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    25 February 2014, Volume 34 Issue 1
    Establishment and Preliminary Phenotypic Analysis of Pten Knockout Mouse Model in Spermatogonial Stem Cells
    ZHOU Wei, ZHANG Di, YU Zhuo, ZHA Wei-wei, FENG Li-xin
    2014, 34(1):  1-5.  DOI: 10.3969/j.issn.1674-5817.2014.01.001
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    Objective To establish Pten conditional knockout (KO) mouse model and investigate the role and related mechanism of Pten in spermatogonial stem cells(SSCs) and preliminary animal phenotype. Methods Cre/Loxp system was adopted to establish the Pten conditional knockout(KO) mouse model in spermatogonial stem cells. The wild type littermates were used as control and animal phenotype was observed by dissection and Hematoxylin and Eosin (HE) Staining. Then the expression of Plzf, a kind of SSCs marker, was detected by immunofluorescence stain (IF) and the positive cells in IF were counted. Results Pten KO mice shown postnatal developmental retardation and some died around 13 days. No difference in the size of testes was found comparing to the wild type mice, but the result of cell counting indicated that less Plzf positive cells were detected in KO mice (P<0.01). Conclusion Pten conditional knockout mouse model using spermatogonia stem cells is successfully established and Pten might affect SSCs development by regulating the expression of Plzf. This study, to some extent, provides some scientific evidence and new thoughtfor further exploring the complicated regulatory network in SSCs.
    Improvement and Comparison of Two Methods for Establishing Hemi-sected Spinal Cord Injury Model of Rats
    ZHAO Fu-sheng, ZHANG Xiao-li, WU Geng, CHEN Bin, LI Yue-zhen, JIN Xiu-dong, ZHANG Ji-fei
    2014, 34(1):  6-11.  DOI: 10.3969/j.issn.1674-5817.2014.01.002
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    Objective Comparing the characteristics and application of two methods for establishing the hemi-sected spinal cord injury model of rats and provide suitable spinal cord injury models for clinical research. Methods Sixty Sprague-Dawley rats were randomly divided into transverse-hemisected group (A group, n=20), wedge-shaped-block-hemisected group (B group, n=20) and normal control group (C group, n=20). After operation, the stability of the two hemi-sected models were comprehensive assessed by Basso-Beattie-Bresnahan (BBB) scores, inclined plane method, HE staining and MRI detection. Results After operation, the BBB scores and maximum angle of inclined Plane method of rats in group A and group B were significantly lower than group C (P<0.05), but these indexes had no significant difference between group A and group B; In addition, HE staining and MRI detection show a significant morphological change between group A and group B 28 days after operation. Conclusion Both methods of transverse-hemisected and wedge-shaped block-hemisected can establish a stable and reliable spinal cord injury model, and wedge-shaped-block-hemisected model is more suitable to assess graft treatment of spinal cord injury.
    Protective Effect of Neosperidin dihydrochalcone on Influenza Virus-induced Air-blood Barrier Injury in Mice
    MAO Dong-sen, CHEN Qin, YU Chen-huan, HU Yi-xiang, ZHANG Cai-qiao
    2014, 34(1):  12-17.  DOI: 10.3969/j.issn.1674-5817.2014.01.003
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    Objective To explore protective effects of neosperidin dihydrochalcone (NHDC) on air-blood barrier injury in influenza virus infected mouse model. Methods The pneumonia model was established by infection of influenza virus. Model mice were treated with NHDC at different dose by gavage. The pulmonary index, wet/dry mass (W/D) ratio and the pathological changes of lung tissue stained with HE were investigated. The levels of serum IL-6, LTB4 and ICAM-1 were determined by ELISA, while expression of aquaporin 5 (AQP5) was detected by immunohistochemistry. Results After challenge, the infected mice had obvious increase in pulmonary index and W/D ratio, which manifested by accumlation of significant numbers of neutrophils, accompanied by marked pulmonary edema and high levels of serum IL-6, LTB4 and ICAM-1. The expression of AQP5 was also significantly increased in the lung of infected mice. However, oral administration of NHDC significantly reversed those adverse factors. Conclusion Influenza virus could cause air-blood barrier damage in the lung of infected mice, and it may be related to the expression of AQP5. NHDC shows obvious effect to the injury.
    Studies on Chronic Toxicity of Hypericum Perforatum Extract on Beagle Dogs
    XIE Jia-jun, ZHOU Guang-xing, QIAO Zheng-dong, WANG Jian-xin
    2014, 34(1):  18-24.  DOI: 10.3969/j.issn.1674-5817.2014.01.004
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    Objective To investigate the chronic toxicity of Hypericum perforatum extract through oral administration for 9 months on Beagle dogs. Methods Twenty-eight Beagle dogs were randomly divided into blank control group and low, middle and high dose groups,which were treated respectively with Hypericum perforatum extract (0.3, 0.55, 1.0 g/kg) by oral administration, once a day, 6 times weekly for 38 weeks. During the whole experimental period, the Beagle dogs were treated with Hypericum perforatum extract through deglutiatur administration once a day for successive 38 weeks, meanwhile the activities of the dogs were observed once a day, the body weight were weighed weekly. Electrocardiographic examination, blood counts determination, serum biochemical measurement, urinalysis test and histomorphologic observation were conducted on the 10th, 20th, 30th, 38th week with successive administration and on the 46th week without administration for 8 weeks. Some animal in each group were sacrificed, whose organs and tissues were harvested, of which the main ones were weighted and examined histopathologically on the 20th, the 38th week with administration and the 46th week without administration for 8 weeks. Results During the whole experimental period, the animal activities, electrocardiographic examination, blood counts determination and urinalysis test showed no obvious abnormalities. But ALP increased in blood of the Beagle dogs in the 1.0 g/kg groups on the 20th and the 30th week with administration, TBIL increased in the 0.55 g/kg and 1.0 g/kg groups on the 38th week (P<0.05 or P<0.01). Meanwhile, liver coefficient increased in the 1.0 g/kg groups (P<0.05) on the 20 th, 38th week after administration. Pathological examination showed that slightly granular degeneration appeared in the liver cells of 2 animals (1♀,1♂) and in the renal tubules of 1 animal (♀) in the 1.0 g/kg groups on the 20th and 38th weeks, regardless of sex. There was no significant difference between the rest of the treatment groups and the corresponding control group in hematological and serum biochemical indexes, the organ coefficient, the morphological changes (P>0.05). Conclusion Some mild toxic effects on the liver and kidney function or organization were found on the 20th and 38th week when the Beagle dogs were treated with Hypericum perforatum extract (0.55, 1.0 g/kg) through successive oral administration once a day for 38 weeks , but could disappear gradually over time after withdrawn. The degree of toxicity was positively correlated with the dose of administration and the medication cycle. The safe dosage of Hypericum perforatum extract by intragastric administration is 0.3 g/kg, equivalent to 40 times of clinical depression treatment dose.
    Comparison on Immunotoxicity Difference of Breviscapine Injection between Rat and Guinea Pig
    LUO Yong-wei, YANG Chun-lei, WANG Zhong-hui, WANG Yong, WANG Rong, JIANG Juan, YAN Han, SUN Zu-yue
    2014, 34(1):  25-28.  DOI: 10.3969/j.issn.1674-5817.2014.01.005
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    Objective To study the immunotoxicity difference of Breviscapine Injection between rat and guinea pig. Methods According to the Chinese traditional medicine, natural medicine immunity toxicity study technical directed principle, the PCA and ASA tests were carried out in order to compare the immunotoxicity difference between the two animals. Results The results of the allergy testing showed that no positive symptom in SD rat occurred at all the doses of 5 mg/kg、50 mg/kg and 500 mg/kg of Breviscapine injection group in PCA toxicity study. While in ASA toxicity study, 50 percent animals appeared weak positive and 50 percent animals appeared positive at the dose of 5 mg/kg of Breviscapine injection group; and all appeared the positive symptoms at the high dose of 50 mg/kg group. Conclusions Compared with the allergic response in animal study, there are significant difference between the PCA and the ASA toxicity study and the ASA is more sensitive and more conformity to the application in clinical.
    Analysis of the Polymorphism of Mitochondrial DNA Control Region in Three Closed Colonies of Rabbits
    SHEN Xing-jiao, YUE Bing-fei, MA Li-ying
    2014, 34(1):  29-34.  DOI: 10.3969/j.issn.1674-5817.2014.01.006
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    Objective To discuss the genetic diversity and uniformity in three groups of closed colony rabbits based on mitochondrial DNA control region. Methods Seventy-one blood samples from New Zealand rabbit, Japanese White rabbit and Chinchilla rabbit were collected for amplifying ,purifying and sequencing the part of sequences of mitochondrial DNA control region by PCR and Genetic Analyzer. The variation sites, Parsimony informative sites, haplotypes, haplotype diversity, nucleotide diversity were analyzed by Mega 4.0 and DNA SP. Results 6 diversity sites and 9 haplotypes were found in the seventy-one rabbits, and most mutations were C→G. Every population had its own haplotypes. The polymorphism of Mitochondrial DNA Control Region and population diversity in three group were both low. Conclusions More sequence information about mtDNA sequences and population structure of laboratory rabbits were supplied,which was helpful to genetic breeding.
    Development and Preliminary Application of RT-PCR Method for Detection of Mouse Adenovirus
    WANG Ji, FU Rui, WEI Li, LI Xiao-bo, FENG Yu-fang, WANG Shu-jing, GONG Wei, YUE Bing-fei, HE Zheng-ming
    2014, 34(1):  35-41.  DOI: 10.3969/j.issn.1674-5817.2014.01.007
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    Objective To develop a RT-PCR method for detection of Mouse Adenovirus (MAdV) in Mongolian gerbils and Laboratory mice. Methods The primers were designed and synthesized according to the published MAdv specific sequences of E1B gene . PCR method is established, and the specificity, sensitivity, stability test were carried out. The method is used to detect 65 Mongolian gerbils, and 12 mice. Results The developed PCR method was no cross reaction with Minute virus, Polyoma virus, its minimum detection limit ,using the recombinant plasmid containing MAdV gene as a template, was 1.67 pg/μl, and the lowest detection virus titer is 10-7/ml. The MAdV cDNA maintained at -30℃ for 12 months, still can enlarge the size of about 606 bp visible mesh belt. The 65 Mongolian gerbils and 12 mice were all negative about MAdv detected by PCR. Conclusion The developed PCR method is good in specificity, sensitivity, stability,and may be used for detecting the MAdv in laboratory animal, such as Mongolian gerbils and mice.
    Expression of nm23-M1 Gene in Mouse Myelomonocytic Leukemia
    DONG Zheng, SUN Xue-dong, MAN Qiu-hong, YAO Bo, LI Yu-fang, LIU Tie-qiang, HUANG Shan, LIU Gguang-xian, Ai Hui-sheng, GUO Mei
    2014, 34(1):  42-45.  DOI: 10.3969/j.issn.1674-5817.2014.01.008
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    Objective To investigate the expression level of nm23-M1 gene in mouse myelomonocytic leukemia. Methods The mice were inoculated intravenously with myelomonocytic leukemia cells of WEHI-3 cells, and selected a control group of normal mice. Bone marrow samples were collected in two groups. The nm23-M1 mRNA expression were detected by TaqMan quantitative real-time reverse transcription polymerase chain reaction(qRT-PCR) and analysis of its relationship with the occurrence and development of leukemia. Results The expression of nm23-M1 gene in bone marrow cells of mice with leukemia is 24 times of the normal control group. There was a significant difference between the two groups (P<0.05). Conclusion The nm23-M1 gene overexpressed .in mouse myelomonocytic leukemia, and had close relationship between and peripheral white blood cell count, the number of original cells in bone marrow and organ against the leukemia cases, regardless of the gender, age and leukemia in mice.
    Analysis on Femur Bone Parameters in Female Mice by Using Micro-CT
    GENG Qin, ZHU Miao-xin, SUN Lei, CUI Yong-qi, KONG Han-wei, YAO Ming, LIU Lei
    2014, 34(1):  46-49.  DOI: 10.3969/j.issn.1674-5817.2014.01.009
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    Objective To obtain femur bone datas about female BALB/c-nu/nu and BALB/c mice by using micro-CT. Methods The femur from fifteen female nude mice with BALB/c background and BALB/c mice respectively, aged 10-12weeks, were scanned with micro-CT. Results The parameters of female BALB/c-nu/nu and BALB/c mice femur bone were detected by using micro-CT, there are differences of femur bone in some parameters between BALB/c-nu/nu and BALB/c mice. In the vortical bone, there is significant difference in BV/TV. In the trabecular bone, BV/TV, BS/TV and BMD have significant difference. Conclusions Micro-CT can be used for quantitative analysis of mice femur bone parameters.
    Establishment and Evaluation of Chronic Allergic Asthma Models in Rat
    QIAO ming, LIU Lan-ying, WANG He-sheng
    2014, 34(1):  79-82.  DOI: 10.3969/j.issn.1674-5817.2014.01.019
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    To establishment the chronic allergic asthma model, which was similar to chronic human asthma, this paper summarized the rat models of chronic allergic asthma established in recent years, from several aspects such as rat strains, methods, and evaluation of model.