Abstract:Objective To establish a gata4 gene knockout zebrafish model of congenital heart disease, and construct transcription activator-like effector nuclease (TALEN) vectors targeting gata4 gene. Method We construct TALEN vectors targeting zabrafish gata4 gene using unit assembly method and the in vitro-transcribed TALEN mRNAs were microinjected into one-cell stage zebrafish embryos. The efficiency of TALEN was identified by injected embryos, and mutations of zebrafish were screened and confirmed the different types through PCR and enzyme digestion.Results We successfully constructed correct targeting vectors by enzyme digestion and sequencing, and the gene knockout efficiency was 35.18%. We screened the mutant zebrafish and confirmed different types of gata4 gene mutations.Conclusions A gata4 knockout zebrafish model is successfully established, it can provide a good animal model for further research of congenital heart diseases.

Abstract:Objective To observe the effect of a traditional Chinese medicine, BuyangHuanwu Decoction, on the ethology in rat models of post-stroke depression (PSD).Methods Sixty male SD rats, after open-field ethology scoring, were randomly divided into five groups (12 rats in each group): sham operation group, cerebral ischemic stroke group, post-stroke depression (PSD) group, BuyangHuanwu Decoction group, and fluoxetine group. The rat PSD models were established with separately breeding and chronic unpredictable moderate stress (CUMS) after middle cerebral artery occlusion. Behavior changes of the rats were detected by body weight measurement, neurological scoring, sugar consumption test and open-field test. Results Compared with the sham operation group and cerebral ischemic stroke group, the body weight,sugar consumption and open-field spontaneous activity in the PSD model group were decreased, and the neurological function deficits became more severe (P<0.01 or P<0.05). Compared with the PSD group, Both BuyangHuanwu Decoction group and fluoxetine group got significantly higher scores of body weight,sugar consumption and spontaneous activity,and improved neurological function deficits(P<0.01 or P<0.05).Conclusions PSD rats exhibit abnormal behavior.BuyangHuanwu Decoction can significantly improve the behavioral impairment of PSD rats. The antidepressant effect of BuyangHuanwu Decoction is comparable to that of fluoxetine in the treatment of PSD rats.

Abstract:Objective To observe the contribution of acute normovolemic hemodilution (ANH) in experiment of cryopreserved dog limb replantation. Methods Sixteen healthy Beagle dogs (male:female=1:1) were divided into two groups. Dogs in the experiment group (Group B) received ANH in the limb replantation, and dogs in the control group (Group A) received the same amount of lactate Ringer's solution intravenously during the surgical operation. We recorded and compared the hemodynamic indexes, HB, HCT, the resuscitation time, the first rising head time, the first standing time and the first eating and drinking time between the two groups. Results (1) During the operation, both PaO₂ and PCO₂ in the two groups were normal, as well as the breathing rate. The heart rate in the group B was lower than that in the group A. Before blood transfusion, there was no statistically significant difference in HB and HCT between the two groups, but after transfusion they were significantly higher in the group B than in the group A. (2) The resuscitation time, the first rising head time, the first standing time and the first eating and drinking time of the group B were all better than those in the group A. Conclusions In cryopreserved dog limb replantation experiments, acute normovolemic hemodilution is helpful to improve the general condition and facilitate the recovery of animals after limb replantation.

Abstract:Objective To obtain the basic growth parameters of a self-established F1 hybrid CB6F1 C57-ras transgenic mouse model, and to provide basic information for commercialization of this mouse model. Methods F1 hybrid mice (CB6F1) were produced by crossing C57-ras heterozygous transgenic (c-Ha-ras^+/-) male mice and wild-type BALB/cJ female mice. The average litter size, weaning rate, sex ratio, growth performance and C57-ras transgenic positive rate were recorded and analyzed. Results The average litter size was eight, weaning rate was 90%, and sex ratio was approximately in accordance with prediction. The average body weight of newborn mice was 1.73±0.05 g. The average body weight of 10-week old c-Ha-ras transgenic female and male mice in CB6F1 background was 24.38±1.74 g and 29.42±1.72g, respectively, which had a significant difference (P<0.01). The c-Ha-ras transgenic positive rate was 46.9%. which was in accordance with genetic rules. Conclusions The F1 hybrid mice (CB6F1) produced by crossing C57-ras heterozygous transgenic (c-Ha-ras +/-) male mice and wild-type BALB/cJ female mice show normal growth performance and development characteristics, and it can be used for large-scale commercial supply.

Abstract:Objective To explore the protective effect of 17β-estradiol on traumatic brain injury in rats. Methods A total of 45 adult male SD rats were divided into 3 groups using the random digit table, 15 rats in each group: the control group only exposed but not injured the brain, the injury group received traumatic brain injury (TBI) by Feeney's method, and the treatment group received the same handling with injury group, and pretreatment with 17β-estradiol peritoneal injection, 1 mg/kg per day for one week. The other two groups were given the same volume of castor oil. At 6 h, 24 h and 48 h after injury, the brain tissue water content, malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured. Results At 6 h, 24 h and 48 h after injury, the levels of brain tissue water content in the injury group and treatment group were significantly higher than those in the control group (P<0.05). At 6 h after injury, the brain tissue water contents in the injury group and treatment group were (99.83±0.40)% and (99.53±0.41)%, respectively, with a non-significant difference between the two groups (P>0.05). At 24 h and 48 h after injury, the brain tissue water contents in the injury group was (105.17±0.43)% and (107.54±0.39)%, in the treatment group was (103.26±0.42)% and (105.89±0.43)%, respectively, showing a significant difference between the two groups (P<0.05). At 6 h after injury, the levels of MDA in the injury group and treatment group increased and maintained at a higher level, and the levels of SOD decreased. Compared with the control group, there had a significant difference (P<0.05). The levels of MDA and SOD in the injury group and treatment group at 6 h after injury had a non-significant difference (P>0.05). But at 24 h and 48 h after injury, the levels of MDA in the treatment group were significantly lower than those in the injury group [(130.39±7.02) μmol/g vs. (149.41±8.25) μmol/g, (125.41±6.59) μmol/g vs. (157.72±8.93) μmol/L], and the levels of SOD in the treatment group were significantly higher than thoset in the injury group [(88.46±7.17)U/g vs. (80.10±4.87)U/mg,(97.31±7.89)U/g vs. (84.29±6.13)U/g], with a significant difference (P<0.05) between the two groups. Conclusions 17β-estradiol has a protective effect on traumatic brain injury.

Abstract:Objective The aim of this experiment was to study the improving effects of a ginsenoside hydrolysis product DS-1227 on scopolamine-induced learning and memory impairment in mice. Methods Sixty healthy 5-6-week old male ICR mice (body weight 22±2 g) were randomly divided into control group, model group, three DS-1227 groups (25 mg/kg, 50 mg/kg, 100 mg/kg), and positive control group (0.3 mg/kg). Fourteen days after oral administration of DS-1227, an open-field test was conduct to determine the mouse locomotor activity. Fifteen days after oral administration of DS-1227, all experimental animals were intraperitoneally administered scopolamine (0.75 mg/kg) and the mice of control group received the same volume of saline. In addition to scopolamine, the mice of positive control group received intraperitoneal injection of physostigmine in a dose of 0.3 mg/kg. Twenty minutes after completion of all the drug administration, object recognition test and Morris water maze test were conducted to evaluate the learning and memory abilities of the mice. Results DS-1227 had no significant effect on locomotor activity of the mice. Scopolamine obviously decreased the discrimination indexes in object recognition test, and prolonged the escape latency of water maze place navigation test. While 25 mg/kg, 50 mg/kg, 100 mg/kg of DS-1227 increased the discrimination indexes and decreased the escape latency of place navigation in the mice. Conclusion DS-1227 can improve the learning and memory impairment induced by scopolamine in mice.

Abstract:Objective To compare the Bama minipig and Juema minipig models of high altitude multi-organ dysfunction syndrome. Methods Six plateau-origin Juema minipigs and plain-origin Bama minpigs in each group received intravenous infusion of 0.35 mg/kg lipopolysaccharide (LPS), respectively. Blood samples were taken at 0 h, 3 h, 6 h, 12 h, 24 h, 48 h and 72 h after LPS infusion. Routine blood test was performed, blood CK, AST, ALT, TBIL, CRE were assayed, and histopathological examination of the lung tissues was performed at 24 h, 48 h after LPS infusion. Results The mortality of Bama minipigs was 33.3%, higher than that of 16.7% of Juema minipigs. The trend of physiological and biochemical changes was similar, but was milder in the Juema minipigs than in Bama minipigs. The lung injuries of the Bama minipigs at 24 h and 48 h were more severe than those in the Juema minipigs. Conclusions Both Bama and Juema minipig models of high altitude multi-organ dysfunction syndrome can be successfully established. Juema minipig models can be more closely and safely established, due to its own plateau biological properties, and avoid the influence by extrinsic injurious effects of plateau environment.

Abstract:Objective To explore the effect of high fat diet on serum biochemical parameters and histopathology of main organs in Mongolian gerbils. Methods Forty-eight healthy adult male Mongolian gerbils were randomly and equally divided into model and normal groups.The gerbils in the model group were fed with high fat diet while the normal group with standard diet. Eight gerbils in each group were killed at the end of 4th,8th and 16th week,respectively,and the body weight,serum levels of Glu,TG,CHOL,HDL-C, LDL-C, UA, CREA,BUN,TBil,TP,ALB,ALT,AST and AMS were determined. The histopathological changes of main organs were observed. Results Compared with the normal group,the blood lipid of the model gerbils was significantly increased,the liver function was impaired,the blood uric acid level was higher, and the blood glucose was decreased at the end of 16th week. The AMS was increased at the end of 16th week,but the renal function showed no significant changes. The liver tissue of the model group gradually showed steatosis,inflammation,fibrosis and cirrhosis, accompanied by splenomegalia. The lung tissue and myocardium showed fatty degeneration and obvious damages in the later period,the pancreatic islets were enlarged and the amount of endocrine cells was increased,and the small intestine and kidney didn't show any distinct changes. Conclusions A gerbil models of hyperlipidemia and nonalcoholic steatohepatitis and cirrhosis can be well established by high fat diet feeding,and may serve as good models for research of hyperlipidemia-related hyperuricemia, and lung and myocardial damages.

Abstract:Objective To explore the effects of the Chinese medicine, modified Erchen decoction, on the serum lipid spectrum of ApoE^-/- mice, and to explore its possible anti-atherosclerotic mechanism. Methods Forty-four male 7-8-week old ApoE^-/- mice were used in this experiment. ApoE^-/- mouse models of atherosclerosis were generated by high-cholesterol diet for 4 weeks. And then, they were given simvastatin or modified Erchen decoction by gavage. The body weight of mice was recorded every week, The mice were sacrificed after treated with the drugs for 8 weeks continuously, and the plasma lipid was determined by enzymatic method. The aortic valves and arches were stained with oil red O to depict atherosclerotic plaques and liver structural changes of the mice were examined by pathology. Results Modified Erchen decoction lowered plasma lipid(including TCHOL and LDL-C)significantly (P<0.01). The body weight was increased in the mice of all groups, but it was more pronounced in the mice of model group than in the blank and modified Erchen decoction groups. The serum CHOL and LDL-C levels were significantly lowered in the modified Erchen decoction group (P<0.01). The area of atherosclerotic plaques in the aortic wall was significantly reduced in the mice of modified Erchen decoction group as shown by oil red O staining. The pathological changes of hepatocytes were less severe and the structure of hepatic lobules was better preserved in the mice of modified Erchen decoction group. Conclusions The Chinese medicin modified Erchen decoction can effectively reduce serum lipids, regulate lipid metabolism, and ameliorate the process of atherosclerosis in ApoE^-/- mice.

Abstract:Objective To explore the effects of Chinese herbal compound, Tengmei decoction, on type II collagen-induced arthritis (CIA) in rats, and to examine the changes of arthritis index (AI), limb swelling, joint tissue inflammatory infiltration, and the effects on immune-inflammatory factors. Methods Sprague-Dawley rat models of arthritis were successfully established by intradermal injection of type II collagen and Freund's complete adjuvant. The model rats were randomly divided into model group, positive drug group, and high- and low-dose Chinese medicine groups, 6 rats in each group. The intervention and treatment period was 12 weeks. To measure weekly the anteroposterior and transverse diameters of the rear ankles and wrists, the transverse diameter of the claw foot palm pad, the thickness and the highest point width of hind limb plantar joint swelling, and to evaluate the integrated scores of joints and limbs swelling using a vernier caliper. Results ① Compared with the normal group, the total arthritis scores and hind limbs AI scores of the model group were significantly increased (P<0.05 or P<0.01). The left forelimb AI scores were significantly increased during 10-12 weeks (P<0.05). The anteroposterior and transverse diameters of the left hind limb, the thickness of the highest point measurement of the left hind foot pad metatarsal were significantly increased (P<0.05 or P<0.01) in different time periods between 1-12 weeks. Compared with the model group, the total scores and the left hind limb joints AI scores of the high- and low-dose drug groups were decreased after 6 weeks (P<0.05). ② Compared with the normal control group, levels of mRNA transcription and protein expression of IL-6 and TNF-α were significantly up-regulated (P<0.01) in the model group. Compared with the model group, the levels of mRNA transcription and the expression of IL-6 and TNF-α proteins were significantly down-regulated in the positive group and Chinese medicine groups (P<0.01). ③ Histological examination showed that the low-dose TCM significantly improved the CIA synovial hyperplasia and inflammatory cell infiltration. Conclusions The molecular mechanism of Chinese herbal compound Tengmei decotion in improving joint pathological injury of CIA rat models may be related to its inhibitory effect on the high expression of immune-inflammatory factors in the synovial tissue of CIA rats.

Abstract:Objective To observe the protective effect of the traditional Chinese medicine prescription, Jiu Nao Yi Zhi water extract, on human neuroblastoma SH-SY5Y cell line, its effect on expression of insulin signal transduction pathway, and to explore the related mechanisms. Methods SH-SY5Y cells cultured in vitro, were divided into control group, Jiu Nao Yi Zhi No.1 prescription group and No.3 prescription group. The doses were 0.0625 mg/mL, 0.125 mg/mL, 0.25 mg/mL, 0.5 mg/mL and 1 mg/mL. The thiazolyl blue (MTT) metabolic rate of each group was determined. The dose of 0.125 mg/mL was chosen for cell immunofluorescence analysis, and to observe the expression of insulin receptor substrates-1(IRS-1), cAMP response element binding protein (CREB), and the factors of insulin signal transduction pathway. Results Compared with the control group, MTT metabolic rates of the Jiu Nao Yi Zhi groups were significantly increased (P<0.05), and the cell morphology was much better in those groups, cell body more plump, well-adherent and neurite extensions were observed. The expressions of IRS-1 and CREB were higher than that in the control group.Conclusions The traditional Chinese medicine prescription Jiu Nao Yi Zhi water extract can protect neurons by promoting nerve cell growth, and improving the expression of IRS-1 and CREB, the factors of insulin signal transduction pathway.

Abstract:Objective To isolate and identify the rat amniotic cells and to explore their stem cell characteristics, so as to find a new cell source for stem cell transplantation. Methods Rat amnion tissue was mechanically separated from 18-18.5-day old SPF pregnant rats and 0.25% trysin-EDTA digestion was used to obtain amniotic cells. The isolated cells were cultured with DMEM D-glucose added with 10 μg/L EGF. Flow cytometry was used to detect the mesenchyme stem cellsurface markers. Neural stem cell surface markers of the rat amniotic cells were detected using immunofluorescence staining. Results The rat amniotic membrane tissue was separated and mesenchymal stem cells were cultured successfully. The cells were ameboid-shaped and showed adherent growth, and can stably proliferate in short term. After culture, the cells expressed stem cell markers e.g. Oct-4 and Sox-2, mesenchymal stem cell markers e.g. vimentin, embryonic stem cell markers e.g. SSEA-4, neural stem cell marker e.g. nestin, and could also express neurotrophic factors, such as BDNF and NGF. Conclusions Rat amniotic cells express mesenchymal stem cells markers, neural stem cell markers and neurotrophic factors, therefore, provide an experiment basis for further research and application of rat amniotic cells in stem cell transplantation.

Abstract:Objective To establish a method for isolation of cynomolgus monkey umbilical cord mesenchymal stem cells.Methods Fresh cynomolgus monkey umbilical cord was directly minced into pasty fine pieces, and the pieces were cultured in tissue flask with DMEM/F12 medium supplemented with 10% fetal bovine serum.The morphological characteristics of the resulting cells were examined, and their expression of mesenchymal cell surface markers were analyzed by flow cytometry.The multidifferentiation potential was examined in vitro, too.Results The fibroblast-like cells were successfully isolated from the fresh umbilical cord by an adherent culture procedure.These adherent cells expressed mesenchymal markers including CD29, CD44, and CD90, and also could be induced to differentiate into adipocytes, osteoblasts and chondrocytes.Conclusion Mesenchymal stem cells can be isolated from fresh cynomolgus monkey umbilical cord by using an adherent culture procedure.

Abstract:Alcohol is closely related to the occurrence and prognosis of cerebral ischemia. Researches on the relationship between alcohol and cerebral ischemia using different animal models draw different conclusions. This paper summarizes the common alcohol intervention methods in making animal models of cerebral ischemia, and evaluates the advantages and disadvantages of these models to provide reference for the future research.

Abstract:Parabiosis is an experimental animal model in medical studies. This kind of animal models can realize the exchange of blood and body fluid between the two united animals very well, and makes it possible to observe all influences of these exchange in different aspects, ranging from blood to immunity. Over the past few years, based on parabiosis studies, heterochronic parabiosis animals are preferred to be used as conjoint models and to provide experimental basis for researches in cancer, endocrine, transplantation, neurology and stomatology and so on.

Abstract:Objective To collect more than 200 kinds of species totaling more than 26,000 existing laboratory animals data worldwide to establish a largest and most comprehensive database of laboratory animal strains, based on the existing laboratory animal strain databases and related publications or references. Methods Based on MySQL database software to set up medium sized databases, and build search interface, input interface and database interface. Results In this work, a laboratory animal strain database was established, including data of 21596 mouse strains, 2062 rat strains, 13 monkey strains, 2 hamster strains, 5 dog strains, and 5 rabbit strains. We will gradually further improve the data to achieve regularly updated database of animal strains in the future. Discussions The establishment of this database will facilitate scientific researchers in our country to access and use domestic and foreign resources of laboratory animal strains, and provide links to introduce laboratory animal strain resources from European, American and other developed countries.