The proliferating cell nuclear antigen (PCNA) gene expression was blocked and retinal pigment epithelium (RPE) proliferation was inhibited by using antisense oligonucleotides (AS-ODN) mediated by liposome, to find a new genetic therapy of proliferative vitreoretinopathy (PVR). RPE cells cultured in vitro were transfected with synthetic fluorescence labled AS-ODN mediated by liposome-Lipofectamine, and the intracellular distribution and persistence time of AS-ODN were dynamically observed. AS-ODN (0.07, 0.28 and 1.12 micro mol/L and sense oligonucleotides (S-ODN with the same concentrations as AS-ODN) mediated by liposome were delivered to the RPE cells cultured in vitro, and CPM values were measured by 3H-TdR incorporation assay and analyzed statistically by variance by comparison with blank control group. Expression of PCNA mRNA in RPE cells was detected by in situ hybridization after the treatment of different concentrations of PCNA AS-ODN and S-ODN, and the average optic density (AOD) was measured by image analysis system and was subjected to q-test and correlation analysis with CPM. Our results showed that AS-ODN mediated by liposome could quickly aggregate in cellular plasma and nuclei in 30 min and 6 h, and stayed for as long as 6 days. AS-ODN (0.28 and 1.12 micro mol/L) markedly suppressed proliferation of RPE cells in a dose-dependent manner with the difference being statistically significant (P < 0.05 and P < 0.01, repectively) as compared with blank control group. AOD was well correlated with CPM (r = 0.975). It is concluded that liposome could increase transfection efficiency of AS-ODN in RPE cells, and AS-ODN could sequence-specifically suppress PCNA mRNA expression and proliferation of human RPE cells.

AIM: To evaluate the effects and safety of intravitreal injection of triamcinolone acetonide ( TA ) or conbercept combined with macular laser grid photocoagulation in the treatment of macular edema secondary to retinal vein occlusion( RVO) . ·METHODS: Fifty cases ( 50 eyes ) with macular edema secondary to retinal vein occlusion were selected and assigned to 2 groups: intravitreal injection of TA or conbercept, and laser photocoagulation after 7d. Best corrected visual acuity ( BCVA ) , fundus examination, optical coherence tomography ( OCT ) and intraocular pressure ( IOP ) were examined before intravitreous injection and 14d, 1 and 3mo after laser, fundus fluorescein angiography(FFA) were examined 3mo after treatment. The postoperative results at each time point were compared with preoperative values. · RESULTS: Two kinds of treatment compared with preoperative, the BCVA all increased in various degrees. At 14d after intravitreous injection, 1 and 3mo after laser, the ratio of vision improved in TA group was 76%, 80%, 68%, conbercept group was 88%, 92%, 88%, BCVA of two groups in each period all had varying degrees of increase than preoperative. The best BCVA acquired at 1mo after treatment. The macular thickness after treatment was significantly lower than preoperative in two groups. At preoperative, 14d, 1 and 3mo after treatment, the macular thickness in TA group was 557. 5 ± 150. 9,301. 7±120. 1, 262. 7 ± 131. 2, 338. 1 ± 146. 5μm; the macular thickness in conbercept group was 569. 4 ± 135. 9, 282. 3 ± 133. 5, 259. 5 ± 116. 4, 307. 8 ± 122. 6μm. The macular thickness of the two groups were significantly different between preoperative and postoperative. · CONCLUSION: The combination of intravitreous injection of TA or conbercept with macular laser grid photocoagulation can be an effective method in the treatment of macular edema secondary to RVO, conbercept treatment is more effective and security.

OBJECTIVETo explore the effect of combined application of psoralen (PSO, an extract from psoralea) and ultraviolet A (UVA) for inducing the apoptosis of human leukemic cell line NB4 and its impact on the Fas/FasL gene expressions.

METHODSAccording to factorial design, changes of apoptosis rate and ultrastructure of NB4 cells, as well as the gene and protein expressions of Fas/FasL were observed after cells were treated with PSO in different concentrations and irradiated by UVA of 360 nm wavelength for different times, using flow cytometry, transmission electron microscopy and quantitative polymerase chain reaction (PCR), and the outcomes were treated with variable analysis.

RESULTS(1) After treatment of PSO in concentration of 10, 20, 40, 80 microg/mL combined with a 5-min exposure of UVA, the NB4 cells apoptosis rate induced were 26.57% +/- 0.42%, 30.67% +/- 0.11%, 34.90% +/- 0.30% and 24.63% +/- 0.38% respectively. The effects were dose- and time-dependent, and an interaction was shown between the two actors. (2) After being treated by PSO plus UVA, obvious ultrastructure changes with apoptosis characteristics were shown in NB4 cell under electron microscope. (3) PSO plus UVA showed up-regulatory effect on gene and protein expressions of Fas, and down-regulatory effect on gene and protein expressions of FasL in a dose- and time-dependent manner, with the interaction between the two actors in altering Fas gene expression, also in altering FasL gene and protein expressions.

CONCLUSIONCombined application of PSO and UVA can induce the apoptosis of NB4 cells, and the Fas/FasL system is one of the pathways for apoptosis inducing.

Apoptosis ; drug effects ; radiation effects ; Cell Line, Tumor ; Fas Ligand Protein ; metabolism ; Ficusin ; pharmacology ; Gene Expression ; Gene Expression Regulation, Leukemic ; Humans ; Ultraviolet Rays ; fas Receptor ; metabolism

With the development of modern imaging technologies,three-dimensional (3D) reconstruction techniques based on the computed tomography (CT) or magnetic resonance imaging (MRI) data has been gradually applied in clinical diagnosis and treatment of hepatobiliary diseases.In order to study the characteristics of the 3 D model of portal branches and collateral circulation in various types of portal hypertension,the data of 64-slice spiral CT scan of5 patients with portal hypertension were collected and imported to the abdominal medical image three-dimensional visualization system (MI-3DVS) for sequence segmenting and 3D reconstruction,and then the results of 3D models were compared with operation findings.The 3D models were vividly,visually and clearly displayed the portal system and the collateral circulation,which is helpful in the diagnosis and treatment of the portal hypertension.

Objective To study the medical image three dimensional visualization system(MI-3DVS)in the diagnosis and treatment of hepatolithiasis.Methods The data of 64-slice spiral computed tomography of 54 patients with hepatolithiasis who were admitted to the Zhujiang Hospital of the Southern Medical University from August 2008 to August 2010 were collected.The liver and bile duct were three dimensionally(3D)constructed.Preoperative diagnosis and pathological classification were made according to the results of the 3D model of liver and bile duct.The optimal surgical procedure was determined by simulating operations based on the 3D model.The compliance of simulated operation and actual operation was observed,and residual stones were detected by cholangiography.Results Of the 54 patients,11 were with type Ⅰ,5 with type Ⅱ(including 2 patients with type Ⅱ a and 3 with Ⅱ b),38 with type E.There were 23 patients with intrahepatic bile duct stricture and 27 with atrophy-hypertrophy syndrome complex.The anatomy of intra-and extrahepatic bile duct,dilation and stricture of the bile duct,site,size and number of the bile duct stones were clearly displayed in the MI-3DVS.The compliance rate of simulated surgery and actual surgery was 94%(51/54).There was no residual stones in 51 patients who received elective surgery and the rate of residual stone of the 54 patients was 6%(3/54).Conclusion Acurate preoperative diagnosis and intraoperative precise operation can be achieved and the rate of residual stone can be reduced by using the MI-3DVS.

BACKGROUND:It is reported that bone marrow mesenchymal stem cell(BMSC)transplantation might be a promising treatment for liver fibrosis.But the mechanism is still unclear.OBJECTIVE:To observe the hepatic stellate cells apoptosis induced by BMSC transplantation,and to study the mechanism of BMSC in treating hepatic fibrosis in vivo.METHODS:CCl_4 subcutaneous injection was performed to induce rat liver fibrosis.After 8 weeks of CCU injection,20 rats which underwent successful model establishment were randomly divided into experimental group and control group,10 in each group.The experimental group received MSC transplantation via tail vein injection,and the control group were given DMEM instead.The rats were killed and the livers were harvested at three time point,the day of MSC transplantation,3 days after transplantation,and 7 days after transplantation.The hydroxyproline content was detected by HE and Masson staining,and the expression changes of α-smooth muscle actin(α-SMA)proteins were determined using immunohistochemistry.The apoptosis of hepatic stellate cells were determined by α-SMA and TUNEL(terminal dUTP nick-end labeling)dual-staining.RESULTS AND CONCLUSION:After 8 weeks of CCU injection,the hydroxyproline content increased and histology indicated progress of liver fibrosis.At 7 days after MSC transplantation,the hydroxyproline in the liver was decreased,and the liver fibrosis was alleviated in the experimental group but aggravated in the control group.Immunohistochemistry indicated that α-SMA positive cells were increased at 8 weeks after CCU injection.At day 7 after transplantation,α-SMA positive cells in the experimental group were significantly less than control group(P ＜ 0.05).At 3 days after transplantation,the hepatic stellate cells apoptosis in the experimental group was significantly aggravated compared with control group(P ＜ 0.05).This suggested that MSC transplant was an effective treatment for liver fibrosis.MSC inducing hepatic stellate cells apoptosis may be one of the mechanisms.

Objective To study the pathophysiology of ankylosing spondylitis (AS)-related osteoporosis by investigating the relation between bone mineral density (BMD) and bone turnover markers.Methods Fortysix AS patients were included in this study,including 35 male and 11 female patients.Twenty-five gender and age matched healthy subjects were served as the healthy control group.Informed consents were obtained from all subjects.BMD of lumbar spine (L2-4),fermoral neck and radius were tested using dual-energy X-ray absorption method.Data about BASDAi,ESR,Ig was collected.Bone formation markers including procollagen type Ⅰ N-terminal peptide (PINP) and osteocalcin (OC) were included for analysis,the formal was measured by radioimmunoassay and the later was measured by immunoradiometric assay and bone resorption marker.β-Crosslaps was measured by electrochemiluminescence immunoassay.Independent-samples t test was used to compare normal distributed data between the two groups.Analysis of variance was used to compare the data between the three groups.For those data which were not normally distributed,Rank sum test was performed.Correlations were determined by Pearson or Spearman's ranking.Results ① 48％ of AS patients had bone loss,while the percentage in the healthy control group was 20％ (x2=5.32,P=0.021).BMD of lumbar spine,fermoral neck and radius of the AS patients were lower than the controls (t=-3.73,-3.68,-5.24; P＜0.05).② Bone density (BMD) of lumbar spine in patients at the late stage was higher than patients at early disease stage (t=2.26,P=0.034),however,the BMD in the femoral neck was not.③ The procollagen (PINP),OC and β-CrossLaps were not significantly different in patients at different stage of diseases (P＞0.05).④BMD of lumbar spine was negatively correlated with the β-CrossLaps and ESR(r=-0.325,P=0.047; r=-0.314,P=0.046),The BMD in fermoral neck was negatively correlated with β-CrossLaps and OC (r=-0.387,P=0.024; r=-0.371,P=0.034),but they were not correlated with disease duration and BADSAI.Conclusion Osteoporosis is common in AS.Bone turnover markers including bone formation and bone resorption are increased in AS.β-CrossLaps is likely to predict bone loss in AS.

Objective: To observe the effect of acupuncture plus thunder-fire moxibustion on the expressions of matrix metalloproteinase-3 (MMP-3), tissue inhibitor of metalloproteinase-1 (TIMP-1) and transforming growth factor-β1 (TGF-β1) in cartilage of knee osteoarthritis (KOA) rats, and to explore the mechanism of acupuncture plus thunder-fire moxibustion in the treatment of KOA. Methods:Thirty Sprague-Dawley (SD) rats were randomly divided into a blank control group, a model group and an acupuncture-moxibustion group by random digits table, 10 rats in each group. Rats in the model group and the acupuncture-moxibustion group were injected with papain in the right posterior knee joint to prepare the models. The levels of MMP-3 and TIMP-1 in rat synovium of each group were measured by enzyme-linked immunosorbent assay (ELISA) after 2 weeks of treatment. The level of TGF-β1 was determined by Motic B5 Micro-camera system. Results:The levels of MMP-3 and TIMP-1 in the cartilage of the model group were significantly higher than those in the blank control group (allP<0.01); the levels of MMP-3 and TIMP-1 in the acupuncture-moxibustion group were lower than those in the model group, and the between-group differences were statistically significant (all P<0.05). The levels of MMP-3 and TIMP-1 in the acupuncture-moxibustion group were higher than those in the blank control group, and the differences were statistically significant (all P<0.05). The level of TGF-β1 in cartilage tissues of the model group was significantly lower than that in the blank control group (P<0.01); the level of TGF-β1 in the acupuncture-moxibustion group was higher than that in the model group (P<0.05), but it was lower than that in the blank control group, and the between-group difference was statistically significant (P<0.05). Conclusion: Acupuncture plus thunder-fire moxibustion can effectively recover the abnormal expressions of MMP-3 and TIMP-1 in KOA model rats and somewhat up-regulate TGF-β1, which may be one of its mechanisms of acupuncture plus thunder-fire for KOA.

Objective Despite regular treatment,antineutrophil eytoplasmie antibodies(ANCA)asso- ciated systemic vasculitis(AASV),in which the role of Fc?Rs has been established,are still associated with significant long-term mortality and remain an important cause of end-stage renal failure.ANCA plays an im- portant role in the pathogenisis of primary systemic small vessel vasculitis(PSV)by their potential to activate neutrophils.Because the interaction between ANCA and its receptors on the Fc portion of immunoglobulins (Fc?R)on neutrophils is essential in the activation process,we investigate the inhibitory,effect of tg19320 on ANCA induced activation of neutrophils,which is a tetrameric tripeptide that interferes with IgG/Fe?Rs in- teraction.Methods We prepared tg19320 by solid-phase peptide syntbesis.The binding between tg19320 and human IgG was assessed by enzyme-linked immunosorbent assay.The biological activity of tg19320 to intefere with FcF?receptor recognition was identified by rosette formation assay.ANCA IgG was prepared from the sera of active Wegener's granulomatosis(WG)and microscopic polyangiitis(MPA)patients.Neu- trophils isolated from the blood of healthy volunteers were primed with TNF-?(2 ng/ml)and then incubated with ANCA IgG(200?g/ml),or pretreated with tg19320(2.5 mg/ml)and then added with ANCA IgG.Su- peroxide burst of neutrophils was determined by Ferri-cytochrome reduction assay.Results We found that tg19320 bound tightly to human IgG in a dose dependent manner and the inhibition of the rosette formation between SRBC-IgG and U937 cells was statistically significant(20.3% vs 53.2%,P

?ig-h_3 was first identified as a transforming growth factor-beta1-inducible gene in human lung adenocarcinoma cell line. It encodes for a secreted extracellular matrix (ECM) protein, which is thought to act on cell attachment and ECM composition. Previous study showed that ?ig-h_3 were highly expressed in human hepatoma cell lines and lowly expressed in human normal hepatic cells. The present study aimed to transfect ?ig-h_3 into 7721 cells to investigate its effect on secretion of MMPs in the transfected human hepatoma cells. Full-length ?ig-h_3 gene,cloned by reverse transcription polymerase chain reaction (RT-PCR) was inserted into the eukaryotic expression vector pEGFP-C_2. The recombinant plasmid was transfected into 7721 cells with Lipofectamine2000 and Gelatin-Zymography were adopted to detect the production of MMPs in the transfected cells. Results showed that ?ig-h_3/pEGFP-C_2 recombinant expression plasmid was successfully constructed and achieved high transfection efficiency. MMPs expression of the transfected cells was promoted significantly. These results suggest that overexpression of ?ig-h_3 promoted the production of MMPs, indicating that ?ig-h_3 may play roles in the invasive and metastatic processes of hepatoma.