Table of Content

25 June 2016, Volume 36 Issue 3

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Expression and Role of KIP1 Ubiquitylation-promoting Complex 1 in Neointimal after Carotid Arteries Balloon Injury in Rats

YANG Bo, LIN Xin-duo, WANG Xing-ren, TANG Jun-ming, YANG Jian-ye, ZHANG Lei, CAO Teng, JIANG Feng-bo, WANG Jia-ning

2016, 36(3):  161-167.  DOI: 10.3969/j.issn.1674-5817.2016.03.001

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Objective To study the expression of Kip1 ubiquitylation-promoting complex 1(KPC1) protein after carotid arteries balloon injury in SD rats and to explore its biological function in neointimal formation process. Methods The adult male SD rats (450-500 g) were randomly divided into sham group and carotid artery injury model group. The left damaged carotid arteries were considered as the experimental group after modelling, with the left normal carotid arteries of sham group being as control group. After 1, 3, 7 ,14 and 28 days of balloon injury, SD rats of the experimental group were sacrificed and harvested for histomorphology and molecular markers assay. Hematoxylin-eosin (HE)staining was used to assess the changes of neointimal formation in the carotid artery while Western blot and qRT-PCR were used to evaluate the expression of KPC1. What’s more, Western blot was also used to assess the levels of proliferating cell nuclear antigen (PCNA). The proliferation model was established by culturing vascular smooth muscle cells (VSMCs) in vitro, then detected the KPC1 protein, KPC1 mRNA expression by Western blot and qRT-PCR. Results Western blot and qRT-PCR showed that the KPC1 expression levels of the experimental group were significantly lower than that of control group (P<0.05), while the PCNA expression was significantly higher than that of control group (P<0.05). Compared to 1 day and 3 days after carotid arteries injury, for VSMC hyperplasia, neointimal generation and luminal stenosis were observed at 7 days by HE staining. With VSMCs hyperplasia, neointimal and luminal stenosis were getting more and more serious, the thickness of intimal exceeded medial at 14 days. At 28 days, the irregular intimal hyperplasia was continued and the degree of vascular stenosis was more than 50%. After stimulation of VSMCs proliferation by platelet-derived growth factor BB (PDGF-BB) in vitro, then Western blot and qRT-PCR data showed a gradual decreasing trend of KPC1 at protein and mRNA levels. Conclusion Inhibiting KPC1 expression can significantly decrease intima hyperplasia in this model, and its mechanism may be related to the promotion of VSMCs proliferation.



Impacts of Chronic Restraint Stress on Cognitive Function and Astrocytes in Different Subregions of Hippocampus in Mice

WANG Yan-yong, ZHANG Zhong-xia, SUN Mei-yu, WANG Ming-wei

2016, 36(3):  168-173.  DOI: 10.3969/j.issn.1674-5817.2016.03.002

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Objective To explore the impact of chronic restraint stress (CRS) on the cognitive function of mice, and observe the activation of astrocyte (AS) in different subregions of hippocampus (CA1, CA3 and DG), thus to make clear whether the damage of chronic stress in hippocampus has specific targets. Methods According to the body mass, 24 male Kunming mice were randomly divided into control group and stress group, with 12 mice in each group. A chronic stress model of mouse was established by using chronic restraint method. The cognitive function was evaluated by novel object recognition test (NORT) and Morris water maze (MWM), cell morphology changes were observed by HE staining, while the glial fibrillary acidic protein (GFAP) as marker of AS in hippocampal CA1, CA3 and DG areas were observed by immunohistochemistry staining. The AS number and GFAP expression in subregions of hippocampus were counted and analyzed by using the microscope and image analysis software respectively. Results Compared with the control group, the NORT and MWM scores of stress group were both decreased significantly (P<0.05), AS numbers and GFAP expression in CA1, CA3 areas were both significantly increased (P<0.05), and in DG area there was no significant changes (P>0.05). Conclusion CRS could lead to cognitive dysfunction in mice, and the activation of AS in hippocampus CA1, CA3 regions may be one of the mechanisms.



Dynamic Observation on In vivo Biofluorescent Imaging of Orthotopic Transplant Model of Human Colon Cancer in Nude Mice

ZHAO Yong-jiang, ZHU Miao-xin, YUAN Li-xin, SUN Lei, GENG Qin, LI Jing, YAO Ming, YAN Ming-xia

2016, 36(3):  174-179.  DOI: 10.3969/j.issn.1674-5817.2016.03.003

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Objective To establish an orthotopic transplant nude mice model of human colon cancer and discuss the application of in vivo biofluorescent imaging system in animal model. Methods Human colon cancer cells were tranfected with green fluorescent protein/Luciferase gene by lentiviral vector, they were orthotopically transplanted into the cecum of nude mice by using tumor tissues or cell culture method. The tumor growth and metastasis were dynamically observed by using in vivo biofluorescent imaging system. Results The human colon cancer cell line and orthotopic transplant nude mice model were successfully established, which could display stable high-level GFP/Luc expression in vitro and in vivo. In vivo biofluorescent images showed that the fluorescent areas and intensities were increased gradually with the increase of tumor volume in nude mice during 7 weeks of dynamic observation. Conclusions In vivo biofluorescent imaging system was fitted for evaluation and quantitation of the tumor growth and metastasis of colon cancer. It would provide effective reference for the observation and study of in vivo deep tumors.



Non-invasive Assessment of Fulminant Liver Injury with Contrast-Enhanced Micro CT in Mice Models

LI Da-wei, HUA Xiang-wei, ZHANG Jiang, YAO Ju-fang, DAI Hui-li, Kong Xian-ming

2016, 36(3):  180-185.  DOI: 10.3969/j.issn.1674-5817.2016.03.004

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Objective To explore the potential use and mechanism of MicroCT using ExiTron nano6000 in the assessment of fulminant liver injury in mouse models. Methods Five male C57BL/6 mice were subjected to administration of LPS/D-GalN to develop model of fulminant liver injury. The mice were scanned with MicroCT before the treatment and at 3 h and 6 h after the establishment of liver injury model. The average CT value of liver were analyzed with software for future study. Twenty male mice were divided into 4 groups, 5 mice in each group: Nano 6000 group, the mice were treated with Nano 6000; PBS group, treated with PBS as control; model 3 h group and 6 h group, injected with LPS/D-GalN and sacrificed at 3 h, 6 h after the injection. Pathological damage of the liver were observed by microscope and serum ALT and AST were detected. Results MicroCT with ExiTron nano6000 as contrast agent could provided specific and clear 2 dimensions and 3 dimensions structure of liver and spleen in mice. The average CT value of liver was significantly increased at 3 h after LPS/D-GalN treatment and drop to baseline level at 6 h. In vitro experiments showed that LPS stimulation could enhance the endocytotic ability of macrophages to uptake ExiTron nano6000. Conclusions The Micro CT provided a rapid noninvasive longitudinal monitoring of fulminant liver injury in mice models and may be had good application in future research.



Establishment of Stress Induced Cardiomyopathy Model by Stimulation of Vagus Nerve with Electricity Signal in Rabbit

GUO Rui-wei, YANG Li-xia, XI Yin-yan, KUANG Chen-wei

2016, 36(3):  186-189.  DOI: 10.3969/j.issn.1674-5817.2016.03.005

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Objective To establish the stress induced cardiomyopathy (SIC) model in rabbit via the stimulation of vagus nerve by using electricity signal. Methods Ten femaleNew Zealand White rabbits at the age of 2 months were randomly divided into control group and experimental group. The vagus nerve of the experimental group rabbits were exposured and stimulated intermittently by using electrical signal. The rabbit model were evaluated by electrocardiogram (ECG), necropsy and pathological observation of myocardial tissue. Results The ECG at 1 day after operation showed that heart rate of rabbit in experimental group was faster than that of the control group. Meanwhile, apical myocardial in the experimental group became significant thinner than that in control group at one month after operation(0.83±0.12 mm vs 1.55±0.11 mm, P<0.05). In addition, myocardial tissue of rabbits in experimental group were showed typical changes of stress induced cardiomyopathy. Conclusion The SIC rabbit model can be successfully established via stimulating the vagus nerve by using electricity signal.



Establishing Facial Paralysis Model in Rats by Clipped Facial Nerve

LI Yao, GAO Xiao-li, CHEN Jian-kang, WANG Si-wang, XIE Yan-hua

2016, 36(3):  190-194.  DOI: 10.3969/j.issn.1674-5817.2016.03.006

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Objective To establish a high morbidity and low mortality of animal model for facial paralysis in rats, which is easy to operate. Methods Fifty SD rats with half male and female were randomly divided into five groups, each group with 10. The facial nerve trunk of rats were clipped with mosquito forceps closing three tooth for 6 min, 8 min, 10 min, 12 min and 14 min. The compound muscle action potential (CMAP) amplitude of facial nerve trunk was measured by biological function experiment system, and the general behavior indices of all the rats were observed next day after injury. Results The CMAP amplitude of rats after facial nerve trunk clipped for 6 min and 8 min respectively were the same as none clipped rats, but it decreased obviously and tended to zero after clipped for 10 min, 12 min and 14 min. In addition, ethology score results showed that the models of 6 min, 8 min rats performed incomplete paralysis symptoms, the models of 10 min,12 min,14 min rats were facial paralysis completely and the models of 14 min and 12 min rats had no obvious difference compared with the model of 10 min. Conclusion The decreased CMAP amplitude of facial nerve trunk and ethology score results showed that clipping the facial nerve trunk with the mosquito forceps closing three tooth for 10 min was the best way to establish rats model of facial paralysis.



Intragastric Estrogen Administration Induced Guinea Pig Model of Uterine Leiomyoma

ZHANG Yue-han, WEN Jing, ZHU Hai-tao, ZHAO Guang-zong, LI Xiao-yu

2016, 36(3):  195-199.  DOI: 10.3969/j.issn.1674-5817.2016.03.007

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Objective To establish the guinea pig model of uterine leiomyoma by intragastric administration of estrogen and to discuss the effect of different cycles of administration on formation of uterine leiomyoma. Method Mature female guinea pigs were divided into short term group and long term group, and intragastric administrated with estradiol valerate at 0.1 mg/100 g body weight twice a week for 6 weeks and 8 weeks respectively. The control was intragastric administrated with normal saline with same volume. After 6 or 8 weeks, the guinea pigs were sacrificed. The serum levels of estrogen and progesterone were measured by ELISA and the leiomyoma forming rate was measured. Histological changes were compared between treatment group and control group with HE staining. Results The leiomyoma forming rate in short term group was 30% and long term group was 40%. The average total rate was 35%. The serum level of estrogen in model animals was increased significantly and the progesterone decreased in long term group. Histopathological examination confirmed that the guinea pig model of uterine leiomyoma were established successfully. Conclusions Intragastric estrogen administration was a simple and effective method for establishing uterine leiomyoma guinea pigs model.



Comparative Study on Mucosal Thickness of Maxillary Sinus and Frontal Sinus in Goats

LI Yan-feng, HAN Yi-shi, REN Huan, LU Yun-long, DONG Xin-ming, LIU Le, HU Pin, WANG Ning

2016, 36(3):  200-203.  DOI: 10.3969/j.issn.1674-5817.2016.03.008

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Objective To explore the differences of mucosal thickness in goat paranasal sinus, and analyse the underlying reasons. The discussion can provide theoretical basis and guidance for issues related to mucosa in the practice of maxillary sinus floor augmentation. Methods The mucosa was cut into desired size from every sinus mucosa and fixed in formalin to get the tissue specimens, and observed under optical microscope with scaled eyepiece. Randomly selected three fields under the microscope from the samples and then measured and recorded. Paired t-test was used to determine the difference between the parameters, P<0.05 was regarded as statistically significant. Results Goat maxillary sinus mucosal thickness of the top was 410.03±65.97 μm, the bottom was 461.33±91.37 μm, the frontal sinus mucosa was 216.90±46.47 μm. The mean differences of mucosal thickness between the top of the maxillary sinus and the frontal sinus, the bottom and the frontal, the top and the bottom were statistically significant (P<0.05). Conclusions The top and bottom mucosal thickness of maxillary sinus was thicker than the frontal sinus mucosa, and the bottom mucosal thickness was slightly thicker than the top.



Experimental Study on Treatment of Rheumatoid Arthritis in Rabbits by Injection of Deer Melon Polypeptide

FENG Yan-qin, AO Ji-bo, YANG Min, CHEN Jian-ming, CHEN De-sen

2016, 36(3):  204-207.  DOI: 10.3969/j.issn.1674-5817.2016.03.009

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Objective To study the mechanism of the treatment on rheumatoid arthrist (RA) in rabbits by deer melon polypeptide. Methods Calf cartilage II collagen+Freund’s complete adjuvant (0.1 mL) was injected intra-articular in right knee of rabbits to prepare RA model. The model rabbits were randomly divided into model group, low and high dosage deer melon polypeptide group (Group A and B) with 1 mg/d or 2 mg/d. The paw swelling were calculated at 1, 7, 14, 21 days after treatment, the hyaluronic acid (HA), interleukin-1β (IL-1β), matrix metalloproteinase (MMP-3) and bone morphogenetic protein (BMP) content in joint effusion were measured. Results Compared with the model group, paw swelling were occurred in the rabbits of group A and B, MMP-3 and IL-1β were significantly decreased, BMP and HA significantly increased(P<0.05), but no difference between groups A and B (P<0.05). Conclusion The deer melon polypeptide had the treatment effects on RA in rabbit by improving the RA synthesis and BMP secretion in articular, and inhibiting the erosion of MMP-3 and IL-1β on synovial.

Influence of Reduced Glutathione Added Medium on in vitro Fertilization in C57BL/6J Mouse

ZHAO Chen, ZHANG Shuang-qin, Wang Li-feng, LI Wei

2016, 36(3):  208-211.  DOI: 10.3969/j.issn.1674-5817.2016.03.010

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Objective To study the influence of reduced glutathione (GSH) in fertilization medium on the in vitro fertilization (IVF) in C57BL/6J mouse. Methods The fertilization medium without reduced glutathione was prepared as control group, and the different concentrations of GSH (0.25 mmol/L, 0.50 mmol/L, 0.75mmol/L, 1.00mmol/L, 1.25mmol/L and 1.50mmol/L) were added to the fertilization medium during IVF, which were divided into 6 experimental groups, to investigate its effect on the fertilization and early embryo development of C57BL/6J inbred mice. In addition, 5 strains of genetically engineered mice having a genetic background of C57BL/6J were chosen to verify the influence of GSH. Results Compared with the control, GSH increased the fertilization rate in a dose-dependent manner. Statistically significant increased fertilization was first observed at 0.5 mmol/L GSH (P<0.05), reaching maximum values at 1.25 mmol/L(P<0.01). The blastocyst developmental rates have no significant difference from that of the control group (P>0.05). Fertilization and development rates of the genetically engineered mice were similar to those of C57BL/6 inbred mice. Conclusion In a certain range of concentration of GSH during the IVF procedure, it can enhance the fertility of C57BL/6J mouse and the genetically engineered mice having a genetic background of C57BL/6J.

Influence of Microwave Radiation on Reproductive Function in Laboratory Animals

GENG De-jun, LV Zhao-hui, LI Xi-ping

2016, 36(3):  231-236.  DOI: 10.3969/j.issn.1674-5817.2016.03.015

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It is inevitable in today’s world to use electronics. Electromagnetic radiation(EMF) from different sources, such as microwave ovens, wireless communication, etc, is the main part of indoor-outdoor electromagnetic field exposure spectrum. Whether or not EMF, especially microwave may do harm to human health is not sure. A systematic review was therefore conducted, by reading papers published in recent years, to determine whether exposure to microwave affects fertility of laboratory animals. Most of the studies agree that microwave is harmful to female or male animals. It leads to abnormality of ovary, implantation, pregnancy, fetus, hormone or testicle, and a decrease in sperm parameters along with an increase in apoptosis-positive cells. The injury mechanism is by inducing oxidative stress. But several of studies didn’t supported it. Further study is required to give a certain conclusion.

Progress in Research of Laboratory Animal Behavior and Environment

TIAN Li-li, WEI Xiao-feng, GAO Cheng

2016, 36(3):  237-242.  DOI: 10.3969/j.issn.1674-5817.2016.03.016

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Laboratory animal behavior colligates the wild and wonderful ways in which animals interact with each other living beings, and with the environment. These behavior is the product of an individual’s innate qualities (“nature”) versus those shaped by personal experience (“nurture”). Environment contains all the resources with which the animals come directly in contact. This article reviews the effects of environment factors on laboratory animal behavior. Based on the related standards, the paper explores the relationship between environment control and laboratory animal holding, which will benefit the animal welfare.