Objective To explore and protect the gene resources of Sika deer in China in oroder to develop recombinant-peptide-drugs derived from Sika deer genes. Methods The total RNA was isolated from Sika deer spleen cells by using the Trizol reagent and the total mRNA was purified, complement DNA(cDNA) and the second DNA were obtained by means of reverse transcriptase-PCR(RT-PCR) and this cDNA was used as the template respectively. The two strains of DNA was subcloned into the pSPORT1. The recombinant plasmids of pSPORT1 were transformed into the E.coli by means of electric transcription. The cDNA library was obtained. The positive recombinant-clone was selected and the sequences of ESTS were analyzed. Results Sika deer spleen cell cDNA library was established successfully; the clones in it was sequenced, during the process some expresion sequence tags were discovered that shared high homology with known cDNAs deposited in the GenBank. EST-1 was homologous to human retinoblastoma binding protein 4. The protein sequence of EST-2 was 26% homologous to human vitamin K-dependent protein precursor. Conclusion Some valuable gene fragments from Sika deer spleen cell cDNA library were obstained.

Objective To obtain the E2 genes from Neimeng strain of classical swine fever virus and construct fusion genes. Methods E2 DNA fragment was acquired from spleen of infected pig and amplified through RT-PCR technique. After being sequenced, the E2 gene was ligated to Chaperone 10 to form the fusion gene Chap 10-E2, the fusion gene was ligated to expressing vector. Results There were differences in four bases between E2 gene from Neimeng strain of swine fever virus and E2 gene from HCLV strain. The fusion gene Chap 10-E2 was successfully constructed. Conclusion The difference of E2 gene of swine fever virus from different strains is not significant.

Objective To clone the sequence of the cDNA of mouse B7-2 gene and construct its expression vectors. Methods Using mouse splenocyte mRNA as template to obtain full length B7-2 with the technique of RT-PCR followed by automatic sequencing of pMD18T-B7-2 and to construct recombinant plasmids containing CMV, Egr-1 and B7-2 gene with recombinant DNA technique. Results Sequencing proved the cloned B7-2 cDNA to be essentially identical with that reported in the literature and the recombinant plasmids containing CMV, Egr-1 and B7-2 gene were constructed successfully. Conclusion B7-2 cDNA was successfully cloned and two expression vectors pcDNA3.1-CMV-B7-2 and pcDNA3.1-Egr-B7-2 were constructed.

Objective To explore the mechanism of proconditioning to hypoxia. Methods The changes of NOS neuron of mouse brain in proconditioning to hypoxia were detected by histochemistry method. Results Compared with control mice, the number, colour, shape, dendrites and axons number and length of NOS positive neurons in cerebral cortex was not changed after mice were exposed to hypoxia for one time, but the dendrites and axons of NOS neuron became thicker. Compared with control mice, NOS neurons in cerebral cortex was not changed after mice were exposed to hypoxia for four times. There were little NOS neurons in hippocampus of control mice and NOS neurons were light positive. Compared with control mice, the number of NOS positive neurons in hippocampus increased and NOS neuron became strong positive after mice were exposed to hypoxia for one time. Compared with mice exposed to hypoxia for one time, the number of NOS positive neurons in hippocampus decreased and their colour became light after mice were exposed to hypoxia for four times. Conclusion The changes of NOS neurons can enhance generation of proconditioning to hypoxia.

Objective To study the effects of methylmercury chloride (MMC) on NMDA receptor (NMDAR) 2A subunit (NR2A) in hippocampus of newborn rats chronically exposed to MMC during developmental stages. Methods Female Wistar rats (120±20 g) were randomly divided into control group and experimental group. In experimental group,the rats were fed on forage containing different dosage of MMC (0.75，1.50 and 3.00 mg•kg^-1) each day for 90 days. NR2A proteins were exacted from hippocampus of newborn rats on P (postnatal days)7,P14,P21, and P28. NR2A protein expression was investigated with Western blotting. Results NR2A protein was detected in both control and experimental groups. Expression of the NR2A reached the peak at P14. A decreasing tendency appeared in amount of NR2A from hippocampus of MMC-exposed rats on P7, 14, 21, and 28 compared with control group. Conclusion NMDAR in rat brain involves in brain developmental process. Decrease of NR2A expression in rat exposed to MMC may be the key molecular mechanism of MMC damage of the developing brains.

Objective To study the effect of homocysteine(Hcy) on collagen synthesis and the expression of type Ⅰand Ⅲ collagen mRNA of vascular smooth muscle cells(VSMCs) in culture. Methods The VSMCs of rats were cultured in Hcy with different concentrations in vitro. The effects of Hcy on collagen synthesis and on the expressions of alpha 1 (Ⅰ)　and (Ⅲ) collagen mRNA of VSMCs were observed by determining hydroxyproline contents in the culture medium and semi-quantitative RT-PCR. Results Hcy accelerated collagen synthesis and up-regulated the expressions of alpha 1 (Ⅰ)and (Ⅲ) collagen mRNA of VSMCs in a dose-dependent manner. Conclusion Hcy can promote collagen synthesis through up-regulating alpha 1(Ⅰ)and (Ⅲ) collagen mRNA expressions in VSMCs in culture.

Objective To investigate the effects of excessive fluoride on osteoclastic activity in rats. Methods The tibia slices of chronic fluorotic rats were stained with HE and tartrate-resistant acid phosphatase (TRAP)，and the changes of osteoclasts were observed；At the same time，the protein was extracted from metaphysis of femurs in chronic fluorotic rats，then the expression of matrix metalloproteinase-9 (MMP-9) was investigated using Western blotting technique. Results The number of osteoclasts in tibia envelope and metaphysis increased in fluoride groups and compact bone of the cortex became porosity. Compared with stock diet group，the number of TRAP positive osteoclasts increased in stock diet group treated with fluoride， low calcium diet group，and low calcium diet group treated with fluoride. But there was no much statistical difference (P>0.05). Compared with stock diet group, the increase of MMP-9 expression in stock diet group treated with fluoride was statistically significant(P<0.05). Statistical difference also showed between low calcium diet group and low calcium diet group treated with fluoride (P<0.05)，and there were significant diffiences when both above were compared with stock diet group (P<0.05). Conclusion The results prove that excessive fluoride can induce accelerative bone turnover and active bone resorption, and low calcium diet is an important promotive factor of skeletal fluorosis. Fluoride can enhance the activity of osteoclastic resorption by increasing the expression of MMP-9.

Objective To explore the effects of diabetes on anti-oxidation levels of key organs and tissues in rats. Methods Twenty-four Wistar rats for 90 days were randomly divided into three groups : control group (C), diabetes treated with insulin group (ID) and diabetes group(D). There were 8 rats in each group. The changes of levels of GSH-Px, GSH, and GR in rat organs and tissues were detected by colorimetric method. Results Compared with group C, GSH-Px levels in heart, kidney, and pancreas in group D were significantly ascended (P<0.01 or P<0.05); it was ascended in liver, but there wasn′t statistical significance (P>0.05), and the GSH-Px level was not changed in spleen. GSH and GR levels in group D were significantly descended in heart, liver, kidney, spleen, and pancreas compared with group C (P<0.01 or P<0.05); but they were alleviated after insulin was administered. Conclusion The anti-oxidation levels in important organs and tissues of diabetic rats may be improved after insulin therapy.

Objective To study the effects of mature dendritic cells (DCs) derived from mouse bone marrow on T cell proliferation in vitro, and provide techinological methods for tumor therapy. Methods With IL-4 and GM-CSF, the cells from mouse bone marrow were cultured for 5-7 days. The morphological changes were observed under light microscope on 3rd and 5th day after culture; immature DCs on 5th or 6th day after culture were loaded with tumor-antigen for 1-2 days; CD83 and CD86 were tested by flowcytometric analysis; T-cell-stimulatory activity in DCs derived from mouse bone marrow and loaded with tumor-antigen was analysed by γ- scintillomete. Results Morphological changes were found and dendritic protrusion appeared on 3rd day after culture; dendritic protrusion prolonged on 5th day after culture; flow-cytometric analysis showed that expressions of CD83 and CD86 of IL-4+GM-CSF+TNFα group (61.68% and 71.25%) and IL-4+GM-CSF+tumor-antigen group (63.1% and 76.88%) had significant differences compared with control group (2.41% and 3.88%) and IL-4+GM-CSF group (21.86% and 28.69%) (P<0.001), and IL-4+GM-CSF+tumor-antigen group had also significant difference compared with IL-4+GM-CSF+TNFα group(P<0.01); proliferation of T cells in IL-4+GM-CSF+tumor-antigen group strongly enhanced compared with other groups (P<0.001,P<0.05). Conclusion Sufficient DCs generated from bone marrow cells are induced in vitro and strongly stimulate proliferation of T cells after tumor antigen are loaded.

Objective To study the common chemotherapy effect and its mechanisms of the adaptive response induced by low dose BLMA₅ (0.08 mg•kg^-1, D₁) on a Kunming S180 sarcoma mouse model made through subcutaneous injection. Methods D₁ was given to tumor-bearing mice before chemotherapy, the growth of tumor was observed, chromosomal samples of bone marrow cells of mice were prepared and the activities of free radicals of liver were measured. Results The grow velocity was significantly decreased (P<0.05~0.001), the chromosome aberrations of bone marrow cells were significantly reduced (P<0.01), SOD activity of liver was increased markedly (P<0.05), the activity of GSH -Px also had the trend of increasing and the content of MDA decreased significantly (P<0.01) when D₁ was given to the tumor-bearing mouse before larger dose BLMA₅(D₂) chemotherapy. Conclusion The chemotherapy effect enhanced with low dose BLMA₅ is associated with the increase of antioxidant capacity and the elimination capacity of free radicals products.

Objective To investigate the effects of interferon-panaxadiol saponins (IFN-PDS) on the function of T-lymphocyte proliferation in influenza virus-infected mice. Methods 60 Kunming mice were divided into 5 groups: control group(ConG),influenza virus-infected group (VIG),PDS treatment group(PDSG),IFN treatment group (IFNG),IFN-PDS treatment group(IFN-PDSG). Mouse adopted influenza virus A was used to construct virus infected model. The functions of T lymphocyte proliferation in spleen and thymus gland were observed by MTT assay. Results ①The spleen index of VIG was much greater than that of ConG（P<0.01）. The mitogenesis induced by ConA of thymocyte and splenocyte was higher than that in ConG （P<0.05）. ②Self-proliferations of splenocytes in PDSG，IFNG, and IFN-PDSG were significantly higher than that in VIG（P<0.01,P<0.05, P<0.05）. The mitogenesis induced by ConA of splenocytes in PDS and IFN-PDSG were lower than that in VIG （P<0.05, P<0.01）.③The mitogenesis induced by ConA of thymocytes in PDSG,IFNG， and IFN-PDSG were lower than that in VIG （P<0.01,P<0.05, P<0.05）,had no difference compared with ConG（P>0.05）. Conclusion Influenza virus can stimulate the proliferation of spleen and thymic T lymphocyte directly. It can increase the T lymphocyte mitogenesis induced by ConA obviously. PDS and IFN-PDS treatment can increase the self-proliferation of splenocyte in influenza virus-infected mice, and decrease the excessive mitogenesis of splenocyte and thymocyte induced by ConA. It has two-way modulation on the function of T lymphocyte.

Objective To investigate the distribution and ecology of Sporothrix schenckii and dematiacious fungi in Jilin District of Jilin Province. Methods Seventy samples of rotten wood, soil, corn stalk and rice stalk from natural environment were collected. Suspected fungi were isolated by SDA mixed with actidione. Then, they were cultured and subcultured by SDA. By this means, Sporothrix schenckii and pathogenic dematiacious fungi were isolated and identified. Results In seventy samples, thirty-seven strains of Sporothrix schenckii and dematiacious fungi had been identified, the positive rate was 52.86%. There were 6 strains of Sporothrix schenckii (8.57%), Exophiala 14 (20%), Alternaria 3 (4.29%), Fonsecaea 5 (7.14%), Phialophora verrucosa 4 (5.71%), unidentified fungi 5 (7.14%) among them.　Opportunistic pathogen, namely Fusarium,　Aspergillus, Penicilliun had also been isolated. Conclusion The study is of great significance to learn about the distribution and ecology of Sporothrix schenckii and dematiacious fungi. It is also important to the etiology, pathogenesis and prevention of sporothichosis, chromomycosis, and phaeohyphomycosis.

Objective To study the biocompatibility and biodegradation characteristics of poly(caprolactone-b-ethylene oxide) (PCL-EO), a kind of biodegradable material, used for urological stent in vivo and in vitro. Methods Pellets made from PCL-EO were implanted in the dorsal muscle of Wistar rats or dipped into flowing urine. Tissue blocks surrounding the biomaterials and the PCL-EO pellets were took out after 2, 4, 6, 8, 10, 12, 14 and 16 weeks， respectively. Histological study was taken to investigate the tissue reaction of the material and evaluate its biocompatibility. Scanning electron microscopy (SEM) and measurement of molecular weight and gravity were used to make out the biodegradation characteristics of PCL-EO both in vivo and in vitro. Results The PCL-EO test pieces degraded into small particles that could be excreted with the urine flow after 16 weeks. The biodegradation pattern in vivo was similar to that in vitro. There was no abscess formation and tissue necrosis surrounding the implanted materials. Non-infectious reactions due to materials implanted were observed. Conclusion The biodegradable material PCL-EO has good biocompatibility and appropriate biodegradation characteristics and will meet the request for urological stent.

Objective To study the effects of morphine on proliferation of cultured C6 glioma cells in rats and its mechanism. Methods ³H-TdR incorporation and MTT colorimetry were used to study the effect of morphine on cell proliferation. Immunohistochemistry method was applied to determine the effect of morphine on P53 and Bcl-2 protein level. Results After different doses of morphine (0.01, 0.1, 1.0, 10 and 100 mg•L^-1) were acted on cultured C6 cells, the level of ³H-TdR incorporation decreased in a dose-dependent manner, compared with control and compared with each other among every two adjacent-dose groups. There were significant differences(P<0.01), and the inhibitory rate of morphine on C6 cell examined by MTT colorimetry was over 30%. The levels of P53 protein and Bcl-2 protein in C6 glioma cells decreased after exposed to morphine with the dose of 10 mg•L^-1 for 24 hours(P<0.01). Conclusion Morphine has inhibitory effect on proliferation of C6 glioma cells in rats and can inhibit P53 and Bcl-2 protein level.

Objective To construt a recombinant eukaryotic expressing plasmid containing dense granules antigen (GRA1) gene of Toxoplasma gondii. Methods A pair of primers were designed for PCR according to the known sequence of GRA1 gene.The full fragment of GRA1 gene obtained by PCR amplification from genomic DNA of RH strain of Toxoplasma gondii was cloned into plasmid pcDNA3 orientatedly.The constructed recombinant plasmid pcDNA3-GRA1 was transferred into E.coli DH5α.The transformatants were screened and identified by restriction endonuclease digestion and PCR. The nucleotide sequences of the cloned genes were determined. Results A 775 bp GRA1 gene PCR product was obtained from genomic DNA of the RH strain of Toxoplasma gondii,in which a 135 bp inserted sequence was found.Therefore the PCR product was longer than expected (628 bp).The frameshift mutation for GRA1 gene resulted from the insertion of exogenous sequence. Conclusion The inserted variation of GRA1 gene of the RH strain of Toxoplasma gondii is reported and its recombinant expressing plasmid is construted.

Objective To study the effect of LaCl₃ on the growth of CBRH-7919 cells. Methods The growth, clone inhibitory rate and general histological changes of CBRH-7919 cells were observed at different time (1th,3rd and 5th day) after treated with different doses (0.01,0.10 and 1.00 mmol•L^-1) of LaCl₃ in vitro. At the same time, the changes of cell cycle were detected by flow cytometry. Results The evident inhibition of LaCl₃ to CBRH-7919 cells growth occurred at 3rd day after treatment (P<0.01), gradually strengthened following the increase of dose and time (P<0.01). The result of clone formation assay expressed that the clone formations of CBRH-7919 cells were inhibited distinctly at the concentration of 0.10 and 1.00 mmol•L^-1 LaCl₃ (P<0.01). The result indicated that treated by LaCl₃, the distribution of cell cycle changed as followed: the cell percentage of G₀/G₁ period was increased, the ratios of groups of 0.01, 0.1 and 1.00 mmol•L^-1 LaCl₃ were evidently increased compared with the control and presented density-dependent relation (P<0.01); The cell percentage of S period was reduced, the rations of groups of 0.10 and 1.00 mmol•L^-1 LaCl₃ were significantly reduced compared with the matched control (P<0.05). Conclusion LaCl₃ can inhibit the growth of CBRH-7919 cells obviously.

Objective To investigate the effects of HGF/SF and VEGF on vascular endothelial cells proliferation and capillary sprouts formation. Methods Endothelial cells proliferation test and capillary sprouts formation test were used to evaluate the effects of HGF/SF and VEGF on human umbilical artery endothelial cells (HUAECs) proliferation and capillary sprouts formation. ELISA was used to measure the contents of HGF/SF and VEGF in the media. The rat cornea neovascularization was used to evaluate the effects of HGF/SF and VEGF on angiogenesis in vivo. Results HGF/SF and VEGF had obvious effects on angiogenesis and endothelial cells proliferation and capillary sprouts formation. The VEGF was found in HGF/SF media. Conclusion The combination use of HGF/SF and VEGF has cooperative effects on angiogenesis in tumor, HGF/SF may have the effect to stimulate vascular endothelial cells to produce VEGF.

Objective To study the effect of CrCl₃ on blood glucose in experimental diabetic mice. Methods The model of experimental diabetic mice was established by abdominal cavity injection of alloxan with the dose of 200 mg•kg^-1. The animals were orally administered with CrCl₃, in the doses of 40,20 and 10 μg•kg^-1•d^-1, respectively, for 4 weeks. The level of blood glucose was determined with the method of glucose oxidase. Results The body weights of mice in various treatment groups were significantly higher than that in diabetic group (P<0.01). The levels of blood glucose as well as the food and water intakes were obviously decreased（P<0.05 or P<0.01）. Conclusion CrCl₃ can reduce the level of blood glucose in experimental diabetic mice.

Objective To explore the protective effects of Ginaton on cerebral ischemia reperfusion injury and its influence on the exprssions of Fas and Bcl-2 protein in rats. Methods The global cerebral ischemia reperfusion model was established to investigate the expressions of Fas and Bcl-2 protein and the changes after given Ginaton were observed with immunochemistry technique. Results The expression of Fas protein appeared in simple ischemic group and reperfusion-3-hours group, and peaked 6 hours after reperfusion ,and decreased 24 hours after reperfusion. The expression of Bcl-2 peaked 3 hours after reperfusion, and gradually decreased 6 hours after reperfusion, and its expression reduced obviously 24 hours after reperfusion.In Ginaton group, the expression of Fas protein obviously decreased and Bcl-2 protein increased at corresponding time. Conclusion Ginaton can reduce the apoptosis following cerebral ischemia reperfusion and has protective effect on the injury of ischemia and reperfusion.

Objective To observe the effects of Acanthopanax senticosus saponins（ASS） on hemorrheology and platelet function in experimental cerebral ischemia rats. Methods The cerebral ischemia model induced by bilateral common carotid artery clamping and controlled hemorrhage to a mean arterial pressure of 50 mmHg was produced in rats treated by continuous intraperitoneal injection of ASS with dosages of 25,50,100 mg•kg^-1 for 3 days. The effects of ASS on the hemorrheology and platelet aggregation and adhesion functions were measured after 15 min. Results ASS with dosages of 50 and 100 mg•kg^-1 decreased the blood viscosity,plasma viscosity, plasma fibrinogen concentration,erythrocyte sedimentation,hematocrit, erythrocyte aggregation index, and erythrocyte rigidity index markedly,and inhibited platelet aggregation and adhesion functions. Conclusion ASS can improve evidently abnormal changes of hemorrheology and platelet functions in experimental cerebral ischemia rats.

Objective To establish the isolation and primary cultivation system of porcine hepatocytes in accordance with the requirement of hybrid bioartificial liver support system (HBLSS) of cell in vitro. Methods Porcine hepatocytes were isolated with digestive method of collagenase perfusion with the improved Seglen method. Morphological changes of the hepatocytes were observed. Results The average yield of hepatocytes was 2×10^10 cells per pig and the average viability was 92.5%. Conclusion The digestive method of collagenase perfusion is the basic approach for obtaining a mass of hepatocytes with high viability. Porcine hepatocytes are the prime cells source for HBLSS.

Objective The characteristics of the proliferation and differentiation of fetus mice skin MSCs were observed in order to provide an ideal method for bone tissue engineering science and treatment for osteoporosis. Methods Fetus mice skin MSCs were purified with the method of cell culture in vitro and potential of differentiation were observed in culture fluid with the osteogenic inducer. Results The repeated adherent passage cultured cells that were originated from the fetus mice skin cells still had a strong proliferation capacity after 15 generation′s passage purification culture. It was clarified that the cultured cells were originated from stem cells. The adherent cultured cells increased the activity of ALP, ALP staining reaction and calcium nodus alizarin-red staining reaction were positive with the osteogenic inducer. Conclusion The skin MSCs existing in the fetus mice have the potential of self-proliferation and can be differentiated into osteoblasts with the osteogenic inducer.

Objective To observe the effects of Shexiang Anti-thrombosis Pill (SATP) on platelet function and blood viscosity in experimental cerebral ischemia rats. Methods The cerebral ischemia model induced by bilateral common carotid artery clamping and controlled hemorrhage to a mean arterial pressure of 50 mmHg was established in rats treated by continuous intragastric administration of SATP 1.5,3.0,6.0 g•kg^-1 for 14 days. The effects of SATP on the blood viscosity (ηB), plasma viscosity (ηP), platelet aggregation and adhesion functions and thrombus stretch picture were measured after 15 minutes. Results SATP 3.0 g•kg^-1 and 6.0 g•kg^-1 decreased the ηB and ηP markedly,and inhibited platelet aggregation and adhesion functions,but had not significant effect on thrombus stretch picture. Conclusion SATP can protect the cerebral tissue from ischemia injury by decreasing the ηB and ηP and platelet aggregation and adhesion functions. After the conversion of honey pill into water pill, the pharmacological effect of SATP is increased significantly.

Objective To explore the role of microsatellite instability （MIN） in the development of laryngeal cancer. Methods The MIN in 50 cases of laryngeal cancer were detected by the methods of PCR- denature PAGE polymorphous markers BAT-26 and BAT-40. Results Twenty-six percent of patients with laryngeal cancer had MIN. MIN were not related to the grade of differentiation and clinical stage laryngeal cancer. And MIN was associated with lymph node metastasis（P<0.05）. Conclusion MIN seemly acts as a positive regulator in the lymph node metastasis of laryngeal cancer.

Objective To investigate the expression of nerve growth factor (NGF) gene in astrocytes cultured in chemical 1,25-(OH)₂D₃ defined medium. Methods The astrocytes prepared from brain of neonatal rats were cultured in a chemical 1,25-(OH)₂D₃ defined serum-free medium with various doses of 1,25-(OH)₂D₃ （0，10_-7，10^-8-8，10^-9-9，10^-10，10^-11 and 10^-12 mol•L^-1 ）. NGF mRNA transcripts were detected by RNA-based polymerase chain reaction method. Secretory NGF protein quantity were measured with a double-site ELISA with monoclonal anti-NGF antibody. Results NGF synthesis and secretion were induced in astrocytes cultured in chemical 1,25-(OH)₂D₃ defined serum-free medium. 1,25-(OH)₂D₃ were effective in the concentration range of 10_-7 to 10^-11 mol•L^-1. The expression of mRNA was detected 6 hours after cultured in 1,25-(OH)₂D₃ medium,the level of expression of mRNA approached to peak value at 12th hour, and NGF mRNA was not found after 36 hours. The reaction period of ELISA of NGF protein was above 48 hours.Conclusion NGF gene in central nervous system (CNS) can be regulated by 1,25-(OH)₂D₃.

Objective To study the anticoagulability and antithrombotic activity of the thrombolysin to evaluate its biochmical pharmacodynamics. Methods The effects of thrombolysin on coagulation time in mice were observed by slide method and capillary method. The effects of thrombolysin on antithrombus formation were observed by thrombosis experiment in rats and rabbits. Its effects on prothrombin time (PT),activated partial thormboplastin time (APTT) and the fibrinogen (Fbg) contents were also measured by reagent kits in rats. Results At the dose of 0.25,0.5 and 1.0 mg•kg^-1,it could significantly prolong the coagulation time in mice;At 0.2,0.4, and 0.8 mg•kg^-1 , thrombolysin inhibited thrombosis of rats in vivo and in vitro. At 0.2 mg•kg^-1,it inhibited thrombosis of rabbits in vitro. At the dose of 0.2,0.4,and 0.8 mg•kg^-1, it could decrease the Fbg content in rats,and had no effects on PT and APTT. Conclusion The thrombolysin possesses anticoagulability and antithrombotic activity, and it may be a promising medicine for diagnosis of thrombotic diseases.

Objective To study the role of heat shock protein65-MUC1 (HSP65-MUC1) fusion protein in the priming of IFNγ producing cells from immunized mice by detecting their IFNγ secretion. Methods The lymphocytes from spleen and lymph nodes of C57BL/6 mice immunized three times with HSP65-MUC1 fusion protein were prepared and then stimulated by HSP65-MUC1 fusion protein, MUC1 peptides or nonspecific stimulator such as CpG ODN and Con A in vitro were used to evaluate their IFNγ secretion through enzyme linked immunospot technique. Results The lymphocytes from the mice immunized by HSP65-MUC1 fusion protein produced much more IFNγ compared with those from control mice in the presence of specific or nonspecific stimulators. Conclusion HSP65-MUC1 fusion protein plays an important role in the priming of mice lymphocytes for the secretion of IFNγ.

Objective To investigate the inhibitory effects of aminoguanidine(AG),a selective inhibitor of inducible nitric oxide synthase(iNOS), on mice stomach cancer and its′mechanism. Methods The mice stomach cancer cell lines MFC were implanted subcutaneously in Kunming mice.Fifty mice were randomly divided into 5 groups: Control group (saline solution), MMC group (Mitomycin, twice a week 0.7 mg•kg^-1 i.p.),low dosage AG group (AG_L, 50 mg•kg^-1•d^-1 i.p.), high dosage AG group (AG_H, 150 mg•kg^-1•d^-1 i.p.) and combined treatment of both MMC and high dosage AG group(MMC+AG_H).All drugs were given by intraperitoneal injection. Two weeks after implantation, the mice were sacrificed, and the tumor weight,inhibitory rates,intratumoral microvessel density (MVD),the positive rate of vascular endothelial growth factor (VEGF) and iNOS were evaluated,respectively. In addition,serum were collected and nitrite levels were tested using Greiss assay. Results Compared with the negative control group,the growth of the orthotopically implanted tumor was significantly inhibited due to the reduced weight and the inhibitory rates of tumor in AG_H group and MMC+AG_H group were 47.1% and 52.9%,respectively. The MVD was decreased significantly in MMC+AG_H group and AG_H group ［(21.2±12.4)% and (8.8±2.6)%］,compared with control group ［(68.3±10.6)%］ (P<0.01). The positive rates of VEGF and iNOS in negative control group were (10.3±1.6)% and (11.3±1.3)%. However,they were (4.8±1.6)%,(3.8±0.9)% and (2.1±1.4)%,(2.4±1.1)% in AG_H group and MMC+AG_H group,respectively. Conclusion AG can significantly restrain the development of mice stomach cancer through inhibiting the expression of iNOS,VEGF and decreasing MVD. Combination treatment can improve the inhibitory effect.

Objective To observe the effect of polyclonal antibodies (Pabs) against glucosyltransferase on the colonization of Streptococcus sobrinus (S.sobrinus). Methods The 30 rats, 25 days old, were divided into 2 groups: control group and experimental group. 50 μL phosphate-buffered saline (PBS)(control group) and 50 μL Pabs (experimental group) were smeared on the surface of the molars 32, 36, 40, 44, 48 and 52 days after birth. After smeared 3 times, all rats were inoculated with S.sobrinus subsequently, they were fed on a cariogenic diet. The rats were killed 40 days after inoculation and the molars were extracted in order to determine the numbers of S.sobrinus by MS plates. The proportions of S.sobrinus expressed as a percentage of the total cultivable microflora. Results Statistical analysis showed that the level of colonization of rats in experimental group ［(37.01±12.08)%］ was significantly lowered than that in control group ［(48.20±14.78)%］(P<0.01). Conclusion Local application of Pabs against glucosyltransferase may significantly reduce the colonization level of implanted S.sobrinus.

Objective To construct hepatitis A virus vaccine with attenuated Salmonella typhimurium as vector, and study its immune response. Methods The plasmid pHAV5 containing hepatitis A virus genome cDNA was digested by restrict endonucleic acid enzyme, the 5′-non coding region was deleted, the remains were inserted into the high level expressive plasmid pBV221 and then were transformed into E.coli DH5α. The positive clone was screened and identified by DNA-probe and restrict endonucleic acid enzyme. The postive recombinant plasmid was used to transform attenuated Salmonella typhimurium BRD509 by electroporation. The transformed Salmonella typhimurium was screened in Amp-control medium. The aim-gene was expressed in Salmonella typhimurium by temperature-induced method. Results The expressed proteins reacted specifically with HAVIgG antibody were demonstrated by ELISA and Western blotting. The transformed Salmonella typhimurium was administrated to mice by oral or abdominal injection, HAVIgG antibody was found in the mice sera. Conclusion Hepatitis A vaccine with attenuated Salmonella typhimurium as vector can induce immune response.

Objective To evaluate the osteocompatibility of titania ceramics in vitro. Methods Primary osteoblast culture was established from Wistar rat calvaria. Cell adherence and proliferation on titania ceramics for 28 days were examined. Morphological changes of the cells were observed by scanning electron microscopy. The activity of alkaline phosphatase of cells in various phrases were measured. Results The number of osteoblasts attached on the surface of titania ceramics was similar to that in control group. Osteoblasts could attach, spread and proliferate on titania ceramics. The activity of alkaline phosphatase of the cells increased with the culture time. Conclusion Titania ceramics may promote the growth and differentiation of osteoblasts in vitro.

Objective To improve metal-porcelain interfacial bond strength through treating the metal bottom with wiping crystal off and roughening it. Methods Crystals (NH₄Cl,NaCl and KMnP₄) with different shapes (globose, rectangular, cubical)，different granularitis（the diameter of 0.1,0.2, and 0.3 mm）and different proportions (the respective content of crystal in which was 30%,50%, and 80%) were chosen.After wax pattern was shaped,routinely imbedded ,casted and fused ,the drawing strength ,the cut strength,and the pressing strength were tested. Results The crystal (NH₄Cl) with globose shape was the best of the three shapes;the crystal (NH₄Cl) with the diameter of 0.2 mm was better than the other two and the content of 50% of crystal was the most appropriate proportion in the three kinds of crystal. Conclusion This method can improve the metal-porcelain interfacial bond strength greatly.

Objective To investigate the effect of endoscopic sinus surgery(ESS) and endoscopic endonasal dacryocystorhinostomy on the nasolacrimal duct and lacrimal sac fossa. Methods Using square caliper, compass and protractor,23 heads of adult cadaver specimens were examined including the shape, direction and the thickness of wall of lacrimal sac fossa. Results In fossa the proportion of the frontal portion of the maxillary bone was bigger than that of the lacrimal bone. There was about 20° between the rear wall of the fossa and the coronal plane. The superior sections of nasolacrimal ducts were circle or sector, there were different directions that each wall of nasolacrimal duct to the hard palate, the front wall of the nasolacrimal duct was thick and the other were thinner, less than 0.2 mm. Conclusion Before the endoscopic endonasal dacryocystorhinostomy was applied,the anterior portion of the medial orbital wall should be scanning, the gap should be enlarged enough, especially in lengthwise；the hiatus maxillaries shouldn′t be enlarged forward too much to avoid injuring nasolacrimal duct.

Objective To study the irradiation doses in several kinds of ⁶0Co radiotherapy patients in order to improve the treatment project, reduce and prevent radio-cystitis. Methods The method of thermoluminescence with a model of imitating human was adopted, radiotherapy course of ⁶0Co was imitated,the irradiation dose of bladder in patients was monitored. Results In a radical cure course of treatment, the irradiation-induced bladder doses of anterior-posterior (AP) and posterior-anterior (PA) of pelvis cavity， AP and PA of chest cavity and front region of lower neck were 42.8，27.5，0.33，0.17, and 0.06 Gy, respectively. Conclusion The irradiation doses of bladder in AP and PA of pelvis cavity are bigger than those in other mode, which may be the main reason for radio-cystitis.

Objective To study the relationship between chimerism of peripheral blood and donor-specific transfusion (DST) and acute kidney transplantion rejection. Methods The subjects were divided into 3 groups: group A, 35 subjects with corpse donor kidney transplantation who did not undertake DST before operation (2 or 3 HLA-mismatched); group B, 3 subjects with live donor kidney transplantation who did not undertake DST before operation (none was HLA-mismatched); group C, 22 subjects who undertook DST before operation (1 or 2 HLA-mismatched). PCR-SSP was used to check the HLA-DRB1 gene in peripheral blood. Results 12 subjects (34.3%) in group A, 3 subjects (100%) group B and 19 subjects (86.3%) in group C were found donor HLA-DRB1 gene. Chimerism was confirmed. 41.2% (14/34) subjects with chimerism and 38.5% (10/26) subjects without chimerism were found acute rejection,but there was no significant difference between two groups (P>0.05), as well as the frequency and times of rejction. Moreover, chimerism disppeared in all cases in group C and there was no rejection. Conclusion DST can lengthen the survival of the donor kidney by improving the formation of micro-chimerism. There is no accountable relationship between the disappearance of chimerism and acute rejection.

Objective To study the action of oxygen free radical (OFR) in pathogenesis of acute viral myocarditis (AVMC) and the therapeutic effect of selenium (Se) on myocardium damaged by OFR. Methods Se in serum, superoxide dismutase (SOD), glutathione peroxidase (GSH-P_X)，malonadelye (MDA) were detected in children, which included 60 cases of AVMC children and 60 healthy children as control, respectively. Children with AVMC were randomly divided into routine treatment group (30 cases) and Se-supplemented group (30 cases). Se-supplemented and routine treatment (anti-viruses, immune system modulation, myocardium nourishing) were used in Se-supplemented group. Only routine treatment was used in routine treatment group. The treatment period was 4 weeks. Through observation of changes of Se in serum, GSH-P_X，SOD，MDA，enzyme of myocardium, ECG and clinical symbols before and after treatment in two groups, the curative effects were assessed. Results OFR and LPO reactions appeared pathological changes in children with AVMC. Se in serum，SOD，GSH-P_X were decreased (P<0.001) and MDA was increased (P<0.001) in children with AVMC. After treatment with Se-supplemented, Se in serum, SOD，GSH-P_X activities were significantly increased (P<0.01, P<0.01,P<0.001)and MDA content was significantly decreased (P<0.001)， compared with before treatment, and all of the clinical symbols were disappeared. Enzymes of myocardium were normal. The percentage of ECG normalization was significantly higher than that of the routine treatment group (P<0.05). The curative rate of AVMC was increased. Conclusion Se is effective to clear OFR and able to protect myocardium through inhibiting OFR and LPO reaction. On the basis of routine treatment for viral myocarditis, Se-supplemented is more effective than routine treatment.

Objective To study the significance of proliferation and expansion of lymphatic capillary in pericancerous tissue in lymphatic metastasis of laryngeal carcinoma. Methods Immunohistochemistry method and automatic image analysis technique were applied to observe the states and correlated parameters of lymphatic capillaries in 75 laryngeal carcinoma tissues,75 pericancerous tissues and 8 normal laryngeal tissues. Results The lymphatic capillary was not seen in laryngeal carcinoma nests. The density of lymphatic capillary in pericancerous tissue was obviously higher than that in normal laryngeal tissue (P<0.01)，and had expansion with different degree, with parts of the walls of some lymphatic capillary distroyed .The density,diameter, and area of lymphatic capillary in pericancerous tissue were obviously higher than that in normal laryngeal tissue (P<0.01). The density,diameter, and area of lymphatic capillary in pericancerous tissue with cervical lymph node metastasis was obviously higher than that without cervical lymph node metastasis (P<0.01). Conclusion The proliferation and expansion of lymphatic capillary in pericancerous tissue are important factors in lymphatic metastasis of laryngeal carcinoma. This research offers theoretic basis for laryngeal carcinoma metastasis process, mechanism and prevention.

Objective To evaluate the efficacy of neoadjuvant chemotherapy (NAC) with nedaplatin, ifosfamide and peplomycin (SIP) for patients with locally advanced squamous cell carcinoma of cervix, and to determine whether or not the mitotic index and the Ki-67 labeling index (LI) obtained from biopsy specimens were useful as predictors of NAC response. Methods Twelve cases (stage Ib:3 and II:9) of cervical carcinoma with more than 4 cm in diameter were treated with 1 to 2 courses of SIP therapy followed by surgery. The response to SIP was measured by magnetic resonance imaging (MRI).　The relationship of response and serum SCC, and the changes of mitotic index (MI) and the Ki-67 LI in the biopsy specimens were investiged before and after SIP therapy. Results Seven cases resulted in partial response (PR) and 5 cases did not change (NC), the effective rate was 58.8%. In cases with PR, tumor size reduced markedly after the first course of SIP and serum SCC decreased. MI values in PR cases decreased, and Ki-67 LI was unchanged. On the other hand, in NC cases, MI values did not decrease and Ki-67 LI tended to increase. Conclusion NAC with nedaplatin, ifosfamide and peplomycin was effective for patients with locally advanced squamous cell carcinoma of cervix. Alterations of MRI, serum SCC, MI and Ki-67 LI can be predictors for NAC.

Objective To investigate the role of adrenomedullin (ADM) and endothelin-1 (ET-1) in the pathogenesis of pregnancy-induced hypertension (PIH). Methods The plasma concentrations of ADM in human umbilical vein of PIH patients（n=30） and normal late trimester pregnancy women （n=12）were measured by radioimmunoassay. The expressions of ADM and ET-1 in umbilical cord vessel of PIH patients（n=40）and normal late trimester pregnancy women （n=12） were detected by immunohistochemistry (SABC) . Results ①The plasma concentration of ADM in human umbilical vein of PIH patients was significantly higher than that of normal late trimester pregnancy women (P<0.05). The plasma concentration of ET-1 in human umbilical vein of PIH patients was similar to that of normal late trimester pregnancy women （P>0.05）. ②The expression of ADM was found in endothelium and smooth muscle cell of umbilical cord vessel，and it increased with the serious degree of PIH. The expression of ET-1 was only found in endothelium of umbilical cord vessel, and it decreased with the serious degree of PIH. Conclusion The changes of ADM and ET-1 in umbilical cord plasma and vessel may related to regulation of fetoplacental circulation in PIH.

Objective To study the effects of loop electrosurgical excision procedure (LEEP) on diagnosis and treatment for cervical intraepithelial neoplasia (CIN). Methods 212 patients with CIN, identified by colposcopy, cytology and pathological biopsy, were randomly divided into two groups:CO₂ laser excision treated (116 cases) and LEEP treated (96 cases), all excised tissues were examined with pathological method. Results After LEEP treatment, 94 cases of CIN were pathologically confirmed, 94.8% confidence interval in specimens, 89 out of these were healed with a cure rate of 94.7%. Two cases of total (2.1%) were diagnosed as cervical microinvasive carcinoma. There were 5 cases of postoperative recurrence with a recurrence rate of 8.8%. Among them, 4 cases demonstrated readable resection margin while 1 case didn′t. In the group of CO₂ laser excision, all pathological postoperative diagnoses referred to CIN₁-2. 103 cases were healed with a cure rate of 88.8%, and 13 cases observed as postoperative recurrence with the recurrence rate of 11.2% and the confidence interval in pathological specimens was 60.3%. There was significant difference of the confidence interval in pathological speciment postoperation between two groups (P<0.001).Conclusion Application of LEEP is effective and reliable in CIN diagnosis and treatment, which is also applicable to diagnosis of cervical microinvasive carcinoma.

Objective To study the characteristics of diabetes with mitochondrial DNA 16189 (T/C transition) mutation of population in North China. Methods PCR and RELP techniques were used to examine 155 patients with type 2 diabetes (diabetic group) and 164 heathy people (control group) to analyse the mitochondrial gene. Results ①The prevalence of mitochondrial DNA (mtNDA) 16189 T/C mutation in diabetic group ［32.26%(50/155)］ was significantly higher than that in the control group ［15.24%(25/164)］; ②No significant difference in fasting plasma glucose level, fasting C-peptide, postprandial C-peptide, GHbA₁c between those with mutation and those without mutation in diabetic group; ③ Clinical characteristics of diabetes with mitochondrial DNA 16189 T/C mutation: there was a late onset at diabetes (about 50 years old),there was no significant difference in sexuality(1.3:1); 74% diabetics with mutation had higher BMI; Blood lipid was found abnormal in 65% diabetics with mutation ; the prevalence of the mutation was high in diabetics with maternal family history. Conclusion The mitochondrial DNA 16189(T/C transition) mutation may be only a polymorphism in population.