Effect of calcium-sensing receptor on apoptosis of neonatal rat cardiomyocytes / 国际儿科学杂志

ABSTRACT

ObjectiveTo observe the influence of calcium - sensing receptor (CaSR) on Fas/Fas ligand (Fas L) pathway during anoxia/reoxygenation (A/R)- induced cardiomyocytes apoptosis in neonatal rat. MethodsSingle cells were dissociated from minced hearts of 2 - day - old Wistar rats with a 0. 25％ solution of crude trypsin and then cultured as monolayers at a density of 5 x 104cells/cm2 in DMEM medium equilibrated with humidified air containing 5％ CO2 at 37C. Three days after the cells were seeded, the cultured cardiomyocytes were randomly divided into three groups. ( 1 ) control group cardiomyocytes were continuously cultured for 26 hours in DMEM medium.(2) A/R group cardiomyocytes underwent anoxia for 2 hours and reoxygenation for 24 hours.(3) GdCl3 group 300μmol/L GdCl3 was added to the culture medium at the beginning of reoxygenation. Apoptosis of cardiomyocytes was assessed by flow cytometer and morphological alterations were observed with transmission electron microscope.The expression of CaSR, Fas and Fas L were analyzed by Western blot.Results The result of flow cytometer showed that cardiomyocytes apoptosis was 12. 18％ ± 1.54％ in A/R group,and was higher than that in the control group (P ＜0. 01 ). At the same time, mitochondrial cristae dissolution and disappearance could be detected. Compared with A/R group, GdCl3, a specific activator of CaSR, further enhanced cardiomyocytes apoptosis to 20. 25％ + 2. 87％ ( P ＜ 0. 01 ), along with an increment in CaSR, Fas and Fas L expressions ( P ＜ 0. 05 ). ConclusionCaSR is closely involved in cardiomyocytes apoptosia during anoxia/reoxygenation. CaSR could induce apoptosis of neonatal rat cardiomyocytes through Fas/Fas L receptor pathway.