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Antimicrobial susceptibility testing (AST) plays a vital role in anti-To investigate the characteristics of heteroresistance inA total of 704 gastric biopsies were selected for 23S rRNA and gyrA gene sequencing, 470For the 699 biopsies that were positive for 23S rRNA gene, 98 (14.0%) showed a heteroresistance genotype, and a wild type (WT) combined with A2143G (86.7%) genotype was found in most samples. For the 694 biopsies that were positive for gyrA gene, 99 (14.3%) showed a heteroresistance genotype, and a WT combined with 87K (26.3%) or WT combined with 91N (23.2%) genotype was predominant. According to the E-test results, the resistance rates of heteroresistance genotype samples for clarithromycin and levofloxacin were 36.2% and 68.1%, respectively. When dividing the heteroresistance samples into different groups according to the sequencing profile peaks of the mutation position, the resistance rates were higher along with mutation peaks at the mutation position. In addition, patients infected with mutated or heteroresistant strains showed lower peptic ulcer detection rates than those infected with the WT strain (p0.05).Heteroresistance genotypes for clarithromycin and levofloxacin were not rare in

Air pollution is a world public health problem. Particulate matter (PM), a mix of solid and liquid particles in the air, becomes an increasing concern in the social and economic development of China. For decades, epidemiological studies have confirmed the association between fine particle pollutants and respiratory diseases. It has been reported in different populations that increased fine particulate matter (PM2.5) concentrations cause elevated susceptibility to respiratory diseases, including acute respiratory distress, asthma, chronic obstructive pulmonary disease, and lung cancer. This review will discuss the pathophysiology of PM2.5 in respiratory diseases, which are helpful for the prevention of air pollution and treatment of respiratory tract inflammatory diseases. Keywords: Air pollution, Particulate matter, Lung diseases, Chronic obstructive pulmonary disease

Air pollution is a global health threat and causes millions of human deaths annually. The late onset of respiratory diseases in children and adults due to prenatal or perinatal exposure to air pollutants is emerging as a critical concern in human health. Pregnancy and fetal development stages are highly susceptible to environmental exposure and tend to develop a long-term impact in later life. In this review, we briefly glance at the direct impact of outdoor and indoor air pollutants on lung diseases and pregnancy disorders. We further focus on lung complications in later life with early exposure to air pollutants. Epidemiological evidence is provided to show the association of prenatal or perinatal exposure to air pollutants with various adverse birth outcomes, such as preterm birth, lower birth weight, and lung developmental defects, which further associate with respiratory diseases and reduced lung function in children and adults. Mechanistic evidence is also discussed to support that air pollutants impact various cellular and molecular targets at early life, which link to the pathogenesis and altered immune responses related to abnormal respiratory functions and lung diseases in later life. Keywords: Air pollutants, Polycyclic aromatic hydrocarbon, Particulate matter, Early disease origin, Respiratory diseases

The health impact of airborne particulate matter (PM) has long been a concern to clinicians, biologists, and the general public. With many epidemiological studies confirming the association of PM with allergic respiratory diseases, an increasing number of follow-up empirical studies are being conducted to investigate the mechanisms underlying the toxic effects of PM on asthma and allergic rhinitis. In this review, we have briefly introduced the characteristics of PM and discussed its effects on public health. Subsequently, we have focused on recent studies to elucidate the association between PM and the allergic symptoms of human respiratory diseases. Specifically, we have discussed the mechanism of action of PM in allergic respiratory diseases according to different subtypes: coarse PM (PM2.5-10), fine PM (PM2.5), and ultrafine PM. Keywords: Particulate matter, Allergic respiratory diseases, Coarse PM, Fine PM, Ultrafine PM

A dicyanoisophorone-based probe with two-photon absorption and NIR emission was developed for the in vivo fluorescence imaging of amyloid-β plaques, which exhibited high selectivity toward Aβ aggregates over other intracellular proteins. The detection limit was calculated to be as low as 109 nM. In vivo imaging studies indicated that the probe could penetrate the blood–brain barrier and label Aβ plaques in the living transgenic mice, and its specific binding to cerebral Aβ plaques was further confirmed by one- and two-photon ex vivo fluorescence imaging. All these results featured its promising application prospects for amyloid-β sensing in basic research and biomedical research.

A highly sensitive colorimetric and red-emitting fluorescent dual probe was synthesized and applied to imaging palladium species based on the Tsuji–Trost reaction. Notably, it displayed specific and turn-on fluorescent response toward Pd0 with a vivid color change from yellow to purple in aqueous media. The detection limit was calculated to be as low as 0.78 nM. Furthermore, the probe was successfully applied for fluorescence imaging of palladium in living cells. All these properties made this probe a promising useful tool not only in environmental detection but also more widely in bioanalytical fields.

A tailor-made colorimetric and NIR fluorescent probe for Pd0 was developed by introducing allyl chloroformate into the methylene blue (MB) fluorophore. The probe exhibited vivid color change and significant fluorescence enhancement towards Pd0, accompanied with excellent selectivity and sensitivity. No absorption and fluorescence response could be observed towards other metal ions. The fluorescence intensity of this probe showed a linear response to Pd0 in the concentration range of 0–2 μM with a detection limit of 5.7 nM. Moreover, living cell imaging results indicated that this probe holds promising application prospects for detecting intracellular Pd0 species based on its low cytotoxicity and specific turn-on NIR emission.

Carbon monoxide (CO) has captivated great attention in part due to the discovery of the therapeutic and cell-signalling role that CO plays in biological systems. The ever-increasing interest in CO has resulted in the development of rapid and facile approaches for the sensitive and selective detection of CO, which still remains challenging. Herein, the first example of a colorimetric and near-infrared fluorescent dual probe for carbon monoxide (CO) has been developed by assembling an allyl chloroformate moiety with a naphthofluorescein fluorophore, which enables the label-free and visual detection of CO. The living cell imaging results indicate that the probe shows great potential in sensing intracellular CO.

Midkine antisense oligonucleotide (MK-ASODN) nanoliposomes have previously been shown to have inhibitory activity against hepatocellular carcinoma growth. Herein we report the 4-week sub-chronic toxicity of MK-ASODN nanoliposomes in SD rats. The adverse effects included loss of body weight gain and food consumption, peri-rhinal bleeding, piloerection, peri-anal filth, and kidney, liver, spleen, thymus, lung, and injection site lesions at high doses. Macroscopic changes were observed in the kidneys of the high-dose group, accompanied by a variation in urine protein and white blood cells, blood urea nitrogen, and serum creatinine. The increased spleen and liver coefficient, and the variation in circulating white blood cells, lymphocytes, and eosinophils in the high-dose group demonstrated that inflammation was caused by MK-ASODN nanoliposomes and was consistent with the macroscopic changes in the spleen and liver. The main necropsy findings of the animals that died were macroscopic changes in the lung. No severe toxic effects or mortalities occurred in the low- and medium-dose groups. However, a No Adverse Effect Level (NOAEL) was not identified since there were changes in organs deemed to be adverse at all dose levels. Thus, the maximum tolerated dose of MK-ASODN nanoliposomes for rats was considered to be 6 mg/kg/day.

Air pollution is a world public health problem. Particulate matter (PM), a mix of solid and liquid particles in the air, becomes an increasing concern in the social and economic development of China. For decades, epidemiological studies have confirmed the association between fine particle pollutants and respiratory diseases. It has been reported in different populations that increased Fine particulate matter (PM2.5) concentrations cause elevated susceptibility to respiratory diseases, including acute respiratory distress, asthma, chronic obstructive pulmonary disease, and lung cancer. This review will discuss the pathophysiology of PM2.5 in respiratory diseases, which are helpful for the prevention of air pollution and treatment of respiratory tract inflammatory diseases.

The midkine antisense oligonucleotide (MK-ASODN, 5′-CCC CGG GCC GCC CTT CTT CA-3′) nanoliposomes have been identified to suppress hepatocellular carcinoma (HCC) growth effectively, and have a great potential to be an effective target drug for HCC. In this study, a facile and reproducible method for large-scale preparation of MK-ASODN nanoliposomes followed by lyophilization has been developed successfully. Meanwhile, the MK-ASODN nanoliposomes characteristics, storage stability and their antitumor efficiency were studied. The mean particle size of MK-ASODN nanoliposomes were 229.43 ± 15.11 nm, and the zeta potential were 29.7 ± 1.1 mV. High entrapment efficiency values were achieved around 90%. Transmission electron microscopy images revealed spherical shaped nanoliposomes. Nanoliposomes allowed sustained MK-ASODN release for as long as 14 days. During 180 days of storage, freeze-dried nanoliposomes showed no significant change in the mean size, zeta potential, entrapment efficiency and drug release ratio. Regarding their antitumor efficiency, the in vitro proliferation of human liver cancer cells were significantly inhibited by the MK-ASODN nanoliposomes. Furthermore, the MK-ASOND nanoliposomes also significantly inhibited the growth of HCC in the mouse model. In summary, the results confirmed that this large-scale preparation of MK-ASOND nanoliposomes was facile and reproducible, and potentially, could speed up the application process of our MK-ASOND nanoliposomes for HCC therapy.

Thymic stromal lymphopoietin (TSLP), an interleukin 7-like cytokine, can trigger dendritic cell (DC)-mediated T-helper type 2 (Th2) inflammatory responses. Recent evidence demonstrates that cytokines TSLP and OX40 (CD134)/OX40 ligand seem to be important players in the maintenance of Th2 memory pool in the pathogenesis of asthma. Accumulating data reveal that the pathogenic T cells involved in asthma are likely to be inflammatory Th2 cells. TSLP is involved in the development of asthma through crosstalk with nuclear factor NF-ĸB. Progression of skin fibrosis in atopic dermatitis occurs via TSLP/TSLP receptor. TSLP-mediated dermal inflammation aggravates experimental allergic asthma. Also, TSLP polymorphisms are associated with susceptibility to asthma, atopic dermatitis, and eczema herpeticum. These findings suggest a master switch of TSLP in the initiation of allergic and adaptive inflammation through innate pathways at the epithelial cell-DC interface. The TSLP pathway is therefore a promising target for immunotherapy of allergic diseases.

Background and aims Epithelial barrier dysfunction plays a critical role in the initiation of a number of immune diseases; the causative factors are not fully understood. The present study aimed to elucidate the mechanism by which the eosinophil-derived interferon (IFN)-lambda induced the gut epithelial barrier dysfunction. Methods The duodenal biopsies were obtained from patients with or without food allergies. The eosinophils and IFNλ expression were observed by immune staining. Intestinal epithelial cell line, T84 cells, and a mouse model were employed to observe the effect of IFNλ on the epithelial barrier function and the initiation of skewed T helper (Th)2 polarization in the mouse intestine. Results IFNλ expression was observed in over 80% human eosinophils of the subjects with or without food allergies. Exposure to microbial products, lipopolysaccharide or peptidoglycan, could induce eosinophils to release IFNλ. Exposure to IFNλ could induce intestinal epithelial barrier dysfunction via inducing the epithelial cell apoptosis. Concurrent exposure to microbial products and food antigens could induce aberrant antigen specific Th2 polarization and Th2 pattern inflammation in the intestine. Conclusions Eosinophils express IFNλ that can induce intestinal epithelial barrier dysfunction and promotes the initiation of the aberrant Th2 polarization in the intestine.

Our previous studies have showed that ginsenoside (GS)-Rd, a mono-compound isolated from traditional Chinese herb panax ginseng, has the neuroprotective effects following ischemic stroke. However, the underlying mechanisms are still largely unknown. Our latest study showed that GS-Rd could block calcium influx in cultured cortical neurons after excitotoxic injury, indicating that GS-Rd may act on cation channels. To explore this possibility, in this study, we used a rat middle cerebral artery occlusion (MCAO) model to examine the effects of GS-Rd on the expression of non-selective cation channels, including transient receptor potential melastatin (TRPM) and acid sensing ion channels (ASIC), and cation channels, including N-methyl-D-aspartate (NMDA) receptors, which all play essential roles in ischemic stroke. Our results showed that both TRPM and ASIC channels were expressed in the brain. At 24 h following MCAO insult, mRNA and protein expression levels of TRPM7, ASIC1a and ASIC2a were significantly increased. Pretreatment of 10 mg/kg GS-Rd attenuated MCAO-induced expression of TRPM7 and ASIC1a but promoted that of ASIC2a. In contrast, GS-Rd had no significant effects on the expression of NMDA receptors. Thus, our results suggest that GS-Rd neuroprotection following cerebral ischemia may be at least due to its effects on the expression of TRPM7, ASIC1a and ASIC2a.

Sialic acid-binding immunoglobulin-like lectin-8 (Siglec-8) promotes the apoptosis of eosinophils and inhibits FcvarepsilonRI-dependent mediator release from mast cells. We investigated the genetic association between sequence variants in Siglec-8 and diagnosis of asthma, total levels of serum IgE (tIgE), and diagnosis of eosinophilic esophagitis (EE) in diverse populations. The effect of sequence variants on Siglec-8 glycan ligand-binding activity was also examined. Significant association with asthma was observed for SNP rs36498 (odds ratios (OR), 0.69, P=8.8 x 10(-5)) among African Americans and for SNP rs10409962 (Ser/Pro) in the Japanese population (OR, 0.69, P=0.019). Supporting this finding, we observed association between SNP rs36498 and current asthma among Brazilian families (P=0.013). Significant association with tIgE was observed for SNP rs6509541 among African Americans (P=0.016), and replicated among the Brazilian families (P=0.02). In contrast, no association was observed with EE in Caucasians. By using a synthetic polymer decorated with 6'-sulfo-sLe(x), a known Siglec-8 glycan ligand, we did not find any differences between the ligand-binding activity of HEK293 cells stably transfected with the rs10409962 risk allele or the WT allele. However, our association results suggest that the Siglec8 gene may be a susceptibility locus for asthma.

A novel recombinant exendin-4 human serum albumin fusion protein (rEx-4/HSA) expressed in Pichia pastoris was prepared and characterized. Ex-4 is a 39-amino acid peptide isolated from the salivary gland of the lizard Heloderma suspectum and is thought to be a novel therapeutic agent for type 2 diabetes. But to gain a continued effect, the peptide has to be injected twice a day owing to its short plasma half-life (t1/2 = 2.4 h). To extend the half-life of Ex-4 molecule in vivo, we designed a genetically engineered Ex-4/HSA fusion protein. Between Ex-4 and HSA, a peptide linker GGGGS was inserted and the fusion protein was expressed in methylotrophic yeast P. pastoris with native HSA secretion signal sequence. The recombinant protein was secreted correctly and was obtained with high purity (typically > 98%) by a three-step purification procedure. cAMP assay demonstrated that the fusion protein had a bioactivity similar to Ex-4 for interaction with GLP-1 receptors in vitro. Results from oral glucose tolerance test indicated that rEx-4/HSA could effectively improve glucose tolerance in diabetic db/db mice. Pharmacokinetics studies in cynomologus monkeys also showed that rEx-4/HSA had a much longer plasma half-life. Therefore, rEx-4/HSA fusion protein could potentially be used as a new recombinant biodrug for type 2 diabetes therapy. Copyright © 2007 European Peptide Society and John Wiley & Sons, Ltd.

A long-lasting recombinant human serum albumin-interferon-alpha2b fusion protein (rHSA/IFNalpha2b) was prepared and its structure and biological activities were studied. rHSA/IFNalpha2b was expressed in methylotrophic yeast Pichia pastoris with HSA's natural signal peptide and purified by dye affinity chromatography, hydrophobic interaction chromatography, ion exchange chromatography and Sephadex G25. Purity of the prepared rHSA/IFNalpha2b was greater than 97% analyzed by non-reduced SDS-PAGE and RP-HPLC. Structure and biological activities of the prepared rHSA/IFNalpha2b were characterized by physical, chemical and biological methods. Its pI was 5.3 and showed a single band on IEF gel. Molecular weight determined by MALDI-TOF was 86004.3+/-29.2. Amino-terminal and carboxyl-terminal amino acid sequences were identical to predicted sequence. Its specific activity in vitro was 6.3+/-0.8x10(5) IU/mg fusion protein, retaining about 1.4% of that of unmodified rIFNalpha on a molar basis. After administered in monkeys, significant increases of 2',5'-oligoadenylate synthetase activity relative to IFN-alpha were maintained for 14 days in serum and the rHSA/IFNalpha2b showed more potent biological activity than IFN-alpha on a molar basis. Therefore, markedly improved in vivo biological activity of rHSA/IFNalpha2b could exhibit more potent antiviral activity than IFNalpha2b in future clinical trials.

Herein, MAPK/JNK signalling is proposed as a potential autophagy regulation pathway for the transcription-dependent or independent role., Autophagy refers to a lysosomal degradative pathway or a process of self-cannibalization. This pathway maintains nutrients levels for vital cellular functions during periods of starvation and it provides cells with survival advantages under various stress situations. However, the mechanisms responsible for the induction and regulation of autophagy are poorly understood. The c-Jun NH2-terminal kinase (JNK) signal transduction pathway functions to induce defence mechanisms that protect organisms against acute oxidative and xenobiotic insults. This pathway has also been repeatedly linked to the molecular events involved in autophagy regulation. The present review will focus on recent advances in understanding of the relationship between mitogen-activated protein kinase (MAPK)/JNK signalling and autophagic cell death.

This study aims to investigate the effect of Golgi Protein 73 (GP73) on autophagy in human hepatoma line cells HepG2. We investigated the functional effects of GP73 on autophagy in hepatoma cell line HepG2 using immunofluoscence staining, Western blotting and real-time PCR. Our data showed that specific small interference RNA (siRNA) notably induced formation of autophagic vacuoles. In addition, upregulation of GP73 significantly inhibited formation of starvation-induced LC3-positive structures. We provide the first experimental evidence to show that GP73 may play an important role in the inhibitory regulation of autophagy. Therefore, our data suggest a new molecular mechanism for GP73-related hepatoma progression.

To evaluate the effectiveness and safety of interventional treatment in the removal of endobronchial hamartoma by flexible bronchoscopy.A retrospective analysis was conducted in 8 inpatients with histologically confirmed endobronchial hamartoma , diagnosed between May 2009 to January 2012 in the First Affiliated Hospital of Nanjing Medical University. The clinical, radiological and bronchoscopic features of hamartoma, and the clinical outcomes after bronchoscopic intervention were described. The endoscopic interventional treatments included resection by electrosurgical snare, electrocautery, argon plasma coagulation (APC) and cryotherapy. Thoracic computed tomography(CT)and bronchoscopy were used to evaluate the airway stenosis during follow-up.The 8 patients, 7 males and 1 female, aged (62 ± 8) years, underwent 13 times of interventional treatment for endobronchial hamartoma. Four patients were cured after receiving a single endoscopic treatment, while 3 patients had recurrence after initial interventional treatment but were cured after the second treatment. Three times of interventional treatment was carried out in 1 patient who had two relapses but later became stable with a 40% stenosis of the airway lumen. The rates of cure and effectiveness were 87.5% and 100%, respectively. Following interventional treatment, pneumothorax occurred in 1 patient who was cured after oxygen therapy. There were no serious complications such as massive haemorrhage, airway perforation, airway ignition and suffocation.Interventional treatments through flexible bronchoscopy appear to be safe and effective for removing endobronchial hamartoma.

A novel multiplex real-time RT-PCR kit was developed to detect EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control to avoids false negatives, which used for hand, foot and mouth disease in the clinical diagnosis and epidemiological surveillance.Design specific primers and probes of EV71, CA16, other intestinal virus and internal amplification control, improve the extraction method of virus nucleic acid. Optimization the detection system of real-time quantitative PCR. Research the products of the accuracy, stability, precision, amplification efficiency and detection of linear range.The primers and probes had high spicificity. The Viral RNA extraction effect of this Kit is as same as that of QIAamp Viral RNA mini Kit (QIAGEN company), but less reagent cost. The optimal concentrations of primers and probes are 0.2 micromol/L for all the upstream and downstream primers, 0.06 micromol/L for probes of other human enteroviruse, 0.08 micromol/L for probes of EV71 and CA16 respectively. The kit has good stability, accuracy and precision. The amplification efficiencies of EV71, CoxA16 and other human enteroviruses are 106% ,101% and 105% and the detection of linear range is from 10(9) copies/microl-10(2) copies/microl.The novel multiplex real-time RT-PCR kit for detecting EV71, CoxA16 and other human enteroviruses simultaneously with an internal amplification control has good stability, accuracy, precision and amplification efficiencies. So it has great value in clinical application.

The aim of our study was to investigate the chronic obstructive pulmonary disease (COPD) assessment test (CAT), serum copeptin, procalcitonin and C-reactive protein (CRP) levels as potential predictive factors for recurrence of acute exacerbation and all-cause mortality in 6 months of COPD inpatients. One hundred fifty-nine patients who met the inclusion criteria were enrolled and followed up for 6 months. The CAT scores, serum copeptin, procalcitonin and CRP levels were measured on admission and 14 days and 3 months later in all patients. The primary endpoint was recurrence of acute exacerbation in 6 months. The secondary endpoint was all-cause mortality after 6 months. The CAT scores, serum copeptin, procalcitonin and CRP levels were significantly elevated on admission and stabilized at 14 days (P < 0.01). In a univariate logistic regression analysis, CAT scores (odds ratio [OR] = 1.10), forced expiratory volume in 1 second % (OR = 1.01), serum copeptin (OR = 1.32) and CRP levels (OR = 1.01) were significantly related to recurrence of acute exacerbation in 6 months (P < 0.05). In a multivariate logistic regression model, increasing CAT scores (OR = 1.10) and serum copeptin levels (OR = 1.29) were still associated with an increased odds of exacerbation (P < 0.05). In a univariate logistic regression analysis, increasing CAT scores (OR = 1.19), forced expiratory volume in 1 second % (OR = 1.05), serum copeptin levels (OR = 1.44) and hospitalization in the previous years (OR = 1.24) were significant determinants of death over a follow-up period of 6 months (P < 0.05). But only serum copeptin (OR = 1.53) and CAT scores (OR = 1.37) were associated with mortality in multivariate logistic regression analysis. Hence, high CAT scores and serum copeptin levels link with recurrence of acute exacerbation and all-cause mortality during 6 months in patients with acute exacerbation of COPD.

Golgi glycoprotein 73(GP73) is a transmembrane glycoprotein residing in the cis-Golgi complex, which is strongly expressed in hepatocellular carcinoma (HCC) and secreted into the blood. It has been regarded as a promising serum tumor marker for the detection of HCC with higher sensitivity and specificity than AFP. GP73 is also significantly elevated in kidney cancer, prostate cancer, lung adenocarcinoma, esophageal cancer and seminomas; therefore, it would be helpful for the diagnosis of these diseases. However, the function of GP73 and the regulatory mechanism for its expression are unclear. In this article, the physical-chemical properties, the regulation of its expression, the relation with various cancers and the clinical applications of GP73 are reviewed.

To explore the effect of asthmatic and healthy serum on differentiation and function of monocyte-derived dendritic cells (MDDC) in a transendothelial trafficking model.The sera and peripheral blood mononuclear cells (PBMC) were separated from 12 asthmatic patients and 12 healthy volunteers, and monocytes were selected from PBMC using magnetic beads. The trypsin-digested human umbilical vein endothelial cells (HUVEC) at passage 2 from 5 healthy lying-in women were used to construct the transendothelial trafficking model under asthmatic or healthy serum, wherein MDDC were identified by silver nitrate staining and scanning electron microscopy. Nuclear factor κB (NF-κB) activity was determined by electrophoretic mobility shift assay. Flow cytometry, ELISA and mixed leukocyte reaction were relevantly utilized to detect the phenotype, cytokine and T cell proliferation.(1) Monocytes traversed through HUVEC monolayer after 2 h, and reverse-transmigrated to develop into DC 48 h later. (2) The healthy serum stimulated monocytes into immature MDDC with lower CD(14) [(20 ± 5)%] (F = 49.01, P0.05), and higher HLA-DR, CD(80), CD(86) and CD(83) [(43 ± 4)%, (17.9 ± 3.5)%, (43 ± 11)% and (6.7 ± 1.8)%, respectively] (F = 10.35 - 40.17, all P0.05) than monocytes did before transmigration at 0 h [CD(14) (81 ± 6)%, HLA-DR (24 ± 5)%, CD(80) (2.8 ± 2.0)%, CD(86) (14 ± 4)% and CD(83) (0.9 ± 0.8)%, respectively]. (3) The asthmatic serum stimulated monocytes into mature MDDC, characteristic of dendrites, with similar HLA-DR and CD(86) [(55 ± 6)% and (59 ± 12)%] (F = 15.29 and 35.97, all P0.05), higher CD(80) and CD(83) [(49.7 ± 10.2)% and (30.2 ± 6.8)%] (F = 4.01 and 20.68, all P0.05), accompanied by increased levels of NF-κB activity, IL-12 p70 and T cell proliferation [(100 ± 11)%, (568 ± 43) ng/L and (2033 ± 198) cpm, respectively] (F = 49.23 - 350.84, all P0.05) relative to the healthy serum-stimulated immature MDDC [(12 ± 3)%, (220 ± 35) ng/L and (952 ± 64) cpm, respectively].The asthmatic serum induces mature MDDC in association with NF-κB overactivation in the transendothelial trafficking model, which provides a promising experimental platform for both investigation of immunological mechanisms in asthma and screening of novel anti-asthma drugs in vitro.

Background & Aims Oral tolerance is an important component of gastrointestinal homeostasis, but mechanisms of its development are not fully understood. Loss of oral tolerance occurs during food allergen–related inflammation in the gastrointestinal tract. Interferon (IFN)-λ regulates immunity, but its role in oral tolerance is not clear. We investigated the role and the mechanism of IFN-λ in the development of oral tolerance and its effect on antigen-induced, T-helper (Th)-2 cell–mediated inflammation in the intestine. Methods Expression of IFN-λ and its receptor were analyzed by immunohistochemical, flow cytometric, or immunoblot analyses. Tolerogenic dendritic cells (DCs) and regulatory T cells were examined in vitro and in vivo. A mouse model of antigen-induced, Th2 cell–mediated intestinal inflammation was used to examine the role of IFN-λ and T cells in oral tolerance in the intestine. Results CD3 + cells expressed the IFN-λ receptor, which was up-regulated following antigen-specific or nonspecific activation. Interaction between IFN-λ and its receptor induced apoptosis of T cells and their subsequent phagocytosis by DCs. This led to the generation of tolerogenic DCs and T regulatory cells in vitro and in vivo. Passive transfer of IFN-λ–primed CD3 + cells inhibited Th2 cell–mediated inflammation in the intestine. Conclusions IFN-λ is involved in development and maintenance of oral tolerance in the intestines of mice; it might be used to suppress antigen-specific Th2 cell–mediated inflammation in patients.

To investigate the inhibitory effects and mechanism of apigenin (APG) on dominant response of Th2 cells in asthma model of mice.Thirty-two 6-week-old healthy BALB/c mice, SPF grade, were randomly divided into four groups equally, the normal control group (A), the asthma model group (B), and the two APG groups (C and D) consisted of asthma model mice treated respectively with high-dose (20 mg/kg per day) and low-dose (2 mg/kg per day) APG given by dissolving in 1% dimethyl sulphoxide via intraperitoneal injection. The murine asthma model was established by ovalbumin (OVA) sensitization and provocation. Twenty-four hours after the last airway provocation, acetylcholine (Ach) was administered via caudalis vein for measuring airway resistance by pulmonary function detector; levels of IL- 4 and IL-13 in bronchoalveolar lavage fluid (BALF) and total IgE in serum were determined by enzyme-linked immunosorbent assay (ELISA); total and differential cell counts in BALF were measured by light microscopy; the airway inflammatory infiltration was detected by haematoxylin and eosin (HE) staining; and the signal transducer and activator of transcription 3 (GATA-3) in the lung tissue was determined by Western blot analysis.As compared with Group A, the airway hyper-reactivity, airway inflammation, cell count and eosinophil percentage in BALF, levels of total serum IgE and BALF IL-4 and IL-13, and GATA-3 protein expression in the lung tissue were significantly increased in Group B (P0.05). As compared with Group B, all the above-mentioned indices in Group C and D were lower, showing respective significant difference (P0.05), and significant difference was also shown between the two APG treated groups (P0.05).APG could reduce the airway inflammation and hyper-reactivity by down-regulating the expressions of pulmonary GATA-3 and Th, cytokines, which is a potential drug for asthma therapy.

The scaffold protein IQGAP1 shows elevated levels in several cancer types, but its expression in hepatocellular carcinoma is unknown. We found that 58% of human hepatocellular carcinoma tissue samples had increased IQGAP1 expression compared to adjacent normal tissue. Overexpressing IQGAP1 raised the in vivo tumorigenicity of hepatocellular carcinoma cells, and forced overexpression of IQGAP1 in vitro stimulated cell proliferation. Cell growth was reduced by knockdown or mutation of IQGAP1, or by treatment of cells with a phosphotidylinositol 3-kinase inhibitor. To determine the mechanism by which IQGAP1 overexpression affected hepatocellular carcinoma cells, we confirmed its interaction in these cells with mammalian target of rapamycin (mTOR), a serine/threonine kinase that integrates signals about nutrient and energy status with downstream effectors that influence cell division. In addition, we discovered a new interaction involving IQGAP1, mTOR and Akt, which is a downstream target of mTOR. Akt phosphorylation on Ser-473, which is catalyzed by mTOR and required for Akt activation, increased with increasing amounts of IQGAP1, and decreased with IQGAP1 mutation. We hypothesize that IQGAP1 is a scaffold that facilitates mTOR and Akt interaction.

The plasma half-life of therapeutic proteins is a critical factor in many clinical applications. Therefore, new strategies to prolong plasma half-life of long-acting peptides and protein drugs are in high demand. Here, we designed an artificial gelatin-like protein (GLK) and fused this hydrophilic GLK polymer to granulocyte-colony-stimulating factor (G-CSF) to generate a chimeric GLK/G-CSF fusion protein. The genetically engineered recombinant GLK/G-CSF (rGLK/G-CSF) fusion protein was purified from Pichia pastoris. In vitro studies demonstrated that rGLK/G-CSF possessed an enlarged hydrodynamic radius, improved thermal stability and retained full bioactivity compared to unfused G-CSF. Following a single subcutaneous administration to rats, the rGLK/G-CSF fusion protein displayed a slower plasma clearance rate and stimulated greater and longer lasting increases in circulating white blood cells than G-CSF. Our findings indicate that fusion with this artificial, hydrophilic, GLK polymer provides many advantages in the construction of a potent hematopoietic factor with extended plasma half-life. This approach could be easily applied to other therapeutic proteins and have important clinical applications.

Beta(2)-adrenoceptor (beta(2)AR) desensitization is a common problem in clinical practice. beta(2)AR desensitization proceeds by at least such three mechanisms as heterologous desensitization, homologous desensitization and a kind of agonist-induced rapid phosphorylation by a variety of serine/threonine kinases. It is not clear whether there are other mechanisms. This study aimed to investigate potential mechanisms of beta(2)AR desensitization.Twenty-four BALB/c (6-8 weeks old) mice were divided into three groups, which is, group A, phosphate buffered saline (PBS)-treated; group B, ovalbumin (OVA)-induced; and group C, salbutamol-treated. Inflammatory cell counts, cytokine concentrations of bronchoalveolar lavage fluid (BALF), pathological sections, total serum IgE, airway responsiveness, membrane receptor numbers and total amount of beta(2)AR were observed. Asthmatic mouse model and beta(2)AR desensitization asthmatic mouse model were established. Groups B and C were selected for two-dimensional gel electrophoresis (2DE) analysis so as to find key protein spots related to beta(2)AR desensitization.Asthmatic mouse model and beta(2)AR desensitization asthmatic mouse model were verified by inflammatory cell count, cytokine concentration of BALF, serum IgE level, airway hyperreactivity measurement, radioligand receptor binding assay, Western blot analysis, and pathologic examination. Then the two groups (groups B and C) were subjected to 2DE. Two key protein spots associated with beta(2)AR desensitization, Rho GDP-dissociation inhibitor 2 (RhoGDI(2)) and peroxiredoxin 5, were found by comparative proteomics (2DE and mass spectrum analysis).Oxidative stress and small G protein regulators may play an important role in the process of beta(2)AR desensitization.

Nuclear factor kappaB (NF-kappaB) overactivation plays a crucial role in T-helper 2 (Th2)-biased allergic airway inflammation by increased activation and decreased apoptosis of CD4(+) T cells. We have shown that targeted NF-kappaB suppression in dendritic cells by adenoviral gene transfer of a novel mutated inhibitor of NF-kappaB (IkappaBalpha) (AdIkappaBalphaM) contributes to T-cell tolerance, but the immunosuppressive action of AdIkappaBalphaM on memory (CD45RO(+)) CD4(+) T cells remains enigmatic.CD45RO(+) T cells from Dermatophagoides farinaei-sensitized asthmatic patients, untransfected or transfected with AdIkappaBalphaM or AdLacZ (beta-galactosidase) for 24 h, were stimulated with anti-CD3 (1.0 microg/ml) plus anti-CD28 (0.5 microg/ml) monoclonal antibody for an additional 24 h. IkappaBalphaM transgene expression and NF-kappaB activation were detected by polymerase chain reaction (PCR), reverse transcription-PCR (RT-PCR), Western blot analysis, and electrophoretic mobility shift assay. Phenotype and apoptosis were measured by flow cytometry, annexin V binding, and terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling analyses. Cytokine production and cell proliferation were determined using enzyme-linked immunosorbent assay and [(3)H] thymidine incorporation.A unique 801-bp IkappaBalphaM cDNA and a dose-dependent increase in IkappaBalphaM transgene expression were observed in AdIkappaBalphaM-transfected CD45RO(+) T cells. Significantly, AdIkappaBalphaM inhibited CD3/CD28-mediated NF-kappaB activation in CD45RO(+) T cells, leading to evident apoptosis, reduction of eotaxin, RANTES, Th1 [interferon (IFN)-gamma and interleukin (IL)-2], and Th2 (IL-4, IL-5, and IL-13 despite a slight decrease in IL-10) cytokines and secondary proliferative response. AdIkappaBalphaM also upregulated cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and downregulated CD69 besides no change in CD28.IkappaBalphaM might be beneficial to augment memory CD4(+) T-cell tolerance through modulating B7-CD28/CTLA-4 co-stimulatory pathways and NF-kappaB-dependent cytokine profiles in allergic inflammatory diseases including asthma.

To observe human papillomavirus (HPV) infection in women with cervical lesions in Huzhou area of Zhejiang province.720 samples of cervical secretion or exfoliated cells were collected from women with cervical lesion in Huzhou area. Human papillomavirus was detected by suspension array technique.Positive HPV infection was detected in 25.42% cases (183/720), with 135 cases of single HPV type, 33 of dual HPV types and 15 of multiple HPV types. HPV16 and HPV58 were the most prevalent types in 183 HPV positive cases.The most prevalent high-risk types of HPV are HPV16 and HPV58 in Huzhou area of Zhejiang province.

A novel recombinant exendin-4 human serum albumin fusion protein (rEx-4/HSA) expressed in Pichia pastoris was prepared and characterized. Ex-4 is a 39-amino acid peptide isolated from the salivary gland of the lizard Heloderma suspectum and is thought to be a novel therapeutic agent for type 2 diabetes. But to gain a continued effect, the peptide has to be injected twice a day owing to its short plasma half-life (t(1/2) = 2.4 h). To extend the half-life of Ex-4 molecule in vivo, we designed a genetically engineered Ex-4/HSA fusion protein. Between Ex-4 and HSA, a peptide linker GGGGS was inserted and the fusion protein was expressed in methylotrophic yeast P. pastoris with native HSA secretion signal sequence. The recombinant protein was secreted correctly and was obtained with high purity (typically98%) by a three-step purification procedure. cAMP assay demonstrated that the fusion protein had a bioactivity similar to Ex-4 for interaction with GLP-1 receptors in vitro. Results from oral glucose tolerance test indicated that rEx-4/HSA could effectively improve glucose tolerance in diabetic db/db mice. Pharmacokinetics studies in cynomologus monkeys also showed that rEx-4/HSA had a much longer plasma half-life. Therefore, rEx-4/HSA fusion protein could potentially be used as a new recombinant biodrug for type 2 diabetes therapy.

AIM: To study mutations in the P-type ATPase (ATP7B) gene responsible for Wilson disease (WD) in the Eastern Chinese population, and the possible correlation of specific mutations with clinical characteristics. METHODS: Mutations of the ATP7B gene were sought by means of direct sequencing in 50 Eastern Chinese WD patients of Han ethnic origin. RESULTS: Two novel mutations, Asp96Gly and Asp196Glu, were first identified. We also compared the characterization of mutations in ATP7B with the clinical findings, and a significant correlation with hepatic manifestations between patients carrying the Arg778Leu mutation and those without was found. CONCLUSION: Gene sequencing analysis was shown to have a high detection rate and accuracy. It may become the first priority in screening of WD patients.