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    Cell and Molecules Biology
    Novel expression of spermatogenesis-associated protein 3 gene in mouse spermatogenic cells and its influence upon apoptosis and autophagy in HEK 293T cells
    WANG Yu-jing WEN Li-min BAI Xin-yan CAO Rui WANG Hai-long GUO Rui
    2018, 49 (1):  41-48.  doi: 10.16098/j.issn.0529-1356.2018.01.007
    Abstract ( )   PDF (917KB) ( )  

    Objective To detect the expression of spermatogenesis-associated protein 3(spata3) gene in mouse spermatogenic cells and to analyze its probable function on apoptosis and autophagy in HEK 293T cells over-expressed spata3. The overall aim is to explore the possible significance of spata3 during mouse spermatogenesis. Methods Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blotting were used to detect the expression of spata3 mRNA and its encoded protein in a variety of mouse tissues, respectively. Immunohistochemical staining and indirect immunofluorescent staining were used to locate the SPATA3 protein in spermatogenic cells. The recombinant eukaryotic expression plasmid Plv-EGFP-2 (a) puro-spata3 were transiently transfected into HEK 293T cells using liposome and the expression level of apoptosis-related proteins including cleaved-Caspase3, poly ADP-ribose polymerase(PARP), BAX, Bcl-2 and autophagy-related protein LC3A/B were investigated by Western blotting. Results Spata3 mRNA and its encoded product were specifically expressed in mouse testis. SPATA3 positive signals were mainly located in the cytoplasm and the nucleus of both the round spermatids and the pachytene spermatocytes. Moreover, SPATA3 stainning signals were detected in the cytoplasm of elongating spermatids. The expression levels of cleaved-Caspase-3 and cleaved-PARP were not significant difference between HEK 293T cells over-expressed spata3 and control groups. The content of BAX in 293T cells overexpressed spata3 0.815±0.020 was higher than that in 293T cells 0.469±0.012 and in 293T cells transfected with Plv-EGFP-2(a) puro 0.588±0.018, while the expression of Bcl-2 in HEK 293T cells over-expressed spata3 0.214±0.020 was lower than that in HEK 293T cells 0.507±0.021 and in HEK 293T cells transfected with Plv-EGFP.2(a) puro 0.545±0.024. The amount of LC3A/B-Ⅱin HEK 293T cells over-expressed spata3 0.741±0.037 were detected higher than that in HEK 293T cells 0.136±0.011 and in HEK 293T cells transfected with Plv-EGFP-2(a) puro 0.169±0.012. Conclusion Spata3 is specifically expressed in mouse spermatogenic cells and its over-expression in HEK 293T cells has no significant effect upon apoptosis, but could promote autophagy.

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    Effect of obeticholic acid on expression of farnesoid X receptor and cytochrome 7A1 in nonalcoholic fatty liver rats
    LIN Hao LIANG Da-wei ZNEG Jian-feng LI Shu-jian WANG Jian-fu LI Fei
    2018, 49 (1):  49-55.  doi: 10.16098/j.issn.0529-1356.2018.01.008
    Abstract ( )   PDF (644KB) ( )  

    Objective To investigate the effect of obeticholic acid on protein expression and gene expression of farnesolid X receptor (FXR), cytochrome 7A1 (CYP 7A1) in non-alcohol fatty liver disease (NAFLD) rat and hepatocyteprotective effect. Methods Fifty two SD rats of the non-alcoholic fatty liver model was established by feeding with high-fat diet. The control group, model group,the experimental group(low), the experimental group(middle), the experimental group(high) were set up. After four weeks of administration, the same part of the liver was taken out. The expression of FXR protein was detected by immunohistochemistry method , the mRNA expression of FXR was detected by RT-PCR, the protein CYP7A1 was detected by Western blotting and the mRNA expression of CYP7A1 was detected by RT-PCR. The liver function was assessed by the rat liver index of the concentrations of peripheral blood alanine aminotransferase (ALT), aspartate transaminase(AST) and alkaline phosphatase(ALP), and the observation of liver sections. Results After fed with high-fat diet for 10 weeks, the rats had typical characteristics of NAFLD and the model was established successfully. The expression of FXR protein and FXR mRNA in the model group were significantly lower than that in the control group (P<0.05). There was no significant difference in the expression of FXR protein and FXR mRNA between the experimental group(low) and the experimental group(middle) (P> 0.05). The expression of FXR protein and FXR mRNA in the experimental group(high) was significantly higher than that in model group (P<0.05). The optical density values of model group, the experimental group(low), and the experimental group(middle) were significantly lower than those of control group (P<0.05). The optical density values of the experimental group(high) was significantly higher, and there was no significant difference compared with those of the control group (P>0.05). The CYP7A1 protein and CYP7A1 mRNA expression levels in the model group were significantly higher than those in the control group (P<0.05). Compared with those in the model group, the expression levels of CYP7A1 protein and CYP7A1 mRNA in the experimental group(low) and the experimental group(middle) had no significant difference (P>0.05). The levels of CYP 7A1 protein and CYP 7A1 mRNA in the experimental group(high) were significantly lower than those in the model group (P<0.05). Compared with the control group, the liver index (%) of the model group increased significantly (P<0.05), while the liver index (%) of the experimental group(low) and the experimental group(middle) had no significant difference compared with the model group (P> 0.05). The liver index (%) of the experimental group(high) was significantly lower than that of the model group (P<0.05). Compared with the control group, the concentrations of ALT, AST and ALP in the model group were significantly increased (P<0.05). The levels of ALT, AST and ALP in the experimental group were significantly lower than those in model group (P<0.05), and the concentrations of ALT, AST and ALP gradually decreased with the increase of acyclobacic acid dosage. The liver tissue sections were observed. The result showed that obeticholic acid improved the histopathology of rats in a dose-dependent manner. Conclusion High doses of acyclobacic acid can significantly upregulate the expression of FXR protein and its gene, inhibit the expression of CYP 7A1 protein and its gene, and significantly improve the liver function and pathological status of rats.

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    Anthropology
    Analysis of morphological characteristics of the palms and fingers of Tibetan and Han youths
    ZHONG Hua ZHANG Hai-long XI Hua-jiu LI You-qiong ANGWANG Ge-sang YU Ze-ming
    2018, 49 (1):  113-117.  doi: 10.16098/j.issn.0529-1356.2018.01.019
    Abstract ( )   PDF (210KB) ( )  

    Objective To explore the characteristics of length, width and thickness of the palms in Tibet, and to provide reference data for supplementary sport science, anthropology and clinical or behavior assessments. Methods This study selected young college students who voluntarily participated in Lhasa as the research objects. There were 216 males (64 in Tibetan, 72 in Han) and 80 females (40 in Tibetan and 40 in Han), with an average age of (22.12±3.06) years. The electronic vernier caliper was used to measure the left hand length, finger length and palm length of the subjects, such as 27 direct measurement indicators. The hand area was calculated by hand length and hand width. Results There were significant differences in hand length, thickness at metacarpale and hand index between Tibetan and Han(P<0.05, 0.01). There was a significant difference(P<0.05,0.01) between Tibetan female and Han female in comparison of the hand length, back of hand length, hand breadth at metacarpale, palm length, palm breadth, finger Ⅰ length, finger Ⅱ length,finger Ⅲ length, finger Ⅳ length,finger Ⅴ length, fingerⅠbreadth,maximum finger Ill breadth,finger Ⅲ breadth,maximum finger Ⅳ breadth,maximum finger Ⅲ breadth, maximum finger Ⅳ breadth,maximum finger Ⅲ thickiness and hand area. When Tibetan palm finger shape was compared between the different genders, in addition to thickness at metacarpale, thickness at hypothenar and hand index, other indicators were more in males than in females(P<0.05,0.01). When Han palm finger shape was compared between different sexes, in addition to palm breadth and hand index, other indicators were more in males than in females(P<0.05, 0.01).Conclusion The palms and fingers morphological features of the Tibetan men are more than those of the Han. The length, width and thickness of palms and fingers of Tibetan women are all smaller than those of Han. The palms and fingers morphological features of females are less than that of males in length, width and thickness.

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    Cancer Biology
    Expression characteristics and significance of cathepsin D in the initiation and progression of mouse lung adenocarcinoma
    ZHANG Hui-juan ZHENG Xiao-wei QIAO Ling CHANG Wen-jing LU Feng JIA Cai-yun
    2018, 49 (1):  56-62.  doi: 10.16098/j.issn.0529-1356.2018.01.009
    Abstract ( )   PDF (816KB) ( )  

    Objective To investigate the expression characteristics and significance of cathepsin D(CTSD) in the initiation and progression of mouse lung adenocarcinoma.Methods We used urethane intraperitoneal injection to establish lung adenocarcinoma model in the BALB/c mouse. Differentially expressed proteins were identified using isobaric tag for relative and absolute quantitation(iTRAQ)-based quantitative proteomic technology in tumor group compared with control group. Cathepsin D expression characteristics were detected in different time points by immunohistochemical and Western blotting analysis. Results The expression of cathepsin D was significantly increased in the early-stage lung adenocarcinoma tissue compared with the control group. Immonohistochemical and Western blotting analysis observed that following tumor progression, cathepsin D expression was up-regulated. At the late-stage of lung adenocarcinoma, cathepsin D expression was 13.7-fold significantly higher than that in the control group and showed different molecular weight isoform of cathepsin D in the lung tissue of mouse with liver metastasis. Conclusion Cathepsin D is a critical protein in lung carcinogenesis. Aberrant expression and modification of cathepsin D may play an important role in the occurrence, development and metastasis of lung adenocarcinoma. Cathepsin D may have the potential to become biomarkers for early diagnosis of lung adenocarcinoma.

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    Astragalus polysaccharide inhibiting Lewis lung carcinoma transplanted tumor in mice and influences the expression of Caspase-3 and Smac/Diablo
    ZHUANG Meng-jie LIU Dan CHEN Yan-wen MING Hai-xia
    2018, 49 (1):  63-69.  doi: 10.16098/j.issn.0529-1356.2018.01.010
    Abstract ( )   PDF (718KB) ( )  

    Objective To observe the effect of Astragalus polysaccharides (APS) combined with cisplatin (DDP) on the expressions of Caspase-3 and Smac/Diablo in the mice with transplantated tumors Lewis lung carcinoma (LLC). Methods Ninety C57BL/6 J mice were randomly divided into normal control group, model group, and 50, 100 or 200 mg/L APS group, 6 mg/kg cisplatin group, and 3 mg/kg cisplatin combined with 50, 100 or 200 mg/L APS group; 10 mice per group. Except the mice in normal group, the rest mice were inoculated with LLC cells (1×1010/L) in the right fore axillary subcutaneous tissue to establish a model of tumorbearing mice. In the second day of building the animal model, the mice in the treatment group were given intraperitoneal injection of 0.3 ml of the drug. At the same time, the mice in the cisplatin group were given once a week, and the rest of the group mice once a day. The mice in the normal and model groups were given the same amount of saline injection for 20 days. All mice were killed on the 21st day. Tumor tissue lesions were observed by HE staining. The expression and location of Caspase-3 and Smac/Diablo proteins in transplanted tumor tissues were detected by immunohistochemical staining and image analys is method . Results The weights of mice were decreased in the 100 and 200 mg/L APS group and 3 mg/kg cisplatin combined with 50, 100, 200 mg/L APS group. Compared with the model group, the necrosis of tumor tissues in the 200 mg/L APS combined with 3 mg/kg cisplatin group was most obvious. The expression of Caspase-3 and Smac/Diablo was increased in the treatment group. The increasing of tumor tissues in 200 mg/L APS combined with 3 mg/kg cisplatin group was most obvious. Conclusion APS and APS combined with cisplatin restrain the growth of Lewis lung cancer in C57BL/6 J mouse, which may depend on increase of the expression of Caspase-3 and Smac/Diablo.

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    Effects of dihydroartemisinin on proliferation and apoptosis of human pancreatic cancer cell line
    ZHU Wen-he XU Na XU Jun-jie ZHANG Hong LI Yan Lü Shi-jie ZHANG Wei
    2018, 49 (1):  70-74.  doi: 10.16098/j.issn.0529-1356.2018.01.011
    Abstract ( )   PDF (361KB) ( )  

    Objective To investigate the effect of dihydroartemisinin on proliferation and apoptosis of human pancreatic cancer cell (PANC-1) and explore its mechanism.Methods After PANC-1cells treated with 30, 60, 120, 240 and 480 μmol/L dihydroartemisinin for 48 hours, MTT assay was used to detect the proliferation effect of dihydroartemisinin on PANC-1 cells. The cell cycle and apoptosis were detected by flow cytometry. The expressions of cyclin B1, Cdk1, p21 and apoptosis related protein Bcl-2, Bax, Caspase-3, Caspase-9 and cytochrome C(Cyto C) were detected by Western blotting. Results MTT result showed that the proliferation of PANC-1 cells was inhibited obviously after treatment with dihydroartemisinin by the concentration of 30-480 μmol/L (P<0.05). The result of flow cytometry showed that dihydroartemisinin could block PANC-1 cell cycle in G2/M phase and induce cells apoptosis (P<0.05). The result of Western blotting showed that the expressions of Cdk1 and cyclin B1 were decreased and the expression of P21 increased after treatment with dihydroartemisinin. The expression of Bcl-2 was downregulated, expression of Bax was up-regulated, the ration of Bax/Bcl-2 was increased and the expression of cleaved Caspase-3, cleaved Caspase-9 and Cyto C were up-regulated. Conclusion Dihydroartemisinin can inhibit the proliferation of PANC-1 cells and induce apoptosis. The possible mechanism might be related to mitochondrial pathway.

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    Histology,Embryology and Developmental Biology
    Morphologic characteristics of the atrioventricular canal myocardium and epicardium in the mouse embryonic heart
    LI Qian CAO Xi-mei LI Hai-rong JING Ya QIAO Cong-jin YANG Yan-ping
    2018, 49 (1):  93-97.  doi: 10.16098/j.issn.0529-1356.2018.01.015
    Abstract ( )   PDF (689KB) ( )  

    Objective To investigate the morphological characteristics of the myocardium and epicardium during the septation and remodeling of the atrioventricular canal in the mouse embryonic heart. Methods Serial sections of twenty-five mouse embryos during embryonic day(ED) 10 to ED15 were stained by immunohistochemistry and immunofluorescence with antibodies against myosin light chain 2a (MLC2a), myosin regulatory light chain 2 (MLC-2), T-box transcription factor 3 (Tbx3), and Lymphoid enhancer-binding factor 1 (Lef1). Results Immunohistochemistry and immunofluorescent staining for MLC2a and Tbx3 were positive in the atrioventricular canal myocardium of ED10 to ED15 mice, but for MLC2 was negative. During ED11 to ED12, the epicardium was observed. At ED12 and ED13, the bilateral atrioventricular canal cushions began to fuse with each other and formed the atrioventricular valve. The epicardium derived mesenchymal cells between the atrioventricular myocardium and epicardium were increased, part of which showed the Lef1 immunohistochemistry positive staining. From ED13, some epicardial cells derived from mesenchymal cells transversed through the myocardium to parietal atrioventricular valve. At ED15, the base of the atrioventricular valve was directly connected with the MLC2a positive atrioventricular myocardium. Conclusion The atrioventricular myocardium develops into the myocardium to support the base of the valves of the adult atrioventricular ring. The epicardium provides mesenchymal cells which contribute to the formation of atrioventricular valves.

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    Expression pattern of transcription factor Tbx3 in the second heart field of the mouse embryo
    YANG Yan-ping LI Qian LI Jie CAO Xi-mei LI Hai-rong JING Ya QIAO Cong-jin ZHANG Tao
    2018, 49 (1):  87-92.  doi: 10.16098/j.issn.0529-1356.2018.01.014
    Abstract ( )   PDF (1037KB) ( )  

    Objective To investigate the expression pattern of Tbx3 in the mouse embryonic second heart field (SHF) to provide morphological basis for the study of the regulation mechanism in SHF. Methods Serial sections of 40 mouse embryonic hearts aged from embryonic day(ED)9 to ED15 were stained by immunohistochemistry or double-label immunofluorescence with antibodies against islet-1 (ISL-1), myosin heavy chain (MHC), myosin light chain 2 a (MLC2a), proliferating cell nuclear antigen (PCNA) and T-box transcription factor 3 (Tbx3). Results Immunohistochemical positive staining for Tbx3 and SHF marker ISL-1 was distributed in pharyngeal ventral endoderm of ED9 mouse. During ED10 to ED12, immunohistochemical positive staining for ISL-1 was presented in pharyngeal ventral mesenchymal and staining intensity was increased gradually. Some ISL-1 positive mesenchymal cells were co-expressed with Tbx3. ISL-1 was also shown in the aortic sac wall, pericardial dorsal wall and the distal pole of the outflow tract, where Tbx3 was negative. However, the dorsal mesenchymal protrusion (DMP) contained ISL-1 or Tbx3 positive cells. During ED13 to ED15, DMP was myocardialized and sporadic Tbx3 positive cells were seen in DMP. Conclusion Tbx3 expression is mainly concentrated in pharyngeal ventral mesenchyme, which suggests that Tbx3 may contribute to the survive and proliferation of SHF progenitor cells. However, its function is different in posterior second heart field(pSHF)and in SHF of arterial pole of embryonic heart.

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    PERK-eIF2α-ATF4 signaling pathway is involved in periprosthetic osteolysis induced by tricalcium phosphate wear particles in mice
    CHEN Yue-liang ZHANG Yun LOU Qian-ping WANG Cheng-long DU Kang-hui SHEN Qi-hua MAO Hong-jiao
    2018, 49 (1):  98-103.  doi: 10.16098/j.issn.0529-1356.2018.01.016
    Abstract ( )   PDF (409KB) ( )  

    Objective To explore whether protein kinase R-like ER kinase(PERK)-eukaryotic translation initator factor 2(eIF2α)-activating transcription factor 4(ATF4) signalling pathway is involved in periprosthetic osteolysis induced by tricalcium phosphate(TCP)wear particles in mice. Methods Thirty male ICR mice were randomly divided into sham group (n=10), TCP group (n=10) and salurinal (SAL) group (n=10). A murine calvarial model of osteolysis was established by 30 mg of TCP wear particles implantation onto the surface of bilateral parietal bones following removal of the periosteum. From the second postoperative day, SAL (1 mg/kg) was locally injected to the calvarium under the periosteum three times a week. After 2 weeks, the calvaria and serum were obtained. Western blotting was used to examine the expression of ER stress marker glucoseregulated protein 78(GRP78)and C/EBP homologous protein (CHOP), together with activation of PERK-eIF2α-ATF4 signalling pathway. Tartrate resistant acid phosphatase(TRAP)staining, HE staining and Real-time PCR were performed to observe the effects of SAL on TCP wear particles-induced osteolysis, osteoclastogenesis and mRNA levels of osteoclastogenic genes including TRAP, cathepsin K and c-Fos in the calvaria, respectively. Enzyme-linked immumsorbent as ay (ELISA) was performed to determine the effect of SAL on serum level of tumor necrosis factor-α (TNF-α), prostaglandin E2 (PGE2) and interleukin-1 beta (IL-1β). Results TCP wear particles triggered remarkable ER stress responses in the mouse calvaria, which was confirmed by up-regulation of ER stress markers GRP78 and C/EBP homologous protein (CHOP), and activation of protein kinase RNA-like ER kinase-eIF2α-activating transcription factor 4 pathway (including phosphorylation of both protein kinase RNA-like ER kinase and eIF2α and elevated level of activating transcription factor 4) (P<0.05). Block of the ER stress response with the eIF2α specific inhibitor salubrinal markedly prevented TCP wear particles-induced osteoclastogenesis and osteolysis, and suppressed the increase in mRNA level of osteoclastogenic genes such as TRAP, MMP-9 and c-fos in the mouse calvaria (P<0.05). In addition, SAL significantly inhibited TCP wear particles-induced release of TNF-α, PGE2 and IL-1β (P<0.05). Conclusion PERK-eIF2α-ATF4 pathway is involved in the calvarial osteolysis induced by TCP wear particle in mice, and inhibition of this signalling pathway reduces periprosthetic osteolysis and prostheses caused by TCP wear debris.

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    Effects of brain-derived neurotrophic factor on mouse preimplantation-embryo development and apoptosis under heat shock
    LIU Ben
    2018, 49 (1):  104-107.  doi: 10.16098/j.issn.0529-1356.2018.01.017
    Abstract ( )   PDF (383KB) ( )  

    Objective To investigate the effect of brain derived neurotrophic factor (BDNF) on the development and apoptosis of mouse early blastocyst either cultured continuously at 37 ℃ or treated with heat shock at 41 ℃ for 2 hours. Methods A total of 150 female Kunming mice were used. The early blastocysts were cultured continuously at 37 ℃ or subjected to heat shock for 2 hours at 41 ℃ while being cultured with or without 10μg/L BDNF. The development of early blastocysts was recorded. The TUNEL reaction and Hoechst 33342 method were used to observe the apoptosis cells of blastocysts. The Caspase-9 activity of mouse blastocyst was analysed by fluorescent technique. Results Treatment with BDNF at 37 ℃ promoted the development of early blastocysts. The normal developmental blastocyst cells were increased while apoptosis cells and Caspase-9 activities were decreased in early blastocyst in BDNF treatment group. The development of early blastocyst was inhibited in heat shock treatment group, and TUNEL positive cells number and Caspase-9 activities were increased. There were normal developmental blastocyst in 10μg/L BDNF treatment group. Conclusion Our findings suggest that BDNF signaling plays important roles during mouse early blastocyst development, blastocyst cell numbers, blastocyst apoptosis and the Caspase-9 activity.

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    Expression of transforming growth factor-β and Smad4 in ovarian granulosa cells in menopausal transitional rat
    YUAN Li-juan REN Jun-xu JI Hong-yu ZHANG Jing ZHANG Juan DING Yan
    2018, 49 (1):  108-112.  doi: 10.16098/j.issn.0529-1356.2018.01.018
    Abstract ( )   PDF (438KB) ( )  

    Objective To investigate the expression of transforming growth factor beta(TGF-β) and Smad4 in ovarian granulosa cells of rats during menopausal transition and to analyze their relationship with the decline of ovarian function. Methods Female SD rats were divided into control group (group C, 6 months old, vaginal smear screening, n=9) and menopausal transition(MT) group (MT group, 12-14 months old, vaginal smear screening, n=8), rats were sacrificed quickly after ovariectomy. The ovarian granulosa cells were released by mechanical separation method, and cultured in CO2 incubator. The expression of follicle-stimulating hormone receptor(FSHR) protein was observed by immunocytochemistry to identify ovarian granulosa cells. Immunohistochemistry was used to detect the expression of TGF-β and Smad4 protein in ovarian granulosa cells and images were collected and analyzed; The expression of TGF-β and Smad4 mRNA in granulosa cells was detected by Real-time PCR. Results Immunocytochemistry showed that the purity of granulosa cells was > 95%. The intensities of immunocytochemical positive staining for TGF-β and Smad4 in MT group was lower than those in normal group (P<0.05).Real-time PCR showed that both TGF-β and Smad4 mRNA were expressed. The amount of Real-time PCR for TGF-β and Smad4 mRNA was significantly lower than those of the control group (P<0.05). Conclusion TGF-β and Smad4 may be involved in the decline of ovarian function in rats with menopausal transition. The causes of the functional decline in postmenopausal rats may be partly related to the decrease of TGF-β and Smad4 expression in ovarian granulosa cells.

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    Anatomy
    Attenuating effects and mechanisms of apelin-13 on aortic injuries in diabetic mice
    ZHANG Jia WANG Yang-jia ZENG Xiang-jun
    2018, 49 (1):  81-86.  doi: 10.16098/j.issn.0529-1356.2018.01.013
    Abstract ( )   PDF (757KB) ( )  

    Objective To observe the effects and mechanisms of apelin-13 on aortic fibrosis in diabetic mice. Methods Eighteen mice were divided into three groups: control group: C57/BL6j mice in eight weeks old (n=6), diabetic group: kkAy mice in eight weeks old (n=6), and apelin-13 group: kkAy mice implanted with osmotic pump to release apelin-13 at the rate of 30μg/(g·d).After 28 days, the aortas were harvested and fixed. Morphological changes and collagen deposition were observed with HE and Masson staining. The aortic elastic fibers were observed with elastin staining. Expression of transforming growth factor-β1 (TGF-β1) was measured with immunohistochemical staining. Results Compared to the control group, the wall of aorta in diabetic mice thickened, the collagen deposite in diabetic mice was significantly more than that in control mice(P<0.05). The elastin in diabetic mice was irregularly arranged. The levels of TGF-β1 in the aorta of diabetic mice were significantly more than that in control mice(P<0.05).After apelin-13 treatment, the wall of the aorta in diabetic mice became thinner, the collagen deposite was significantly improved than that in diabetic mice(P<0.05), while apelin-13 reset the elastin of aorta in diabetic mice,and reduced TGF-β1 expression to almost normal levels(P<0.05). Conclusion Apelin-13 may attenuate fibrosis and elastin breakdown in the aorta in diabetic mice by inhibit TGF-β1 expression and proliferation of smooth muscle cells in medium of the aortic wall.

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    Clinical anatomy of parotid surface division associated with tumorigenesis
    XU Xiang-liang SUN Zhi-peng WANG Dian-can WANG Jian-wei ZHANG Wei-guang
    2018, 49 (1):  75-80.  doi: 10.16098/j.issn.0529-1356.2018.01.012
    Abstract ( )   PDF (416KB) ( )  

    Objective To partition human parotid gland on the body surface anatomically based on radiological images of parotid tumors. Methods According to parotid tumors computed tomography (CT) images, the method of partitioning parotid gland was proposed in this study. Line A and B were determined by the several landmarks such as alae nasi, angle of mouth, earlobe and posterior margin of mandibular ramus, which partitioned parotid gland into regions Ⅰ, Ⅱ and Ⅲ. Parotid regions were dissected and observed in cadavers. Regions of tumors shown in parotid CT images were analyzed according to the partition method. Results Branches of facial nerve, great auricular nerve, external jugular vein and retromandibular vein appeared frequently in region Ⅰ of parotid gland. Branches of facial nerve and transverse facial artery appeared frequently in region Ⅱ. Auriculotemporal nerve and superficial temporal artery and vein appeared frequently in region Ⅲ. Based on analyzing the CT images of parotid glands, it could be concluded that tumor incidence in region Ⅰ is the highest rate among the tumors which grew in only one region, that no tumor grew in both region Ⅰ and region Ⅱ, and that the tumors which grew in both two regions are those who have relative large volume, among which 62.5% tumors have larger than 3 cm diameters. Conclusion The method of partitioning parotid gland anatomically has the value of anatomy and tumor biology based on this research, which could be valuable for parotid tumor resection clinically.

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    Neurobiology
    Heat shock protein 70 and Hantavirus persistent infection of cortical astrocytes from newborn rats
    HE Shuai YANG Shou-jing
    2018, 49 (1):  7-13.  doi: 10.16098/j.issn.0529-1356.2018.01.002
    Abstract ( )   PDF (522KB) ( )  

    Objective To define that heat shock protein 70(HSP70)overpression might contributed to the persistent infection of Hantaviruses (HTNV). Methods The expression of HSP70 in cortical astrocytes was examined by Western blotting and RT-PCR during Hantaviruses persistent infection. The following apoptosis was detected by immunofluorescence microscopy, AnnexinV assays and RT-PCR assay. Results Compared to control group ,the expression of HSP70 protein in astrocytes infected with HTNV 76-118 or SEOV L99 increased, peak expression of HSP70 at day 3 following HTNV 76-118 infection or at day 2 following SEOV infection (n=15, P<0.01, P<0.05), and the expression of HSP70 gene in astrocytes infected with HTNV 76-118 or SEOV L99 up-regulated, peak expression of HSP70 at day 2-5 post HTNV76-118 infection and at day 2-4 post SEOV L99 infection (n=15, P<0.01, P<0.05). In the course of HTNV76-118 or SEOV L99 infection, any cytopathic effect was not shown in astrocytes and cell apoptosis was not induced by viral replication. Conclusion HSP70 induced by Hantaviruses infection might inhibit apoptosis and contribute to the viral persistent infection.

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    Screening and identification of the glial cell activation associated microRNA after corticospinal tract injury
    LIU Xiao-dong WANG Yi-ru Lü Jin-yang XU Xiao-yan ZHANG Zhi-yi WANG Xin Lü Guang-ming
    2018, 49 (1):  14-19.  doi: 10.16098/j.issn.0529-1356.2018.01.003
    Abstract ( )   PDF (550KB) ( )  

    Objective To detect changes in microRNA and target mRNA at the head and tail of the corticospinal tract (CST) injury side, and to investigate the function of microRNA in spinal cord injury (SCI) and repairment, which could provide a theoretical basis and experimental evidence in regeneration of the CST after injury.Methods The left side of the CST in 30 rats was cut at the level of the pyramid of medullar oblongata. The BBB test was used to assess the motor function. After the CST injury, the bioinformatics method was conducted to screen differences in microRNA target gene prediction. The intersection of differentially expressed mRNA was selected to conduct the microRNA and mRNA co-expression analysis. Real-time PCR was applied to detect the gene expression of miR-342-5p and Irf8 in 12 rats. Western blotting was applied to detect the protein expression of Irf8 and Iba1 in 6 rats. Results The limb movement after CST semitransection was uncoordinated, and motor function was limited according to BBB test but not completely paralyzed. There were statistical significances(P<0.05)after comparing 2 hours, 1 day, 3 days, 5 days and 7 days with the control group, indicating the successful establishment of model. Comparison of the inrostraland caudal region between 2 hours and 5 days, we focused on biological processes in regeneration. Data analysis from Realtime PCR experiments indicated that caudal miR-342-5p expression was significantly different(P<0.01) from 2 hours to 5 days, with a reverse correlation with Irf8 expression, whereas no change was observed in rostral miR-342-5p(P>0.05). Real-time PCR data analysis showed that the expression of Irf8 was consistent with the chip result , statistically significant difference compared with control groups (P<0.01). The result of Western blotting indicated that, comparing tissue from 5 days with 2 hours, both Irf8 and Iba1 were significantly creased in caudal samples (P<0.01). Conclusion After CST injury, there were significant up-regulated or down-regulated differentially expressed genes in rostral and caudal regions. In the caudal region, the miR-342-5p and Irf8 reverse expression and miR-342-5p was closely related to SCI repair and regeneration by glial cell activation.

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    Expression of DDX3 and casein kinase 1ε in the brain stem of amyotrophic lateral sclerosis transgenic mice
    ZHANG Ya-wen WANG Qing YUAN Meng LIU Huan-cai WANG Qiao-zhen DING Hao-yu ZHOU Feng-hua CHEN Yan-chun
    2018, 49 (1):  1-6.  doi: 10.16098/j.issn.0529-1356.2018.01.001
    Abstract ( )   PDF (814KB) ( )  

    Objective To detect the expression of DDX3 and casein kinase 1ε (CK1ε)in the brain stem of amyotrophic lateral sclerosis (ALS) transgenic mice and study the role of DDX3 and CK1ε in the degeneration of brain stem motor neurons of amyotrophic lateral sclerosis (ALS). Methods Thirty-three ALS transgenic mice were used in this study. The brain stem was dissected and collected at the early (95 days), middle (108 days) or late (122 days) stages. The expression of DDX3 and CK1ε in the brain stem of ALS transgenic mice, the distribution and co-localization of positive cells in the hypoglossal nucleus and facial nucleus of the brain stem were detected by RT-PCR, Western blotting and immunofluorescence technology. In each group, the same number of wild type littermates were selected as controls. Results The result of RT-PCR and Western blotting showed that compared with the wild type mice, the expression of DDX3 and CK1ε mRNA in the brain stem of ALS mice remained unchanged at day 95, day 108 and day 122. DDX3 and CK1ε protein levels were up-regulated at day 95 and day 108 but down-regulated at day 122 in the ALS mice brain stem group (P<0.01, P<0.001). The result of immunofluorescence showed that DDX3 and CK1ε positive cells were detected in the sublingual nerve and facial nerve of ALS mice and wild type mice brain stem. DDX3 and CK1ε were expressed in neurons, but not in astrocytes. The immunoreactivity of both DDX3 and CK1ε of ALS mice and wild type mice was different. Conclusion The abnormal expression of DDX3 and CK1ε in the brain stem is closely related to the pathogenesis of ALS.

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    Transferrin receptor 1’s neuroprotection in amyloid precusor protein/presenilin-1 transgenic mice
    WANG Qian FAN Wen-juan SUN Yi-zheng WANG Lai CHENG Yan-hong DENG Jin-bo
    2018, 49 (1):  20-28.  doi: 10.16098/j.issn.0529-1356.2018.01.004
    Abstract ( )   PDF (976KB) ( )  

    Objective To investigate the neuroprotection effect of transferrin receptor 1(TfR1) in Alzheimer’s disease(AD). Methods Immunofluorescence and Western blotting were used to detect the expression of TfR1 in the amyloid precursor protein(APP)/presenilin-1(PS1)transgenic mice from postnatal 0 day(P0) to P360. With primary cultured hippocampal neurons, TfR1 expression and amyloid betapeptides (Aβ) secretion were detected with Western blotting and ELISA assay, respectively. The cultured neurons and their processes were labeled with TfR1 and microtubule associated protein 2 (MAP2) immunolabeling, and the TfR1-mediated axonal vesicles were observed with FM1-43 staining, after TfR1 shRNA interference. Results TfR1 expression in AD mice (APP/PS1 transgenic mice) decreased significantly after postnatal 6 months (P 6 M) compared with the wild type(WT) mice. Similarly, in cultured cells, after TfR1 gene silence, the neuronal processes became long and thin, and the axonal vesicle transportation was blocked. Conclusion APP and PS1 gene mutation can decrease expression of TfR1. TfR1 shRNA interference can increase the amount of Aβ1-42 secretion and impact neurite growth and axonal vesicle transportation. Therefore, we conclude that TfR1 may play an important role in neuroprotection.

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    Effects of vasoactive intestinal peptide on synapse of MPTP-induced mice with Parkinson’s disease
    Lü E FEI Xue-chao LI Kan ZHAN Tong-xia LI Feng-jie FU Wen-yu
    2018, 49 (1):  29-34.  doi: 10.16098/j.issn.0529-1356.2018.01.005
    Abstract ( )   PDF (626KB) ( )  

    Objective To study the effects of vasoactive intestinal peptide (VIP) on synaptic and behavioral alterations in the mice model of Parkinson’s disease (PD) induced by 1-methyl-4-phenyl-1,2,3,6tetrahydropyridine (MPTP). Methods Thirty male C57BL/6 J mice were randomly divided into normal saline (NS) group, MPTP group and MPTP + VIP group. The behavioral changes of mice were measured by the Tru Scan system. The expression of tyrosine hydroxylase (TH) in the substantia nigra (SN) and striatum and vesicle-associated membrane protein 2 (VAMP2) and synapsin 1 (SYN1) in the striatum were examined by immunohistochemistry, and the levels of VAMP2 and SYN1 were analyzed by Western blotting. The synaptic structural changes in the SN were observed by transmission electron microscopy. Results In the behavioral test, the MPTP group had a significantly lower vertical movement distance than that of the control group, while the MPTP group was markedly higher than that of MPTP group. Compared with the MPTP group, VIP treatment obviously increased the average absorbance (AA) of TH, VAMP2 and SYN1 immunolabeling in striatum and the number of dopaminergic neurons in the SN, while the levels of the VAMP2 and SYN1 proteins also elevated. Synapse density in the model group was less than that in the control group. The amount of synaptic vesicles was decreased in the model group compared to the control, but normalized in the MPTP + VIP group. Conclusion VIP administration can up-regulate the expression of VAMP2 and SYN1, mitigate synaptic damage, protect dopaminergic neurons and improve movement in the MPTP model mice.

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    Effects of autophagy on apoptosis in rat hippocampal neurons following cardiac arrest and cardiopulmonary resuscitation
    MA Lei WANG Rong LI Hai-long MA Xiao-long CHEN Wei YANG Li-shan
    2018, 49 (1):  35-40.  doi: 10.16098/j.issn.0529-1356.2018.01.006
    Abstract ( )   PDF (393KB) ( )  

    Objective To explore the effect of autophagy on apoptosis in rat hippocampal neurons following cardiac arrest and cardiopulmonary resuscitation (CA/CPR).Methods Forty Sprague-Dawley(SD)rats were randomly divided into 4 groups: sham, CA/CPR model, rapamycin (CA/CPR +Rapa) and 3-Methyladenine (CA/CPR +3-MA) groups. The CA/CPR model was established by the asphyxial method . The rapamycin (autophagy agonist) or 3-methyladenine (autophagy inhibitor) were used to treated the corresponding group, respectively. The neurological deficit score (NDS) was used to evaluate the neurological function of CA/CPR rats. The TUNEL staining method was used to detect the apoptosis rate of hippocampal neurons of CA/CPR rats. The reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting method were used to detect the expression levels of microtubuleassociated protein light chain 3(LC3), Beclin-1, Bax, Bcl-2 and Caspase-3 mRNA and protein in hippocampus of CA/CPR rats. Results Compared with the sham group, the NDS of rats was decreased in the CA/CPR group; the apoptosis rate of the hippocampal neurons in CA/CPR group was increased; the expression levels of LC3, Beclin-1, Caspase-3, Bax were up-regulated, and Bcl-2 was down-regulated in the hippocampus of CA/CPR group (P<0.05, P<0.01). Compared with the model group, Rapa decreased NDS of CA/CPR rats, promoted the apoptosis rate of the hippocampal neurons, up-regulated the expression levels of LC3, Beclin-1, Caspase-3 and Bax in hippocampal tissue, and down-regulated the expression of Bcl-2; 3-MA increased CA/CPR rats’ NDS, inhibited the apoptosis rate of the hippocampal neurons, down-regulated the expression levels of LC3, Beclin-1, Caspase-3 and Bax in hippocampal tissue, and up-regulated the expression of Bcl-2 (P<0.05, P<0.01).Conclusion The increased level of autophagy may promote the apoptosis of CA/CPR rats’ hippocampal neurons. The inhibited level of autophagy may suppress the apoptosis of CA/CPR rats’ hippocampal neurons. The interaction of these two effects may be involved in the pathological process of CA/CPR.

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    Review
    Function of apelin/APJ signal in liver cirrhosis and hepatocellular carcinoma
    LIN Bo PAN Wei-tong KONG Jing Lü Xin-rui
    2018, 49 (1):  127-131.  doi: 10.16098/j.issn.0529-1356.2018.01.022
    Abstract ( )   PDF (264KB) ( )  

    Apelin/APJ is a G-protein-coupled receptors signal, which is widespread in human and animal tissues including brain, lung, heart, vessels, gastrointestinal tract and kidney, et al. The system plays a vital role in regulating the physiological activities of the human body by participating in multitudinous cell signaling transduction process. Apelin/APJ function mainly includes the regulation of the cardiovascular system homeostasis, maintaining water and electrolyte balance, promoting pituitary hormone release and regulating biological rhythm, and regulating the immune system as an immunomodulatory factor. However, apelin/APJ function is still unknown. In liver, apelin-APJ signal may promote the development of liver fibrosis, cirrhosis and cancer and may play an inhibitory role in liver regeneration. In this review, we summarise the function of apelin/APJ system in hepatic disease.

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    Androgen, androgen receptor and polycystic ovary syndrome
    YANG Mei LIN Hai-wei ZHANG Hong-quan XU Xiao-ming CHEN Xi
    2018, 49 (1):  132-136.  doi: 10.16098/j.issn.0529-1356.2018.01.023
    Abstract ( )   PDF (213KB) ( )  

    Polycystic ovary syndrome (PCOS), with a prevalence of 5%-10%, is the most common endocrine and metabolic dysfunction of reproductive age women. However, the underlying mechanism has not been fully elucidated. Hyperandrogenism is the leading defect in PCOS patients, especially local androgen secretion by the ovary is the key element for PCOS pathogenesis. Androgen activates androgen receptor (AR), so the role of AR is an indispensable factor. This review focuses on the functions of androgen and AR in PCOS pathogenesis.

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    Medical Education
    A new pedagogy for topographical anatomy: the application of minimally invasive surgery to topographical anatomy teaching
    LIU Ping ZHANG Yan JIANG Dong MA Cai-hong WANG Yi-ting HE Mei-hua ZHANG Wei-guang
    2018, 49 (1):  124-126.  doi: 10.16098/j.issn.0529-1356.2018.01.021
    Abstract ( )   PDF (148KB) ( )  

    Objective To examine the effects of a new pedagogy of application of minimally invasive surgery (MIS) to clinical medical students in topographical anatomy.Methods Clinical medical students were randomly divided into two groups, traditional group taught by traditional topographical anatomy pedagogy and minimally invasive surgery group taught by the new pedagogy of clinical minimally invasive topographical anatomy. Results Compared to students in traditional group, students in minimally invasive topographical anatomy group have better results in many ways including learning interest, learning motivation, knowledge fastness, operation ability, clinical way of thinking, integration of anatomical and clinical knowledge, and communication skills etc. Conclusion The application of minimally invasive surgery in topographical anatomy teaching can fulfill the requirements of early contact of the minimally invasive surgery and related surgical appliances used in clinics for medical students, especially clinical medical students and can also facilitate anatomy teacher to understand the development of clinical surgery and teaching purpose, as well as enhance the communication between anatomy teaching and clinical practice.

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    Bioengineering
    Soluble expression and purification of VP1 protein from O-type foot-and-mouth disease virus in Escherichia coli and its nanometer structure observation
    GUO Yu-kun MING Sheng-li GUO Wan-ying YANG Guo-yu GUO Yu-jie
    2018, 49 (1):  118-123.  doi: 10.16098/j.issn.0529-1356.2018.01.020
    Abstract ( )   PDF (603KB) ( )  

    Objective To establish a method to express the soluble VP1 protein from foot-and-mouth disease virus type O in Escherichia coli, (E. coli) and to observe its nanoparticles. Methods VP1 gene from FMDV O/MYA/7/98 strain was obtained according to the sequence of O-type foot-and-mouth disease virus. Truncation and optimization were performed. The fusion tages were screened to obtain soluble expression of SeFnt16798 protein. The 73 aa ferritin protein was isolated from Salmonella enteric and linked with the VP1 protein, which was named SeFnt16798. Prokaryotic expression vectors were fused with nine different fusion tages(Grifin, GST, MBP, Sumo, Thioredoxin, γ-crystallin, ArsC, PpiB and CeHSP17)fragments were constructed. These recombinant plasmids were transformed into Escherichia coli BL21 (DE3) and induced by isopropyl β-D-thiogalactiside(IPTG). SDS-PAGE analysis was used to identify the soluble expression of recombinant protein after ultrasonic decomposition. The fusion tag that could significantly promote soluble expression of SeFnt16798 protein was selected. Abundantly inducible expression recombinant protein was purified by Ni-NTA purification system and detected by electron microscopy. Results Recombinant expression of SeFnt16798 was successfully constructed and had the high purity MBP-SeFnt16798. The result of electron microscopy showed that MBP-SeFnt16798 formed nano-sized particles. Conclusion We established a stable method to obtain recombinant protein of SeFnt16798, which may lay a foundation for the development and subsequent study of FMDV structure vaccine.

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