Abstract:Objective To investigate the effects of miR-182 mimics on cardiac function in type 1 diabetic mice and explore the mechanisms of action.Methods Forty 8-week-old C57 mice were randomly divided into 4 groups: control group (n=5), miR-182 mimics control groups group (n=5), type 1 diabetes group (n=15) and type 1 diabetes treated by miR-182 mimics (n=15). Eight weeks after modeling, the cardiac function was evaluated by a high-resolution ultrasonic imaging system for small animals (Vevo 2100). The samples of cardiac tissue was used to observe the ultrastructural changes by electron microscopy and to detect the expression levels of β-MHC, α-MHC, ANP, Col Ⅰ, Col Ⅲ and miR-182 mRNA.Results ① MiR-182 mimics improved the cadiac function of type 1 diabetic mice with an increase of left ventricular function and left ventricular short axis fractional shortening (P<0.01);② MiR-182 mimics significantly decreased the expression level of ANP, Col Ⅰ, Col Ⅲ protein and the ratio of β/α-MHC in cardiac tissue;③MiR-182 mimics induced ultrastructural changes of cardiac tissue in type 1 diabetic mice and the phenomenon of autophagy.Conclusions MiR-182 mimics may improve the cardiac function of type 1 diabetic mice, which is associated with decreased expression of hypertrophy-related gene ANP,β/α-MHC ratio and expression of Col Ⅰ, Col Ⅲ, reduction of mitochondrial damage and autophagy.

Abstract:Objective Maternal dietary modifications determine the susceptibility to metabolic diseases in adult life. However, whether maternal high-fat diet can induce early-onset aberrant glucose and lipid metabolism in offspring mice is less understood. Materials and Methods We examined the effects of feeding the pregnant mice with either a high fat diet or nornal chow diet throughout pregnancy and lactation on the glucose and lipid metabolism in their offsprings. Some glucose and lipid metabolism biomarkers and hepatic histology were analyzed. Results At weaning, for males of dams fed high-fat diet, they had heavier body weight, the blood glucose levels were significantly higher at 30 min (P<0.001) and 60 min (P<0.01) after intraperitoneal glucose administration, and the AUC was also significantly larger(P=0.0016) and HOMA-IR was higher(P<0.05). For females of dams fed high-fat diet, they had higher blood glucose levels at 30 min (P<0.01) after intraperitoneal glucose administration. No significant differences were observed of AUC and HOMA-IR. Both males and females of dams fed high-fat diet had elevated total cholesterol levels(P<0.0001 and P=0.0004, respectively). Triglyceride levels showed no significant differences between the two groups of males and females. The offsprings of high-fat diet group indicated hepatic steatosis and there were no differences between males and females. Conclusions Maternal high fat diet during pregnancy and lactation can induce early-onset aberrant glucose and lipid metabolism in offspring mice, and the male offsprings are more prone to metabolic changes, such as obesity, glucose intolerance and insulin resistance.

Abstract:Objective To explore the feasibility of establishing mini-pig model of type 1 diabetes by single intravenous injection of high dose streptozotocin. Methods Eight male Zhonghua mini-pigs (22.6±1.8 kg) were administrated with high-dose streptozotocin (150 mg/kg) into the ear vein. Before and 10, 30, 90 minutes, 1, 3 and 7 days after administration of streptozotocin, blood samples were obtained respectively, and used to dynamically monitor the fasting blood-glucose. C peptide and insulin levels were evaluated by IVGTT test. Results Since 24 hours after STZ administration, the fasting blood-glucose level was increased significantly compared with that of pre-administration and maintained at 16.7-20.6 mmol/L, while the C peptide and insulin levels were decreased significantly. IVGTT results showed that blood sugar levels at 1 h after intravenous injection of 50% glucose were much higher than 11.1 mmol/L and failed to restore to fasting glucose levels until 2 h, insulin and c-peptide did no response after injection of glucose, always kept at a trace level. Conclusion A single high-dose streptozotocin injection can be used to establish a mini-pig model of type 1 diabetes successfully.

Abstract:Objective To evaluate the rat model of type 2 diabetes mellitus (T2DM) associated with depression by body weight, fasting blood glucose, manifestations, and open field test. Methods The T2DM rat model was induced by high fat diet and low dose of STZ injection, and in addition, the T2DM rats were made restraint stress for 21 days. 32 Wistar rats were randomly divided into three groups: normal group (group N), T2DM group (group T) and T2 DM with depression group (group T+D), with 8 rats in each group. After the model was established, to measure the body weight, fasting blood glucose at day 0, 7, 14, and 21, and observe the gross manifestations, drinking and diet, feces, urine, and mental state, and test the rat depression by open field test. Results After establishment of the T+D rat model, the rats in group T+D showed some symptoms, including messy dark and gloomy hair, slow movement, increasing drinking, diet, feces and urine and mental fatigue. At day 0, 7, 14, and 21, compared with the group N, the body weight of the group T and group T+D were decreased, showing a significant difference (P<0.05;P<0.01). At day 0, 7, 14, and 21, compared with the group N, and the fasting blood glucose in the groups T and T+D were increased, with a significant difference (P<0.01). After 21 days of restraint stress, the fasting blood glucose in the group T+D was significantly higher than that in the group T (P<0.01). Compared with the group N, the total movement distance in 5 minutes in the group T+D was reduced, but without a significant difference (1532.6±126.8 cm vs. 940.5±208.3 cm, P>0.05). Compared with the group N, the movement speed in 5 minutes in the group T was significantly slower than that in the group T, with a significant difference (5.1±0.4 cm/s vs. 2.9±0.6 cm/s, P<0.05), and even more slower than that in the group T+D, with a significant difference (5.1±0.4 cm/s vs. 2.4±0.5 cm/s, P<0.01). Conclusions A rat model of type 2 diabetes mellitus associated with depression has been successfully established by high fat diet and injection of low dose streptozotocin in combination with restraint stress for 21 days. This rat model is useful for further relevant studies.

Abstract:Objective To investigate the application of metabolomics in research of the metabolic profiling of spontaneously hypertensive rats (SHR) and Wistar rats using NMR combined with PLS-DA. Methods SHRs were fed with normal diet for 1 week and then the urines of normal rats and SHR hypertensive model rats were collected at 1, 2, 3, and 4 weeks. The metabolic profiles were analyzed using ¹H-NMR. Partial least squares discriminant analysis (PLS-DA) was used to determine the differential biomarkers. Results The systolic blood pressure (SBP) of SHRs were significantly higher than that of normal rats. Using PLS-DA and orthogonal partial least squares discriminant analysis (OPLS-DA), it was also possible to distinguish them in the principal components score plot. In the SHRs, the levels of some amino acids and some small molecular substances were higher than those in the control rats. Conclusions NMR combined with PLS-DA is a promising approach to provide the information on metabolic changes related to the pathophysiological processes in spontaneously hypertensive rats.

Abstract:Objective To explore the humoral and cellular immune responses of a vaccine of S fragments from a new type reovirus R4 strain in mice. Methods Four recombinant plasmids were constructed by respectively cloning S1, S2, S3,S4 genes into pcDNA3.1⁺, and mice were intramuscularly immunized with the recombinant plasmids in a dose of 100 μg/mouse. Control vector pcDNA3.1⁺ and phosphate buffered saline (PBS) were used as negative controls. The specific antibody level and IgG subclass (IgG1, IgG2a, and IgG2b) were detected by ELISA, and cellular immune responses to R4 were assessed using an interferon (IFN)-γ ELISpot assay. Results All recombinant plasmids induced significantly higher levels of anti-R4 IgG compared with that of the controls (pcDNA3.1⁺ and PBS), and the titers were highest in the mice immunized with S1 and S3. On the other hand, S1 gene induced highest IgG2a antibody and the cellular immune response was best. Conclusions After the mice immunized with S1 gene recombinant plasmid, this plasmid can initiate both cellular and humoral immune responses in mice. S1 gene recombinant plasmid is a promising vaccine candidate.

Abstract:Objective To conduct morphological observation and gene identification of the strain of flagellate isolated from Cricetulus migratorius in the Xinjiang Research Center for Experimental Animals. Methods The ileocecal contents of C. migratorius were microscopically examined on direct smear with Wright-Giemsa staining, and the total RNA isolated from Xinjiang C. migratorius was extracted and 16S rRNA was amplified by PCR, and then sequenced. Furthermore the homology was compared and the phylogenetic tree was developed using MEGA5.22 software. Results Morphological observation indicated that the isolated flagellate was Tritrichomonas muris. The 16S rRNA gene sequence of the Xinjiang C. migratorius isolate shared highly homology with that of other Tritrichomonas. Phylogenetic tree analysis indicated that the 16S rRNA gene of Xinjiang C. migratorius isolate was classified into a subgroup with T. muris 16S rRNA (U85966.1), but was relatively distant relative from other related tritrichomonas. Conclusions The flagellate isolated from Xinjiang C. migratorius is identified to be T. muris by both morphological observation and 16S rRNA gene analysis.

Abstract:Objective The aim of this study was to investigate the masculinizing effects of progesterone (PRO) exposure at different concentrations on the morphology of female mosquitofish (Gambusia affinis). Methods Immature female G. affinis individuals were put into static water with 0.5, 5, 50 and 500 nmol/L progesterone (n=56), respectively. In addition, control group and parallel groups were set up. After 42-day-long exposure, we measured four main indexes of the G. affinis: the body length, body weight, morphological changes in the anal fin 3rd fin and the 14, 15 and 16 vertebral ribs. We also observed the state of ovarian development. Results The body lengths (BL) of experimental groups exposed at concentration of 50 and 500 nmol/L progesterone for 42 d showed significant differences, respectively (P<0.01 for both), when compared with those of the control group. The body weights (BW) of experimental groups exposed to progesterone at all concentrations were significantly or very significantly decreased (P<0.05 or P<0.01). Only the progesterone at 5 nmol/L concentration showed significant effect on the body health index (CF) (P<0.05). The section number (FJ) of the anal fin 3rd fin, the section length (FL)and the widest portion width (FW)in the experimental groups had significant difference (P<0.05), respectively, showing significant changes in morphology after the exposure to 500 nmol/L progesterone. However, there was no significant difference between the values resulted by progesterone in other concentrations (P> 0.05). When exposed to 500 nmol/L PRO, the L, D and L:D values of the 14th, 15th and 16th vertebral ribs of the female G. affinis had very significant difference (P<0.01 for all), respectively, indicating significant changes of morphological masculinization. Histological examination revealed that the developmental status of the ovaries of G. affinis in the experimental groups were to different degrees inhibited, and the nuclei appeared swelling in stage Ⅱ and Ⅲ oocytes. Conclusions The results of this study indicate apparent masculinizing effects of progesterone on immature female mosquitofish.

Abstract:Objective The aim of this study was to observe the dynamic changes of serum and bone marrow levels of alkaline phosphatase (ALP) in ovariectomized (OVX) rats. Methods Eighty 3-month-old non-pregnant female Sprague Dawley rats were randomly divided into following groups by body weight: baseline group, sham-operation (Sham), and ovariectomized (OVX) rat groups. The rats were killed at 0, 3, 6, 12 or 24 weeks after operation. The changes of serum and bone marrow alkaline phosphatase (ALP) levels were detected with photometer-721 and the number of ALP-positive bone marrow cells was detected with a research grade upright fluorescence microscope. Results In the sham group, the serum ALP levels were significantly increased in the weeks 3 until the week 6. However, the serum ALP levels were significantly decreased in the week 12 until the week 24. The ALP-positive cell number in the bone marrow was significantly increased in the week 3 until week 12, but significantly decreased in the week 24. In the OVX group, the serum ALP levels were significantly increased in the week 3, but significantly decreased in the week 6 until the week 24. The number of ALP-positive bone marrow cells was significantly decreased in the week 3 until the week 24. In addition, the serum ALP levels in the OVX group were significantly higher than that of the sham group. However, the number of ALP-positive bone marrow cells was significantly lower than that of the sham group. Conclusions The change tendency of serum ALP in the sham group is similar with that of bone marrow cells. But the change tendency of serum ALP in OVX rats is different from that of bone marrow cells.

Abstract:Objective To explore the effect of electroacupuncture on CD34⁺VEGFR2⁺endothelial progenitor cell (EPC)-derived vessels and stromal cell-derived factor-1α (SDF-1α)/CXCR4, and study its mechanism of promoting angiogenesis in hippocampus after focal cerebral ischemia/reperfusion. Methods A total of 180 healthy male adult Sprague Dawley (SD) rats were randomly divided into sham operation (sham) group, model (I/R) group, electroacupuncture (I/RE) group, I/RE plus AMD3100 (A specific antagonist of CXCR4) group (I/REA) and AMD3100 (I/RA) group. The rats received filament occlusion of the right middle cerebral artery for 2 hours followed by reperfusion. Electroacupuncture was applied to "Baihui" (GV20)/"Siguan" (Hegu LI 4/Taichong LR 3) acupoints for 30 min, once a day. The mRNA expression of SDF-1α and CXCR4 were detected by reverse transcription-polymerase chain reaction (RT-PCR). Double immunofluorescence was used to stain CD34⁺VEGFR2⁺ EPC-derived vessels. Results Compared with the sham group, the mRNA expressions of SDF-1α and CXCR4 were significantly upregulated in I/R and I/RE group(P<0.05), but that in I/RE group was more significantly increased than I/R group(P<0.05). In addition, the mRNA expression of SDF-1α and CXCR4 were highly increased on day 1 in the I/REA group than that of I/RE group, but decreased than that of I/RE group on day 7 after reperfusion (P<0.01). CD34⁺VEGFR2⁺ EPCs-derived vessels were obviously increased on 3d and 7d in the I/RE group compared with that of the I/R group, and significantly decreased on 7d in the I/REA group compared with that of the I/RE group (P<0.01). Conclusions Electroacupuncture can effectively promote angiogenesis through upregulating the expression of SDF-1α and CXCR4 in rat ischemic hippocampus after focal cerebral ischemia/reperfusion.

Abstract:Objective To establish an animal model of calpastatin (CAST) transgenic mice by inserting the full human CAST into the genome of C57BL/6J mice. Methods Recombinant transgenic vector pRP.EX3d-EF1A-CAST-IRES-eGFP was constructed by Gateway technology. It was injected into the fertilized eggs from C57BL/6J mice. The injected eggs were transplanted into the oviduct of pseudopregnant mice. Tail DNA PCR screening was performed to identify the positive founder mice. The expressions of CAST mRNA and protein in tissues of the transgenic mice were detected by RT-PCR and Western blotting. Results Ninty eggs were transplanted into the oviducts of 3 recipients. The transplantation success rate was 100%. 23 viable offsprings were born from the recipients. Tail DNA PCR screening showed that two of the offsprings were positive transgenic mice. The positive rate of transgenic mice was 9%. RT-PCR assay revealed that CAST mRNA expressions were present in the heart, liver, kidney, lung, spleen, brain and skeletal muscle of the transgenic mice. Additionally, the CAST protein expression was significantly increased in the transgenic mice. Conclusion CAST transgenic mice have been successfully established and provide a good animal model support for further studies on the CAST function.

Abstract:Objective The aim of this study was to establish a mouse model of chronic hepatic injury induced by low dose carbon tetrachloride (CCl₄). Methods Twenty SPF male B/C mice (body weight 18-20 g) were randomly divided into three groups including the CCl₄-treated group, oil-treated group and non-treated control group (n=5/group). Mice in the CCl₄-treated group were intraperitoneally injected with 0.5% CCl₄ prepared in oil. Mice in the oil group received intraperitoneal injection of oil. Mice in the non-treated control group were left untreated. After 6 weeks, the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured, as well structure, cellular morphology and degree of fibrosis of the hepatic tissues were examined by histology with HE and Masson staining. Results After low dose CCL₄ treatment, the serum ALT and AST were significantly increased(P =0.00). Histology with HE staining showed extensive vacuolar degeneration of hepatic epithelial cells and large number of necrotic foci. Histology with Masson staining revealed fibrous hyperplasia mainly located around hepatic lobules. Quantitative analysis of the fibrosis showed that the degree of fibrosis and the integrated optical density of fibrosis were significantly increased after CCl₄ induction(P=0.00). Conclusion Low dose carbon tetrachloride can induce hepatic injury in B/C mouse models presenting pathological changes of hepatic injury and fibrosis.

Abstract:Objective To compare the effects of different transplantation sites on the outcome of testicular grafts in mice, and to provide a basis for development and application of this technique in relevant research. Method 5-day old and 4-week old SPF male C57BL/6J mice were used in this study. Three groups of testicular transplantation, i.e. dorsal subcutaneous transplantation (5 mice, 40 testes), transplantation inside the testicular tunica albuginea (6 mice, 12 testes), and were subrenal capsule transplantation (10 mice,15 testes) groups were set up for evaluating the effect of transplantation site on the outcomes in mice. Sham operation (4 mice) and castration (4 mice) groups were also used in this study. The mice were sacrificed at 8 weeks after transplantation and the transplanted testes were collected for analysis of their weight, transplant survival, weight gain, and germ cell differentiation. Results There were significant differences of the testicular germ cell differentiation in different transplantation groups. The germ cell differentiation was best in the intra-tunica albuginea transplantation group, and were similar to that in the sham operation group. The germ cell differentiation rate was 100% in the intra-tunica albuginea transplantation group, 29.2% in the subcutaneous transplantation group and 0% in the subrenal capsule transplantation group. Conclusions Transplantation beneath the testicular tunica albuginea is the most favorable site for germ cell differentiation, and dorsal subcutaneous transplantation is an alternative choice. Subrenal capsule transplantation is not appropriate for preservation of male reproductive organs in mice.

Abstract:Objective To study the effect of cryopreservation on some biological properties of rabbit adipose-derived mesenchymal stem cells (rADMSCs). Methods rADMSCs culture was isolated by tissue explants adherent method. Morphology of the primary cells was observed by inverted microscopy. Immunophenotypes of the rADMSCs were determined using flow cytometry. The third passage cells were preserved in liquid nitrogen for 6 months, and then were thawed, and subcultured to passage 7. The growth curves of the cryopreserved cells were analyzed by MTT assay, and the cryopreserved cells were cultured in adipogenic and osteogenic medium, with non-cryopreserved rADMSCs as a control group. The adipogenic and osteogenic abilities of the rADMSCs were evaluated by oil red O staining, alizarin red staining and alkaline phosphatase activity assay, respectively. Results The rADMSCs cultured in vitro exhibited a spindle-shaped appearance and rapid growth expansion. Flow cytometry analysis revealed that the third passage rADMSCs were CD44-and CD90-positive, but negative for hematopoietic cells surface marker CD45. The growth curves of cells in the experimental and control groups were "S" shaped, showing a non-significant difference between the two groups (P>0.05). The oil red O staining and alizarin red staining results were positive at 2 weeks after adipogenic and osteogenic induction. The ALP activities of the two groups were increased with osteogenic induction time, with a non-significant difference (P> 0.05). Conclusions Cryopreservation does not affect the growth and differentiation pluripotency of rADMSCs significantly.

Abstract:Objective To analyze the genetic variation among white hair black eyes (WHBE) rabbit, Japanese white (JW) rabbit and New Zealand white (NZW) rabbit using random amplified polymorphic DNA (RAPD) technique. Methods Thirty rabbits (male/female 1:1) of each strain were used in this study. The genomic DNA was extracted from 90 rabbits. Sixty arbitrary primers were used to amplify DNA of rabbits with RAPD-PCR method. Based on the preliminary experiments, polymorphic primers were selected to analyze the genetic variation among the three rabbit strains. The experimental data were analyzed using Popgene 3.2 software. Results (1) Twenty-five polymorphic primers were selected among 60 arbitrary primers. 493 amplified fragments were detected ranging from 100 bp to 1800 bp. Sixteen primers among 25 arbitrary primers could not only amplify the common DNA bands of 3 rabbit breeds, but also amplify particular alleles in the WHBE rabbit. (2) 234 RAPD sites were detected by agarose gel electrophoresis in WHBE rabbit, among which 166 sites were polymorphic, accounting for 70.94%. 228 RAPD sites were detected by agarose gel electrophoresis in the JW rabbit, while 122 sites of them were polymorphic, accounting for 53.51%. 231 RAPD sites were detected by agarose gel electrophoresis in the NZW rabbits, with 94 sites being polymorphic, accounting for 40.69%. (3) The Shannon genetic diversity index of WHBE rabbit, JW rabbit and NZW rabbit was 0.3385, 0.2222 and 0.1905, respectively. (4) The genetic similarity between JW rabbit and NZW rabbit was highest among the three rabbit breeds (0.8443), followed by that between WHBE rabbit and JW rabbit (0.8204), and the genetic similarity between WHBE rabbit and NZW rabbit (0.7862) was the lowest. Conclusions Our results demonstrate that there are both genetic similarities and genetic variations among WHBE rabbit, JW rabbit and NZW rabbit. The RAPD technique can be used to delect the genetic relationships among different breeds and different individuals of the same breed of rabbits.

Abstract:Mouse and rat models of type 2 diabetes mellitus play a key role in basic and clinical translational studies. Different animal models should match the determined investigational objects and methods. In this review, the establishments and diabetes-specific changes of various animal models were described, which will be helpful for better use of animal models in research of diabetes mellitus.

Abstract:Traditionally it has been widely accepted that our genes together with adult lifestyle factors determine our risk of developing noncommunicable diseases such as type 2 diabetes mellitus,cardiovascular disease and obesity in later life. Currently, there is substantial evidence that the early life environment plays a key role in determining our susceptible to such diseases in later life. As an increased focus on the understanding of central insulin resistance, central nerve system plays an extremely important role in regulating the peripheral glucose homeostasis. Substantial studies have indicated that it may be regulated by epigenetic mechanisms. Epigenetics can be defined as the study of heritable changes in gene expression that do not involve alterations in the DNA sequence.Epigenetic processes play a central role in regulating tissue specific gene expression and hence alterations in these processes can induce long-term changes in gene expression and metabolism which persist throughout the lifecourse. This review will focus on the epigenetic machanisms of central neuroregulation between early life environment and glucose metabolism in adulthood.

Abstract:Hyperuricemia is closely associated with abdominal obesity. The prevalence of hyperuricemia combined with abdominal obesity has been increased significantly in recent years, along with the improvement of daily life and the changes in dietary structure. The state of hyperuricemia combined with abdominal obesity is most harmful, and becomes a common and high risk metabolic disease. Animal model with hyperuricemia combined with abdominal obesity is very important for the research of pathomechanism and treatment of this disease.

Abstract:Cardiovascular complication of diabetes (CCD) is a leading cause of mortality in diabetic patients, and diabetic cardiomyopathy(DC) vastly accounts for heart failure in those CCD patients. In the research of pathogenesis, early diagnosis and better treatment of DC, cardiac function evaluation plays a key bridging role. Addtionally, animal models may well simulate the development of diabetes. The purpose of this article is to review the important methods in evaluating heart function of diabetes animal models.

Abstract:The establishment of rat models of type 2 diabetes mellitus can provide an ideal animal experimental technology platform for the research of pathogenesis, prevention, diagnosis and treatment of diabetes mellitus and its complications. The establishment of type 2 diabetic rat models may be affected by various factors. In this paper, the main factors, streptozotocin (STZ) application, high fat and calorie diet, selection and breeding of rats are reviewed, which will provide an important reference for the development of rat models type 2 diabetes.

Abstract:Ningxiang pigs fat-type breeds have been increasingly used in biomedical research due to the characteristics of genetic stability, high food value, fast growth, adaptability and strong reproductive ability. The recent development in research on Ningxiang pigs in a closed group training, inbred cultivation, biological characteristics and medical application is reviewed in this article. Ningxiang pigs have been closed-bred for fourteen years, and may be applied in biomedical research such as artery atherosclerosis and diabetes mellitus.

Abstract: