Objective We used the DNA methylation microarrays to investigate the differential methylation genes and loci sites in Kashin-Beck disease (KBD),to study the relationship between DNA methylation and KBD pathogenesis.Methods Totally 12 KBD adults and 12 healthy adults were selected and peripheral blood samples were collected and DNA was extracted.Illumina 450K bead-chip was applied to detect methylation status in KBD and healthy controls.Aberrant hyper-methylated sites were filtrated according to the P value after correction and methylation differences,together with GenomeStudio soft.Screened genes were validated using bisulfite sequencing polymerase chain reaction (BSP) technology.Results A total of 484 948 loci sites were analyzed and compared,93 differential methylated loci were found by comparing KBD and normal people,including 34 hypermethylated sites and 59 hypomethylated sites.There were 50 genes corresponding to the loci,43 genes not reported in literature.According to gene ontology analysis,the genes were involved in the immune response,antigen processing,phosphate and phosphoric acid metabolism and phosphorylation and the process of metal ions in combination.However,in the verification test using BSP method,there was no significant difference in methylation rate in human leukocyte antigen (HLA)-DRB1 between the case and the control group (48％ vs 70％,x2 =3.688,P ＞ 0.05).Conclusions The high and low differentially methylated sites in peripheral blood DNA of KBD patients are significantly different from those of the health control.HLA-DRB1 locus is not significantly different between the BSP verification test and methylation chip.

Transforming growth factor-β is an important cytokine with various bioactivities, including embryonic development, wound healing, chemotaxis and cell cycle regulation. Epithelial-mesenchymal transition (EMT) is the main pathway of tumor cell to obtain the ability of invasion and metastasis. The TGF-β is the key factor known to induce EMT in cancer cells and plays an important role in the process. In recent years, some progress has been obtained. Some TGF-β inhibitors have approved in the market or in clinical trials. TGF-β inhibitors can play an important role on the treatment of tumors, glaucoma, liver and kidney fibrosis disease and scar repair. Novel TGF-β inhibitors reported in recent years were reviewed in this article.
Epithelial-Mesenchymal Transition ; Humans ; Neoplasms ; Transforming Growth Factor beta ; antagonists & inhibitors ; Wound Healing

BACKGROUND: It has reported that nicotinamide is capable of protecting intervertebral disc (IVD) against interleukin-1β (IL-1β) or tumor necrosis factor-alpha (TNF-α) induced degeneration. However, the protective mechanism of nicotinamide on IVD cells apoptosis and proliferation remains unclear.OBJECTIVE: To investigate regulatory effects of nicotinamide on rabbit nucleus pulposus cell apoptosis and proliferation in vitro.DESIGN, TIME AND SETTING: Randomized control grouping design, which was carried out in the Laboratory of Orthopaedics and Stem Cell Center, Union Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology from April to October 2007.MATERIALS: Ten Japanese white rabbits (aged 2-3 months weighing 1.5-2.0 kg) were used in this study. Furthermore, nucleus pulposus cells obtained from L1-6 lumbar spine were harvested and cultured for further experiments.METHODS: The NP cells were divided into 6 groups, including control group (without any drug as control), nicotinamide group (0.5 g/L nicotinamide), IL-1β group (10 μg/L IL-1β), IL-1β + caspase group (10 μg/L IL-1β and non-specific caspase inhibitor Z-VAD-FMK), IL-1β + small-dose nicotinamide group (10 μg/L IL-1β and 0.05 g/L nicotinamide), and IL-1β + large-dose nicotinamide group (10 μg/L IL-1β and 0.5 g/L nicotinamide). After 3 days of culture, the cells were examined with Annexin V-PI staining, caspase-3, 8 and 9 activity staining and MTT assay.MAIN OUTCOME MEASURES: The apoptotic rates, the positive rates of caspase-3, 8 and 9 activity staining and the absorbance of MTT assay of each group.RESULTS: ① As compared to IL-1β group, the apoptotic rates were decreased in the IL-1β + caspase group and IL-1β + large-dose nicotinamide group (P < 0.01). ②As compared to IL-1β group, the positive rates of caspase-3, 8 and 9 activity staining were decreased in the IL-1β + caspase group, IL-1β + large-dose and small-dose nicotinamide groups (P < 0.01 or P < 0.01). ③As compared to IL-1β group, the absorbance was increased in the IL-1β + caspase group and IL-1β + large-dose nicotinamide group (P < 0.01).CONCLUSION: Nicotinamide is capable of promoting cell proliferation and inhibiting IL-1β induced apoptosis of nucleus pulposus cells in vitro. The inhibition of apoptosis mainly acts via inhibition of the mitochondrial pathway.

Objective To explore the effect of exogenous adrenomedullin(ADM) on expressions of glutathion in plasma and brain tissue inneonatal rats with hypoxic-ischemia reperfusion brain damage(HIRBD) and the mechanism of action.Methods Fifty-six cases of 7 d SD rats were randomly divided into 4 groups including normal control group(without any treatment),HIRBD group(the model with hypoxia for 2 h and ischemia for 1 h),primed group(abdomen infusion of ADM at 0.5 h before making model,the other was same to the HIRBD group)and treatment group(abdomen infusion of ADM at onces after making model,the other was same to the HIRBD group).The neonatal rats in 4 groups were derived blood and brain tissue after decapitation at either 4 h or 24 h after reperfusion.The levels of gtutathion(GSH) in plasma and brain tissue were determined by using chromatometry.Results In HIRBD group,the affection areas at 24 h after reperfusion enlarged compared with those at 4 h after reperfusion.Meanwhile,the affection of punctiform degeneration or necrosis at 4 h after reperfusion transformed into the affection of lamellar or diffuse degeneration or necrosis at 24 h after reperfusion.The levels of GSH in plasma and brain tissue in HIRBD group at either 4 h or 24 h after reperfusion were significantly lower than that in normal control group(Pa0.05).Meanwhile the pathology score of brain section in primed group and treatment group were significantly lower than that in HIRBD group at either 4 h or 24 h after reperfusion(Pa0.05).Conclusion Exogenous ADM can induce the neuroprotection in HIRBD by adjusting the expression of GSH.

OBJECTIVE:To optimize the formulation of Methotrexate thermosensitive hydrogels,and to study the in vitro re-lease property of it. METHODS:Taking PLGA-PEG-PLGA as matrix and mannitol as viscosity modifiers,Methotrexate thermosen-sitive hydrogels were prepared. Using the absolute value of the deviation of gelation temperature between 35 ℃,its formulation was optimized by orthogonal test using the concentrations of PLGA-PEG-PLGA,methotrexate and mannitol as factors. The dialysis bag method was used to investigate accumulative release rate of Methotrexate thermosensitive hydrogels and methotrexate solution within 336 h in vitro. RESULTS:The optimal formulation was as PLGA-PEG-PLGA of 20%,methotrexate of 0.10% and mannitol of 0.10%;gelation temperature were 35.1,35.0,35.0℃. The average drug-loading rate was more than 90%. 12 d accumulative re-lease rate was more than 90%(r=3). Methotrexate solution has been substantially completely relased within 240 h. CONCLU-SIONS:Methotrexate thermosensitive hydrogels with sustained-release are prepared successfully,and the preparation technology is simple and practical.

Objective To prepare triamcinolone acetonide acetate(TAA)thermosensitive hydrogel for intra-articular injection,and to investigate its release in vitro. Methods TAA suspending thermosensitive hydrogel was prepared by using poly lactic-co-glycolic acid(PLGA)-polyethylene glycol(PEG)-PLGA as gel matrices,xanthan gum as suspending agent and NaCl as flocculant. It was evaluated preliminarily by determining its contents and its in vitro release. Results The best compositions for preparation of TAA suspending thermosensitive hydrogel were 25% PLGA-PEG-PLGA,0.05% xanthan gum, 0.9% NaCl and 4 mg·mL-1 TAA.The gelation temperature was 35.3 ℃ .The average recovery was(98.98±0.31)%. By the method of membraneless dissolution,the accumulative drug release was up to 83. 31% at 16 days. The drug release followed Ritger-Peppas methematical model. Conclusion The TAA thermosensitive hydrogel with obviously sustained release is expected to become a new drug delivery system for intra-articular injection.

Objective:To investigate the radiosensitizing effect of three flavonoids on ovarian cancer SKOV3 cells under hypoxia. Methods:The SKOV3 cells were divided into normoxic group and hypoxic group. The hypoxic SKOV3 cellular model in vitro was es-tablished and tested by measuring the expression profile of HIF-1αprotein in SKOV3 cells. Colony-forming assay was used to detect the radiosensitivity of normoxic and hypoxic SKOV3 cells. The cytotoxicity and radiosensitizing effects of flavonoids were evaluated on the basis of cell death and MTT assay. Results:The Western blot results showed that the gray intensity ratio of HIF-1α/β-Actin in hypoxia group was significantly higher than that in normoxia group (1. 068>0. 117). Radiosensitivity of hypoxic SKOV3 cells in hypoxia group was significantly lower than that in normoxia group. The survival rate of SKOV3 cells was decreased with the increase of concentration. When the concentration was increased, D0 and Dqin chrysin group and quercetin group were significantly decreased (P<0. 01), and SER was increased (P<0. 05). SER showed no significant difference in breviscapine group (P>0. 05). The overall radiobiological parameters of hypoxia group were higher than those of normoxia group. Conclusion: Hypoxia can induce the expression of HIF-1α in SKOV3 cells, which results in the decrease of radiosensitivity. Chrysin and quercetin can enhance the radiosensitivity of SKOV3 cells, and the enhancement is significant under hypoxia, while breviscapine is without such effect. The radiosensitizing effect may be achieved by the level decrease of HIF-1α in SKOV3 cells and inhibition of DNA damage repair.

Objective To investigate surgical technique and clinical efficacy of Sky bone ex- pander kyphoplasty in the treatment of osteoporotic vertebral body compression fractures.Methods Eighteen cases with osteoporotic vertebral body compression fractures were treated with Sky bone expander kyphoplasty from August 2004 to November 2005.Under the local anesthesia,3.5-5ml of bone cements were injected into each pathologic vertebral body through unipedicle approach after reduction procedure was done with Sky bone expander.Results The postoperative follow-up ranged from 3 to 11 months, with an average of 4.5 months.Back pain was effectively relieved after the operation in all cases.No complications occurred.Conclusion The Sky bone expander kyphoplasty has the advantages of safe- ty,easy operation,minimal invasion,effective restoration of the vertebral body height and fast relief of pain.

BACKGROUND:Recent studies have demonstrated that Niacinamide is capable of promoting the proliferation of intervertebral cells and improving intervertebral disc degeneration.Overloading is thought to the main cause of intervertebral disc degeneration.However,the protective effects of Niacinamide in loading induced intervertebral disc degeneration remains uncertain,OBJECTIVE:To investigate the protective effects of Niacinamide against axial loading induced degeneration of rabbit lumbar disc.DESIGN,TIME AND SETTING:A randomized controlled experiment was carded out in the Central Laboratory and the Laboratory of Department of Orthopaedics,Union Hospital Affiliated to Tongji Medical College,Huazhong University of Science and Technology from November 2008 to April 2009.MATERIALS:Twenty-four Japanese white rabbits (4 months old,weighing 2.0 kg).Niacinamide was supplied by Tianjin Damao Chemical Reagent Factory.METHODS:Twenty-four Japanese white rabbits were randomly divided into 6 groups.The controllable axial loading induced rabbit lumbar disc degeneration model was adopted to impose 98N pressure on the rabbit discs to induce degeneration.Various doses of Niacinamide were given intragastrically to the rabbits in different groups:2 rabbits in group 1,the loading device was installed without pressing,and no Niacinamide was given;2 rabbits in group 2,given 50 mg/kg Niacinamide for 1 week;5 rabbits in group 3,loaded with 98N for 1 week;5 rabbits in group 4,loaded with 98N for 1 week,then the pressure was released for another week's recovery;5 rabbits in group 5,loaded with 98N and given 50 mg/kg Niacinamide for 1 week;5 rabbits in group 6,loaded with 98N for 1 week and then the pressure was released for another week's recovery,50 mg/kg Niacinamide was continually given during the 2 weeks.MAIN OUTCOME MEASURES:Magnetic resonance image and Thompson's grading system were used to assess degeneration degree of the discs;hematoxylin and eosin staining,immunohistochemical staining for type Ⅱ collagen,and Safranin O staining were used to evaluate histological changes;immunohistochemical staining for P161NK4A was used to evaluate cell proliferation and senescence.RESULTS：①According to the Thompson's grading system，there was no disc exhibited degeneration in group 2；5 rabbits graded Ⅱ in group 3；4 rabbits graded Ⅱ and 1 rabbit graded Ⅲ in group 4；2 rabbits graded Ⅰ and 3 rabbits graded Ⅱ in group 5；3 rabbits graded Ⅰ and 2 rabbits graded Ⅱ in group 6.MRI results revealed the alleviated degeneration in Niacinamide given groups.②The content of type Ⅱ collagen of annulus fibrosus of group 6 was 53.2% higher than that of group 4 (P＜0.01).③Safranin O-Fast Green staining density of group 2 was higher than that of group 1；The staining density of nucleus pulposus and annulus fibrosus of groups 5 and 6 was higher than the corresponding parts of group 4，especially that of nucleus pulposus (P＜0.01，P＜0.01)，and group 6 exhibited slightly increased levels than group 5.④P16INK4A positive staining rates decreased with the extension of Niacinamide administration time.CONCLUSION：Niacinamide can help to alleviate overloading-caused damage to intervertebral disc，and can benefit the recovery of damaged intervertebral disc.

OBJECTIVE:To compare the efficacy,safety,vascular endothelial growth factor(VEGF)and matrix metallopro-teinase-2 (MMP-2) of docetaxel combined with carboplatin and paclitaxel combined with cisplatin (DDP) in the treatment of ad-vanced ovarian cancer. METHODS:120 patients with advanced ovarian cancer were randomly divided into docetaxel combined with carboplatin group(60 cases)and paclitaxel combined with DDP group(60 cases). Docetaxel combined with carboplatin group received 70 mg/m2 Docetaxel injection,intravenous infusion of 1 h,d1;50 mg/m2 carboplatin injection,intravenous infusion of 1 h,d2. Paclitaxel combined with DDP group received 135 mg/m2 Paclitaxel injection,intravenous infusion of 24 h,d1;30 mg/m2 DDP for injection,intravenous infusion,d3;60 mg/m2 Paclitaxel injection (a maximum of 2.0 m2) by intraperitoneal infusion,d8. 3-week was regarded as 1 treatment course,and it lasted 6 courses. Clinical efficacy,VEGF,MMP-2,progression-free survival, overall survival before and after treatment,mortality rate within 2 years of treatment and the incidence of adverse reactions in 2 groups were compared. RESULTS:There were no significant differences in the objective response rate,disease control rate,mortal-ity rate,incidence of adverse reactions between 2 groups(P>0.05). The progression-free survival in docetaxel combined with car-boplatin group was significantly longer than paclitaxel combined with DDP group,the difference was statistically significant (P<0.05). Before treatment,there were no significant differences in VEGF and MMP-2 level between 2 groups(P>0.05). After treat-ment,VEGF and MMP-2 level in 2 groups were significantly lower than before,and VEGF at different time points and MMP-2 level after 4 weeks,8 weeks and 12 weeks of treatment in docetaxel combined with carboplatin group were lower than paclitaxel combined with DDP group,the differences were statistically significant(P<0.05). CONCLUSIONS:Docetaxel combined with car-boplatin and paclitaxel combined with DDP shows similar efficacy and safety in the treatment of advanced ovarian cancer,but docetaxel carboplatin combined with is superior to paclitaxel combined with DDP in reducing VEGF and MMP-2 and improving pro-gression-free survival.