[ 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 .

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 .

BACKGROUND:Recent studies have suggested that intranasal administration is one of the ways to target drugdelivery, and can effectively make the drug that cannot pass through the blood brain barrier by other pathways to bypass the blood brain barrier, resulting in targeted delivery to the brain. It provides a promising route for the treatment of central nervous system diseases. OBJECTIVE: To study the pharmacokinetic and brain-targeted channel-tropism tissue distribution character of cimicifugoside H-1 after nasal and intravenous administration in plasma and tissues in rats, in order to evaluate the feasibility of developing brain-targeted nasal delivery system of cimicifugoside H-1 by the passage between nose and brain in nasal olfactory area. METHODS: After intravenous injection and nasal administration of cimicifugoside H-1, the drug concentrations of plasma and channel-tropism organs (lung, spleen, stomach, large intestine, liver, kidney, brain, brain, cerebelum, cerebrospinal fluid, olfactory bulb and olfactory region) were detected. Drug-time curve was drawn. DAS program was used to select apartment model and pharmacokinetics parameters. RESULTS AND CONCLUSION:(1) The pharmacokinetics characters of cimicifugoside H-1 are rapidly absorbed and extensively distribution. Among major channel-tropism organs, drug concentrations were higher in the lung and brain than in the other organs. (2) Cimicifugoside H-1 could be straightly transported into brain by the intranasal administration. The molecule through olfactory mucosa in nasal cavity entered into olfactory bulb in arachno-hypostegal cavity, and then entered into olfactory region, cerebrospinal fluid, cerebrum and cerebelum gradualy. Olfactory bulb was the only way for drug molecule to go through nasal cavity into brain. (3) Compared with the intravenous injection, cimicifugoside H-1 through the intranasal administration has a significant

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.

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: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.

BACKGROUND:Cuscuta chinensis is a mature seed of Cuscutachinensis Lam., can warm kidney. Previous studies demonstrated that kidney compound composed of Cuscuta chinensis could apparently inhibit bone loss and improve bone density.
OBJECTIVE:To explore the influence of flavonoids from Cuscuta chinensis on bone mineral density of femur, 1,25(OH)2D3 levels of serum and kidney, the expression of smal intestine CaBp-D9K mRNA in model rats with ovariectomized osteoporosis.
METHODS:A total of 72 Sprague-Dawley female rats were equal y and randomly divided into six groups (n=12):sham surgery group, model group, vitamin D3 group and low-, moderate-and high-dose flavonoids groups. The sham surgery group only received sham operation and the other five groups were ovariectomized respectively. One week after ovariectomy, the rats were given flavonoids from low-, moderate-and high-dose Cuscuta chinensis and vitamin D3 (2 mg/kg) by intragastric administration for 3 consecutive months. Blood was obtained from the abdominal aorta. Serum was isolated. The kidney was obtained. Enzyme linked immunosorbent assay was used to determine 1,25(OH)2D3 contents of renal and serum. Rats were sacrificed at the end of experiment. The thighbone was taken out to determine bone mineral density. The second lumbar vertebra was taken out to measure the expression of lumbar vertebra and renal vitamin D receptor mRNA using real-time fluorescent reverse transcription-polymerase chain reaction. The smal intestine was taken out to measure the expression of smal intestine CaBp-D9K mRNA using real-time fluorescent reverse transcription-polymerase chain reaction.
RESULTS AND CONCLUSION:Compared with the sham operation group, bone mineral density of femur, and 1,25(OH)2D3 levels of serum and kidney and the expression of lumbar vertebra vitamin D receptor mRNA significantly decreased in model group, and the expression of smal intestine CaBp-D9K mRNA significantly decreased in model group. Compared with the model group, bone mineral density of femur, and 1,25(OH)2D3 levels of serum and kidney, and the expression of the second lumbar vertebra vitamin D receptor mRNA, and the expression of smal intestine CaBp-D9K mRNA were increased in moderate-and high-dose flavonoids groups and vitamin D3 group. Results indicated that flavonoids from Cuscuta chinensis could significantly raise bone mineral density of femur, and 1,25(OH)2D3 levels of serum and kidney and the expression of lumbar vertebra vitamin D receptor mRNA and the expression of smal intestine CaBp-D9K mRNA, accelerate intestinal calcium absorption and osteoblast activity, and reinforce quality of the bone.

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.

OBJECTIVETo study the relationship between tissue quantitative distribution and pharmacokinetics of 3H-achyranthes bidentata ecdysterone and the channel-tropism of herbal drugs in mice.

METHOD3H-achyranthes bidentata ecdysterone was used as a tracer agent and injected into mice by the caudal vein. In 36 hours, the contents of the tracer agent of samples involving 9 different tracing phases and organ or tissue were determined in order to observe the dynamic quantitative distribution and excretion and pharmacokinetics of 3H-achyranthes bidentata ecdysterone and to understand the channel-tropism of herbal drugs achyranthes bidentata.

RESULT3H-achyranthes bidentata ecdysterone of same organs in different tracing phases and the contents of 3H-achyranthes bidentata ecdysterone in same tracing phases of different organs were significantly different (P<0.01). 3H-achyranthes bidentata ecdysterone was mainly distributed, in the liver, kidney, adrenal gland, small intestine and lung. The concentration-time profiles of achyranthes bidentata ecdysterone in rats injected into mice by the caudal vein were shown to fit a two-compartment open model with half-lives of (778.65 +/- 12.36) min, the elimination of achyranthes bidentata ecdysterone from plasma was found to be in accord with linear kinetics.

CONCLUSIONThe above mentioned selective distribution of 3H-achyranthes bidentata ecdysterone basically coincides with the meridian affinity and zang fu selection of the traditional Chinese medicine drug Achyranthes bidentata. This study will provide a scientific basis for the channel-tropism of A. bidentata.

Achyranthes ; chemistry ; Animals ; Drugs, Chinese Herbal ; metabolism ; pharmacokinetics ; Ecdysterone ; metabolism ; pharmacokinetics ; Isotope Labeling ; Male ; Meridians ; Mice ; Organ Specificity ; Tissue Distribution ; Tritium ; chemistry

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.