Abstract:Objective To explore the proliferation characteristics of primary small intestinal epithelial cells of tree shrews and the characteristics of human rotavirus (RV) G1P[8] infection to these cells, and establish a model of tree shrew primary small intestinal epithelial cells infected with human rotavirus G1P[8]. Methods The primary small intestinal epithelial cells were obtained by collagenase XI and dispase I digestion from tree shrew. After purification and identification, the obtained primary small intestinal epithelial cells were infected with RV. Then, culture supernatants of infected cells were collected every 12 hours after infection. Viral titer and viral load were subsequently determined. Western blot and indirect immunofluorescence observation were used to detect the expression of RV protein VP6 in the primary cells. The infectivity of RV to the tree shrew primary cells was finally evaluated.Results After purification and identification of primary epithelial cells from the tree shrew, high purity above 90% primary tree shrew small intestinal epithelial cells was obtained. These primary small intestinal epithelial cells could be infected with RV virus by comparing the virus infectivity to primary renal cells, HCT116 cells and MA104 cells. The virus titer reached to 2.0×10⁵TCID₅₀/mL at 72 h after infection. Using Western blot and indirect immunofluorescence observation, the specific viral protein of VP6 was determined to be expressed in the tree shrew primary small intestinal epithelial cells, and were located in the cytoplasm from days 1 to 5. Conclusions The separation, purification and cultivation methods of tree shrew primary small intestinal epithelial cells are successful, and the tree shrew model of RV-infected the tree shrew primary small intestinal epithelial cells is successfully established.

Abstract:Objective To establish an enterovirus 71 (EV71) infection model of tree shrew primary renal cells. Methods Tree shrew primary renal cells were obtained by trypsin digestion. After subculture and purification, EV71 virus was used to infect these primary cells. The culture supernatant of these EV71-infected cells was collected for virus titer detection at 1, 2, 4, 6 and 8 days post-infection. The cells were collected for detection of EV71 VP1 protein by Western blot assay. Furthermore, the expression and location of VP1 protein in the infected cells were detected by indirect immunofluorescence assay. Vero cells were taken as positive control to evaluate the infectivity of EV71 virus to tree shrew primary renal cells. Results Morphologically, the cultured cells were proved to be majorly consisted of the primary renal cells after subculture and purification. The obtained primary cells were infected by EV71 virus. The virus titer was up to 1.3 × 10⁶ TCID₅₀/mL during 48-96 h post-infection, proving that EV71 virus infected and proliferated in the tree shrew primary renal cells. Western blot showed that the viral VP1 protein was detected from infected primary cells at 2 to 8 d post infection. VP1 protein was also observed in the cytoplasm at 2 to 6 d post infection by indirect immunofluorescence. Compared with Vero cells, the infectivity of EV71 virus to tree shrew primary renal cells and its proliferation were confirmed. Conclusions Based on the successful establishment of cell culture of tree shrew primary renal cells, the infectivity to the obtained cells and proliferation of EV71 virus in the cells are confirmed. The model of EV71 virus-infected tree shrew primary renal cells is initially established.

Abstract:Objective To preliminarily explore the feasibility of tree shrew as a new kind of animal model in research of amblyopia, to discuss the primary visual cortex plasticity mechanism of form deprivation in tree shrew, and to provide a theoretical basis for further understanding the mechanism of amblyopia formation and recovery. Methods Sixty 30-days old tree shrews were divided into five groups, 12 in each group: the group A had the right eye sutured for 1 month; the group B had the right eye sutured for 2 months; the group C had the left eye sutured for 1 month and then opened and the righ eye was sutured for 1 month, in other words, the group C was performed by alternating suture; the tree shrews of control group 1 (D1) were in the same age as the the group A, but fed in normal breeding environment; the tree shrews of control group 2 (D2) were at the same age of groups B and C, but fed with a normal diet. Samples of the visual cortex were taken after the completion of modeling, and were processed to observe the histology and ultrastructure of the visual cortex, the neuron apoptosis, and the c-fos protein expression in the tree shrews of different groups. Results Damages to different degrees were found by histological and electron microscopic examination of the visual cortex in each experimental group, and they were more obvious in the group sutured for 2 months. TUNEL staining showed that there were no significant differences between the apoptosis in the experimental and control groups. The expression of c-fos mRNA and protein in the experimental groups was decreased, and it was the lowest in the group sutured for 2 months. There was a small increase in the c-fos expression after the alternate suture, and no significant difference of c-fos expression was found in the control groups. Conclusions Different degrees of deprivation amblyopia lead to different histopathological changes. There is a plasticity in the neurons affected by amblyopia. Tree shrew can be used as an ideal animal model for the studies of form deprivation amblyopia.

Abstract:Objective The aim of this study was to differentiate and identify all the variants of APP mRNA in tree shrew, describe the characteristics of APP genes, and determine its distribution in different tissues. Method Based on the known human APP sequence and predicted tree shrew APP gene, we designed a pair of common specific primer. We extracted total RNA from different tissues respectively, using RT-PCR to get the targeted cDNA. We differentiated the variants by electrophoresis. Finally we recollected the RT-PCR products for sequencing. Combined with the qPCR results, we confirmed the quantitative distribution of the variants in different tissues. Results Our sequencing results showed that the length of the tree shrew APP spliceosome was 3514 bp with a 109 bp 5'-UTR and a 1092 bp3'-UTP. APP genes in the investigated 9 species were highly homologous and conservative, tree shrews and primates show a very close genetic relationship. The tree shrew APP genes shared four domains by 3D modeling. We also confirmed the distribution pattern of 4 spliceosomes derived from exon jumping. All existed APP variants, including APP770, APP695, APP751, and APP677, were all expressed in the lung, kidney and intestine. The highest expression levels were in the lung, muscle, and testicles. Conclusions This study helps to promote the studies of tree shrews as an Alzheimer's disease model and the mechanism of its pathogenesis and drug development.

Abstract:Objective To investigate the expressions of metalloproteinase-9 (MMP-9)in serum, brain and aorta matrix and tissue inhibitor of metalloproteinase-1(TIMP-1) in renovascular hypertensive rats (RHR),and to evaluate the association between blood pressure and levels of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1(TIMP-1).Methods Eighty healthy male SD rats were randomly divided into RHR group (n=40) and sham-operated group (n=40). Hypertension was induced by two-kidney, two-clip (2K-2C)clamps.Systolic blood pressure (SBP) was measured every 2 weeks during 12 weeks using a tail pressure meter. Stroke was confirmed by Longa's five-point scale and pathological examination. The expressions of MMP- 9 and TIMP-1 in the brain and aorta tissues were detected by Western blot and immunohistochemistry. The levels of serum MMP-9 and TIMP-1 were measured by double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). Results Compared with the sham-operated group, SBP stayed significantly elevated in the RHR group at 2, 4, 6, 8, 10 and 12weeks after the operation [(157±9.0) vs. (128±7.0), (176±10.0) vs. (122±6.0), (194±8.0)vs. (117±6.5), (202±12.0)vs. (124±8.0), (218±15.0) vs. (126.±8.5),and (224±20.0)vs. (129.±9.0) mm Hg, all P < 0.05]. 12 weeks after the surgery, the level of serum MMP-9 in the RHR group was kept significantly higher than that in the sham-operated group [(783.4±109.79)vs. (573.4±109.59) ng/mL, P < 0.05],and the serum TIMP-1 level was lower in the RHR group than that in the sham-operated group[(313.02±83.9) vs. (976.19±191.1) pg/mL, P < 0.05]. MMP-9 expressions were significantly higher in the brain and aorta in the RHR group than that in the sham-operated group(both P < 0.05), and TIMP-1 expressions were lower than that in the sham-operated group(both P < 0.05).The Pearson correlation analysis showed that MMP-9 levels in serum,brain and aorta were positively correlated with systolic blood pressure (r=0.557,r=0.774 and r=0.661,all P < 0.05), and TIMP-1 levels were negatively correlated with systolic blood pressure(r=-0.481,r=-0.535 and r=-0.685,all P < 0.01). Conclusions Hypertension induces increased MMP-9 and decreased TIMP-1 in serum, brain and aorta in renovascular hypertensive rats. There are consistent alterations of circulating and tissue MMP-9 and TIMP-1 levels in renovascular hypertensive rats. There is a relationship between increased blood pressure and high MMP-9 and low TIMP-1 in serum and tissues.

Abstract:Objectives To study the role of protein serine/threonine phosphatase1γ (PP1γ) and DNA methylation in learning and memory. Methods The mice and cells were treated with 5-aza-cdR, a DNA methyltransferase (DNMT) inhibitor, and to observe the changes of ability of learning and memory and expression levels of PP1γ in mice. The ability of learning and memory in mice was assessed by Morris Water maze test. The mRNA expression levels of DNMTs and PP1γ in the mouse hippocampus were determined by real-time PCR and the protein level of PP1γ was measured by Western blot. To further investigate their role, NG108-15 cell line was treated with 5-aza-cdR. Flow cytometry, xCelligence and luciferase were used to detect the effects of 5-aza-cdR on the proliferation, apoptosis and PP1γ transcription in the NG108-15 cells. Results The ability of learning and memory was enhanced in the mice after administration of 5-aza-cdR injection. The expressions of DNMTs and PP1γ were decreased in the mouse hippocampus after injection of 5-aza-cdR. On the other side, 10 μmol/L 5-aza-cdR inhibited cell proliferation and decreased PP1γ transcription without inducing apoptosis. Conclusions Our data demonstrate that 5-aza-cdR inhibits the expression of PP1γ which is related to learning and memory in mice.

Abstract:Objective To establish a visualization model of CT three-dimensional reconstruction of tree shrew skeletal system and to provide a basis for diagnosis of tree shrew skeletal system diseases. Methods We used Toshiba Aquillon One 320 row helical CT scan (voltage: 100 kV, electric current: 80 mA, bulb revolution: 0.35 s/roll, pitch: 1.35, image set: 0.5 mm, layer spacing: 0.5 mm, algorithm: bone reconstruction method, and we used volume imaging (VR), multi-planar imaging (MPR) and surface imaging technology by choosing Musculoskeletal CT option of Vitrea package in the parallel computing environment for 3D reconstruction. Results The reconstruction of visual model structure is distinct, clear, allowing to truly reappear the tree shrew skeletal system in the 3D computer model. In this model, we identified five ridges on the back of the skull, and located four big holes in the lateral view. In addition, we showed some smaller skull holes, such as optic foramen and submandibular foramen. We determined the quantitative data of the tree shrew skeleton system more precisely and comprehensively. We also found some abnormal data in tree shrews, such as pelvic asymmetry, increased thorax, bone fracture and calcification in the synovial bursa. Conclusions The established CT 3-D visualization technique can determine special features of the skeletal system in tree shrews non-invasively, which is very important for ecological classification, identification, and evolution analysis of tree shrews.

Abstract:Objective To determine the reproductive physiology and blood physiological and biochemical characteristics of SPF Yorkshire and Landrace swine. Methods Ten reproductive physiology parameters, 19 blood physiological parameters and 18 blood biochemical parameters in SPF Yorkshire and Landrace swine were measured using conventional methods and the differences between population, between age groups and between both sexes were analyzed. Results There were no significant differences (P > 0.05) in reproductive physiology parameters and most blood physiological and biochemical parameters of the SPF Yorkshire and Landrace swine. A few of parameters, such as blood physiological indices GRAN, HGB, RDW, PLT, PCT, and blood biochemical indices ALKP, CHOL, TBIL, BUN, showed significant difference (P < 0.05) between populations, between age groups and between both sexes, however, the values of difference were rather small, deviated from the normal range. Conclusion The physiological and biochemical characteristics of SPF Yorkshire and Landrace swine are basically stable and there is no significant difference compared with other laboratory miniature pigs. This study will provide valuable basic data for raising velvet yield, establishment of animal models and evaluating the genetic quality of closed colony.

Abstract:Objective To establish a pure planarian Dugesia asexual strain in China. Methods The planarian worms were collected from a wild stream, and then made the worm grow after amputation in the lab. To establish an asexual strain through cutting and culturing for the single worms. Results After ten years and more than ten thousand experiments, an asexual pure planarian strain Dugesia ZB-1 originated from Shandong Zibo area was established. It grows stably under laboratory controlled conditions. Conclusions The establishment of Dugesia ZB-1 Priovides a solid foundation for further experiments and promoting planarian research in our country and participation in the field of the international planarian research.

Abstract:Objective To establish a breeding method of Myodes rufocanus in the laboratory, collect their growth and reproduction data, and provide a basis for carrying out the experimental animalization.Methods Wild Myodes rufocanus caught in the Moranbong woodland were brought back to the laboratory. They were bred artificially in a large hard wall rodent negative pressure isolator. Their growth and reproduction data were recorded for evaluating the results of breeding. Results The Myodes rufocanus were successfully bred in the laboratory. The pregnancy rate was 54.55%.The average pregnancy length was 20.4 days(8 to 22 days). During one breeding period,they gave birth 2.9 times on average. The maximum number of births was 7 times,far more than the number tested under field conditions.The average litter size was 4.3±1.22. The highest litter number of a single nest was 8. The weaning rate of pups was 94.8%. The growth and development of pups were good. Conclusions The breeding method for Myodes rufocanus is established. The growth and reproduction data are tested too. The results of our study laid a foundation for the experimental animalization of Myodes rufocanus.

Abstract:Objective To establish a porcine model of liver failure after different percent hepatectomy. Methods The porcine models of liver failure 75%, 85%, 95% hepatectomy were developed and the living conditions and survival time were recorded. The blood samples of pre-surgery, post-hepatectomy d1, d3, d5 and post-hepatectomy 1 week, 2 weeks, and 3 weeks were collected for hepatic function analysis. Histological examination of liver tissues was performed using HE staining. Liver injury histology was interpreted and scored in the terminal samples. Results The average survival time of pigs with post-hepatectomy liver failure after 75%, 85%, 95% hepatectomy was 19.0±5.6 days, 17.3±5.5 days, 1.3±1.5 days, respectively. Their pathological scores were 5.67±0.52, 8.17±0.82 and 8.50±0.71, respectively. With the increase of percent hepatic resection, the incidence of hepatic failure was increasing. ALT, AST, ALP, LDH and TBA were dramatically increased in the pigs after 85% hepatectomy. Conclusions The pig model of acute liver failure by 85% hepatectomy is successfully established, which can cause typical acute liver failure in Bama miniature pigs.

Abstract:Objective To investigate the effects of sand therapy temperature on bone reconstruction and numerical study on the heat transfer. Methods CT scan imaging was performed respectively for four times (before and after OA model was set up, and after the first and second weeks during the sand therapy). Import the scan data to MIMICS software, and the changes of femoral bone mass layer were analyzed. After combining the muscle, femur, bone marrow and meshing, the established three-dimensional model of the STL format was introduced into the COMSOL software for heat transfer simulation and effect of stress on bone remodeling induced by temperature field. Results Changes of four CT scan data were analyzed. The soft bone volume was reduced, while the dense and hard bone volume were increased. Numerical simulation on the heat transfer showed the temperature distribution of the thigh and the femur. Conclusions The thermal stress produced by sand therapy temperature exert promoting effect on the femur bone remodeling.

Abstract:Objective To observe the tumorigenicity of High Five insect cell line in Balb/c nude mice, and make sure the safety of the cells when used in vaccine production. Methods Balb/c nude mice were randomly divided into 5 groups: the basic cell bank of High Five group, the highest limited passages of High Five group, HEp-2 cell group as positive control, CEF cell group as negative control, and blank control. Except of the blank control, cell suspension was injected subcutaneously into the nude mice in the different groups, respectively. At 3 and 12 weeks after injection, anatomical observation and histopathologic examination were performed to detect the tumor formation. Results At 3 and 12 weeks after injection, the tumorigenicity study results showed that no tumor developed at the transplantation site in the blank control group, negative group, and High Five groups. Histopathological examinations also showed no abnormality in these groups. Otherwise, squamous cell carcinoma was developed in the positive group at 3 weeks after injection. Conclusions High Five cells of basic cell bank and highest limited passages are not tumorigenic, and can be used in vaccine production safely.

Abstract:Objectives To systematically evaluate the effect of music given to pregnant rats on the development of brain functions in the offspring rats and to provide scientific evidence for the application of antenatal musical training and the promotion of welfare for laboratory animals.Methods We comprehensively retrieved and collected the research literatures related to the effect of music on brain function development in offsprings of the pregnant rats from Pubmed, Web of Science, Cochrane Library, Wanfang, Weipu, CNKI and CBMdisc. The retrieval time was set from the foundation date of databases to 2 April, 2016. We selected literatures according to the inclusion and exclusion criteria, evaluated their utilities, then extracted and qualitatively described the data. Results Seven experimental studies were selected in this study including 4 published in Chinese and 3 in English. The object laboratory animals of those studies were Wistar or SD rats. Music materials involved comfort music, classic music, violin concerto(Liangzhu/The butterfly lovers). Intervention were given to the pregnant rats roundly from the gestation until parturition. These results showed that, to some extent,music stimulations during gestation may promote the development of brain function and improve spatial memory of the offspring rats. However,expressions of some functional receptors were not significantly altered.Conclusions Appropriate music provided to the pregnant rats promote the development of brain functions in their offspring.

Abstract:Objective To establish a Wuzhishan minipig model of atherosclerosis (AS) induced by high fat/cholesterol diet, and observe the changes of expression of lipoprotein associated phospholipase A₂ (Lp-PLA₂) in plasma and plaques.Methods 10 Wuzhishan minipigs were randomly divided into 2 groups: The normal control (Ctr, n=4) group was fed with normal diet, and AS model (n=6) group fed with high fat/cholesterol diet for 24 weeks. After the modeling for 24 weeks, the changes of total cholesterol (TC), low density lipoprotein (LDL-C), high density lipoprotein (HDL-C), triglyceride (TG), C-reactive protein (CRP), Lp-PLA₂ activity and composition were detected. The changes of vascular lipid deposition and plaques were assessed by pathology using oil red O staining and HE staining, respectively, and immunohistochemical staining for IL-6 protein expression. Moreover, the expression of Lp-PLA₂ mRNA determined by RT-PCR and protein by Western blot were observed in the abdominal aortic tissues. Results Compared with the control group, the body weight, body mass index (BMI), TC, LDL-C, HDL-C, CRP, Lp-PLA₂ activity and composition and aortic lipid deposition were significantly increased, and AS plaque formation was observed in the AS model group (P < 0.05, P < 0.01). The expression of Lp-PLA₂mRNA and protein and IL-6 protein in abdominal aortic tissues were also significantly increased (P < 0.05). Conclusions Long-term high fat/cholesterol diet feeding for 24 weeks can induce atherosclerosis in Wuzhishan minipigs, and Lp-PLA2 plays a key role in the vascular inflammation and plaque formation.

Abstract:Objective To observe the changes of S-opsin expression in guinea pigs with flickering light-induced and form-deprived myopia, and to investigate the causes. Methods Thirty-six two-week-old healthy guinea pigs were randomly assigned to three groups: Flickering light-induced myopia group (FLM group,n=13), form-deprived myopia group (FDM group,n=12) and control group (n=11).For the FLM group, the cages were equipped with astroboscope (0.5 Hz), and LEDs were used as the light source. The right eyes of the guinea pigs in FDM group wore translucent goggles which did not interfere with the normal activity of their eyelids. No special treatment was given to the guinea pigs in the normal groups.All measurements were performed prior to and then after 6 weeks of treatment. The first measurement day was recorded as 0 week. Biological parameters, such as the refraction, axial length (AL) and corneal radius of curvature (CRC), were measured and fundus photography is performed before and after 6 weeks of the treatment. The expression of S-opsin was observed and analyzed by immunofluorescence technique and image analysis system. Results Before the treatment, no significant difference was found in three biometric measurements including refraction, AL and CRC between the groups at 0 week (P > 0.05). After the treatment for 6 weeks, significant differences were found in changes of both the biometric measurements between the FLM and control groups, and between the FDM and control groups (P < 0.05). However, no significant differences were found in the changes of CRC among the FLM, FDM and control groups(P=0.358), indicating that myopia models were established successfully. No significant differences were found in the changes of values of refraction, AL and CRC between the FLM and FDM groups (P > 0.05). Expression of S-opsin differed in the FLM and FDM groups. For the mean gray values of green channel,compared with the control group respectively, significant differences were found in both the FLM and FDM group (P < 0.001). The mean gray value of green channel of the FLM group was higher, however the mean gray value of green channel of the FDM group was lower. Conclusions Both guinea pig models of flickering light-induced and form-deprived myopia can be established successfully. S-opsin is increased in the flickering light-induced myopia model and decreased in the form-deprived myopia model, indicating that the mechanisms of formation of these two experimental myopia models may be different.

Abstract:Objective This study was designed to explore the therapeutic effect of psoralen on type II collagen-induced rheumatoid arthritis in mice and its molecular mechanism. Methods DBA/1J mice were immunized with type II bovine collagen to induce rheumatoid arthritis. The model mice were randomly divided into Psoralen group (PSO), methotrexate group (MTX) and model group (Vehicle). Clinical signs of arthritis in the mice were monitored. The spleen index was assessed. Splenic Th1 and Th2 cells were counted by flow cytometry. ELISA was used to detect the levels of inflammation-associated factors TNF-α, IL-6 and IL-1β in the serum. Results Compared with the vehicle group, the ankle swelling and limitation of joint activity in the PSO group were significantly reduced, the spleen index and Th1 cell percentage were significantly decreased, and the Th2 cell percentage showed no significant change in the PSO group. Expression of TNF-α, IL-6 and IL-1β in serum was notably decreased in the PSO group. All the indexes showed no significant difference between the PSO and MTX groups. Conclusions Psoralen may attenuate the severity of type II collagen-induced rheumatoid arthritis in mice by regulating the balance of Th1/Th2 cells and inhibiting the expression of TNF-α, IL-6 and IL-1β.

Abstract:Objective To investigate the effects of lipopolysaccharide (LPS) on myelin phagocytosis during Wallerian degeneration after early peripheral nerve injury in rats. Methods Fifty male Wistar rats were recruited and randomly divided into LPS group (n=20), model group (n=20) and sham group (n=10). The right sciatic nerves of rats in the LPS and model groups were cut and sutured end-to-end, while the sciatic nerve of sham group rats were only exposed. Immediately after surgery, the rats in LPS group were given microinjections of LPS (2 g/L) into the surgical site in a final volume of 1 μL, and the rats in other two groups were injected with the same volume of saline. The sciatic nerves were taken at 1.5 h, 24 h and 7d after surgery. Real-time quantitative PCR (qRT-PCR) was applied to detect the dynamic expressions of IL-1β mRNA and MCP-1 mRNA. Immunofluorescence staining was used to test the expression of CD68⁺ macrophages in sciatic nerves. HE staining was used to observe the pathological alterations of sciatic nerves tissue. ORO staining was used to observe sciatic nerves demyelination. LFB staining was used to detect the sciatic nerves myelin. Sciatic function index was used to evaluate the recovery of motor function in rats. Results Compared with the model group, qRT-PCR indicated that the expression of IL-1β and MCP-1 from LPS group were increased at 1.5 h and 24 h after surgery (P < 0.001,P < 0.001), respectively. Compared with the model group, the expression of CD68⁺ cells was increased significantly at 7th day after surgery (P < 0.05). Histological examination showed that compared with the model group, a lot of inflammatory cells and Schwann cells were found at sciatic nerve stump in the LPS group at 7th day after operation. ORO staining showed that the degree of demyelination in the LPS group was higher than that in the model group. LFB staining showed that the sciatic nerve stump demyelination appeared in both model group and the LPS group at 7th day after operation, but compared with the model group, myelin debris clearance in the LPS group was significantly accelerated (P < 0.05). Finally, compared with the model group, the SFI in the LPS group was increased significantly at 20 d after surgery (P < 0.05). Conclusions The results confirm that LPS is possible to manipulate the innate immune response to accelerate myelin clearance during Wallerian degeneration after early peripheral nerve injury in rats.

Abstract:Objective To provide a basis for clinical diagnosis, a serum metabonomic dynamic study was carried out on the Tg2576 mouse model at different stages of Alzheimer's disease (AD) whose pathological progress is similar to that of human AD patients. Methods Serum samples of Tg2576 mice were collected at the early(6 months) and late(12 months) stages of Alzheimer's disease.The ¹H NMR spectra of the serum samples were collected and the metabolic characteristics were analyzed by multivariate analysis.Results Significant differences in serum metabonomics were found in the transgenic Tg2576 mice and C57 mice at 6 and 12 months of age, and there were significant metabolic changes in Tg2576 mice at different stages of Alzheimer's disease. Compared with C57 mice, the Tg2576 mice at early stage of Alzheimer's disease showed higher levels of serum lactate,myo-inositol and amino acids (such as leucine, isoleucine, alanine), and lower levels of lipids, choline, phosphorylcholine, glycerol phosphorglcholine, betaine, glycine and glucose.At the late stage of Alzheimer's disease, the transgenic Tg2576 mice had higher levels of lactate, myo-inositol and alanine,while the serum levels of lipids, choline, phosphorylcholine, glycerophosphorylcholine, betaine, and glycine continued to drop. Meanwhile glutamine and creatine levels started to decline. By comparing the early and late serum metabolites of Alzheimer's disease, serum metabonomic profiles of the late stage of Alzheimer's disease indicated an up-regulation of lactate, myo-inositol and alanine, and a down-regulation of lipids,choline, phosphorylcholine and glycerophosphorylcholinelevels.Moreover, the levels of lactate, lipids, choline, phosphorylcholine and glycerophosphorylcholine showed statistical significance at the early stage of AD, and they were closely correlated with the severity of Alzheimer's disease. Conclusions The above results show that the changes of lactate, myo-inositol and alanine are positively-correlated with the development of AD, while the serum levels of lipids, choline, phosphorylcholine and glycerophosphorylcholine are inversely-proportional to the severity of AD. These metabolites are dynamically and progressively changed along with the disease progression, which hopefully may serve as early metabolic markers for the diagnosis of AD in clinical practice.

Abstract:Objective To observe and identify the microorganism isolated from diseased and dead Ctenogobius gymnauchen cultured in seawater near the Daya Bay of south China sea. Methods GDLAMI-1210 strain was isolated from the diseased Ctenogobius gymnauchen (Bleeker). We applied physiological and biochemical characteristics in the bacterial classification. In order to confirm the results, we amplified a 1438 bp sequence of GDLAMI-1210's 16 S rRNA(HM 362434)and compared with other sequence in GenBank, and followed by artificial infection. Results The GDLAMI-1210 strain was Gram-negative and in a shape of short rod with single polar flagellum. The homology analysis and phylogenetic study showed that the 16 S rRNA sequence of GDLAMI-1210 has the highest similarity to Vibrio sp. espec Vibrio vulnificus, showing 99% identity. Conclusions To our knowledge, this is the first report that the causative pathogen, Vibrio sp, leads to the mortality of Ctenogobius gymnauchen (Bleeker).