The effect and underlying mechanism of Bu-Shen-An-Tai recipe on ovarian apoptosis in mice with controlled ovarian hyperstimulation (COH) implantation dysfunction were studied.The COH implantation dysfunction model in mice was established by intraperitoneal injection of 7.5 IU pregnant mare's serum gonadotrophin (PMSG),followed by 7.5 IU human chorionic gonadotrophin (HCG) 48 h later.Then the female mice were mated with male at a ratio of 2:l in the same cage at 6:00 p.m.The female mice from normal group were injected intraperitoneally with normal saline and mated at the corresponding time.Day 1 of pregnancy was recorded by examining its vaginal smears at 8:00 a.m.of the next day.Fifty successfully pregnant mice were equally randomly divided into 5 groups:normal control pregnant group (NC),COH implantation dysfunction model group (COH),low dosage of Bu-Shen-An-Tai recipe group (LOW),middle dosage of Bu-Shen-An-Tai recipe group (MID) and high dosage of Bu-Shen-An-Tai recipe group (HIGH).Then from day 1,the mice in different groups were respectively intragastrically given corresponding treatments at 9:00 a.m.for 5 consecutive days.The concentrations of 17β-estradiol (E2) and progesterone (P4) were determined by radioimmunoassay (RIA).The ultrastructural changes of ovarian tissues were observed by transmission electron microscope (TEM).The histopathological changes of ovarian tissues were observed by HE staining.The number of atretic follicles and pregnant corpus luteum were also recorded.TUNEL was applied to measure apoptotic cells of ovarian tissues.Western blotting was used to detect the protein expression of apoptosis-related factors like Bax,Bcl-2 and cleaved-caspase-3 in ovarian tissue of mice.The results showed that ovarian weight,the concentrations of E2 and P4,the number of atretic follicles and pregnant corpus luteum,as well as the apoptosis of granulosa cells were significantly increased in the COH group.The ultrastructures of ovarian tissues in the COH group showed that chromatin in granulosa cells was increased,agglutinated,aggregated or crescent-shaped.The focal cavitation and the typical apoptotic bodies could be seen in granulosa cells in the late stage of apoptosis.After the treatment with different doses of Bu-Shen-An-Tai recipe,the ultrastructural changes of ovarian granulosa cells apoptosis were dramatically improved and even disappeared under TEM.Visible mitochondria and mitochondrial cristae were increased and vacuoles were significantly reduced.The lipid dropltes were shown in a circluar or oval shape.The protein expression levels of Bax and cleaved-caspase-3 were decreased,and the expression of Bcl-2 protein was increased after treatment.It was concluded that Bu-Shen-An-Tai recipe can inhibit the apoptosis of ovarian granulosa cells,probably by up-regulating the protein expression of Bcl-2 and down-regulating Bax and cleaved-caspase-3,which contributes to the formation and maintenance of ovarian corpus luteum.It's helpful to promote the embryonic implantation,to reduce embryo loss and ultimately to improve the success rate of pregnancy.

OBJECTIVETo explore the association between the effort-reward imbalance at work and depressive symptoms among healthcare workers.

METHODSThe effort-reward imbalance at work was conceptualized in terms of the Chinese version of the effort-reward imbalance (ERI) model. Depressive symptoms were assessed by the Chinese version of the Center of Epidemiology Survey Depression (CES-D) scale. The data came from the cross-sectional survey of 1 179 healthcare workers aged between 18 and 73 employed in 6 affiliated hospitals of Zhejiang University. The questionnaire comprised questions on the effort-reward at work, over-commitment, the full CES-D scale of depression and a range of other characteristics. Univariate analyses were used with Spearman's correlation, Mann-Whitney test, Pearson chi(2) test and likelihood chi(2) test. Multivariate logistic regression analyses was used to discover factors associated with depressive symptoms.

RESULTSThe prevalence of depressive symptoms among healthcare workers was 48.12% (95% CI: 45.08% to 51.16%). The prevalence of depressive symptoms among nurses was 52.40% (95% CI: 47.87% to 56.93%) higher than doctors' 44.70% (95% CI: 10.64% to 48.77%) with the significant difference (chi(2) = 6.077, P = 0.014). Positive associations were found between the high effort-low reward, level of work-related over commitment and depressive symptoms (OR = 1.859, 95% CI: 1.337 to 2.585; OR = 2.207, 95% CI: 1.656 to 2.942) among healthcare workers, respectively.

CONCLUSIONThe high effort-low reward and the work-related over-commitment have a negative impact on healthcare workers' health.

Adolescent ; Adult ; Aged ; Depression ; epidemiology ; psychology ; Female ; Humans ; Logistic Models ; Male ; Medical Staff, Hospital ; psychology ; Middle Aged ; Models, Psychological ; Occupational Health ; Prevalence ; Surveys and Questionnaires

Objective To research CpG and Al(OH)3 adjuvants enhancing immunogenicity of hepatitis C virus(HCV) recombinant ptotein combined vaccine(TIE).Methods BALB/c mice were immunized with candidate vaccine TFE using CpG,Al(OH)3,Al(OH) 3 + CpG,or freund's adjuvant(FA) as the adjuvant.Five mice were sacrificed after 10 d of the last immunization.Specific antibodies in sera were tested by enzyme-linked immunosorbent assay(ELISA).Splenic cells were isolated and levels of IFN-γ,IL-4 and cytotoxic T lymphocyte(CTL) cytotoxicity assay were messuredin vitro.The remaining mice were subcutaneouly injected with 1 × 106 SP2/0-NS3 cells on the back to investigate the protective effects.The differences of means between groups were compared by LSD-t test.Results The specific CTL activity of TFE + A1(OH) 3 + CpG group was higher than TFE + FA group and TFE + CpG group(P ＜ 0.05).The level of IFN-γsecreting cells in TFE + Al(OH)3 + CpG group was higher than that in TFE + M(OH)3 group or TFE + CpG group(P ＜ 0.05).Conclusion Combining Al(OH) 3 and CpG could enhance specific cellular immunogenicity of candidate HCV vaccine TFE.TFE + M(OH) 3 + CpG could effetively prevent the attack of tumor cell SP2/0-NS3 expressing nonstructural protein NS3 of HCV.

Objective: To observe the clinical efficacy of 'Tong Du Yun Pi' (Governor Vessel-unblocking and spleen-promoting) manipulation in treating infantile diarrhea in autumn. Methods: Eighty-four kids were divided into a control group and an observation group using the random number table method, with 42 cases in each group. The control group was intervened by oral administration of montmorillonite powder, and the observation group was given additional 'Tong Du Yun Pi' pediatric massage (tuina) treatment. After treatment, the traditional Chinese medicine (TCM) symptoms scores, symptom improvement time, clinical efficacy and immune function indicators were compared between the two groups. Results: After treatment, the total effective rate was 95.2％ in the observation group versus 76.2％ in the control group, and the between-group difference was statistically significant (P<0.05); each item score in TCM symptoms was notably lower in the observation group than in the control group (all P<0.05); among the effective cases, the times to restore normal defecation, relieve abdominal bloating, arrest vomiting, and bring down the fever were markedly shorter in the observation group than in the control group, and the between-group differences were statistically significant (all P<0.05); the levels of immunoglobulin (Ig) G, IgM, CD4+ and CD4+/CD8+ were significantly higher and CD8+ was significantly lower in the observation group than in the control group (all P<0.05). Conclusion: In the treatment of infantile diarrhea in autumn, based on oral administration of montmorillonite powder, 'Tong Du Yun Pi' manipulation can notably improve diarrheal symptoms, shorten disease duration, and strengthen the immunity of kids, producing more significant efficacy than oral administration of montmorillonite powder.

OBJECTIVETo detect humoral immune response against different function regions of hepatitis C virus (HCV) in chronic patients, and further to investigate the correlativity between anti-HCV antibody titers and HCV RNA concentration.

METHODSUsing recombinant dominate epitope antigens, e.g. HCV Core, NS3, NS4, NS5 and chimeric HVR1, a set of ELISA test reagents was formulated. Then, titers of antibodies against HCV different regions and the RNA concentration of HCV in chronic patient sera were detected by ELISA and quantitative RT-PCR technique, respectively.

RESULTSGreat differences have been noted in antibody titers and positive rate of different HCV function regions in chronic patients. Antibodies against HCV Core and HVR1 have the highest positive rate, then NS3, NS4, and NS5 in sequence.

CONCLUSIONThe titer of antibodies against different regions of HCV in chronic patients has good correlation with HCV RNA concentration.

Hepatitis C Antibodies ; blood ; Hepatitis C, Chronic ; immunology ; virology ; Humans ; RNA, Viral ; blood

OBJECTIVETo investigate the association of the polymorphism in manganese superoxide dismutase (Mn-SOD) gene in Chinese type 2 diabetic patients with diabetic retinopathy.

METHODSThe Ala(-9)Val polymorphism of the Mn-SOD gene was determined by polymerase chain reaction and direct sequencing in 198 normal control subjects and 264 patients with type 2 diabetes mellitus, among them there were 139 non-diabetic retinopathy (NDR) subjects and 125 subjects with diabetic retinopathy (DR).

RESULTSThere was no statistic difference in the frequencies of VV genotype and V allele between the type 2 diabetic group and the control group. However, the frequencies of VV genotype and V allele were significantly higher in the DR group than that in the NDR group (chi-square (2)=5.015, P=0.025; chi(2)=10.253, P=0.001),but there was no statistic difference in the NDR group compared with the control group (P > 0.05). The presence of V allele was shown to be associated with diabetic retinopathy (OR=1.96, 95%CI: 1.29-2.97). Furthermore, the subjects carrying the VV genotype had lower serum Mn-SOD level (P=0.025) and had a tendency of higher total serum SOD activity, but this tendency had no statistic significance.

CONCLUSIONThe Ala(-9)Val polymorphism in the Mn-SOD gene may not be related to the etiology of type 2 diabetes, but it seems to contribute to the development of diabetic retinopathy in Chinese type 2 diabetic patients.

Alleles ; Asian Continental Ancestry Group ; genetics ; DNA ; analysis ; Diabetes Mellitus, Type 2 ; complications ; genetics ; Diabetic Retinopathy ; etiology ; genetics ; Female ; Genotype ; Humans ; Male ; Middle Aged ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Polymorphism, Single Nucleotide ; genetics ; Superoxide Dismutase ; genetics

OBJECTIVETo clone the entire coding sequence and analyze the function of P2X7 receptor of J6-1 human leukemia cells.

METHODSThe entire coding sequence of P2X7 receptor was amplified by RT-PCR and then inserted into pTARGET plasmid to construct an eukaryotic expressing plasmid followed by DNA sequencing. HEK293 cells stably expressing P2X7 receptor were obtained after transfection and screening, and confirmed by RT-PCR and Western blotting. The bleb formation upon agonist stimulation was observed under phase contrast microscope.

RESULTSThe entire coding sequence of P2X7 receptor of J6-1 cells was successfully cloned. DNA sequencing analysis revealed a substitution of G559, for A559, causing a substitution of Glu187 for Gln187. The P2X7 receptor derived from J6-1 cells could be functionally expressed in HEK293 cells, in which bleb formation could be detected upon stimulation.

CONCLUSIONSThe entire coding sequence of P2X7 receptors was successfully cloned from J6-1 leukemia cells. Other unknown mechanism may contribute to the dysfunction of P2X7 receptor in these cells.

Cell Line, Tumor ; Cloning, Molecular ; DNA, Complementary ; genetics ; Gene Expression ; Humans ; Leukemia ; genetics ; metabolism ; Receptors, Purinergic P2 ; genetics ; physiology ; Receptors, Purinergic P2X7 ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

OBJECTIVETo screen hepatocellular carcinoma (HCC) autoantibodies as diagnostic biomarkers or therapy targets by serologic proteome analysis (SERPA).

METHODSTotal proteins extracted from human HCC cell line HCCLM3 were separated by two-dimensional electrophoresis (2-DE) and then transferred onto PVDF membranes, which were subsequently incubated with sera from HCC, hepatitis B virus (HBV) infected patients or healthy volunteers. All immuno-reactive protein spots on blot films were matched to those on 2-DE gel maps by image analysis and identified by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS/MS).

RESULTS2-DE gel maps of HCCLM3 and corresponding blot films of good quality and reproducibility were established. The number of spots on HCCLM3 2-DE reference gel totaled 603 and those on HCC, HBV and healthy sera blotted films were 70.75+/-24.25, 68.5+/-23.44 and 41.38+/-15.05, respectively. Blot films of HCC and HBV groups had more spots than those of the healthy group (P < 0.05) while no significance was found between films of HCC and HBV groups. By identification, those HCC autoantibodies could be classified as nuclear proteins, cytoskeleton proteins, heat shock proteins and metabolic enzymes.

CONCLUSIONSerological proteome analysis is a high throughput technique for screening tumor autoantibodies. Those newly identified HCC associated tumor antigens and corresponding autoantibodies can be used in the early diagnosis or immuno-therapy of HCC.

Antibodies, Neoplasm ; analysis ; Autoantibodies ; analysis ; Carcinoma, Hepatocellular ; immunology ; Electrophoresis, Gel, Two-Dimensional ; Humans ; Liver Neoplasms ; immunology ; Proteomics ; methods ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Tumor Cells, Cultured

This study was purposed to investigate the effect of crocin on the proliferation in vitro and immune function of dendritic cells (DC) derived from the bone marrow of children with acute leukemia. The mononuclear cells were isolated from bone marrow of leukemia children by Ficoll-Hypaque. The experiment was divided into six groups: blank control group (A), crocin 1.25 mg/ml group (B), cytokines (rhGM-CSF 75 ng/ml+rhIL-4 75 ng/ml+rhTNF-α 50 ng/ml) group (C), cytokines+crocin 0.3125, 1.25 or 5.0 mg/ml groups (D, E, F). The numbers of DC were counted and the phenotypes of DC were determined by flow cytometry on the ninth day of culture. The DC of different groups were mixed with T cells just separated from peripheral blood of another children with acute lymphoblastic leukemia, and cultured with rhIL-2 200 U/ml for 5 d. The function of DC was detected by mixed lymphocyte reaction (MLR). The results indicated that the test groups and control group all obtained a certain amount of typical DC, but the DC numbers in test groups were all higher than those in control group (P < 0.01). Cultured for 9 days, the rates of CD1a(+), CD83(+), and HLA-DR(+) in group C, D, E, F were higher than group A (P < 0.01). There was no statistically significant difference between A and B groups (P > 0.05). MLR showed that with the increasing of DC, the stimulation index of T cells in group A and B was not rising (P > 0.05); the stimulated index of T cells in group C and E was significantly rising, there was statistically significant difference between them (P < 0.01). When the number of stimulated cells was the same, the stimulation index of T cell in group E was the highest (P < 0.01). It is concluded that the capability of DC proliferation promoted by crocin alone is lower than that of its combination with rhGM-CSF, rhIL-4 and rhTNF-α, but the crocin can synergically promote the maturity of DC cooperating with rhGM-CSF, rhIL-4 and rhTNF-α. The DC induced by crocin can particularly enhance the proliferation of T cells.
Bone Marrow Cells ; cytology ; drug effects ; Carotenoids ; pharmacology ; Cell Proliferation ; drug effects ; Child ; Dendritic Cells ; cytology ; drug effects ; Humans ; Leukemia ; pathology ; Lymphocyte Culture Test, Mixed ; T-Lymphocytes ; cytology ; Tumor Cells, Cultured

Objective To investigate the role of microRNA-10b (miR-10b) in the proliferation and invasion potential of osteosarcoma cell lines MG-63 and the exact underlying mechanism.Methods The expression level of miR-10b in human osteosarcoma tissue samples and adjacent normal bone tissues were detected by relative quantitative real-time PCR (qRT-PCR).miR-10b mimic and siRNA against Twist (Twist siRNA) were transfected into human osteosarcoma cell lines MG-63 respectively using lipofactamine 2000, and RT-PCR was used to detect the mRNA expression levels of miR-10b and Twist, and Twist protein expression level was detected by Western blot.The effect of miR-10b mimic and Twist siRNA on proliferation of MG-63 were detected by MTT[3- (4, 5-dimethylthiazol-2-yl) -5- (3-carboxymethoxyphenyl) -2- (4-sulfophenyl) -2 H-tetrazolium].The in-vitro cell invasion ability was determined by Transwell invasion assays after up-regulating miR-10b or knocking down of Twist.Results The expression levels of miR-10b was higher in human osteosarcoma tissue samples compared with adjacent normal bone tissues, the differences were extremely statistical significance (P＜0.01).miR-10b directly up regulated the mRNA and protein expression levels of Twist, the differences were significant (P＜0.05).In addition, miR-10b had enhanced the cell invasion and the proliferation (P＜0.05), whereas the proliferation and invasion ability of MG-63 which transfected by both miR-10b mimic and Twist siRNA were significantly reduced than that transfected by miR-10b mimic (P＜0.05).Conclusion miR-10b in MG-63 promotes the proliferation and invasion potential of human osteosarcoma cell lines MG-63, at least partly through the upregulation of Twist gene.