Acute Disease ; Adolescent ; Asthma ; immunology ; Child ; Child, Preschool ; Female ; Humans ; Infant ; Male ; RNA, Viral ; genetics ; Respiratory Syncytial Viruses ; genetics ; Respiratory Tract Infections ; immunology ; virology ; Reverse Transcriptase Polymerase Chain Reaction

Objective To clone the gene encoding protein of EspA and Stx2B from EHEC OI57:H7 by DNA recombinant technology, construct prokaryotic expression vector pET-28a ( + )-espAstx2B, express fusion protein of EspA-Stx2B and to analyze the biological and immunological characteristics of the fusion protein. Methods the sequence encoding the protein of EspA and Stx2B was amplified by PCR from the enterohemorrhagic Escherichia coli strain. The amplified products were connected with linker by recombinant technology and cloned into pET-28a( + ) vector. The vector was then transferred to the host cells E. Coli BL21 strain (DE3). Following, the protein expression was induced by IPTG. The expression quantities and style of fusion protein was then determined by SDS-PAGE. Its immunoreactivity was analyzed by Western blot. Finally, BALB/c mice were injected with the preliminarily purified recombination protein EspA-Stx2B, then oral challenged these mice with EHEC O157-SMR2 and counteracted toxic substances with O157 ultrasonic supernatant. Results The determination of the sequence encoding of the espA-stx2B fusion gene has 100% of consistency with the sequence from GenBank Sakai strain and contrivable linker. This fusion protein EspA-Stx2B was expressed as inclusion body formation and the percentage is approximately 40%. Western blot suggested the fusion protein has excellent immunoreactivity. Titer of antiserum of the mice to EspA-Stx2B increased evidently. EspA-Stx2B could not decrease bacterial number attached to the intestinal tract of mice based on fecal shedding of Oi57 in mice. In the test of death of BAI,B/c causing by conteracting toxic substances with O157 ultrasonic supernatant, immunoprotection of EspA-Stx2B rate was 66.7%. Conclusion A recombinant plasmid that has high performance on expression of EspA-Stx2B prorein was successfully constructed in present study, and the fusion protein has excellent immunoreactivity and immunogenicity. EspA-Stx2B could not decrease bacteria] number attached to the intestinal tract of mice based on fecal shedding of O157 in mice, but evidently decrease the mortality rate of the mice. The antiEspA and anti-Stx2B had immunoprotection effect by different means. These results may provide the foundation for the further development on EHEC O157:H7 double subunit vaccine.

OBJECTIVESTo study the relationship between HBV genotypes and the efficacy of antiviral therapies.

METHODSHBV genotypes of 90 hepatitis B e antigen positive patients with chronic hepatitis B (CHB) were determined by PCR sandwich hybridization-ELISA technique. Forty-one patients with CHB were treated with lamivudine (100 mg/day) for 48 weeks and 49 patients with CHB were given alpha-interferon (3 MU/QOD) therapy for 48 weeks. The serological, biochemical and virological symbols were measured before, during and after treatment for all the patients.

RESULTSOf the 90 patients, genotype B HBV was found in 16 and C in 74. There was no difference in the rate of response to lamivudine treatment between patients with genotype B or C HBV (33.3% vs. 20.0%) after 48 weeks treatment with lamivudine in the 41 patients. Of the 49 HBeAg positive CHB patients treated with alpha-interferon for 48 weeks, in HBV genotype B and C patients the rates of normalization of ALT were 60.0% and 20.5%; the rate of HBeAg turning to negativity was 50.0% and 17.9%; and the rate of HBV DNA undetectability was 50.0% and 17.9%. The rate of response to the interferon treatment was significantly higher in patients with HBV genotype B compared to those with genotype C.

CONCLUSIONSOur study shows that there is no influence on the lamivudine treatment effects for the HBV genotype B and C CHB patients, but the alpha-interferon treatment for HBV genotype B CHB patients is more effective than that for the genotype C ones.

Adolescent ; Adult ; Antiviral Agents ; pharmacology ; therapeutic use ; Female ; Genome, Viral ; Genotype ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; drug effects ; genetics ; Hepatitis B, Chronic ; drug therapy ; virology ; Humans ; Interferon-alpha ; pharmacology ; therapeutic use ; Lamivudine ; pharmacology ; therapeutic use ; Male ; Young Adult

OBJECTIVETo investigate the expressions of small heterodimer partner (SHP) and target gene cholesterol-7-hydroxylase (CYP7A1) in livers of rats with intrahepatic cholestasis of pregnancy (ICP), and to study the mechanism of ICP.

METHODSThirty SD rats (pregnant for 15 days) were equally and randomly divided into two groups: an estradiol benzoate (EB) group and a normal saline (NS) group. Two ml blood was drawn from each rat before and on the 5th day after medicine administration to measure the levels of ALT, AST, ALP, TBA, TBIL, and DBIL. After delivery, the histopathological changes of the mother rat livers were studied. The mRNA and protein expressions of SHP and CYP7A1 in the livers were determined by RT-PCR and Western blot.

RESULTS(1) In the EB group, the serum levels of ALT, AST, ALP, TBA, TBil, and DBil after EB administration increased significantly (P less than 0.01), but there were no significant changes in the NS group (P more than 0.05); (2) Intrahepatic cholestasis appeared in the EB group, but not in the NS group; (3) The mRNA expressions of SHP and CYP7A1 were significantly higher in the EB group than in the NS group [(SHPmRNA: NS 0.365+/-0.0317 vs EB 0.4865+/-0.0237, P less than 0.01), (CYP7A1 mRNA: NS 0.3570+/-0.0175 vs EB 0.4802+/-0.0217, P less than 0.01)]; (4) The protein expressions of SHP and CYP7A1 were also higher in the EB group than that in the NS group [(SHP: NS 0.3762+/-0.0284 vs EB 0.5033+/-0.0274, P less than 0.01), (CYP7A1: NS 0.3570+/-0.0175 vs EB 0.4802+/-0.0217, P less than 0.01)].

CONCLUSIONEstrogen induces ICP in rats. The mRNA and protein expressions of SHP and CYP7A1 in livers of the ICP rats were increased, which causes more bile acids to be synthesized. This may be one of the mechanisms of ICP.

Animals ; Cholestasis, Intrahepatic ; chemically induced ; metabolism ; Cholesterol 7-alpha-Hydroxylase ; metabolism ; Estradiol ; analogs & derivatives ; pharmacology ; Female ; Liver ; metabolism ; Pregnancy ; Pregnancy Complications ; chemically induced ; metabolism ; Rats ; Receptors, Cytoplasmic and Nuclear ; metabolism

OBJECTIVETo investigate the effects and mechanism of farnesoid X receptor (FXR) and its ligands on the metabolism of bile acids in rats with estrogen-induced intrahepatic cholestasis of pregnancy (ICP).

METHODSAn ICP rat model was established with estradiol benzoate (EB) injections. Then FXR ligand chenodeoxycholic acid (CDCA) was administrated (100 mg/kg daily) to ICP rats for 5 days. The serum TBA and expression of FXR and bile salt export pump (BSEP) in the rat livers were examined by immunohistochemistry and reverse transcription PCR.

RESULTSThe levels of TBA in the CDCA group rats were significantly lower than the untreated rats [(17.2+/-4.1)micromol/L vs (29.3+/-6.4)micromol/L], and the expressions of mRNA and protein of FXR were significantly higher [(0.76+/-0.09 vs 0.53+/-0.06, P<0.05 and 2.35+/-0.06 vs 1.83+/-0.05, P<0.017, respectively)], and the expressions of BSEP were also higher [(0.99+/-0.21 vs 0.76+/-0.07, P<0.017 and 1.88+/-0.03 vs 1.46+/-0.06, P<0.017, respectively)].

CONCLUSIONSFXR plays an important role in modulating the metabolism of bile acids. CDCA can lower the levels of serum TBA by upregulating the expression of FXR and BSEP and then increasing the transport of the bile acids. These facts might present a new idea and target for the treatment of ICP.

Animals ; Bile Acids and Salts ; metabolism ; Chenodeoxycholic Acid ; pharmacology ; Cholestasis, Intrahepatic ; chemically induced ; genetics ; metabolism ; Estrogens ; pharmacology ; Female ; Pregnancy ; Pregnancy Complications ; chemically induced ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Cytoplasmic and Nuclear ; agonists

Objective To evaluate the short-term efficacy and safety of sevelamer hydrochloride in treating maintenance hemodialysis (MHD) patients with hyperphosphemia.Methods A multicenter,open-labeled,self-control study was performed.Phosphate binders were discontinued during a two-week washout period.Patients with more than 1.78 mmol/L serum phosphorus after two-week washout period were eligible for the trial.The dose was adjusted every two weeks as necessary to achieve serum phosphorus control. Sevelamer hydrochloride was administered to 138 MHD patients for 10 weeks and a second two-week washout period followed.Results A total of 111 from 138 patients fulfilled the whole 14-week study. Mean serum phosphorus and calcium-phosphate products starte to decline after two-week sevelamer hydrochloride treatment. By the end of 10-week sevelamer hydrochloride treatment, mean serum level of phosphorus [(1.85±0.50) vs (2.57±0.54) mmol/L,P＜0.01],calcium-phosphate product [(4.16± 1.72) vs (5.79 ± 1.50) mmol2/L2,P＜0.01 ] and low density lipoprotein [(1.64±0.76) vs (2.31 ±0.87) mmol/L,P＜0.01] were significantly decreased,while the adjusted serum level of calcium and serum intact parathyroid hormone kept steady.Both serum phosphorus and calcium-phosphrus product increased after the second washout period, but the levels were still lower as compared to pre-treatment [(2.26±0.71) vs (2.57±0.54) mmol/L; (5.12±1.63) vs (5.79±1.50) mmol2/L2,P＜0.01].Of the 138 patients involved,214 episodes in 106 patients and 121 episodes in 89 patients were reported as adverse events and adverse drug reaction respectively. Gastrointestinal symptoms,of which most were mild or moderate,happened to 68.1％ (94/138) patients. Conclusions Sevelamer hydrochloride can control serum phosphorus and reduce the levels of calcium-phosphorus product and cholesterol.Slight gastrointestinal symptoms like constipation are common during the treatment.

OBJECTIVETo evaluate their long-term outcome and the efficacy and economic significance of antiviral drugs by investigating the long-term health-related quality of life (HQL) in chronic hepatitis B (CHB) patients.

METHODSThe HQL of 101 CHB patients with biopsy-proven 6 to 18 years ago and 105 persons of general population as control was studied with revised SF-36 questionnaire.

RESULTSThe HQL in CHB patients was lower than that in general population in physical functioning, role physical, general health, mental health, and specific symptoms (mu > or = 2.10, P<0.05).

CONCLUSIONSThe long-term HQL in chronic hepatitis B patients is poor.

Adolescent ; Adult ; Antiviral Agents ; therapeutic use ; Cost of Illness ; Female ; Follow-Up Studies ; Hepatitis B, Chronic ; drug therapy ; economics ; Humans ; Male ; Middle Aged ; Quality of Life ; Surveys and Questionnaires

The objective of this study was to investigate the expression of exogenous hPDGF-A and hBD(2) in gene-modified bone marrow mesenchymal stem cells (BM-MSCs) in vitro and in vivo. Recombinant adenovirus vector expressing hPDGF-A/hBD(2) genes was constructed and packaged into virion. Primary isolated and cultured BM-MSCs were transfected by using hPDGF-A hBD(2), then the expressions of exogenous hPDGF-A/hBD(2) were detected by immunocytochemical staining in vitro. The conditioned medium (serum-free cultured supernatant of BM-MSCs transfected with recombinant adenovirus) collected from gene-modified BM-MSCs was applied to scratch wound on monolayer cells of multipotential cell line 10T1/2 in order to confirm the stimulative effect of hPDGF-A on cell migration. Gene-modified BM-MSCs were topically transplanted on wound of rats with radiation and skin excision combined injury. The distribution of BM-MSCs and expression of hPDGF-A/hBD(2) on the wound was observed by fluorescent microscopy and immunohistochemical staining respectively. The results indicated that the rat BM-MSCs transfected with recombinant adenovirus could express the EGFP in vitro. The immunofluorescent cytochemistry assay showed that the gene-modified BM-MSCs expressed the hPDGF-A and hBD(2). The scratch test confirmed that the percentage of healing area of wound in cultured supernatant group of gene-modified BM-MSCs was significant higher than that in control group on 8, 12, 24 and 48 hours (p < 0.05). The fluorescence microscopy of exogenous gene-modified BM-MSCs transplanted on wound revealed that the gene-modified BM-MSCs could higher express exogenous genes of EGFP at least within 2 weeks. The immunohistochemistry staining of wound indicated that the expression of exogenous genes began from day 3, reached to peak on day 7, and still visible on day 21 even though the expression became weak because of the possible dilution of the exogenous genes during cell division. It is concluded that efficient expression of exogenous hPDGF-A/hBD(2) in gene-modified BM-MSCs are demonstrated both in vitro and in vivo, which suggests that the molecular mechanism underlying chronic wound-healing accelerated by the strategy combining cell therapy with gene therapy.
Animals ; Bone Marrow Cells ; cytology ; metabolism ; Gene Expression ; Genetic Vectors ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Platelet-Derived Growth Factor ; genetics ; Rats ; Rats, Sprague-Dawley ; Transfection ; beta-Defensins ; genetics

BACKGROUNDHuman epididymis secretory protein 4 (HE4) has been proved to be a promising novel biomarker for the detection of epithelial ovarian carcinomas. Compared with CA125, HE4 assay demonstrated an improved ability to discriminate between pelvic mass with malignant and benign disease. Though it is well known that HE4 is overexpressed in ovarian cancer, however, the role of HE4 in the carcinogenesis and progression of ovarian cancer remains unkown.

METHODSIn this study, we explored the role of HE4 in the carcinogenesis and progression of ovarian cancer. We screened nine ovarian cancer cell lines for HE4 expression, and using RNA interference (RNAi), we silenced HE4 gene expression in CaoV3 and SKOV3.ip1 ovarian cancer cell lines. We assessed the effect of HE4 gene silencing on the transformed phenotype by examining the cell cycle, apoptosis, proliferation and transwell migration/invasion in vitro.

RESULTSHE4 gene silencing induces G0/G1 arrest and blocks the progression from the G1 to S phase in CaoV3 and SKOV3.ip1 cells. HE4 knockdown also inhibited cell proliferation, migration and invasion in SKOV3.ip1 cells in vitro.

CONCLUSIONHE4 may be involved in the regulation of the cell cycle and promote ovarian cancer migration and invasion.

Biomarkers, Tumor ; analysis ; Cell Line, Tumor ; Disease Progression ; Epididymal Secretory Proteins ; analysis ; genetics ; physiology ; Female ; Gene Silencing ; physiology ; Humans ; Ovarian Neoplasms ; pathology ; RNA Interference

【Objective】To measure the orbital fat fraction（FF）of Graves ophthalmopathy（GO）patients and normal individuals with MR Water- fat separation（Dixon） technology ，and to explore the value of orbit MR quantitative fat analysis in diagnosis and treatment for GO patients.【Methods】Forty GO patients and 20 normal individuals who underwent orbital MR imaging were analyzed prospectively from February 2017 to February 2019 in the First Affiliated Hospital of Sun Yat- sen University. All participants received bilateral orbital MR examination with oblique coronary FSE T2- weighted imaging combined with two-point Dixon technique，and then the signal intensity values of ipsilateral extraocular muscles and lacrimal glands on water and fat phase images were measured to calculate FF. The differences of mean FF between the two groups were evaluated，and the correlation between FF of GO and TRAb，CAS score，and the course of GO were conducted by Spearman rank correlation analysis. 【Results】 The median FF of extraocular muscles in GO patients was higher than that of normal individuals，and the result showed a statistically significant difference（P < 0.05）， while the FF of lacrimal glands in the two groups showed no statistical difference. The FF of extraocular muscles in the GO group were correlated with TRAb，CAS score and the course of GO（P < 0.05），especially the course was more correlative.【Conclusions】The fat content of extraocular muscles in GO patients were higher than that of normal individuals ，and there was a positive correlation in the fat content among TRAb，CAS score and the course of GO，suggesting that MR fat quantitative analysis may provide a new reference indexes for the evaluation of clinical staging ，curative effect assessment in GO patients.