Abstract:Objective To Study of in situ polymerase chain reaction assay for the detection of canine parvovirus(CPV) in Feline Kidney F 81 cells.Method Feline kidney F 81 cells were cultured in slide and infected with CPV. The cells infected with CPV in different stages were detected by direct in situ PCR, which used biotin labelled primers, and immunohistochemical method. Result In situ PCR method, a lot of signals were found in cells infected from 12 to 48 hours. Some signals were found in cytoplasm, and some in nucleus. In immunohistochemical method, no signals were found in cells infected for 12 hours, and sporadic signals in cells infected for 48 hours. The signals detected by these two methods showed significant difference. Conclusion The results indicate that the in situ PCR not only has high sensitivity, but also can locate the position of virus.

Abstract:Objective To investigate human SR AI transgenic integration and reproduction and its effect on mice breeding. Methods Human SR AI transgenic mice models 2, 3534,3560,3638,3639 have been bred by means of creating the system similar to the system ancestry. PCR and Southern blot techniques were used to test the DNA samples of tail tissues in the 5 lines of mice. Results In 431 baby mice of the 5 lines, 178 were tested by PCR to be positive (41.2%) while the positive rates of generations F 1, F 2, F 3 in line 3639 and pure zygote transgnic mice were 47.8%,71.3%,75% and 100% respectively. Conclusion Human SR AI gene can steadily heritable in the mice sub generation and doesn't influence obviously their reproduction and growth.

Abstract:Objective To establish a model of xenografted human ovarian carcinoma in severe combined immunodificiency (SCID) mice and homologous cell lines in vitro. Method A human ovarian serous papillary carcinoma derived from a surgical specimen was transplanted in the subcutis of SCID mice. After growth they were passaged mice to mice, meanwhile a human ovarian carcinoma cell isolated from xenografted tumor in mice was cultured in vitro and coloned for six months. The biological characteristics of xenografted tumor and homologous cells were determined by cell and molecular biological techniques. Results The xenografted tumor in SCID was successively passaged for 5 generations and a successful inoculation rate (90%) was observed during a period of 14 months. A tumor cell line (OVA 319) was established in 6 months and grown steadily, which isolated from the xenografted tumor. Histology and ultrastructure of microscopy showed that xenograted tumor retained its original growth characteristics and morphological properties. The chromosomal analysis of xenografted tumor cells and OVA 319 cells revealed aneuploid pattern of 12 46 in number, exhibiting features of human carcinoma. Furthermore, the FCM analysis and RT PCR technique showed same features as those of patient with original tumor tissue and xenografted tumor and OVA 319 cells, including distribution of cell cycle and it grew rapidly, and the expression of MAGE 2 gene at the mRNA level was observed. Conclusion Establishment of xenografted human ovarian carcinoma in SCID mice and OVA 319 cell lines could be considered as a model and provided experimental material for further investigation of human ovarian tumor.

Abstract:目的用辣根过氧化酶(HRP)逆行追踪技术探讨异种神经移植后神经纤维的再生.方法将多次冻融处理后的兔胫神经移植于大鼠坐骨神经,术后第2、4、6、8和10周,将HRP注人大鼠坐骨神经吻合部远侧端.结果移植术后第4周起在L4～5脊神经节见到HRP标记细胞,从第6周在腰段脊髓前角内见到标记细胞,其数量随术后存活期延长而增多.术后4周在移植神经内见少量再生神经纤维,6周后再生神经纤维穿过异种移植神经进入大鼠坐骨神经远侧端.结论自移植术后4周起,移植神经内已有再生纤维并部分恢复了轴浆流,证实了用HRP法可反映移植后神经纤维的再生情况.

Abstract:Objective To establish RT PCR method for detecting Sendai virus in live vaccine and vaccine producing medium. Methods The virus RNA was extracted from allantoic fluid of 9 days old chicken embryo inoculated with Sendai virus E17 strain after 72 hours, and was reverse transcribed into cDNA. Then cDNA was amplified by outer primer sets and inner primer sets respectively, designed according to the NP gene sequence of Sendai virus.The PCR products were cloned into T vector and sequenced. The sensitivity experiment was performed by serially diluting allantoic fluid, extracting RNA and then amplifying by RT PCR. This method was used to detect Sendai virus in Japanese encephalitis attenuated live vaccine and the kidney of nurturing hamster, which was used for producing vaccine in China for years. Results Two fragments, 684bp and 248bp, were amplified by using outer primer sets and inner primer sets respectively. The sequencing result of PCR products of outer primer sets was completely homology with the nucleotide sequence reported in Genbank. The sensitivity experiment indicated that 10 -4 virus titer was detected by the first PCR with the outer primer sets and 10 -7 virus titer by nested PCR with the inner primer sets. The results of detecting Japanese encephalitis attenuated live vaccine and the kidney of nurturing hamster were negative. Conclusions The RT PCR method for detecting Sendai virus was highly sensitive and special.

Abstract:目的建立在猫肾F81细胞中犬细小病毒原位PCR的检测方法.方法在猫肾F81细胞上感染犬细小病毒,设计特异性引物,用直接原位PCR法在染毒12h,24h,48h细胞片上检测出犬细小病毒,并与常规免疫组化的方法进行了比较.结果在染毒48h的细胞片上,用阳性记分法将两种检测方法得到的阳性细胞进行统计比较,差异极显著(P＜0.001),用原位PCR法所得出的阳性率高.结论原位PCR法检测犬细小病毒具有敏感性高和组织定位的优点.

Abstract:Objective To investigate whether HCMV infection can cause ocular tissue lesions in rats and mice.Methods Twenty rats and ten mice were inoculated with HCMV AD 169 strain by vein and development of ocular disease was observed.HCMV DNA of tissues from inoculated animals were examined by in situ hybridization.Results Some animals expressed much secretion,turbidity in eyes even losing sight at different time post\|inoculation.HCMV DNA sections were identified in the corneal endothelium cell,rod cell and cone cell of the rats and mice inoculated. Conclusion HCMV AD 169 strain can infect rats and mice and cause ocular disease.\;

Abstract:Objective To replicate the acute hypoxic ischemic brain damaged animal model.Methods The uterine arteries of full\|term pregnant mice were ligated to interrupt the blood supply by cesarean section.The baby mice with asphyxia were observed for their physical and behavior development and their pathological changes in their brains.Results With prolonging the duration of interrupting the blood supply,the morbidity in fetal mice increased.The relationship between those was positive linear correlation ( P <0.05).The development of body weight and behavior in baby mice was reatrded.The pathologic changes in the brains were apparent,which were similar to the neonatal.Conclusion The animal model can be used for the study of the neonatal hypoxic ischemic brain damage.\;

Abstract:目的目前的虚寒证模型存在两个环节的不足一是可能未能在寒证的基础上达到寒邪伤阳而致虚;二是虚证上未达到"精气夺则虚”.本实验尝试通过加大造模量改进这一模型.方法雌性WistarⅡ级大鼠,按照动物体温分为对照组和虚寒证组.虚寒证用较典型的造模药物灌胃造模,但用量较大,时间较长.造模结束检测有关指标.结果与对照组比较,虚寒证组动物外观差,舌质淡嫩,明显腹泻,体重增长减缓,体温较低,食量减少,游泳时间缩短,受激排便增多.血浆cGMP含量、乳酸脱氢酶活性、肾脏指数、脾脏指数、肝脏指数、肾上腺指数显著增高,皮质醇含量、T含量、卵巢指数、血清乳酸含量显著降低,cAMP/cGMP比值,E2含量、E2/T比值、胸腺指数有降低趋势.血清IL-2含量、血清木糖吸收率、血清TT3含量无显著差异.结论本模型有程度较深的寒证表现,并出现虚证.

Abstract:Objective To improve the efficiency of producing transgenic mice.Methods The major steps were the preparation of DNA fragments for microinjection,the preparation of eggs with high quality,and the microinjection and implantation optimized.Results The born mice were examined by PCR and Southern Blot.At the beginning,we got 2 transgenic mice in 146 born mice.After improvement,we got 10 transgenic mice from 34 born mice.The ratio of transgenic founder mice was improved from 1.4% to 30%.Conclusion The technical platform for transgenic research has been establised due to the great improvement of the efficiency of producing transgenic mice.

Abstract:Objective To observe the injury and permeability of intestine, together with the changes of peroxidation and TNF and IL 6 concentrations after hemorrhagic shock in rats. Methods At 0, 2, 6, 12, 24 and 48 hours after being subjected to sham shock, or 70 minutes of 30mmHg shock, rats were sacrificed. The SOD activity and the MDA, TNF and IL 6 concentrations in serum and intestine were detected, the histology and bacteria translocation of intestine were also observed. Results The serum MDA concentration was increased at 0-2h after shock and SOD activity was increased almost all the detected time. In the intestine, the MDA concentration increased at 0-24h, and SOD activity decreased at 0-12h. The serum TNF concentration was increased at 6-48h after shock while increased at 0-12h in intestine. No change was observed of IL 6 concentration in both serum and intestine. Extensive sheding of mucosa epithelia were observed at 2h after shock and the bacteria were observed at 6-12h in lamina propria. The indigenous bacteria of intestine were detected at mesenteric lymph nodes and other visceral organs at 6-48h after shock. Conclusion The SOD activity decrease was one of the reasons in intestine injury after hemorrhagic shock. The early phase increase of serum TNF came from the injured intestine.

Abstract:Objective To study the effect of exogenous basic fibroblast growth factor (bFGF) on delayed peripheral nerve regeneration. Method Transection was made on the right sciatic nerve of 50 Sprague Dawley rats. Reparing operation was made 12 weeks after transection. bFGF was injected into 25 rats as experimental group and only saline into other 25 rats as control group.All animals were observed through neuroelectro physiological examinations and histological examinations at 2,4,6,8 and 12 weeks after the reparing operation. Results In both experiment and control groups, there could be found significant nerve regeneration. In experiment group, more axons grew in the distal nerve segment at the fourth week postoperatively. The motor nerve conductive velocity,the muscle evoked action potential, the thickness of myelin, and the diameter and cross section area of regenerating axons in experimental group were better than those in the control group with a great difference. Conclusion bFGF can enhance the delayed peripheral nerve regneration.

Abstract:Objective To investigate the degradation of Human\|hair keratin artificial tendon (HHKAT) material and evaluate its biocompatibility in body. Methods The HHKAT materials were divided into two groups: Z and F according to disposed time, with normal human hair used as control group O. The Z, F and O samples were implanted into the two sides of spine muscle in 12 rabbits. The rabbits were killed 2,6,12,24 weeks after operation, observing the degradation progress of HHKAT and its tissue reaction. Result The degradation time of HHKAT was different according to disposed time: group F absorbed completely, group Z absorbed a little in 24weeks, while O groups showed no absorption. Histologic investigation showed that HHKAT and normal human hair did not evoke apparent inflammation reaction, and the histologic reaction decreased with degradation of HHKAT. Conclusion HHKAT is a kind of material with good biocompatibility, and can be in degradation according to requirement.

Abstract:Objective To clarify the G banding pattern and spontaneous aberration frequencies of a new inbred line mouse IRM 2. Methods The bone marrow chromosome preparation and G banding pattern for mouse were applied.Results Of the 50 G banding cells from 20 mice,10 cells were selected for banding analysis. The chromosomes were distinguished by characteristic banding patterns. The relative length and standard deviation were depicted. Some discerned points between similar banding of chromosomes were presented, such as 1, 6 and X, 4 and 5, 9, 13 and 14. In addition, 1200 metaphase cells were analysed by conventional Giemas stain. The results showed that break was 0.33%, acentric aberration and translocation were 0.08%. The spon taneous aberration frequencies were very low. Conclusion The discernment of G banding pattern of a new inbred line mouse IRM 2 is for providing scientific basis to study on abnormal structure, radiation effect, cancer research and gene map.

Abstract:目的为进一步深入研究Smad3基因在脊椎动物发育中的重要作用,对Smad3基因剔除小鼠进行保种和繁育研究.方法采用基因剔除杂合子小鼠进行保种,通过PCR和Southern杂交对杂合子小鼠交配所产生的后代进行基因型鉴定,纯合子小鼠和野生型小鼠用于表型分析,杂合子小鼠用于留种和繁殖生产.结果采用PCR方法对278只子代小鼠进行了基因型鉴定,83只为野生型,133只为杂合子,62只为纯合子.结论Smad3基因剔除突变能稳定遗传.采用杂合子小鼠保种,子代小鼠三种基因型比例符合孟德尔遗传定律.

Abstract:目的建立雌激素替代治疗动物模型,观察其血清中一氧化氮(NO)含量的变化.方法取40只雌性大鼠切除双侧卵巢,随机分成四组A组假手术组;B组单纯去卵巢组;C组去卵巢+雌激素治疗组(隔日肌注苯甲酸雌二醇5μg);D组去卵巢+雌孕激素治疗组(隔日肌注苯甲酸雌二醇5μg,黄体酮1mg),两月后检测血清中雌二醇(E2)、孕酮(P)及NO2-/NO3-的含量.结果(1)卵巢切除后血清E2、P和NO2-/NO3-浓度显著降低;(2)雌激素治疗组和雌孕激素治疗组血清E2和NO2-/NO3-的含量及雌孕激素治疗组血清P的含量,略低于A组而显著高于去卵巢组.结论所建立的动物模型可以模拟绝经及绝经后的小剂量雌激素替代治疗,动物模型体内雌激素浓度与具有抗衰老作用的NO浓度呈相互平行的变化关系,雌激素可能通过NO达到延缓衰老作用.

Abstract:目的采用体外无血清培养方法,研究雌激素(E2)对恒河猴睾丸Leydig细胞睾酮分泌的影响.方法恒河猴睾丸活体组织,经胶原酶消化,BSA密度梯度重力沉降分离细胞,得到的Leydig细胞置二氧化碳培养箱培养,收集培养液,应用放射免疫测定法测定培养液中的睾酮含量.结果不同剂量的睾酮前身物孕烯醇酮、孕酮和17α-羟孕烯醇酮都能促进睾酮的含量升高,其中以17α-羟孕烯醇酮的作用最为显著;当孕烯醇酮、孕酮和17α-羟孕烯醇酮分别与E2合并应用时,有明显地抑制作用,同样以17α-羟孕烯醇酮的抑制作用最为显著.结论E2抑制恒河猴Leydig细胞睾酮的分泌是由于影响了3β-羟类固醇脱氢酶、17α-羟化酶以及17,20-碳裂酶的活性所致.