Objective To observe the PML protein expression of hepatocellar carcinoma tissue and cells lines and As 2 O3 regulate its expression .Methods Immunohistochemistry was used to examine the PML protein expression of hepatocellar carcinoma tissue . Western blot analysis were used to observe PML protein expression of hepatocellar carcinoma tissue of 12 cases ,5 hepatocellar car-cinoma cell lines ,such as HuH7 ,HepG2 ,Hep3B ,SMMC-7721 ,MHC97H .Western blot analysis was used to detected the PML pro-tein expression of these hepatocellar carcinoma cell lines after 72-96 h treated with 0 .25 μg/mL of As2 O3 .Results Immunohisch-enmical staining showed that the PML protein was expressed in both cytoplasm and nucleus ,did not well-distributed in hepatocellar carcinoma cells .There was no significant differences of PML protein expressed among differently differentiated stages of hepatocel-lar carcinoma cells .Western blot analysis found that hepatocellar carcinoma tissues of 12 cases with hepatocellar carcinoma ex-pressed PML protein ,and there was significant difference of PML protein expressed among 12 cases suffer with hepatocellar carci-noma .hepatocellar carcinoma cell lines ,such as HuH7 ,HepG2 ,Hep3B ,SMMC-7721 and MHC97H all expressed PML protein ,and there was little difference of PML protein expressed among hepatocellar carcinoma cell lines .The PML protein expression of HuH7 ,HepG2 ,Hep3B ,SMMC-7721 and MHC97H cell after 72-96 h treated with 0 .25 μg/mL of As2O3 significant decreased . Conclusion Hepatocellar carcinoma tissue and cells may express PML protein ,and As2 O3 may regulate this protein expression as well .PML protein may be the target molecule of As2 O3 treating HCC .

Objective To investigate the safety of left atrial appendage occlusion by silk thread ligation during open heart op‐eration in patients with rheumatic atrial fibrillation ,and to evaluate its effectiveness for prevention of cerebral embolism .Methods From April 2012 to March 2014 ,129 patients with rheumatic atrial fibrillation were undergone mitral valve replacement and left at‐rial appendage occlusion by ligation using two silk threads from the outside of the heart (ligation group) .The indexes related to the operation ,postoperative complications incidence ,and cerebral embolism incidence during the follow‐up period of ligation group were compared with the indexes of another 129 patients without ligation of left atrial appendage over the same period (control group) . Results The operation time ,the cardiopulmonary bypass time ,the clamp time ,the intensive care unit stay time ,the postoperative hospitalization time in ligation group were (235 ± 50)min ,(88 ± 24)min ,(57 ± 16)min ,(26 .5 ± 9 .3)h and (12 .4 ± 7 .5)d respective‐ly ,and significant difference was not found compared with control group (P>0 .05) .The thoracotomy for hemostasis(1 cases) ,low cardiac output syndrome(2 cases) ,acute renal failure(2 cases) ,pulmonary infection(3 cases) ,sternal wound dehiscence(2 cases) and other complications in ligation group had no significant difference ,compared with control group(P>0 .05);2 cases died in liga‐tion group ,3 patients died in control group ,the differences had no statistical significance(P>0 .05) .No cerebral embolism occurred in ligation group with 127 patients following‐up (23 .6 ± 11 .3) months ,but 5 patients suffered from cerebral embolism in control group with 126 patients following‐up (22 .9 ± 12 .1) months ,the difference had statistical significance(P<0 .05) .Conclusion Left atrial appendage occlusion by silk thread ligation during open heart operation in patients with rheumatic atrial fibrillation is simple and safe ,can reduce cerebral embolism incidence .

Background. Our previous studies indicated that the increased arginine vasopressin(AVP) in ischemic brain regions of gerbils could exacerbate the ischemic brain edema. This experiments is further clarify the relation between AVP and cerebral ischemia at the molecular level. Methods. The contents of AVP, AVP mRNA, AVP immunoreactive(ir) neurons in supraoptic nucleus(SON)and paraventricular nucleus(PVN) after cerebral ischemia and reperfusion were respectively determined by radioim-munoassay(RIA), immunocytochemistry( Ⅱ C), situ hybridization and computed image pattem analysis. Results. The contents of AVP in SON, PVN were increased, and the AVP ir positive neurons in SON and PVN were also significantly increased as compared with the controls after ischemia and reperfusion. And there were very light staining of AVP ir positive neurons in the other brain areas such as suprachiasmatic nucleus (SC) and periven-tricular hypothalamic nucleus (PE), but these have no significant changes as compared with the controls. During dif-ferent periods of cerebral ischemia (30～ 120 min) and reperfusion (30 min), AVP mRNA expression in SON and PVN were more markedly increased than the controls. Condusions. The transcription of AVP gene elevated, then promoting synthesis and release of AVP in SON,PVN. Under the specific condition of cerebral ischemia and repeffusion, the activity and contents of central AVP in-creased abnormally is one of the important factors which causes ischemia brain damage.

This paper was aim to screen microorganisms with attenualed efficiency for Chinese medicine containing aristolochic acid A by liquid-state fermentation. Twelve Chinese medicine were detected by UPLC and aristolochic acid A was only founded in four species of Aristolochia, those were Caulis Aristolochiae Manshuriensis, Aristolochiae Radix, Aistolochia Contorta Bunge and Herba Aristolochiae Mollissima,but not in the others. With the four Chinese medicine containing aristolochic acid A as raw material, ten microorganisms were tested, and the content of aristolochic acid A was detected by UPLC. The results showed that one microorganism can decrease content of aristolochic acid A in all those four Chinese medicine.
Aristolochic Acids ; analysis ; metabolism ; Bacteria ; metabolism ; Biotransformation ; Drugs, Chinese Herbal ; analysis ; metabolism ; Fungi ; metabolism ; Plants, Medicinal ; chemistry ; microbiology

At present time, the widely used hepatitis B virus( HBV) vaccines consist of only the small hepatitis B surface antigen expressed in yeast or CHO cells. The frequency of non-responders to these vaccines has increased the demand for a more immunogenic vaccine. Some studies have suggested that the addition of preS region to the vaccine will improve its efficacy. However, the large protein (L) containing the whole preS region can not be effectively expressed in vitro. To overcome this problem, two chimeric contructs, SS1, surface gene containing preS1 region at C-terminus and SS2, surface gene containing preS2 region at C-terminus, were constructed and effectively expressed in our previous studies. Here we further constructed an expression vector containing both SS1 and SS2 expression cassettes by separation and ligation the SS2 cassette to a linearized SS1 expression vector pAO815-SS1. The recombinant vector was transformed into Pichia pastoris GS115 by electroporation. A high-level expression strain (GS115-SS1S2) was established by primary screening for His+ transformants and further analysis for induction products. ELISA results demonstrated that the expressed protein had S, preS1 and preS2 antigenicities simultaneously. Western blotting showed that the product can bind to all of the three antibodies, anti-S, anti-preS1 and anti-preS2. The expression protein was present in the form of particles of 20-35 nm diameter and the yield of recombinant particles reached 300-600 mg/L by fermentation. The SS1 and SS2 polypeptides kept intact in purified particles, suggesting that the stability of preS region has been significantly improved.
Animals ; CHO Cells ; Cricetinae ; Cricetulus ; Epitopes ; genetics ; immunology ; Hepatitis B Surface Antigens ; genetics ; immunology ; Hepatitis B Vaccines ; immunology ; Peptide Fragments ; genetics ; immunology ; Pichia ; genetics ; Protein Precursors ; genetics ; immunology ; Vaccines, Synthetic ; immunology

The preparation process and immunogenicity of a novel hepatitis B vaccine containing preS1, preS2 and S epitopes were investigated in this study. A Pichia pastoris stain GS115-SS1S2 harbouring two chimeric HBsAg gene constructs, SS1 and SS2 was cultivated by high-density fermentation. 300-600 mg/L of the expression level was achieved through 48-72 h methanol induction. SSIS2 antigen was extracted and purified by silica adsorption, HIC and SEC to 99% purity from the harvested cells. 82 mg purified antigen could be achieved from one liter of fermentation culture. The immunogenicity of the purified antigen was evaluated in NIH mice. Three groups of female NIH mice, 14-16 g in weight, were injected once intraperitoneally with 2.5, 0.625, 0.156 microg of each of the two vaccines: SS1S2 or a commercially available S vaccine. Part of the mice were bled in 30 days after injection to compare the ED50 of the two vaccines. For the SSIS2 vaccine, the ED50 is 0.46, 0.29 and 0.84 microg respectively for the preS1, preS2 and S antigens. For the S vaccine, the ED50 is 0.99 microg for the S antigen. Another part of the mice were bleed in 7 or 14 days to detect preS1, preS2 and S antibodies. Higher ratios of mice were seroconverted for preS1 and preS2 antibodies as compared to the S antibody in these two time points. These results suggest that the SS1S2 vaccine may be more immunogenic than the conventional S vaccines.
Animals ; Epitopes ; immunology ; Female ; Hepatitis B Surface Antigens ; immunology ; Hepatitis B Vaccines ; biosynthesis ; immunology ; Mice ; Pichia ; genetics ; metabolism ; Protein Engineering ; Protein Precursors ; immunology ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; Viral Envelope Proteins ; immunology

Objective To investigate the effects of different dialysates on expression of protein kinase C-δ (PKCδ) and apoptosis of U937 cell line. Methods Different dialysates were added into culture fluid with U937 cell line at exponential phase of growth, and groups were divided: fluid A+fluid B group (dialysate A+dialysate B), fluid A+fluid B+rottlerin (PKCδ specific inhibitor)group, fluid A+powder B group (dialysate A+powder B) and fluid A+powder B + rottlerin group. Besides, blank control group and normal control group were established. Cells were harvested 24 h and 48 h after treatment, morphological changes were observed by Hoechst33258 fluorescence staining, cell apoptosis was measured by Annexin-V-FITC/PI double staining, and expression of PKCδ mRNA and protein was detected by RT-PCR and Western blotting, respectively. Results Cell apoptosis significantly increased in fluid A+powder B group, with typical morphology of apoptosis. After treatment for 24 h and 48 h, cell apoptosis rates in fluid A+powder B group were significantly higher than those at corresponding time points in blank control group , normal control group and fluid A+powder B+rottlerin group (P<0.05). Compared with normal control group, blank control group and fluid A+powder B+rottlerin group, the expression of PKCδ mRNA and protein of U937 cells in fluid A+powder B group were significantly increased (P<0.05). There was no significant difference in cell apoptosis rates and expression of PKCδ mRNA and protein between fluid A+fluid B group and blank control group, normal control group and fluid A+fluid B+rottlerin group (P＞0.05). Conclusion Fluid A+powder B can significantly increase apoptosis of U937 cell line, the mechanism of which may be associated with the up-regulation of expression of PKCδ. Compared with fluid A+powder B, fluid A+fluid B is superior in reducing apoptosis of peripheral blood monouclear cells.

OBJECTIVESTo study the efficacy of proximal embolic protection device in preventing intracranial artery embolization during carotid artery stenting (CAS) and to evaluate its security and maneuverability.

METHODSFrom October 2007 to July 2008, 23 patients with carotid artery stenosis who were suitable for surgical therapy according to the standards of NASCET or ACAS were enrolled in this clinical research. Among them 19 patients (82.6%) were symptomatic, 6 patients (26.1%) with 50%-70% stenosis and 17 cases (73.9%) with > 70% stenosis. All the patients received carotid angioplasty and stenting under the protection of MO. MA system (one kind of proximal embolic protection device). We recorded the cerebral ischemic time during the procedure and observed neurologic events within 30 days.

RESULTSAll the procedures were performed successfully, the mean carotid artery blocking time was (5.3 +/- 1.2) min. No death or stroke occurred during perioperative period. Two cases of patients developed transient loss of consciousness combined with contralateral limb convulsion, while the common carotid artery was occluded by balloon. Two cases of patients developed bradycardia, sustained 6 hours and 1 week. Plaque debris in the withdrawal blood from carotid artery were found in 9 cases. At 30-day follow-up after CAS, TIA occurred in 1 case, new contralateral stroke occurred in 1 case, the incidence of 30-day stroke and death rate was 4.3%.

CONCLUSIONThe application of proximal embolic protection device in CAS procedure for preventing neurologic complications is safe and effective, especially for severe stenosis and unstable plaque in carotid artery stenting.

Aged ; Aged, 80 and over ; Angioplasty, Balloon ; instrumentation ; methods ; Carotid Stenosis ; surgery ; Embolic Protection Devices ; Female ; Follow-Up Studies ; Humans ; Intracranial Embolism ; etiology ; prevention & control ; Male ; Postoperative Complications ; prevention & control ; Stents ; Treatment Outcome

OBJECTIVESTo investigate the internal links between immune responses and Tregs and cytokine by the expression of T regulatory cells (Tregs), Foxp3 mRNA of different response groups and the detection of cytokine secretion after hepatitis B vaccination.

METHODSBlood samples were collected in different response groups. Real-time fluorescence quantitative PCR was used to detect the expression of Foxp3 mRNA of peripheral blood mononuclear cells; The surface markers CD4 and CD25 in peripheral-blood mononuclear cells were determined by flow cytometry; ELISA tests were used to detect the production level of phytohemagglutinin (PHA) in peripheral blood mononuclear cells, IL -4, IL-12, IL-18 stimulated by HBsAg and (IFN) gamma.

RESULTS(1) Foxp3 expressions in response group and non-response group were higher before or after PHA and HBsAg were stimulated. Differences were statistically significant (P value less than 0.05) ; (2) In peripheral blood, the percentage of CD4+CD25+ Treg of CD4+ T cells in response group (0.59%+/-0.46%) was obviously lower than those in control group (1.30%+/-1.44%) ; (3) Peripheral blood mononuclear cells stimulated by PHA and HbsAg in each group, the concentration of IFNgamma in non-response group [(11.00+/-9.03) IU/ml] was markedly lower than those in response group [(38.88+/-28.16) IU/ml],and differences were statistically significant (P value less than 0.01); (4) In PHA- or HBsAg-stimulated peripheral-blood mononuclear cells, the concentrations of IL-18, IL-4 and IL-12 had no significant difference.

CONCLUSIONSTo some extent, CD4+CD25+Foxp3+ Treg cells may be involved in negative regulation of the immune responses to HBV vaccination. Immune non-response to HBV vaccination may be connected to insufficient secretion of IFNgamma; There was no correlation between the titer of anti-HBs and the expressions of IFNgamma and CD4+CD25+ Foxp3.

Adolescent ; Antibody Formation ; CD4 Antigens ; metabolism ; Female ; Forkhead Transcription Factors ; immunology ; Hepatitis B ; immunology ; prevention & control ; Hepatitis B Vaccines ; immunology ; Hepatitis B virus ; immunology ; Humans ; Interferon-gamma ; immunology ; Interleukin-12 ; immunology ; Interleukin-18 ; immunology ; Interleukin-2 Receptor alpha Subunit ; metabolism ; Interleukin-4 ; immunology ; Male ; T-Lymphocytes, Regulatory ; immunology ; Young Adult

Biliary Fistula ; diagnosis ; Bronchial Fistula ; diagnosis ; Female ; Humans ; Middle Aged