Figure 3

Hypoxia and pro-inflammatory mediators upregulate IL-6 secretion by ADSC. A, B) Quantitative RT-PCR analysis of gene transcript levels of IL-6 normalized to GAPDH expression. Gene transcript levels of IL-6 were strongly induced by stimulation with pro-inflammatory mediators (IL-1β) both under normoxia and hypoxia conditions within 24h and 48h (p < 0.05). The increase of IL-6 gene transcript level was far stronger after IL-1β stimulation (p < 0.05). C, D). Secreted IL-6, quantified by ELISA, was significantly increased after pre-stimulation of ADSC with the pro-inflammatory cytokine IL-1β (p < 0.05). Graphs represent triplicates (with SEM) data from n = 3 independent experiments from 3 donors. E, F) IL-6 is a positive regulator of cardiomyocyte proliferation. In serum-free medium, treatment with IL-6 restored the proliferation of rnCM and HL-1 murine cardiomyocytes compared to serum-free and serum (10% FBS) controls (p < 0.05). Similarly, pretreatment of human ADSC with IL-1β, yielded conditioned media that increase proliferation of rnCM and fully restored the proliferation of HL-1 cardiomyocytes in the absence of serum (p < 0.05). Addition of IL-6 neutralizing antibody to the IL-6 or conditioned medium of ADSC treated serum-free rnCM and HL-1 cardiomyocytes resulted in a significant decreased proliferation of rnCM and HL-1 cardiomyocyte (p < 0.05). Graphs represent triplicates (with SEM) data from n = 3 independent experiments, normalized to ADSC cultures before the treatment (0h).