Objective To evaluate the effect of mild hypothermia combined with hydrogen-rich saline on cerebral injury after cardiac arrest and resuscitation in rats.Methods Healthy male Sprague-Dawley rats,aged 7-8 weeks,weighing 280-320 g,were divided into 5 groups (n=33 each) using a random number table method:sham operation group (group S),cardiac arrest and resuscitation group (group CAR),hydrogen-rich saline group (group H2),mild hypothermia group (group MH),and mild hypothermia plus hydrogen-rich saline group (group MH+H2).Cardiac arrest was induced with transoesophageal cardiac pacing followed by cardiopulmonary resuscitation to establish the cerebral injury model.Hydrogen-rich saline 5 ml/kg was intraperitoneally injected immediately after return of spontaneous circulation (ROSC) in H2 and MH+H2 groups,while the equal volume of normal saline was given instead in the other groups.The body temperature of rats was cooled down to 32-34℃ within 15 min starting from the time point immediately after ROSC and maintained for 4 h in MH and MH+H2 groups.Fifteen rats were selected at 24 h after ROSC to assess the neurological function score (NDS).Eighteen rats in each group were sacrificed at 24 h after ROSC,and brains were removed for microscopic examination of the pathological changes in hippocampal CA1 region after hematoxylin and eosin staining and for determination of pyramidal cell count and expression of glucose-regulated protein 78 (GRP78),C/EBP-homologous protein (CHOP),caspase-12,caspase-3,Bcl-2 and Bax in hippocampal CA1 region (by Western blot).Results Compared with group S,the NDS was significantly decreased,the pyramidal cell count was reduced,the expression of GRP78,CHOP,caspase-12,caspase-3 and Bax was up-regulated,and the expression of Bcl-2 was down-regulated in the other four groups (P＜0.05).Compared with group CA-R,the NDS and pyramidal cell count were significantly increased,the expression of GRP78 and Bcl-2 was up-regulated,and the expression of CHOP,caspase-12,caspase-3 and Bax was down-regulated in H2,MH and MH+H2 groups (P＜0.05).Compared with group H2 or group MH,the NDS and pyramidal cell count were significantly increased,the expression of caspase-3 and Bax was down-regulated,the expression of Bcl-2 was up-regulated (P＜0.05),and no significant change was found in the expression of GRP78,CHOP and caspase-12 in group MH+H2 (P＞ 0.05).Conclusion Combination of mild hypothermia and hydrogen-rich saline offers enhanced efficacy in reducing cerebral injury after cardiac arrest and resuscitation over mild hypothermia or hydrogen-rich saline alone in rats.

Objective To explore the immune-microenvironment of the retinas at different stages of retinal degeneration in Royal College of Surgeon (RCS) rats. Methods RCS-rdy--P+(RCS) rats at early stage (P20), middle stage (P40) and late stage (P60) were involved,12 rats at each post-natal day,RCS-rdy+-P+rats severed as control. Relative concentrations of rat cytokines in rat retina homogenate were detected by using Bio-Plex Suspension Array System. Relative expressions of interleukin-2 (IL-2),C-C motif ligand 2 (CCL2),chemokine (C-X-C motif) ligand 9 (CXCL9),CXCL10,CXCL11 and interferon-γ(IFN-γ) mRNA in rat retina were analyzed by real-time PCR. Expressions of IFN-γ and immune cells surface marker CD4,CD8 and CD161 in the retinas were detected by immunohistochemical staining. Percentage of IFN-γ positive T lymphocytes and natural killer(NK) cells in rat retina were analyzed by flow cytometry. The concentrations of IFN-γ in rat retina homogenate were evaluated by enzyme-linked immunosorbent assay ( ELISA ). The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission. Results Lymphocytes related cytokines and chemokines mRNA expression levels in the RCS rat retinas showed increase trends with the extension of time. The expression levels of IL-2,CCL2,CXCL9,CXCL10,CXCL11 and IFN-γ mRNA in P60 RCS rat retinas were significantly increased than those in the P20 RCS rat retinas and the control rat retinas (all at P<0.05).The positive rates of CD4,CD8 and CD161 cells in the retinas of P60 RCS rats was (9.09±0.89)%, (18.77±0.38)% and (9.41±0.38)% ,respectively. The proportion of IFN-γ positive cells in the retinas of P60 RCS rats was (8.29±0.27)%,which was significantly higher than that of the control rats ([0.28±0.02]%),with a significant difference between them (t=29.03,P=0.00). CD4+,CD8+and CD161+lymphocytes were mainly distributed in the retinas of P60 RCS rats, and the expressions of IFN-γ were co-located with lymphocyte surface markers. There were significant differences in the concentrations of IFN-γ in the retinas of RCS rats and control rats at different day ages (Fgroup=16.49,P<0.01; Ftime=21.05,P<0.01),the concentration of IFN-γ in retinas of P60 RCS rats was significantly higher than that of P20 RCS rats, P40 RCS rats and control rats, and the differences were statistically significant ( all at P<0.05). Conclusions Along with the process of retinal degeneration,immune privilege balance in the retinas is disrupted, the expressions of lymphocytes related chemokines and cytokines are elevated. Lymphocytes infiltration and activation are appeared in the retina highly activated at the late stage of RP, leading to the significant up-regulation of inflammatory cytokine IFN-γ in microenvironment, which indicates that lymphocytes mediated immune response may take part in retinal degeneration.

Objective To acquire the prevalence and molecular identification data on Syphacia muris and provide reference for the revision of national standard. Methods 923 batches of 5199 SPF animals ( including one batch of 5 monkeys, 3 batches of 25 mini?pigs, 28 batches of 55 rabbits, 13 batches of 248 hamsters, 37 batches of 198 guinea pigs, 93 batches of 459 rats, 742 batches of 4179 mice, 5 batches of 25 chickens and one batch of 5 ducks) and 145 batches of 1389 clean animals ( including one batch of 3 rabbits, 4 batches of 31 hamsters, 16 batches of 157 guinea pigs, 32 batches of 268 rats and 92 batches of 930 mice ) came from 50 different manufactures in China. Direct microscopy real?time dynamic video recording techniques in combination with morphological identification method were applied to screen the Syphacia muris infestation. A multiple polymerase chain reaction ( multiple?PCR ) testing of the isolate based on amplification of the conserved portions of the Syphacia muris internal transcribed spacer (ITS), 28S ribosomal RNA (28S rRNA), NADH dehydrogenase subunits 1 (nad1) and cytochrome c oxidase subunit 1 (cox1) genes, and the molecular sequencing of the multiple?PCR amplicons was used to confirm the Syphacia muris infection. Results Syphacia muris eggs, larvae and adults were detected by using direct microscopy real?time dynamic video recording technique. Syphacia muris were detected based on the morphology and size of ovum, larvae, and female and male adult worms. Multiple?PCR and sequencing were performed to identify ITS, 28S rRNA, nad1 and cox1 genes of DNA extracted from the single egg, larva and adult parasite Syphacia muris. This approach allowed the specific identification with no amplicon being amplified from heterogeneous DNA samples, and sequencing confirmed the identity of the amplified sequences. Molecular characterization by multiple?PCR amplification and sequencing of the ITS, 28S rRNA, nad1 and cox1 genes demonstrated the presence of Syphacia muris. Multiple?PCR followed by sequencing confirmed 285 of 5199 SPF and 135 of 1389 clean animal samples classified as positive by using direct microscopy real?time dynamic video recording technique in the study as containing Syphacia muris?specific DNA. Comparison of the partial sequences of the ITS, 28S rRNA, nad1 and cox1 genes revealed 100% similarity amongst Syphacia muris from different animals. The prevalence of Syphacia muris infection in SPF and clean animals were 5?5% (285/5199) and 9?7% (135/1389), respectively. Conclusions Direct microscopy real?time dynamic video recording technique, multiple?PCR and sequencing can be used to rapidly detect and accurately identify Syphacia muris. The zoonotic nature of Syphacia muris can be regard as a public health alter, hence the good quality control of animal has an important role in protecting human health and safeguarding people safety. This is the first molecular identification and infection investigation of Syphacia muris in SPF and clean animals in China.

Objective To analyze and evaluate the population genetic quality of 3 subbreeds of China Agricultural University miniature pigs in Beijing.Method According to the local standard DB11/T828.3 -2011, 25 pairs of microsatellite primers were used in 3 subbreeds of China Agricultural University miniature pigs, and software Popgen32 was used to process the data.Results 24 microsatellite loci shared 130, 122 and 138 alleles in the China Agricultural University miniature pigs I, II, III, respectively. The average heterozygosity was 0.6759, 0.5967 and 0.6779, respectively, while the average polymorphism information content ( PIC) was 0.6344, 0.5540 and 0.6403, respectively. The genetic distance between China Agricultural University miniature pig II and III was 0.4251, while the genetic distance between China Agricultural University miniature pig I and II was 0.2084.Conclusions In the 3 subbreeds, China Agricultural University miniature pigs II and III have genetic stability and genetic diversity, and both of which satisfy with the genetic characteristics of closed colony laboratory animal.

Objective To evaluate the therapeutic effects of methamphetamine (MA) dependence and the repairment of DA neuronal function by SPECT corpus striatum DAT visual digital neural molecular imaging techniques.Methods 25 MA dependent patients (BPRS score ≥ 35) were treated by self-designed treatment program for more than 6 months.The clinical therapeutic effects were scored with reducing rate of BPRS.MA dependent patients were examined by SPECT corpus striatum DAT imaging before and after treatment,while healthy volunteers were examined only once.The SPECT corpus striatum DAT images were analyzed visually and quantitatively.Results The reducing rate of BPRS showed that the total effective rate was 80.0％.Visual analysis of SPECT corpus striatum DAT images showed that the distribution of DAT in the corpus striatum was regionally reduced or defected in various degrees before treatment,and was significantly increased after treatment.Quantitative analysis showed that the bilateral striatal V ((19.26 ± 2.85) cm3),m((20.22±2.99) g) and Ra(4.78±0.79) ％) of MA dependent patients were significantly lower compared with those of the healthy volunteers(respectively (35.39±4.42) cm3,(37.16±4.64) g and (7.93± 0.86) ％) (all P＜ 0.01) before treatment and were significantly improved (P＜ 0.01) after treatment (V:(22.80±4.28) cm3,m:(23.93± 4.49) g and Ra:(5.64 ± 0.99) ％) with a 76.0％ corpus striatum DAT improvement rate.However,the bilateral striatal V,m and Ra of MA dependent patients after treatment were still lower than those of the healthy volunteers (P＜0.01).There was no significant difference between the striatal DAT improvement rate and the BPRS reduction rate (P＞ 0.05).Conclusion SPECT corpus striatum DAT visual digital neural molecular imaging techniques are reliable in the evaluation of the treatment programs for MA dependence and the repair of DA neuronal function.

Objective Giardia lamblia is an important pathogen of zoonosis giardiasis , it poses a potential threat to the quality of SPF (specific pathogen-free) laboratory animals cannot be ignored.The aim of this study is to establish the method of rapid diagnosis of Giardia lamblia, and analyze the test results of 516 batches form 17 manufactures.Methods Direct microscopy, Giemsa-fast staining and multiplex polymerase chain reaction (multiplex PCR) were applied to detect Giardia lamblia.Results Numerous of Giardia lamblia trophozoites and cysts were detected in SPF laboratory animals by using direct microscopy and Giemsa-fast staining, and multiplex PCR were performed to identify 18S rDNA,β-giardin, TPI and GDH genes of DNA extracted from these trophozoites and cysts identified Giardia lamblia.Direct microscopy, Giemsa-fast staining, and multiplex PCR methods can be used to detect Giardia lamblia.Of the 2562 SPF laboratory animals studied, 22.9%(586/2562) were positive for Giardia lamblia.Conclusions Direct microscopy , Giemsa-fast staining , and multiplex PCR were effective techniques with high sensitivity and specificity for rapid diagnosis of Giardia lambliain.It is not satisfactory that the results of Giardia lamblia examination in 516 batches form 17 manufactures failed to meet the requirements 100%.

To monitor and analyze Cricetulus migratorius fungal diversity ,60 adult Cricetulus migratorius brought from Xinjiang region of China were dissected after being euthanized and the specimens were collected .Fungal diversity research was carried out by TaqMan MGB probe real-time fluorescence quantitative PCR ,the ribosome cloning sequencing ,and fungal cul-ture identification techniques .The 60 fungal isolates were characterized from Cricetulus migratorius ,including Candida albi-cans ,Trichosporonasahii ,Aspergillus fumigatus ,Aspergillusniger ,Aspergillussydowii ,Aspergillus japonicus ,Asper-gillus ustus ,Aspergillus versicolor ,Penicillium chrysogenum ,Paecilomyces variotii ,Penicillium aurantiogriseum ,Neuros-pora sitophila ,Neurospora intermedia ,and Cladosporium cladosporioides .Many of them associated with grey hamster as zoonotic pathogens .The results showed that the most dominant fungal group was Aspergillus ,and Penicillium followed by it . These fungi were susceptible to nystatin ,clotrimazole and voriconazole .It’s indicated that application of TaqMan MGB probe real-time fluorescence quantitative PCR ,the ribosome cloning sequencing ,and fungal culture identification techniques could ef-fectively analyze Cricetulus migratorius .The results could provide a scientific basis for Cricetulus migratorius microbiological monitoring and quality standard establishment in China .Overall ,the findings of the present study constitute ,to the authors’ knowledge ,the first extensive report on the diversity of fungal flora associated with Cricetulus migratorius .

Objective To diagnosis tumor transplanted nude mice Strongyloidiasis .Methods Postmortem microscopic examination of the tumor transplanted nude mice detected Strongyloides stercoralis for morphological identification and double polymerase chain reaction ( PCR ) assay for molecular diagnosis of Strongyloides stercoralis infection in tumor transplanted nude mice .Results Presence of numerous S .stercoralis in autopsy in tumor transplanted nude mice samples preliminary determined the diagnosis of strongyloidiasis .Confirmed diagnosis of Strongyloides stercoralis infection by double PCR detection of specific DNA in tumor transplanted nude mice samples .Conclusion The most important clue to prevent such serious consequences is early diagnosis .Tumor transplanted recipients and donors should be screened for parasitic infections including strongyloidiasis .To the authors ’ knowledge , this study is the first extensive report on diagnosis tumor transplanted nude mice Strongyloidiasis .

Objective According to the basic theory of Yin and Yang,acute urticaria should be treated with divergent medicine instead of convergence medicine.In order to prove the theory,we carry out the quantifiable and repeatability traditional Chinese medicine screening tests in acute urticaria patients,with double controls of experimental group (divergent Chinese medicine),control group (convergence Chinese medicine)end instrumental quality-control standard substance MEBE.Methods Germany MORA-Super bio-resonance instrument (medical mode) as adopted as objective index.32 cases of acute phase urticaria were selected and treated with divergent or convergence Chinese traditional medicine,to observe the results whether conform to traditional Chinese medicine basic theory of Yin and Yang (That is whether the acute phase urticaria should be treated with divergent Chinese medicine).Results Through the 32 cases repeat experiments,22 out of 23 kinds of Divergent Chinese medicine have been screened out in experimental group; while none has been screened out in 12 kinds of convergence Chinese medicine in control group.There was significant difference between two groups (P＜0.01).The result was entirely consistent with TCM basic theory of opposite relationship between yin and yang.The top five divergent Chinese medicines were divaricate saposhnikovia root,cicada slough,catnip,shrub chastetree rruit,and lily magnolia.Conclusion Treated acute urticaria with divergent Chinese medicine conforms to the basic theory of traditional Chinese medicine.

Objective To develop a TaqMan MGB probe-based,sensitive and specific real-time fluorescence quantitative PCR assay for rapid detection of Helicobacter hepaticus.Methods Primers and probes specific toflaB gene of Helicobacter hepaticus were designed.A TaqMan MGB probe-based,real-time fluorescence quantitative PCR was established.The specificity,sensitivity and stability of the assay were assassed.Then,the established TaqMan MGB probe real-time fluorescence quantitative PCR assay was applied to detect Helicobacter hepaticus in 1081 clinical specimens during 2008-2011,compared with bacterial isolation and culture method and conventional PCR assay.Results The specificity of this established TaqMan MGB probe-based real-time fluorescence quantitative PCR was high and there were no cross-reactivity with Helicobacter pylori,Campylobacter jejuni,Clostridium piliforme,Pasteurella pneumotropica,Escherichia coli,Pseudomonas aeruginosa.The detection limits was 8.3 copies.The correlation coefficient and slope value of standard curve were 0.999 and -3.227,respectively and the efficiency of TaqMan MGB-based probe realtime fluorescence quantitative PCR assay was 100％.The TaqMan MGB-based probe real-time fluorescence quantitative PCR and conventional PCR were preformed to detect Helicobacter hepaticus in 1081 clinical specimens,a total of 86 specimens were positive for Helicobacter hepaticus.However,there was only 4 specimens were positive by bacteria isolation and culture method.The results showed that TaqMan MGB -based probe real-time fluorescence quantitative PCR for Helicobacter hepaticas was more sensitive than bacteria isolation and culture method,and it could detect Helicobacter hepaticus DNA from clinical specimens directly,and detection time is only 2 hours.Conclusion The TaqMan MGB-based probe real-time fluorescence quantitative PCR assay was a reliable,specific,sensitive and useful tool for rapid detection of Helicobacter hepaticus.