Table of Content

28 December 2018, Volume 35 Issue 06

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Localization of ND3 and NF-κB on Livers and Kidneys From Hereditary Diabetic Mongolian gerbils

2018, 35(06):  1. 

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To investigate the localization and expression of NADH dehydrogenase 3 (ND3) and nuclear factor κB (NF-κB) in livers and kidneys of spontaneous hereditary diabetic and non-diabetic gerbils. We selected 6 diabetic and 6 non-diabetic control gerbils, and collected livers and kidneys from each animal, respectively. Immunohistochemistry was conducted to localize and semi-quantify ND3 and NF-κB in diabetic and non-diabetic control groups. Both ND3 and NF-κB were expressed in hepatocytes. And compared with the control group, NF-κB expression in livers was elevated significantly in the diabetic group. ND3 was also localized in renal tubular epithelial cells of kidneys. And ND3 was up-regulated in kidneys of diabetic gerbils, relative to the control gerbils. In conclusion , ND3 expressed in renal tubular epithelial cells was significantly up-regulated in kidneys of diabetic gerbils, and NF-κB expressed in hepatocytes was also increased in livers of diabetic gerbils, suggesting that the aberrant expression of ND3 in kidneys and NF-κB in livers may relate with the initiation and development of diabetes in gerbils.

Animal Experiment Research on the Dynamic Change of Chronic Urinary Tract Infection Disease

2018, 35(06):  7. 

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In order to establish the animal model of chronic urinary tract infection(CUTI)and study the dynamic changes of the relevant indicators, the CUTI model was induced by injecting Escherichia coli into the bladder of the female rat, and treated with levofloxacin hydrochloride at the same time. Indicators such as body temperature, number of urine bacteria, blood routine and BUN, Cr value were detected on day 0, 7, 14, 21 and 28. On day 31, number of bacterium in kidney and bladder was counted and histopathological examination was performed. Under the study, we discovered that on day 7 and 14, the body temperature elevated significantly(P<0.05)and the number of bacterium in urine, kidney and bladder increased obviously(P<0.05)in the model group rather than in the sham-operated group. Inflammation of bladder and kidney could be observed either. However, administrated with levofloxacin hydrochloride, the number of bacterium was significantly decreased no matter in urine or bladder or kidney. What’s more, inflammation of bladder and kidney were attenuated. In a word, the method carried out in this study is suitable for building CUTI animal model and pharmacodynamics observation.

The Effecct of Huoxue Xiaoyi Fang on Pelvic Adhesions and TGF-β1 of Endometriosis Rats

2018, 35(06):  15. 

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Objective To observe the therapeutic effect of Huoxue Xiaoyi Fang on pelvic adhesions and TGF-β1 in rats with endometriosis. Method To establish rat model of endometriosis pelvic adhesions by adopting the method of autologous transplantation that endometrial is transplanted on mesenteric.Rats were randomly divided into blank group, sham operation group,model group, Huoxue Xiaoyi Fang group and GnRH-α group.And then all rats were treated 28 days in a row.TGF-β1 protein and gene expression of rats’ adhesions peritoneal tissue or normal peritoneal tissue were detected. Result 1.The pelvic adhesions marks of model group, Huoxue Xiaoyi Fang group and GnRH-α group were all significantly higher than the sham operation group(P0.05).2.TGF-β1 protein and mRNA of model group were higher than the blank group and the sham operation group(P0.05). Conclusion TGF-β1 is involved in the occurrence and development of pelvic adhesions in endometriosis. Huoxue Xiaoyi Fang can improve the pelvic adhesions of endometriosis and inhibit the expression of TGF-β1 gene and protein in adhesion tissues.The curative effect of Huoxue Xiaoyi Fang is comparable to that of GnRH-α.

SCD1 Deletion Affects Organ Coefficients and Hematology Indexes of Mice

2018, 35(06):  22. 

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Objective To investigate the effect of SCD1 gene deletion on organ coefficients and hematology indexes of mice. Method Organ coefficients are observed in 8 weeks SCD1-/- mice and heterozygous SCD1mice as well as hematology indexes of different ages mice are compared between the two group. Result Liver coefficient of 6-8 weeks SCD1-/- mice markedly increase compared with heterozygous mice. RBC,MPV,PCT of SCD1-/- mice are significantly lower (P<0.05),while MCV,MCH,RDW,PDW,PLT,LYM are remarkbly higher(P<0.05) than those of the control group. ALB,TC,TG also obviously go down and ALP,ALT,UA clearly go up(P<0.01)in 6-8 weeks experiment group. MCHC, PLT, PCT,ALP, TC of 16-18 weeks significantly decrease compared with those of SCD1+/- mice. At the same time,RDW、HCT and MCV significantly increases(P<0.01). Conclusion SCD1 gene deletion impairs the organ coefficients of mice. Moreover, gene SCD1 deletion has impact on some hematology indexes.

he Feasibility of Recovering Strains Used for Transportation and Accidental Death of the Mice by Refrigerating Male Mouse Body

2018, 35(06):  28. 

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To investigate the feasibility of recovering strains used in the transportation and accidental death of the mice by refrigerating male mouse body at low temperature, based on the efficiency of in vitro fertilization (IVF) at different time length of mice were refrigerated at 4 ℃. All experimental C57BL/6 J male mice were 5 months old. Taking the mice cauda epididymis sperm fertilization in vitro after executed and refrigerated at 4 ℃ in 24 h, 48 h, 72 h and 96 h. The experiment of C57BL/6 J male mice executed and fertilization in vitro as the Control group. Statistical groups of the fertilization rate were calculated by groups. After 24 hours of refrigerated, the spermatozoa of the cauda epididymides had the capacity of normal fertilization, and the fertilization rates were 73.0%, 60.0% and 77.0%. There was the obvious difference within the group after 48 hours of refrigerated. The fertilization rates were 0%, 55.4% and 3.2%. After 72 hours of refrigerated, the fertilization rate was 3.2%, 0%, and 1.7%. After 96 hours of refrigerated, The sperm vitality, but no fertilization ability. All fertilized eggs can develop to 2-cell embryos. The pregnancy rates were 60% and 66% respectively by transplanting the 2-cell embryos of 24 h group and control group. It was proved that there was no effect on the early development of fertilized eggs at 4 ℃.C57BL/6 J mice had the stability of the fertilization rate in 24 h at 4 ℃, can be used for transporation and strain recovery after accidental death.

Study on the Stability of 5 Standard Substances for the Evaluation of Genetic Quality in Laboratory Animals

2018, 35(06):  32. 

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Objective To investigate the stability of 5 standard substances in order to obtain the storage condition and transportation condition of standard substance. Method On the basis of the national standard GB/T14927.1—2008, standard substances were prepared, stability test were carried on 4 temperatures(-20 ℃，4℃，room temperature and 37 ℃) and 8 time points (1 day、2 days、3 days、7 days、14 days、30 days、60 days and 90 days). Result 5 standard substances can be stably preserved during 90 days at 4 ℃ and -20 ℃. Catalase 2 standard substance can be preserved during 3 days at 37 ℃, the other 4 standard substances can be preserved only in 1 day at 37 ℃. At room temperature, malic enzyme 1, isocitric dehydrogenase 1 and catalase 2 can be preserved during 3 days. Alkaline phosphatase 1 can be preserved during 5 days. Peptidase 3 can be preserved in 14 days. Conclusion The preservation and transportation conditions of the standard substances must be controlled strictly during genetic quality evaluation of laboratory animals.

NF-κB Expression and Partial Behavioral Changes in Welfare Impaired Rats Caused by Restrain-stress

2018, 35(06):  36. 

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Objective To study the behavioral and NF-κB expression changes in welfare impaired rats caused by restrain-stress. Method Wrap restraint stress (WRS) were administrated by using tubular container within male SD rats (± 200 g). At time points of 0, 1, 3, 7, 10 and 14 days after WRS administration, open filed test and elevated o-maze test were employed to analyze the physical behavior changes of SD rats. NF-κB p65 mRNA transcription levels and protein expression in all SD rats were examined as well. Result Significant expression enhancements were observed both in NF-κB p65 mRNA transcription and NF-κB p65 protein expression at the 3rd day after WRS administration compared with pre-procedure (P<0.05), but no behavior changes could be found. At the 7th day after WRS administration, significant decline (P<0.05) in physical activity capacity and activity intension were noticed. All SD rats exhibited symptoms of depression and dread. Conclusion Changes in NF-κB mRNA and protein expression may prior to behavioral changes in welfare impaired rats.

Study on the Characteristics of Murine Model with Food Butter Induced Hyperlipidemia

2018, 35(06):  41. 

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Objective To establish a murine model with the butter induced hyperlipidemia, and to observe and analyze its characteristics. Method Twenty littermate male C57BL/6 mice were randomly divided into 2 groups with 10 mice in each group. Mice in the control group were fed the normal diet (ND) without the butter,and the others were fed the high fat diet with butter (HFD). The experiment lasted for 120 days.The changes of body weight and blood lipid were monitored over time; Organ weight and the histopathological changes in the liver were measured at the end of the experiment. Result At day 120, (1) compared with the control group, the body weight, serum total cholesterol (TC) and low density lipoprotein (LDL) were significantly increased in the HFD group (P<0.001), while the level of serum total triglyceride (TG) was not distinctly changed; (2) the weight coefficient of abdomen fat and liver in the HFD group was significantly increased (P<0.001); and (3) the liver of the HFD group appeared the severe fatty degeneration and inflammatory infiltration. Conclusion Hypercholesterolemia characteristics with the high TC without the change of TG in serum exist in murine model by the butter induced hyperlipidemias, suggesting that the milk butter in diet plays an important role in animal hypercholesterolemia model.

Subcutaneous Transplanted Tumor Model of 615 Mice Gastric Cancer with Pleural Puncture Needle and OB Anastomosis Glue

2018, 35(06):  47. 

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Objective A subcutaneous xenograft model of 615 mice was established by using pleural puncture needle and OB anastomosis glue. Method Mouse Forestomach Carcinoma Cell (MFC) was cultured under aseptic conditions to obtain a cell suspension with a concentration of 1×107 cells/mL. The suspension was inoculated into the right armpit of the mouse to establish a model of subcutaneous xenograft tumor of gastric cancer. The 615 mice subcutaneous xenograft model was built through puncture and transplation pleural needle method and by blocking three times in vivo passage. At the same time, a control group(without the tumor block) was used. The tumorigenesis of the mice was evaluated based on the appearance morphology and HE staining. Result Model mice lost weight. The model group grew faster since the 12th day and the model success rate was 100%. In terms of appearance morphology, tumor formation rate of 10 mice in the model group was 100%；The average tumor weight (7.7160 ± 0.2937)g; The tumor rate of the control group was 0, thus the difference between the model group and the control group was significant(P<0.01).Pathological result show the typical morphological characteristics of gastric cancer tissue in that the nodules in the model group were larger in size, irregular in surface, round in nucleus, ovate or irregularly shaped. Besides,nucleolus did not appear and mitotic cells were more obvious. Conclusion The 615 mice gastric cancer subcutaneous xenograft model, built with pleural puncture needle combined with OB anastomosis gel is characterized by high success rate, high efficiency and convenience, thus it is worth further research and promotion.

Establishment of Experimental Allergic Encephalomyelitis Model in C57BL/6 Mice

2018, 35(06):  52. 

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The establishment of an Experimental Allergic Encephalomyelitis(EAE) model in C57BL/6 mice provides a better understanding to the mechanism of multiple sclerosis(MS) and a development of new drugs with good effects for MS treatment. Our study applied ten C57BL/6 mice to immunize with both MOG53-58 and Mycobacterium tuberculosis (MT) in incomplete Freund’s adjuvant(IFA). And then they were treated with 0.2μg/mice Pertussis toxin (PTX) at day0 and day2 by i.p. injection to induce EAE model. Ten C57BL/6 mice were just treated with normal saline as control group. Histological change in central nervous were investigated by H&E staining, immunohistochemistry assay and Luxol fast blue(LFB)/Periodic Acid-Schiff (PAS) double staining. The result comprise the mice in EAE group started to develop clinical symptoms from 10th day post immunization, and all mice developed EAE disease. But no mice in control group developed EAE disease. Histological analysis showed abundant inflammatory cells filtrated into brain and spinal cord. Thus, the established EAE model is stable with high incidence, and this model could be used to study the mechanism of MS and screen the drugs for the prevention and treatment of MS.

The Sensitivity of Coxsackievirus A16 in Different Cells

2018, 35(06):  57. 

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Objective In order to isolate enterovirus CVA16, we selected two kinds of cells and screened out the most sensitive cells of enterovirus CVA16. Method Twenty samples were identified as positive for CVA16 by nucleic acid test. The cell sensitivity was compared by comparing the isolation effect of RD and VERO cells. Result Nine of the twenty samples were identified as CVA16, six strains were isolated from RD cells and eight strains were isolated from VERO cells. Conclusion The sensitivity of CVA16 to VERO cells is higher than that of RD cells. In the experiment, the VERO cells were selected.

Culture and Identification of Thoracic Aortic Vascular Endothelial Cells Isolated from Tree Shrews

2018, 35(06):  62. 

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Objective To study the isolation and culture of cultured vascular endothelial cells (VECs) from tree shrews (Tupaia belangeri), and to identify the cultured cells, and establish an in vitro model for the study of new tree shrews. Method The thoracic aorta was isolated from the tree shrews by tissue culture and trypsin digestion, culturing in complete culture medium, and cells morphology was observed by inverted microscopy, identified by Ⅷ factor Immunofluorescence assay. Result Tissue culture method was used to separate the cultured vascular endothelial cells, 3 d cells adherent, 7 d after the cells from the tissue block in a large number of long, cells were confluent to monolayer at 15 days. The cells were digested by enzyme digestion and adhered at 12 h, and grew at 3-7 days. The cells were treated with microscopic observation and showed that the cells were typical paving stone. The two cells were isolated and cultured in vitro. The cell growth was good and the positive expression was confirmed by Immunofluorescence assay using the vascular endothelial cell marker molecule Ⅷ. Conclusion VECs were successfully isolated from the tree shrews, and the method of isolation for tree shrews’ intestinal epithelial cells was established, which makes it available for VECs-related study.

Effects of Morroniside on the Expression of Cell-cycle Proteins in Hippocampus of Rats with Focal Cerebral Ischemia/Reperfusion Injury

2018, 35(06):  66. 

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Objective To explore the effects of morroniside on cell cycle-related proteins in a rat model of focal cerebral ischemia-reperfusion. Method After the model of middle cerebral artery occlusion (MCAO) was established, Sprague-Dawley rats were randomly divided into sham group, model group, morroniside low dose (30 mg/kg) group, morroniside middle dose (90 mg/kg) group and morroniside high dose (270 mg/kg) group. Western blot analysis was used to measure the expression of CyclinD1 and Cdk6 in ischemic ipsilateral hippocampus. Result Compared with sham group, the expression of CyclinD1 was increased obviously in vehicle-treated ischemic-reperfusion rat. Compared with model group, the expression of CyclinD1 was significantly reduced by treatment of morroniside at dose of 90 mg/kg and 270 mg/kg after ischemia-reperfusion. Compared with sham group, the expression of CDK6 was increased obviously in vehicle-treated ischemic-reperfusion rat. Compared with model group, the expression of CDK6 was significantly reduced by treatment of morroniside at dose of 90 mg/kg and 270 mg/kg after ischemia-reperfusion. Conclusion Morroniside could inhibit the inappropriate activation of cell cycle-related proteins and play neuroprotective effects by downregulating the expression of CyclinD1 and Cdk6 after cerebral ischemia-reperfusion.

How to Get a Higher Success Rate of Adventitious Virus Agent Test in the Mouse and Suckling Mouse Detection Method

2018, 35(06):  70. 

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The safety of biological products has attracted more and more attention because of its widespread clinical application. According to the regulations of the state, adventitious virus agents of biological products which applied in the clinic should be detected referring the 2015 China Pharmacopeia. However, the mice and the neonatal mice inspection method are hard to achieve successfully leading to the false-negative or the false-positive result . Hence, this article aims at generalizing the critical point of the mice and the neonatal mice inspection method and providing the efficient and practical operating method to get a higher success rate of adventitious virus agent test on the basic of the rich experience of the Guangdong medical laboratory animal center.

Brief Introduction of American Veterinary Education and Laboratory Animal Specialty Training

2018, 35(06):  73. 

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The veterinary education in the United States of America is widely agreed to be the most advanced. The degree for veterinary medicine is accredited as a professional doctorate degree by the Ministry of Education of the US. To become a veterinarian in the U.S., one needs to meet the strict requirements for admission, graduation and practice entrance examinations. There has already been a sophisticated system post the doctorate education of veterinary medicine in the U.S., including internship trainings, residency trainings and certifications. This article illustrated the processes before the admission of the colleges of veterinary medicine, and the postdoctoral specialty training and certification with the example of laboratory animal veterinarians.

Correlation Factors and Evaluation Criteria of OA Animal Model

2018, 35(06):  77. 

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This review focuses on the related factors and evaluation criteria of osteoarthritis in animal models, so as to seek a better research plan and experimental method for the treatment of osteoarthritis.

Preparation and Evaluation of Amblyopia Animal Model

2018, 35(06):  81. 

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Amblyopia is a common eye disease in children，which is a great problem for children’s life. It is important to establish an ideal animal model to study amblyopia. At present，the main method of making animal model of amblyopia are：suture of eyelid，cutting of rectus muscle，feeding in dark environment，and retinal detachment. This article reviewes the establishment of models，including animals， method, characteristics and discussed the frequently used models. This review will provide help for researchers in research design of animal models of amblyopia．