Effect Of Serum Isolated From Osteoporotic Human On The Growth And Differentiation Of Normal Human Osteoblast

ABSTRACT

Osteoporosis is a condition in which an imbalance appears between bone resorption and formation,with bone resorption exceeding formation. Recent studies have shown that bone formation abnormalities in various forms of osteopenia result mainly from defective recruitment of osteoblastic cells. These abnormalities in osteoblast function and bone formation are associated with alterations in the expression or production of several growth factors, such as TGF-beta which modulate the proliferation and activity of bone-forming cells. Bone transplantation is an absolute requirement in several patholigical conditions. The growth factors, such as TGF-beta ,PDGF,were the effective in promoting growth of the bone in vitro and in animal models. We have investigated the effects of PDGF and TGF-beta on the proliferation and differentiation of the normal human osteoblast in vitro culture.The normal human osteoblast from iliac bone were primarily cultured. The serums obtained from the osteoporotic patients and the normal was used to quantify PDGF and TGF-beta from the osteoporotic patients serum and the normal serum.To clarify the effects of the various different the culture conditions such as 1 x 10(4)cells/ml,2.5 x 10(4)cells/ml, 5 x 10(4)cells/ml, 10 x 10(4)cells/ml (0.22 x 10(4)cells/well) of the osteoblast at 24 hours, 48 hours, 72 hours under 10%FBS, 10%normal humanserum, 10%osteoporotic human serum, 3% normal human PRP, 3%osteoporotic human PRP. The cell proliferation and differentiation was determined by [3H ]-thymidine and SRB assay, and the magnitude of differentiation to osteoblast was confirmed by von Kossa staining and alkaline phosphatase stain and measuring the alkaline phosphatase activity during 48 hours and 72 hours. Statistical differences were evaluated using the scheffe test. The ANOVA procedure of the SAS system. The obtained results were as follows ; 1.The age distribution of normal human was 36.9 +/- 4.4 old, osteoporotic human was 72.5 +/- 0.2 old with statistical significant difference(p<0.05). 2. The quantification of TGF-beta of the normal human serum was 39658.38 +/- 11630.43 pg/ml,the osteoporotic human serum was 30459.40 +/- 1704.92 pg/ml with statistical significant difference. The quantification of PDGF of the normal human serum was 3064.13 +/- 709.51 pg/ml, the osteoporotic human serum was 2514.13 +/- 140.21 pg/ml with no statistical significant difference(p<0.05). 3. The DNA synthesis and protein assay of human osteoblast at 24 hours, 48 hours, 72 hours was similar increased to 10%normal human seurm, 10%osteoporotic human serum.There was no statistical significant difference between the normal human and the osteoporotic patients in 3%normal human PRP and 3%osteoporotic human PRP. 4.The optimal cell concentration was 5 X 10(4)cells/ml among 1 x 10(4)cells/ml, 2.5 x 10(4)cells/ml, 5 x 10(4)cells/ml, and 10 x 10(4)cells/ml.The DNA synthesis was decreased after 72 hours in the normal human serum and PRP,the osteoportic serum and PRP. 5.The alkaline phosphatase activity was as the same result 10%FBS, 10%osteoportic serum and 10% normal human serum at 48 hours with no statistical significant, but the alkaline phosphatase activity was increased in 10%osteoportic human serum and 10%normal human serum except 10% FBS at 72 hours. From above result,the amount of TGF-beta of the normal human growth factor was higher than the osteoporotic patients, but the growth factors of the osteoportic patients were enough the proliferation and differentiation of normal human osteoblasts such like the same effects of normal human growth factors.