The use of animals in research will be necessary for scientific advances in the basic and biomedical sciences for the foreseeable future.As we learn more about the ability of animals to experience pain,suffering,and distress,and particularly for mammals,it becomes the responsibility of scientists,institutions,animal caregivers,and veterinarians to seek ways to improve the lives of research animals and refine their care and use.Refinement is one of the three R's emphasized by Russell and Burch,and refers to modification of procedures to minimise the potential for pain,suffering and distress. It may also refer to procedures used to enhance animal comfort. This paper summarizes considerations for refinements in research animal.

Objective to develop a rotation monitoring device to monitor animal rotation during experiments.Methods Use serial port controls,USB and rotary encoder et al.modern computer cience,with the combination of traditionally experimental methods and the software designed in VC language.Results A mini animal rotation monitor was developed,which could continuously monitor animal rotation and intelligently display the rotaion.conclusion Statistic analysis of experimental data proofs the rationality and effectiveness of the apparatus.The rotational behavior monitor can play an importance role in research into the pathogenesis of Parkinson's Disease(PD)and screening the anti-PD drugs.

Objective To achieve the fusion expression of the entire human beta-defensin-3(hBD-3) gene. Method We synthesized two oligonucleotide primers accor ding to the codon preference of Escherichia coli. The gene was cloned into p GEX -4T-2 to establish the pGEX-4T-2-hBD-3 as the fusion expression vector by PCR. Transformed into E.coli strain DH5α, the express vector was induced an d ex pressed by IPTG. The fusion protein GST-hBD-3 was obtained by repeated cycles of freezing and thawing, cut by thrombin to attain the recombinant hBD-3 protei n. Result The result of the antibacterial peptide agarose diffu sion assay shows the antibacterial activity of the rhBD-3 against the S.aureu s exists, and it reaches 0.843U. Conclusion The fusion expr ession of the hBD-3 gene is successful.

This paper reported the history of avian flu outbreaks in China from 1996 to 2006, directed the characteristics of avian influenza outbreaks in recent years. China is one of the biggest poultry producers in the world and many migratory birds go through China, China faces high risk of pandemic bird flu. The continuing occurrence of sporadic human cases indicates that the virus is continuing to circulate in birds in some parts of the country. In the fighting again avian flu virus, China has developed some possible practice on precautionary and control measure. China enhances international cooperation and scientific involvement in the prevention and control of the avian flu. The State Council and Ministry of Agriculture issued the laws and regulations on avian flu. Wild birds may have played a role in getting domestic fowl and human beings infected with bird flu. Poultry manure is considered to be another key source of the spread routes of H5N1 virus, but the bio - security processing in free range poultry farming and waste products is very poor disposition in China. Epidemic control at grassroots level still needs to be strengthened.

Multiple sclerosis ( MS) is an autoimmune disease of the central nervous system ( CNS) characterized by demyelination and inflammation lesions.MS predominantly affects young adults with a high incidence of disability. However, the exact pathogenesis of MS is still not clear.Studies found that microglia polarization tending to pro-inflammatory M1-like state during the onset of MS, causing the M1/M2 ratio imbalance, forming pro-inflammatory microenvironment state, and which further leading to nervous tissue damage ultimately.Microglia polarization may be considered as the initiator of pathologic alterations by releasing pro-inflammatory cytokines and secondarily trigger the initial microglia response.Given the pivotal role of imbalanced microglia polarization in MS initiation, a critical review of microglia polarization is presented here, in order to elucidate the pathogenesis of MS and highlight the noteworthy candidate therapeutic targets for clinic treatment.

Objective To evaluate the feasibility and safety of transjugular liver biopsy( TJLB) by using the LABS 200 liver access and biopsy set ( Cook Inc, USA) .Methods Five minipigs were operated though TJLB puncture under the imaging guidance.The liver biopsies were analyzed by histological examination.Results Technical success of TJLB was achieved in all the 5 minipigs.No procedure-related complications occurred, and sufficient amount of specimen for histological examination was obtained in all cases.Conclusions Our preliminary results indicate that transjugular liver biopsy with the use of Cook LABS 200 liver access and biopsy set is clinically safe and feasible, and provide technical support for its clinical application.

Objective To establish a stable and fast method for primary culture of mouse cardiomyocytes. Methods Dishes were coated with polylysine firstly.A two-step approach was used to isolate and digest mouse cardiomyocytes cells (0.25%trypsin in 4°C overnight and 0.5 mg/mL to 1.0 mg/mL collagenase +5 mg/mL albumin collagen digestion liquid in 37°C for short-time digestion), then the cardiomyocytes were purified through differential adhesion for 70 min and 5-bromodeoxyuridine ( BrdU) .The cell morphology was observed under an inverted microscope. The survival rate of cardiacmyocytes was detected by trypan-blue staining and their purity was identified by α-actinin immunofluorescence staining.Results The cardiomyocytes were in good shape and pulsed spontaneously.The survival rate of the cardiomyocytes reached 98%and the purity was 95%.Conclusions This method described in this study is an ideal method for primary culture of mouse cardiomyocytes with a high survival rate and high purity.

Objective To investigate the accumulation of mutations and single nucleotide polymorphisms ( SNPs) in the displacement loop ( D-loop ) of mitochondrial DNA ( mtDNA ) might be associated with cancer risk and disease outcome.Methods We obtained cancerous and noncancerous liver tissues from 49 HBV-related HCC patients at the Fourth Hospital of Hebei Medical University.mtDNA of the liver tissues was extracted with Mitochondrial DNA Extraction Kit.Mutation and polymorphism were confirmed by repeated analysis.We assessed the prediction power of D-loop SNPs in hepatocellular carcinoma ( HCC) patients.Results No mutation in these HCC patients had prediction power for post-operational survival, whereas one SNP site ( nucleotide 150 C/T ) was identified by the log-rank test for statistically significant prediction of HCC survival.In an overall multivariate analysis, allele 150 was identified as an independent predictor of HCC outcome.The length of survival of patients with allele 150C was significantly shorter than that of patients with allele 150T (relative risk, 0.246;95% CI, 0.070–0.861; P=0.028).Conclusions The analysis of genetic polymorphisms in the mitochondrial D-loop helps to identify patient subgroups at high risk of a poor disease outcome.

Objective To decrease the p53 gene expression at cellular and animal levels in marmoset using RNA interference technique.Methods The shRNA interference sequences were designed and inserted into the adeno-associated virus vector plasmid after bioinformatics analysis.The plasmids were transfected into African green monkey kidney cos-7 cells.The suppression of p53 mRNA was detected by real-time PCR, and the changes of p53 protein expression were detected by Western bolt.The adeno-associated virus-8 was injected through the hind leg vein.The changes of p53 protein expression in the liver tissue was detected by Western blot and immunohistochemistry.Results We screened two RNA interference effective arget sequences.The expression of p53 mRNA was suppressed ( 82.7 ±8.1 )% and ( 80.7 ± 7.5)%, respectively (P<0.05), and the expression of p53 protein was decreased (77.3 ±11.5)% and (73.7 ± 10.7)%, respectively (P<0.05).The two marmosets after virus infection showed that there were virus distributions in the liver, testes, and neck detected by in vivo fluorescence imaging.The expression of p53 in the marmoset liver was detected by western blot, immunohistochemistry analysis showing no obvious changes.Conclusions In the present study, the decrease of P53 gene expression at cellular level is achieved, however, the liver P53 protein in the marmoset liver is not significantly changes.Further optimization of the way of infection is needed in the future.

Objective To develop a more convenient and stable method for assessment of drug sensitivity of prostate cancer based on real-time cell analysis system as a reference for clinical treatment.Methods Human prostate cancer VCaP, DU145, PC-3, PC-3M-2B4 and PC-3M-IE8 cells were chosen to detect the sensitivity to three drugs, docetaxel, cabazitaxel and abiraterone acetate.Serial dilutions of the three drugs were used to treat the cell culture for 24 hours.The drug-induced effects on the cell lines after an incubation of 24 hours were recorded by the real-time cell analysis system to determine the half maximal inhibitory concentration (IC50).Results Docetaxel showd IC50 of 8.81 nmol/L, 11.61 nmol/L, 1.78 nmol/L, 1.44 nmol/L, 8.69 nmol/L for VCaP, DU145, PC-3, PC-3M-2B4, PC-3M-IE8 cells, respectively.Cabazitaxel showed IC50 of 3.73 nmol/L, 3.96 nmol/L, 10.41 nmol/L, 5.43 nmol/L, and 7.37 nmol/L, respectively, for the five cell lines.Abiraterone acetate showed IC50 of 8.34 μmol/L, 8.60 μmol/L, 24.20 μmol/L, 8.59 μmol/L, and 13.21 μmol/L for the five cell lines.Conclusions PC-3M-2B4 and DU145, VCaP and PC-3 cells can be used as control for docetaxel, cabazitaxel and abiraterone acetate to establish cell models for the drug screening in vitro and to provide reference for clinical applications.