Laboratory Animal Science

The Research on Immunoregulation Effects of Guben Qingfei Tiegao

LI Xian-Hua, DU Jia-Lin, SONG Da-Fu, CHEN He, AN Ran, GUO Zhen-Wu

2011, 28(05): 1-02.

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Objective To research the immunoregulation effects of Guben Qingfei Tiegao. Methods chicken’s erythrocytes is used as immunogen, the blood-serum erythrocytolysin test is used to observe the humoral-mediated effects of Guben Qingfei Tiegao; the delayed hypersensitive reaction in mice test which is caused by 2,4- dinitrochlorobenzene is used to observe the cell-mediated immunity effects of Guben Qingfei Tiegao .Results Guben Qingfei Tiegao could promote the form of blood-serum erythrocytolysin, restrain the delayed hypersensitive reaction in mice which is caused by 2,4- dinitrochlorobenzene. Conclusion Guben Qingfei Tiegao has the effects of immunoregulation

Behavioural Comparison of Fmr1 Knockout Mice at 30 Days Age in Step-down Test

2011, 28(05): 3-06.

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Objective This study was designed to observe the cognition of Fmr1 knockout mice at 30 days Age in step-down test. Method Fmr1 knockout mice were identified using the PCR technical and step-down test were used in the study. Animals were tested for two days. The latency and the number of errors were recorded. The data was analyzed with Multifactor Variance Analysis. Result KO mice obviously had the shorter latency than WT mice, and KO mice obviously had more errors than WT mice (P<0.05); On the first day, the latency and the number of errors of KO mice had no difference compared with the second day (P>0.05); On the first day, the latency and number of errors of WT mice had significant difference compared with the second day (P<0.05). Conclusion Fmr1 knockout mice displayed cognitive impairment in the step- down test.

The Study of Randomly Amplified Polymorphic DNA Characteristic in HFJ and MIJ Inbred Rats

CAI Yue-Hua, LI Jian-Hui, ZHENG Long, LIU Jun-Xu, WANG Jun-Xia, XU Zeng-Nian, SHI Rong-Zhen, LIU Fu-Ying

2011, 28(05): 7-11.

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Objective To establish the fingerprint of HFJ and MIJ inbred rats by randomly amplified polymorphic DNA (RAPD) analysis. And to compare the DNA polymorphism of the four inbreeds rats including HFJ, MIJ, Lewis and F344. Methods To isolate rats genomic DNA by standard procedure of phenol/chloroform. Sixty primers were used to amplify rats DNA with RAPD-PCR. Thirty primers could produce clear fragments and were selected to analyze the DNA polymorphism between HFJ and MIJ inbred rats. Twelve primers were selected to analyze the DNA polymorphism among HFJ, MIJ, Lewis and F344 inbred rats. Results 1-7 clear bands were detected by electrophoresis on agarose gel in HFJ and MIJ inbred rats with thirty primers respectively. All amplified bands by random oligonucleotide primers were located between 200bp-1100bp except primer O-08, which located over 2000bp and there was a distinct band near the origin of electrophoresis. The results of RAPD indicated that there were identical completely between HFJ and MIJ inbred rats or within the two strains. DNA polymorphisms among HFJ, MIJ, Lewis and F344 inbred rats were studied by RAPD using twelve primers. The results indicated that 5 primers could to obtain difference fragments in all twelve primers. Conclusion With the thirty random primers to establish RAPD fingerprint of HFJ and MIJ inbred rats. Since the two inbred strains originated from a pair of Wistar rats, DNA polymorphism between HFJ and MIJ inbred rats were no detected, although the two inbred strains had remarkable genetic difference. By RAPD analysis, DNA polymorphism among HFJ, MIJ, Lewis and F344 inbred rats were detected.

Establishment of Cell Culture System for Kilham Rat Virus (KRV) using Rat Glial Cell Line C6

LIU Xian-Ju, TONG Wei, ZHANG Li-Fang, WANG Yan-Rong, GAO Zi-Qi, ZHANG Hui-Min, RONG Rong, LIU Yun-Bo

2011, 28(05): 12-14.

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Objective To establish a culture method for Kilham Rat Virus (KRV) using the Rat glioma cell line (C6) instead of primary rat embryo cells (RE). Methods 500 TCID50 KRV was inoculated into C6 (The C6 cells were diluted to 2×105/ml and then cultivated overnight before infection) and the infected cells were cultivated until the CPE (cytopathic effect) reached “++~+++”. Then, the viral antigen was identified by immunofluorescence (FITC) assay and the viral titer in the supernatant of culture was detected by hemagglutination test (HA). In addition, the virus was identified by DNA sequencing of the conserved regions of FRV genome. And the virulence of viral culture (TCID50) was determined by the 96 wells cell culture method. Results The CPE of C6 cells reached “+++” at 4~5 day post infection with KRV. The result of FITC indicated that the viral antigen was positive for KRV, and HA titer of the viral culture reached 1:5,120.The cultivated virus was identified as KRV, as the sequence of it had 97% similarity with KRV, and the virulence of KRV obtained by infection on C6 cells was 104.6 TCID50/0.1mL. Conclusion The infection of KRV on C6 cells and subsequently identification experiments indicated that KRV could be maintained in C6 cells instead of RE cell.

Primary Culture of Tracheal Smooth Muscle Cells of Guinea Pigs

HAO Zhi-Hui, ZHAO Hou-De, ZHANG Yu-Yang, WANG Xiao-Xiao

2011, 28(05): 15-18.

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Objective To establish a method of primary culture tracheal smooth muscle (TSM) cells of guinea pigs. Methods Cultured TSM cells of guinea pigs by tissue cultivation and identified TSM cells by the method of immunocytochemistry stain with specific α- smooth muscle actin antibody. Results Cells in fusiform or polygon shape erupted around the pieces of smooth muscle tissue and grew and proliferated radiat with a nucleus lying in the middle of cytoplasm in 5―8 days. The immunocytochemical stain of the monoclonal antibody of anti-smooth muscle α-actin was positive and the purity of the cells was above 95%, which indicated that most of cultured cells were smooth muscle cells. Conclusions The method of culture TSM cells of guinea pig is successful and can be applied.

Comparisonof Two Different Material Affecting Osteablast on Sticking to the Wall and Growth

LI Jian-Ying, XU Yong-Hua, ZHANG Dong-Hui, XU Qin, SHI Wen-Hui, LI Jia-Jia, MA Na, DONG Xiang

2011, 28(05): 19-21.

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Objective To compare polylysine and d collogen made by rat’s tail affecting on sticking to the wall of culture dishes in vitro. Methods We paved culture dish’s wall with polylysine and collagen made by rat’s tail, then to planted osteoblast. At 24h 48h 72h, we observed, under microscope, the cell’s morphous and area percentage which they stuck to the wall of culture dishes. Result In the polylysine group, at 24h, 48h, 72h, osteoblast adherence displayed hypodispersion and adherence area ratio is 30.6%±12.9%, 43.8%±14.5%, 56.3%±15.4%, respectively. In the collagen made by rat’s tail group, at 24h 48h 72h, the cell adherence area ratio is 42.5%±15.7%, 61.3%±17.1%, 94.4%±14.1%, respectively. At 24h, the cell adherence displayed mass; at 48h, the cell displayed mass and conjunction each other; at 72h, the cell displayed plaque. Two groups have significant difference. Conclusion The results showed that collagen made by rat’s tail is much suitable for osteoblast sticking wall growth.

Cardioprotection of Cardiomyopeptidin in a Mini-pig Model of Acute Myocardial Ischemia-reperfusion Injury

TANG Yue, QIAN Xin, WU Ai-Li, TIAN Yi, YUAN Wei-Min, PENG Peng, ZHANG Bao-Jie, SUN Jing, RUAN Ying-Mao

2011, 28(05): 22-25.

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Objective To observe whether Cardiomyopeptidin can protect myocardium in a mini-pig model of acute myocardial ischemia-reperfusion injury. Methods Twelve mini-pigs were used. A balloon was inflated in the left artery descending to block the blood and result in myocardial ischemia for 90 minutes. Then the balloon was deflated to induce reperfusion injury. Sixty minutes after the balloon inflation, Cardiomyopeptidin was administered by intravenous drop infusion in test group (n=6) while 0.9% sodium chloride was used in control group (n=6). Before ischemia and at the end of and 24 hrs after administration, lactate dehydrogenase (LDH), creatine kinase (CK), and creatine kinase isoenzyme (CK-MB) in serum were detected, echocardiography was performed. Twenty-four hrs after administration, animals were sacrificed. Necrotic area was measured by triphenyhetrazolium chloride (TTC) staining and histopathology was described and scored. Apoptosis was detected by TUNEL. Results Five mini-pig of each group survived. There was no statistical difference between the two groups (P>0.05) in LDH, CK and CK-MB. The left ventricular end diastolic volume (LVEDV) of test group was increased and higher than the control group at 24 hrs after administration (P<0.05). There was no statistical difference (P>0.05) between the two groups in the rate of infarction area to all myocardium area by TTC staining and the score of myocardial injury. There were fewer apoptotic myocardial cells in test group than in control group (P<0.05). Conclusion In a mini-swine model of acute myocardial ischemia-reperfusion injury, Cardiomyopeptidin may protect myocardium by decreasing myocardial cells went apoptosis. However, the infarction area doesn’t be reduced and the function of heart doesn’t be proved only by Cardiomyopeptidin in a short-term when the myocardial injury is severe.

Establish an acute renal failure model induced by cisplatin in Guizhou miniature pigs

DONG Xiao-Jun, DING Dou, WU Shu-Guang, QIAN Ning

2011, 28(05): 26-28.

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Objective To establish an acute renal failure model induced by cisplatin in Guizhou miniature pigs. Methods Guizhou miniature pigs were randomly divided into the control group and the model group. Animals of the model group were given cisplatin(4 mg/kg) by a single peritoneal injection. Urine samples were collected by a bladder fistula. Urinary output, urine level of creatinine and urea nitrogen were dynamically observed on the 1st hour before injection and the 1st，3rd，6th，24th，48th，96th hours after cisplatin treatment. The excretions of creatinine and urea nitrogen were calculated on the basis of the above determination. Results Urinary output，excretion of creatinine and urea nitrogen of the model group were significantly lower than that of the control group. Conclusions This study established successfully the acute renal failure model induced by cisplatin (4 mg/kg) in Guizhou miniature pigs.

Quality Status of Conventional Rabbit and the Corresponding Managerial Measure of Animal Experiment Department in Present

FAN Wen-Ping, HE Zheng-Ming

2011, 28(05): 29-34.

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The animal experiment is an important link node on the laboratory animals industry chain, the animal experiment managers are most familiar the situation of animal use in the domain, are most sensitive to the laboratory animal quality too. this article from the animal experiment department's observation angle elaborated the quality status of our country conventional rabbit, analyzed the concrete reason created this present situation, and proposed some concrete management measures to guarantee animal experimental quality.

An Exploration on Reconstruction of Animal Laboratory

HAO Zhi-Hui, LIANG Jian-Kun, KONG Li-Juan

2011, 28(05): 41-42.

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Think of Laboratory Animal Status and Development for Qinghai Province

FAN Wei

2011, 28(05): 43-47.

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Acute-Toxicity Test of WHPI on KM Mice

XU Di, YE Ming-Xia, XIONG Mei-Yun, KONG Li-Jia

2011, 28(05): 66.

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A Novel Surgical Model Established-Collects Cerebrospinal Fluid in Beagle

ZHANG Sheng, XIN Yan-Fei, GU Li-Qiang, XUAN Yao-Xian

2011, 28(05): 69.

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Objective To establish a novel surgical model of multi-collecting cerebrospinal fluid （CSF） in Beagle dog and assess the surgical effect. Method With investigating the construction of skull of beagle dog, deciding the position of surgical point, completed the collecting process by skull puncture, collection tube covering and cerebrospinal fluid multi-drawing. Result We successfully explore a novel surgery of collecting cerebrospinal fluid, with the successful rate up to 70%. Conclusion The new model to cerebrospinal collect is favor of multi-draw CSF. It is a potential model to be popular method in colleting CSF.

A Modification of Rabbit Cag

JI Lian, MA Tie, SUN Jian

2011, 28(05): 72.

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objective: To design plastic film method, compared plastic film method with other rabbit breeding methods of the pros and cons. Method: After proving the flush type method, using the plastic film method, flush type method, dry raise type method feeding rabbit, determination of different methods rabbit feed room of humidity, and compares three cages work style characteristics. Results: 1. compared the plastic film method to the flush type method, it could reduce the humidity18% 。 2. No difference among the three methods in regard to feeding rabbit. 3. Using the flush type method the workload can reduced by92%, in comparison with the dry raise method. Conclusion: plastic film method has its unique advantages; it can reduce the humidity of the feeding room of the flush type method, and also lighter the workload of the plastic film method. The plastic film method easily elaborate the advantage of the method of both, but also complementary the defect of the both.

Anesthesia Salvage of Beagle Dog and Cynomolgus Monkey

HUANG Zhen-Xing, LI Hong-Xia

2011, 28(05): 75.

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During the procedure of drug safety evaluation, many manipulations need animal anesthesia. But anesthesia is a complicated process which accompanied with risk. Anesthetic accident happened sometimes. Two accidents were met in 2010, one was beagle dog which was salvaged continuously for 8 hours, and another was cynomolgus monkey which was salvaged for 2 days. Finally, two animals were recovered.

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