Autophagy modulation altered differentiation capacity of CD146+ cells toward endothelial cells, pericytes, and cardiomyocytes

Abstract Background: To date, many attempts are employed to increase the regenerative potential of stem cells. In this study, we evaluated the hypothesis whether an autophagy modulation could induce/reduce CD146+ cells differentiation into mature pericyte, endothelial and cardiomyocyte lineage. Methods In this study, CD146+ cells were enriched from human bone marrow aspirates and trans-differentiated into mature EC, PC and CM after exposure to autophagy stimulator (50 µM Met)/inhibitor (15 µM HCQ). The protein levels of autophagy proteins were monitored by western blotting. NO content was measured using the Griess assay. Using real-time PCR assay and western blotting, we monitored the lineage protein and gene levels. The fatty acid change was determined by gas chromatography. Pro-inflammatory cytokine and angiocrine factors were measured by ELISA. The exosome secretion capacity was assessed by AChE activity and real-time PCR assay. Result Data revealed the modulation of autophagy factors, Beclin-1, P62 and LC3 II/I ratio in differentiating CD146+ cells after exposure to Met and HCQ (p<0.05). The inhibition of autophagy increased released NO content and decreased intracellular NO content compared to the Met-treated cells (p<0.05). Real-time PCR analysis showed that the treatment of CD146+ cells with autophagy modulators altered the expression of VE-cadherin, cTnI and α-SMA. Our data demonstrated that the stimulation of autophagy signaling in CD146+ cells with Met increased the expression of VE-cadherin, α-SMA, and cTnI compared to HCQ-treated cells (p<0.05) while western blotting revealed the protein synthesis of all lineage-specific proteins in under the stimulation and inhibition of autophagy. Fatty acid profile analysis revealed the increase of unsaturated fatty acids after exposure to HCQ (p<0.05). None statistically significant differences were found in the levels of Tie-1, Tie-2, VEGFR-1 and VEGFR-2 after autophagy modulation. The treatment of cells with HCQ increased the levels of TNF-α and IL-6 compared to the Met-treated cells. Data revealed the increase of exosome biogenesis and secretion to the supernatant in cells treated with HCQ compared to the Met groups (p<0.05).ConclusionsIn summary, autophagy modulation could be altered differentiation potency of CD146+ cells and could be novel and applicable cardiac cell therapy in the cardiac regeneration field.