Table of Content

25 October 2018, Volume 38 Issue 5

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Cyclosporin A on Cardioprotective Effect of Sevoflurane Postconditioning under High Glucose Concentration in Rat

YU Jin, ZHAN Hai-ting, CHENG Hu, LI Yu-qian, ZHENG Hong

2018, 38(5):  329-335.  DOI: 10.3969/j.issn.1674-5817.2018.05.002

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Objective To evaluate whether cyclosporin A (CsA) restore the cardioprotective effect of sevoflurane postconditioning (SPostC) against hypoxia /reoxygenation (H/R) injury in rat under high concentration of glucose through inhibiting mitochondrial permeability transition pore opening and maintaining mitochondrial morphology. Methods Primary cultures of neonatal rat cardiomyocytes in low and high concentrations of glucose for 48 h were subjected to HR (3 hr hypoxia followed by 3 hr reoxygenation), treated with SPostC before perfusion or CsA before gypoxia, or combined use of these two interventions were performed before hypoxia. Cell death, lactate dehydrogenase (LDH) level, mitochondrial morphology and mitochondrial membrane potential were measured after H/R injury. Results Both SPostC and CsA decreased cell death, LDH level, increased mitochondrial interconnectivity and mitochondrial membrane potential following H/R (P<0.05), combination of these two interventions did not further enhance the protective effects. High concentration of glucose (35 mmol/L) eliminated the cardioprotective effect mentioned above. As compared with high glucose group, SPostC or CsA, or combined use of these two interventions did not affect the cell death, LDH level, mitochondrial interconnectivity and mitochondrial membrane potential following H/R (P<0.05). Conclusion Both SPostC and CsA can protect cardiomyocytes against H/R injury through maintaining mitochondrial morphology. High concentration of glucose eliminates the cardioprotective effect, and CsA can not restore the cardioprotective effect of SPostC under high concentration of glucose.



The Expression of Myelin Related Inhibitory Factor in Microenvironment after Spinal Cord Injury in SD Rats

JIA Xu-feng, Long Miao, JI Yong, HUANG Guang-ping, ZHOU Yu, FENG Da-xiong

2018, 38(5):  336-342.  DOI: 10.3969/j.issn.1674-5817.2018.05.003

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Objective To observe the expression of myelin related inhibitory factor in Sprague-Dawley (SD) rats at different time points in different injury potential energy of Allen's model and spinal cord total cut model with the microenvironment. Methods One hundred and twenty five adult female healthy SD rats were selected and randomly divided into 5 groups with 25 in each, group A (sham operation group), group B [(20 g×5 cm, injury potential energy 100 gram-cm force (gfc)], group C (20 g×10 cm, injury potential energy 200 gcf) , group D (20 g×15 cm, injury potential energy 300 gcf); group E (total spinal cord resection group).The expression of myelin related inhibitory factors of neurite outgrowth inhibitor-A (Nogo-A), the myelin associated glycoprotein(MAG), and the oligodendrocyte myelin glycoprotein (OMgp) were analyzed by Western blotting after spinal cord injury (SCI) at 1 d, 5 d, 7 d, 14 d and 28 d after successful modeling. Results Twenty-four cases (16.67%) of SD rats were died in the whole experiment. The expression of Nogo-A, MAG and OMgp was observed at the first day after SCI in group B, C, D and E. The expression of Nogo-A and MAG reached the peak at the 7th day and returned to the levels of sham operation group on the 28th day. The expression of OMgp in myelin was observed at the first day after SCI, and reached the peak at day 5, and returned to the levels of sham operation group on the 28th day. Conclusion The Nogo-A, MAG and OMgp are highly expressed after SCI and those indexes are reached the peak 5-7 days after SCI.



Visualized Detection of Ectromelia Virus by Loop-Mediated Isothermal Amplification Assay

ZHOU Jie, TAO Ling-yun, ZHAO Li-juan, HU Jian-hua, GAO Cheng

2018, 38(5):  343-349.  DOI: 10.3969/j.issn.1674-5817.2018.05.004

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Objective To established a loop-mediated isothermal amplification (LAMP) assay for rapid visual detection of ectromelia virus (ECTV). Methods According to the published GenBank sequences (NC_004105.1), 9 pairs of primers were designed targeting the conserved region of ECTV. The amplification was detected with LAMP Real Time Turbidimeter LA-302, and the visudlized detection method of ECTV was established. Meanwhile, the amplified products were colored by fluorescence detection reagent after completion of the reaction, so that the amplification could be detected with naked eyes. Then, methodological evaluation of the LAMP was tested, and the samples were tested for artificial infection. Results The method of LAMP shows a highly efficient amplification for ECTV viral target gene which performed at 63 ℃ for 60 min by the LAMP Real Time Turbidimeter LA-302. The detection limit was 530 fg/µL, was 10³ times higher than that of PCR, and no cross-reaction with other RNA and DNA of viruses from mice. The results of ECTV LAMP reaction visualized the tube directly with naked eyes by the addition of fluorescence detection reagent are consistent with the results detected by Tubidimeter real-time. Conclusion The established LAMP detection method for ECTV is rapid, specificity, high sensitivity, and be easy of operation under simple conditions, which may be suitable for rapid detection of ECTV.



Identification of the Target miR-191 for the Biological Detection of Toxoplasma Gondii

WEN Fu-li, ZHENG He-ping, Dang Yuan, Xue Lai-en, ZHAO Dong-yue

2018, 38(5):  350-355.  DOI: 10.3969/j.issn.1674-5817.2018.05.005

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Objective To establish the quantitative detection method of microRNAs (miRNAs) probe of Toxoplasma gondii. Methods High-throughput sequencing technology was used to screen out the high-level and non-significant potential targets between different strains of Toxoplasma gondii, and the expression and diagnostic efficiency of peripheral blood at different time points (0 d, 1 d, 3 d, 5 d, 7 d) were studied. Results High-throughput sequencing revealed that miR-252, miR-3641, miR-509-5p, miR-1285, miR-140, miR128-1, miR-191 and miR-3426 had potential as biological detection targets for Toxoplasma gondii. The miR-191 diagnostic efficiency of mice infected with Toxoplasma gondii RH strain and M49 strain showed that the specificity were 100%, and the sensitivity were 85% and 95%, respectively. Nanoscale gold probes were prepared by miR-191. The quantitative analysis of target miR-191 in blood of rats infected with Toxoplasma gondii RH strain and ME49 strain at different time points showed that miR-191 expression was relatively stable within 7 days after infection.The infection models of mice with Toxoplasma RH strain, Toxoplasma ME49 strain, Plasmodium bergi, Plasmodium yoelii, Cryptosporidium, mouse hepatitis virus and Staphylococcus aureus were made respectively. Compared with those of in control group, the miR-191 was highly expressed in mice infected with RH and ME49 strains of Toxoplasma gondii and no expression was found in mouse models infected with bacteria, virus and other parasites. Conclusions The miR-191 of Toxoplasma gondii has the application value as a detection target.



Comparison of Traditional Ultrasound with Speckle Tracking Imaging Technique by Myocardial Infarction Model in Mice

LU Jin, YAN Guo-feng, ZHOU Jin, XIAO Zhong-biao, ZHU Yin-qiu, ZHU Lian, WANG Jing, LI Yao, ZUO Yong

2018, 38(5):  356-364.  DOI: 10.3969/j.issn.1674-5817.2018.05.006

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Objective By analyzing and comparing conventional echocardiography (ECG) to speckle tracking imaging (STI) technique on mice myocardial infarction (MI) model, a more precise in vivo monitoring method is able to be demonstrated. Methods The left anterior descending artery (LAD) ligation (MI group) or sham operation (Sham group) were randomly performed in male C57BL/6 mice at 16 weeks age. Same ultrasound parameters were performed on both groups. Followed conventional ECG method to record ejection fraction (EF%) and fraction shortening (FS%) from left ventricle and utilized STI technique to analyze cardiac cycle velocity, displacement, strain and strain rate of each segment in both short axe and long axe. Compared the results with conventional ECG and STI technique. Results Conventional ECG is able to point out MI status in mice, but STI technique can give more information related to myocardial motor ability and mechanics properties. Thus, it's more sensitive and informative than conventional ECG. Conclusion STI technique can be used as a supplementary method for MI monitoring in mice that will improve the precision of evaluation.



A Minimally Invasive Method of Electrophysiological Examination and Arrhythmia Models in Rabbits

ZHOU Zhi-wen, ZHAO Li-fang, CAO Jia-qi, ZHENG Hong-chao, DING Yue-you, ZHU Fu, CHANG Wei

2018, 38(5):  365-371.  DOI: 10.3969/j.issn.1674-5817.2018.05.007

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Objective To explore a minimally invasive technique for electrophysiological examination and arrhythmia model by using rabbits. Methods After general anesthesia, rabbit's jugular vein was isolated. The tip of multipolar electrode was inserted slowly to the right atrium or right ventricle. Also rabbit's carotid artery was isolated, and 5 F arterial sheath was inserted in the isolated artery by using Seldinger's method. The tip of multipolar electrode was send to the left ventricle through the arterial sheath. The multipolar electrode was connected with the cardiac stimulation system and chest lead of electrocardiogram (ECG). The location of the tip of multipolar electrode can be detected by the endocardial and pacing ECG. After determining the threshold of stimulation for pacing, the electrodes were fixed, and electrophysiological program stimulation was performed. Arrhythmias can also be induced by electrophysiological program stimulation. Results Twenty rabbits were used in the study. The tip of multipolar electrode can be placed easily into the desirable place in the heart. A wave and V wave can be shown in the ECG clearly. The tip of multipolar electrode can be replaced to the desirable place according to the electrocardiogram signal of cardiac pacing without the help of X-ray. Through atrial stimulation program, it can be used to study the sinus-node recovery time, atrial refractory period, and forward conduction function of atrioventricular node. Atrial fibrillation and atrial flutter can be induced by atrial stimulation. Combination with the intravenous of acetylcholine, the induction rate of atrial fibrillation and atrial flutter can reach over 95%. Through ventricular stimulation program, the ventricular refractory period and atrioventricular junction reverse transmission function can be studied. Ventricular tachycardia and ventricular fibrillation can be induced by ventricular stimulation as well. Combination with the intravenous of isopropyl, the induction rate of ventricular tachycardia and ventricular fibrillation can reach over 90%. Conclusions Our newly established minimally invasive technique can be used for electrophysiological examination and arrhythmia model in rabbits without the help of X-ray.



Killing Effect Investigation to Schistosomula through In Vitro Co-Culturing with Four Kinds of Fibroblasts Culture Supernatant from Microtus fortis

WU Xia, CHENG Gang, LI Huan, LI Hai-xia, TANG Zhi-hong

2018, 38(5):  372-376.  DOI: 10.3969/j.issn.1674-5817.2018.05.008

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Objective To identify the kill effect of Microtus fortis culture supernatants against Schistosoma japonicum. Method Co-culture schistosomula with supernatants of wild Microtus fortis embryos, skin, lungs and peritoneal fibroblasts were collected at the logarithmic phase in this study. The death rate of schistosomula was observed and calculated in 96 h. We used 20% M. fortis serum as a positive control and DMEM medium as a blank control. Result After co-culture with lungs and peritoneal fibroblasts culture supernatant 72 h, schistosomula show death sign, including: shrinking skin, constriction. After 96 h, schistosomula dissolved, and the death state of the schistosomula was slightly different with the positive control group. The death rate is 8.3%±2.1%, 9.3%±1.38%, 17.2%±3.27% and 15.1%±4.35% respectively after 96h co-culture with Microtus fortis skin, embryos, peritoneal fibroblasts and lungs. Conclusion Killing effect of lungs and peritoneal fibroblasts culture supernatant of M. fortis was higher than that of the same treated skin and embryonic cell group in vitro (P<0.05).



Effect of He Dan Tablet on Chronic Non-alcohol Fatty Liver Disease Model Rat

XU Li-ying, LI Yan-shu, ZHENG Guo-an, WU Jin-hua, DING Qi, XIAO Xiao-hua

2018, 38(5):  382-386.  DOI: 10.3969/j.issn.1674-5817.2018.05.010

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Objective To explore the effect of He Dan tablets on chronic non-alcohol fatty liver disease (NAFLD) induced by high fat emulsion in rat. Methods The NAFLD rats model was made with intragastric administration of high fat emulsion and high glucose solution every day for 4 weeks. Then the model rats were intragastrically administrated with high fat emulsion and high glucose solution in the morning and He Dan tablets, Xuezhikang Capsule and Fenofibrate tablets solution in the afternoon for 4 weeks. At the end of experiment, the concentration of the serum alanine aminotransferase (ALT), triglyceride(TG), total cholesterol (TCH), low density lipoprotein cholesterol ( LDL-C), high density lipoprotein cholesterol (HDL-C) and the contents of reduced glutathione (GSH), malondialdehyde (MDA), TCH, TG in liver, and liver organ coefficient were measured. Results Compared with model group, the concentration of the serum ALT, TCH, LDL were significantly lower and serum HDL-C was significantly higher in 3.53 g/kg, 7.06 g/kg, 14.12 g/kg He Dan tablets groups. Moreover, the serum TG and liver MDA, TCH, TG were significantly reduced in 7.06 g/kg、14.12 g/kg He Dan tablets groups compared with model group. Besides, all the rats were examined with pathologic histology, the dose of 7.06 g/kg, 14.12 g/kg could reduce the hepatocyte swelling, steatosis, necrosis and degree of infiltration of inflammatory cells in chronic NAFLD rats. Conclusion He Dan tablets can improve the related indicators in serum and liver of NAFLD rats, and also can alleviate the hepatocyte swelling, steatosis, necrosis and degree of infiltration of inflammatory cells in NAFLD rats.



The Hypoglycemic Effect of Gardenia Yellow Monomers on Diabetic Mice

LI Yan-shu, XU Li-ying, ZHOU Yan-yan, DI Qi, LI Yan, LI Yu-yun, XIAO Xiao-hua

2018, 38(5):  387-389.  DOI: 10.3969/j.issn.1674-5817.2018.05.011

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Objective To explore the hypoglycemic effect of Gardenia yellow monomers on diabetic mice induced by alloxan monohydrate. Methods The effect of gardenia yellow monomers (crocin Ⅰ, crocin Ⅲ, crocetin) on blood sugar in diabetic mice with starch loading and sucrose loading was determined. Diabetic mice were prepared by intravenous injection of alloxan monohydrate. Results In comparing with model group, the blood sugar were significantly decreased in 100 mg/kg crocin Ⅰ, 100 mg/kg crocetin , 50 mg/kg crocin Ⅲ groups mice with starch loading. The blood sugar were significantly decreased in 100 mg/kg crocin Ⅰ, 50 mg/kg and 100 mg/kg crocetin , 100 mg/kg crocin Ⅲ groups mice with sucrose loading than the model mice. Conclusion Gardenia yellow monomers crocin Ⅰ, crocin and crocin Ⅲ all have hypoglycemic effect on starch loading and sucrose loading diabetic mice.

Preliminary Observation on Micro-electrolyzed Water as Drinking Water in SPF Animals

ZHANG Wei, TANG Bin, LI Fu-rong, ZHAO Yong

2018, 38(5):  394-398.  DOI: 10.3969/j.issn.1674-5817.2018.05.013

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Objective To monitor the bacteria in the pipe network and the water used in microbiological sterile water system in barrier breeding facilities, and to observe the effect of drinking micro-electrolysis sterile water on blood biochemical in SPF-grade SD rats. Methods From the period of 2015 to 2017, 6 outlet points of the micro-electrolysis water system were sampled and detected; 21-day-old SD 20 rats were divided into 2 groups, half male and half female, drinking micro-electrolyzed water and high pressure water respectively. After drinking Sterilized water 1 to 8 months, Blood was taken from the abdominal aorta to detect blood biochemical. Results The microelectrolytic water system has been detected for more than 2 years. Using the current detection technology, no bacteria have been detected in the micro-electrolyzed water and the pipe network. There was no difference in blood biochemical indicators after drinking 1 to 8 months of water in 2 groups. Conclusion The application of micro-electrolysis sterile water system in barrier breeding facilities has a continuous disinfection and bactericidal effect on the pipe network and the water, and can kept sterile for a long time. No effect on blood biochemical indicators were observed in 21-day-old SD rats drinking micro-electrolyzed water for 8 months .

Current Situation and Countermeasures of Laboratory Animal Welfare in Medical Research in China

LUO Xiao-quan, LI Long-xue, LI Shan-shan, LI Zhong-lian, XU Peng

2018, 38(5):  403-406.  DOI: 10.3969/j.issn.1674-5817.2018.05.015

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The results of medical research are usually obtained by means of effective animal experiments, and at the expense of laboratory animals themselves being hurt or suffering. How to improve the welfare of laboratory animals on the premise of ensuring the scientific and reliable results of animal experiments is a problem that every medical researcher should consider. This paper, starting from the present situation of animal welfare in China's medical scientific research, points out the differences between China and the developed countries in Europe and the United States, and analyzes the reasons for the lagging of animal welfare in scientific research and experiment in China. In order to promote the rapid improvement of the welfare level of experimental animals in China, the measures of speeding up the standards and rules of the laboratory animal welfare technology, strengthening the publicity and education of experimental animal welfare and establishing a scientific animal welfare ethics review system are proposed.