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ObjectiveThe mechanism of histone phosphorylation modification in oocyte meiosis is less studied. This study is designed to investigate the pattern of histone H3 phophorylation and regulation of maturation process in the porcine oocytes.MethodsThe histone H3Ser10 (H3S10) phosphorylation expression was examined on the porcine oocyte meiotic process. The porcine cumulus oocyte complexes (COCs) were divided into four groups, one group was cultured as control group, and the other 3 groups were supplemented with 5, 10, and 30 μmol/L ZM447439, and cultured in vitro for 27 h, respectively, 5, 10, and 30 μmol/L ZM4474349 treatment group. The proportion of each meiotic stage was counted. The phosphorylation pattern of histone H3S10 and the expression level of protein kinase Aurora B were detected at the porcine oocytes.ResultsCompared with histone H3S10 phosphorylation level of oocyte GVBD phase, the MI and AI phases were significantly increased (P<0.05), and H3S10 phosphorylation level of AI phase was remarkedly higher than that of MII phase (P<0.05). Compared with the control group, the proportion of oocytes at the GVBD phase in the 10 and 30 μmol/L ZM4447439 treatment group [(32.14±0.51)%, (95.34±0.59)%]was higher than that of the control group [(2.56±0.03)%, P<0.05], the proportion of oocytes at the MI phase [(66.88±0.13)%, (4.66±0.04)%] significantly decreased than that of the control group [(87.42±0.14)%, P<0.05], and the proportion of oocytes at the AI stage [(1.01±0.03)%, (0.000±0.00)%] significantly decreased compared with the control group[(10.02 ± 0.21)%, P<0.05]. Compared with the control group (0), oocytes H3S10 dephosphorylation modification ratio in the 10 μmol/L and 30 μmol/L ZM4474349 treatment group [(35.2±0.39)%, (95.4±0.65)%]significantly increased (P<0.05). Compared with the control group, the relative expression level of Aurora B in the 10 and 30 μmol/L ZM4447439 treatment group was significantly reduced (P<0.05).ConclusionHstone H3S10 phosphorylation plays arolein the maturation of mammalian oocytes. AuroraB kinase inhibitors (ZM447439) treatment can reduce H3S10 phosphorylation and Aurora B expression level and lead to oocytesmaturation disorder.
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Objective To analyze differentially expressed proteins in A375 melanoma cells before and after short hairpin RNA (shRNA)-mediated Cbl-b gene silencing.Methods The label-free quantitative proteomics approach was performed to identify differentially expressed proteins between A375 cells transfected with lentiviral vectors containing Cbl-b shRNA (Cbl-b shRNA group) and those with control lentiviral vectors (control group).Then,the properties of differentially expressed proteins were analyzed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genome (KEGG) enrichment analysis.Western blot analysis was conducted to determine the expression of differential proteins (EphA2 and GSK3β) and phosphorylated protein kinase (p-AKT) after shRNA-mediated Cbl-b gene silencing.Statistical analysis was carried out by t test of two independent-samples for comparison of protein expression abundance between the two groups with SPSS 23.0 software.Additionally,the results of GO and KEGG enrichment analysis were analyzed by Fisher's exact test.Results A total of 3 449 proteins were identified and quantified,and 74 of them were differentially expressed between the Cbl-b shRNA group and control group.Compared with the control group,52 proteins were up-regulated and 22 were down-regulated in the Cbl-b shRNA group.GO enrichment analysis of differential proteins revealed that the top five significantly enriched biological processes were integrin-mediated cell adhesion,single-organism metabolic process,regulation of integrin-mediated cell adhesion,regulation of protein-targeting mitochondria and nucleic acid metabolic process.The top five significantly enriched molecular functions included DNA binding,2-iron,2-sulfur cluster binding,signaling receptor activity,cadherin binding and cell adhesion molecule binding.The top five significantly enriched cell components included nucleosome,DNA packaging complex,photoreceptor connecting cilium,DNA-protein complex and extracellular region part.KEGG enrichment analysis demonstrated that the top five significantly enriched melanoma-related signaling pathways were folate biosynthesis,axon guidance,extracellular matrix-receptor interaction,adherens junction and Wnt signaling pathways.As Western blot analysis revealed,the Cbl-b shRNA group showed lower protein expression of EphA2 (0.369),but higher protein expression of GSK3β (3.524) compared with the control group (1),which were consistent with the results of proteomics analysis.Additionally,the protein expression of p-AKT was down-regulated in Cbl-b shRNA group (0.453) compared with the control group (1).Conclusion Cbl-b may be involved in the occurrence of melanoma through a variety of biological pathways,and the EphA2/PI3K/AKT signaling pathway may be one important pathway.
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Objective To construct a recombinant lentiviral vector carrying the casitas B-lineage lymphoma (Cbl)-b short hairpin RNA (shRNA),and to evaluate its effect on the biological behavior of A375 melanoma cells in vitro.Methods Three specific shRNAs targeting Cbl-b gene and a negative control shRNA were designed and synthesized,and recombinant lentiviral vectors were constructed.A375 cells were divided into 5 groups to be transfected with 3 kinds of lentiviral vector expressing Cbl-b genespecific shRNAs (CBLB-shRNA-1 group,CBLB-shRNA-2 group and CBLB-shRNA-3 group),a lentiviral vector containing negative control shRNA (negative control group),and an empty vector (blank control group).Real-time fluorescence-based quantitative PCR and Western blot analysis were performed to determine the silencing efficiency at 72 hours after transfection.Cell counting kit-8 (CCK-8)assay was conducted to evaluate cellular proliferative activity at 24,48,72 and 96 hours after transfection,flow cytometry to detect cell apoptosis and cell cycle at 72 hours after transfection,and Transwell invasion assay to assess cellular invasive activity at 72 hours after transfection.Results Three recombinant lentiviral vectors containing Cbl-b shRNA were constructed successfully.As Western blot analysis revealed,the CBLB-shRNA-3 showed the highest silencing efficiency.CCK-8 assay indicated that the proliferative activity of A375 cells was significantly lower in the CBLB-shRNA-3 group than in the negative control group and blank control group at 72 and 96 hours after transfection(all P ＜ 0.01).Flow cytometry showed that the apoptosis rate of A375 cells was significantly higher in the CBLB-shRNA-3 group (22.73％ ± 6.58％) than in the negative control group (6.08％ ± 1.35％,P ＜ 0.01) and blank control group (6.34％ ± 1.07％,P ＜ 0.01).The CBLB-shRNA-3 group showed a significantly higher proportion of A375 cells at G1 phase,but a significantly lower proportion of A375 cells at S phase compared with the negative control group and blank control group(all P ＜ 0.01).Transwell assay showed that there were significant differences in the number of A375 cells crossing the artificial basement membrane (matrigel) at 72 hours after transfection among the negative control group,blank control group and CBLB-shRNA-3 group (76.60 ± 1.82,73.20 ± 3.83,19.60 ± 1.14,respectively;F =794.50,P ＜ 0.01).Conclusions A recombinant CBLB-shRNA-3-expressing lentiviral vector which can efficiently silence Cbl-b gene has been successfully constructed.It can inhibit the proliferation,cell cycle progression and invasive activity of A375 cells,but promote the apoptosis of A375 cells.
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Objective To observe the clinical efficacy of Fu San Zhen (acupuncture at three abdominal acupoints) plus Xing Nao Kai Qiao (brain-awakening and orifice-opening) needling in treating urinary incontinence after cerebral infarction. Method Sixty patients with urinary incontinence after cerebral infarction were randomized into an experimental group of 20 cases and a control group of 20 cases. The experimental group was intervened by Fu San Zhen [Qihai (CV6) and bilateral Shuidao (ST28)] plus Xing Nao Kai Qiao needling by selecting Shuigou (GV26), Neiguan (PC6) and Sanyinjiao (SP6). The control group was intervened by ordinary acupuncture [Zhongji (CV3), Qihai (CV6), Shenshu (BL23) and Pangguangshu (BL28)] plus Xing Nao Kai Qiao needling method by selecting Shuigou (GV26), Neiguan (PC6) and Sanyinjiao (SP6). The maximum bladder capacity, residual urine volume and the score of International Consultation on Incontinence Questionnaire Short-form (ICIQ-SF) of the two groups were observed before and after the intervention, and the clinical efficacies were also compared. Result After the intervention, the maximum bladder capacity, residual urine volume and the score of ICIQ-SF were significantly changed in both groups (P<0.01). There were significantly statistical differences in comparing the maximum bladder capacity, residual urine volume and the score of ICIQ-SF between the two groups after the treatment (P<0.01). The total effective rate was 95.0％ in the experimental group versus 75.0％ in the control group, and the difference was statistically significant (P<0.01). Conclusion Fu San Zhen plus Xing Nao Kai Qiao needling can produce a more significant efficacy in treating urinary incontinence compared to ordinary acupuncture plus Xing Nao Kai Qiao needling.
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Biomarkers are the detectable changes associated with physiological or pathological changes. Urine as excreta of the body, without the mechanisms to maintain a homeostatic internal environment, can reflect a variety of changes in the body. Using animal models can simulate human disease processes, monitor disease changes, and provide clues to early diagnosis. Rats as commonly used model animals are not the dominant models for all disease, thus comparing the urinary proteins of rats with other animals to provide clues to the selection of other animal dominant models. In this study, urinary proteins were digested and profiled by liquid chromatography and tandem mass spectrometry (LC-MS/MS). The urinary proteins of rats, guinea pigs and golden hamsters were compared. The results showed that the number of urine proteins in the three different animals was different, and also different in every system of the body. This provides a basis for selecting the best animal models for different diseases.
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Objective To investigate the clinical therapeutic effect of biological information infrared liver therapeutic appa-ratus(BILT)combined with praziquantel in the treatment of patients with chronic schistosomiasis. Methods A case-control study was conducted. A total of 142 chronic schistosomiasis patients were divided into an experimental group(BILT combined with praziquantel)with 64 cases and a control group(routine treatment with praziquantel alone)with 78 cases on the basis of the age,gender,disease duration and liver function as paired condition. Fatigue,diarrhea,abdominal distension,liver func-tion,hyaluronic acid(HA)and laminin(LN)were as observation indexes and the observation results were compared between two groups. Results Before the treatment,there were no significant differences between the two groups in the indexes above-mentioned(P>0.05). After the treatment,the incidence rates of fatigue,diarrhea,abdominal distension,abnormal liver func-tion,and the levels of HA and LN in the experimental group were significantly lower than those in the control group(P<0.01). Conclusion BILT combined with praziquantel can significantly alleviate the short-term clinical symptoms,restore liver func-tion and also alleviate hepatic fibrosis of the patients with chronic schistosomiasis.
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Objective To analyze the podiatric conditions of athletes with intellectual disabilities.Methods From May eleventh to 17th, 2015,779 athletes who participated in the sixth National Special Olympics were enrolled partly.Their lower limbs and feet were evaluated with Fit Feet Healthy Athletes Screening Form.Results Common podiatric conditions found in Special Olympics athletes included flat feet (52.2% in left side and 51.7% in right side), over pronation (56.1% in left side and 56.9% in right side), callus (44.7%), wrong nail cut (46.6%),ingrown nail(21.1%),and cross over toes(6.8%),etc.Conclusion Special Olympics athletes have podiatric structural,functional, nail and skin problems,which may relate to intellectual disabilities.
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Objective To investigate the clinical features,pathological features and prognostic factors of young patients with endometrial cancer.Methods One hundred and ten cases of endometrial cancer under 40 in Jiangning Hospital Affiliated to Nanjing Medical University from January 1st 2008 to January 1st 2012 were selected as the study group while one hundred and ten cases of cervical cancer over 40 in the same period as the control group.The clinical features and pathological features were compared between the two groups.Univariate and multivariate analysis were used to determine the prognostic factors.Results The occurrence rate of abnormal vaginal bleeding (74.5% (82/110)),vaginal drainage (22.7% (25/110)) and infertility (38.2% (42/110)) in the study group were significantly lower than those in the control group (59.1%(65/110),4.5%(5/110),3.6%(4/110),χ2=4.30,12.23,16.53,P0.05).There were differences in histology,PR,muscle,and lymph node metastases (χ2=5.39,4.29,4.89,P<0.05).Multivariate analysis of prognostic factors in young patients with endometrial cancer showed that histology,PR and muscle metastasis were independent risk factors for the prognosis of young endometrial cancer patients.Conclusion There are differences in clinical features and pathological features between young and elderly endometrial cancer patients.The census and prevention among young women should be strengthened and the individualized treatment and the prevention and control system should be upgraded.
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Objective:To explore the function of IL-9 and PU.1 on genesis and development of pulmonary fibrosis,and the effect of active vitamin D3[1,25(OH)2VD3] on the expression levels of this two factors during the pathogenesis of fibrosis.Methods: 90 male SD rats were randomly divided into model group,treatment group,control group (n=30).Bleomycin(5 mg/kg) was injected into the trachea of rats to establish the model of pulmonary fibrosis in the model group and treatment group,while the control group was injected with isopyknic sterile saline.The treatment group,the model group and the control group were injected intraperitoneally with active vitamin D3,solvent of vitamin D3 (propylene glycol) and sterile saline on the 2nd day after surgery respectively.All injections were carried out once every other day.10 rats were euthanized at 14th,21st and 28th day in each group in turn.After obtaining lung tissues from experimental rats,the pathological change of lung was compared by hematoxylin-eosin staining.The difference of collagen fiber and hydroxyproline content were compared by the Masson staining and basic-hydrolysis method respectively.The mRNA and protein expression of IL-9 and PU.1 in lung tissue were detected by Real-time PCR and immunohistochemical technology respectively.The expression of IL-9 in serum was detected by ELISA.Results: Fibrosis appeared in lungs of experimental rats treated with bleomycin after 14 days,and more and more aggravated with time.At three time points,the hydroxyproline content in model group and treatment group were significantly higher than that of control group,and the treatment group was significantly lower than the model group.At three time points,the expression of IL-9 and PU.1 in model group and treatment group were risen gradually,and obviously higher than that in control group.On the 14th and 21st day,the expression of two factors in treatment group was significantly lower than model group;on the 28th day,there was no statistically significant difference between treatment group and model group(P>0.05).In model group and treatment group,the expression of two factors on 21st day was significantly higher than that on 14th day;there was no statistically significant difference between the 28th day and the 21st day.Conclusion: IL-9 and PU.1 may play a profibrotic role at early stage of pulmonary fibrosis induced by bleomycin.The active vitamin D3 may lower the expression level of PU.1,and then reduce the secretion of IL-9,thus may play an inhibiting effect on genesis and development of pulmonary fibrosis in rats.
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@#: Objective To investigate the general physical conditions of Special Olympics athletes in China. Methods May 11th to 17th, 2015, 777 Special Olympics athletes of Sixth National Special Olympics accepted FUNFitness survey to be evaluated in muscle flexibility, muscle strength, balance and aerobic fitness. Results There were 727 Special Olympics athletes needing more muscle flexibility training guidance from physical therapists, 365 needing muscle strength training guidance, 452 needing balance training guidance and 272 needing aerobic fitness training guidance. Conclusion Special Olympics athletes need more care during the training. They need special training guidance in flexibility, endurance and balance from physical therapists.
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Objective Idiopathic pulmonary fibrosis ( IPF) is a chronic inflammatory disease with unknown etiology and is lack of effective therapy. The aim of this study is to explore the function of Ca2+ and PI3K/AKT/mTOR pathway in the pathogenesis of IPF, and the impact of 1,25?( OH) 2 D3 on Ca2+ and PI3K/AKT/mTOR pathway in type Ⅱalveolar epithelial cells of rat with IPF. Methods 150 male SD rats were randomly divided into 2 groups: prevention group ( control groupⅠ, model groupⅠ, medication groupⅠ) and treatment group ( control groupⅡ, model groupⅡ, medication groupⅡ) . The tracheal exposure surgery was operated in control groupⅠ/Ⅱ, and then 200μL sterile physiological saline was administered by intraperitoneal injection of each rats 2 days and 14 days after surgery, separately. Bleomycin(BLM)(5 mg/kg) was in?jected into the trachea of model groupⅠ/Ⅱ, and then vitamin D3 solvent(0.1%ethanol and 99.9%glycol propylene, 1μL/g) was ad?ministered by intraperitoneal injection 2 days and 14 days after surger?y, separately. Bleomycin( BLM) ( 5 mg/kg) was injected into the tra?chea of medication groupⅠ/Ⅱ, and then 1,25?( OH) 2 D3( 2μg/kg) was administered by intraperitoneal injection 2 days and 14 days after surgery, separately. IPF model was built by injecting Bleomycin into the trachea of rats, 1,25?(OH)2D3(2μg/kg) was used to prevent and treat IPF by intraperitoneal injection in medication group. The hydroxyproline content of lung tissue in each group was measured, type Ⅱalveolar epithelial cells were separated from lung tissue and labeled with Fluo?3AM, then concentration of Ca2+ was detected by Laser scanning confocal microscope. The mRNA levels of PI3K, AKT and mTOR in the typeⅡalveolar epithelial cells were tested by RT?PCR. Results Compared with control groupⅠ/Ⅱ at each time point, hydroxyproline content of lung tissue, Ca2+ concentration and expression of PI3K, AKT and mTOR in typeⅡalveolar epithelial cells in model groupⅠ/Ⅱand medication groupⅠ/Ⅱwere sig?nificantly raised( P<0.05 or P<0.01) , but these were significantly reduced in medication groupⅠ/Ⅱcompared with model groupⅠ/Ⅱ( P<0.05 or P<0.01) . Correlation analysis showed that there is significant positive correlation between Ca2+ concentration and mRNA expression levels of PI3K, AKT and mTOR in model groupⅠ/Ⅱ(r=0.5988, r=0.6230, r=0.6603,P<0.01)and medication groupⅠ/Ⅱ( r=0.701 2, r=0.632 3,r=0.740 3,P<0.01) . Conclusion The PI3K/AKT/mTOR pathway plays an important role in devel?opment of IPF. 1,25?( OH) 2 D3 is able to reduce Ca2+concentration in typeⅡalveolar epithelial cells and inhibit the PI3K/AKT/mTOR pathway, and then inhibit the development of IPF.
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Point Dingchuan (Ex-B1) pertains to extraordinary points and often produces a good therapeutic effect on respiratory system diseases. Ancient Chinese medical books do not record point Dingchuan. Modern Chinese medical textbooks describe the location and efficacy of and indications for this point but do not state their provenances. In order to further popularize the clinical application of point Dingchuan, this article verifies the origin and development of the name and location of point Dingchuan so as to provide a certain reference for clinically correct selection and use of this point. The results show that point Dingchuan followed the course of development from ashi point to new point and then to extraordinary point. The development of point Dingchuan to the same acupoint name and location as now international use went mainly through four stages, that is, similar name and different location, similar name and location, same name and different location, and same name and location.
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Objective:To compare the sealing ability and marginal adaptation of three obturation techniques when used in root canal obsturation of c-shaped canals of mandibular second molars. Methods:Twenty-one extracted mandibular second molars with C-shaped canals were randomly divided into three groups. After instrumentation, the canals were filled using three different techniques: ObturaⅡ(group A), cold lateral condensation (group B) and Thermafil (group C). Then, gaps,obturated lateral canals and reticular apical triangles on the X-ray were counted. After dyeing for 7 days in the ink, dyeing lines were measured to reflect the microleakage. The percentages of gutta-percha, sealer and gaps on the root canal surface were determined by analyzing the images of 3 sections per tooth. Results:The reticular apical triangles were most frequently observed in group C(P
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With the wide application of triazophos in agricultural production, increasing attention has been paid to the contamination of this pesticide to the environment. The studies on triazophos residue and degradation have made certain progress in recent years.This paper gives an outline of micro-organisms degrading triazophos and the possible pathways by which they degrade this pesticide. The reaction kinetics and mechanisms of triazophos hydrolysis and photolysis are also reviewed briefly.
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A bacterium strain BFHM2002 is isolated from Lake Donghu, Wuhan. BFHM2002 has advantages that it can produce melanin with a high rate and high yield in the absence of tyrosine. Induced by tyrosine, melanin yield can be dramatically increased. BFHM2002 may be identified as a new strain in Bacillus firmus, for melanin-production.