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  • Table of Content
      28 May 2020, Volume 36 Issue 3 Previous Issue    Next Issue
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    Original Articles
    Spermine attenuates high glucose-induced myocardial fibrosis by regulating the cell cycle
    HU Jing, LI Fa-dong, SHAO Xiao-ting, LI Si-wei, ZHANG Xin-ying, ZHAO Bing-bing, XU Chang-qing, WEI Can
    CJAP. 2020, 36 (3): 193-196.   DOI: 10.12047/j.cjap.5952.2020.043
    Abstract   PDF (1236KB) ( 327 )
    Objective: To investigate the protective effect of spermine (Sp) on diabetic cardiomyopathy (DCM) and high glucose-induced cardiac fibroblasts (CFs), and to explore its mechanism.Methods: ①Animal experiments: 24 male Wistar rats were randomly divided into control group, type 1 diabetes group (TID) and spermine group (TID+Sp, each group n=8). TID rats were induced by streptozocin (STZ, 60 mg/kg), and TID+Sp rat were pretreated with spermine (Sp, 5 mg/(kg·d)) for 2 weeks before STZ injection. After 12 weeks of modeling, blood glucose, insulin levels, ejection fraction (EF) and shortening fraction (FS) were measured, and Masson staining and Sirius red staining were performed in the rat cardiac tissues. ②Cell experiments: primary CFs were extracted from newborn (1-3 d) Wistar rat hearts, and were randomly divided into control group, high-glucose group (HG) and HG+Sp group (n=6 per group). HG group was treated with 40 mmol/L glucose, and the HG+Sp group was pretreated with 5 μmol/L Sp for 30 min before HG treatment. The cell viability of CFs was detected by CCK8, the content of collagen in culture medium was analyzed by ELISA, and protein expressions of cell cycle related proteins (PCNA, CyclinD1 and P27) were detected by Western blot. Results: Compared with control group, the blood glucose and collagen content were increased, and the insulin level and heart function were decreased in the T1D group. Meanwhile, HG induced an increasing of the cell viability, the collagen content in the medium and the expressions of PCNA and CyclinD1, while the expression of P27 was down-regulated. Spermine could reduce the above changes, manifested as improving the cardiac function, regulating the expression of cyclin and reducing the level of myocardial fibrosis. Conclusion: Spermine can alleviate myocardial fibrosis in diabetic cardiomyopathy, which mechanism is related to the regulation of cell cycle.
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    Effects of leptin on lipid metabolism and inflammatory factors in diabetic rats
    ZHU Jian-zhong, ZHAO Can, SUI Yue-lin, LIU Yuan-yuan, QIAO Yue-bing
    CJAP. 2020, 36 (3): 197-201.   DOI: 10.12047/j.cjap.5960.2020.044
    Abstract   PDF (967KB) ( 257 )
    Objective: To investigate the effects of leptin on glucose metabolism and related inflammatory factors in diabetic rats. Methods: Ten healthy male Wistar rats were randomly selected as the control group. Fifty rats were fed with high sugar and high fat diet and injected with streptozotocin (STZ, 25 mg/kg) intraperitoneally. They were randomly divided into model group, leptin low, middle and high dose group. The rats in the low, middle and high dose group were fed with leptin at the doses of 20, 50 and 100 μg/kg for 5 d respectively. Blood glucose (FBG) was measured by GOD-PAP method, insulin content (INS) was tested by radioimmunoassay, the serum levels of triglyceride (TG), total cholesterol (TC), low density lipoprotein (LDL-C) and high density lipoprotein (HDL-C) were determined by automatic biochemical analyzer, the contents of malondialdehyde (MDA), interleukin-6 (IL-6) and tumor necrosis factor (TNF-α) were detected by enzyme-linked immunosorbent assay (ELISA). Western blot was used to detect the expression of leptin in adipose tissue of diabetic rats. Results: Compared with the control group, the blood glucose levels of other groups were increased significantly (P<0.01). Compared with the model group, the blood glucose levels of middle and high dose leptin rats decreased significantly (P<0.05, P<0.01). The insulin level of high dose leptin group decreased significantly (P<0.01). There was no significant difference in FBG and INS among the three groups (P>0.05). Compared with the model group, TC levels of middle and high dose leptin group were decreased significantly (P<0.05, P<0.01). TG and LDL-C levels of high dose leptin group were decreased significantly (P<0.05), HDL-C level of high dose group was increased significantly (P<0.01). Compared with different dose groups, the high dose of leptin (100 μg/kg) could decrease the levels of TC, TG and LDL-C, and increase the level of HDL-C, which was better than those of the middle and low dose of leptin (P<0.05) Compared with the model group (52.27±10.93), the levels of leptin in low, middle and high dose group were (47.35±12.09), (44.68±10.23) and (40.13±9.87) respectively, which could be decreased by leptin in a dose-dependent manner. Conclusion: The abnormal secretion of leptin is one of the factors inducing diabetes mellitus. Under the intervention of a certain concentration of exogenous leptin (100 μg/kg), it can significantly reduce the level of MDA, TNF-α, and improve the level of IL-6. The mechanism may be closely related to the reduction of inflammatory response, oxidative stress and correction of dyslipidemia. Leptin also reduces the risk of disease progression in diabetes treatment.
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    Effects of aerobic exercise and resveratrol on the expressions of JAK2 and TGF-β1 in renal tissue of type 2 diabetes rats
    SUN Xiao-juan, FENG Wu-long, HOU Na, LI Na, JIANG Wei-li
    CJAP. 2020, 36 (3): 202-206.   DOI: 10.12047/j.cjap.5918.2020.045
    Abstract   PDF (1454KB) ( 374 )
    Objective: To investigate the effects of aerobic exercise and resveratrol on janus kinase 2(JAK2) and transforming growth factor-β1(TGF-β1) in renal tissue of type 2 diabetes rats and its mechanism. Methods: The model of type 2 diabetic rats was established through SD rats fed high-fat diet for 5 weeks together with intraperitoneal infecting after a low dose of STZ. The rats were randomly divided into diabetic control group(DC), diabetic exercise group(DE), diabetic resveratrol group(DR), diabetic exercise and resveratrol group(DER), normal control group(NC), 12 rats in each group. Exercise-related groups performed 8 weeks treadmill exercise (20 m/min, 60 min/day). Resveratrol was administered to drug-related groups for 8 weeks (45 mg/kg, 7 day/week). Eight weeks later, we examined blood glucose concentrations, 24 h microalbuminuria(UA), serum creatinine(Scr), blood urea nitrogen(BUN), and the expressions of TGF-β1, janus kinase 2(JAK2) and JAK2 mRNA in renal tissue. Results: After eight weeks of intervention, compared with NC group, the concentrations blood glucose, 24 h UA, Scr, BUN, the expressions of TGF-β1, JAK2 and JAK2 mRNA were increased significantly in DC group(P<0.05). Compared with DC group, the concentrations of blood glucose, 24 h UA, Scr, BUN, the expressions of TGF-β1, JAK2 and JAK2 mRNA were decreased significantly in DE group, DR group and DER group(P<0.05). Conclusion: Exercise, resveratrol and combined intervention may decrease the expressions of JAK2 mRNA, JAK2 and TGF-β1, which further attenuate renal injury for type 2 diabetes. The renal protective effect produced by exercise and resveratrol combined intervention is better than that produced by exercise or resveratrol intervention alone.
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    Effect of exogenous spermine pretreatment on alleviating renal fibrosis in diabetic nephropathy mice and its related mechanism
    SHAO Xiao-ting, HU Jing, ZHANG Xin-ying, ZHAO Bing-bing, LI Si-wei, GAO Ping, XU Chang-qing, WEI Can
    CJAP. 2020, 36 (3): 207-210.   DOI: 10.12047/j.cjap.5973.2020.046
    Abstract   PDF (1213KB) ( 262 )
    Objective: To observe the protective effects of exogenous spermine on renal fibrosis induced by diabetic nephropathy (DN) and to explore its mechanism.Methods: Twenty-four male C57 mice were randomly divided into control group, type 1 diabetes group (TID) and spermine pretreatment group (TID+Sp, n=8 in each group). TID mice were induced by STZ (60 mg/kg), and TID+Sp mice were pretreated with spermine (5 mg/(kg·d)) for 2 weeks before STZ injection. The mice were killed at the 12th week. The renal function was determined by serum creatinine and urea nitrogen. HE, PAS and Masson staining were used to evaluate renal tissue injury and fibrosis. The expressions of matrix metalloproteinase (MMP-2, MMP-9) and collagen IV (Coll-IV) in the kidney of mice were detected by Western blot. Results: Compared with the control group, the blood glucose (5.67±0.22 vs 28.40±0.57 mmol/L), creatinine (14.33±1.22 vs 30.67±4.73 μmol/L) and urea nitrogen (6.93±4.94 vs 22.00±1.04 mmol/L) in the T1D group were increased significantly (P<0.05), the glomerular basement membrane was thickened, the collagen was significantly increased, the expressions of MMP-2, MMP-9 and Coll-IV protein were increased (0.57±0.07 vs 1.06±0.20, 47.00±0.04 vs 1.29±0.09 and 0.42±0.16 vs 0.95±0.18,P<0.05). Exogenous spermine significantly alleviates the above-mentioned changes. Conclusion: Exogenous spermine pretreatment could significantly alleviate renal fibrosis in diabetic mice by regulating the balance between MMPs and collagen.
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    Overexpression of miR-31 regulates TLR4/NF-κB signaling pathway and apoptotic protein in colitis model mice
    LIU Kai-li, DU Xin-xin, ZHANG Wen-qin, WU Ya-li, WANG Xi, WANG Chun-fang, CUI Xiang-li
    CJAP. 2020, 36 (3): 211-215.   DOI: 10.12047/j.cjap.5973.2020.047
    Abstract   PDF (5436KB) ( 330 )
    Objective: To investigate the effects of miR-31 on TLR4/NF-κB signaling pathway and apoptosis-related proteins in dextran sulfate sodium (DSS) induced mouse colon colitis. Methods: ① Mouse model of colon colitis: 1% DSS was used to induce mouse ulcerative colitis (UC). Fourteen FVB non-transgenic mice were randomly divided into control group (n= 6), DSS group (n= 8), and 16 FVB miR-31 transgenic mice were randomly divided into miR-31 overexpression group (n= 8), miR-31 overexpression +DSS group (n= 8). DSS was dissolved in water and administered to mice by drinking water. The DSS group and miR-31+DSS group drank 1% DSS water in the first week, normal sterilized water in the second week, and 1% DSS water in the third week, after 5 weeks, the modeling was completed, then the colon tissues of the mice were collected. Western blot and IHC were used to detect the expressions of NF-κB p65, TLR4, Bax and Bcl-2 proteins in mouse colon tissue, TUNEL was used to detect apoptosis of mouse colon tissues. ② Cell culture experiments: Transfection of miR-31mimic and inhibitor by lipofectamine resulted in overexpression or knockdown of miR-31 in human colon epithelial cell line HCT 116 cells, each group was repeated three times and cells were collected 48 h later, Western blot was used to detect the expressions of NF-κB p65 and TLR4 protein. Results: ① In animal experiments, compared with the control group, the expression levels of NF-κB p65, TLR4 protein and apoptotic cell index in the DSS group and miR-31 overexpression group in mouse colon tissue were significantly increased (P<0.05 or P<0.01), and the Bcl-2 / Bax ratio was significantly reduced (P<0.05 or P<0.01); and compared with the DSS group, the expression levels of NF-κB p65, TLR4 protein and apoptotic cell index in the miR-31+DSS group were significantly increased (P<0.01), while the Bcl-2/Bax ratio was significantly decreased (P<0.01). ② In cell experiments, compared with the control group, the expression levels of NF-κB p65 and TLR4 protein in the over-expressed miR-31 group of HCT 116 cells were significantly increased (P<0.05 or P<0.01), the expressions of NF-κB p65 and TLR4 protein in miR-31 knockdown group were decreased (P<0.05). Conclusion: miR-31 promotes the development of colitis by promoting TLR4/NF-κB signaling pathway and mediating apoptosis of intestinal epithelial cells.
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    Effect of small dose capsaicin for treatment of pulmonary fibrosis in mice and its mechanism
    LU Lin-ming, YU Ting-ting, HE Xiao-wei, TANG Juan, LI Xian-wei
    CJAP. 2020, 36 (3): 216-222.   DOI: 10.12047/j.cjap.5974.2020.048
    Abstract   PDF (1711KB) ( 348 )
    Objective: To observe whether the mechanism of small dose capsaicin (Cap) against pulmonary fibrosis in mouse is mediated by agitating transient receptor potential vanilloid 1 (TRPV1). Methods: A total of 60 BALB/c mice were randomly divided into control (CON) group, bleomycin (BLM)group, Cap (0.5, 1,2 mg/kg) groups and Cap (2 mg/kg) plus SB-452533 (2.5 mg/kg) group. C57BL/6 mice were intratracheally injected with 3.5 mg/kg BLM to induce pulmonary fibrosis model. Animals for drugs treatment received daily drug via subcutaneous injection for 21 days. The morphological changes and collagen deposition in lung tissues were analysed by HE staining, Masson staining and immunohistochemistry. The concentration of calcitonin gene-related peptide (CGRP) in plasma was determined by ELISA. The mRNA and (or) proteins levels of α-CGRP, β-CGRP, collagen I, collagen III, E-Cadherin, zonula occludens-1 (ZO-1), vimentin, alpha smooth muscle actin (α-SMA), TRPV1, p-ERK1/2 and eukaryotic initiation factor 3a (eIF3a) were detected by qPCR and (or) Western blot. Results: Compared with the BLM group, small dose Cap significantly reduced bleomycin-induced pulmonary fibrosis in mice and obviously reversed alveolar epithelial cells epithelial-mesenchymal transition (EMT) (the expression of E-cadherin and ZO-1 were increased(P<0.05 or P<0.01)and the expression of α-SMA and Vimentin were decreased (P<0.05 or P<0.01) after drugs treatment for 21 day, concomitantly with the increase the expressions of TRPV1 and CGRP (P<0.05 or P<0.01), and inhibiting ERK1/2 phosphorylation and eIF3a expression (P<0.05 or P<0.01). These effects of small dose Cap were abolished in the presence of TRPV1 receptor antagonist SB-452533. Conclusion: The results suggest that small dose Cap can reverse alveolar epithelial cells EMT and alleviate bleomycin-induced pulmonary fibrosis in mice by inhibiting ERK1/2/eIF3asignaling pathway, which is related to agitating TRPV1 receptor and releasing of CGRP.
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    Effects of hypothermia on the repolarization duration of ventricular myocytes in rats and its mechanism
    FU Xiao-kui, LIU Yan-qiu, GAO Hong, WANG Gui-long, LI Hua-yu, DAI Dong-jun
    CJAP. 2020, 36 (3): 228-231.   DOI: 10.12047/j.cjap.5914.2020.050
    Abstract   PDF (1025KB) ( 112 )
    Objective: To observe the effects of hypothermia on the repolarization duration and the expression of Kir2.1 protein of ventricular myocytes in isolated rat heart and explore the role of Kir2.1 protein.Methods: Eighteen healthy adult male Sprague-Dawley rats were randomly divided into three groups (n=6 per group): Control group (C group), 35℃ group (H1 group), 32℃ group (H2 group). Langendorff isolated heart models were established. After 15 min 37℃ K-H fluid banlanced perfusion, C group continued to perfuse the K-H solution at 37℃ for 30 minutes, H1 group continued to perfuse the K-H solution at 35℃ for 30 minutes, H2 group continued to perfuse the K-H solution at 32℃ for 30 minutes. At 15 min of balanced perfusion (T1), and 30 min of continuous perfusion (T2), the heart rate,and the MAP in the three layers of the left ventricular anterior wall were recorded, the action potential duration at 50% repolarization (MAPD50), the action potential duration at 90% repolarization (MAPD90) and transmural dispersion of repolarization(TDR) were calculated. At the same time, the occurrence of arrhythmia was recorded. The expression of Kir2.1 protein was measured by Western blot. The average optical density (AOD) and the distribution of Kir2.1 protein were measured by immunohistochemistry in the ventricular tissue measured by electrophysiology. Results: Compared with T0, the heart rate was decreased, MAPD50 and MAPD90 were prolonged significantly (P<0.05), and TDR was increased significantly (P<0.05) in H1 group, H2 group at T1. Compared with C group, the HR was decreased, the MAPD90 was prolonged significantly (P<0.05), TDR was increased significantly (P<0.05),the expression and the AOD of Kir2.1 protein were decreased significantly (P<0.05) in H1group, H2group at T1. Compared with H1 group, the heart rate of H2 group was decreased significantly (P<0.05), MAPD50 and MAPD90 were prolonged significantly (P<0.05), and TDR was increased significantly (P<0.05) at T1. The distribution of Kir2.1 protein in group C was normal, while the distribution of Kir2.1 in H1 group and H2 group was disordered. Conclusion: Hypothermia prolonged the ventricular duration of repolarization and increased the dispersion of repolarization. The mechanism is related to the down-regulation the expression of Kir2.1 protein and the disorder of the distribution of Kir2.1 protein.
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    Effects of NaHS on MBP and learning and memory in hippocampus of mice with spinocerebellar ataxia
    JIANG Hong-bo, DONG Jia-xing, QIN Yu-fei, LIU Jia-cong, JIANG Wan-ju, LI Ruo-nan, LIU Lan-ci, TIAN Yi-dan, XU Yu-ming, DU Ai-lin
    CJAP. 2020, 36 (3): 235-239.   DOI: 10.12047/j.cjap.5953.2020.052
    Abstract   PDF (1219KB) ( 221 )
    Objective: To investigate the effects of exogenous NaHS on myelin basic protein (MBP) and learning and memory of hippocampal neurons in mice with spinocerebellar ataxia type 3 (SCA3) and its therapeutic significance.Methods: Twelve male normal mice were randomly selected as normal control group (NC Group), and 48 SCA3 mice were randomly selected as SCA3 model group (M Group), low dose group (NL Group, 10 μmol/kg), medium dose group (NM Group, 50μmol/kg) and high dose group (NH Group, 100 μmol/kg), 12 rats in each group. The drug treated groups were injected with NaHS intraperitoneally once a day for 4 weeks. The changes of learning and memory ability of SCA3 mice before and after the intervention of different doses of NaHS were determined by Morris water maze, the content of hydrogen sulfide (H2S) in hippocampus was measured by spectrophotometry, the expression of MBP was detected by immunohistochemistry, and the morphological changes of neuron myelin sheath were observed by electron microscope. Results: Compared with the control group, the learning and memory ability of SCA3 mice was decreased significantly (P<0.05), and the content of H2S in hippocampus was decreased (P<0.05). After different doses of exogenous NaHS treatment, the learning and memory ability was improved in different degrees (P<0.05), and the contents of H2S and MBP in hippocampus of SCA3 mice were also improved in different degrees (P<0.05). Conclusion: Exogenous NaHS may increase the contents of H2S and MBP in the hippocampus of SCA3 mice, which may have a protective effect on the neurons, and then improve the learning and memory ability of SCA3 mice, and provide a new idea for the treatment of SCA3.
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    Inflammatory mechanism of hippocampal tissue injury induced by PM2.5 in nasal drip in mice
    FANG Zhen, HU Xi-hou, LI Kang, HAN Jie, TIAN Lei, YAN Jun, ZHANG Wei, LAI Wen-qing, LIN Ben-cheng, LIU Xiao-hua, XI Zhu-ge
    CJAP. 2020, 36 (3): 240-244.   DOI: 10.12047/j.cjap.5916.2020.053
    Abstract   PDF (2070KB) ( 215 )
    Objective: To investigate the inflammatory mechanism of nasal instillation of fine particulate matter (PM2.5)on hippocampal tissue injury in mice.Methods: Thirty C57BL/6J mice were randomly divided into 3 groups(n=10):control group, low-dose group, high-dose group. The nasal instillation doses of PM2.5 in the low-dose group and the high-dose group were 1.5 mg/kg BW and 7.5 mg/kg BW, respectively, and the control group was given saline with an equal volume. Saline was sprayed once every other time for 12 times. The serum levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) were determined by ELISA method. HE staining and electron microscopy were used to observe the pathological changes and ultrastructure of lung tissue and hippocampus. The inflammatory cytokine levels in hippocampus were detected by antibody chip technique. Results: There was no significant effect of PM2.5 nasal instillation on serum TNF-α, IL-1β and IL-6 levels (P>0.05), and there was no obvious pathological changes in lung tissue structure. In hippocampus, low-dose and high-dose PM2.5 exposure could lead to disordered neuronal arrangement in the hippocampal CA3 region, and there were neurological changes around the neuron cells and ultrastructural changes such as edema around small blood vessels. Compared with the control group, the levels of inflammatory cytokines such as CX3CL1, CSF2 and TECK in the low-dose group were increased significantly (P <0.05), while sTNFR1 was decreased significantly (P<0.05); the inflammatory factors CX3CL1, CSF2, and TCA-3 were significantly increased in the high-dose group (P<0.05), while leptin, MIG, and FASLG were significantly decreased (P<0.05). Conclusion: Nasal instillation of PM2.5 can induce tissue damage in the hippocampus of mice, and its mechanism of action may be the olfactory brain pathway. The increasing of TNF-α and IL-6 and the decreasing of sTNFR1 and FASLG may be involved in inflammatory mechanisms.
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    Effects of repeated immobilization stress on hypothalamic-pituitary- ovarian axis in female rats
    HAN Jie, LIU Zi-yi, FANG Zhen, TIAN Lei, YANG Dan-feng, XI Zhu-ge, LIU Xiao-hua
    CJAP. 2020, 36 (3): 245-249.   DOI: 10.12047/j.cjap.5927.2020.054
    Abstract   PDF (2307KB) ( 203 )
    Objective: To explore the effects of repeated immobilization stress on hypothalamic-pituitary-ovarian axis in female rats. Methods: Forty female SD rats were randomly divided into two groups: control group (n=20) and experimental group (n=20). One group was fed normally, the other group was subjected to incremental load restraint stress. Brake stress once a day in the retainer (starting at 9: 00 a.m.), braking for 2 hours on the first day, increasing load by 0.5 hours a day for two weeks. Body weight, estrous cycle, sex hormone, organ coefficient, pathology and expression of related genes were detected to explore the harm of hypothalamic-pituitary-ovarian axis. Results: Repeated immobilization stress caused weight loss, prolonged estrous cycle, and changed the organ coefficient and morphology of ovaries and uterus. QPCR technique was used to detect the related genes. It was found that the expressions of gonadotropin releasing hormone, pituitary gonadotropin releasing hormone receptor, follicle stimulating hormone and luteinizing hormone mRNA were decreased significantly, while the expressions of ovarian follicle stimulating hormone and luteinizing hormone receptor mRNA were increased significantly. The expression of estrogen receptor mRNA in ovary and uterus was decreased significantly. Conclusion: Repeated immobilization stress may disrupt the estrous cycle by interfering with the endocrine regulation of the hypothalamic-pituitary-ovarian axis, thus damaging the gonadal and reproductive endocrine function of female animals.
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    Potential toxic effects of TDCIPP on the thyroid in female SD rats
    SUN Jing-ran, FAN Mi-mi, ZHANG Zhen, WU Jin, HAN Dian-peng, BAI Jia-lei, DU Lian-qun, FANG Yan-jun
    CJAP. 2020, 36 (3): 250-254.   DOI: 10.12047/j.cjap.5979.2020.055
    Abstract   PDF (2006KB) ( 201 )
    Objective: To investigate the potential toxic effects and mechanisms of Tris(1; 3-dichloro-2-propyl) phosphate (TDCIPP) on thyroid in female SD rats.Methods: Thirty-two 3-weeks-old female SD rats were randomly divided into normal group(treated with corn oil ), and low/moderate/high-dose group treated with TDCIPP (dissolved in corn oil )(n=8). All rats were treated with corn oil or TDCIPP (50, 100, 250 mg/(kg·d)) once a day during a 21-day period. All rats were sacrificed after the last administration. Serum thyroid stimulating hormone (TSH), 3,3’,5-triiodothyronine (T3), 3,3’,5,5’-tetraiodothyronine (T4), free 3,3’,5,5’-tetraiodothyronine (FT4) were detected with ELISA kit. Morphology of thyroid was observed with hematoxylin and eosin (HE) staining. Expressions of genes and proteins correlate with thyroid were measured respectively by real-time fluorescence quantitative PCR and Western blot. Results: Compared with control group, morphology of thyroid showed follicles irregular arrangement, hypocolloid, and follicular hyperplasia in TDCIPP treatment groups. The levels of serum TSH in low-dose TDCIPP group and T3 in high-dose TDCIPP group were significantly higher than those in control group(P<0.05). Thyroid stimulating hormone receptor (TSHR) mRNA expression was decreased distinctly in low-dose TDCIPP group, while the expression of thyroperoxidase (TPO) mRNA was increased notably in moderate and high-dose TDCIPP groups(P<0.05,P<0.01). Compared with control group, the level of TRβ protein was decreased significantly in moderate and high-dose TDCIPP groups, while the expressions of udp-glucuronosyl-transferases (UGTs) and cytochrome-p450-3A1 (CYP3A1) proteins were upregulated notably in TDCIPP treatment groups(P<0.05). Conclusion: Treated with 50 mg/(kg·d) TDCIPP can cause thyroid hyperplasia, change the levels of thyroid hormones, and disturb thyroid function, therefore, it has toxic effects on the thyroid.
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    Effect of psoralen on rat osteoblasts injuries induced by TCP wear particles in vitro and its mechanism
    CHEN Yu-feng, DONG Fan-he, LOU Yun-wei, SHOU Jin-hao, ZHANG Hui-ting, ZHOU Yi-chao, YAN Ming, MAO Hong-jiao, ZHANG Yun
    CJAP. 2020, 36 (3): 255-260.   DOI: 10.12047/j.cjap.5900.2020.056
    Abstract   PDF (1337KB) ( 181 )
    Objective: To investigate the effect and mechanism of psoralen on calvarial osteoblasts injuries caused by tricalcium phosphate (TCP) wear particles in vitro.Methods: Primary osteoblasts were obtained from the calvaria of neonatal SD rat by the series of digestion and were identified with ALP staining. Calvarial osteoblasts were treated with TCP wear particles for 48 h to establish the in vitro model of osteoblasts injuries. The rat osteoblasts were randomly divided into control group, TCP wear particles (0.1 mg/ml) group, psoralen treated (at the concentrations of 10-7, 10-6, 10-5 mol/L) groups. WST assay and the flow cytometry were used to detect the cell viability of osteoblasts and apoptosis, respectively. Chemical colorimetry was performed to examine ALP activity of osteobalsts. When the osteoblasts were treated for 14 day, mineral nodules formation was observed with alizarin red S staining. Western blot was applied to examine protein expressions of glucose regulated protein78/94(GRP78/94), inositol dependent enzyme 1 alpha (IREα), spliced X-box binding protein 1 (XBP1s) and phosphorylated c-Jun N-terminal kinase (p-JNK) in calvarial osteoblasts. Results: Compared with control group, the cell viability of osteoblasts, ALP activity and mineral nodules formation in TCP group were decreased significantly (P<0.05), while the percentage of apoptosis and protein expressions of GRP78/94, IRE1α, XBP1 and p-JNK were obviously increased in calvarial osteoblasts (P<0.05). Compared with TCP group, the injuries of calvarial osteoblasts and cell apoptosis in psoralen treated groups were obviously decreased (P<0.05), and the expression levels of GRP78/94, IRE1α, XBP1 and p-JNK were down-regulated remarkably (P<0.05). Conclusion: Psoralen prevents osteoblasts injuries caused by TCP wear particles through IRE1α-XBP1s-JNK signaling pathway activation.
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    Effects of liraglutide combined with vitamin D on non-alcoholic fatty liver induced by high fat in mice and its mechanism
    WANG Feng, ZHANG Hai-feng
    CJAP. 2020, 36 (3): 261-264.   DOI: 10.12047/j.cjap.5911.2020.057
    Abstract   PDF (1003KB) ( 287 )
    Objective: To investigate the effects of liraglutide combined with vitamin D on high-fat-induced non-alcoholic fatty liver disease (NAFLD) mice and its potential mechanism. Methods: C57BL/6 mice were divided into control group, NAFLD model group, liraglutide group, vitamin D group and liraglutide combined with vitamin D group. Each group consisted of 10 mice. The control group was fed with normal diet for 12 weeks; the model group was fed with high-fat diet for 12 weeks; the liraglutide group, vitamin D group and combined group were fed with high-fat diet for 12 weeks, From the 9th week, the three groups of mice were intraperitoneally injected with liraglutide (0.6 mg/kg), vitamin D(250 mg/(kg·d) ) by gavage, and combination. After 12 weeks of feeding, the blood and liver tissues of mice in each group were collected for biochemical and pathological examination, and the phosphorylation level of AMP-activated protein kinase (AMPK) in liver tissues of mice in each group was detected by immunoblotting. Results: Liraglutide or vitamin D alone or in combination could improve liver lipid accumulation (triglycerides: 6.0±0.7 vs 3.8±0.3, 3.9±0.3 and 2.1±0.2, all P<0.05; cholesterol: 1.4±0.5 vs 0.9±0.2, 0.8±0.2 and 0.5±0.1, all P<0.05) and steatosis (NAFLD activity score: 2.4±0.3 vs 1.0±0.2, 0.9±0.1 and 0.6±0.1, all P<0.05) in NAFLD mice. In addition, compared with liraglutide or vitamin D group, liraglutide combined with vitamin D treatment was more effective, and might be related to the regulation of insulin resistance and AMPK phosphorylation. Conclusion: The results showed that vitamin D could enhance the therapeutic effect of liraglutide on NAFLD induced by high fat, and may be related to the regulation of insulin resistance and AMPK phosphorylation.
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    Effects of obatoclax combined with gemcitabine on breast cancer cells under hypoxia condition
    SONG Hai-yan, ZHANG Yi-min, LIAN Hui, ZHOU Li
    CJAP. 2020, 36 (3): 268-272.   DOI: 10.12047/j.cjap.5955.2020.059
    Abstract   PDF (1181KB) ( 171 )
    Objective: To explore the effects of obatoclax(OBX) combined with gemcitabine(GEM) on breast cancer cells MCF-7 and BT-20 cell activity, migration, invasion and apoptosis under hypoxia condition.Methods: Breast cancer cells MCF-7 and BT-20 were divided into normal group, hypoxia group, GEM group, OBX+GEM group. Normal group: Cells were cultured at 37℃, 5% CO2 for 24 h and 48 h; Hypoxia group: Cells were cultured at 37℃, 1% O2, 5% CO2, 94% N2 for 24 h and 48 h; GEM group: Cells were cultured at 37℃, 1% O2, 5% CO2, 94% N2, adding 10 μmol/L GEM for 24 h and 48 h; OBX + GEM group: Cells were cultured at 37℃, 1% O2, 5% CO2, 94% N2, adding 10 μmol/L GEM and 50 nmol/L OBX for 24 h and 48 h. Western blot method was used to detect the expressions of HIF-1α in MCF-7 and BT-20 cells under normal oxygen and hypoxia condition. CCK-8 method was used to detect cancer cell activity, each group was provided with 15 compound holes. Scratch experiment was used to detect cells migration ability, each group was provided with 6 compound holes. Western blot method was used to detect the expressions of vimentin, E-Cadherin and p53 protein in cells of each group. Results: Under hypoxia condition, the expression of HIF-1α in MCF-7 and BT-20 cells was much higher than that under normal oxygen(P<0.05). Compared with hypoxia group, GEM could reduce MCF-7 and BT-20 cells migration ability(P<0.01)and cell activity(P<0.05), while decrease the expression of vimentin protein(P<0.01)and promote the expressions of E-Cadherin (P<0.01)and p53 protein(P<0.01) in tumor cells under hypoxia condition. In OBX combined with GEM group, the cell activity and the migration ability of MCF-7 and BT-20 were reduced significantly(P<0.01). The expression of vimentin in cells was further reduced(P<0.01). The expressions of E-Cadherin(P<0.01)and p53(P<0.01) protein were increased significantly compared with GEM group. Conclusion: Under hypoxia condition, OBX combined with a low-dose of GEM can significantly inhibit the growth, migration and invasion of breast cancer cells, and enhance the pro-apoptotic effect of GEM, but the specific mechanism needs further study.
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    Effects of propofol on TGF-β1 induced activation of hepatic stellate cells and its mechanism
    ZHANG Qian-lu, KUANG Xin
    CJAP. 2020, 36 (3): 273-278.   DOI: 10.12047/j.cjap.5929.2020.060
    Abstract   PDF (1240KB) ( 209 )
    Objective: To observe the effects of propofol on the activation of hepatic stellate cell line HSC2-T6 induced by transforming growth factor-beta 1 (TGF-β1) and explore its possible mechanism.Methods: The cells were divided into control group, TGF-β1 group, propofol group, TGF-β1 + propofol group, rapamycin group, TGF-β1 + propofol + rapamycin group. Cells were treated with rapamycin (5 μmol/L) for 1 hour, propofol (100 μmol/L) for 1 hour, then TGF-β1 (5 ng/ml) was added to co-culture for 24 hours. Cell proliferation was measured by MTT assay. The concentrations of hyaluronic acid (HA), collagen IV (IV-C) and laminin (LN) in the supernatant of cell culture medium were measured by ELISA. The ultrastructure of cells was observed by transmission electron microscopy. The expressions of alpha-smooth muscle actin (α-SMA), mammalian rapamycin target protein (mTOR), phosphorylated mTOR (p-mTOR) and the autophagy related gene Beclin 1, LC3 and p62 were measured by Western blot. Results: Compared with control group, cell proliferation, the expression of α-SMA, the concentrations of HA, IV-C and LN in culture supernatant, the number of autophages, the expressions of Beclin-1 and LC3-II, the ratio of LC3-II/LC3-I in HSC2-T6 cells were increased significantly, while the expression of p-mTOR, the ratio of p-mTOR/mTOR and the expression of p62 protein were decreased significantly in TGF-β1 group (All P<0.05). Compared with TGF-β1 group, cell proliferation, the expression of α-SMA, the concentrations of HA, IV-C and LN in culture supernatant, the number of autophages, the expressions of Beclin-1 and LC3-II, the ratio of LC3-II/LC3-I in HSC2-T6 cells in TGF-β1 group were decreased significantly, and the expression of p-mTOR, the ratio of p-mTOR/mTOR and expression of p62 protein were increased significantly in TGF-β1 + propofol group (All P<0.05). Conclusion: Propofol inhibits the activation of hepatic stellate cells induced by TGF-beta 1, and its mechanism involves the mTOR-autophagy pathway.
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    Effects of artesunate on the inhibition of hepatic fibrosis through ceramide synthase-ceramide pathway
    RUAN Jian-jia, DU Yan
    CJAP. 2020, 36 (3): 279-282.   DOI: 10.12047/j.cjap.5931.2020.061
    Abstract   PDF (939KB) ( 270 )
    Objective: To investigate the effects of ceramide pathway on the inhibition of artesunate (Art) to hepatic fibrosis.Methods: LX-2 cells were divided into control group, Art treated group with 350 μmol/L, fumonisin B1 (FB1) treated group with 6 μmol/L, and Co-administration group of artesunate 350 μmol/L and fumonisin B1 6 μmol/L. There were 7 compound holes in each group. After 24 hours of treatment, the cells and supernatant were collected and detected. The expressions of homo sapiens longevity assurance homologue 2 (LASS2), peroxisome proliferators-activated receptors-γ (PPAR-γ) and Caspase-3 were evaluated by Western blot, the content of ceramide was evaluated by HPLC-FLD method, MTT assay was adopted to measure the rate of proliferation of LX-2 cells. The content of hydroxyproline was determined by digestive method. Results: Compared with the control group, the expression of ceramide synthase protein and the ceramide content were increased significantly, the proliferation of LX-2 cells was inhibited significantly, the expressions of PPAR-γ and Caspase-3 protein were up-regulated and the secretion of hydroxyproline was inhibited in Art treated group (P<0.05). In FB1 treated group, the protein expression of ceramide synthase and the ceramide content were decreased significantly, the proliferation of LX-2 cells was increased significantly, the expressions of PPAR-γ and Caspase-3 protein were down-regulated, and the secretion of hydroxyproline was increased (P<0.05). Compared with the Art alone group, the combination of the two drugs could significantly reduce the effects of Art on the expression of ceramide synthase protein and the increase of ceramide content, and attenuate the effects of Art on the cell proliferation , PPAR-γ, Caspase-3 protein expression and hydroxyproline level of LX-2 cells (P<0.05). Conclusion: Artesunate could inhibit hepatic fibrosis by increasing the content of ceramide through the ceramide synthase-ceramide pathway.
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    Technical Notes
    The optimization and assessment of the method for inducing hyperuricemia in rats
    SHI Hui, LIANG Xiao-shan, HUANG Li-wen, LUO Zhi-gang, TAN Long
    CJAP. 2020, 36 (3): 223-227.   DOI: 10.12047/j.cjap.5933.2020.049
    Abstract   PDF (1200KB) ( 212 )
    Objective: To explore an effective method for inducing a rat model with hyperuricemia in a short period and assess the effects of the model. Methods: Sprague-Dawley rats were adopted as donors and randomly divided into control group (CT group, n=6) and 5 model groups (M1-M5 groups, n=8 in each group). M1 group (gavage with 10 g/kg yeast extracts and 100 mg/kg adenine, twice per day, 300 mg/kg oxonic acid potassium by intraperitoneal injection, in the 7th day of model inducing), M2 group (gavage with 10 g/kg yeast extracts and 100 mg/kg adenine, twice per day, 300 mg/kg oxonic acid potassium by intraperitoneal injection, in the 1st, 3rd and 7th day of model inducing),M3 group (gavage with 10 g/kg yeast extracts and 100 mg/kg adenine, twice per day, 300 mg/kg oxonic acid potassium by intraperitoneal injection, once per day during the model inducing), M4 group (gavage with 20 g/kg yeast extracts and 100 mg/kg adenine, twice per day, 300 mg/kg oxonic acid potassium by intraperitoneal injection, once per day during the model inducing), M5 group (gavage with 30 g/kg yeast extracts and 100 mg/kg adenine, twice per day, 300 mg/kg oxonic acid potassium by intraperitoneal injection, once per day during the model inducing), and group CT (gavaged with equal volume sterilized water and intraperitoneal injected with normal saline according to the weight and at the same frequency as the model groups). The model inducing lasted for 7 days. After the inducing was finished, blood and 24-hour urine were sampled for uric acid and creatinine determination. Then rats were maintained for 2 weeks and blood and 24-hour urine samples were collected, the concentration of uric acid and creatinine were detected. The kidney and stomach were weighed,morphological changes in kidney were observed. Results: After model inducing, the body weight of rats in all model groups was lower than that of the control group (P<0.01). Deaths occurred in all the rats with model treatments except M2. M4 and M5 groups were failed to be analyzed because of the high mortality, model 1 and 3 groups had 4 and 2 deaths, respectively. The uric acid levels in blood and urine of the model groups were significantly elevated (P< 0.01) at the end of model inducing. The model 2 group’s blood uric acid was highest among the model groups (P<0.05). It sustained a higher concentration than CT group in the three model groups after 2 weeks feeding (P<0.05). The kidneys in model groups obviously swelling and were heavier than CT group (P<0.01). The inflammation and structural damages were observed in kidneys of all model groups.Conclusion: The yeast extract (10 g/kg), adenine (100 mg/kg) gavage combined with intraperitoneal injections(the 1st, 3rd, 7th day during inducing) of potassium oxonate can be an rapid and effective method for inducing the rat model with hyperuricemia, which can be suggested to the related research.
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    A new design puncture needle and a device of microcatheter protection for lumbar intrathecal catheterization in rats
    XU Chang-shun, SUN Pei, LIN Chun
    CJAP. 2020, 36 (3): 283-288.   DOI: 10.12047/j.cjap.5938.2020.062
    Abstract   PDF (1214KB) ( 196 )
    Objective: To introduce a new design needle and a device of microcatheter protection for lumbar intrathecal catheterization in rats,and evaluate its feasibility and effectiveness.Methods: Sixty pathogen-free adult male Sprogue-Dawley rats were randomly divided into two groups(n=30 in each group), the control group (group C) and the modification group(group M). The traditional puncture device, 20G needle, was used in the group C without extemal shielding protection. The new design puncture needle and the microinjection cock were used in the group M. All rats were assessed for motor function on postoperative. The motor function was evaluated 1 day afteroperation. Lidocaine was injected in the catheter at 1st,3rd,7th,14th,21st day post-catheterization, methylene blue was injected in intrathecal at 30th day after operation, and the catheter location was observed. The paw withdrawal threshold(PWT) was measured at 1st,3rd,7th,14th,21st,30th day after operation, open-field test was tested at preoperative and one week postoperative for the purpose of evaluating the autonomous behavior of rats. Results: About motor function:level Ⅰ 75.9%,level Ⅱ 20.7%,level Ⅲ 3.4% in group C, and level Ⅰ 96.7%,level Ⅱ 3.3% in group M, Compared with group C,group M had higher percentage of the level Ⅰ in motor function (P<0.05);Lidocaine test and methylen blue location showed that each one case of catheter was removed on the 14th and 21st day after intubation in group C, and total four cases were removed till the 30th day, while all catheters were in normal location in group M. There was significant difference between two groups in protection of the extemal portion of catheter(P<0.05); The time of intrathecal injection in group M was only 1 minute, and it spent more than 3 minutes in group C. Compared with group C,the time of intrathecal injection is significantly shorter in group M(P<0.01);PWT was reduced to the lowest on the third day after catheterization, and there was significant difference compared with preoperative(P<0.05), PWT recovered on the 7th day and there were no significant difference between two groups; Compared with preoperative, there was no significant difference in the parameters of the group M in the open field test, neither between two groups. Conclusion: The new design puncture needle by its less injury and higher efficiency can be used in intrathecal catheterization. The microinjection cock is reliable and convenient for repeat injection with a perfect protection function of the external portion of catheter, meanwhile it has no impact on rats’ autonomous behavior so that it is worthy of further promoting.
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  2019-03-15

Sponsor
Chinese Association for Physiological Sciences Academy of Military Medical Sciences Institute of Health and Environmental Medicine
Associate Sponsor
Institute of Basic Medical Sciences
Editor in Chief
WANG Hai
Edited and Published by
Editorial Board,Chinese Journal of Applide Physiology;Dali Dao,Tinanjin 300050,China



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