Cytokines ; metabolism ; Humans ; Leukocytes, Mononuclear ; immunology ; secretion ; Ubiquitin ; blood

We investigated the genetic diversity and evolution of the M gene of human influenza A viruses in Hangzhou (Zhejiang province, China) from 2009 to 2013, including subtypes of A(H1N1) pdm09 strains and seasonal A(H3N2) strains. Subtypes of analyzed viruses were identified by cell culture and real-time reverse transcription-polymerase chain reaction, followed by cloning, sequencing and phylogenetic analyses of the M gene. Assessment of 5675 throat swabs revealed a positive rate for the influenza virus of 20.46%, and 827 cases were diagnosed as. infections due to influenza A viruses. Seventy-six influenza-A strains were selected randomly from nine stages during six phases of a virus epidemic. Sequences of the M gene showed high homology among six epidemics with identities of amino-acid sequences of 98.98-100%. All strains contained the adamantine-resistant mutation S31N in its M2 protein. Two of the A(H1N1)pdm09 strains had double mutants of V27A/S31N or V271/S31N. One of the seasonal A(H3N2) viruses had another form of double-mutant R45H/S31N. Evolutionary rate of the M gene was much lower than that of the HA gene and NA gene. Compared with A(H3N2) strains, higher positive pressure on the M1 and M2 proteins of A(H1N1) pdm09 viruses was observed. Separate analyses of M1 and M2 proteins revealed very different selection pressures. Knowledge of the genetic diversity and evolution of the M gene of human influenza-A viruses will be valuable for the control and prevention of diseases.
Amino Acid Substitution ; China ; epidemiology ; Evolution, Molecular ; Genetic Variation ; Humans ; Influenza A Virus, H1N1 Subtype ; classification ; genetics ; isolation & purification ; Influenza A Virus, H3N2 Subtype ; classification ; genetics ; isolation & purification ; Influenza, Human ; epidemiology ; virology ; Phylogeny ; Selection, Genetic ; Viral Matrix Proteins ; genetics ; Viral Proteins ; chemistry ; genetics

Cochlear Implantation ; Cochlear Implants ; Guidelines as Topic

Pseudorabies virus (PRV) is a swine herpesvirus of the Alphaherpesvirinae subfamily and a pathogen of swine resulting in devastating disease and economic losses worldwide. Cre/loxP site-specific system has the character of site specific, time specific, tissue specific and high efficiency in recombination, which makes this system universal in vivo and in vitro recombination of bacteria, fungus, plants, insects and mammals. A recombinant PRV which contain a loxP site in TK locus by using Cre/LoxP recombinant system was construsted. A pair of primers were synthesized according to the pEGFP-C1 sequence published on GenBank, and were used to amplify the EGFP gene expression cassette with two loxP sites flanking each side. This target gene was cloned into pSKLR, the resulting transfer vector pSKLR-GFP-loxP was then cotransfected into 293T cells with PRV SH strain genomic DNA. The recombinant virus rPRV1 was selected and purified in TK-143 cells by choosing fluorescent expressing plaques. Cre expression vector pOG231 was cotransfected into 293T cells with rPRV1 genomic DNA. The second recombinant virus rPRV2 was obtained, which contains only one loxP site in TK locus. Sequencing results of rPRV2 TK gene indicated that 34bp loxP site was inserted into rPRV2 genome and there were 270bp deletion in TK gene. PCR amplifying different generations of rPRV2 TK gene showed that the mutant was stable when passages in RK-13 cells. TCID_ 50 assay indicated that rPRV2 grows well on RK-13 cells. The LD_ 50 test results on BALB/C mice suggested that the virulence of rPRV2 was reduced. As a conclusion, the report gene GFP expression cassette was removed successfully from rPRV1 genome and only one LoxP site was leaved in rPRV2 genome by using Cre/LoxP recombinant system.

OBJECTIVETo investigate regulation function of anodonta glucan HBP-A on chondrocytes through Wnt pathway in vitro.

METHODSRat chondrocytes were cultured and differentiated induced with IL-1beta (10 ng/ml) in vitro. Chondrocytes were divided into five groups:IL-13 group,IL-1beta + IWP-2 (5 microM,Wnt pathway inhibitor) group, IL-1beta + HBP-A (0.3 mg/ml) group and IL-1beta + IWP-2 + HBP-A group. Wnt-3a, beta-catenin (24 h,48 h,72 h) and MMP-13(72 h) genes expression were detected by Rt-PCR, while beta-catenin, MMP-13, Sox-9 and coll-II (48 h) protein expression were measured by Western-blot.

RESULTSAfter induction of IL-1beta, gene expression of Wnt-3a, beta-catenin and MMP-13 were increased,so were the protein expression of beta-catenin and MMP-13. In contrast,protein expression of Sox-9 and Coll-II were declined. Following addition of HBP-A, Wnt-3a, beta-catenin and MMP-13 were shown as induction of IL-1beta, but protein expression of Sox-9 and Coll-II were upgraded. Combining HBP-A with IWP-2 led to the lowest level in Wnt-3a, beta-catenin gene and beta-catenin protein expression and highest expression of Sox-9 protein.

CONCLUSIONHBP-A could not only delay the differentiation of chondrocytes through downgrading the signal expression of Wnt/beta-catenin,but also adjust the expression of Wnt-3a, beta-catenin and Sox-9 when combinated with the Wnt inhibitor.

Animals ; Anodonta ; chemistry ; Cell Differentiation ; drug effects ; Cells, Cultured ; Chondrocytes ; cytology ; drug effects ; metabolism ; Glucans ; pharmacology ; Interleukin-1beta ; metabolism ; Rats ; Wnt Signaling Pathway ; drug effects ; Wnt3A Protein ; genetics ; metabolism ; beta Catenin ; metabolism

This study was aimed to study the potential effects of alloreactive NK cells (allo-NKs) in therapy of relapsed lung cancer after haploidentical hematopoietic stem cell transplantation using donor lymphocyte infusion (DLI). The F1 donors derived-NK cells were purified with MACS magnetic separation system, in which the proportion of the alloreactive Ly49A(+) cells was detected by flowcytometry and alloreactivity was measured by LDH method. The relapse model of lung cancer after haploidentical-HSCT was established. The distribution kinetic of infused donor lymphocytes in vivo was analyzed. The inhibition of relapse tumor, infiltration of lymphocytes in situ and fluctuation of 22 kinds of cytokines in serum after DLI were compared among different groups. The results showed that the infused donor cells of allo-NK group were accumulated mostly in lung, spleen and kidney for more than 48 hours with considerable higher levels according to the distribution kinetic curve. The sizes of relapse tumors between chemotherapy + PBS group and chemotherapy + DLI group showed no difference. However, the relapsed tumors in allo-NK + DLI group were significantly smaller than that in chemotherapy + DLI group or allo-NK + PBS group, in which increased infiltration of lymphocytes were defined in situ. The levels of cytokines such as MCP-1, IL-17, IL-12 and MCP-5 in serum of allo-NK + DLI group ascended compared with control group, though the level of IL-10 declined simultaneously. It is concluded that allo-NKs prolong the survival time of infused donor lymphocytes in vivo, promote the secretion of inflammatory cytokines and Th1-type of cytokines, and further improve the antitumor effects of DLI against relapse after transplantation.
Animals ; Cytokines ; blood ; Hematopoietic Stem Cell Transplantation ; methods ; Killer Cells, Natural ; cytology ; Lung Neoplasms ; therapy ; Lymphocyte Transfusion ; methods ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Neoplasm Recurrence, Local ; therapy ; Transplantation Conditioning ; methods

Objective To elucidate the association of genetic polymorphisms of key molecules in JAK/STAT signaling pathway with susceptibility of hepatocellular carcinoma (HCC).Methods A total of 367 HCC patients and 367 healthy controls were recruited in this sex- and age-matched case-control study.Genetic polymorphisms of IL-6 (rs1800796,-572C＞G),STAT3 (rs744166,+ 26312T＞C; rs3816769,+ 42240T＞C; rs6503695,+ 40980T＞C),EGFR (rs11543848,+ 142530A＞G),and mTOR (rs7211818,+ 170278A＞G; rs9674559,+ 196983A＞G; rs11653499,+65678G＞A) were genotyped using a mass spectrometry method.Odds ratio (OR) and 95％ confidence interval (CI) were calculated.Results Genotype frequency of the 8 polymorphisms of IL-6,STAT3,EGFR,and mTOR were not significantly different between the patients with HCC and the controls.When stratified by sex,the female subjects who carried STAT3 +26312CC,+ 42240CC,or + 40980CC had a decreased risk of HCC when compared to those who carried TT allele (OR=0.192,95％CI:0.047-0.784; OR=0.180,95％CI:0.045-0.725;OR=0.198,95％ CI:0.049-0.806,respectively).When compared with AA genotype on the site of EGFR + 142530,the (AG+ GG) genotype reduced the risk of HCC in women (OR=0.422,95％CI:0.179-0.994).Conclusion The polymorphisms of IL-6 (rs1800796) and mTOR (rs7211818,rs9674559,and rs11653499) were not associated with the HCC susceptibility.Those carrying CC allele in three loci (rs744166,rs3816769,and rs6503695) of STAT3 and (AG + GG) in rs11543848 of EGFR had a decreased risk of HCC in women.However,these results need to be validated using larger sample size.

·Long non-coding RNAs (lncRNAs) refer to a kind of non-coding RNA which is longer than 200 nucleotides, with the characteristic of its numerous, diversity of types and modes of action. The biological functions of lncRNA involved in genomic imprinting, chromatin remodeling, translational control of mRNA, cell cycle and cell differentiation control, immune surveillance, constituting the skeleton of nuclear sub structure, etc. LncRNA plays an important role in individual development and human diseases. This paper mainly reviewed those lncRNAs that have been published, and closely related to eye development and diseases.

AIM To explore the mechanism of Yiqi Wenyang Huwei Decoction on airway inflammation improvement of rats with bronchial asthma based on IL-25/NF-κB signaling pathway.METHODS 60 rats were randomly divided into the control group,the model group,the dexamethasone group(0.2 mg/mL),the low-dose,medium-dose and high-dose Yiqi Wenyang Huwei Decoction groups(1,2,4 g/mL),with 10 rats in each group.Intraperitoneal injection of ovalbumin(OVA)and aluminum hydroxide suspension was applied to establish the rat asthma model,followed by 2-week corresponding dosing of the drugs.The rats of each group had their daily diet,mental status,hair growth and respiration observed;their differential count of inflammatory cells in bronchoalveolar lavage fluid(BALF)detected by automatic hematology analyzer;their pathological changes of lung tissue observed by HE staining;their pulmonary IL-25 protein expression detected by immunohistochemistry(IHC);their levels of IL-4,IL-5 and IL-13 in BALF measured by ELISA;their pulmonary expression of IL-25 and TRAF6 mRNA detected by RT-qPCR;and their pulmonary protein expressions of IL-25,TRAF6,IκBα,p-IκBα,NF-κB p65 and p-NF-κB p65 detected by Western blot.RESULTS Compared with the control group,the model group displayed severe damage of the lung tissue and infiltration of a large number of inflammatory cells;increased number of inflammatory cells and levels of IL-4,IL-5 and IL-13 in BALF(P<0.01);increased mRNA expressions of IL-25 and TRAF6,and pulmonary protein expressions of IL-25,TRAF6,p-IκBα/IκBα and p-NF-κB p65/NF-κB p65(P<0.01).Compared with the model group,all of the Yiqi Wenyang Huwei Decoction groups shared improved pulmonary infiltration of inflammatory cells;decreased number of inflammatory cells and levels of IL-4,IL-5 and IL-13 in BALF(P<0.05,P<0.01);and decreased mRNA expressions of IL-25 and TRAF6,and pulmonary protein expressions of IL-25,TRAF6,p-IκBα/IκBα and p-NF-κB p65/NF-κB p65(P<0.01).CONCLUSION Yiqi Wenyang Huwei Decoction can inhibit the airway inflammation in the rat model of bronchial asthma,which may be related to the inhibited activation of IL-25/NF-κB signaling pathway and the reduced expression of inflammatory factors.

OBJECTIVETo explore the diagnostic value of whole-organ magnetic resonance imaging score (WORMS) in knee osteoarthritis (KOA).

METHODSFrom November 2009 to January 2011,70 patients with KOA combined with knee effusion among outpatient and inpatient were analyzed retrospectively. Among the patients, 12 patients were male, 58 patients were female,ranging in age from 46 to 75 years,with a mean age of (59.66 +/- 9.93) years. The clinical symptoms were evaluated by WOMAC, the imaging of KOA was assessed by K-L score and WORMS, and COMP and CTX- II were measured respectively by ELISA. The correlation analyses and multiple linear regression analysis were studied to determine associations among biomarkers, clinical variables and radiographic findings of knee joints.

RESULTSThe average scores of WOMAC and WORMS were (57.50 +/- 8.20) and (64.54 +/- 16.45) respectively. The median of CTX- II nd COMP were 2.42 ng/ml and 4.56 ng/ml respectively. Grouped by less than the lowest quartile and more than the highest quartile of WORMS, COMP was significantly different (Z=2.04, P=0.039), but there was no significant difference in CTX-II (Z=0.79, P=0.427). WORMS were positively correlated with WOMAC and K-L score (r=0.777, P<0.01; r=0.716, P<0.01; respectively); WOMAC was also positively correlated with K-L score (r=0.692, P<0.01). WORMS's cartilage, osteophytes and synovitis were positively correlated with WOMAC, K-L score and COMP respectively (r=0.771, P<0.01; r=0.509, P<0.01; r=0.917, P<0.01). It was determined by stepwise regression that the KOA was mainly affected by WORMS, K-L score (P=0.015, P=0.025 respectively) when WOMAC as a dependent variable, age, gender, K-L score, WORMS, COMP and CTX- II as independent variables (F=20.327, P<0.01).

CONCLUSIONWORMS has a better reference value for diagnosis of KOA. The expression of COMP is high in the synovial fluid when WORMS at the high point. The clinical symptoms of knee osteoarthritis are mainly affected by WORMS and K-L score.

Aged ; Cartilage Oligomeric Matrix Protein ; Collagen Type I ; analysis ; Extracellular Matrix Proteins ; analysis ; Female ; Glycoproteins ; analysis ; Humans ; Magnetic Resonance Imaging ; methods ; Male ; Matrilin Proteins ; Middle Aged ; Osteoarthritis, Knee ; diagnosis ; metabolism ; physiopathology ; Peptides ; analysis