BACKGROUND: Transplantation of Akt-over-expressing mesenchymal stem cells (Akt-MSCs) has been shown to repair infarcted myocardium and improve cardiac function. However, little is known about the therapeutic effects of Akt-MSCs on cardiac autonomic neuropathy in chronic heart failure (CHF). OBJECTIVE: The present study used adriamycin-induced CHF rat models to observe the effect of Akt-MSCs on cardiac autonomic nervous regeneration and the factors mediating this effect. DESIGN, TIME AND SETTING: A randomized, controlled animal experiment was performed at the Central Laboratory of Basic Medical College, China Medical University, between September 2008 and April 2009.MATERIALS: Rabbit anti-choline acetyltransferase (ChAT), growth associated protein-43 (GAP-43), synaptophysin (SYN) polyclonal antibodies and the secondary antibody (goat anti-rabbit IgG) were purchased from Boster, China. Cat-A-Kit assay system was provided by Amersham, USA.METHODS: (1) Adult rat MSCs were isolated and cultured for the preparation of Akt-MSCs. (2) Forty male Wistar rats were intramyocardially administered adriamycin at 2 mg/kg over 3 days for a total of five times and once a week for additional five times thereafter to establish CHF models. At 2 weeks after final adriamycin treatment, 34 successful CHF rat models were randomized to three groups: Akt-MSCs (n = 11), simple MSCs (s-MSCs, n =11), and control (n = 12). Each group was intravenously administered Akt-MSCs (2×10~6 cells in 100 μL PBS), s-MSCs (2×10~6 cells in 100 μL PBS) or equal volume of phosphate buffered saline, once a day for a total of three times. MAIN OUTCOME MEASURES: At 4 weeks after final adriamycin treatment, myocardial norepinephrine (NE) content was detected using a Cat-A-Kit assay system. Myocardial ChAT, SYN and GAP-43 were performed by immunohistochemistry and Western blot analysis. Prior to, 2 and 4 weeks after adriamycin treatment, echocardiographic examination was performed and left ventricular ejection fraction (LVEF) was determined. RESULTS: Myocardial NE content, as well as SYN-positive and GAP-43-positive nerve fiber density and expression, and LVEF, was the greatest in the Akt-MSCs group, followed by the s-MSCs group, and lastly the control group (P < 0.05 or P < 0.01). ChAT expression was similar between Akt-MSCs and s-MSCs groups, but it was higher compared with the control group (P < 0.05). NE contents were negatively correlated to LVEF (r = -0.64, P = 0.015). CONCLUSION: Transplantation of MSCs, in particular Akt-MSCs, promotes cardiac nervous regeneration in failing heart, which might be mediated by GAP-43.