| Objective: The rate of diabetic osteoporosis is increasing with the improvement of people's life quality. Loss of bone mineral content has been recognized as one reason of the type 1 diabetic osteoporosis. Osteoporosis is a bone condition defined by low bone mass,increased fragility, decreased bone quality, and an increased fracture risk. The mechanism of bone loss in type 1 diabetes is still unknown, although osteopenia is a recognized complication of diabetes mellitus which could be due to abnormal bone turnover or disturbances in the calcium/parathyroid hormone/vitamin D axis or insulin deficiency or both..Osteoclasts are one of the most highly specialized cells in the human body, whose sole function is to degrade bone. We investigated the effect of ovariectomy and type 1 diabetes on the derivation of bone marrow stem cells to osteoclast-like cells(OLC) in rats by bone marrow cells culture in vitro, the effect of alendronate on osteoclasts was also studied in vitro. At the celluar level, we tried to find pathogenesis of osteoporosis in type 1 diabetic osteoporosis.Methods: Sixty 2.5 to 3-month-old female Wistar rats were divided into six groups: normal control group(NC),normal sham-ovariectomy group (NS),normal ovariectomy(NOVX), type 1 diabetes control group (DC), type 1 diabetes sham-ovariectomy group (DS), type 1 diabetes ovariectomy group (DOVX).Each normal group contained 8 rats, each group of type 1 diabetes contained 12 rats. No statistically significant difference was found between any groups in body weight. Rats in type 1 diabetes groups were administered a single celioinjection of streptozotocin(STZ,60mg/kg body weight) to induce type 1 diabetes, while animals in normal groups were given a sham injection of citrate buffer. The levels of blood glucose were determined 2 days after the injection of STZ or vehicle, and rats with blood glucose levels ≥16.7 mmol/L were used as diabetic rats. Rats in group NOVX and group DOVX were subjected to be ovariectomized, while rats in group NS and group DS subjected to sham-operation. All rats were allowed free access to food and water. On the day of surgery (day 0),2 rats were killed and served as baseline.2 weeks,4 weeks and also 8 weeks after ovariectomy, body weight and blood glucose were measured in every group. At the same time,6 rats chosen from 6 groups were killed, femur bone marrow were collected and cultured, They were cultured in the presence of macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor kappa-B ligand(RANKL),alendronate was added to culture medium to the final concentrations of 10-7mol/L. Bone ends of rat were cut off and marrow cavity flushed with (-MEM .The marrow cells were washed twice with (-MEM and counted. 1.5(106cells/ml in 24-well-plate were cultured in (-MEM, containing 10%FCS and 30ng/ml sRANKL,10ng/ml M-CSF.Fresh medium every second day. Cultures were stopped at day 7 by fixation and staining.The OLC formed are multinucleated, TRAP-positive containing three nuclei or more. The number of OLCs was measured under light microscopy. The experimental data was demonstrated in mean±standard deviation and the paired-samples t test and analysis of variance were used to the significance test and the relation about the data was measured with linear correlation by SPSS 10.0,a statistic software.Results: During the animal experiment, the blood glucose in diabetic groups was always higher than that in normal groups(p<0.01),but there was no significant difference between diabetic groups (p>0.05).It was found that rats with type 1 diabetes had significantly lower body weight than in healthy controls(p<0.01).At the end of 8 weeks experiment, the body weight of group NOVX(282.00±21.49) and group DOVX(200.00±10.32) increased more significant than group NC(239.60±10.11,p<0.01),NS(241.67±12.86,p<0.01) and group DC(171.33±10.44, p<0.01), DS(170.40±9.84,p<0.01). All the numbers of OLCs in normal and in type 1 diabetes rats rose time-dependent in the experiment of 8 weeks. In the whole group... |
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