P < .0001) and protein (P < .001) expressions were significantly elevated in failing myocardium (Fig. 1A and B); although binding activity of GSTP-1 with TRAF2 was not significantly changed compared with control (Fig. 1C). Heart failure was also associated with significantly higher protein expression of active JNK1 and p38 (P < .0001; Fig. 1D) but with reduced binding activity of JNK1 and p38 with GSTP-1 (P < .0001; Fig. 1E). We then examined whether exogenously supplemented GSTP1 affects its own binding activity in failing myocardial tissue in vitro, and we found a significant concentration-dependent inhibition of JNK1 and p38 following GSTP1 treatment in both failing and control human cardiac tissue compared with untreated control samples (P < .001; Fig. 2A). Of note, GSTP1 treatment resulted in a significantly greater binding activity of itself to TRAF2 and JNK1 in failing cardiac tissue cultures but not to p38 in vitro (P < .01; Fig. 2B). TNF-α treatment significantly up-regulated active JNK1 and p38 and reduced GSTP-1 binding activity to JNK1 and p38 in both failing and control cardiac tissue cultures compared with untreated control samples (P < .01; Fig. 2C and D). The subsequent addition of recombinant GSTP1 effectively suppressed the TNF-α–induced JNK1 and p38 activation in tissue cultures (P < .01; Fig. 2C). On the other hand, GSTP1 treatment enhanced its own binding activity to TRAF2 and JNK1 that had been suppressed by TNF-α pretreatment (P < .01; Fig. 2D). Dylight 488–labeled GSTP1 was abundantly present in rat cardiac macrophages and skeletal myoblasts and fibroblasts, whereas Dylight-labeled BSA was not detectable in those cells (Fig. 3A). A very high myocardial uptake was found after [123I]GSTP injection. At 120 minutes after injection, there was only slight [123I] myocardial activity measured in the control animals. In contrast, animals injected with [123I]GSTP showed a 15-fold higher [123I] signal (P < .001; Fig. 3C and D). Collagen staining in serial sections indicated a significant reduction of infarct area (P < .01) and increase of thinning ratio (P = .02) at 21 days after MI in GSTP1-treated rats compared with control subjects (Fig. 4A–C). The apoptotic index was significantly decreased within the AAR of GSTP1-treated group versus control at 24 hours (P = .04; Supplemental Fig. 1A) and at 21 days after MI (P = .02; Fig. 4D). At 24 hours and 21 days after MI, the majority of the apoptotic nuclei in control (95.9 ± 2% and 97.3 ± 1%, respectively) and treated (97.2 ± 0.8% and 96.1 ± 0.9%, respectively) groups were cardiomyocytes (Supplemental Fig. 1B; Fig. 4E). Flow cytometry analysis of Annexin V indicated a significantly decreased apoptosis in GSTP1-treated rats (9.3 ± 1.6% at 24 hours [Supplemental Fig. 1C] and 13.7 ± 3.5% at 21 days [Fig. 4F]) compared with control subjects (13.5 ± 3.1% and 23.1 ± 4.9%, respectively). GSTP1 treatment also significantly down-regulated active caspase-3 and -9 (both P < .02 at 24 hours [Supplemental Fig. 1C] and P < .004 and P < .006, respectively, at 21 days after MI [Fig. 4G]). Moreover, Kaplan-Meier curve revealed that GSTP1 treatment improved animal survival (P < .05; Fig. 4H).