Objective To explore the effect of total flavonoids of Epimedium sagittatum (TFE) on the mRNA expressions of the estrogen receptor alpha (ERα) and estrogen receptor beta(ERβ) in hippocampus and hypothalamus in ovariectomized (OVX) Sprague-Dawley(SD) rats, and the mechanism of TFE against postmenopausal osteoporosis. Methods Forty-eight female SD rats,aged 10-11 months old, were randomly divided into 4 groups: a sham group, an ovariectomy group (rats were bilaterally ovariectomized), a TFE group, and a 17β-estradiol group (rats were fed with TFE and 17β-estradiol for 4 months, respectively). The RT-PCR was performed to determine the mRNA expression of ERα and ERβ in hypothalamus and hippocampus. Results Serum estradiol level, bone mineral density (BMD) of vertebra,wet weight of uterus, and the mRNA expressions of ERα and ERβ in hypothalamus and hippocampus were markedly decreased in OVX rats, all of which were reversed by 17β-estradiol treatment except the mRNA expression of ERβ. Similar results were achieved by TFE treatment except the wet weight of uterus. Conclusion TFE can improve BMD of vertebra in the OVX rats without side effects on the uterus. The mechanism may be related to increasing the mRNA expressions of ERα and ERβ in hypothalamus and hippocampus.

Objective To investigate the relationship between the Schatzker types and postoperative knee function in tibial plateau fractures .Methods Totally 120 patients with tibial plateau fracture from January 2010 to December 2014 were selected and performed the Schatzker classification according to the X‐ray film ,CT and three dimensional reconstruction .They were divided into 6 groups .The postoperative knee function in each group was followed up and the statistical analysis was conducted .Results The excellent grade distribution of knee function by Kruskal‐Wallis test had statistical difference among 6 groups(P<0 .05) ,in the fur‐ther pairwise comparison ,the significant differences were found between the group 1 and 2 with the group 4 ,5 and 6(P<0 .05) . Conclusion The Schatzker types of tibial plateau fractures is closely correlated with the postoperative knee function .The knee function scores of type Ⅳ ,Ⅴ and Ⅵ are obviously poor .

Aim To investigate the effect of total flavonoids of Epimedium sagittatum(TFE)and its metabolites on the proliferation and function expression of newborn rats calvarial osteoblasts(ROB).Methods TFE was supplemented into the culture medium of ROB at 0.2,2,20,100 and 200 mg?L-1 respectively.The serum of rats administered TFES(SRAT) was also added into the medium in a parallel treatment at 2%,4%,8% and 16% respectively.Their effects on cell proliferation and function expression were studied by MTT and the analysis of osteogenic differentiation marks.Results TFE had no effect on cell proliferation and function expression at any concentration.However,2% and 4% SRAT stimulated cell proliferation and,4% SRAT promoted the maturation and function of osteoblast by improving the alkaline phosphatase(ALP) activity,bone gla protein(BGP) secretion,calcium deposition and the number of mineralized nodular structures.Conclusions The effective substance of herba epimedii treating on osteoporosis is not TFE itself but certain of metabolites derived from TFE and the products induced by these metabolites in serum.The metabolites in serum of TFE may enhance the cell function expression and proliferation.

BACKGROUND:Animal and cel studies have shown that the Kidney Recipe can prevent and treat osteoporosis and improve bone metabolism, but this recipe is complicated. Recent studies on compound Chinese medicine mainly focused on serum drug metabolism and pharmacokinetics, which has limitations, and the effective ingredient and pharmaceutical material basis are uncertain. OBJECTIVE:In the different concentrations and time, by using different compatibility proportion of active ingredients of Kidney Recipe, osteoblasts from neonatal Sprague-Dawley rats were subjected to culture intervention. The proliferation and differentiation of osteoblasts were determined so as to identify time-effect and dose-effect relationship of Kidney Recipe on osteoblasts and to provide experimental evidences for prevention and treatment of osteoporosis. METHODS:Primary neonatal 24-hour osteoblasts of Sprague-Dawley rats were cultured in vitro. Herbs“tonic”and“cathartic”active chemical components of different proportion were used. The experiment contained three groups:Tonics Medicine (T)>Cathartic Medicine (C) group, TC group and TC group (P<0.05). At mass concentration of 20μg/L, no significant difference was detected in the T>C and TC group (P<0.05), and the proliferation became weak at 72 hours. At mass concentration of 10μg/L and 72 hours of culture, alkaline phosphatase secretion could be promoted in the T>C group. At mass concentration of 20μg/L and 24 hours of culture, the promoting effect on alkaline phosphatase secretion was most significant in the TC group (P<0.05). Results confirm that active ingredients of Kidney Recipe compatibility proportion can promote osteoblasts proliferation and differentiation most strongly in the T>C group in the time-effect and the dose-effect relationship.

Aim To investigate the effect of catalpol from Radix rehmanniae on the proliferation,differentiation and matrix mineralization of MC3T3-E1 cells in vitro.Methods Catalpol from Radix rehmanniae of different concentration preparations were extracted with ethanol(catalpol ethanol-extract),respectively.A mouse osteoblast cell line MC3T3-E1 was used to determine the potency of catalpol.MTT were applied to determine proliferation of the cell treated by catalpol at different concentrations.Differentiating effects of the catalpol with different concentrations in the cell were evaluated through the examinations of alkaline phosphatase(ALP)activities and bone gla protein(BGP)levels.Von kossa staining method was used to demonstrate the effects of the catalpol on calcification of the cells.Results Catalpol at concentration from 1×10~(-7) to 1×10~(-9) mol·L~(-1) for 24 hours and 48 hours effective promoted the proliferation of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10~(-5) to 1×10~(-6) mol·L~(-1) for 48 hours and 72 hours effective stimulated the activity of ALP of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10~(-5) to 1×10~(-6) mol·L~(-1) for 8 days and 12 days effective increased the synthesis and secretion of bone gla protein(BGP) of osteoblasts of MC3T3-E1 cells line.Catalpol at concentration from 1×10~(-5) to 1×10~(-6) mol·L~(-1) for 19 days effective increased the mineralized bone nodular structure number of osteoblasts of MC3T3-E1 cells line.Conclusion Catalpol could promote proliferation and differentiation of MC3T3-E1 cells in vitro.Catalpol may be one of effective monomer of Radix rehmanniae on treatment of osteoporosis.

BACKGROUND:Chinese nourishing kidney herbs can prevent osteoporosis and improve bone metabolism, which has been proved in animal and cel experiments. But there are few reports on the compatibility of Chinese nourishing kidney herbs, and it is difficult to screen the optimal compatibility, as the interaction of active ingredients and drug substance basis are uncertain. OBJECTIVE: To determine the proliferation, differentiation and Smad4 mRNA expression of neonatal Sprague-Dawley rat osteoblasts cultured by Chinese nourishing kidney herbs with different compatibility so as to find out the optimal compatibility of Chinese nourishing kidney herbs. METHODS: Passage 5 osteoblasts were divided into five groups: group A, 1×10-5mol/L icarin; group B, 1×10-5mol/L icarin+1×10-5 mol/L naringin; group C, 1×10-5mol/L icarin+1×10-5 mol/L diosgenin; group D, 1×10-5mol/L icarin+ 1×10-5mol/L catalpol; group E, 10 μL normal saline (control group). There were six wels in each group. RESULTS AND CONCLUSION: Compared with the group E, the proliferative ability of osteoblasts and expression of Smad4 mRNA were increased in the groups B and C; until the 72nd hour, the proliferative ability of osteoblasts in the group B reached the peak. At 48 hours of culture, the activity of alkaline phosphatase in groups B and C was higher than that in group E; at 72 hours of culture, the activity of alkaline phosphatase in groups B and D was higher than that in group E. These findings indicate that the compatibility of Chinese nourishing kidney herbs can influence the activity of osteoblasts, and icarin+naringin has the strongest effect.

BACKGROUND: In the research, the recipe for kidney tonification is prepared into "topical form" and "oral form". According to the interrelationship of "acupoints-meridans and collaterals-internal organs-target organs", osteoporosis is treated by the approaches of point compress and oral application respectively. According to the traditional theory of Chinese medicine, it is to infer that the herbs bring the adjustment into play by"being attributive to kidney meridian", but,whether does the specialty of herbs on "targeting medication" happenpractically or not? Hormones in "hypothalamus-pituitary- target gland" system is assayed to further analyze estrogen receptor and androgen receptor of target organs,such as kidney,bone, uterus, thyroid gland and testis so as to observe the relativity between "tropism and receptor".DESIGN: Complete random design with the objects of osteoporosis rats, and the control experimental study with kidney tonification recipe administrate with different approaches.OBJECTIVE: To probe into the effect of kidney tonification recipe on pituitary-thyroid axis in experimental osteoporosis.SETTING: College of Traditional Chinese Medicine of Hebei Medical University.MATERIALS: The experiment was performed in Chinese-Western integrated Basic Experiment Room of Hebei Medical University from January 2000 to December 2004. Sixty SD healthy female rats of 3-month old were employed,weighted(300 ± 20) g. The experimental animals were provided by Hebei Experimental Animal Center. After bred routinely for 1 week in Experimental Room, the rats were randomized into 6 groups,namely normal control(10rats), pathological model group(10 rats), infusing group(10 rats) in which kidney tonification recipe was applied based on 0. 8 g/100 g body mass,acupoint paste on bladder meridian group (10 rats), acupoint paste on kidney meridian group(10 rats) and paste on non-meridian group(10 rats). METHODS: Osteoporosis model was prepared with injection of dexamethasonc. Antagonism osteoporosis acupoint paste(AOAP)on acupoints and non-neridian places was administrated to compare with oral recipe for kidney tonification in the treatment of osteoporosis. It took the changes in serum estrogen, bone density, calcitonin/parathyroid hormone(CT/PTH), thyrotropic hormone(TSH), freeT3andfreeT4asthemeasuringindexes. Immunocyto-chemistry stain and morphological measurement of TSH cell,follicular adenoma of thyroid gland(C cell) and thyroid globulin(Tg) were applied in pituitary gland and thyroid gland to observe the therapeutic results.MAIN OUTCOME MEASURES: ① Adjustment of the recipe for kidney tonification on thyroid function. ② Adjustment of the recipe for kidney tonification on bone density.RESULTS: AOAP improved the levels of estrogen, CT/PTH, TSH, free T3and free T4, increased bone density. The counts of TSH cell, C cell and Tg positive reaction substance were increased and they were colored deeply.CONCLUSION: AOPA, by "multiple relevant convergence reaction", adjusts and affects systematically the endocrinal function of bone metabolism,improves the levels of estrogen and CT/PTH, reverses the declining tendency of pituitary-thyroid axis function and improves thyroid function so as to weaken osteocyte in the bony absorption, enhance osteoblast in osteogenesis and increase bone density.

[ ABSTRACT] AIM:To investigate the relationship between morphological changes of autophagy and apoptosis in the PC12 cells induced by oxygen-glucose deprivation and reoxygenation .METHODS:The PC12 cells were randomly di-vided into normal control group , oxygen-glucose deprivation and reoxygenation group , autophagy inhibitor group and auto-phagy activator group .The cells in oxygen-glucose deprivation and reoxygenation group , autophagy inhibitor group and au-tophagy activator group were exposed to reoxygenation (12 h) after 3 h of oxygen-glucose deprivation, and autophagy inhib-itor 3-methyladenine and autophagy activator rapamycin were added into the cells at the same time .Using transmission elec-tron microscope and monodansylcadaverine fluorescence staining , the morphological changes of autophagosome were ob-served.The apoptosis of the PC12 cells were analyzed by flow cytometry with Annexin V-FITC/PI staining and TUNEL method.RESULTS: Compared with normal control group , the numbers of autophagosomes and the apoptotic rates in-creased in oxygen-glucose deprivation and reoxygenation group (P<0.05).Compared with oxygen-glucose deprivation and reoxygenation group , the numbers of autophagosomes decreased obviously ( P<0.05 ) and the apoptotic rates increased markedly in autophagy inhibitor group (P<0.05).The numbers of autophagosomes increased obviously (P<0.05), the apoptotic rates decreased markedly ( P<0.05 ) , the autophagosomes became bigger in size , and autolysosomes was also found in autophagy activator group .CONCLUSION: Oxygen-glucose deprivation and reoxygenation induce autophagy in PC12 cells, and autophagy inhibits cell apoptosis to play a protective role .

Aim To investigate the effects of astragalo-side IV on apoptosis of PC12 cells inducedby hypoxia/hypoglycemia and reoxygenation. Metheds PC12 cells were randomly divided into 4 groups: normal control group,hypoxia/hypoglycemia and reoxygenation group, astragaloside Ⅳ group and vehicle group. Hypoxia/hy-poglycemia and reoxygenation group, astragaloside Ⅳgroup and vehiclegroup were exposed to reoxygenation (12 h) after 3 h of oxygen and glucose deprivation, and astragaloside Ⅳ was added into cells at the same time. Inverted microscope was used to observe the morphological changes ofPC12 cells and MTT method to detect the activities of PC12 cells, and Annexin V-FITC/PI assay and TUNEL staining were used to meas-ure the apoptosis of PC12 cells. Results Compared with normal control group, cells became round or swol-len and its cellula processes were retracted or disap-peared in hypoxia/hypoglycemia and reoxygenation group;a large number of apoptotic cells could also be observed,whose nucleus were shrinkaged, fragmented or deep-stained. The activities of hypoxia/hypoglycemia and reoxygenation group were decreased markedly than those in normalcontrol group(P<0. 05),while the ap-optotic rates of hypoxia/hypoglycemia and reoxygen-ation group were increased obviously than those in nor-malcontrol group( P<0. 05 ) . Compared with hypoxia/hypoglycemia and reoxygenation group, a good cell growth state could be observed and cellula processes could also be observed significantly in astragaloside Ⅳgroup. The activities of astragaloside Ⅳ group were in-creased than those in hypoxia/hypoglycemia and reoxy-genation group(P<0. 05),while the apoptotic rates of astragalosideⅣgroup were decreased than those in hy-poxia/hypoglycemia and reoxygenation group ( P <0. 05 ) . There was no obvious difference between vehi-clegroup and hypoxia/hypoglycemia and reoxygenation group( P >0. 05 ) . Conclusion Astragaloside IV can reduce the damage of PC12 cells induced by hypoxia/hypoglycemia and reoxygenation, increase cell activity and inhibit cell apoptosis.