Establishment of a RILD model in C57BL/6J mice treated with precise radiotherapy / 中国辐射卫生

Objective To provide a reliable and stable animal model for investigating the molecular pathogenesis of radiation-induced liver disease (RILD). Methods Ninety C57BL/6J mice were divided into control, 20 Gy, 25 Gy, 30 Gy and 35 Gy radiation groups. The mice were executed at 4 weeks after radiation and the levels of alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase in the liver serum were measured. HE staining was performed on the pathological liver tissues. Masson staining was performed at 36 weeks after radiation. Results Compared with the control group, the fatality rate was higher in the 30 and 35 Gy radiation groups, and the body weight significantly decreased in the 20 and 25 Gy radiation groups. Compared with the control group, alanine aminotransferase significantly increased in mice exposed to 20 Gy, while aspartate aminotransferase and alanine aminotransferase increased in mice exposed to 25 Gy. No significant changes were observed in the livers of the mice in the 20 and 25 Gy radiation groups, but pathological examination showed liver damage induced by both 20 and 25 Gy radiation. Conclusion A stable and reliable mouse model of RILD was constructed for treatment with linear accelerator. The mouse model of RILD constructed for stereotactic body radiation therapy using linear accelerator has significant research implications for the exploration of RILD.

Effects of modified Zhujing Pill on mitochondrial dynamics related protein in mice with form deprivation myopia / 国际眼科杂志(Guoji Yanke Zazhi)

AIM: To investigate the effects of modified Zhujing Pill on the mitochondrion structure and dynamin-related protein of retinal pigment epithelial cells(RPEs)in mice with form deprivation myopia.METHODS: 3-week-old C57BL/6J mice were randomly divided into control group, model group and Chinese medicine group, with 10 mice in each group. Myopia model of the right eye of mice was established by means of form deprivation in model and Chinese medicine groups. After 4wk, the Chinese medicine group were given intragastric administration of modified Zhujing Pill suspension 0.546g/(kg·d)(0.2mL/d)for 4wk, and same amount of saline was given to mice in other groups at the same time of modeling. The axial length and diopter of the right eye of the mouse were measured before and after the experiment by A-ultrasound and a strip retinoscope respectively. At the end of the experiment, the mitochondrial ultrastructure of RPEs was observed by transmission electron microscope. Western blot, and real-time fluorescent quantitative PCR(q-PCR)were used to detect quantitative and gene expression of mitofusin 1(MFN1), optic atrophy 1(OPA1), and dynamin-related protein 1(DRP1)in retinal tissues respectively.RESULTS: At the beginning of the experiment, there was no statistically significant difference in axial length and diopter of the right eye of the mouse in control, model and Chinese medicine groups(P>0.05). At the end of the experiment, compared with the control group, the mice in the model group and the Chinese medicine group had lower diopter and continuously prolonged axial length(all P<0.05), while the mice in the Chinese medicine group had significantly shorter axial length and higher diopter than the model group(all P<0.05). Western blot and q-PCR results showed that the relative expression of MFN1 and OPA1 decreased and DRP1 increased in both the model group and the Chinese medicine group compared with the control group(all P<0.05), and the relative expression of MFN1 and OPA1 increased in the Chinese medicine group compared with the model group(all P<0.05). The electron microscopic results showed that the mitochondria in the right retina of the mice were only mildly swollen in the Chinese medicine group, while the mitochondria in the model group were obviously swollen and disordered and empty.CONCLUSION: Modified Zhujing Pill could protect the retinal mitochondria by regulating the key proteins of mitochondrial dynamics(MFN1, OPA1, and DRP1), and it has a protective effect on the retina of axial myopic mice.

Effect of gender difference on antigen induced Sjogren's syndrome model in mice / 中国药理学通报

Aim To study the effect of gender differences in C57BL / 6J mice on antigen induced Sjogren's syndrome(SS)model. Methods The submandibular gland protein of C57BL/6J female and male mice was extracted and mixed with the same amount of Freund's complete adjuvant(FCA)for the first three times, the antigen concentration was adjusted to 2.5 g·L-1, mixed with Freund's incomplete adjuvant(FIA)for the fourth time, and the same-sex mouse antigen was injected into the back of mice for a total of four times to induce the mouse SS model. The mouse SS model was induced by multi-point intradermal injection of antigen on the back of mice for four times,the body weight of female and male mice was measured every week, the general condition was observed, the saliva volume of mice was measured at the sixth week of modeling. After the mice were sacrificed, the pathological changes of submandibular gland and the changes of T and B lymphocyte subsets in spleen were detected, and the differences in SS model preparation between female and male mice were compared. Results The SS model of male and female mice was successfully established, and there was no significant difference in general condition, saliva volume, submandibular gland pathology, plasma cells and memory B cells between male and female SS mice. The success rate of SS model was 75% in female mice and 60% in male mice. Compared with normal mice of the same sex, the weight loss of female SS mice was earlier and more obvious than that of male SS mice; the submandibular gland index of male mice was significantly higher than that of female mice. Compared with normal mice of the same sex, the proportion of Th17 and Treg cells in spleen of female SS mice was more statistically significant than that of male SS mice. Conclusions The success rate of SS modeling in female mice is higher than that in male mice. Compared with male SS mice, female SS mice have more significant SS like manifestations and pathological manifestations, which can provide a reference basis for the selection of gender when establishing SS model.

Effects of Total Flavonoids from Chamomile on Lipid Metabolism of Hyperlipidemia Model Mice and Its Mechanism / 中国药房

OBJECTIVE：To st udy t he effects of total flavonoids from chamomile on lipid metabolism of hyperlipidemia model mice and its potential mechanism. METHODS ：Thirty male C 57BL/6J-ApoE－/－ mice were randomly divided into model group ， positive control group（fenofibrate 30 mg/kg）and chamomile total flavonoids low-dose ，medium-dose and high-dose groups （88， 176，352 mg/kg），with 6 mice in each group. In addition ，6 male C 57BL/6J mice were used as normal control group. Mice in normal control group were fed with ordinary diet ，and mice in other groups were fed with high-fat diet for 8 weeks to replicate hyperlipidemia model. At the time of making model ，administration groups were given relevant liquid （using 1％ sodium carboxymethyl cellulose as solvent ）；normal control group and model group were given 1％ sodium carboxymethyl cellulose intragastrically，200 mL per gavage ，once a day ，for consecutive 8 weeks. The body weight of mice in each group was weighed before medication and 8 weeks after medication. The serum contents of total cholesterol （TC），triacylglycerol（TG），low-density lipoprotein cholesterol （LDL-C），high-density lipoprotein cholesterol （HDL-C），aspartate aminotransferase （AST）and alanine aminotransferase （ALT） in mice were detected after last administration ；the contents of superoxide dismutase （SOD） and malondialdehyde（MDA）as well as the protein expressions of peroxisome proliferator-activated receptor α（PPARα），carnitine palmityl transferase 1A（CPT1A）and peroxase acyl-CoA oxidase 1（ACOX1）in liver tissue were determined. The pathological changes i n liver tissue were observed. RESULTS：Compared w ith before medication ，the body weight of each group showed an increasing trend after 8 weeks of medication. Compared with normal control group ，body weight ，the contents of TC ，TG， LDL-C，AST and ALT in serum and MDA content in live r lan- tissue of mice in model group were significantly increased wei516@sina.com after 8 weeks of medication （P＜0.05 or P＜0.01）. The ·2706· China Pharmacy content of HDL-C in serum and the cont ent of SOD in liver tissue ，as well as the protein expressions of PPARα，CPT1A and ACOX1 were significantly decreased （P＜0.05 or P＜0.01），and the structure of liver tissue was disorganized ，with circular fat vacuoles of different sizes and lipid droplets of different sizes in the cytoplasm. Compared with model group ，body weight （except for chamomile total flavonoids low-dose group ）of mice ，serum contents of TC ，TG，LDL-C，AST and ALT ，content of MDA in liver tissue （except for chamomile total flavonoids low-dose and medium-dose groups ）were significantly decreased （P＜0.05 or P＜ 0.01）. Serum content of HDL-C ，content of SOD in liver tissue ，protein expressions of PPARα，CPT1A（except for chamomile total flavonoids low-dose and medium-dose groups ） and ACOX 1 were significantly increased （P＜0.05 or P＜0.01）；liver tissue structure was clear ，and liver fat vacuoles were improved to varying degrees ，and less lipid droplets. The improvement effect of the above indexes was the best in the chamomile total flavonoids high-dose group. CONCLUSIONS ：Chamomile total flavonoids can prevent the occurrence of hyperlipidemia in C57BL/6J-ApoE －/－ mice，the mechanism of which may be associated with up-regulation of PPARα expression，the improvement of liver injury and oxidant stress injury.

Effect of modified Zhujing pill on retinal thickness and apoptosis in form deprivation myopia mice / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To investigate the effect of modified Zhujing pill on retinal thickness and apoptosis in form deprivation myopia(FDM)mice.<p>METHODS: Totally 72 C57BL/6J mice aged 3-week-old were randomly divided into control group 1, model group 1, intervention group 1, control group 2, model group 2 and intervention group 2, with 12 mice in each group. The first three groups were tested for 3wk and the last three groups were tested for 6wk, except for the groups of control 1 and control 2, all the mice used translucent goggles to cover their right eyes for form deprivation. The mice of intervention 1 and intervention 2 were respectively given intragastric administration modified Zhujing pill suspension 0.546g/(kg·d)(0.1mL/d)for 3wk at the beginning and after 3wk of the experiment. Same amount of saline was given to mice in other groups at the same time of modeling. The eye axis was measured before and after the experiment. At the end of the experiment, the eye of mice was taken for HE staining to observe the thickness changes of each layer of retina. Immunohistochemistry(IHC)and western blotting(WB)were used to detect Bcl-2 and Caspase3 expression of protein.<p>RESULTS: At the end of the experiment, the axis of model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), the axis of intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), and the axis of model group 2 was significantly higher than that of control group 2(<i>P</i><0.01), the eye axis of intervention group 2 was significantly lower than that of model group 2(<i>P</i><0.01); HE staining showed that the thickness of NFL and ONL of model group 1 was significantly thinner than that of control group 1(<i>P</i><0.01). The thickness of INL of model group 1 was significantly thinner than that of control group 1(<i>P</i><0.05), and the thickness of NFL, INL and ONL of intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.05); The thickness of NFL, INL and ONL model group 2 was significantly thinner than that of control group 1(<i>P</i><0.01); IHC testing showed that the average optical density of Bcl-2 protein in model group 1 was significantly lower than that of control group 1(<i>P</i><0.05), which in intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.01), and which in the average optical density of Bcl-2 protein of model group 2 was significantly lower than that of control group 2(<i>P</i><0.01), which in intervention group 2 was significantly higher than that of model group 2(<i>P</i><0.01); Caspase 3 protein average optical density of model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), which in model group 2 was significantly higher than that of control group 2(<i>P</i><0.05), which in intervention group 2 was significantly lower than that of model group 2(<i>P</i><0.01); WB test proved that the relative expression level of Bcl-2 protein in model group 1 was significantly lower than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly higher than that of model group 1(<i>P</i><0.01), and which in model group 2 was significantly lower than that of control group 2(<i>P</i><0.01), which in intervention group 2 was significantly higher than that of model group 2(<i>P</i><0.01); The relative expression level of Caspase3 protein in model group 1 was significantly higher than that of control group 1(<i>P</i><0.01), which in intervention group 1 was significantly lower than that of model group 1(<i>P</i><0.01), the intervention group 2 was significantly lower than that of model group 2. <p>CONCLUSION: The results show that the modified Zhujing pill can interfere with the pathological changes of retinal thickness thinning in the process of myopia and formed myopia mice by regulating the expression of apoptosis-related proteins Bcl-2 and Caspase3, and alleviating the apoptosis of retinal cells in myopia formation and myopia mice.

Lotus leaf flavonoids induce apoptosis of human lung cancer A549 cells through the ROS/p38 MAPK pathway

BACKGROUND: Leaves of the natural plant lotus are used in traditional Chinese medicine and tea production. They are rich in flavonoids. METHODS: In this study, lotus leaf flavonoids (LLF) were applied to human lung cancer A549 cells and human small cell lung cancer cells H446 in vitro to verify the effect of LLF on apoptosis in these cells through the ROS/p38 MAPK pathway. RESULTS: LLF had no toxic effect on normal cells at concentrations up to 500 µg/mL, but could significantly inhibit the proliferation of A549 cells and H446 cells. Flow cytometry showed that LLF could induce growth in A549 cells. We also found that LLF could increase ROS and MDA levels, and decrease SOD activity in A549 cells. Furthermore, qRT-PCR and western blot analyses showed that LLF could upregulate the expression of p38 MAPK (p-p38 MAPK), caspase-3, caspase-9, cleaved caspase-3, cleaved caspase-9 and Bax and downregulate the expression of Cu/Zn SOD, CAT, Nrf2, NQO1, HO-1, and Bcl-2 in A549 cells. Results of HPLC showed that LLF mainly contain five active substances: kaemp-feritrin, hyperoside, astragalin, phloridzin, and quercetin. The apoptosis-inducing effect of LLF on A549 cells came from these naturally active compounds. CONCLUSIONS: We have shown in this study that LLF is a bioactive substance that can induce apoptosis in A549 cells in vitro, and merits further research and development.

Enhancement of cellular activity in hyperglycemic mice dermal wounds dressed with chitosan-alginate membranes

The use of specially designed wound dressings could be an important alternative to facilitate the healing process of wounds in the hyperglycemic state. Biocompatible dressings combining chitosan and alginate can speed up wound healing by modulating the inflammatory phase, stimulating fibroblast proliferation, and aiding in remodeling phases. However, this biomaterial has not yet been explored in chronic and acute lesions of diabetic patients. The aim of this study was to evaluate the effect of topical treatment with a chitosan-alginate membrane on acute skin wounds of hyperglycemic mice. Diabetes mellitus was induced by streptozotocin (60 mg · kg-1 · day-1 for 5 days, intraperitoneally) and the cutaneous wound was performed by removing the epidermis using a surgical punch. The results showed that after 10 days of treatment the chitosan and alginate membrane (CAM) group exhibited better organization of collagen fibers. High concentrations of interleukin (IL)-1α, IL-1β, granulocyte colony-stimulating factor (G-CSF), and tumor necrosis factor-alpha (TNF-α) were detected in the first and second days of treatment. G-CSF and TNF-α level decreased after 5 days, as well as the concentrations of TNF-α and IL-10 compared with the control group (CG). In this study, the inflammatory phase of cutaneous lesions of hyperglycemic mice was modulated by the use of CAM, mostly regarding the cytokines IL-1α, IL-1β, TNF-α, G-CSF, and IL-10, resulting in better collagen III deposition. However, further studies are needed to better understand the healing stages associated with CAM use.

Co-transplantation of adipose mesenchymal stem cells and endothelial progenitor cells in ulcerative colitis mice / 中国组织工程研究

BACKGROUND: Mesenchymal stem cell transplantation has been proved to be effective for ulcerative colitis, while the effect of endothelial progenitor cells in ulcerative colitis treatment is still unknown. They are both promising cells for tissue engineering, which can be used for cell transplantation. OBJECTIVE: To explore whether co-transplantation of endothelial progenitor cells and adipose mesenchymal stem cells can relieve ulcerative colitis in a mouse model. METHODS: C57BL/6J mice were randomly divided into six groups (n=24 per group): co-transplantation group, mesenchymal stem cell transplantation group, endothelial progenitor cell transplantation group, glucocorticoid treatment group, transplantation control group and normal control group. Murine ulcerative colitis model was established in all groups except for the normal control group. At 7 and 10 days after modeling, transplantation groups were respectively injected via tail vein with adipose mesenchymal stem cells and/or endothelial progenitor cells, glucocorticoid or PBS. Mice were sacrificed at 12 days after modeling. Colon length, disease activity index, histological score and the serum level of tumor necrosis factor-α were compared between groups. RESULTS AND CONCLUSION: Treatment with glucocorticoid was significantly effective for ulcerative colitis relative to the transplantation control group (P 0.05). Co-transplantation of adipose mesenchymal stem cells and endothelial progenitor cells was better than the other treatments, which significantly improved the shortening of the colon, disease activity index, histological score, and serum level of tumor necrosis factor-α.

Effect of cold exposure on browning of white adipose tissues in mice fed with high fat diet / 中国药理学通报

Aim: To observe the effect of cold exposure on browning of white adipose tissues in mice fed with high fat diet. Methods: Male 8-week-old C57BL/6J mice were randomly divided into HFD + 5 °C group and HFD + RT group, after 8 weeks of high-fat diet. Male 8-week-old C57BL/6J mice were randomly divided into normal + 5 °C group and normal + RT group, after 8 weeks of normal diet. Each group of mice were intervened for 2 h at different temperatures in the same time period for 8 weeks. Parameters, including body weight, food intake, inguinal white adipose tissue (iWAT) mass, brown adipose tissue (BAT) mass, blood glucose, blood lipids, leptin, adiponectin, adipose tissue pathology and in situ expression of uncoupling protein 1(UCP1) and prohibitin protein (PHB) in iWAT and BAT. Results: Compared with HFD + RT group, cold exposure not only significantly reduced body weight, blood glucose, iWAT weight/body weight ratio, TC, TG, LDL-C and leptin, but also increased food intake and BAT weight in high-fat diet mice. HE staining showed that iWAT and BAT cells became smaller and had multiple compartments. The iWAT had a browning trend. Immunohistochemistry showed that UCP1 and PHB protein in iWAT and BAT significantly increased. Conclusions: Cold exposure can counteract the weight gain caused by a high-fat diet, which may be related to the activation of brown adipose tissue and the induction of browning of white adipose tissues, increasing heat production and reducing white fat accumulation.

Generation of a novel HBeAg transgenic mice using CRISPR/Cas9 technique / 南方医科大学学报

OBJECTIVE@#To generate a new strain of HBeAg transgenic mice using CRISPR/Cas9 technique.@*METHODS@#Hepatitis B virus (HBV) HBeAg gene was cloned and inserted in the pliver-HBeAg expression frame at the site of Rosa26 gene using CRISPR/Cas9 and homologous recombination techniques to construct the pliver-HBeAg expression vector containing HBeAg gene. The linear DNA fragment containing HBeAg gene was obtained by enzyme digestion. Cas9 mRNA, gRNA and the donor vector were microinjected into fertilized eggs of C57BL/6J mice, which were then transplanted into the uterus of C57BL/6J female surrogate mice to obtain F0 generation mice. The F0 generation mice were identified by long fragment PCR to obtain F0 transgenic mice with HBeAg gene. The positive F0 generation mice were bred with wild-type C57BL/6J mice to produce the F1 mice, which were identified by PCR and sequencing. The positive F1 transgenic mice carrying HBeAg gene were backcrossed until the homozygous offspring transgenic mice were obtained. The genotypes of the offspring mice were identified. The expressions of HBeAg and HBeAb in the heterozygous and homozygous HBeAg transgenic mice were detected by automatic chemiluminescence immunoassay, immune colloidal gold technique and immunohistochemistry method.@*RESULTS@#A total of 56 F0 mice were obtained, and 2 of them carried homologous recombined HBeAg gene. Six positive F1 mice were obtained, from which 22 homozygous and 29 heterozygous F2 generation HBeAg transgenic mice were obtained. High concentration of HBeAg protein was detected in the peripheral blood of all the positive HBeAg transgenic mice without HBeAb expression. HBeAg expression was detected in the hepatocytes of HBeAg transgenic mice.@*CONCLUSIONS@#We obtained a new strain of HBeAg transgenic mice with stable expression of HBeAg in the hepatocytes and immune tolerance to HBeAg using CRISPR/Cas9 technique, which provide a new animal model for studying HBV.

Animal Model of Lung Adenoma in C57BL/6J Mice Induced by Urethane / 中国药学杂志

OBJECTIVE: To compare and optimize the animal model of lung cancer on C57BL /6J mice by three different experimental methods. METHODS: Female C57BL /6J mice were divided into four groups: control group, model group , Ⅱ and III. The mice in model groupreceived 800 mg•kg-1 urethane by intraperitoneal injection twice a week, followed by 5 weeks. The mice in model group Ⅱ were injected intraperitoneally with 1 000 mg•kg-1 urethane in the same way with model group . The mice in model group III received urethane at the dose of 1 000 mg•kg-1 intraperitoneally, every 5 day continuing for 8 weeks. The control group was given equal volume of sodium chloride solutions in the same way as the model group Ⅱ. The main organs of all the groups were taken such as heart, liver, spleen, lung, kidney, brain and thymus at the 28th week, and their organs index was calculated. At the same time the lung tissues were examined to get the amount of adenomas and the diameters, and developed the lung and thymus histopathological examination. The bronchial alveolar lavage fluids were collected to classify the cells of the control and experimental groups. RESULTS: Compared with the control group, all the three experimental groups could induce lung adenomas absolutely, the number of tumors in model groupand III was 6, while tumors in model Ⅱ group was about 2. In the three experimental groups the index of lung increased and the brain, spleen and thymus decreased compared with the control group. The bronchial alveolar lavage fluids analysis showed that the number of lymphocytes and polymorphonuclear leukocytes in the model group are more than that in control group. CONCLUSION: All the three experimental groups could induce lung adenomas absolutely. Compared with model group Ⅱ, the model III and are more stable and homogeneous with the similar results of experiment.

Establishment of a chronic rhinosinusitis model in C57BL/6J mice / 医学研究生学报

Objective Streptococcus pneumoniae is one of the main pathogenic factors of chronic rhinosinusitis (CRS). This study was to explore the method of establishing a CRS model in C57BL/6J mice.Methods Thirty C57BL/6J mice were equally randomized into a CRS model, a sham operation and a blank control group. The CRS model was made by implanting type-Ⅲ streptococcus pneumoniae in the maxillary sinuses, the mice of the sham operation group underwent opening and suturing of the maxillary sinuses without implantation of any streptococcus pneumonia, while the blank controls did not receive any operation. Two months later, all the mice were sacrificed and the nasal sinus mucosal tissue was harvested, embedded with paraffin, sectioned, and subjected to HE staining, followed by observation of the pathological changes under the microscope.Results The CRS model mice exhibited significant proliferation and disorderly arrangement of epithelial cells in the nasal mucosal tissue, with degeneration, necrosis, exfoliation, ulceration, goblet cell proliferation, gland and tissue hyperplasia, and infiltration of lymphocytes, plasmocytes and monocytes. No obvious inflammation was observed in the sham operation and blank control groups. The pathology index score of the nasal sinus mucosal tissue was significantly higher in the CRS model than in the sham operation and blank control groups (\[14.800±5.200\] vs \[2.000±2.906\] and \[1.800±2.098\], P0.05).Conclusion The CRS model was successfully established in mice, which has provided some reference for the construction of the CRS model in animals.

Establishment of a C57BL/6 J mouse model of metastatic melanoma in the lung / 中国实验动物学报

Objective To investigate the influencing factors involved in the establishment of a C57BL/6 J model of metastatic melanoma in the lung,including the way of tumor inoculation,the number of inoculated cells and the time of tumor formation. Methods Mouse melanoma B16F10 cells were cultured in vitro. 1)Eighteen healthy male C57BL/6 J mice were randomly divided into three groups. Mice in each group received 100 μL cell suspension(including 3 ×106 melanoma cells)via intravenous,intraperitoneal and subcutaneous injection,respectively. After two weeks,the mice were killed and dissected,and the tumor growth and metastasis were observed. 2)Eighteen male mice were randomly divided into three groups. Mice in each group were injected with 3 ×106cells,1 ×106cells, and 3 ×105cells through the tail vein,respectively. After two weeks,mice were killed and dissected,and the tumor growth and metastasis were observed. 3)Eighteen male mice were randomly divided into three groups. Mice in each group were injected with 1×106cells though the tail vein. Mice were killed and dissected after one week, two weeks and three weeks, respectively. The growth and metastasis of tumor were observed. Results 1)The success rate of lung metastasis was 100% in the mice with intravenous injection,but not in mice receiving intraperitoneal injection and subcutaneous injection. 2)The size of metastatic melanoma nodules were moderate in mice inoculated by 1 ×106cells. The number of melanoma metastatic foci was too high in the mice inoculated with 3 ×106cells,but too low in the mice inoculated with 3 ×105cells. 3)Significant metastatic melanoma foci were observed in the mice killed and dissected after two weeks with no death. The number of melanoma foci in the lung was too high in the mice killed after three weeks,while was too low in the mice killed at one week after tumor cell inoculation. Conclusions Intravenous injection of 1×106mouse melanoma cells into C57BL/6 J mice and killed after two weeks is an optimal method for establishment of a mouse model of metastatic melanoma in the lung, and is worth of recommendation.

The beneficial effect of metformin on beta-cell function of type 2 diabetes mice and its possible mechanisms / 药学学报

To investigate the effects of metformin on pancreatic β-cell function and its possible mechanism, high fat diet-induced type 2 diabetic C57BL/6J mice were divided into two groups according to fasting blood glucose (FBG), glucose decreasing rate at 40 min of insulin tolerance test, triglycerides (TG), cholesterol (CHO) and body weight (BW). The C57 mice were gavaged with water or metformin for 58 days. β-Cell function was evaluated by oral glucose tolerance test and hyperglycemic clamp. Genes and proteins related to pancreas proliferation, lipid metabolism and endoplasmic reticulum stress were investigated. Compared with the model group, metformin group exhibited a reduction in the body weight (PPPPPPdx-1, Pβ (Lxr-β, PPPP<0.05) were also down-regulated. These results suggest that metformin could improve the insulin secretion function of type 2 diabetic C57BL/6J mice. The mechanism of the action may rely on its improvement of pancreas cell proliferation, lipid metabolism and amelioration of endoplasmic reticulum stress.

An experimental study of effect of curcumin on improvementof bone microarchitecture and bone quality inhigh-fat-diet C57BL/6J mice and its association with cathepsin K / 中国药理学通报

Aim To investigate the effect of curcumin against high-fat-diet induced C57BL/6J mice bone changes and the correlation between the expression of cathepsin K and curcumin.Methods Curcumin treated C57BL/6J mice had been on high fat diet for 12 weeks.The HE, Alizarin red S staining and Safranin O/fast green staining of femur were employed to evaluate bone microstructure, bone metabolism and bone development.The expressions of cathepsin K were assessed by Western blot and immunohistochemical staining.Results Histopathological results showed that curcumin could improve the destruction of trabecular bone structure, cartilage development and bone calcification.Biomechanical results proved that curcumin could improve the bone strength of the type 2 diabetic mice induced by high fat.The results of immunohistochemistry and Western blot assay indicated that curcumin could significantly inhibit the expression of cathepsin K in bone tissues of mice.Conclusion Curcumin can increase bone strength, improve bone microstructure, and enhance the degree of bone calcification, which may be achieved by inhibiting the expression of cathepsin K.

Establishment of a mouse model of aorta dissection induced by β-aminopropionitrile drinking combined with angiotensin II infusion / 中国实验动物学报

Objective To establish a mouse model of aorta dissection (AD) by β-aminopropionitrile (BAPN) in drinking water + subcutaneously pumped angiotensin II (Ang II) infusion.Methods Forty 3-week-old C57B1/6J male mice were randomly divided into two groups.All animals received 0.1 g/kg/d BAPN in drinking water for 4 weeks.Then the BAPN drinking + saline infusion group and BAPN drinking + Ang II infusion group received continuous saline or Ang II (1,000 ng/kg/min) infusion, respectively, via subcutaneous osmotic minipump for 72 hour.The mice were restricted in a noninvasive computerized tail-cuff system and their arterial systolic blood pressure and heart rate were monitored.Autopsy was performed if a mouse died during the experiment.At the end of the experiment, mice were sacrificed by injection with an overdose of sodium pentobarbital and the aortas were harvested.The formation of aortic false lumen was observed by pathology using hematoxylin-eosin staining.Results The overall incidence of AD in the BAPN drinking administration +Ang II infusion group was 95%, whereas the incidence of AD in the BAPN drinking administration +saline infusion group was only 5%.The mortality from dissecting aneurysm rupture was 24% in the BAPN drinking administration +Ang II infusion group during the experiment.Pathological examination of the aortic cross-sections clearly showed the formation of blood-filled false lumens induced by Ang II.Conclusions A mouse model with high incidence of aortic dissection is successfully established.

A simplified method for isolation and culture of primary mouse hepatocytes / 中国比较医学杂志

Objective To simplify and optimize the method for isolation and culture of primary mouse hepatocytes on the basis of conventional extraction method,and to provide a reference for related research. Methods Mouse liver was reversely perfused with the isolation solution(i.e.,through the vena cava inferior in,and portal vein out),cut into small pieces and digested with enzymes. Then the hepatocytes were isolated by density gradient centrifugation and transferred into culture medium. The cell viability was detected by trypan blue staining. The purity of the hepatocytes was analyzed by flow cytometry and the cell morphology was observed with an inverted microscope. Results The hepatocytes obtained by this improved method showed high viability and purity,with typical characteristics of cell morphology. Conclusions The liver perfusion is facilitated by reversed perfusion, and the isolated hepatocytes are with high viability and purity confirmed by many times of experiments. This optimized procedure is an easy and efficient method for isolation of primary mouse hepatocytes.

Protection of cannabinoid to retinal ganglion cells against oxygen-glucose deprivation damage / 中华实验眼科杂志

Background Acute retinal ischemia anoxic injury is common in eye disorders,such as acute glaucoma,central retinal artery occlusion and ischemic optic neuropathy,etc.This will cause retinal ischemia anoxic injury and induce retinal ganglion cells (RGCs) death in addition.Endogenous cannabinoid (CB) and its receptors are involved in the central nervous system injury,ischemia,inflammation,and poisoning and other physiological and pathological process.Objective This study was to investigate the effect of CB on RGCs damage induced by oxygen-glucose deprivation (OGD).Methods The eyeballs were obtained from 6-week-old normal C57BL/6J mice to prepare retinal frozen sectionsand the expression and distribution of cannabinoid receptors (CB1R and CB2R) in RGCs was detected by immunofluorescence staining.The eyeballs of ten newborn C57BL/6J mice (postnatal 0-3 days) were obtained after immersed by 75％ alcohol and the retinas were isolated in preeooling DMEM for the primary culture of RGCs.The cells were identified by detecting the expression of Brn3a,a marker of RGCs,with immunofluorescence staining.Then the cells cultured for 14 days were divided into normal control group (in complete culture medium+95％ air+5％ CO2) and OGD group (in glucose-free medium+95％ N2 +4％ CO2 + 1％ O2) for 20 hours.The mitochondrial damage and RGCs morphology changed were evaluated by JC-1 staining to observe the mitochondrial membrane potential change.SR141716A (CB1R antagonist,1 μmol/L),SR144528 (CB2R antagonist,1 μmol/L) and 5 or 10 μmol/L WIN 55212-2 (CB1R and CB2R agonist) were added,and the survival rate of RGCs was assayed MTT.Results CBR was positively expressed in various layers of normal mouse retinas.The cells in the normal control group showed uniform size and polygon in shape with the long and thin axons,and the expression of Brn-3a was seen in the cells.However,in the OGD group,cell shrinkage and fragments were found and most of the axons disappeared.The expression of Brn-3a was evidently weakened.The fluorescence intensity of JC-1 was evidently weakened in the OGD group compared with the normal control group,showing the reduce of mitochondrial membrane potential.MTT assay showed that the survival rate of RGCs was (100.00± 13.87)％,which was significantly higher than (89.52-± 18.16)％ in the normal control group (q =8.065,P =0.008).The mean survival rates of RGCs were (116.63±22.21)％ and (112.61 ±19.02)％ in the cells treated by SR141716A and SR144528,and that in the normal cells was (89.52 ± 18.16)％ in the OGD group,with significant differences between SR141716A-or SR144528-treated cells and normal cells (q =29.780,17.391;both at P＜ 0.01).Conclusions Hypoxia and glucose-free up-regulate the expression of CB and activate CB pathway.Inhibition of activation CBR process has a neuroprotection effect under the Hypoxia and glucose-free condition.

Effects of fermented blueberry liquid in high-fat diet-induced obese C57BL/6J mice / 한국영양학회지

PURPOSE: The objective of the present study was to determine whether fermentation can increase the protective effects of blueberry liquid in a high-fat diet-induced obese mice model. METHODS: Male C57BL/6J mice were fed a high-fat diet (HD, 60% fat, w/w,), HD supplemented with 10 ml/kg BW/day of blueberry liquid (BHD, blueberry high-fat diet), or HD supplemented with 10 ml/kg BW/day of fermented blueberry liquid (FBHD, fermented blueberry high-fat diet) for 10 weeks. RESULTS: There were significant decreases in the body, epididymal adipose tissue, and liver weights of blueberry-fed groups compared to HD, whereas there were no significant differences in food intake among the groups. Furthermore, blueberry liquid groups, especially fermented blueberry liquid, significantly attenuated the contents of hepatic triglycerides and total cholesterol induced by HD. Serum LDL-cholesterol was significantly lower in the BHD and FBHD-fed groups, whereas FBHD significantly increased the serum HDL-cholesterol level compared to the control. Concentrations of aspartate transaminase, alanine transaminase, and leptins in serum were also reduced by blueberry liquid supplementation. The mRNA expression of hepatic acetyl CoA carboxylase was significantly reduced in both the BHD and FBHD groups compared to HD. Furthermore, FBHD altered the mRNA expression level of hepatic lipolysis genes. CONCLUSION: In conclusion, these results suggest that blueberry, especially fermented blueberry liquid, may improve obesity-related abnormalities.

Mensurement and analysis of biological characteristics of C57BL/6J mice and their offsprings after frozen-thawed embryo transfer of C57BL/6J mice / 中国比较医学杂志

Objective To investigate the changes of average body weight gain and serum biochemical indexes of C57BL/6J mice (B6 mouse) and their offspings after frozen-thawed embryo transfer of B6 mice.Methods The mice were divided into three groups in this study.In the experimental group I (E-I,30 males and 20 females),2-cell embryos after in-vitro fertilization were collected,and cryopreserved by EFS method,then obtained the offsprings after transplantation of the recovered embryos to oviduct of recipient mice (ICR mouse).In the experimental group II (E-II,26 males and 17 females),when the mice from E-I grew to maturity,the offsprings were obtained from natural mating of mice from E-I.In the control group (20 males and 20 females),the offsprings came from conventional feeding and natural mating.The three groups of mice were raised to 16 weeks old,weighing the body weight at a regular time intervals,and the serum biochemical indexes were obtained from 16-week-old mice.Then the changes of average body weight and serum biochemical indexes of the mice were analyzed.Results The average body weight of E-I mice was significantly higher than that of control group at each week-age (P<0.01).The average body weight of E-II female mice was significantly higher than that of the control group in 12-16-week old mice (P<0.01),but the average body weight of E-II male mice showed no significant differences compared with the control group except for few weeks.The serum biochemical indexes of E-I and E-II mice were changed in all items except for AST,TP and Ca.Conclusions There are some effects on the average body weight gain and serum biochemical indexes of C57BL/6J mice and their offspings after frozen-thawed embryo transfer.