Adolescent ; Asthma ; blood ; Child ; Child, Preschool ; Cough ; blood ; Eosinophil Cationic Protein ; blood ; Female ; Humans ; Immunoglobulin E ; blood ; Infant ; Male

Objective To identify hepatitis C virus(HCV)-specific cytotoxicity T lymphocytes (CTL)epitopes by the combination of T epitopes prediction software and in vitro assays.Methods HCVspecific CTL epitopes were predicted by T epitope prediction software Rankpep and then candidate HCV-specific CTL epitopes were selected.Candidate HCV-specific CTL epitopes were used to stimulate PBMC of HCV-infected patients and healthy volunteers.and then enzyme-linked immunospot(ELISPOT)and intracellular cytokine staining(ICS)were used to measure the frequencies of IFN-γ-producing cells in total PBMC and the percentages of IFN-γ+CD8+T cells in total CD8+T cells,respectively.Results Five candidate CTL epitopes[NS3 450(TVPQDAVSR),NS3 594(GPTLLYRL),Ns4b 78(sMMAFSAAL),NS5a 416(SEENVSVVF)and NS5a 367(TVSSALAEL)]were used to stimulate PBMC of ten HCV-infected patients and two healthy volunteers.PBMC of seven HCV-infected patients secreted IFN-γ while PBMC of healthy volunteers did not produce IFN-γ.The frequencies of peptide-specific IFN-γ-producing cells ranged from 5 to 36 SFC/105 PBMC and the percentages of peptide-specific IFN-γ+CD8+T cells ranged from 0.02%-0.25%.Conclusion Resuhs of ELISPOT assay and ICS assay confirm that these five peptides NS3 450,NS3 594,NS4b 78,NS5a 416 and NS5a 367 are identified as Hovel HCV-specific CTL epitopes.

Objective To explore the role of hydrogen peroxide in mesenteric artery contraction of cirrhotic rats with portal hypertension,which was induced by bile duct ligation.Possible mechanism in RhoA/ROCK signal pathway was also part of the focus.Methods The bile duct ligation-induced cirrhotic rats and normal rats (control group) were treated equally with PEG-catalase(10 000 U/kg-1 · d-1,ip.) or by its vehicle for 8 days.Then the level of H2O2 in mesenteric arteries was detected.The contractile response to norepinephrine of arterioles was analyzed by vascular perfusion system.The protein expressions of the α1 adrenergic receptor,β-arrestin-2 and Rho kinase-1 (ROCK-1),and the activity of ROCK-1 were measured by western blot.In addition,the interaction of α1-adrenergic receptor with β-arrestin-2 was assessed by co-immunoprecipitation.Results Compared to normal rats,the dose-response curve of the mesenteric arterioles in response to norepinephrine shifted to the right,and the EC 50 increased in the rats with portal hypertension.PEG-catalase treatment can decrease the hydrogen peroxide level in arteries,thus significantly lowered EC50 and improved the reactivity to norepinephrine of the mesenteric arterioles in portal hypertension rats.No significant difference in the α1-adrenergic receptor amounts was observed among groups.There was remarkable decreases in the protein expressions of β-arrestin-2 and its interaction with the α1-adrenergic receptor in cirrhotic rats with PEG-catalase treatment.PEG-catalase also increased the amount and activity of ROCK-1 in cirrhotic rats.Conclusions The level of hydrogen peroxide increases in the mesenteric arteries in bile duct ligation-induced cirrhotic rats.And it enhances the β-arrestin-2 expression and its interaction with the α1-adrenergic receptor,which subsequently decreases the amount and activity of ROCK as well as the contractility of mesenteric arteries in response to vasoconstrictors.

Pancreatic ductal adenocarcinoma (PDAC) is one of the five most malignant cancer. ZX-1201 is one of the active constituents in Alismatis Rhizoma,a well-known traditional Chinese medi-cine with a wide variety of pharmacological properties including diuretic,anti-hyperlipidemic,anti-atheroscle-rotic,anti-cancer,anti-inflammatory and anti-oxidative activities.We investigated the inhibitory effect of ZX-1201 on pancreatic cancer and the relevant molecular mechanism in vitro and in vivo. ZX-1201 inhibited the growth and metastasis of PANC-1 cells in BALB/c nude mice significantly.ZX-1201 inhibited the function of AQP1 via directly interaction and involved in the reversion process of ZX-1201 on TGF-β1. CTGF was an important protein in the reversion process of ZX-1201 on TGF-β1.ZX-1201 inhibited the migration of PANC-1 and CPFAC-1 cells induced by TGF-β1in vitro.ZX-1201 reversed the down-regu-lated of epithelial markers and up-regulated of mesenchymal markers, as well as the up-regulated of Snail and p-Smad2/3 induced by TGF-β1.And ZX-1201 reversed Epithelial-Mesenchymal Transition by down-regulating AQP1 and inhibiting translocation of β-catenin, the promotor of CTGF. According to these,ZX-1201 inhibited the migration of pancreatic cancer cells.We concluded that ZX-1201 inhibited the growth and metastasis of PANC-1 cells in vivo significantly.And AQP1,β-catenin and CTGF were the pivotal proteins in the process of ZX-1201 inhibiting PANC-1 cells migration induced by TGF-β1.

The intestinal absorption properties of the main effective components (glycyrrhizic acid, isoliquiritigenin, 6-gingerol, ginsenoside Rb1, atractylode-I) in Lizhong decoction (LZD) extracts were investigated with an in situ single-pass intestinal perfusion model in rats. UPLC-TQ-MS was used to determine the concentration of the five components in the intestinal perfusion. Animal welfare and experimental procedures were in accordance with the regulations of the Animal Ethics Committee of Nanjing University of Chinese Medicine. As evaluation indexes for the intestinal absorption characteristics, the absorption rate constant (K_a) and the apparent permeability coefficient (P_eff) of the five main ingredients were analyzed. Results showed that the best absorption sites for glycyrrhizic acid, isoliquiritin and 6-gingerol were the ileum, colon and duodenum, respectively, and the differences between different intestinal segments were statistically significant (P <0.05). There was no notable difference in K_a and P_eff between ginsenoside Rb1 and atractylode-I in the different intestinal segments (P > 0.05), suggesting that they were absorbed throughout. The five components were well-absorbed in the whole intestine (P_eff > 1.0×10^-3 cm·min^-1), indicating that LZD is suitable for preparing sustained, controlled release and enteric-coated preparations.

Objective To investigate the expression of the interleukin-1 receptor(IL-1R)Ⅰ,IL-1RⅡand IL-1R accessory protein(IL-1RAcP)in osteoarthritis and analyse their biological significance.Methods Immunohistochemistry and reverse transcription-polymerase chain raction(RT-PCR)were adopted to detect the expression of IL-1RⅠ,IL-1RⅡand IL-1RAcP on the synovium of 107 OA patients.Results Immunohis- tochemistry showed strong positive expression of IL-1RⅠand IL-1RAcP,and positive expression of IL-1RⅡ. The expression was distributed in lining cells,monocyts and vascular endothelial cells of the sublining area, but all of them were negative or weak positive in normal synoviums.RT-PCR showed the expression of IL-1RⅠ,IL-1RⅡand IL-1RAcP in OA synoviums was significantly enhanced than normal synoviums (P＜0.05),and the expression of IL-1RⅠwas significantly enhanced than IL-1RⅡ(P＜0.05),but no sig- nificant difference with IL-1RAcP(P＞0.05).In stageⅡandⅢOA synoviums,the expression of IL-1RⅠand IL-1 RAcP had no significant difference with normal synoviums(P＞0.05).The expression of IL-1RⅡin stageⅢOA synoviums was significantly enhanced than normal(P＜0.05).Conclusion IL-1RⅠ,IL-1RⅡand IL-1RAcP play significant roles in the pathogenesis of OA,especially IL-1RⅠand IL-1RAcP.But their increase is only observed in the early stage of OA.These suggest that they may have no association with the development of OA and have no direct association with the severity of OA.OA can be cured by interrupting the signal transduction path in which IL-1 has played biological roles.

Objective: Many physicians and patients still have concerns about the safety of breastfeeding in mothers infected with hepatitis B virus; we evaluate the safety of the newborn and the women with HBeAg positive and high viral load, who received nucleoside analogues to block maternal to child transmisssion and selected postpartum breastfeeding after drug discontinuance. Methods: This prospective, observational study enrolled 60 HBeAg positive patients and HBV-DNA >2*10E+ 5 IU/ml, all patients started antiviral treatment for blocking maternal to child transmission at 24-28 weeks of pregnancy. All the newborns received the active-passive immunization therapy with hepatitis B immunoglobulin (HBIG) and HBVac. After the delivery, patients with normal liver function discontinued the antiviral drug and selected breastfeeding voluntarily. The safety of breastfeeding were compared with patients selected artificial feedings, they were followed up for 7 months. Primary measurements were the proportion of mothers with abormal liver function after stopping the drug and the level of newborn’s anti-HBs at 6 months of age; secondary measurements were the positive rate of neonatal HBsAg and the HBV-DNA value of the patients at 6 weeks postpartum. Results: From December 1, 2015 to May 1, 2017, 415 patients were enrolled in Beijing You’an Hospital Affiliated to the Capital Medical University and all these patients were born following full-term single-child pregnancy. After the delivery and drug withdrawal, there was no significant difference in the incidence of ALT elevation between the breast fed group and the artificially fed group: 29 /220 versus 30/195, (χ²=0.411, P=0.521). Patients continued to take the antiviral medicines between the breast feeding group and the artificial feeding group: 15 /220 versus 20/195, (χ²=1.487, P=0.223), there were no significant differences between them (P> 0.05). At the month 7, there were no significant difference between the breast fed group and the artificially fed group (747.62±374.08 mlU/ml versus 709.76±374.32 mlU/ml, t-value: 0.309, P-value>0.05). At birth, hepatitis B surface antigen (HBsAg) was detected in 15/220 and 20/195 of newborns in the breast feeding and artificially fed groups, respectively. At month 7, an intention-to-treat analysis indicated 0/220 of HBsAg infants from the breast fed versus 0/195 in the artificially fed group (P>0.05) and no significant difference was found in the rate of positive HBsAg between the two groups. In the breast fed group, the mean HBV DNA at baseline was significantly higher than that of the artificially fed group: (1.17±1.82) E+ 8 IU/ml versus (1.12±0.39)E+ 8 IU/ml, and the difference was statistically significant. Conclusions We have not found the relationship between the rate of neonatal infection and the increase of maternal abnormal liver function in HBeAg positive and high viral load patients, who chose breastfeeding after drug discontinuance.

OBJECTIVETo explore the effects of adenovirus-delivered tissue inhibitor of metalloprotein- ases-3 (Ad-TIMP-3) on the irradiation sensitivity of human papillomavirus (HPV)-positive cervical cancer cells.

METHODSAn adenovirus expressing TIMP-3 (Ad-TIMP-3), alone or in combination with irradiation,was used to treat HPV-positive cervical cancer cells HeLa-Luc and CaSki. The effects of Ad-TIMP-3 on the proliferation of HeLa-Luc and CaSki cells were detected with MTT assay. The effect of the combination of Ad-TIMP-3 and X-ray on the proliferation of cells were determined by clone formation assay. Twenty nude mice were equally randomly divided into four groups: normal control group,Ad-TIMP-3 group,X-ray group,and combination group. The size of tumor was measured separately,and tumor growth curves were drawn.

RESULTSAd-TIMP-3 significantly inhibited the proliferation of HPV-positive cervical cancer cells in a dose-dependent manner. Combination of Ad-TIMP-3 and X-ray significantly decreased the clones of HeLa-Luc and CaSki than Ad-TIMP-3 or X-ray alone (P<0.05). The tumor weights were (0.216±0.098), (0.276±0.073), and (0.044±0.043) g, respectively, in Ad-TIMP-3 group, X-ray group,and combination group, which were all significantly lower than that in normal control group [(0.534±0.218) g] (all P<0.05). In addition,the tumor weight in the combination group was significantly lower than that in Ad-TIMP-3 group and X-ray group (both P<0.05). The tumor inhibition rate was 59.60%, 48.30%, and 91.80% in X-ray group, Ad-TIMP-3 group and combination group, respectively.

CONCLUSIONSAd-TIMP-3 can effectively inhibit the proliferation of cervical cancer cells. When combined with X-ray,it can remarkably increase the irradiation sensitivity of HPV-positive cervical cancer cells,and thus suppress the tumorigenesis capability of these cells in vivo.

Adenoviridae ; genetics ; Animals ; Cell Line, Tumor ; Cell Proliferation ; Female ; Genetic Vectors ; Humans ; Mice ; Mice, Nude ; Papillomaviridae ; genetics ; Radiation Tolerance ; Tissue Inhibitor of Metalloproteinase-3 ; genetics ; Transfection ; Uterine Cervical Neoplasms ; pathology ; radiotherapy ; virology

A simple, rapid and sensitive colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was established to detect norovirus genotype GII. The method employed a set of six specially designed primers that recognized eight distinct sequences of RNA-dependant RNA polymerase and capsid protein gene for amplification of nucleic acid under isothermal conditions at 65 degrees C for 60 minutes. The amplification process of RT-LAMP was monitored by the addition of HNB (Hydroxy naphthol blue) dye prior to amplification. A positive reaction was indicated by a color change from violet to sky blue and confirmed by agarose electrophoresis. The specificity of the RT-LAMP was validated by detecting several different diarrhea viruses including norovirus genotype GII. The sensitivity was determined by serial dilutions of RNA molecules from in vitro transcription of norovirus genotype GII in parallel with conventional RT-PCR detection. The assay was further evaluated with 93 clinical specimens of diarrhea patients. The results showed that the sensitivity of RT-LAMP was 1 000 copies/microL with a high specificity and the relative sensitivity was at the same level as that of conventional RT-PCR. Positive rate of RT-LAMP in analysis of clinical specimens was approximately the same as that of conventional RT-PCR as well. This colorimetric RT-LAMP assay was potential for rapid detection of norovirus genotype GII on spot due to the observation of visual result with high specificity and sensitivity, time-saving and cost benefit.
Caliciviridae Infections ; diagnosis ; virology ; Colorimetry ; methods ; Feces ; virology ; Genotype ; Humans ; Norovirus ; genetics ; isolation & purification ; Nucleic Acid Amplification Techniques ; methods