Abstract:Objective To compare the sequence and expression level of tyrosinase gene (TYR) among four rabbit strains covering a wide range in the extent of iris pigmentation and to verify the relationship between the tyrosinase gene and the iris colour of rabbits. Methods Exons of TYR were amplified and directly sequenced. The relative expression level of TYR in different strains of rabbits was analyzed by real-time quantitative PCR. ResultsThe albino strains Japanese White and Rex rabbits showed a specific nucleotide exchange leading to an amino acid exchange (T373K) at the last N-glycosylation site. The silence nucleotide exchange (A→T) at nt870 of the tyrosinase coding sequence was observed in both the white hair black eyes (WHBE) and Chinchilla rabbits. A non-strain-specific amino acid exchange (V31M) was revealed in WHBE and Rex rabbits. The gene expression level of TYR in dark iris rabbits was significantly higher than that in non-pigmented ones. All results were corrected by the household gene GAPDH. ConclusionsThe nucleotide exchange at nt 1118 (C→A) affect the function of tyrosinase, and the findings indicate that the gene expression level of TYR is related with the phenotype of iris color in rabbits．

Abstract:ObjectiveTo apply SYBR green I fluorescent quantitative real-time PCR (QPCR) in detecting 4 vertical transmission viruses of chicken and to evaluate its application value for the quality control of SPF chickens. MethodQuantitative real-time PCR technique was used to detect REV, ALV, ARV and CAV RNA, DNA or provirus DNA in 30 cloacal swab samples and 30 eggs of SPF chicken samples as well as 40 cloacal swab samples and 30 eggs of common chicken samples. Standard curve and melting curve analyses were conducted to obtain the quantitative result. ResultsOut of the SPF chicken samples, 2/30 (3.3%) were ALV-positive, and the three other viruses were negative. For the common chicken samples, 2/70 (2.9%) were REV-positive, and 10/70 (14.3%) were ALV-positive. ConclusionsThese QPCR assays show better sensitivity and specificity for detecting the 4 viruses in SPF chickens with a detection limit of 100 copies of target DNA per-reaction, and have a bright prospect in clinical application.

Abstract:Objective To investigate the influence of different week-age and the hormonal level on superovulation in Mongolian gerbils. Methods Eight groups of Mongolian gerbils at different age of 4-18 weeks were induced to superovulate by intraperitoneal injection of 10 IU PMSG/HCG, respectively. At 16-17 h after the last injection, the amounts of eggs of animals in different groups were counted and analyzed to determine the best week-age of superovulation. Superovulation was induced in the animals of best week-age by intraperitoneal injection of 5,0,15 IU PMSG/HCG to analyze the different amounts of eggs. ResultsCompared with other groups, the amount of eggs obtained in gerbils at age of 6 weeks is highest and there were significant differences among all groups (P<0.05). The amounts of eggs obtained in groups of different PMSG/HCG doses were significantly different and that of the 10 IU group was highest. ConclusionThe age of 6 weeks and the dose of 10 IU PMSG/HCG are the best ages and effective dose to induce superovulation in Mongolian gerbils.

Abstract:ObjectiveTo explore the effect of ginsenoside Rb₃ on chromosome aberration in Chinese hamster lung (CHL) cells. MethodsTo determine the IC₅₀ of ginsenoside Rb₃ in CHL cells, and to establish the range of doses according to the IC50 and induction of chromosome aberration. The results were judged according to the positive response standard. ResultsNegative response was found at 6 h and 24 h after the treatment with ginsenoside Rb₃ and at 6 h after the addition of S₉ mixture. ConclusionsUnder the conditions used in this study, ginsenoside Rb₃ does not induce chromosome aberration in CHL cells in vitro.

Abstract:ObjectiveTo compare the difference of two heart failure models established by constriction of aortic arch and abdominal aorta, and to explore an effective method to establish an animal model of heart failure. MethodsNinety healthy adult male Wistar rats were randomly divided into three groups. The rats in the group 1 had cervical operation to contract the aortic arch, the rats in the group 2 were operated on abdomen to contract the abdominal aorta, and the rats in the group C had surgery but no aorta constriction. All the rats were fed for 4 or 6 weeks. The heart function of the rats was detected with ultrasound and heart intubation. ResultsAt 4 weeks after constriction, ultrasonic results showed a significant increase of the left ventricular wall thickness and internal diameter of the left ventriclular cavity in the aortic arch constriction group, and not significantly increased in the abdominal aorta constriction group, compared with those of the control group. At 6 weeks after constriction, both the left ventriclular wall thickness and internal diameter of the left ventricle were significantly increased in the abdominal aorta constriction group, while only the internal diameter of the left ventricle was significantly increased and without change of the left ventriclular wall thickmess in the aortic arch constriction group. Hemodynamic detection showed that LVESP, LVEDP, LVDP and ±dp/dtmax in the aortic arch constriction group were significantly lower than those of the abdominal aorta constriction group at 6 weeks after contraction. ConclusionThe progression of heart failure caused by aortic arch constriction in the rats occurs obviously faster than that developed after abdominal aorta constricction.

Abstract:ObjectiveTo investigate the reference range of enzymes in urine samples of normal rats.MethodsFour-hour urine samples were collected from 95 normal rats. Urinary enzymes including NAG, ALP, γ-GT, LDH were analyzed using a biochemical analyzer. Reference ranges were shown with upper 95th percentile. ResultsThe reference ranges for female rats:NAG<5.3 U/L,ALP<110.8 U/L,γ-GT<753.0 U/L,LDH<32.6 U/L; for male:NAG<27.3 U/L,ALP<329.0 U/L,γ-GT<769.8 U/L,LDH<40.0 U/L. Reference ranges using the unit/g·Cre for female:NAG<28.9 U/g,ALP<723.5 U/g,γ-GT<1763.2 U/g,LDH<113.1 U/g; for male rats:NAG<84.3 U/g,ALP<680.2 U/g,γ-GT<2522.3 U/g,LDH<121.8 U/g.ConclusionThe reference ranges of urinary enzymes may provide background values for renal toxicity evaluation in the future.

Abstract:ObjectiveTo evaluate the value of MRI pre-scanning technique in stereotactic brain puncture in rats.MethodsTotally 18 SD rats were randomly divided into 2 groups (n=9):control group,in which the brain puncture was performed in strict accordance with the stereotactic brain atlas, and experimental group, in which MRI pre-scanning was performed and brain puncture was guided by the images. Rats in both groups were injected 2μL Gd-DTPA, taking caudate nucleus in the axial section which crosses the center of globus pallidus as target. Then MRI scanning was performed with three-dimensional magnetization-prepared rapid gradient-echo sequence to detect the enhancement of Gd-DTPA and compare the brain puncture accuracy between the two groups. ResultsThe number of rats which were located precisely was 8 in experimental group (88.89%), and 3 in the control group (66.67%). The difference between the two groups was statistically significant (P<0.05). The accuracy of experimental group was much higher than that of control group.ConclusionMRI pre-scanning can improve the accuracy of stereotactic brain puncture, and it is a necessary step to confirm accuracy and repeatability of the stereotactic operations.

Abstract:ObjectiveTo explore the mechanism of neuro-immuno-endocrine system of rats in sub-health state.MethodsThirty-six male SPF Wistar rats were randomly divided into six groups:multi-factor group (MF), multifactor treatment group (MFT), warm water swimming group (WS), sleep deficiency group (SD), purely constraint group (PC) and control group (C), six animals in each group. Respectively, using warm water swimming, dietary restrictions, sleep deprivation and constraint to establish sub-health rat models, and daily orally administered 300 mg/kg anti-aging tablet in the MFT group. At 5 days after modeling, the levels of serum 5-HT, DA, T3, T4, T, CORT, ACTH, IL-1β, IL-2, IL-6, IL-8, IFN-γ and T lymphocyte subsets were detected, and changes of T and B lymphocyte proliferation and NK cell activity in the spleen were also determined. Results(1) Compared with the control group, ① Serum 5-HT in the MF group was significantly reduced (P<0.05), while 5-HT in the WS and PC groups was significantly increased (P<0.01), and the serum DA content in the SD and PC groups was significantly decreased (P<0.05). At the same time, the serum T3,T4, CORT and ACTH contents in each sub-health model rats were significantly increased (P<0.01), T/C ratio was significantly decreased (P<0.01), and the serum T in the SD group was significantly lowered (P<0.05); ② Sserum IL-8 content in the MF group was significantly reduced (P<0.01), while the serum IL-1β, IL-2, IL-6, IL-8 and IFN-γ levels in the WS and PC groups, and IL-1β, IL-2 and IL-6 levels in the SD group were all significantly increased (P<0.05, P<0.01), and the lymphocyte proliferation and T lymphocyte subsets CD₃₊, CD₄₊, CD₄₊/CD₈₊ ratio and NK cell activity were significantly reduced in all sub-health model rats (P<0.05, P<0.01). (2) Compared with the MF group, the administration of anti-aging tablets induced significantly increased serum DA content in the MFT group (P<0.05), and significantly improved the T lymphocyte subsets CD₃₊, CD₄₊, CD₄₊/CD₈₊ ratio (P<0.01), and lowered serum CORT content. ConclusionsSub-health state can lead to imbalances of the rat HPA axis, excessive cortisol release, endocrine disorders, reduced lymphocyte function and immune inhibition, while anti-aging tablets have anti-stress effects, enhancing T cell immune function and ameliorating emotional and endocrine disorders, so as to achieve the purpose of improving the sub-healthy physique.

Abstract:ObjectiveTo establish a sheep model of multiple organ dysfunction syndrome (MODS) at high altitudes induced by lipopolysaccharide (LPS) and observe the pathological characteristics of organs. MethodsSixteen 1-year old sheep (male∶female = 1∶1) were randomly divided into moderate altitude group (1500 m) and high altitude group (3200 m). The MODS models were generated by intravenous infusion of LPS at a dose of 6 μg/kg. After 24 hours, the sheep were anesthetized, and samples of the lung, liver, heart, spleen, kidney and intestine tissues were taken and fixed in 10% formalin for histopathological examination. ResultsThe lung, liver, spleen and intestinal tissues had significant inflammatory changes in both MODS groups, and especially, the pathology scores of the lung and liver at high altitude MODS group were significantly higher than those in the moderate altitude MODS group. ConclusionsPathological damages of MODS at high altitude are more aggravated than those at moderate altitude.

Abstract:ObjectiveTo observe synaptic protein expressions in the brain of permanent middle cerebral artery occlusion (pMCAO) model rats on 2 and 7 after ischemic attack. Methods To establish a rat model of pMCAO. The rats with ischemia were randomly divided into 2-day group and 7-day group, taking rats with no occlusion as the sham group. On d 2 and d 7 post-operation, pathological changes in ischemic brain tissue were observed with H&E staining, and expression of synapsin-I, postsynaptic density protein 95 (PSD-95), α-synuclein were examined with immunohistochemical staining. ResultsCompared with the sham group, there were a large amount of neurons lost, and degeneration and necrosis of scattered neurons in the brain tissue in the model group. On the 2^nd day post-ischemia, in the model group, expressions of synapsin-I in the CA1 and CA3 regions and cortex were significantly reduced (P<0.05 or P<0.01), PSD-95 in CA1 region and cortex was significantly reduced (P<0.05 or P<0.01), while α-synuclein in CA1 region significantly accumulated (P<0.01). On the 7^th day after ischemia, in the model group expression of synapsin-I in CA1 region and cortex were still significantly reduced (P<0.01), PSD-95 in CA1 region and cortex also significantly reduced (P<0.05 or P<0.01), and α-synuclein in CA1, CA3 regions and cortex was significantly increased (P<0.05 or P<0.01). ConclusionsIn the brain tissue of pMCAO model rats, expression of synaptic proteins is significantly changed and varies along with the time-course of ischemia, probably, it might be related to synapse remodeling. The expression of synapse-associated proteins is closely related to the degree of ischemic injury.

Abstract:ObjectiveTo investigate a most suitable method for harvesting mouse embryonic fibroblasts (MEF)). Methods Three different protocols including tissue only, cell-trypsin digestion, and tissue-trypsin digestion were applied for harvesting and culturing MEF. The MEF growth speed and cell morphology were also compared and analyzed respectively among the 3 methods. ResultsThe cell proliferation was slow, cell size was small, but cells of other types were very few in the tissue only method, but the duration of purification was short. In the cell-trypsin digestion method, the cell proliferation was fast, cell size was somewhat large, but there were a little bit more cells of other types and the duration of purification was somehow longer. Finally, in the tissue-trypsin digestion method, the cell proliferation was very slow, cell size was very small, but there were also very few cells of other types. The duration of purification was very short. ConclusionsOur findings indicate that the most efficient method for collecting and culturing mouse embryonic fibroblasts is the protocol of tissue-trypsin digestion.

Abstract:ObjectiveTo develop a new method for rapid detection of canine distemper virus by reverse transcriptase loop-mediated isothermal amplification method (RT-LAMP). MethodAccording to the published GenBank sequence, four strips of primers specific recognizing NP gene of canine distemper virus were designed. We studied the optimal reaction conditions, specificity, sensitivity, and its visualized judging method of products. Then we used 63 clinical specimens to verify its efficiency in field detection. ResultsThe amplication can be performed at a constant temperature 60℃ within 1 hour. Specific primers did not recognize genes of other virus commonly infecting canine. The results of RT-LAMP could be easily observed by naked eyes after SYBR green I staining. In 63 specimens collected from Nanjing and Shangdong, 45 samples(71.4%)were positive, but only 40 samples (63.5%) were positive using RT-PCR. Conclusion The results of our study demonstrate a higher superiority of specificity, sensitivity, efficiency and convenience of the RT-LAMP method, and provide a new detection method for canine distemper virus.

Abstract:ObjectiveTo develop a novel fixator for beagles. Methods To devise a stainless steel frame according to the body size of Beagles, on which the dogs stay in a prone position on a canvas pad of the upper frame, their legs are bound to the lower frame, and their heads can move freely. ResultsConscious Beagles can be fixed on the fixator for a long time and this fixing method is suitable for drug infusion or repeated blood-taking. Conclusion We have successfully developed a simple and convenient fixator for Beagle dogs in laboratory experiments.

Abstract:Objective To develop an instrument that detects the behavior of mice in tail suspension during pharmaceutical experiments. Methods One pair of infrared sensors was used for signal conversion. A diode laser was used for positioning. A 51-series microcontroller was used to control the experimental setup and record the results. Results A microcontroller-based experimental instrument that can automatically detect the behavior of animals in tail suspension during pharmaceutical experiments was developed. ConclusionsOur experimental results with 30 mice indicate that the developed instrument provides an innovative tool for detecting the behavior of animals in tail suspension.

Abstract:Pseudomonas aeruginosa is a common zoonotic opportunistic pathogen, widely distributed in nature, is one of important pathogens causing experimental animal pollution and intra-hospital infection. Molecular typing is an important tool in pathogen epidemiology. It is very effective for determining the source of infection and ways to detect cross-contamination and epidemic strains. In this paper, we summarized the research progress of P. aeruginosa molecular typing methods.

Abstract:The silkworm with no unknown organisms living in or outside its body called gnotobiotic silikworm. The foster methods of gnotobiotic silikworm is much easier then that of mammal ones, and completely overcomed seasonal and regional breeding limits. Gnotobiotic silikworm lay the foundation for its wide application in laboratory animal science. With the development of modern biotechnology, study on the application of gnotobiotic silikworm as a bioreactor generator or genetic engineering animal has been gradually developed into the practical stage.

Abstract:Idiopathic pulmonary fibrosis is one of a group of chronic interstitial lung diseases of unknown causes, and is characterized by alveolar epithelial cell injury, abnormal proliferation of fibroblasts and excessive deposition of extracellular matrix. There are limited therapeutic options, poor prognosis, low long-term survival rate and is associated with increasing incidence since the pathogenic mechanisms are elusive. This demands a better understanding of the molecular mechanisms underlying the pathogenesis. It is most essential to know the onset and progression of this disease, especially in relevance to human health. In this paper, we reviewed the advances in research of idiopathic pulmonary fibrosis.