30 June 2010, Volume 30 Issue 6

    

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  Dynamic Observation of In vivo Bioluminescence Imaging of Orthotopic Implantation of Human Hepatocellular Carcinoma in Nude Mice

  YAN Ming-xia,LIU Lei,ZHU Miao-xin,ZHAO Fang-yu,SA Bing-qing,LIANG Lin-hui,YAO Ming

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 401-405.

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  Objective To establish a human hepatocellular carcinoma cell line with stable and high-level GFP/Luciferase(Luc) expression, and observe the development of tumors in deep organisms of the nude mice by in vivo biofluorescence imaging. Methods The SMMC-7721 -GFP/Luc cell line were established by lentivirus transfection，then were orthotopic implanted in the liver of the BALB/c nude micc, the behaviors of the tumors was detected by in vivo bioluminescence imaging. Results The cell line SMMC-7721-GFP/Luc with stable and high-level GFP/Luc expression had been successfully established. After the cell line had been inoculated in the liver of the nude mice, the tiny tumor masses in early time and the growth of tumors in late time in the deep organisms were observed by in vivo biofluorescence imaging. Conclusion The use of cell line SMMC-7721 -GFP/Luc could directly observe the tumor growth and metastasis in the orthotopic implantation animal model. It would help us to understand the biological behavior of human hepatocellular carcinoma and its characteristics. In view of its sensibility, intuitiveness, dynamics and reliability, it would do some good to further study of human hepatocellular carcinoma.
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  Establishing and E thology Evaluation of Anger-out and Anger-in Inducing Premenstrusl Syndrome Macaque Model

  ZHANG Hui-yun,WEI Sheng,QIAO Ming-qi

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 406-409.

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  Objective Establishing anger-out and anger-in inducing emotional diseases macaque model to provide suitable research object for brain center mechanism of anger-out and anger-in inducing emotional diseases. Methods Using female macaque and inducing anger emotion occurence, then applying “Emotional evaluation scale of female experiment macaque”to score different emotion type of macaque model. We will differentiate anger-out and anger-in types from all anger monkeys based on scale scores and finish evaluating different anger types. Using anger-out and anger-in monkeys individu-ally add to pressurization method to prepare anger-in or anger-out inducing emotional diseases monkey model. Results After modeling stress，anger factors of PMS liver-qi invasion model monkeys increased significantly than control team(P＜0.05), while PMS liver-qi depression model monkeys had a notable change compared with control team(P＜0.05). While there is no difference compared with control team (P＞0.05) after administration. Conclusion It could be inferred that target disease could be induced successfully using social surbodination stress according to results of 30-minute group ad libitum obser-vation and scale scores, subjective combined objective results were regarded as a good measure methods to evaluate target emotional disease occurence on macaque. Idea and method mentioned above provide a suitable research object for occurrence pathogenesy of anger-out and anger-in emotion inducing emotional disorder.
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  Premature Aging in Zebrafish Heterozygous breakdance Mutant

  CHEN Zhen-yue1,2,XU Xiao-lei2,WU Li-qun1,LU Guo-ping1

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 410-414.

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  Objcctivc The breakdance-mutation (bretb 218) has been described as a mutation in the zebrafish orthologe of the human ether-a-go-go-related gene (HERG,kcnh2) with an arrhythmic heart beat in the homozygous. Interestingly, we found the heterozygous (hets) could generate multiple premature aging-like features and distinct electrophysiology. It made possible to develop a vertebrate model for study on premature aging associated to kcnh2. Methods Life span and body shape change were followed up and recorded. Exercise endurance capacity was examined by swim test. ECG test was performed in wild type (WT) and the hets at different adult stages. Results Survival rate dramatically descended and curly body showed from 20 months in the hets. ECG test showed heart rate dramatically decreased along aging process and QTc duration prolonged in the hets. Exercise endurance capacity significantly reduced 56.44% in the hets at 32 months comparing to WT at the same stage.Conclusion The heterozygous breib 218 has shorter life span and showed the presenility on appearance， electrophysiology and the exercise endurance capacity which may associate to the mutation in one allele of zebrafish orthologue of kcnh2.
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  Comparison of Three Detection Methods of Clostridum Piliforme Infections in Laboratory Rats

  DING Cong1,FENG Jie2,XIE Jian-yun2,CHEN Lu3,GAO Cheng2,HU Jian-hua2

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 415-419.

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  Objective To investigate the infection states of Clostridium piliforme in laboratory rats and evaluate the usefulness of indirect fluorescent antibody assay(IFA), nested-polymerase chain reac-tion (nested-PCR), ELISA. Methods Specimens were collected from the rat colonies and detected for C.Piliforme by using IFA, ELISA and nested-PCR methods at the same time. The results of three detection methods were compared and analysis. Results A total 125 Rats，36 out of 125 serum samples (28.8%) are showed to be positive by IFA; 24 out of 125 serum samples(19.2%) are showed to be positive by ELISA;6 out of 125 liver and fecal samples (4.8%) was positive by PCR. The positive rate of detected by IFA and ELISA were higher than PCR. Conclusions IFA method to combination with ELISA method can improve the sensitivity and reliability of the detection results. For the positive samples could be differentiated by PCR method. By contrast, IFA is a quick ,reliable and cheap method. IFA is available for the epidemiological investigation of C.Piliforme.
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  Studies on the Determination of Islets β Cell in Scalded Rats with LPS-injccted and the Effects of GIK Treatment

  WANG Zhan-ke1,2,CHANG Jin1,WANG Shi-liang3,XU Lin-shui3,LIU Long-yan2,HU Yang-tao2,YU Wen2

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 420-423.

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  Objective To study the dysfunction of islets β cell in scalded rats with LPS-injected and. the effects of GIK(glucose-insulin- Potassium) treatment to the islets P cell. Methods Ninety SD scalded rats with LPS-injected were divided randomly into the S+LPS(n=30)，GIK (n=30) and control groups(n=30). The glucose and insulin of the fasting and 30min post of glucose injected of the all groups were determined, and the HOMA-β,△ INS30/△ GLU30 and HOMA-IR were calculated and observed. Results The glucose and insulin of the fasting and 30min post of glucose injected and the HOMA-IR of the S+LPS group were higher than that of the before scalded(P＜0.01), but the HOMA-β and △ INS30/△ GLU30 of the S+LPS group were lower than that of the before scalded(P＜0.01). The HOMA-β and △ INS30/△ GLU30 of the GIK group were higher than that of the control group, there was no difference of HOMA-TR significantly between the GIK and control group. Conclusion The serious burn rats represents dysfunction of early phase and basal insulin release of p cell, GIK treatment would be beneficial for the protect of β cell on this states.
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  The Changes of Cardiac Structure and Function Induced by Hypokalemia with Echocardiography

  LU Min,XU Rong,WANG Yin-chun

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 424-427.

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  Objective To explore the correlation between serum potassium levels and the changes of the heart, the real-time echocardiography was applied to monitor the effects of hypokalemia on the heart structure and function of the experimental rabbits, and for the purpose of early diagnosis and treatment in the clinical practice. Methods The hypokalemia experimental model was established by intramuscu-lar furosemide injection. M-mode echocardiography was used to monitor the cardiac morphology，the sequential variation of flow spectrum and cardiac function. After that, the heart was removed and the tissue was made into pathological section and dyed with HE. Results The cardiac morphology and function of the rabbits showed little change during the early and middle stage. Then, the heart dilated rapidly, and its function decreased until cardiac arrest. The myocardial pathological section showed swell and vacuolization of the myocardial cells. Conclusion There were a series of changes in the heart of rabbit during the decreasing process of serum potassium concentration, and echocardiography is a noninvasive method to monitor these changes, it can observe the heart morphology and evaluate the cardiac function accurately in real time. Thus, echocardiography provides a convenient and reliable method for monitoring the heart complications induced by hypokalemia.
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  Establishment and Biological Characteristics of Orthotopic Transplantion Tumor Model of Hepatocellar Carcinoma Cell line in Nude Rat

  SHEN Yan1,PENG Xiu-hua1,XU Chun-hua1,ZHOU Wen-jiang1,2

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 428-431.

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  Objective To study the growth and metastases of the tumors after orthotopic transplan-tation with hepatocellar carcinoma cell line (Huh-7) in nude rat. Methods Tissue from the tumor inoculated subcutaneously in nude mice with Huh-7 cell line was transferred to the nude rat beneath the skin. Subcutaneous and orthotopic implant models simulating hepatocellar carcinoma was established by suturing histologically intact human tumor tissue on the liver of nude rat. Subsequently,the rate of the orthotopic tumor and their biological properties of tumor growth and metastasis were observed.Tuinorgenecity, invasion，metastasis and morphological characteristics of the implanted tumors were studied by light microscopy, electron microscopy and flow cytometry. Results In orthotopic trans-plant model of nude rat，tumor formation rate was 90.0% and metastasis rate of liver was 50.0%.Conclusions The nude rat is more suitable for orthotopic transplant models for hepatocellar carcinoma，and should be valuable for research on mechanism and new therapies for metastasis.
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  Pathological Observation on Macaca speciose thibetanas with Secondary Infection Caused by Diarrhea

  ZHONG Zhen-dong1,XU Yao-zhong1,YAO Fang1,WEI Jian-jun2,ZHONG Hao1,ZHOU Liang1,HE Yong-liang1,LI Jun-hui1

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 432-435.

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  Objective To probe etiopathogenesis of secondary infection caused by diarrhea in one Macaca speciose thibetanas. Methods The pathological observation on an 8 year-old Macaca speciose thibetanas with historically recurring diarrhea were carried out. Results Macaca speciose thibetanas was in the the dehydrant condition as showed in the clinical symptom. Also serious pathologyical change was observed in the lung and intestine. Histopathologically, granular degeneration and vacuolar degenera-tion were observed in the liver and kidneys. Congestion and hyperemia were observed in the blood vessel of many tissues and organization, and the inervallum of vessel wall was swollen. Pancreas，bladder and brain were normal. Conclusion The characters of diarrhea in Macaca speciose thibetanas are complex etiological factors and seasonal occurrence. The treatments on diarrhea of such animal are suggested to mainly depend on TCM and modern treatments to be adjuvant.
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  Expression of GRP78 mRNA in Obese Tissue of Rats Fed with High Fat Diet

  LIU Jia,DU Jian,PEI Li-na,LUO Hong-fei,GUO Ning-ning,LIN Zhi-nan,Zhang Ke-ying

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 436-439.

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  Objective To observe the expression of GRP78 mRNA in visceral obese of rats fed with high fat diet in order to explore its effects on insulin resistance. Methods Thirty Wistar rats were randomly divided into two groups: normal chow group (NC) was fed with free normal chow for 10 weeks; high fat group (HF) was fed with high fat diet for 10 weeks. The expression of GRP78 mRNA was detected by real time PCR. The total cholesterol triglyceride，the rate of visceral fat (epididymal fat) vs. body weight and fasting insulin were also detected.ResuIts (1) Glucose infusion rate (GIR) in HF group was obviously lower than that in NC group (P＜0.01). Rats in HF group had an elevation of fasting insulin compared with NC group (P＜0.05). (2) The expression of GRP78 mRNA was significantly higher in HF group than that in NC group (P＜0.01). (3) Rats in HF group had an obviously elevation of fasting insulin ,total cholesterol and triglyceride compared with NC group Conclusion After 10 weeks fed with high fat diet, the insulin resistant animal models was established. High fat diet can induce the expression of GRP78 mRNA in visceral obese tissue.
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  The Application of Toluidine Blue in Histological Staining

  DAI Xiao-wei,HUANG Lan,XU Yan-feng,MA Chun-mei

  Laboratory Animal and Comparative Medicine. 2010, 30(6): 440-444.

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  Objective To expand the application of toluidine blue staining. Methods Skin, lung,trachea, cerebrum and spinal cord tissues were collected and fixed by 4% formalin. After routine slicing,the slices were stained by toluidine blue. Lastly the feature of all kinds of tissues under microscope was observed. Results Besides nissl body was clearly showed，the langerhan's cell, melanocytes and merkel cell in skin can also be differentiated. The type I alveolarenhanced by toluidine blue staining.The layers of cerebral cortex were displayed much more clearly than HE stai cell，type II alveolar cell and macrophage in lung and the ciliated cell and goblet cell in trachea were markedly ning. Conclusion Toluidine blue staining may be explored in skin, lung, trachea, cerebrum and spinal cord tissues.