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A new model of rat artery transplantation

Lailong SUN ; Jianping DU ; Chaobing LIU ; Songlin ZHANG

Chinese Journal of Comparative Medicine.2017;27(5):99-102. doi:10.3969.j.issn.1671-7856.2017.05.021

Objective Allograft Vasculopathy(AV), which limits the function and long-term survival of transplanted organs, it is the most urgent problem need to be solved in the field of organ transplantation.Traditional vascular anastomosis requires excellent microsurgical techniques, it is difficult to carry out.To explore a simple and feasible AV model can provide ideas for clinical research.Methods We make vessel cannulas by using vein detained needles, pulling the thoracic aorta of donor rat in vessel cannula, both ends turned over and fixed,and then transplant it in recipient rat abdominal aorta.1, 4 and 8 weeks after transplantation, we take out of the transplanted arteries, HE stain, to observe the histomorphology and measurement the vascular intimal thickness.Results 38 of the 40 recipient rats healthy survival to the detection time points, the legs and tail can move freely, defecating and urinating normally.It is found that transplanted arteries concentric circles thickening, its histomorphology just like the AV by using HE staining.Conclutions The method of rats artery transplantation is simple and feasible, significantly shorten the time of abdominal aortic blocking, reduce surgical trauma, can improve the success rate and the reproducibility of operation.

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Establishment and application of a multiplex PCR method in Staphylococcus aureus、Pseudomonas aeruginosa and Klebsiella pneumoniae

Yanbo ZHU ; Long ZHENG ; Hongyu YOU ; Xuan WANG ; Xiaoliang LIANG ; Jianmin LIU ; Dongming ZHANG ; Weiguang LIAN ; Yanning LI ; Junxia WANG

Chinese Journal of Comparative Medicine.2017;27(5):94-98. doi:10.3969.j.issn.1671-7856.2017.05.020

Objective Aiming at detecting Staphylococcus aureus、Pseudomonas aeruginosa and Klebsiella pneumoniae in laboratory animals,the paper provides a rapid,sensitive and simple test method.Methods According to Staphylococcus aureus nuc gene,Pseudomonas aeruginosa LasI gene,Klebsiella pneumonia PhoE gene and general 16S rRNA gene, designed specific primers;Through the optimization of multiplex PCR primer concentrations and annealing temperature, the specificity and sensitivity of detection, establishing multiplex PCR system.Application of the PCR system test specimens of artificial infections and experiment animal feces is compared with traditional test method.Results Multiplex PCR amplification of Staphylococcus aureus (153 bp), Pseudomonas aeruginosa (600 bp) with Klebsiella pneumoniae (368 bp) and general (520 bp).The multiplex sensitivity for the purpose of 10pg, specificity of detection was not detected from other pathogens.Application of establishing multiplex PCR system to detect the artificial positive samples, and detect 1 Pseudomonas aeruginosa positive case in 76 fecals.Conclusions This paper established the multiplex PCR method which has the advantages of specific,sensitive,simple and rapid, and provides a reliable way for rapid test in laboratory animals microbiology.

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The role of miR-221 in the injury induced by hydrogen peroxide in rat myocardial cells

Lizhen FU ; Maoqin HUANG ; Zhiyong LAO ; Mengsheng XIAO ; Dekang ZHU

Chinese Journal of Comparative Medicine.2017;27(5):83-88. doi:10.3969.j.issn.1671-7856.2017.05.018

Objective To explore the role of miR-221 in the injury induced by hydrogen peroxide (H2O2) in rat myocardial cells (H9c2).Methods The viability of H9c2 cell induced by cell different concentrations of H2O2 was determined by MTT.The expression of miR-221 was detected by RT-PCR method.The miR-221 inhibitor and negative control were transferred into H9c2 cells by Lipofectamine 2000, then the cells were divided into normal control group, model control group (H2O2 group), negative control group (H2O2+ negative control group), inhibition group (H2O2+miR-221 inhibitor group).The cell viability was measured by MTT assay.Cell apoptosis was detected by acridine orange staining method.The expression of Bcl-2, Bax, phosphatase and tensin homolog deleted on chromosome ten (PTEN, p-protein kinase B (AKT) were assayed by Western Blot.Results 0,25,50,100,200,400 μmol/L H2O2 inhibited H9c2 cell activity gradually, of which 200 mol/L inhibition of cell viability moderate, so as a subsequent induction dose.Compared with normal control group, cell viability was decreased (P < 0.01), cell apoptotic rat was increased (P < 0.01), the expression of Bax and PTEN was upregulated (P < 0.01), the expression of Bcl-2 and p-AKT was downregulated (P < 0.01) in model control group and negative control group.Compared with model control group and negative control group, inhibition group proves the contrary.Conclusions Down-expression of miR-221 could significantly inhibit oxidative stress damage in H9c2 cells, which related to regulation of PTEN/AKT signal pathway.

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Study on the protective effect of Guanxinning Tablet on myocardial ischemia reperfusion injury in rats

Fei HU ; Yili RONG ; Keyan ZHU ; Hong LU ; Cheng CHEN ; Minli CHEN ; Yongming PAN

Chinese Journal of Comparative Medicine.2017;27(5):76-82. doi:10.3969.j.issn.1671-7856.2017.05.017

Objective To observe the effect of Guanxinning Tablet (GXNT) on myocardial infarction and cardiac autonomic nervous function in rats with myocardial ischemia reperfusion injury (MI/RI).Methods Seventy SD rats were divided into 7 groups randomly (n=10);the sham group, the MI/RI group, 75 mg/kg, 150 mg/kg, 300 mg/kg and 600 mg/kg GXNT groups and 300 mg/kg Compound Danshen Tablets (DST) group.All rats were administered orally for 7 days, and then the MI/RI model was made by ligating the left anterior descending branch of coronary artery in rats.The changes of electrocardiogram were recorded and the electrocardiogram of J points and heart rate variability (HRV) parameters were analyzed.At the end of reperfusion, the myocardial infarct size was measured by using Evans blue and tetrazolium chloride (TTC) double staining, and pathological changes of myocardium were observed by HE staining.The changes of serum lactate dehydrogenase (LDH), creatine kinase (CK), malondialdehyde (MDA) and nitric oxide (NO) levels were also detected.Results Compared with MI/RI group, GXNT and DST groups were significantly reduced myocardial infarct size and inhibited the rising of serum LDH and CK activities (P < 0.05, P < 0.01), and also reduced the total or average value of J point during reperfusion (P < 0.05, P < 0.01).Meanwhile, GXNT and DST groups were markedly increased HRV and serum NO level as well as decreased serum MDA content (P < 0.05, P < 0.01), and improved myocardial tissue pathology.Conclusions GXNT can reduce the myocardial infarction in rats with MI/RI, and also improve the cardiac autonomic nervous function.

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The effects of paclitaxel on the levels of CD28,CTLA-4 and BAFF in experimental autoimmune encephalomyelitis

Shujiang ZHANG ; Zuoxiao LI

Chinese Journal of Comparative Medicine.2017;27(5):71-75. doi:10.3969.j.issn.1671-7856.2017.05.016

Objective To discuss the effects of Paclitaxel(PTX) on levels of CD28 and CTLA-4,B lymphocyte stimulator(BAFF) in experimental autoimmune encephalomyelitis(EAE).Methods The 50 rats were divided into 5 groups by the random number table, 10 rats in each group,the doses of small group,Middle group, High group were 1 mg/kg,2 mg/kg,4 mg/kg by intraperitoneal injection for 10 consecutive days, the normal group and model group were injected 0.9% NS 2 mL,Using brain tissue score to estimate the neurological dysfunctions of rats.Using flow cytometry to detect the levels of CD28 and CTLA-4,using enzyme-linked immunosorbent (ELISA) to detect the levels of BAFF.Results The brain tissue score in PTX experimental groups were lower than model group,the comparative differences between groups were statistically significant(P < 0.01);The levels of CD28 in PTX groups were lower than EAE group,the comparative differences between groups were statistically significant(P < 0.01).The levels of CTLA-4 in PTX groups were higher than EAE group,the comparative differences between groups were statistically significant(P < 0.01);the content of BAFF in all PTX groups were lower than EAE control group.Conclusions PTX could decrease the brain tissue score,the mechanism may adjust the express of CD28、CTLA-4 in brain and the expression of BAFF.PTX may have preventive and therapeutic effects on EAE rats.

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Synergism and attenuation effects of Pini Pollen on cyclophosphamide

Ping GONG ; Tong SUN ; Xiaoli DAI ; Yukui MA

Chinese Journal of Comparative Medicine.2017;27(5):60-63. doi:10.3969.j.issn.1671-7856.2017.05.014

Objective To investigate the synergism and attenuation effects of Pini Pollen on cyclophosphamide chemotherapy in model mice.Methods The models of mouse were induced with S180 tumor and cyclophosphamide.The tumor inhibiting rates, hematology, Liver function, giant cell phagocytosis, hemolysin level and indexes of thymus and spleen were determined to evaluate the synergism and attenuation effects of Pini Pollen(150,300,600 mg/kg) on cyclophosphamide chemotherapy.Results Compared with cyclophosphamide chemotherapy group,the tumor inhibiting rates,white blood cell number,giant cell phagocytosis, hemolysin level and indexes of thymus and spleen were increased, at the same time liver function damage were alleviated significantly in cyclophosphamide plus Pini Pollen groups.Conclusions Pini Pollen has a remarkable synergism and attenuation effects on cyclophosphamide chemotherapy through its immune adjustment functions.

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Budesonide inhibits murine asthmatic airway inflammation through reduction IL-22 secretion

Minglei WANG ; Wenge WANG ; Junhong ZHANG ; Dawei ZHANG ; Kesong YAN

Chinese Journal of Comparative Medicine.2017;27(5):55-59. doi:10.3969.j.issn.1671-7856.2017.05.013

Objective To study the effect of Interleukin-22 (IL-22) on murine asthmatic airway inflammation and airway remodeling, observe the effect of budesonide on IL-22 of asthmatic mouse model, explore the mechanism of budesonide in the treatment of asthma.Methods Ovalbumin(OVA) was used as an allergen to sensitize and challenge the mice.24 female specific-free (SPF) BALB/c mice aged four weeks were randomly divided into 3 groups:control group, asthma group and budesonide treatment group (BUD group).For Histopathological Examination, HE staining was used to measure the inflammation scores, AB-PAS staining was used to measure the hyperplasia of goblet cells and mucin.Enzyme-linked immunosorbent assay(ELISA) was used to analyze IL-22 levels in bronchoalveolar lavage fluid (BALF).Quantitative Real-time PCR was performed to analyze the effects of budesonide on IL-22 mRNA levels in lung tissue.Results The inflammation scores of asthma group were elevated compared with the control group.An overall change towards less severe asthmatic airway inflammation by the end of the trial was observed in the BUD group.IL-22 levels in BALF were significant decreased after the treatment of budesonide, the mRNA levels of IL-22 were obviously decreased in BUD group, too.A significant positive correlation was observed between the mRNA levels of IL-22 and airway inflammation.Conclusions The increasing IL-22 secretion can lead to the occurrence of airway inflammation of asthma.Budesonide can inhibit the expression of IL-22, thereby Budesonide could inhibit the development of airway inflammation of asthma.

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Effects of intrauterine injection of ethanol on estrous cycle in mice

Yinhong ZHANG ; Chunfang WANG ; Min GUO ; Ruihu ZHANG ; Zhaoyang CHEN

Chinese Journal of Comparative Medicine.2017;27(5):51-54. doi:10.3969.j.issn.1671-7856.2017.05.012

Objective To investigate the effect of uterine cavity injection of absolute ethanol on estrous cycle in mice.Methods Twenty mice with regular estrous cycle were selected by vaginal exfoliated cells staining, and then were injected absolute ethanol into their uterine cavity.The estrous cycle was observed and recorded every day.Ten mice were selected randomly to observe the changes of second estrous cycle every two hours.Results After the injection of absolute ethanol into the uterine cavity of the experimental mice, the estrous cycle was greatly prolonged, and the various degree of disorder was observed in estrus and pre-estrus.Conclusions The disorder was observed in estrous cycle of mice after the injection of absolute ethanol, which maybe provides some references for clinical use of absolute ethanol.

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Validation of the silencing site of marmoset B2m gene at the cellular level

Yichen DENG ; Chen ZHANG ; Zhiguang XIANG ; Yongkang TENG ; Yunbo LIU

Chinese Journal of Comparative Medicine.2017;27(5):37-41. doi:10.3969.j.issn.1671-7856.2017.05.010

Objective To screen and determine the effective silencing targets of β2-microglobulin(B2m)gene at the cellular level in marmoset.Methods By homology comparison of the b2m gene in human and the B2m gene in marmoset, choose homology small hairpin RNA(shRNA)sequences targeting marmoset B2m gene were designed, We choose homology small hairpin RNA(shRNA)sequences targeting designed B2m gene to make homology analysis, and insert into lentivirus-based gene silencing constructs FUGW-TDT.The vectors were transfected into HEK293T cells induced by polyethylenimine(PEI).The suppression of B2m mRNA was detected by real-time PCR.Results Two gene-silencing sequences were screened that lied in 290~310 bp and 665~685 bp of the marmoset B2m mRNA, and have statistical significance in the silencing rate:(46.54±7.91)% (P < 0.05) and(83.22±4.37)%(P < 0.0001).Conclusions Two effective silencing target sequences are screened at cellular level, which can be further used in studies on gene silencing in marmoset.

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The study on the mechanism of platelet in the development of salt-sensitive hypertension via inflammation

Hongya MAO ; Yunpeng LIU ; Zihao WANG ; Xiaoliang JIANG ; Xing LIU ; Zhiwei YANG

Chinese Journal of Comparative Medicine.2017;27(5):23-30. doi:10.3969.j.issn.1671-7856.2017.05.008

Objective To investigate the role and mechanism of platelet in the development of salt-sensitive hypertension.Methods 25 Dahl salt-sensitive rats (Dahl SS) were divided into three groups: low-salt diet (0.12% NaCl, LS), high-salt diet (8%NaCl, HS) and high-salt diet + platelet inhibitor (8%NaCl+busulfan, HS+bus).Blood pressures were measured by tail-cuff method.After six weeks, animals were sacrificed.Platelet p-selectin expression, platelet cytosolic Ca2+ concentration, platelet-leukocyte aggregation (PLA) in peripheral blood, and immune cells infiltrated on aortic walls were assessed by flow cytometry, and serum IL-6 level was tested by ELISA in vivo.Platelets purified from SD rats were treated with normal salt (0.9%NaCl) and high salt (1.3%NaCl), then the cytosolic Ca2+ concentration and p-selectin expression of platelet were detected.Results We found that Dahl SS rats with high-salt diet, relative to low-salt diet, presented with high blood pressure and increased the ratio of platelet p-selectin expression, Ca2+ concentration.IL-6 level and PLA in peripheral blood, and the number of infiltrated immune cells on aortic walls were also significantly elevated in high-salt diet group.The whole events were ameliorated by the platelet inhibitor busulfan.Cytosolic Ca2+ concentration and p-selectin expression were also increased in purified platelets treated with high salt than those treated with low salt (P < 0.05).Conclusions Our findings suggest that high salt induced platelet activation with increased Ca2+ concentration may play an important role in the development of salt-sensitive hypertension via vascular inflammation.However, the detailed mechanisms of platelet activation and development of high blood pressure via inflammation induced by high salt intake remain to be determined.

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