Objective To characterize the effects of progesterone(P) on the expression of membrane type 1 matrix metalloproteinase (MT1 MMP) and gelatinase A activation in human osteoblast like cell strain MG 63 in order to understand the mechanism of progestin acting on the bone. Methods Semi quantitative RT PCR, Western blot, and confocal immunohistochemistry were used to study the actions of P on the expression of mRNA and protein of the MT1 MMP in cultured MG 63 cells. Zymogram of the gelatin and ELISA were performed to examine the effects of P on the activation of gelatinase A. Results Treatment with increasing dose of P in MG 63 cells caused a dose dependent increase in the expression of MT1 MMP mRNA ( P 0 05). Conclusions Our studies suggested that P might promote the bone formation by increasing MT1 MMP production in osteoblastic cells and thus played an important role in maintaining the osteogenic activity of osteoblasts and preparing the bone matrix for the subsequent bone cell migration and the deposition of new matrix.

The effect of 17?-estradiol ( 17?-E2 ) on expression of estrogen receptor ?( ER?) was observed in cultured MC63 cells and human osteoblast-like (HOB) cells. The results show that the expression of ER* is in a dose-dependent manner with 17p-E2 (0-1?10-6 mol/L) in MC63 cells and has an optimal 17?-E2 concentration (1?10-8 mol/L) in HOB cells resulting in maximum expression of ERp protein.

Objective To observe the effect of 17? estradiol (E 2) on the expression of membrane type 1 matrix metalloproteinase (MT1 MMP) in human osteosarcoma cell line MG 63. Methods The expression of MT1 MMP protein in MG 63 cells were assayed with Western blot and immunohistochemical method. The activity of MMP 2 was determined by gelatin zymogram and ELISA. Results Treatment with different concentrations of E 2 in MG 63 cells resulted in a dose dependent enhancement in expression of MT1 MMP protein (P

Objective To study the changes of microarchitecture, bone mineral density (BMD) , and bone mineral content (BMC) in apolipoprtein E knockout( ApoE-/-) mice. Methods Male ApoE-/- mice at 15, 28, and 40-week of age and sex-age-matched wild-type (WT) mice were involved in the study. The trabecular and cortical bone microarchitecture were assessed by micro-CT( μCT) in the right distal femur. The total body BMD of the left femur was determined by dual-energy X-ray absorptiometry ( DXA). The relationships among BMD, microarchitecture, and BMC were analyzed. Results Compared with WT mice,the advancing age ApoE-/- mice showed an increased volumetric BMD (vBMD), tissue BMD (tBMD) , BMC, bone volume fraction (BV/TV), trabecular number (Tb. N ) , trabecular thickness (Tb. Th) with an decreased bone surface fraction ( BS/BV), trabecular separation (Tb. SP) , and the structure mode index (P <0. 05 ) in the cancellous bone of femur. The cortical bone microarchitecture parameters as inner perimeter, outer perimeter, cortical area, marrow area, total area and moment of inertia were also increased, but cortical BMD, cortical bone mineral content (C. BMC) and cortical thickness retained constant. At the age of 28 weeks,the total body BMD in ApoEE-/- mice revealed higher than WT mice (P<0. 05) and there was no changes in 15 and 40-week-old mice compared with the sex-age-matched controls. vBMD was positively correlated with BMC, BV/TV,Tb. Th, BS/BV, and C. BMC, with the correlation coefficients 0.955,0.944,0. 834,0.923, and 0.903 .respectively, and there was no correlation between vBMD and the other parameters. Conclusions ApoEE-/- mice display an increased bone mass, suggesting that ApoE has an important role in bone remodeling.

BACKGROUND:Estrogen/progestins replacement therapy prevents excess bone loss in postmenopausal women.Recently osteoprotegerin (OPG) has been identified in osteoblast and displayed to inhibit bone resorption.OBJECTIVE: To compare the action between 17β-estradiol (E2) and progesterone on OPG expression in cultured normal human osteoblast-like cells (hOB).DESIGN: A comparative investigation.SETTING: Institute of Metabolic Endocrinology, the Second Xiangya Hospital of Central South University.MATERIALS: α-MEM (Sigma Chemical Corp., St. Louis, MO, USA); Type Ⅳ collagenase (Sigma); Fetal bovine serum (Gibco-BRL Corp., Grand Island, NY, USA); Osteocalcin radioimmunoassay kit (DiaSorin Corp., Stillwater, MN, USA).METHODS: The experiments were carried out in the Institute of Metabolic Endocrinology, Second Xiangya Hospital of Central South University from January 2003 to March 2006. The osteoblasts were extracted from the cancelous bone of anterior superior iliac spine of normal people, then cultured. The hOB were treated with E2 and progesterone, and the expressions of OPG mRNA and OPG protein were determined by Northern blot analysis and enzyme-linked immunoabsorbent assay (ELISA) respectively.MAIN OUTCOME MEASURES: ①Characterization of human osteoblast-like cells; ②Effect of E2 and progesterone on OPG mRNA levels by Northern blot analysis; ③ Effect of E2 and progesterone on OPG protein levels in the conditioned medium by ELISA.RESULTS: ① Characterization of hOB in vitro The ALP levels in normal human osteoblasts were (74.3±4.7) U/g protein,and the detectable osteocalcin levels was (3.84±0.39) μg/L protein], which suggested that osteoblasts were the primary cell type found in our bone-derived cell cultures from donors. ② Effects of E2 and progesterone on the levels of OPG mRNA by Northern blot analysis: The OPG mRNA band was week in the control group [(12.3±3.5)%], treatment with 1 × 10-10, 1 ×10-9 1 ×10-8 mol/L E2 caused an increase in the levels of OPG mRNA. The expression of OPG mRNA in the 1×10-8 mol/L E2 group was gradually increased at 12, 24 and 48 hours. Progesterone had no influence on OPG mRNA expression. ③ Effects of E2 and progesterone on OPG protein production in conditioned medium determined with ELISA:ELISA revealed that treatment with 1 ×10-10, 1 ×10-9, 1 ×10-8 mol/L E2 induced obvious increase in the levels of OPG protein in cells media as compared with that in the control group [(1.27±0.26), (2.34±0.35), (3.62±0.23), (0.64±0.14)μg/L, P ＜ 0.01]. In the presence of 1×10-8 mol/L E2, OPG protein production in cells media at 12, 24 and 48 hours were significantly higher than that in the control group [(1.30±0.30), (3.07±0.14), (3.50±0.33), (0.62±0.12) μg/L, P ＜ 0.01]. 1 × 10-10, 1 ×10-9 1 × 10-8 mol/L progesterone had no influence on the OPG protein production after 12-24 hours (P ＞ 0.05).CONCLUSION: The different regulation of OPG production in osteoblasts by E2 and progesterone may contribute to the mechanisms by which estrogen or progestins exerts its different action on bone resorption.

Objective To investigate the signal transduction pathways related to 17?-estradiol-induced membrane type-1 matrix metalloproteinase (MT1-MMP) expression in human osteosarcoma cell line MG63. Methods MG63 cells were incubated with ICI 182 780 for 48 h, then exposed to different concentrations of ICI 182780, 17?-estradiol, and specific antagonists H-89, H-7, PD98059, SB203580, genistein and PTDC of protein kinase (PK) A、PKC、extracellular signal-regulated kinase (ERK)、p38MAPK、protein tyrosine kinase (PTK) and nuclear factor (NF)-?B. The MT1-MMP level was determined by Western blotting and immunohistochemistry. Results Western blotting showed that MT1-MMP level was increased in 17?-E_2 group and decreased in PDTC and genistein groups, and immunohistochemistry revealed that MT1-MMP was located in cytoplasm and cellular membrane, but not in nucleus. Conclusion 17?-estradiol regulates MT1-MMP expression mainly through NF-?B and PTK pathways in MG63 cells.

Objective To determine the relationship between serum testosterone level and lean body mass, body fat content, and bone mineral density (BMD) . Methods The study involved 185 healthy females in Changsha, aged 45 ～81. Fasting serum testosterone was measured by radioimmu-noassay. Hologic QDR 4500A fan beam X-ray bone densitometer was used to measure the BMD of anteroposterior lumber (AP, L_(1～4)) and total hip, to measure the bone mineral content, BMD, body fat content and muscle tissue weight of head, trunk, ribs, pelvis, spine, upper limbs, lower limbs and the total body. Body weight, lean body mass and body fat percentage were calculated. SPSS 11.0 software was used to conduct regression analysis. Results (1) Serum testosterone showed no correlation with lean body mass, body fat content, and body fat percentage. (2) Serum testosterone was positively related with the BMD of lumbar spine and hip, but showed no correlation with the BMD after adjustment of age and years since postmenopause. (3) Lean body mass showed significant positive correlation with the BMD of different sites. Total body fat content showed positive correlation with the BMD of total hip, while body fat percentage showed negative correlation with the BMD of the whole body. Conclusion Keeping lean body mass benefits postmenopausal women to maintain bone mineral content, and taking androgen should still be cautious.

Serum osteoprotegerin (OPG) concentration was measured in 647 healthy female adults (aged 20-81 years), and was compared with that of other races. The serum OPG was positively correlated with age (r = 0.276, P <0.01). The geometric mean value of serum OPG in premenopausal women was significantly lower than those in perimenopansal and postmenopausal women. The serum OPG in middle-aged Chinese women was signifieandy higher than those in middle-aged Austrian and Icelandic, but this was quite contrary to the results obtained in old-aged women.

This in vitro study demonstrated that taurine supplemented culture medium enhanced alkaline phosphatase (ALP) activity, osteocalcin secretion and mineralized matrix formation. Taurine induced activation of ERKI/2 and osteoblast differentiation, which was blocked by pretreatment of osteoblasts with ERKI/2 inhibitor (PD98059), suggesting taurine stimulated osteoblast differentiation via ERKI/2.

Objective To investigate the mechanisms of action of adiponectin on receptor activator of NF-Kb ligand(Rankl) and osteoprotegerin (OPG)expressions in human osteoblasts.Methods Real-time PCR was used to detect the expressions of RANKL and OPG mRNA in cultured human osteoblasts. The phosphorylations of JNK, p38 mitogen-activated protein kinase (MAPK) , ERK1/2 were assayed by Western blot. RNA interference for adiponectin receptor, MAPK inhibitors SB203580 and SP600125 were used for elucidating the mechanism of the action of adiponectin in regulating OPG and RANKL expressions. Results Suppression of adiponectin receptor-1 (AdR1) expression with siRNA abolished the adiponectin-regulated expressions of OPG and RANKL mRNA in human osteoblasts. Furthermore, pretreatment of osteoblasts with MAPK inhibitor SB203580 abolished the expressions of adiponectin-regulated RANKL and OPG mRNA, but SP600125 did not show the effect. Conclusion Adiponectin induces the expression of RANKL and inhibits the expression of OPG in human osteoblasts through AdR1/p38 MAPK pathways.