Table of Content

25 August 2019, Volume 39 Issue 4

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Research Progress on Infection Model in Experimental Pigs

WANG Yu-e, ZHANG He, CHEN Hong-yan

2019, 39(4):  253-259.  DOI: 10.3969/j.issn.1674-5817.2019.04.001

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Traditional biomedical model animal is easy to manage in the experimental facility, which can be quickly and efficiently applied to the basic study of human disease related genes. However, in some cases, they cannot represent the complexity of human physiology. Pig is highly similar to human beings in anatomy and physiology, and widely used in related research on human diseases. In this review, we summarized the present progress of pigs as human infectious disease model, and the characterization of disease infection model.

Isolation and Identification of Oligodendrocyte Precursor Cells from Tree Shrews Cerebral Cortex

LI Ming-xue, LI Xiao-hui, HUANG Xin, WANG Xuan, ZHANG Zhi-cheng, YUAN Yuan, LU Cai-xia, SUN Xiao-mei

2019, 39(4):  260-266.  DOI: 10.3969/j.issn.1674-5817.2019.04.002

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Objective To establish an effective method of isolation, identification and primary culture for oligodendrocyte precursor cells (OPCs) which derived from the cerebral cortex of tree shrews in vitro. Methods Newborn tree shrews within 48 hours were given euthanasia, the cerebral cortex were collected, removed the meninges and brainstem. Then, the cerebral cortex was digested with trypsin and DNase I, and primary cultured. Oligodendrocyte precursor cells were purified by chemical separation and differential adherence. The oligodendrocyte precursor cells were identified by immunofluorescence staining of chondroitin sulfate proteoglycan 4 (NG2) and myelin basic protein (MBP). Results The purified oligodendrocyte precursor cells showed round or oval cell body, with strong refractive and bipolar or tripolar protuberances. The purity of the cells was 97% by immunofluorescence staining of NG2. After adding differentiation medium, oligodendrocyte precursor cells could differentiate and mature, and the immunofluorescence staining of MBP is positive. Conclusion A method for isolation and purification of cerebral cortex oligodendrocyte precursor cells from tree shrews was successfully established.



Investigation on Behavioral and Pathological Characteristics of Tg2576 Mice

GUAN Ya-lun, LIU Shu-hua, HUANG Zhong-qiang, LI Yun-feng, LI Xue-jiao, LI Ge, ZHANG Yu

2019, 39(4):  267-273.  DOI: 10.3969/j.issn.1674-5817.2019.04.003

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Objective Tg2576 transgenic mice with C57BL/6 and SJL hybridization background, which are commonly used in research of Alzheimer’s disease (AD), were used to investigate the potential effect of long-term breeding on the phenotype of AD. Methods Utilizing Morris water maze, immunohistochemistry, ELISA and immunofluorescence, the behavior and pathological phenotype of AD were studied in these Tg2576 mice which were introduced and bred in our laboratory for a long time. Results With the age increase, the learning and memory ability of in these Tg2576 mice was still significantly decreased. The typical beta amyloid (Aβ) deposition was also found in the cerebral cortex of these mice by immunohistochemistry. The content of Aβ in brain tissue was significantly higher than that in control mice. In addtion, the immunofluorescence analysis indicated that a large number of astrocytes gathered around Aβ plaques in the cerebral cortex. Conclusion The long-term bred of Tg2576 mice still have the typical clinical and pathological characteristics of AD mice.



Effects of Fhl1 Knockout on the Differentiation and Autophagy in Gastrocnemius Muscle in Mice of Different Ages

MIAO Jia-ning, LIU Bo, DING Jing-jing, WANG Li-li

2019, 39(4):  274-279.  DOI: 10.3969/j.issn.1674-5817.2019.04.004

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Objective To explore the effects of Fhl1 knockout (Fhl1-ko) on the differentiation and autophagy in gastrocnemius muscle in mice of different ages. Methods Wild-type (wt) and Fhl1-ko type male mice at 2.5-month-old and 8-month-old were studied and protein expressions of autophagy (Beclin-1), autophagic flux (increased LC3B-II/LC3B-I ratios and a lack of p62 accumulation) in gastrocnemius muscle were investigated by Western blotting. Transitions in myofiber phenotype in gastrocnemius muscle of Fhl1-ko mice at different ages were examined by real-time quantitative RT-PCR. Result Compared with wt mice, protein expression data showed that Beclin-1, LC3B-II/LC3B-I ratios were all increased in gastrocnemius muscle in Fhl1-ko mice at 2.5-month age. At the same time, p62 accumulation was not evident. LC3B-II/LC3B-I ratios were increased in gastrocnemius muscles in Fhl1-ko mice at 8-month age, but Beclin-1 was not clearly increase. Furthermore p62 accumulation was clearly evident. Transitions in the myofiber phenotype in the gastrocnemius muscle indicated MYH IIA and MYH IIX expression were decreased and MYH IIB increased in 2.5-month Fhl1-ko mice, compared with wt mice. However, MYH I and MYH IIA were both increased in 8-month Fhl1-ko mice compared with 8-month wt mice. Conclusion Effects of Fhl1-ko on the differentiation and autophagy in 2.5-month and 8-month gastrocnemius muscle were different which indicated regulation of autophagy might be a promising therapeutic target for hereditary myopathies induced by FHL1 and precise treatment should be used for patients of different ages.



Investigation on Radiosensitivity Impact of Lung Cancer A549 Cells by Vascular Endothelial Growth Factor Silencing in vivo

MA Jun, LUO Jun-dong, JING Wen-jiang, ZHANG Shu-lian, WANG Juan-yi, FAN Zhi-gang

2019, 39(4):  280-286.  DOI: 10.3969/j.issn.1674-5817.2019.04.005

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Objective To research the radiosensitivity of A549 cells in tumor-bearing mouse model by knock down vascular endothelial growth factor (VEGF) gene. Methods The tumor-bearing mouse model was established by using A549 cells which was cultured in vitro. The siVEGF/RNAi-MateTM mixture was injected into the tumor of model mice and with radiotherapy. The nude mice were euthanasia on day 24 after radiotherapy. The expressions of VEGF were detected by immuno-histochemical, Q-PCR and Western blotting. Results Both the protein and mRNA levels of VEGF in VEGF knockdown and radiotherapy group were significantly decreased compared with the control group (P<0.05). The volume of tumor was decreased and the necrotic area of tumor were increased distinctly in siVEGF transfected and radiotherapy group compared with control group (P<0.05). Conclusion Knock down of VEGF and radiotherapy can inhibit tumor growth, and increase the radiosensitivity of lung cancer radiotherapy.



Immunization with Leptin Promotes Fat Accumulation and Up-regulates mRNA Expression of PPARγand SREBP-1

WU Li-hong, LI Hong-tao, Guan Hong-bing, GU Wei-wang, Huang Li-wen

2019, 39(4):  287-293.  DOI: 10.3969/j.issn.1674-5817.2019.04.006

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Objectives To assess the effects of intramuscular injection of exogenous leptin on fat deposition and mRNA expression of fat metabolism related genes in subcutaneous, hypothalamus and liver of rat. Methods Thirty female SD rats were injected thrice with leptin intramuscularly. At the end point of this study, the rats were sacrificed, then the body weight, abdomen fat weight and liver weight were measured. The serum were collected and the total cholesterol, high density lipoprotein, low density lipoprotein and triglycerides were detected. Subcutaneous and abdominal fat were blocked,made into paraffin slices and stained with HE. Livers were blocked, made into frozen slices and stained with Oil Red O. Pathological analysis of fat distribution in the subcutaneous, abdominal and liver. The mRNA expression levels of abdominal fat Leptin, hypothalamus peroxisome proliferator-activatedreceptor-gamma (PPARγ) and liver sterol regulatory element binding protein-1(SREBP-1) were qualified by fluorescence quantitative PCR. Results SD rats administered with leptin immunogen displayed significantly increased body weight, total cholesterol, low density lipoprotein, triglycerides and fat deposition in subcutaneous, abdominal and liver. Conclusions Intramuscular injection of exogenous leptin promotes fat accumulation in subcutaneous, abdominal and liver of rat by up-regulating of Leptin, PPARγ and SREBP-1.



Effect of Different Freund's Adjuvant Dose on Establishment of Immune Premature Ovarian Failure Model in B6AF1 Mice

CHEN Yue, WANG Pei-juan, CHEN Si, LU Yan

2019, 39(4):  294-297.  DOI: 10.3969/j.issn.1674-5817.2019.04.007

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Objective To observe the effect on different doses of Fletcher’s adjuvant to B6AF1 mice with autoimmune premature ovarian failure (POF) induced by zona pellucida 3 peptide (pZP3). Methods Autoimmune POF models were established in 60 female B6AF1 mice (6-8 weeks old) by immunization with pZP3 and another 20 mice served as control group. The POF models were randomly divided into incomplete Freund’s adjuvant group, complete Freund’s adjuvant group (1 mg/mL) and incomplete Freund’s adjuvant + tuberculosis (TB) group (3.3 mg/mL). The HE staining ovary histomorphology results were observed by the microscope. Results Compared with those in control group, the mice in the model group showed a smaller volume of the ovary, and larger amount of atretic follicle of ovary. Immunohistochemical studies showed that the model mice had redundant lymphocytes infiltrating around follicle compared with control mice. The immunological effect of incomplete Freund’s adjuvant +TB (3.3 mg/mL) group was the best, followed by complete Freund’s adjuvant group. Conclusion Different concentrations of Freund’s adjuvant combined with zona pellucida 3 polypeptide (pZP3) can induce autoimmune premature ovarian failure (POF) in B6AF1 mice. High concentration of Mycobacterium tuberculosis has the best effect.



Expression of Phospholipase A2 in Different Xenografted Tumor Models of Nude Mice

TU Jue, ZHOU Wei-min, Cai Yue-qin, Ling Yun, Chen Min-li

2019, 39(4):  298-304.  DOI: 10.3969/j.issn.1674-5817.2019.04.008

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Objective To observe the expression of phospholipase A2 (PLA2) in different human cancer xenograft models in nude mice. Methods Different human cancer xenograft models, including hepatocarcinoma, pancreatic cancer, gastric cancer, colon cancer, esophageal cancer, lung cancer, nasopharyngeal carcinoma, ovarian cancer, were established by tumor cell lines in nude mice. After the tumors grow to a volume of 1 cm³, the protein expression of four types of PLA2 enzymes in the tumor tissues, normal tissues, and serum were detected by ELISA, which in the serum of normal nude mice were tested as control. Results In the nude mice xenograft models of respiratory, digestive and reproductive system, the protein expressions of cPLA2, sPLA2 and lp-PLA2 in tumor tissues were significantly higher than those in normal tissues (P<0.05, P<0.01). Similarly, the protein expression of iPLA2 in tumor tissues was higher than that in normal tissues, except for human colorectal cancer cell line HT-29, which showed significant difference (P<0.01). Moreover, in the eight xenograft models, the protein content of four types of PLA2 in serum of tumor-bearing mice were obviously higher than those of normal nude mice (P<0.01). Conclusions Expression of four types of PLA2 in tumor tissues and serum were both up-regulated significantly in all the xenografted models, including respiratory cancer, digestive system cancer and reproductive system cancer, which did not limit to specific cancer type. The results inferred a key role for the PLA2 in the development of xenografted tumors in nude mice.



Establishment of Genetic Quality Standard of Inbred Chinese Hamster

XU Guo-qiang, GAO Ji-ping, LIU Mao-lin, CHEN Chao-yang, SONG Guo-hua

2019, 39(4):  305-309.  DOI: 10.3969/j.issn.1674-5817.2019.04.009

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Objective To establish genetic quality standard of inbred Chinese hamster (Cricetulus griseus). Methods Based on investigating a large number of related literatures, according to Laboratory animal-Genetic quality control of mammalian laboratory animals (GB14923-2010) and the theory of population genetics, the genetic quality standard of inbred Chinese hamster was established. The genetic quality of Chinese hamster was detected by microsatellite markers which were obtained from a Chinese hamster genomic enrichment library and the PCR primers of them were designed. Results The genetic quality standard and the detection method of inbred Chinese hamster were preliminarily established. Conclusion The establishment of genetic quality standard of inbred Chinese hamster will standardize the breeding and reproduction of inbred Chinese hamster and promote the further improvement of genetic quality standard system of laboratory animals in China.

Anesthetic Effect Evaluation of Combined Zoletil 50 with Dexdomitor in BALB/c Mouse

ZOU Lu, DAI Tian-e, LU Xiao, AI Xiao-jie

2019, 39(4):  310-313.  DOI: 10.3969/j.issn.1674-5817.2019.04.010

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Objective To evaluate the anesthetic effect of combined Tiletamine/Zolazepam (Zoletil 50) with Dexmedetomidine (Dexdomitor) in BALB/c mouse. Methods Nine male and 9 female BALB/c mice were randomly divided into 2 groups. A group mice were assigned to Zoletil 50 and Dexdomitor. B group mice were assigned to same dosage, 1 hour after stable anesthesia, added with Antipamezole (Antisedan). Results The anesthetic effect of combined Zoletil 50with Dexdomitor present well in BALB/c mouse. The recovery time of group B was shorter than group A (P<0.05). Conclusion Combined Zoletil 50 with Dexdomitor is an excellent anesthetic method for BALB/c mouse.

Study on Angiography in Mice with Barium Sulfate

QIAN Feng-tao, WANG Cheng-ji, WANG Kai, LI Jun, ZHAO Yi-yun, YUAN Yi-ting

2019, 39(4):  319-322.  DOI: 10.3969/j.issn.1674-5817.2019.04.012

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Objective To optimize the formulation of barium sulfate contrast agent and try to perform angiography in mice. Methods The arterial angiography of mice was performed with different ratios of hand-free disinfection gel-saline-barium sulfate contrast agent , and the imaging results were compared. Results After repeated tests, the optimum formula of barium sulfate suspension was obtained as follows: barium sulfate 4 g, hand disinfecting gel 1 mL, and saline 5 mL, with temperature 25 ℃ and perfusion volume 0.5~0.8 mL/25 g. Conclusion The optimized barium sulfate contrast agent can be used in mouse angiography and can obtain good results.



Molecular Genetical Research of Developmental Language Disorders in Foxp2 Mouse Models

LI Hui

2019, 39(4):  331-336.  DOI: 10.3969/j.issn.1674-5817.2019.04.015

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FOXP2 was the first gene discovered to be related to a fairly specific speech and language phenotype, called developmental verbal dyspraxia (DVD). This mechanism discovery opened up a new direction for the research of the relevant neural pathway. FOXP2 showed significantly high conservation of sequence and expression in the neural system of a variety of vertebrates, one typical example was that the expression patterns in human fetal brains were highly similar to those in mice of a comparable embryonic stage. Foxp2 mouse models include knock-out mice, mice that carry the etiological mutations that mirror those of the KE family or other patients, and mice in which the murine Foxp2 gene has been ‘humanized’ by including the two human-specific amino acids. The study of the Foxp2 mouse models was reviewed in this paper from molecular networks, sensory processing and motor skill learning, which aimed to shed light on the genetical mechanism of the language disorders.