Objective To explore the detailed underlying molecular and signaling mechanisms in the effects of icariin on bone formation by an in vitro cell model. Methods The proliferation of MC3T3-E1 osteoblast-like cells was evaluated by MTT, and gene expression of cell cycle related proteins in MC3T3-E1 cells after icariin treatment was detected by real-time PCR. The phosphorylation of MAPK signals, including ERK, P38, and JNK was determined by Western blot, and then the inhibitors of MAPK signals were used to treat cells with icariin alone or together to determine the role of MAPKs in the process of icariin treatment on MC3T3-E1 cell proliferation. Alkaline phosphatase and Alizarin red staining were used to detect the formation of mineralization nodules, and gene expressions of alkaline phosphatase, type Ⅰ collagen, and osteocalcin in osteoblasts after being treated by icariin were evaluated by real-time PCR. ICI182780, and nilutamide was used to decide the participation of estrogen and androgen receptor signals in the process of icariin treatment on the differentiation and mineralization of MC3T3-E1 cells. Results Treatment with icariin promoted MC3T3-E1 cell growth in a time- and dose-dependent manner. This treatment also revealed that icariin increased the expression of mRNAs encoding both cyclin E and PCNA, positive regulators of cell growth, but decreased levels of mRNAs encoding Cdkn2b, a negative regulator of cell cycle progression. When MC3T3-E1 cells were cultured in a differentiated condition, icariin enhanced mineralized nodule formation and increased the expression of mRNAs encoding alkaline phosphatase, type Ⅰ collagen, and osteocalcin. Treatment with icariin significantly induced phosphorylation of both ERK and JNK and this phosphorylated effect occurred very rapidly within 5 minutes and reached peak at 15 minutes. Furthermore, the stimulated effects of icariin on proliferation and gene expression of cyclin E, PCNA, and Cdkn2b in MC3T3-E1 cells were dramatically attenuated by treatment with both U0126 and SP600125, inhibitors of MAPKs. Interestingly, such stimulating effects of icariin were at least partly reduced by treatment with ICI182780, an inhibitor of estrogen receptor. Icariin induced mineralized nodule formation and gene expression of alkaline phosphatase, type Ⅰ collagen, and osteocalcin in MC3T3-E1 cells were also partly reduced when the cells were treated with ICI182780. Conclusions Our findings indicate that the anabolic effect of icariin on bone formation is, at least partly, mediated through the MAPK signaling pathway in order to modulate osteoblast proliferation and differentiation.

Objective To assess the coordinated regulation and the molecular mechanisms of 17β-estradiol and 1,25-dihydroxyvitamin D3 [ 1,25-( OH ) 2 D3 ] on the proliferation and differentiation of osteoblastic MC3T3-E1 cells.Methods MC3T3-E1 cells were cultured in phenol-red free α-MEM medium supplemented with 10％ FBS,MTT assay was performed to determine the effects of 17β-estradiol and 1,25-( OH )2 D3 on MC3T3-E1 cells proliferation.After cells were treated with different agents,cell cycle related genes [ cyclin E,proliferation cell nuclear antigen ( PCNA ),and cyclin-dependent kinase inhibitor 2b ( Cdkn2b ) ] and markers of osteoblastic differentiation [ type Ⅰ collagen ( COL Ⅰ ),alkaline phosphatase ( ALP),osteopontin ( OPN ) ] were detected with SYBR green-based quantitative PCR.ALP activity was detected with BCIP/NBT method.Results 17β-estradiol could promote proliferation of MC3T3-E1 cells,which was accordant to its ability to increase cyclin E and PCNA and to inhibit Cdkn2b mRNA expression in MC3T3-E1 cells.However,1,25-( OH)2D3 had no effect on the proliferation of MC3T3-E1 cells and also did not enhance the proliferation of MC3T3-E1 cells stimulated by 17β-estradiol.On the other hand,17β-estradiol promoted the gene expression of differentiation markers Col Ⅰ,ALP,and OPN,and 1,25-(OH) 2 D3 synergistically increased the expression of these genes with 17 β-estradiol.Conclusion As two of the most important hormones which regulate bone metabolism,estrogen and vitamin D may coordinately promote osteoblast differentiation,but may not regulate osteoblasts proliferation synergistically.

Objective:To investigate the relationship of hypertension with osteoporosis(OP)and bone mineral density(BMD) in elderly type 2 diabetic women.Methods: A total of 179 elderly type 2 diabetic women were categorized into hypertension group(n=124)and non-hypertension group(n=55).Lumbar and hip BMD were measured by dual-energy X-ray absorptiometry.Prevalences of osteoporosis were compared between the two groups using Chi-square test,and BMD was compared using covariance analysis.The correlation between the osteoporosis and risk factors was analyzed using binary logistic regression method.Results: The prevalences of osteoporosis(total OP,lumber OP and hip OP) in non-hypertension group were higher than that of the hypertension group(Pt=0.037,Pl=0.042 and Ph=0.051,respectively).Lumbar and hip BMD in the hypertension group was higher than that in the non-hypertension group,and there were significant differences in L3 and L4 BMDs between the two groups(P3=0.040,P4=0.021);however,the difference disappeared after adjustment for body mass index(BMD).After adjustment for C peptide,the P value also increased,but not as obvious as after adjustment for BMI.Binary logistic regression analysis showed that OP was associated with age,BMI and osteocalcin,but not with hypertension.Conclusion: OP is associated with the age,BMI,and osteocalcin,but not with hypertension in elderly type 2 diabetic women.

OBJECTIVEThis study investigated the effects of hydroxyapatite (HA)/chitosan (CS)-transforming growth factor-β1 (TGF-β1) composite coatings on titanium surfaces, as well as on the attachment and proliferation of osteoblasts.

METHODSHA/CS-TGF-β1 composite coatings were prepared on titanium surfaces by physical, chemical, and biological modifications. Scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transform-infrared spectroscopy (FTIR), and other methods were employed to analyze the chemical composition and surface topography of the composite coatings. CCK-8 and immunofluorescence assays were used to analyze the effects of the coatings on the attachment and proliferation of osteoblasts.

RESULTSHA/CS-TGF-β1 composite coatings were successfully prepared. Their contact angle was almost zero. These composite coatings were applied in vitro, with a drug released early and a burst release effect. The growth of osteoblasts was not inhibited on it and it had obvious promoting effect on the adhesion and early proliferation of osteoblasts.

CONCLUSIONThe composite coatings significantly promote the adhesion and early proliferation of osteoblasts in vitro. This finding shows that the proposed method demonstrates a good prospective application in surface modification of titanium.

Cell Proliferation ; Chitosan ; chemistry ; Dental Bonding ; methods ; Durapatite ; chemistry ; Microscopy, Electron, Scanning ; Osteoblasts ; physiology ; Prospective Studies ; Spectroscopy, Fourier Transform Infrared ; Surface Properties ; Titanium ; chemistry ; Transforming Growth Factor beta ; chemistry ; Transforming Growth Factors ; X-Ray Diffraction

Objective: To discuss the conversion coefficient of morphine injection with continuous intravenous pump delivery or subcutaneous injection for the patients with advanced cancer pain demanding high dose of opioids.Methods: Using a retrospective survey, the patients with advanced cancer pain demanding high dose of opioids with poor efficacy were divided into 3∶1 group and 2.5∶1 group, and the conversion coefficient of 3∶1 or 2.5∶1 was used for the opioids equivalently conversed to intravenous or subcutaneous injection of morphine.After the conversion, the degree of pain relief, the analgesic efficiency in the conversion process, titration time, daily oral morphine equivalent amount at stable pain, morphine related adverse reactions and the other indicators were studied to evaluate the analgesic effect of morphine injection with different conversion coefficient.Results: There was no statistical significant difference between the two groups in the degree of pain relief, the effective rate of analgesia and the daily oral akministration amount of morphine at sable pain(P>0.05).The adjustment times for morphine in the two groups was (1.57±0.93) and (1.0±0.00), respectively, and the difference was statistically significant (P<0.05).The daily oral administration amount of morphine at stable pain in the two groups respectively was (226.67±69.74) mg and (258.67±101.34) mg;the morphine related adverse reactions were mainly constipation, and there was no significant difference in the incidence (P>0.05).Conclusion: Giving morphine injection to the patients with terminal cancer pain demanding high dose of opioids with poor effect, the use of PCA pump through intravenous or subcutaneous injection can effectively relieve pain.Using the conversion coefficient of 2.5:1 can quickly complete the titration process, and safely achieve the effective analgesia.

Objective:To explore the pharmacodynamic differences of india madder root before and after charcoal in antiinflammatory,ease pain,promoting blood circulation for removing blood stasis and hemostasis function.Methods: The india madder root and india madder root charcoal decoction pieces were processed by the same one operation,then the water decoction of them were given to the mouse by intragastric administration in different dosages.The method of auricle tumefaction was adopted to compare the antiinflammatory function,body wrings was adopted to compare the ease pain function,to compare the hemostasis function of india madder root before and after charcoal by snipping off the mouse’s tail and capillary method.Blood stasis model was made by injecting Dexamethasone Sodium Phosphate,then to compare the promoting blood circulation for removing blood stasis function of india madder root before and after charcoal of india madder root before and after charcoal.Results: India madder root decoction pieces is more effective than india madder root charcoal decoction pieces in antiinflammatory,ease pain,promoting blood circulation for removing blood stasis,but less effective in hemostasis.Conclusion: The function of antiinflammatory,ease pain,promoting blood circulation for removing blood stasis of india madder root were less effective after charcoal,but the fuction of hemostasis was more effective.

Objective To investigate the effects of icarrin on the activity and protein expression of core binding factor otl(Cbfa1) in rat osteoblasts cultured in vitro,and to explore whether mitogen-activated protein kinase (MAPK) pathway is involved in this process.Methods Calvarial osteoblasts were obtained from newborn (<24 h) SD rats by trypsin-coUagenase digestion method.The second generation osteoblasts were cultured in the medium containing icariin (10 ng/ml) or estradiol (10-8 mol/L) with or without extracellular-signal regulated kinase (ERK) inhibitor (UO126) or p38MAPK inhibitor (SB203580).Nuclear protein was extracted from osteoblasts.And then the activity of Cbfa1 was detected by ELISA.The amounts of Cbfa1 protein were detected by Western blot.Results Calvarial osteoblasts were obtained successfully and were used in this study after indentified by alkaline phosphatase and mineralized nodus staining.Cbfa1 expression and the activity in osteoblasts were up-regulated by both icariin and estradiol (P<0.05).The effects were partly inhibited by addition of U0126or SB203580 (P<0.05).Conclusions Either icarrin or estradiol can stimulate the proliferation and maturation of cultured osteoblasts in vitro via up-regulating the activity and expression of Cbfal.The MAPK signal pathway inhibitor seems to partly decrease Cbfa1 activity.It suggests that MAPK pathway may be involved in the transduction of icariin's impact on proliferation and mineralization of osteoblasts.

Objective To study the changes of bone mineral density(BMD)and bone turnover in postmenopausal osteoporotic patients treated with salmon calcitonin nasal spray. Methods Sixty-seven postmenopausal osteoporotic patients were enrolled in our trial. All of them received calcium and vitamin D; 37patients were treated with salmon calcitonin nasal spray for 12 months and the other 30 patients received calcium and vitamin D only. Dual-energy X-ray absorptiometry(DEXA)and measurements of a series of bone turnover indices were performed before and after medication for 6 and 12 months. Results After treatment with salmon calcitonin nasal spray for6 months, BMD in lumbar spine 2-4 increased but no change occurred in femoral neck. However, after treatment for 12 months, BMD in both lumbar spine 2-4 and femoral neck increased. In the control group, BMD in lumbar spine 2-4 decreased after treatment for 6 and 12 months, but BMD in femoral neck decreased only after 12months. Comparing with the control group, after treatment with salmon calcitonin nasal spray, BMD in lumbar spine 2-4 and femoral neck were increased obviously. The level of TRACP-5b and NTX/Cr decreased after treatment with salmon calcitonin nasal spray for6 months and 12 months, while BALP increased only after treatment for 12 months. In the control group, BALP decreased after treatment for 12 months. The level of 25-(OH)vitamin D increased after treatment for 6 months and 12 months in both groups. Conclusions Long-term treatment with salmon calcitonin nasal spray prevents bone loss and may increase bone mass.

Objective To evaluate the efficacy and safety profile of S-1 combined with oxaliplatin L-OHP (SOX) in the treatment of locally advanced or metastatic colorectal cancer.Methods 70 patients with advanced or metastatic colorectal cancer were randomly divided into trial group (35 cases) and control group (35 cases).The trail group was administered with dose of 130 mg/m2 L-OHP,plus S-1 which was given orally with body surface area (BSA) (BSA＜1.25 m2,80 mg/d; BSA≥ 1.25 m2 and ＜1.5 m2,100 mg/d; BSA≥ 1.50 m2 and ＜1.8 m2,120 mg/d; BSA＞1.8 m2,140 mg/d).This schedule was repeated every 3 weeks.The control group treated by FOLFOX4 regimen (L-OHP was given on d1 with 80 mg/m2 through intravenous,leucovorin was intravenously on d1,2,with 200 mg/m2,5-Fu was intravenously injected on d1,2,with 400 mg/m2,and was administered intravenously 44 hours with 1 200 mg/m2 on d1).This schedule was repeated every 2 weeks.Results The total clinical effective rate had no significant difference in the trail group and control group (51.4 ％,18/35 vs 45.7 ％,16/35) (x2 =0.229,P =0.632).Toxicity,nausea and vomiting rate in the trail group were lower than those in the control group (48.5 ％,16/35 vs 71.4 ％,25/35,68.6 ％,24/35 vs 88.6 ％,31/35,P ＜ 0.05),but hand-foot syndrome and peripheral neurotoxicity rates had no significant difference between two groups (P ＞ 0.05).Weight increased significantly after chemotherapy treatment in the two groups (t =2.702 5,P =0.003 9).Conclusion SOX regimen is feasible and safe for advanced colorectal cancer.

OBJECTIVETo investigate the expression of PTEN and Caspase-3 in malignant lymphoma of the stomach and explore their role in progression of primary gastric malignant lymphoma.

METHODSFormalin-fixed paraffin embedded tissues from 56 cases of primary gastric malignant lymphoma and their adjacent non-tumor mucosa were evaluated for PTEN and Caspase-3 protein expression by streptavidin-biotin-complex (SABC) immunohistochemistry. Their expression was compared with clinical tumor parameters with the relationship between PTEN and Caspase-3 expression concerned as well.

RESULTSThe positive rate of PTEN expression in primary gastric lymphomas (50.0%, 28/56) was significantly lower than that in adjacent non-tumor gastric mucosa (96.4%, 27/28) (P < 0.05). Meanwhile, 43 of 56 (76.8%) gastric lymphomas indicated Caspase-3 expression, less than that in adjacent non-tumor mucosa (93.5%, 29/31) (P < 0.05). The expression of PTEN was negatively correlated with invasion and lymph node metastasis of gastric lymphoma (P < 0.05), while the Caspase-3 expression was negatively associated with the latter one (P < 0.05). Additionally, the PTEN expression was positively correlated with Caspase-3 expression in the primary gastric malignant lymphoma (P < 0.05).

CONCLUSIONSThe down-regulated expression of PTEN and Caspase-3 played an important role in progression of primary malignant gastric lymphoma. PTEN, as a molecular marker of pathobiological behaviors of tumor, contributes to tumor progression by increasing cell mobility and angiogenesis, as well as decreasing cell adhesion and apoptosis.

Adult ; Aged ; Biomarkers, Tumor ; metabolism ; Caspase 3 ; Caspases ; metabolism ; Down-Regulation ; Female ; Gastric Mucosa ; enzymology ; Humans ; Lymphatic Metastasis ; Lymphoma ; enzymology ; pathology ; Male ; Middle Aged ; Neoplasm Invasiveness ; PTEN Phosphohydrolase ; Phosphoric Monoester Hydrolases ; metabolism ; Stomach Neoplasms ; enzymology ; pathology ; Tumor Suppressor Proteins ; metabolism