OBJECTIVETo study the killing effects of the liposome-mediated thymidine kinase (TK)/ganciclovir (GCV) system on MG-63 osteosarcoma (OS) cells and its bystander effects.

METHODSLiposome-mediated TK gene transfected into MG-63 OS cells, the efficiency of transfection was analyzed by flow cytometry and observed under inverted fluorescence microscope. Non-transfected osteosarcoma MG-63 cells were divided into three groups,in the experimental group 1 transfected TK/GCV cells cultured in solutiona liquid mixture by supernatant by 1/10,1/7,1/5,1/2 ratio to original broth; in the experimental group 2 transfected cells cultured in solutiona liquid mixture of supernatant filtered through 0.22 microm filter by 1/10,1/7, 1/5, 1/2 ratio to original broth, in control group the transfection cells cultured in original culture solution. Cell growth inhibition rate and osteosarcoma cell sensitivity to TK/GCV system were measured by MTT assay in each group.

RESULTSThe TK gene was transfected into MG-63 OS cells successfully by liposome-mediated, flow cytometry instrument detection TK gene transfection cell transfection efficiency can reach 75.5%. Six days later the MTT assay showed that in the experimental group 1 inhibition rate of all concentration ratio of the mixed culture fluid were statistically significant as compared with the control group (P < 0.05), and in the experimental group 2 that of the 1/10 and 1/7 of concentration ratio of mixed culture medium was not statistically significant as compared with the control group (P > 0.05). TK gene transfected MG-63 cells increased with the the GCV concentration,the cell apoptosis rate increased.

CONCLUSIONThe experiment demonstrated that the MG-63 OS cells are sensitive to the liposome-mediated TK/GCV system and bystander effects are significant.

Apoptosis ; drug effects ; Bone Neoplasms ; enzymology ; genetics ; physiopathology ; Bystander Effect ; drug effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; Ganciclovir ; toxicity ; Humans ; Osteosarcoma ; enzymology ; genetics ; physiopathology ; Thymidine Kinase ; genetics ; metabolism ; toxicity

Objective To investigate the efficacy and safety as well as the effects of rituximab on B-lymphocytes and anti-platelet glycoprotein-specific antibodies,in patients with steroid-resistant idiopathic thrombocytopenic purpura(ITP).Methods Twelve steroid-resistant ITP patients,16 to 54 years old,received intravenous rituximab at the dose of 375 mg/m2 once-weekly for 4 weeks.Lab studies included CBC,serum concentrations of IgG,IgM and IgA.CD+3,CD+4,CD+8,CD+19,CD+20 cell numbers were assayed by flow cytometry and anti-platelet glycoprotein-specific antibodies(GP Ⅱ b/Ⅲ a,GP Ⅰ b/Ⅸ)were assayed by monoclonal antibody-specific immobilisation of platelet antigens prior to and following rituximab therapy.Results A complete response(platelet counts ≥100×109/L)was observed in 4 cases,a partial response (platelet counts between 50 and 100×109/L)in 3 cases,a minor response(platelet counts between 30 and 50×109/L)in 2 cases,and non response(platelet counts＜30×109/L)in 3 cases.Responses were sustained 0.5 to 12 months(median 5 months).After 4 weeks of rituximab therapy,anti-platelet glycoprotein-specific antibodies(GP Ⅱ b/Ⅲ a,GP Ⅰ b/Ⅸ)disappeared except one NR patient and CD+19/CD+20 cells were almost depleted in all patients(295.0±86.4)×106/L vs(4.1±2.2)×106/L(P＜0.01).As expected,the T cell counts,and the serum concentrations of IgG,IgM and IgA were not changed after therapy.No severe side effects were observed.Conclusion Rituximab may be an effective and safe treatment for adults with steroid-resistant ITP.

Objective To estalish an accurate method for determining the content of astragaloside Ⅳ in Shouqing Granula. Method TLC scanner was selected to detect astragaloside Ⅳ with silica gela thin layer. The sample was separated by using chloroform-methanol-water (13 : 6 : 2) below 10 ℃ overnight with the under layer solution, ?_s=530 nm, ?_R=700 nm. Results The linear ranges of astragaloside Ⅳ was 0.22～ 0.73 ?g, the average recovery was 97.72%, RSD=0.69%. Conclusion The method was simple and accurate, and can be used for the quality control of Shouqing Granula.

OBJECTIVE To investigate the neuroprotective effect and possible mechanisms of lute-olin-7-O-β-D-glucuronide (LGU) against focalcerebral ischemic injury. METHODS The focal cerebral ischemic injury model was established by middle cerebral artery occlusion (MCAO). Male Sprague Dawley rats were randomly divided into sham group,model group(MCAO),LGU group(0.24,0.72 and 2.16 mg·kg-1)and positive control group(Edaravone at 5 mg·kg-1).LGU was injected intravenously 30 min after MCAO.Neurological severity score,infarct volume and brain water content were detected 24 h after MCAO and the levels of Na+-K+ATPase,Ca2+ATPase,TNF-α and IL-1β were detected to explore the possible mechanisms.For the therapeutic time window test,LGU(0.72 mg·kg-1)was injected intrave-nously 0.5, 2, 4, 6, 8, 10 and 12 h respectively after MCAO. To evaluate motion behavior, LGU were injected intravenously 30 min after MCAO and once per day during detection period. The changes of motor coordination were detected by rotating rod method and grip strength analysis, and the changes of gaits were detected using DigiGait Imaging System. RESULTS LGU improved the neurological severity score, infarct volume ratio and brain water content. The therapeutic time window of LGU for cerebral infarction and brain edema was at least 6 h and for neurological dysfunction was 12 h.LGU also prolonged the latency on rotarod, increased the forelimb tension and improved 8 gait parameters, including stance duration,stride length,stance width,paw area,paw area variability,gait symmetry,ataxia coefficient and tau propulsion.Furthermore,LGU increased Na+-K+-ATPase and Ca2+-ATPase levels in the cortex and hippocampus in the ischemic side,reduced the levels of TNF-α and IL-1β in the serum. CONCLUSION LGU has a significant neuroprotective effect against cerebral ischemic injury via improving energy metabolism and reducing inflammation.

OBJECTIVETo investigate the relationship of virological breakthrough and production of neutralizing anti-interferon antibody (NAb) in chronic hepatitis B patients treated with recombinant interferon-alpha (rIFN-alpha).

METHODFour hundred eighty-five patients with histological proven chronic hepatitis B were treated with 5 MU recombinant interferon-alpha 1b (rIFN-alpha1b) thrice weekly for 6-37 months (median 10). Serum HBV DNA, HBeAg and NAb levels of the patients were detected by fluorescent-quantitative PCR, enzymoimmunoassay and antiviral neutralizing biological assay respectively during the therapy.

RESULTSVirological breakthrough occurred in 66 patients (13.6%), and NAb was found in 98 patients (20.2%) of the total 485 patients. The rate of NAb positivity was higher in patients with viral breakthrough than those without it (68.2%, 45/66, vs 12.6%, 53/419, chi(2)=109.06, P < 0.01), and viral breakthrough occurred more in patients with positive NAb than with negative NAb (45.9%, 45/98, vs 5.4%, 21/387, chi(2)=109.06, P < 0.01). The time of the viral breakthrough occurrence and the time of NAb production had a significant correlation (P < 0.01). The occurrence of viral breakthrough was also influenced by the age of patients (P < 0.05) and HBeAg status (P < 0.01) before they were treated.

CONCLUSIONViral breakthrough occurred in 13.6% of our 485 chronic hepatitis B patients treated with recombinant interferon-alpha. Their viral breakthrough and production of NAb production had a significant correlation.

Adult ; Antibodies, Neutralizing ; biosynthesis ; Female ; Hepatitis B Antibodies ; biosynthesis ; Hepatitis B virus ; immunology ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Interferon Type I ; therapeutic use ; Male ; Recombinant Proteins ; Young Adult

OBJECTIVETo study the causes of poorer antiviral response to neutralizing anti-interferon-alpha antibodies (NA) in male chronic hepatitis B patients treated with recombinant interferon-alpha (rIFN-alpha).

METHODSTwo hundred sixty-nine patients (198 males and 71 females) with histologically proven chronic hepatitis B were treated with 5 MU recombinant interferon-alpha 1b (rIFN-alpha 1b) subcutaneously thrice weekly for 6-37 (median 10.0) months. For each patient, serum HBV DNA levels were detected with fluorescent-quantitative PCR, HBeAg with enzymoimmunoassay, and NA with an antiviral neutralizing biological assay during therapy.

RESULTSNA was found in 70 (35.4%) of the 198 males and in 15 (21.1%) of the 71 females during treatment (x2 = 4.894, P = 0.027). At the end of treatment combined-response was achieved in 21 (24.7%) of the 85 NA-positive patients and in 100 (54.3%) of the 184 NA-negative cases (x2 = 20.642). Stratification analysis by NA showed that combined-response rate was significantly lower in males than in females (18.6%, 13/70 vs. 53.3%, 8/15, x2 = 8.024) among NA-positive patients while it was similar in males and in females (50.8%, 65/128, vs. 62.5%, 35/56, x2 = 2.156) among NA-negative patients. In stratification analysis by gender, it was significantly lower in NA-positive patients than in NA-negative ones (18.6%, 13/70 vs. 53.3%, 8/15, x2 = 8.024) among males but there was no significant difference between combined-response rates among females.

CONCLUSIONThe poorer antiviral response to recombinant interferon-alpha in male chronic hepatitis B patients than in female patients is related to the neutralizing anti-interferon antibodies.

Antibodies ; blood ; Antiviral Agents ; immunology ; therapeutic use ; DNA, Viral ; blood ; Female ; Hepatitis B, Chronic ; drug therapy ; immunology ; Humans ; Interferon Type I ; immunology ; therapeutic use ; Male ; Neutralization Tests ; Recombinant Proteins ; Sex Factors ; Treatment Outcome

OBJECTIVETo evaluate the clinical significance of MAIPA test in diagnosis of idiopathic thrombocytopenic purpura (ITP) and in the differential diagnosis of antoimmune thrombocytopenias from nonimmune thrombocytopenias.

METHODSA total of 321 thrombocytopenic patients (118 males, 203 females) from 14 centers were studied. A modified monoclonal antibody immobilization of platelet antigen (MAIPA) method was used to detect the platelet glycoprotein-specific autoantibodies (anti-GP IIb/IIIa, anti-GPIb/IX) to double-blindly evaluate its sensitivity and specificity for the diagnosis of ITP and to investigate the impact of the antibodies on platelet count.

RESULTSThe results showed that for the diagnosis of ITP, anti-GPIIb/IIIa, anti-GPIb/IX and both of them had the sensitivity of 39.75%, 32.64% and 55.23%; the specificity of 97.56%, 93.94% and 92.68%; the positive predictive value of 97.94%, 93.98% and 95.65%; the negative predictive value of 35.71%, 32.35% and 41.53%; and the total efficiency of 54.51%, 48.29% and 64.80%, respectively. The positivity of the autoantibodies in immune thrombocytopenias was incredibly higher than that in nonimmune thrombocytopenias. The platelet counts in the immune thrombocytopenias with autoantibody positivities were significantly lower than those without the autoantibodies. The platelet counts were negatively correlated with the concentration of the autoantibodies. The levels of anti-GPIIb/IIIa or anti-GPIb/IX or both of them dropped or disappeared in patients being responsive to steroid therapy.

CONCLUSIONMAIPA assay is proved to be of great value for the diagnosis of ITP and for differential diagnosis of immune thrombocytopenias from nonimmune thrombocytopenias.

Antibodies, Monoclonal ; immunology ; Antigens, Human Platelet ; immunology ; Autoantibodies ; immunology ; Blood Platelets ; Humans ; Prospective Studies ; Purpura, Thrombocytopenic, Idiopathic ; immunology

OBJECTIVETo establish a cell model of secreted alkaline phosphatase (SEAP) co-controlled by HCV 5'NCR and NS3 serine protease in an effort to develop new antiviral agents.

METHODSThe fragments of HCV 5'NCR and NS3/4A-SEAP were amplified by PCR. They were fused into pBluescript SK+ to generate 5'NCR-NS3/4A-SEAP chimeric plasmid. The resulting chimeric gene was subcloned into HindIII/Bsu36 I site of pSEAP2-Control (a SEAP eukaryotic expression plasmid), to generate pNCR-NS3/4A-SEAP, in which the SEAP was fused in-frame to the downstream of NS4A/4B cleavage site. The SEAP activity in the culture media of transiently transfected cells was monitored quantitatively. The regulatory effect of HCV 5'NCR and NS3 serine protease on SEAP expression was measured by treatment of transfected cells with antisense oligodeoxynucleotide (ASODN) against HCV 5'NCR and TPCK, a irreversible serine protease inhibitor.

RESULTSThe SEAP activity in the culture media reached 80801+/-4794 RLU, and was significantly inhibited by 5 micromol/L, 10 micromol/L of ASODN (t=4.315, p<0.01; t=6.985, p<0.001) and 100 micromol/L of TPCK (t=6.949, P<0.001).

CONCLUSIONA cell model of SEAP co-controlled by HCV 5'NCR and NS3 serine protease has been successfully established. This might promote the screening of anti-viral drugs

Alkaline Phosphatase ; secretion ; Antiviral Agents ; Drug Evaluation, Preclinical ; Hepacivirus ; genetics ; Hepatocytes ; enzymology ; virology ; Humans ; Recombinant Proteins ; biosynthesis ; genetics ; Serine Endopeptidases ; biosynthesis ; genetics ; Viral Nonstructural Proteins ; biosynthesis ; genetics

OBJECTIVETo investigate the Malytea Scurfpea fruit (MSF) on melanocyte adhesion and migration.

METHODHuman epidermal melanocytes were treated with MSF and Ginger respectively, then adhesion to bovine serum fibronectin-coated culture dishes was checked. Control and treated cells were also examined for migration into micropore filters coated with the same protein.

RESULTCompared with control, MSF treated melanocytes were obviously easier to adhere to the dishes and move into the filters in a dose-dependent manner. When the dose of MSF was 200 mg x L(-1), it could not reincrease melanocyte adhesion and migration. At 10 mg x L(-1), under every other concentrations of MSF, there was no marked difference among MSF-treated, Ginger-treated and untreated melanocytes (P < 0.05) when adhesion test were studied. But to migration, even at 10 mg x L(-1) MSF, there was obvious increased migration compared with MSF-untreated or Ginger-treated melanocytes (P < 0.01).

CONCLUSIONMSF has effect on melanocyte adhesion and migration, which can explain, in part, the capacity of MSF to modulate melanocyte function in vitiligo lesions.

Cell Adhesion ; drug effects ; Cell Movement ; drug effects ; Cells, Cultured ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Fruit ; chemistry ; Ginger ; chemistry ; Humans ; Melanocytes ; cytology ; drug effects ; Plants, Medicinal ; chemistry ; Psoralea ; chemistry

OBJECTIVETo summarize the experience of diagnosis and treatment of esophageal leiomyoma.

METHODSClinical data of 52 patients with esophageal leiomyoma were analyzed from 1993 to 2002.

RESULTSAbout 54% patients in this group had difficulty of food intake. The diagnostic accuracy of gastrointestinal barium meal series, computed tomography, gastric endoscope and endoscopic ultrasonography (EUS) for esophageal leiomyoma was 64% 44% 27% and 90% respectively. All patients received operation, resection of esophageal leiomyoma by videothoracoscopy (VAS) and endoscope were performed in 6, 9 patients respectively. The remaining 37 patients received regular open operation,in whom 32 cases received enucleation of esophageal leiomyoma, 5 cases received partial esophageal resection and esophageal-gastric anastomosis. No serious complications occurred except only one case needed operation again because of bleeding.

CONCLUSIONEUS is an effective method for diagnosing esophageal leiomyoma. VAS and endoscopic treatment should be considered for suitable cases in order to reduce the trauma.

Esophageal Neoplasms ; diagnostic imaging ; surgery ; Female ; Follow-Up Studies ; Humans ; Leiomyoma ; diagnostic imaging ; surgery ; Male ; Middle Aged ; Thoracoscopy ; Ultrasonography