Fig. 4.

SDF-1 induced-TNF-α production. a, b To address whether SDF-1 induces TNF-α production, freshly isolated adult rat cardiomyocyte were treated for 24 h with either 10, 50,100, 200, 300 or 500 ng/ml SDF-1 and the TNF-α protein expression was assessed in either cell culture supernatant (a) or cell lysates (b) using ELISA assay. c To conform whether SDF-1 mediated apoptosis is TNF-α-dependent, cardiomyocytes were pretreated with neutralizing antibody against TNF-α (5 μg/ml of anti-murine TNF-α) followed by SDF-1 treatment (300 ng/ml) for 24 h. IgG isotype match was used as negative control. Apoptosis was detected by using TUNEL assay and percentage (%) of TUNEL positive cells was determined (i.e., number of TUNEL-positive myocytes/total number of myocytes × 100). Three independent experiments were performed, in each experiment, each condition was performed in triplicate (i.e., coverslips from three separate dishes were counted per treatment condition, a total of 15 fields) (n = 15; *p < 0.05)