Mesenchymal stem cell (MSC) therapy for myocardial injury has inherent limitations due to the poor viability of MSCs after cell transplantation. In this study, we directly delivered Hsp70, a protein with protective functions against stress, into MSCs, using the Hph-1 protein transduction domain ex vivo for high transfection efficiency and low cytotoxicity. Compared to control MSCs in in vitro hypoxic conditions, MSCs delivered with Hph-1-Hsp70 (Hph-1-Hsp70-MSCs) displayed higher viability and anti-apoptotic properties, including Bcl2 increase, reduction of Bax, JNK phosphorylation and caspase-3 activity. Hsp70 delivery also attenuated cellular ATPdepleting stress. Eight animals per group were used for in vivo experiments after occlusion of the left coronary artery.Transplantation of Hph-1-Hsp70-MSCs led to a decrease in the fibrotic heart area, and significantly reduced the apoptotic positive index by 19.5 6 2%, compared to no-treatment controls. Hph-1-Hsp70-MSCs were well-integrated into the infarcted host myocardium. The mean microvessel count per field in the infarcted myocardium of the Hph-1-Hsp70-MSCtreated group (122.1 6 13.5) increased relative to the MSCtreated group (75.9 6 10.4). By echocardiography, transplantation of Hph-1-Hsp70-MSCs resulted in additional increases in heart function, compared to the MSCs-transplanted group. Our results may help formulate better clinical strategies for in vivo MSC cell therapy for myocardial damage.
PurposeThiazolidinediones (TZDs) are known to inhibit the proliferation of vascular smooth muscle cell (VSMC) by increasing the activity of p27Kip1 and retinoblastoma protein (RB). However, the upstream signaling mechanisms associated with this pathway have not been elucidated. The Akt-mTOR-P70S6 kinase pathway is the central regulator of cell growth and proliferation, and increases cell proliferation by inhibiting the activities of p27Kip1 and retinoblastoma protein (RB). Therefore, we hypothesized in this study that rosiglitazone inhibits VSMC proliferation through the inhibition of the Akt-TOR-P70S6K signaling pathway.Materials and MethodsRat aortic smooth muscle cells (RAoSMCs) were treated with 10 µM of rosiglitazone 24 hours before the addition of insulin as a mitogenic stimulus. Western blot analysis was performed to determine the inhibitory effect of rosiglitazone treatment on the Akt-mTOR-P70S6K signaling pathway. Carotid balloon injury was also performed in Otsuka Long-Evans Tokushima Fatty (OLETF) diabetic rats that were pretreated with 3 mg/kg of rosiglitazone.ResultsWestern blot analysis demonstrated
significant inhibition of activation of p-Akt, p-m-TOR, and p-p70S6K in cells treated with rosiglitazone. The inhibition of the activation of the p-mTOR-p-p70S6K pathway seemed to be mediated by both the upstream PI3K pathway and MEK-ERK complex.ConclusionThe inhibitory effect of rosiglitazone on RAoSMC proliferation in vitro and in vivo is mediated by the inhibition of the Akt-mTOR-P70S6K pathway.
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