Table of Content

25 December 2019, Volume 39 Issue 6

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Influence of Strain, Gender and Hair Coat of Mice on Establishing Bioluminescent Imaging Pseudovirus Mouse Model

LEI Shan, LIU Qiang, HUANG Wei-jin, WANG You-chun

2019, 39(6):  423-428.  DOI: 10.3969/j.issn.1674-5817.2019.06.001

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Objective To investigate the effects of strain, gender, and hair coat of mice upon establishing a bioluminescent imaging based pseudovirus mouse model. Methods C57BL/6J and BALB/c mice were infected with pseudotyped Marburg virus by intraperitoneal (ip) injection. The bioluminescent signal of mice was collected before and after removing hair to show the influence of hair upon the bioluminescent imaging signal. Meanwhile, C57BL/6J and BALB/c mice of different gender were infected with pseudotyped Ebola virus by ip and the dynamic changing process of bioluminescence over time was observed. Results The black hair coat of C57BL/6J mice blocked 82% of the bioluminescent signal while the white hair coat of BALB/c mice blocked 32%, the blocking rates are statistically different between C57BL/6J mice and BALB/c mice (P<0.001). The bioluminescent signal of male mice was stronger (P<0.05) and more homogeneous than the females. The dynamic changing process of bioluminescence showed that the peak of signals of BALB/c mice were reached on the fourth day after infection while those of C57BL/6J mice were observed on the sixth day after infection. Conclusion The strain, gender and hair of mice have significant influence upon the bioluminescent imaging based pseudovirus mouse model.



Comparative Studies on Behavior of C57BL/6J、129/SvJ and Their Hybrid Mice

WAN Ying-han, SHEN Rui-ling, WANG Jue, GONG Hui, LI Jun, DONG Ru, FEI Jian

2019, 39(6):  429-436.  DOI: 10.3969/j.issn.1674-5817.2019.06.002

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Objective To compare the genetic backgrounds of C57BL/6J and 129/SvJ and the behavioral phenotypes with their hybrid (F1) mice. Methods C57BL/6J and 129/SvJ were cross mated and bred to obtain two F1 (129B6F1 and B6129F1). These F1 mice with C57BL/6J and 129/SvJ were divided into 8 groups of experimental mice according to different gender. Numbers of mice in each group were no less than 12. Behavioral studies such as the open field test, the tail suspension test, the morris water maze, the passive avoidance test, the hot plate were conducted to compare the behavioral performance of C57BL/6J, 129/SvJ and F1 from the paramters of anxiety, depression, learning and memory, and heat pain response respectively. Results Compared with 129/SvJ, F1 had significant difference in peripheral and marginal activities in the open field activities (P<0.01). There was no significant difference in the behavior of F1 and parent mice in the tail suspension activities. Compared with C57BL/6J, the latency of finding the platform and entering the dark box of F1 were significantly different in the morris water maze and the passive avoidance activities (P<0.05). Conclusion Different behavior phenotypes of F1 are influenced by different genetic backgrounds of their parents.



Protective Effect of Naoxintong Capsule on Brain Cell Injury in Rats with Ketamine Addiction

LIU Xiao-bo, YANG Mei, Song Jia

2019, 39(6):  437-442.  DOI: 10.3969/j.issn.1674-5817.2019.06.003

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Objective To investigate the protective effect of naoxintong capsule on brain cell injury in rats with ketamine addiction. Methods Sixty SD rats were randomly divided into control, model, Naoxintong capsule with high, medium and low dose, and positive control group, with 10 rats in each group. The model group, Naoxintong capsule high, medium and low dose group and positive control group was intraperitoneally injected with 50 mg/mL ketamine hydrochloride injection (10 mg/kg). Naoxintong Capsule high, medium and low dose group was gavaged with Naoxintong Capsule (500 mg/kg, 250 mg/kg and 100 mg/kg). Positive control group was intraperitoneally injected with L-tetrahydropalmatine (20 mg/kg). Control group was intraperitoneally injected with same amount of normal saline, once a day for 7 days. The conditional position preference (CPP) of each rat was detected before and after the experiment. After experiment, expression of brain tissue c-fos gene was detected by immunohistochemistry. The expression of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax) and cysteinyl aspartate specific proteinase-3 (Caspase-3) protein in brain tissue was determined by Western blotting. HE staining was performed to observe pathological changes of brain tissue lesions. Results Compared with the control group, the CPP of the model group, Naoxintong capsule high, medium and low dose group and positive control group were significantly increased (P<0.05), c-fos gene, expressions of Caspase-3 and Bax protein in brain tissue were significantly increased (P<0.05), Bcl-2 expression was significantly decreased (P<0.05). Compared with the model group, CPP of Naoxintong capsule high and medium dose group and positive control group was significantly decreased (P<0.05), c-fos gene, expression of Caspase-3 and Bax protein were significantly decreased (P<0.05), and Bcl-2 expression was significantly increased (P<0.05). HE staining showed that degree of brain cell lesions was reduced. Conclusion Naoxintong Capsule may protect brain cells injury of rats with ketamine addiction by regulating apoptosis of brain tissues.



Experimental Observation on Effect of Astragalus Polysaccharides on Learning Ability in Rats with Post-stroke Depression

WANG Yuan-yuan, DENG Wei-ping, LI Xiao-qiang, PENG Jia-xia

2019, 39(6):  443-448.  DOI: 10.3969/j.issn.1674-5817.2019.06.004

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Objective The effects of Astragalus polysaccharides on learning and memory ability and depression-like behavior in rats with post-stroke depression (PSD) model were observed and analyzed. Methods Forty-eight Wistar rats were randomly divided into sham operation group, depression model group, astragalus polysaccharide low dose group and high dose group (n=12). PSD models were constructed except for the sham operation group. Rats in the low-dose group and the high-dose group were intragastrically administered with the corresponding dose of Astragalus polysaccharide. The depression model group and the sham operation group were given the same volume of saline as the control, and the 4 groups were all treated for 4 weeks. The edema intake experiment and Zero water maze analysis system were used to evaluate the depression behavior of rats. The patch clamp technique was used to record the spontaneous action potential (AP) and group peak potential (PS) in hippocampal CA1 area to evaluate the learning ability of Astragalus polysaccharides in PSD model rats. Behavioral effects; serum cytokines interleukin-1β (IL-1β) and interleukin-6 (IL-6) levels were detected by double antibody sandwich ELISA; nuclear transcription factor-κB (NF-κB) was detected by Western blotting in hippocampal CA1 region Peroxisome proliferator-activated receptor-gamma (PPAR-γ) protein expression, real-time quantitative (RT-PCR) detection of NF-κB and PPAR-γ gene expression. Results In the low-dose and high-dose groups of Astragalus polysaccharides, the sucrose intake increased significantly, the escape latency was shortened, and the number of crossovers increased. The AP and PS membrane voltages and serum IL-1β and IL-6 levels in CA1 area were significantly decreased. NF-κB protein and NF-κB mRNA were down-regulated in CA1 region, while PPAR-γ protein and PPAR-γ mRNA were highly expressed, and the difference was statistically significant (P<0.05). The difference was statistically significant (P<0.05). Conclusion Astragalus polysaccharide can alleviate the depression-like behavior of PSD model rats and improve learning and memory ability. The mechanism may be to reduce the release of cytokines IL-1β and IL-6 regulated by NF-κB signaling pathway and reduce middle cerebral artery occlusion (MCAO). Traumatic stress and inflammatory response are associated with increased membrane stability in the CA1 region.

Establishment of Necrotizing Enterocolitis Model in SD Rats with Different Ages

PAN Zhao-jun, TIAN Zhao-fang, WU Rong

2019, 39(6):  449-453.  DOI: 10.3969/j.issn.1674-5817.2019.06.005

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Objective To establish and compare two models of necrotizing enterocolitis (NEC) in neonatal SD rats at different ages. Methods New born SD rats within postal 2 hours and 48 hours (each time point 24 rats) were randomly divided into three groups respectively according to body weight as the artificial feeding group (groups A, B), the artificial feeding + lipopolysaccharide (LPS) group (groups C, D), and control group (groups E, F), namely postal 2 hours (groups A, C, E) and postal 48 hours (group B, D, F). The rats in 4 groups A, B, C and D were treated with hypoxia plus cold stimulation twice a day, group C and D were also given LPS once a day by gavage, and the control group was fed only by female rats. The rats were executed 84 hours after the experiment time point with fasting. The proximal ileocecal ileum was removed about 1 cm for histomorphological observation. The intestinal histomorphology was observed and the intestinal histopathological score was made. Results The rats in group A, B, C and D had yellow-green mucus defecation, abdominal distension, slow weight growth, gastric retention, vomiting, lethargy, decreased activity and gradually aggravated. Neonatal rats in two control groups had better milk intake, good growth and development. The histological scores and the incidence of NEC in groups A, B, C, and D were significantly higher than those of control groups. Conclusion NEC-compliant models both can be induced in neonatal SD rats within postal 2 hours and 48 hours by artificial nasal feeding, low body temperature and hypoxia, LPS gavage can aggravate the clinical manifestations and histopathological changes.



Effects of Different Concentrations of Chromium-containing Bedding Materials on Toxity of Blood and Organs in BALB/c Nude Mice and KM Mice

JIA Huan-huan, ZENG Ye-wen, LUO Ting, GONG Bao-yong, MAI Dong-mei, PAN Ying-chun, ZHAO Wei-bo

2019, 39(6):  454-461.  DOI: 10.3969/j.issn.1674-5817.2019.06.006

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Objective To evaluated the toxic effects of different concentrations of chromium-containing bedding materials on blood and organs of BALB/c nude mice and KM mice. Methods The stable different concentrations of chromium-containing bedding materials were prepared, 48 BALB/c nude mice and 48 KM mice were randomly divided into 4 groups respectively, 12 mice per group with half males and females, the dosage of chromium in control, low-dose, middle-does and high-does groups were (2.30±1.12) mg/kg, (15.50±0.71) mg/kg, (71.50±3.54) mg/kg and (347.00±35.36) mg/kg, respectively. BALB/c nude mice and KM mice were raised for 15 weeks with different bedding materials which were replaced once a week, general condition of mice were observed daily, the mice were dissected at the 15th week and the blood was collected. Besides, haematological and biochemical indexes of blood plasma, organ coefficient and organ pathology were detected. Results After 15 weeks with different bedding materials, no abnormalities and death were observed in BALB/c nude mice and KM mice, no significant differences of haematological index and organ coefficient were found between the four groups, and no abnormal anatomy was observed. In BALB/c nude mice, the alanine aminotransferase (ALT) and blood urea nitrogen (BUN) levels significantly increased in low, middle and high-dose groups, alkaline phosphatase(ALP) and creatinine (Crea) levels significantly increased in middle and high-dose groups, Triglyceride (TG) levels significantly decreased in low and high-dose groups. In KM mice groups, no significant differences of biochemical index between four groups were observed. The organ pathology showed varying degrees of focal inflammatory infiltrates and cellular swelling in the liver, inflammatory infiltrates in kedney, atrophic seminiferous tubular and decreasing corpus luteum in both BALB/c nude mice and KM mice. Conclusions Hexavalent chromium content greater than (15.50±0.71) mg/kg can cause varying degrees of toxicity in liver, kidney, testis and ovary of BALB/c nude mice and KM mice.



Analysis of Goiter in Zebrafish

Li Ying-niang, Dai Wei, Sheng Jian

2019, 39(6):  462-466.  DOI: 10.3969/j.issn.1674-5817.2019.06.007

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Objective To investigate the causes of goiter in zebrafish, and explore whether the disease is caused by iodine deficiency. Methods The enlarged tissue in the lower jaw of zebrafish was observed through HE staining and electron microscopy. Meanwhile, both iodine-fed and iodine-free feeding groups were set up, and the expressions of five thyroid-related genes were detected at different periods. Result According to HE staining, goiter was confirmed. Through electron microscopy observation, the mitochondria was enlarged and lysosomes was increased in the diseased tissues. The expression of corticotropinreleasing factor (CRF) was different between the two groups at 7 days post fertilization (7dpf). The expression of CRF in iodine group was significantly higher than that in non-iodine group. Conclusion Iodine deficiency can cause goiter in zebrafish,iodine deficiency causes a decrease in CRF expression.



Establishment of a Purified Hippocampal Neuron Model in Naked Mole-rat and Preliminary Study on Its Hypoxic Tolerance

YANG Wen-jing, LI Yu, FENG Yan, LI Lei-chen, XU Yue-rong, LIU Pan, LIN Li-fang, SUN Wei, CUI Shu-fang

2019, 39(6):  467-472.  DOI: 10.3969/j.issn.1674-5817.2019.06.008

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Objective To establish a method for purifying and culturing hippocampal neurons of naked mole-rats and to preliminarily explore their hypoxic tolerance characteristics. Methods Cells in hippocampus of naked mole-rats during 60-65 days of gestation were collected, and purified by the medium containing Neurobasal+2%B27+10 μmol/L 5-fluorouracil. Hippocampal neurons were maintained by neurobasal medium containing 2% B27. Cell growth was observed by inverted microscope after 6-7 days of adherent culture in vitro. Neuron-specific antibody was used to identify neuron-specific antibody. CCK8 kits were used to study on hypoxic tolerance of hippocampal neurons. Axonal length of naked mole-rat hippocampal neurons were measured by Image-Pro Plus 6.0. QPCR analysis was used to detect the hypoxia-inducible factor 1α (HIF1α) mRNA levels of hippocampal neurons under normoxia and hypoxia conditions. Results The hippocampal neurons obtained by this method exibits typical morphological characteristics of neurons, and immunocytochemical staining shows that the purified cells are NeuN-positive. The cell survival rate under hypoxia is more than twice of that under conditions with normal oxygen, and the axon lengths of cells in hypoxia conditions were smaller than those under nomoxia conditions. HIF1α levels of naked mole-rat hippocampal neurons were higher than those under normoxia conditions. There are statistically differences among these results (P<0.05). Conclusion The method established in this study is a simple and practical method for isolating and culturing hippocampal neurons of naked mole-rats in vitro, and has strong hypoxic tolerance.



Comparative Study of Effect of Poly I:C on PKR/eIF2α Signal Path in Naked Mole-rats and Mouse Macrophages

LIN Li-fang, ZHANG Cheng-cai, LI Yu, ZHANG Qian-qian, CHEN Chao, LI Lei-cheng, YANG Wen-jing, LI Li, CUI Shu-fang

2019, 39(6):  473-478.  DOI: 10.3969/j.issn.1674-5817.2019.06.009

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Objective To compare the affect of polyinosinic-polycytidylic acid(Poly I:C) on double-stranded RNA-dependent protein kinase R (PKR)/eukaryotic initiation factor-2α (EIF2α) signaling pathway in macrophages of naked mole-rats and mouse. Methods The naked mole-rat and C57BL/6J mice which physiological age-matched were randomly divided into control group, Poly I:C administration group, 2,6-Diaminopurine (2-AP) administration group and Poly I:C+2-AP administration group. After administration, the expression of phosphorylated PKR (Pho-PKR), PKR, phosphorylated EIF2α (Pho-EIF2α), EIF2α and Caspase-8 protein in the cells were detected by Western blotting. Results After Poly I:C administration, the phosphorylation levels of PKR and eIF2α in mice macrophages were significantly decreased (P<0.01), while those in naked mole-rat were significantly increased (P<0.01).This phenomelon indicated that Poly I:C could inhibit PKR activity in mice, but activite PKR in naked mole-rat; Poly I:C and 2-AP intervention group showed that PKR and phosphorylated PKR were down-regulated both in mice and naked mole-rats (P<0.01), especially PKR in naked mole-rats. Active PKR and active eIF2α was significantly decreased (P<0.01) in mouse macrophages, after 2-AP administration. Active PKR in naked mole-rat was significantly decreased (P<0.01) too. After inhibition of PKR activity by 2-AP, Caspase-8 expression levels in naked mole-rat were significantly reduced (P<0.01), but there were no significant differences in mouse cells. Conclusion Compared to mouse, naked mole-rat's cells were highly resistant to Poly I:C stimulation,PKR in naked mole-rats was more sensitive to 2-AP, PKR activity in naked-mole rat cells promoted the expression of Caspase-8.



Comparative Study on Radiation Resistance of Naked Mole-rat and Mice

CHEN Chao, CONG Wei, YANG Wen-jing, CUI Shu-fang

2019, 39(6):  479-483.  DOI: 10.3969/j.issn.1674-5817.2019.06.010

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Objective The radiation resistance of naked mole-rats and mice and their skin fibroblasts were investigated by gamma ray irradiation at different doses and frequencies. Methods Adult mice (8 weeks old) and adult naked mole-rats (52 weeks old) were treated with ⁶⁰Co gamma ray irradiation in low doses by multi-times or high doses one-time, respectively, to analyze their basic vital signs, changes in peripheral blood lymphocytes and pathological changes in major organs. Skin fibroblasts from naked mole-rats and mice were irradiated with different dosages to detect the damage of reactive oxygen species (ROS). Results Thirty days after one-time irradiation with 6 Gy, the survival rate of mice was 30%, and the ratio of helper T lymphocytes and killer lymphocytes (CD4+/CD8+) and dendritic cells (DC) in their peripheral blood showed no significant change. However, the survival rate of naked mole-rats on day 30 was 100%, CD4+/CD8+ in peripheral blood increased significantly on day 4 after irradiation, and DC percentage increased first and then decreased sharply. After repeated irradiation with low dosage, the bone marrow, thymus, liver and spleen and other major tissues and organs of mice all showed different degrees of lesions, while the corresponding organs of naked mole-rats showed no obvious abnormalities. There was not any significant change in ROS in skin fibroblasts of naked mole-rats and mice irradiated by 6 Gy dose. But, under radiation of 50 Gy doses, the oxygen injury of skin fibroblasts of naked mole-rats was significantly lower than that of mouse skin fibroblasts. Conclusion Naked mole rats have natural resistance to ionizing radiation. Their vital signs, morphology and functions of tissues and organs have not changed markedly due to external radiation factors, and their immune level is activated. Skin fibroblasts of naked mole-rats also exhibit anti-ionizing radiation properties.



The Characteristics of Anti-chemical Cancer Inducing Factors in Naked Mole-rat

CHEN Chao, CONG Wei, YANG Wen-jing, CUI Shu-fang

2019, 39(6):  484-488.  DOI: 10.3969/j.issn.1674-5817.2019.06.011

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Objective Chemical carcinogens were used to induce naked mole-rats and mice, respectively, to study the characteristics of anti-chemical carcinogens in naked mole-rats. Methods In the adult naked mole-rats (52 weeks) and the adult mice (8 weeks), urethane and o-aminoazotoluene (OAAT) were respectively used to induce tumor formation in the two groups of animals for 3 months.The incidence of tumor in animals were observed, the leukocyte count and body weight of the animals were detected after urethane injection. The pathological changes in the main organs of the animals were observed after OAAT smear. Results After 3 months of urethane mutagenesis, lung tumors could be observed in the mice, and the average body mass continued to decline and the content of white blood cells significantly decreased. Two months after the end of mutagenesis, the incidence of the tumor was more than 50%. However, the naked mole-rat group still maintained a healthy physiological state, with no significant changes in average body weight, white blood cell indicators. Three months after OAAT mutagenesis, obvious pathological changes occurred in thymus, spleen, liver, small intestine and bone marrow of mice, while no obvious lesions were found in the corresponding organs and tissues of naked mole-rat. Conclusion Naked mole-rats have natural resistance to chemical carcinogens, and their living conditions, biochemical indicators, and the morphology and functions of tissues and organs cannot be affected by external chemical carcinogens.



Six Kinds of Commonly Used Disinfectants for Laboratory Animal Barrier Facilities

PAN Ming-ming, CHEN Wang-wang, WANG Yan, WANG Jing, ZHOU Zheng-yu

2019, 39(6):  489-495.  DOI: 10.3969/j.issn.1674-5817.2019.06.012

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Control of pathogenic micro-organisms in laboratory animal barrier facilities is not only related to the quality of laboratory animals, the credibility of experimental results, but also the health of personnel and the safety of the environment. However, during the daily operation of laboratory animal barrier facilities, pathogenic micro-organisms may be introduced via the entry and exit of personnel and articles, etc, so daily sanitation and disinfection of the barrier environment is very important. In this paper, 6 kinds of commonly used disinfectants for laboratory animal barrier facilities were selected, and their properties, mechanism of action, toxicity, occupational health and safety, effects and other parameters were summarized, so as to provide reference for selecting reasonable and effective disinfectants for laboratory animal barrier facilities.

Application of Patient-Derived Xenograft Models for Personalized Treatment of Pancreatic Cancer

HU Bin-quan, CHU Hai-meng, SHI Chang-hong

2019, 39(6):  499-504.  DOI: 10.3969/j.issn.1674-5817.2019.06.014

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This paper introduces the basic characteristics and problems in modeling and the application of patient-derived xenograft (PDX) models for pancreatic cancer,and expounds the feasibility of PDX models in the study of individualized drug screening and drug resistance mechanism of pancreatic cancer. Conventional standard chemotherapy and combination therapy regimens for pancreatic cancer are less effective. Therefore, it is necessary to pay attention to the importance of targeting and remodeling the anti-tumor study of pancreatic cancer microenvironment so that it can provide new ideas for researchers, reasonably use this model to carry out pancreatic cancer research,and provide more efficient personalized treatment for patients with pancreatic cancer.