Abstract:目的　使用免疫外科法分离克隆BALB c小鼠胚胎干细胞 (embryonicstemcells,ES细胞 ) ,为进一步建立BALB c小鼠ES细胞系打下基础。方法　用免疫外科法从 4 5枚BALB c小鼠囊胚中分离得到 2 0枚去除滋养层细胞的ICM ,接种在MEF饲养层上 ,使用DMEM(高糖 ) 15 ?S 0 1mmol Lβ 巯基乙醇 0 0 1mmol L非必需氨基酸 10 0 0IU mlLIF 10 0IU mL青霉素 10 0IU ml链霉素培养液 ,17枚形成典型的ICM集落 (85 0 % ) ,有一枚胚胎传至第 10代。用于全胚培养的BALB c小鼠胚胎共 10 2枚 ,使用与免疫外科相同的培养方法 ,6 6枚形成典型的ICM集落 (6 4 7% ) ,其中一枚胚胎传至第 8代。结果　免疫外科法较全胚培养法有利于小鼠ES细胞的分离与克隆 ;添加LIF(10 0 0IU ml)有利于小鼠ES细胞的分离与传代 (P <0 0 5 ) ;0 0 5 %胰酶 0 0 0 8?TA是较好的ES细胞消化液 ,对细胞综合损伤力小 ,且传代后ES细胞集落形成能力也较高 (P <0 0 5 )。结论　分离得到的ES细胞经形态学观察 ,AKP染色 ,体外分化实验 ,核型分析等证明其具有胚胎干细胞的诸多特性。

Abstract:Objective To study the change of ultrastructure of bone and the change of serum biochemical indexes after spinal cord injury (SCI). Whether the bones above and below the level of trauma are affected differently?To know well to what extent the secondary osteoporosis after SCI is. Method 110 Wistar rats 300-320 g weight ,4-5 months old were used. The control group only removed lamina with dura intact , without spinal cord injury at the level of the tenth thoracic vertebrae. The experiment group removed lamina , then , the SCI model of rats was made by the method of Allen's (60gcm). The experiment group's BBB (Basso, Beattie and Bresnahan ) score was 0 or 1 two weeks after operation. We observed the change of the serum concentration of calcium, phosphorus and alkaline phophatase (ALP) , the change of ultrastructure of osteocytes at the sites of tibial plateau and the surgical neck of the humerus. Results and Conclusion The bones above and below the level of trauma are both affected after SCI, but different extent of osteoporosis can be seen in different bones .The change of ultrastructure of osteocytes is remarkable when osteoporosis has happened.

Abstract:Objective To evaluate the usefulness of nested-polymerase chain reaction (nested-PCR), indirect fluorescent antibody assay (IFA), immunosuppressive provocation tests, and histopathological diagnosis assays for detecting Clostridum Piliforme (C. Piliforme) infections in laboratory rats. Methods In this study, the nested-PCR primers, which we synthesized to amplify C. Piliforme 16S ribosomal DNA (rDNA) sequences as an identification system for this organism, were performed by using C. Piliforme and 15 non-C. Piliforme strains. Specimens taken from 20 immunosuppressive Wistar rats and 5 non-immunosuppressive SD rats and submitted to detect C. Piliforme by nested-PCR. Results The C. Piliforme reference strain revealed positive result indicated by a 196 bp specific fragment on gel, while 15 non-C. Piliforme standard strains gave negative results demonstrating that the inner primers should be specific for C. Piliforme tested. The nested-PCR would be able to detect as little as 10 pg C. Piliforme-DNA. Tissues samples taken from rats and submitted to detect C. Piliforme by nested-PCR were found negative results, and the results of nested-PCR and consistent with those of conventional bacteria identification tests and histopathological diagnosis assays. 25 serum samples taken from rats were submitted to detect C. Piliforme antibodies by IFA were found heterologous cross-reactive antigens such as some strains of Clostridia and Bacillus. Conclusion IFA is best used for colony surveillance, and follow-up testing may include immunosuppressive provaction tests. Confirmation of C. Piliforme infections is best performed by PCR, bacteria identification tests, and histopathological diagnosis assays. In our study, the results of nested-PCR are consistent with those of bacteria identification tests and histopathological diagnosis assays. The data have also demonstrated that the nested-PCR established here is a quick, reliable, sensitive and specific method for detecting C. Piliforme infections in laboratory rats.

Abstract:Objective To discuss the effect of in vitro fertilization (IVF), embryos and sperm cryopreservation in various mouse strains. Methods IVF was performed in 13 mouse strains(C 57 BL/6J,BALB/c,C3H/HeJ,ICR,KM,FVB,MRL,NOD,CBA,DBA/2,CD-1,BDF1,B6C3F1) in two laboratories of CARD and SLAC to compare the IVF, embryos thawed and embryos transfer. Results The fertility rates of fresh spermatozoa by in vitro were from 15.1% to 87.9%, and frozen sperm were about 8%-80%, the thawed rates of frozen embryos were from 42.6% to 83.9%, and 17.8% to 51.8% offspring were procreated after morphological normal 2-cell embryos were transferred to the oviducts of pseudopregnant recipient in all strains. Conclusion These results suggested that there were distinct differences in the rate of fertilization, thawed and transfer in different genetic background. But no significant differences(P>0.05) were found in each strain in CARD versus SLAC in spite of the high fertility rates(P<0.01) of frozen sperm in the former.

Abstract:Objective To study the influence of tonifying kidney plus strengthening vital energy recipe on the nonspecific immunity function in H 22 tumor transplanted mice treated with cyclophosphamide(CY). Methods The low immunity model was established in H 22 tumor transplanted mice treated with CY. The activity of natural killer cells and macrophages was measured by LDH release test. The expression of IL-12 mRNA in spleen cells was examined by RT-PCR. Results The recipe could markedly improve the activity of natural killer cells and macrophages in low immunity mice treated with CY and the expression of IL-12 mRNA in spleen cells. Conclusion The recipe can greatly increase the nonspecific immunity function treated with CY in tumor transplanted mice.

Abstract:Objective To investigate the metabolic derivatives of the glyceryl trinitrate (GTN). Methods Rabbits were used in this work. Blood pressures were measured by dynamic analyzer, methemoglobin (MetHb) by blood gas analyzer and alanine transferase (ALT) by auto-biochemist analyzer. Nitrate, nitrite and reduced glutathione (GSH) were measured with colorimetry. Results Blood pressures and GSHs in red blood cells showed significant decrease, nitrite showed significant increase, MetHb and ALT showed no remarkable difference before and after GTN administration. Conclusion Metabolic GTN exhausts thiols and yields nitrite, and it has no effects on hemoglobin and liver.

Abstract:目的　应用快速心室起搏的方法制备扩张型心肌病心力衰竭犬模型 ,并对其方法学进行改进。方法　 6只成年健康杂种犬麻醉后先通过射频消融希氏束致Ⅲ度房室传导阻滞 ,再经静脉植入心内膜起搏电极 ,脉冲发射器为技术处理后的人用永久起搏器。起搏频率为 2 5 0次 min ,起搏时间持续 (2 3 6± 2 5 7)d。结果　除有相应症状、体征外 ,所有动物均成功制备为扩张型心肌病心力衰竭模型 ;B超和病理解剖检查证实有心脏扩大、心室壁变薄以及肝瘀血、肺瘀血 ;病理切片显示心肌细胞空泡变性、肝窦扩张充血、肺泡内含铁血黄素沉积。结论　改进后的快速心室起搏制备充血性心力衰竭动物模型的方法更为安全、可靠、实用。

Abstract:Objective In order to obtain a feeder layer for ES cells transfected by pTRE-Ins-human gene by means of developmed of the hygromycin-resistant 3T3 cell line. Methods The plasmid pHyg containing hygromycin gene was purified and transfected into 3T3 cells with Lipofectin.The transfected cells would be survived in further culture medium containing hygromycin B antibiotic as the hygromycin gene could express a hygromycin resistant products.The identification of the hygromycin resistant 3T3 cell line was conducted by polymerase chain reaction with the specific primers of hygromycin gene. Results The hygromycin R 3T3 cell line was established successfully.There were no differences among hygromycin R 3T3 cells and normal 3T3 cells morphologically and propagatively. The hyg gene DNA fragment could be amplified by PCR with specific primers in the genomic DNA of hygromycin R 3T3 cells. Conclusion The hygromycin R 3T3 cells were established with transfection of plasmid pHyg by Lipfectin,which may be useful for selecting ES cells transfected by pTRE-Ins-human gene.

Abstract:Objective To study the way of infection with eperythrozoon, try to set up the model of infecting animal with eperythrozoon. Method Mice were infected through abdominal cavity, subcutaneous way with the blood from infected ruminants, suspension from infected viscera of mice,touching with infected mice. Result A regularity of the eperythrozoon in mice appeared. The time of appearance, peak and disappearance of eperythrozoon in mice was respectively 8th days,15th-18th days,19th days after infected. During the experiment no obvious clinical symptom was found in infected mice. Conclusion The model of infection with eperythrozoon was established in mice.

Abstract:目的　观察α 半乳糖苷酶对猕猴类人B抗原的酶解效果 ,探讨α 半乳糖苷酶酶解对猕猴红细胞结构、功能的影响。方法　采用热吸收放散试验从 30只华南猕猴中选取类人ABO血型抗原较强的 2只A型、3只B型猕猴做为实验对象 ,以基因重组的α 半乳糖苷酶体外酶解猕猴类人B型血抗原 ,并回输到A型猕猴体内 ,测定红细胞脆性、自身溶血率、胆固醇、高铁血红蛋白、乙酰胆碱脂酶、ATP等红细胞的结构功能指标。结果　经α 半乳糖苷酶酶解后 ,猕猴红细胞胞膜完整、携氧能力正常 ,酶解后的“通用”型血回输给受体猕猴无任何输血反应发生。结论　α 半乳糖苷酶酶解对于猕猴红细胞的形态、结构、功能无不良影响 ,且在实验动物体内是安全的。

Abstract:目的　观察实验性Ⅰ型糖尿病恒河猴的组织病理学变化。方法　 7只恒河猴分别静脉注射不同剂量的链脲佐菌素 (streptozotocin ,STZ) ,建立STZ性Ⅰ型糖尿病恒河猴模型 ,观察其心脏、肾脏、胰脏、脾脏等组织病理学变化。结果　动物胰岛数量明显减少 ,分布稀疏 ,残存胰岛萎缩 ,胰岛大小不等 ,成纤维细胞增生。肾小球增大 ,毛细血管壁增厚、僵硬 ,肾小管内膜细胞玻璃样变性 ,肾小球球囊内皮细胞增生。心肌变性、坏死、淤血 ,心肌细胞肥大 ,中、小血管壁增厚 ,血管壁纤维组织增加 ,冠状动脉内膜局部增厚 ,内皮细胞增生。脾脏小动脉硬化。结论　通过对STZ诱发的Ⅰ型糖尿病模型胰岛、肾脏和心脏等结构病理观察说明 ,该动物模型可用于糖尿病组织病理研究和药物疗效的评价。

Abstract:Objective To explore the influences of MMP-1,MMP-9 on metrorrhagia induced by bacterial infection in experiment Rhesus monkey. Methods The immunohistochemical S-P were performed to estimate express level of MMP-1,9 and TIMP-1 in the endometrium of model group and normal control group. Result Compared with control group, MMP-1 and MMP-9 were much expressed in endometrial stroma and gland and TIMP-1 are expressed similarly. Conclusion The over-expression of MMP-1,9 may participate in metrorrhagia induced by endometritis in Rhesus macaques.

Abstract:Objective To test protective effect of inactivated HA vaccine on marmosets.Methods Inactivated HA vaccine was prepared by culturing the strain YN5 of HA virus in Vero cells. Its immunogenicity and protection was studied in marmosets(Saguinus). Results Specific anti-HAV antibodies were induced in 7 common marmosets, and no elevation of serum enzyme activity or pathological exchange of liver tissue were observed. The vaccine protected marmosets against the intravenous challenge with virulent HAV(Han-ZD). But the elevation of serum enzyme activity and pathological exchange of liver tissue were showed in all the control marmoets. Conclusion The inactivated HA vaccine (YN5) had good immunogenicity and protective effect.

Abstract:目的　观察新药“蜈龙抗癌颗粒”对四种小鼠移植性肿瘤的治疗效果 ,初步探讨该制剂对小鼠的毒副作用。方法　在小鼠急性毒性试验的基础上 ,选择三个剂量对小鼠移植性肿瘤Lewis肺癌、P388白血病、S1 80 肉瘤和H2 2 肝癌进行治疗实验 ,同时检查一批治疗Lewis肺癌实验小鼠的 15项血液学指标和脾脏、胸腺的相对重量。结果与结论　“蜈龙抗癌颗粒”在生药剂量为每日每公斤体重 10、2 5、4 0g(Lewis肺癌、P388白血病 )和每日每公斤体重 15、30、5 0g(S1 80 肉瘤和H2 2 肝癌 )时 ,对小鼠移植性肿瘤有肯定的治疗效果 ,并呈良好的量效关系 ,对小鼠Lewis肺癌的治疗效果最明显。该制剂大剂量时可能对小鼠的造血系统和免疫系统有明显影响。

Abstract:Objective To study the changes of SOD, MDA and GSH-Px in sera of Monglian gerbils.Methods The gerbils at the age of 1 month, 3 months and 24 months were used in the experiment, and the sera were collected for determining the SOD,MDA and GSH-Px. Results At the age of 3 months, MDA was the lowest, but GSH-Px was the highest, while the SOD was ascending with growing. Conclusion The changes of GSH-Px showed the direct relation to MDA, while the changes of the SOD was irrespective to the changes of MDA.