Effect of erythropoietin on the proliferation and apoptosis of neonatal porcine islet cells.
OBJECTIVE
To investigate the effect of erythropoietin on the proliferation,differentiation, and apoptosis of the cultured neonatal porcine islet cells in vitro.


METHODS
Neonatal porcine islet cells were separated and pured from neonatal pigs with collagenase digestion and tissue culture, and their viability and purity were tested. The neonatal porcine islet cells were divided into a control group and an experimental group.The experimental group was treated with erythropoietin but not the control group, and the insulin secretion responsiveness induced by low and high glucose stimulation in the islet was tested after 5 days. Cells were counted and the activation of amplification was determined by MTT chromatometry. The rates of cell apoptosis were observed by ethidium bromide/acridine orange (EB/AO) of fluorescent light staining and flow cytometry, and the cell cycle was analyzed by flow cytometry. The expression of bcl-2, bax, caspase-3, glucose transporter 2 (GlUT-2), and pancreatic duodenal homeobox-1 (PDX-1) mRNA was tested by RT-PCR.


RESULTS
After erythropoietin was treated in the cell culture, the neonatal porcine islet cells had normal morphology, function, and reaction of insulin secretion to the glucose stimulation. Cell count showed more cells in the experimental group than in the control group (P<0.05). MTT chromatometry showed the optical absorbance tended to increase with time, and compared with the control group, the optical absorbance was higher in the experimental group (P<0.05), the expression of PDX-1 mRNA was slightly up-regulated (P<0.05). The expression of GLUT-2 mRNA had no difference in the 2 groups (P=0.34). In the experimental group, the apoptisis rate was lower than that in the control group by flow cytometry and EB/AO fluoscence staining (P<0.01), and the expression of bcl-2 mRNA was higher. Howerer bax mRNA and caspase-3 mRNA were obviously lower than those in the control group (P<0.01).


CONCLUSIONS
Erythropoietin can promote the proliferation but has no effect on the function of neonatal porcine islet cells in vitro. Erythropoietin can protect neonatal porcine islet cells from apoptosis through up-regulating bcl-2 mRNA and downreguling bax and caspase-3 mRNA.
Department of Endocrinology, Third Xiangya Hospital, Central South University, Changsha 410013,China.
Honghui
H
He
Tianhui
T
Wu
Jing
J
Xiong
Ke
K
Chen
Zhaohui
Z
Mo
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D004921:Erythropoietin / Q000494:pharmacology*; D007515:Islets of Langerhans / Q000166:cytology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D013552:Swine; D051028:bcl-2-Associated X Protein / Q000235:genetics / Q000378:metabolism
D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D004921:Erythropoietin; D053148:Caspase 3