Objective To study the effects of As_2O_3 on tumor model of hepatocarcinoma.Methods HepAgrafed hepatocarcinoma mouse model was established by subcutaneously injection of mouse hepatoma cells(1×10~6)into the oxter of mice.After treated by As_2O_3,the volume change of tumor and tumor inhibition rates were observed.The expression of vascular endothelial growth factor(VEGF) was detected by immunohistochemical and calculated the difference of MVD.Results The volume of tumor and the tumor inhibition rates were significantly decreased in As_2O_3 group compared with control group(P<0.05).The As_2O_3 could inhibit angiogenesis of xenograft tumor,depress expression of VEGF and decrease microvascular density(MVD).Conclusion As_2O_3 can inhibit the growth of tumor,inhibit the expression of VEGF and decrease MVD.

To understand the immunogenicity of purified inactivated severe fever with thrombocytopenia syndrome bunyavirus (SFTSV), concentration by ultrafiltration as well as molecular-sieve chromatography (MSC) were used for purification of inactivated SFTSVs. Inactivated viruses in purified samples were analyzed and identified by western blotting and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the glycoprotein (GP) and nucleoprotein (NP) antigen titers of which were detected using a double-sandwich enzyme-linked immunosorbent assay (ELISA). Purified inactivated SFTSVs were enriched and observed by electron microscopy, and the total protein concentration detected using the bicinchoninic acid assay. Purified inactivated SFTSVs were applied to New Zealand rabbits via two immunization programs to evaluate immunogenicity and to compare the immune effect. After SFTSVs were inactivated and concentrated by ultrafiltration, MSC revealed two typical elution peaks. The sample of one peak was identified as inactivated virions, in which GP and NP were detected by SDS-PAGE, western blotting and ELISA. Main corponent of the other peak was NP. After concentration by ultrafiltration, purified inactivated SFTSVs with purity >90% and total protein concentration of 1. 1 mg/mL were obtained, and the typical electron microscopy of bunyavirus was observed. In the sera of animals immunized with purified inactivated SFTSVs, SFTSV-specific IgG antibody and neutralizing antibody were detected at high titers. However, antibody titers were affected by the immunization program. Effect of immunization on days 0, 14 and 28 was significantly better than that on days 0, 7 and 28. Our work revealed that cultivation of SFTSVs contained intact virus particles and large amounts of free NP. Using MSC, purified inactivated SFTSVs of high purity could be obtained. Purified inactivated SFTSVs induced high titers of neutralizing antibody and virus-specific IgG antibody showing satisfactory immunogenicity, which provides important clues for further study on a vaccine for the inactivated virus.
Animals ; Antibodies, Neutralizing ; immunology ; Antibodies, Viral ; immunology ; Bunyaviridae Infections ; immunology ; virology ; Humans ; Neutralization Tests ; Phlebovirus ; classification ; genetics ; immunology ; isolation & purification ; Rabbits

To prepare monoclonal antibodies (mAbs) against structural proteins of severe fever with thrombocytopenia syndrome bunyavirus (SFTSV), BALB/c mice were immunized using purified inactivated SFTSV virions as the antigens. Subsequently, hybridoma cell lines that secreted monoclonal antibodies against nucleoprotein (NP) and glycoproteins (GP) were obtained using a hybridoma technique. The antigen specificities of prepared mAbs were examined by indirect immunofluorescence and immunoprecipitation assays. Functional analyses were then performed,including the detection of IFA antibody titers,the levels of neutralizing activity and antibody affinities. After cell fusion and cloning,13 hybridoma cell lines secreted mAbs specifically against SFTSV-GP and 7 hybridoma cell lines secreted mAbs specifically against SFTSV-NP. Immunofluorescence and immunoprecipitation assays showed that the mAbs had high levels of antigen specificity. Among the 13 anti-SFTSV-GP mAbs,6 recognized Gn,whereas the others reacted with Gc. IFA titers of most anti-SFTSV-GP mAbs were between 1,280 and 20,480, and four anti-SFTSV-Gn mAbs showed neutralizing activity. Seven of the obtained anti-SFTSV-NP mAbs reacted specifically with NP,of which the IFA titers ranged from 5,120 to 20,480 with no observed neutralizing activity. Furthermore, two anti-SFTSV-GP mAbs, 1C8 and 1G8, showed high levels of affinity via a non-competitive ELISA. Our study lays the foundation for the development of further diagnostic assays and basic research into SFTSV.
Animals ; Antibodies, Monoclonal ; immunology ; Antibodies, Viral ; immunology ; Antibody Specificity ; Bunyaviridae Infections ; immunology ; virology ; Female ; Humans ; Hybridomas ; immunology ; Mice ; Mice, Inbred BALB C ; Phlebovirus ; immunology ; Viral Structural Proteins ; immunology

This article aimed to study the antigenicity of nucleocapsid proteins (NPs) in six pathogenic phleboviruses and to provide theoretical evidence for the development of serological diagnostic reagents. NPs of six pathogenic phleboviruses were expressed and purified using a prokaryotic expression system and rabbits were immunized with individual recombinant NPs. Cross-reactions among NPs and rabbit sera were determined by both indirect ELISA and Western blotting analyses, and the sera titer was determined by indirect ELISA. Furthermore, sera from SFTS patients were also detected by each recombinant NP as a coating antigen using indirect ELISA. The cross-reactions and the sera titer were subsequently determined. Both the concentration and purity of recombinant NPs of six pathogenic phleboviruses met the standards for immunization and detection. The results of indirect ELISA and Western blotting showed that each anti-phlebovirus NP rabbit immune serum had potential serological cross-reactivity with the other five virus NP antigens. Furthermore, the sera from SFTS patients also had cross-reactivity with the other five NP antigens to a certain extent. Our preliminary study evaluated the antigenicity and immune reactivity of six pathogenic phleboviruses NPs and laid the foundation for the development of diagnostic reagents.
Animals ; Antibodies, Viral ; immunology ; Antigens, Viral ; genetics ; immunology ; Cross Reactions ; Humans ; Nucleocapsid Proteins ; genetics ; immunology ; Phlebotomus Fever ; diagnosis ; immunology ; virology ; Phlebovirus ; classification ; genetics ; immunology ; isolation & purification ; Rabbits

Acetates ; therapeutic use ; Acute Disease ; Anti-Asthmatic Agents ; therapeutic use ; Asthma ; prevention & control ; Child ; Child, Preschool ; Female ; Humans ; Leukotriene Antagonists ; therapeutic use ; Male ; Quinolines ; therapeutic use

Three pSIREN-siRNA plasmids were constructed using a pSIREN-RetroQ vector to silence the expression of muhidrug resistance-associated protein (MRP1) gene, and subsequently characterized by restriction endonuclease digestion and DNA sequencing. A truncated MRP1 and a full-length MRP1 were cloned into pEGFP-N2 and PeDNA3.1 respectively as pEGFP-MRP1T and pcDNA-MRP1. The plasmid pEGFP-MRP1T was co-transferred with each of the three pSIREN-siRNAs into HEK293 cells for MRP1T-GFP targeted silencing, and pSIREN-siRNA1 was used as the negative control, pSIREN-siRNA2 and pSIREN-siRNA3 appeared to be more effective to silence MRP1T-GFP compared to pSIREN-siRNA1 as shown by fluorescence microscopy. For the silencing of full-length MRP1 expression, HEK293 ceils were co-transferred with pcDNA-MRP1 and either of the three pSIREN-siRNAs, then subjected for Western blot analysis and MTT assays, pSIREN-siRNA2 and pSIREN-siRNA3 were able to inhibit the expression of 190 kD MRP1, but not pSIREN-siRNA1. The MDR of MRP1-transfected HEK293 ceils was abolished with pSIREN-siRNA2 or pSIREN-siRNA3 transfections. RNA secondary structure predictions demonstrated that the mRNA local free energy (△G) of the siRNA1 targeted sequence was lower, as the GC content and Tm value of siRNA1 were higher than those of siRNA2 and siRNA3. These data suggest that the local structure siRNAs and target mRNA may influence the silencing efficiency of MRP1 expression.

Objective To investigate the role of UDP-glucuronosyltransferase 1A (UGT1A),nuclear factor erythroid-2-related factor 2 (Nrf2) and kelch-like ECH-associated protein 1 ( Keapl ) in the tumorigenesis of colonic carcinoma.Methods The expressions of UGT1 A,Nrf2 and Keapl were detected in normal colonic mucosa(24 cases),adenoma tissue (30 cases) and adenocarcinoma tissue (77 cases) by immunohistochemistry,and the relationship between their expressions and the clinical pathological characteristics was analyzed.Results The positive rates of UGT1 A in normal colonic mucosa,adenoma and adenocarcinoma tissue were 83.3％ ( 20/24),80.0％ ( 24/30 ) and 53.2％ ( 41/77 ),respectively.The positive rate of UGT1A in adenocarcinoma was lower than those in colonic mucosa and adenoma ( all P ＜0.05 ).On the contrary,the positive rates of Nrf2 in adenoma [70.0％ (21/30) ] and adenocarcinoma tissue [ 87.0％ (67/77) ] were higher than that in normal colonic tissue [ 41.7％ (10/24),all P =0.000 ].The positive rates of Keapl in normal colonic mucosa,adenoma and adenocarcinoma tissue were 54.2％ ( 13/24),70.0％ (21/30) and 61.0％ (47/77),respectively ( normal colonic tissue vs adenocarcinoma tissue,P =0.040 ; adenoma vs adenocarcinoma,P =0.002 ).There was no correlation between the expression of UGT1 A,Nrf2 and the clinicopathologic features of colon carcinoma,while the differences of Keapl positive rates in the various degrees of tumor differentiation [ moderately-well differentiated vs poorly differentiated:70.0％ (35/50) vs 44.4％ (12/27) ] and invasion [T1-T2 vs T3-T4:78.8％ (26/33) vs 47.7％ (21/44) ]were statistically significant (all P ＜ 0.05 ).Conclusion The decreased expression of UGT1A and the dysregulation of Nrf2/Keapl system may play a role in colonic tumorigenesis.

This study was aimed to explore the antitumor effect of volatile oil of X ihuang pill and its immune mecha-nism in order to screen the antitumor active site of Xihuang pill. Among 70 female Wistar rats, 10 rats were random-ly selected as the blank control group; and the other 60 rats was used in the establishment of walker 256 breast can-cer cell tumor-bearing rat models. The model rats were randomly divided into the negative control group (model group), high-dose volatile oil group, middle-dose volatile oil group, low-dose volatile oil group, high-dose Xihuang pill group, and lentinan group (positive control group), with 10 rats in each group. The intragastric administration was given twice a day for 14 days. Blood was taken from the abdominal aorta. Tumor tissues was removed and weighed to calculate the tumor inhibitory rate. ELISA method was used to detect the level of IL-2, IL-6, IL-10, IFN-γ and TGF-β in peripheral blood. The flow cytometry was used to detect the content of CD3+ T cell, CD4+ T cell, CD8+ T cell, and B7-1 cell (CD80). The results showed that the tumor inhibitory rates of volatile oil of high-dose group and middle-dose group were 28.4% and 24.1%, respectively. Compared with the model group, the average level of IL-2 and IFN-γ of volatile oil of high-dose group and middle-dose group and CD3+ T cell, CD8+ T cell, B7-1 cell con-tent were obviously increased (P< 0.05). It was concluded that volatile oil of Xihuang pill had certain antitumor ef-fect, which was one of the antitumor active sites of Xihuang pill. The volatile oil of Xihuang pill upregulates the lev-els of IL-2 and IFN-γ, as well as the contents of CD3+ T cell, CD8+ T cell, B7-1 cell in order to increase the im-mune clearance function of tumor-bearing rats.

Objective To evaluate the effect of dexmedetomidine on hippocampal neuronal apoptosis in endotoxemic rats.Methods Twenty-four pathogen-free male Sprague-Dawley rats,aged 6 weeks,weighing 200-250 g,were randomly divided into 3 groups(n=8 each)using a random number table:control group (group C),endotoxemia group (group E),and dexmedetomidine group (group D).Dexmedetomidine 50 μg/kg was injected intraperitoneally,and lipopolysaccharide 5 mg/kg was injected via the caudal vein 30 min later in group D.Normal saline 2 ml was injected intraperitoneally,and lipopolysaccharide 5 mg/kg was injected via the caudal vein 30 min later in group E.Normal saline 2 ml was injected intraperitoneally,and normal saline 2 ml was injected via the caudal vein 30 min later in group C.At 12 h after the model was successfully established,Morris water maze test was used to assess the cognitive function.The escape latency,swimming distance and frequency of crossing the original platform were recorded,and the swimming speed was calculated.The rats were then sacrificed,and hippocampi were harvest for determination of neuronal apoptosis (by TUNEL) and expression of glucose-regulated protein 78 (GRP78),CAAT/enhancer-binding protein homologous protein (CHOP) and caspase-12 (by immunohistochemistry).The apoptosis index (AI) was calculated.Results There was no significant difference in the swimming speed between the 3 groups (P＞0.05).Compared with group C,the escape latency and swimming distance were significantly prolonged,the frequency of crossing the original platform was significantly decreased,and the AI was significantly increased,and the expression of GRP78,CHOP,and caspase-12 was significantly up-regulated in E and D groups (P＜0.05).Compared with group E,the escape latency and swimming distance were significantly shortened,the frequency of crossing the original platform was significantly increased,and the AI was significantly decreased,and the expression of GRP78,CHOP,and caspase-12 was significantly down-regulated in group D (P ＜ 0.05).Conclusion Dexmedetomidine reduces cognitive dysfunction probably through reducing hippocampal neuronal apoptosis mediated by endoplasmic reticulum stress in endotoxemic rats.

ObjectiveTo discuss the application value of high frequency ultrasound (HFUS) characteristics, acoustic radiation force impulse (ARFI) imaging technology and contrast-enhanced ultrasound (CEUS) in the diagnosis of soft tissue hemangioma.MethodsTo retrospectively analyze the clinical data of 44 cases of soft tissue hemangioma that were treated in Capital Medical University Affiliate Beijing Chaoyang Hospital from August, 2013 to May, 2014, and to analyze the difference between the characteristics of HFUS, ARFI and CEUS in soft tissue hemangioma and normal surrounding tissue.ResultsHFUS shows the features of morphological diversity of sinus shape expansion tube structure, unclear boundary, irregular configuration, compressibility and partial strong echo in the phlebolith. Color Doppler ultrasound (CDFI), detects abundant interphase red-blue bloodstream signal slowly and consistently. The blood signal is strengthened after partial compression. CDFI shows more vein spectrum in the lesion. The discrepancy of comparison between VTQ and SWV value of soft tissue hemangioma and surrounding muscular tissue possesses statistical significance [(1.082±0.183) m/svs (1.414±0.331) m/s,P<0.01]. Ultrasound contrast can show the relationship between diseased region and surrounding tissue clearly, which is beneficial to the selection of operation method and prognosis.ConclusionThrough conventional two-dimensional ultrasound and high frequency CDFI, and further combining the acoustic pulse radiation ARFI technology and CEUS technology, the soft tissue hemangioma can’t only be more accurately diagnosed, but also provides more reliable diagnostic basis for clinic.