Methotrexate (MTX)was linked covalently to monoclonal IgG antibody (AU_(14-1)) to a mouse uterine cervical cancer with the use of dextran T-40 as the intermediate carrier. The molar ratio of IgG: MTX was 1: 20 in the conjugate. The conjugate retained full antibody binding activity at the AU_(14-1) concentration of 3.75?10~(-8) M measured by an indirect membrane immunofluorescence assay. The in vitro experiments confirmed that the conjugate not only showed target-selective antitumor effecacy but also was proved to be more effective in inhibiting the growth of HeLa cells than the effect of U_(14) cells, so that it confirmed the "evolutionary antigen" theory. It is suggested that AU_(14-1)-Dex-MTX conjugate may have good potentiality in clinical application.

BACKGROUND: Researches find that grape seed proanthocyanidins (GSP) can eliminate free radicals, protect heart against ischemia-reperfusion injury and enhance learning and memory abilities in experimental animal, but their effects on the cerebral ischemia-reperfusion injury remain unclear.OBJECTIVE: To study the protective effects of proanthocyanidins derived from grape seeds on the cerebral ischemic reperfused brain by measuring the total antioxidative capacity (T-AOC), nitric oxide synthase activities and malondialdehyde (MDA) content in brain tissue of mice.DESIGN: A completely randomized and controlled study.SETTING: Department of Pathophysiology and Functional Central Laboratory, Jinzhou Medical College; Department of Neurology, the First Affiliated Hospital of Jinzhou Medical College.MATERIALS: The experiment was conducted in the Department of Functional Central Laboratory, Jinzhou Medical College from March to August 2004. Forty Kunming mice, provided by the Experimental Animal Center, Jinzhou Medical College, were randomly divided into five groups: sham control group, cerebral ischemia-reperfusion group (IR group) and cerebral ischemia-reperfusion treated with low or high dose of GSP or nimdipine (IR+GSP or IR+Nim) group with eight mice in each group.METHODS: ① Animal model establishment: The animals were anesthetized with ether. Then they were incised through median incision of the neck. The bilateral common carotid arteries were then occluded by microaneurysm clips for 30 minutes. After removing the clips, return of flow was visualized in the arteries. ② Model group and control group:The mice in low or high dose of GSP treated group or nimdipine treated group were injected GSP or nimdipine 10, 40, 2 mg/kg body mass respectively during the common carotid arteries occlusion and again at 24hours after reperfusion, while the mice in sham control group were injected the same volume distilled water with 40 mg/kg body mass. After 72-hour reperfusion, nitric oxide synthase activities, the total antioxidative capacity and MDA content in brain tissue of mice in each group were detected with chemical chromatometry. ③ The results were assessed by t test.MAIN OUTCOME MEASURES: Nitric oxide synthase activities, the total antioxidative capacity and MDA content in brain tissue of mice in each group were detected.RESULTS: Data of forty Kunming mice was entered the results analysis without any loss. ① Total antioxidative capacity: Total antioxidative capacity in cerebral ischemia-reperfusion group was obvious lower than that in the sham control group (t=8.145, P=0.000) while total antioxidative capacity in low or high dose of GSP treated group and nimdipine treated group was obvious higher than that in the cerebral ischemia-reperfusion group (t=6.313, 8.956, 4.14, P ＜ 0.01). ② Nitric oxide synthase activities: Nitric oxide synthase activities in cerebral ischemia-reperfusion group was obvious higherthan that in the sham control group (t=12.541, P ＜ 0.01), while nitric oxide synthase activities in low or high dose of GSP treated group and nimdipine treated group was obvious lower than that in the cerebral ischemia-reperfusion group (t=2.231, 8.956, 7.260, P ＜ 0.05-0.01). ③ MDA content: MDA content in cerebral ischemia-reperfusion group was obvious higher than that in the sham control group (t=7.883, P ＜ 0.01), while high dose of GSP treated group and nimdipine treated group was obvious lower than that in the cerebral ischemia-reperfusion group (t =5.234,4.518, P ＜ 0.01).CONCLUSION: GSP exerted a protective effect on the cerebral ischemic reperfused brain by enhancing total antioxidative capacity and reducing lipid peroxidantion and nitric oxide synthase activities.

AIM: To investigate the effect of monoclonal antibody(McAb) AU_(14-1) mediated cytotoxicity against cervical cancer U_(14) cell in vitro.METHODS: MTT colorimetric assay was applied to study the McAb AU_(14-1) mediated cytotoxicity of effector cells including splenocytes,peritoneal macrophages and LAK to U_(14) cells.RESULTS: The cytotoxicity mediated by each effector cells to the U_(14) cells treated with McAb AU_(14-1) was significantly higher than those not treated with it(P

AIM: To study the effect of chronic hypoxia(CH) on the intracellular calcium([Ca~(2+)]i) in pulmonary artery smooth muscle cells(PASMCs) and the role of L-type calcium channel and calcium store. METHODS: The rat chronic hypoxia model was set up and intervene the PASMCs with normal PSS,calcium-free PSS,nifedipine,and heparine respectively.The resting [Ca~(2+)]i was determined with the Fura-2/AM calcium imaging technique.RESULTS:(1) The [Ca~(2+)]i in CH group in normal PSS was higher than that in control group in normal PSS(P

OBJECTIVE:To optimize the extraction technology of polysaccharides from Gossypium herbaceum L. by pectinase hydrolysis. METHODS:Polysaccharides were extracted from G. herbaceum L. by pectinase hydrolysis combined with traditional hot water extraction. With the extraction rate of polysaccharides as the evaluated index,single factor test and orthogonal design were applied to investigate the effects of 4 factors including enzymolysis temperature,enzymolysis time,the amount of enzyme and pH value on polysaccharides extraction rate,and verification tests were conducted. RESULTS:The optimal extraction technolo-gy was as follows as enzymolysis temperature of 40 ℃,enzymolysis time of 150 min,enzyme accounting for 2.0%,pH value for enzymolysis of 4.6. Under the above conditions,the average extraction rate of polysaccharides from G. herbaceum L. was 2.474%(RSD=3.34%,n=5). CONCLUSIONS:Pectinase hydrolysis is an effective method to extract polysaccharides from G. herbace-um L.. The optimal extraction technology is reasonable and feasible.

Objective:To investigate the onset mechanism of cervical vertigo from the proprioceptive sensation and provide clinical basis for its treatment.Methods:Among the 121 cases that conformed to the diagnostic criteria of cervical vertigo,70 cases who presented with negative neck-rotation test and cervical vertigo without obvious vascular factors by transcranial Doppler(TCD)were assigned to the observation group,while 51 cases who presented with positive neck-rotation test and cervical vertigo due to spasm of vertebral basal artery or insufficient blood supply by TCD were assigned to the control group.The cases in the two groups were treated once every day,5 days make up one treatment course and the resuIts were statistically analyrzed after one treatment course.Results:The skull triaxial spatial offset of the cases in the two groups were significantly reduced after the treatment (P<＜0.01).However, there was no statistically significant difference between the skull triaxial spatial offset between the two groups (p＞0.05).It is not conclusive that the therapeutic effect in the two groups was significantly different after one treatment course.Conclusion:Tuina manipulation therapy can improve the skull spatial ofrset repositioning ability of the Patients.

BACKGROUND: Recombinant adeno-associated virus 2(rAAV-2) has attracted considerable attention due to its nonpathogenic nature in contrast to other viral vectors such as adenoviral and retroviral vectors in gene therapy attempts.OBJECTIVE: To explore rAAV-2 transduction to bone marrow mesenchymalstem cell(BMSC) in vitro and evaluate the possibility of using rAAV-2 as a vector for gene therapy of acute myelogenous leukemia(AML).DESIGN: An open experiment with cells as the observational subjects.SETTING: Department of Hematology, Nanfang Hospital, Southern Medical University.MATERIALS: The experiment was conducted in the Department of Hematology, Nanfang Hospital, Southern Medical University from February to July 2004. We used passages 3 to 5 BMSCs derived from six de novo AML patients and four healthy volunteers in this study.METHODS: BMSC was isolated from 6 to 10 mL of bone marrow aspirates obtained from the iliac crests of the patients who had been diagnosed as having de novo AML and from those of healthy volunteers. The acquired BMSC was infected by rAAV-2 which contained enhanced green fluorescent protein (rAAV-2-eGFP) at different multiplicity of infection(MOI) (MOI = 1 × 102,1 × 103, 1 × 104, 1 × 105, 1 × 106, 1 × 107) . Then we observed through phase contrast fluorescent microscope and flow cytometer to evaluate green fluorescent protein(GFP) expression 10 to 14 days after transduction. GFP expression was observed as the rAAV-2-eGFP transduced BMSC cultured in vitro. We also observed the in vitro gene expression profile of GFP in rAAV-2-eGFP transduced BMSC which was selected by neomycin ( G418). First, we confirmed GFP expression in BMSC through phase contrast fluorescent microscope, then on flow cytometer to detect the percentage of GFP expression.MAIN OUTCOME MEASURES: The efficiency of rAAV-2-eGFP transduction to BMSC. GFP expression was observed through phase contrast fluorescent microscope and flow cytometer at different time points after transduction.rAAV-2-eGFP to BMSC derived from normal volunteers and AML patients had no significant differences. GFP began to express 10 to 14 days after transduction, and the transduction efficiency ranged from 0. 3% to 1.4%. By changing infection condition, we could not make a higher transduction efficiency( P ＞ 0.05) . One round infection of BMSC by rAAV-2-eGFP at a MOI of 1 × 105 was ( 1. 030 ± 0. 034) %, 3 rounds of infection of BMSC by rAAV-2-eGFP at a MOI of 1 × 105 was (1. 140 ±0. 036)%, and coinfected by LipofectAMINE was (1. 380 ± 0. 054)%. However, 293 cell line which was the package cell of rAAV-2 could be efficiently transduced by AML patients transduced by rAAV-2-eGFP at MOI = 1 × 105: The percentage of GFP expression cell gradually decreased from 1.14% at day 12 after transduction to 0. 6% as cell passaged from 2 to 3, and maintained at a level of 0. 5% to 0. 6% later on till 61 days after transduction. After selected by neomycin(G418) 1 month later, rAAV-2-eGFP transduced BMSCs could maintain a long-term GFP expression at a level of 6.0% in vitro without significant decay within 100 days of observation period after transduction.CONCLUSION: The advantages of rAAV-2 mediated gene transduction lie in safety, no immune response to the host, and long-term expression maintained by the target gene. rAAV-2 and BMSC can be used for in vitro gene therapy, and as a systemic gene delivery system, it might be an alternative for systemic gene therapy in the future.

Objective To explore the mechanism of drug resistance of ovarian cancer cells to TRAIL-induced apoptosis.Methods We collected 3AO cells and CAOV3 cells,respectively,at 18,24,48 and 72 hour under 12.5,25,50 and 100 ng/mL concentrations of TRAIL.The rate of cell growth inhibition was checked by methyl thiazolyl tetrazolium (MTT)assay to evaluate the effect of TRAIL.Morphology of apoptotic cells was observed by TdT-mediated-dUTP nick end labeling (TUNEL).The apoptosis rate was detected by flow cytometry (FCM)and C-FLIP protein was determined by Western blotting.Results TRAIL inhibited the growth of 3AO and CAOV3 cells.The rate of growth inhibition at 24 hour was 28% in 3AO cells and 10% in CAOV3 cells.TRAIL induced apoptosis of cells.The apoptosis rate at 24 hour was 8.5% in 3AO cells,which was higher than 5.5% in CAOV3 cells.The expression level of C-FLIP protein was higher in CAOV3 cells than in 3AO cells.Conclusion C-FLIP protein is an important protein that regulates drug resistance of ovarian cancer cells to TRAIL-induced apoptosis.

AIM and METHODS: A study of radioimmunoimaging was carried out on Kcnming mice - uterine cervical cancer (U14) using 99mTc labeled monoclonal Au14-1 with a modified Schwartaz method. RESULTS: The bio - distribution showed that radioactivity accumulated in tumor tissue at 12h after 99mTc - Au14- 1 injection in tail vein. The uptake by tumor was 4. 12 % ID/g at 2h and 8. 79 % ID/g at 24h respectively. The tumor/non - tumor (T/NT) radiocativity ratios for organs except kidneys were ranged from 2.02 to 6.71 at 24h post - injection. The image of tumor showed at 12h and clearer at 24h after injection. CONCLUSION: The quality of tumor image was relevant to the T/NT radioactivity ratios. It was demonstrated that 99mTc- Au14-1 has a good capability of localization for tumor.