To investi gate if various ratios of ADSC influence cardiomyocyte density, lentivirally eGFP tagged ADSC were co cultured with lentivirally dTomato tagged HL 1 cardiomyocytes. The HL 1 cell density doubled in a 1 1 and 1 2 ratio and further increased in a 1 3 and 1 4 ratio compared to HL 1 cardiomyocytes alone. ADSC enhanced HL 1 cardiomyocyte selleck screening library proliferation rate in all ratios, no significant differences were found between various ratios of ADSC to HL 1 cardiomyocytes. Conditioned medium of ADSC promotes the rate of proliferation of HL 1 cardiomyocytes Possibly, secreted factors of ADSC could cause the enhanced proliferation rate of cardiomyocytes. The puta tive beneficial influence of conditioned media from ADSC was assessed on rnCM and HL 1 cardiomyocytes subjected to hypo ia and inflammation.
In serum containing media, appro i mately 10% and 12% of the rnCM proliferated respectively under normo ia and hypo ia. Serum starvation reduced the rate of proliferating rnCM to appro imately 8% irrespectively of additional inflammatory factors. Normo ic conditioned medium of ADSC did not change the rate of rnCM proliferation in high serum. Entinostat Yet, after serum starvation the proliferation rate of rnCM increased 1. 4 fold after treat ment with normo ic conditioned medium of ADSC. The pre conditioning of ADSC with TNF or IL 1B for the formation of the primed conditioned medium of ADSC resulted in respectively 1. 2 fold increase in the proliferation rate of rnCM compared to TNF or IL 1B primed rnCM under hypo ia.
To confirm the positive effect of the conditioned medium of ADSC on the enhancement of the cardiomyocyte proliferation rate, we performed the readout on the pure cardiomyo cytes HL 1 cells. In normal culture medium, appro imately 85% of the HL 1 cardiomyocytes proliferated under both normo ic and hypo ic conditions. Serum starvation reduced the fraction of proliferating HL 1 cardiomyocytes almost two fold under normo ia or hypo ia. Treatment of serum free HL 1 cardiomyocytes with TNF or IL 1B did not alter proliferation, irrespective of o y gen concentration. Serum free conditioned medium from normo ically cultured ADSC increased the proliferation rate of serum free HL 1 cardiomyocytes by 18% compared to serum free HL 1 cardiomyocytes control.
The proliferation rate of HL 1 cardiomyocytes under normo ic conditions was even further stimulated upon incubation with conditioned medium from ADSC prestimulated with TNF or IL 1B compared to TNF or IL 1B stimulated serum free HL 1 cardiomyocytes controls. The pro inflammatory stimulation of ADSC with TNF or IL 1B to obtain primed http://www.selleckchem.com/products/Enzastaurin.html ADSC conditioned medium ameliorated the cardiomyocyte proliferation rate as well. Furthermore, IL 1B primed conditioned medium of ADSC significantly increased the HL 1 proliferation rate compared to nonstimulated conditioned medium of ADSC.