| ObjectiveIn order to study the effect of leptin and adiponectin on proliferation ,cell cycle distribution and apoptosis of human breast cancer cells MCF-7 and T47D. And explore the role of leptin and adiponectin in the occurrence and development of breast cancer.Methods(1) MCF-7 and T47D cell lines were grown in RPMI-1640 medium supplemented with 10% and 15% FBS respectively. Trypan Blue was used to count the number of T47D cell line.The growth of the cell line was also observed by invert microscope every 24h.The experiment lasted for three days.(2) The cell lines were treated with several concentrations of leptin for 24h,48h and 72h.The effect of leptin on proliferation of MCF-7 and T47D cell lines were determined by means of MTT method.(3) After staining by PI ,the effect of leptin for 48h on distribution of cell cycle of MCF-7 and T47D cell lines were determined by FCM.The effect of adiponectin on distribution of cell cycle of T47D cell line was also determined by FCM.(4) By Annexin V/PI double fluorescent staining we use FCM to estimated the effect of leptin and adiponectin on apoptosis of MCF-7 and T47D cell lines.Results(1) By invert microscope we found that the T47D cells grew well . It was monolayer cultured and the morphous was irregular. The.The number of the cells was increasing from 1.7×105/ml to 4.2×105/ml in 72h.(2) When treated with (25-400)ng/ml leptin for 24h,48h,72h, the leptin could induce the proliferation of the MCF-7 cells. There was not interaction between concentration of leptin and action time(F=0.919,P=0.523). The main effect of concentration of leptin and action time was statistically significant(P<0.01,respectively). Compared 200ng/ml and 400ng/ml with the control group,we found the difference was statistically significant by multiple comparison (P<0.01, respectivelly).And the effect was most markedly at treated with 400ng/ml leptin for 72h.The proliferation rate was 16.6%. The difference was also statistically significant among action time by multiple comparison(P<0.01,respectivelly).(3) When treated with the same concentration and action time,we found the leptin could induce the proliferation of the T47D cells. There was not interaction between concentration of leptin and action time(F=0.620,P=0.787). The main effect of concentration of leptin and action time was statistically significant(P<0.01,respectively). Compared 200ng/ml and 400ng/ml with the control group,we found the difference was statistically significant by multiple comparison (P<0.01, respectivelly).And the effect was most markedly at treated with 400ng/ml leptin for 48h.The proliferation rate was 21.75%. The difference was also statistically significant among action time by multiple comparison(P<0.01,respectivelly).(4) After treated with 100ng/ml and 400ng/ml leptin on MCF-7 cells for 48h,the proportion of G0/G1 phase was(60.26±2.26)%,(54.72±2.55)%,respectively. Compared with the control group(63.94±0.81)% ,the difference was statistically significant (F=10.464,P=0.044).The proportion of S phase was(32.24±0.94)%, (36.62±0.75)%,respectively. Compared with the control group(29.97±0.04)% ,the difference was also statistically significant(F=47.361,P=0.005). But the difference of G2/M phase was not statistically significant(F=1.77,P=0.311). And the same concentration of leptin could not change the G0/G1,S,and G2/M phase of the cell cycle of T47D cells(P>0.05).(5) After treated with 500ng/ml and 2000ng/ml adiponectin on T47D cells for 48h,the proportion of G0/G1 phase was(91.07±0.63)%,(91.60±0.98)%,respectively. Compared with the control group(85.31±1.07)% ,the difference was statistically significant (F=29.277,P=0.011). But the difference of S and G2/M phase was not statistically significant(P>0.05).(6) After treated with 100ng/ml and 400ng/ml leptin on MCF-7 cells for 24h, the difference of early apoptosis rate and total apoptosis rate among control group and them was not statistically significant(P>0.05). After treated with the same concentration of leptin on T47D cells for 24h, the early apoptosis rate was (1.77±0.47)%,(1.08±0.03)%,respectively. Compared with the control group (6.54±1.27)%,the difference was statistically significant (F=28.213, P=0.011). And the total apoptosis rate was (2.13±0.74)%, (1.55±0.16)%,respectively. Compared with the control group (7.48±1.39)%, the difference was also statistically significant (F=25.642,P=0.013).(7) After treated with 500ng/ml and 2000ng/ml adiponectin on MCF-7 cells for 24h, the total apoptosis was (17.20±0.31)%,(16.25±0.29)%,respectively. Compared with the control group (12.61±0.62)%, the difference was statistically significant (F=62.049,P=0.004). But the difference of early apoptosis rate among control group,500ng/ml and 2000ng/ml adiponectin was not statistically significant(P>0.05). The same concentration of adiponectin could not induce the early apoptosis and total apoptosis of T47D cells(P>0.05,respectively).Conclusions(1) The leptin may induce the proliferation of MCF-7 and T47D cell lines.(2) The leptin could change the distribution of cell cycle of MCF-7 cells.It may induce the DNA synthesis through promoting G0/G1 enter to S phase. The leptin may inhibit apoptosis of T47D cells. The leptin may serve as a risk factor of breast cancer development.(3) The adiponectin may regulate the distribution of cell cycle.It may induce G0/G1 phase arrest of T47D cells and reduce the DNA synthesis. The adiponectin may induce midlate apoptosis of MCF-7 cells. It suggest that adiponectin may serve as a protective factor of breast cancer development.
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