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OBJECTIVE@#To study the effect of intermittent versus daily inhalation of budesonide on pulmonary function and fractional exhaled nitric oxide (FeNO) in children with mild persistent asthma.@*METHODS@#A total of 120 children, aged 6-14 years, with mild persistent asthma who attended the hospital from January 2016 to January 2018 were enrolled. The children were divided into an intermittent inhalation group with 60 children (inhalation of budesonide 200 μg/day for 6 weeks when symptoms of asthma appeared) and a daily inhalation group with 60 children (continuous inhalation of budesonide 200 μg/day) by stratified randomization. The children were followed up at months 3, 6, 9, and 12 of treatment. The two groups were compared in terms of baseline data, changes in FeNO and pulmonary function parameters, amount of glucocorticoid used, number of asthma attacks, and asthma control.@*RESULTS@#At the start of treatment, there were no significant differences in baseline data, FeNO, and pulmonary function between the two groups (P>0.05). Over the time of treatment, FeNO gradually decreased and pulmonary function parameters were gradually improved in both groups (P0.05). Compared with the daily inhalation group, the intermittent inhalation group had a significantly lower amount of budesonide inhaled (P<0.05) and a significantly higher number of asthma attacks (P<0.05).@*CONCLUSIONS@#Intermittent inhalation and daily inhalation of budesonide can achieve the same level of asthma control in children with mild persistent asthma and both have no influence on the increases in body height and body weight. Daily inhalation of budesonide can produce a better efficiency in reduing FeNO and increasing FEV1%pred. Although intermittent inhalation can reduce the amount of glucocorticoid used, it may lead to a higher risk of asthma attacks.
Subject(s)
Adolescent , Child , Humans , Administration, Inhalation , Asthma , Drug Therapy , Budesonide , Therapeutic Uses , Forced Expiratory Volume , Nitric OxideABSTRACT
This study aims to develop multifunctional drug delivery system based on hollow mesoporous copper sulfide (HMCuS) nanoparticles. This type of nanoparticles is expected to achieve the synergistic treatment of tumor by targeted phototherapy and chemotherapy. The carrier was synthesized by a substitution method, and the anti-cancer drug doxorubicin (DOX) was loaded and then modified by hyaluronic acid (HA) to prepare the anti-cancer drug system DOX/HMCuS-HA. The results suggested that DOX/HMCuS-HA presented uniform spherical structure, with the drug loading efficiency of 33.6%, the particle size and zeta potential being 113.8 ± 6.9 nm and 18.4 ± 2.8 mV, respectively. When 100 μg·mL-1 HMCuS was irradiated under 808 nm laser (2 W·cm-2) for 8 min, the temperature can heat up 51 ℃, demonstrating high photothermal conversion efficacy. Electron spin resonance (ESR) tests and methylene blue degradation experiments showed that HMCuS nanoparticles could simultaneously produce hydroxyl radical (•OH) mediated photodynamic therapy. In addition, HA was responsible for minimizing premature drug release and increasing tumor targeting efficiency by acting as a smart gatekeeper with tumor specific targeting moiety. In vitro drug release experiments showed that the coated HA could be degraded by intracellular lysosomal enzyme hyaluronidase, which facilitated DOX release. The acidic micro-environment of tumor cell and external near infrared (NIR) stimulus could trigger further release of DOX from the nanoparticles. These results point to a new strategy for timely and effective anti-tumor treatment.
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OBJECTIVE@#To investigate the effect of P53 expression on prognosis of patients with double expressor lymphoma(DEL) and the interaction between the expression of MYC, BCL2 and P53 in diffuse large B-cell lymphoma(DLBCL).@*METHODS@#Eighty-eight patients with newly diagnosed DLBCL from 1st September 2012 to 31th May 2018 in Shanxi Dayi Hospital affiliated to Shanxi Medical University were selected. The expressions of MYC、BCL2、P53、CD10、BCL6、MUM and Ki-67 were tested by immunohistochemistry method. The overall survival of patients was analyzed by the Kaplan-Meier method and compared by the log-rank test. The prognostic effect of MYC, BCL2 and P53 expression was analyzed by univariate and multivariate analysis.@*RESULTS@#Compared with patients without P53 expression, the patients with P53 expression had higher LDH level, higher NCCN-IPI scores, lower response to chemotherapy，poorer overall survival(OS) and a higher rate of death(P0.05). Among lymphoma patients with MYC/P53, MYC/BCL2 and BCL2/P53 co-expression, the patients with MYC/P53 co-expression had the worse OS (3 year OS rate：31.6%), followed by the subgroup of patients with MYC/BCL2/P53(3 year OS rate：46.2%), patients with MYC/BCL2/P53 expression(3 year OS rate： 636%) showed a longer OS compared with the other two subgroups（P<0.05）. Multivariate analysis demonstrated that P53 expression and NCCN-IPI were independent prognostic factors in this patient cohort.@*CONCLUSION@#P53 and MYC expressions have a synergistically negative prognostic effect in DLBCL patients. P53 expression augments the negative prognostic effect of MYC/BCL2 double expression. Patients with MYC/P53 co-expression have a worse prognosis in comparison with the patients with MYC/BCL2 double expression.
Subject(s)
Humans , Lymphoma, Large B-Cell, Diffuse , Genetics , Prognosis , Proto-Oncogene Proteins c-bcl-2 , Proto-Oncogene Proteins c-myc , Tumor Suppressor Protein p53 , GeneticsABSTRACT
BACKGROUND: Thyroid cancer stem cells are essential to the recurrence and metastasis of thyroid carcinoma. Leukemia inhibitory factor receptor (LIFR) shows a downward trend in a variety of malignant tumors, and its overexpression can inhibit the recurrence and metastasis of malignant tumors. OBJECTIVE:To explore the effect of LIFR on the stemness maintenance and lung metastasis of thyroid cancer stem cells in vivo. METHODS: Primary thyroid cancer cells TCLM were isolated from the lung metastases of a metastatic thyroid cancer patient. Serum-free suspension culture was used to form tumor cell balls. Flow cytometry was used to screen CD133+phenotype of metastatic thyroid cancer stem cell subpopulation TCLM-S. The overexpressed recombinant lentiviral plasmid containing LIFR and its negative control containing the empty plasmid were infected into thyroid cancer stem cells TCLM-S at the ratio of virus/cell number=20, and screened with 2.0 mg/L puromycin to construct TCLM-SLIFRand TCLM-Scontrolstem cells which stably expressed LIFR and its control. Real-time quantitative PCR (qRT-PCR) was used to detect the expression of LIFR in TCLM-SLIFRand TCLM-Scontrolstem cells. Flow cytometry was used to detect the percentage of CD133+phenotype cell subsets, western blot assay was used to detect the expression of tumor stemness related factors SOX2, Oct4, Nanog and tumor invasion and metastasis related proteins E-cadherin, matrix metalloproteinase (MMP)-2, MMP-7 in TCLM-SLIFRand TCLM-Scontrol stem cells. TCLM-SLIFRand TCLM-Scontrolstem cells were respectively injected into BALB/c nude mice by tail vein, and the lung metastasis model of thyroid cancer stem cells was constructed. The effect of LIFR overexpression on lung metastasis was observed. RESULTS AND CONCLUSION: Compared with TCLM-Scontrolcells, the expression of LIFR in TCLM-SLIFRcells was significantly increased, the proportion of CD133+phenotype stem cell subsets was significantly decreased, the expression of SOX2, Oct4 and Nanog were significantly decreased, the expression of E-cadherin was significantly increased, and the expression of MMP-2 and MMP-7 was significantly decreased. Moreover, the number of lung metastasis in nude mice given TCLM-SLIFRcells was significantly decreased as compared with those given TCLM-Scontrol cells.To conclude,LIFR overexpression can decrease the stemness and ability of lung metastasis in vivo.
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AIM: To investigate and analyze the changes of corneal biomechanics of normal eyes,forme frusta keratoconus eyes,subclinical keratoconus eyes and clinical keratoconus eyes by Corneal visualization Scheimpflug technology (Corvis ST),and provide clinical basis for early diagnosis of keratoconus.METHODS: Case-control study.We randomly selected 40 normal eyes as normal group,15 forme frusta keratoconus eyes as forme frusta keratoconus group,23 subclinical keratoconus eyes as subclinical keratoconus group,and 40 clinical keratoconus eyes as keratoconus group.The biomechanical parameters of each group were measured by Corvis ST.The receiver operating characteristic(ROC) curves was plotted to distinguish keratoconus from the normal cornea.RESULTS: There was no significant difference in the parameters of biomechanics between normal group and forme frusta keratoconus group (P>0.05).Compared to normal group and subclinical keratoconus group,the parameters second applanation length(AL2),first velocity of applanation (AV1),central curvature radius at highest concavity (HC-radius),deformation amplitude (DA) were revealed statistically significant differences(P<0.05).The biomechanical parameters of the keratoconic group were significantly different from those of normal group except for the second velocity of applanation (AV2),time from the start until the highest concavity(HC-time),peak distance (PD).ROC curve showed that the DA(area under the curve:0.891±0.028) was the best predictive parameter to distinguish keratoconus from the normal eyes.CONCLUSION: The corneal biomechanical parameters of forme frusta keratoconus group are not changed compared with normal group.The changes between normal group and subclinical keratoconus group should combine with other technology to further improve subclinical keratoconic screening.Compared with normal corneas,keratoconus has a great change in biomechanics,which DA diagnosis of the highest efficiency.
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Long-term compliance with regular surveillance is important for the prevention and timely management of chronic hepatitis B (CHB). However, there are no researches focusing on the compliance of hepatitis B virus infected patients in regular surveillance so far. The purpose of our study was to investigate the outpatient compliance with long-term regular surveillance in China. Data of 3257 CHB outpatients was pooled and analyzed to assess the outpatient's compliance with the long-term regular surveillance plan. In all outpatients, the non-follow-up and the follow-up group accounted for 73.2% and 26.8%, respectively. Among the follow-up outpatient's, only 48.9% received ongoing-follow-up and 51.1% were finally lost to follow-up; the median length of visiting duration was 25 months; and the predictive 1-, 2-, 3-, 4- and 5-year ongoing follow-up rate was 72.7%, 52.5%, 42.4%, 33.8%, and 26.3%, respectively. In conclusion, our survey proved that the regular long-term surveillance on Chinese chronic HBV carrier is difficult to be fully implemented. A large proportion of outpatients do not receive routine follow-up and are at risk of treatment delay due to various social reasons.
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Long-term compliance with regular surveillance is important for the prevention and timely management of chronic hepatitis B (CHB). However, there are no researches focusing on the compliance of hepatitis B virus infected patients in regular surveillance so far. The purpose of our study was to investigate the outpatient compliance with long-term regular surveillance in China. Data of 3257 CHB outpatients was pooled and analyzed to assess the outpatient's compliance with the long-term regular surveillance plan. In all outpatients, the non-follow-up and the follow-up group accounted for 73.2% and 26.8%, respectively. Among the follow-up outpatient's, only 48.9% received ongoing-follow-up and 51.1% were finally lost to follow-up; the median length of visiting duration was 25 months; and the predictive 1-, 2-, 3-, 4- and 5-year ongoing follow-up rate was 72.7%, 52.5%, 42.4%, 33.8%, and 26.3%, respectively. In conclusion, our survey proved that the regular long-term surveillance on Chinese chronic HBV carrier is difficult to be fully implemented. A large proportion of outpatients do not receive routine follow-up and are at risk of treatment delay due to various social reasons.
Subject(s)
Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Young Adult , Carrier State , Diagnosis , Epidemiology , Therapeutics , China , Chronic Disease , Hepatitis B , Diagnosis , Epidemiology , Therapeutics , Longitudinal Studies , Patient Compliance , Population Surveillance , Methods , PrevalenceABSTRACT
A growing body of evidence suggests that p300 histone acetyltransferase plays important roles in cancer cell differentiation and proliferation. Here, we employed structure-based hierarchical virtual screening method to identify novel lead compounds of p300 histone acetyltransferase. From a screening library containing approximate 100 000 diverse druglike compounds, 33 compounds were chosen for experimental testing and one compound, 4-acetyl-2-methyl-N-morpholino-3,4-dihydro-2H-benzo[b][1, 4]thiazine-7-sulfonamide (17), showed as micromolar inhibitor. Based on its predicted binding pose, we investigated its binding characteristics by designing two series of structural modifications. The obtained structure-activity relationship results are consistent with the predicted binding model. We expect that the identified novel p300 histone acetyltransferase inhibitors will serve as starting points for further development of more potent and specific histone acetyltransferase inhibitors.
Subject(s)
Drug Design , Enzyme Inhibitors , Chemistry , Molecular Structure , Morpholines , Chemistry , Structure-Activity Relationship , Sulfonamides , Chemistry , p300-CBP Transcription Factors , ChemistryABSTRACT
Background The special pathological change of diabetic retinopathy(DR)is microvascular disorder.Angiopoietin-like protein 2(Ang-2)is a new protein associated with genesis of blood vessels.Quercetin has multiple pharmacological action,including improving the microcircularion and the permeability of blood capillary.However,the action mechanism of Ang-2 on DR was unclear.Objective The present study was to investigate the effects of quercetin on Ang-2 and its receptor Tie2 expression in retina with diabetes mellitus.Methods Sixty clean male Wistar rats were randomized into 7 groups and 10 rats for each group,and 10 rats served as blank control group.Streptozotocin of 35 mg/kg was intraperitoneally injected in 60 rats to establish the diabetic models.Quercetins encapsulated by liposome with the doses of 50,150 and 250 mg/(kg · d)(3-5 ml)were used to gavage in different groups of models for 12 weeks,and normal saline solution and calcium dobesilate were used at the same fashion as the negative control group and positive control group,respectively.Twelve weeks later,the animals were sacrificed and retinas were isolated.Expressions of Ang-2 protein and Tie2 mRNA in retinas were detected by ELISA and RT-PCR,respectively.The usage and rearing of the animals complied with the Regulations for the Administration of Affair Concerning Experimental Animals by State Sciences and Technology Commission.Results ELISA showed that the A450 of Ang-2 in 150 and 250 mg/(kg · d)quercetin groups was 0.796±0.057 and 0.842±0.043 respectively and was lower than that negative group(1.012±0.046),showing statistically significant differences(q =2.95,2.698,P＜0.05).RT-PCR assay showed that expression of Tie2 mRNA(Tie2 mRNA/GAPDH mRNA)in retinas was 0.712±0.092 and 0.821±0.087,presenting statistically significant differences in comparison with negative group(1.182±0.098)(q =3.497,2.852,P＜0.05).The expression levels of Ang-2 and Tie2 mRNA in retina were lowest in 150 mg/(kg · d)quercetin group.Conclusions Quercetin can improve the retinal microcirculation by downregulating the expressions of Ang-2 and its receptor in early period of diabetic rats.
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<p><b>OBJECTIVE</b>To assess the effects of LiCl on prostate cancer growth and to explore the underlying mechanisms.</p><p><b>METHODS</b>Effects of LiCl on cell growth in vitro and in vivo were determined by cell counting and xenografts of prostate cancer cells. Alterations in cell proliferation and the expression of DNA replication-related protein were determined by MTT assay, BrdU incorporation and Western blot.</p><p><b>RESULTS</b>Compared to PBS control group, the number of prostate cancer cells (PC-3) were lower treated with 10 mmol/L LiCl, the number was 1.9×10(5), 4.8×10(5) and the difference was significant (P < 0.05). The inhibition rate of cellular proliferation were 50%, 95% and 98%, respectively, in LiCl group, NaCl and KCl control group, the difference was significant (P < 0.05). The A-Value of BrdU incorporation was 1.5, 1.3 treated with 10 mmol/L, 30 mmol/L LiCl, while the A-value of BrdU incorporation was 4 in PBS control group, the difference was significant (P < 0.05). On the protein level, LiCl downregulates expression of cdc 6, cyclins A and cyclins E, and cdc 25C, and upregulates expression of the CDK inhibitor p21(CIP1). The mean volume and weight of xenograft tumor were 50 mm(3) and 296 mg after LiCl intraperitoneal injection, But PBS control group were 180 mm(3) and 957 mg, the difference was significant (P < 0.05).</p><p><b>CONCLUSION</b>LiCl disrupts DNA replication and suppresses tumor growth of prostate cancer cells in vitro and in vivo.</p>
Subject(s)
Animals , Humans , Male , Mice , Antineoplastic Agents , Pharmacology , Cell Cycle Proteins , Metabolism , Cell Line, Tumor , Cell Proliferation , Cyclin A , Metabolism , Cyclin E , Metabolism , Cyclin-Dependent Kinase Inhibitor p21 , Metabolism , DNA Replication , Lithium Chloride , Pharmacology , Mice, Inbred BALB C , Mice, Nude , Neoplasm Transplantation , Nuclear Proteins , Metabolism , Prostatic Neoplasms , Metabolism , Pathology , Tumor Burden , cdc25 Phosphatases , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the properties of HBsAb in occult hepatitis B virus infection and its affinity to different serotypes of hepatitis B virus surface antigen (HBsAg).</p><p><b>METHODS</b>Long-term follow-up was conducted in 2 HBsAb positive patients with occult hepatitis B virus infection. HBsAg was detected using multiple diagnostic kits and the HBsAb subtype was determined by performing neutralization experiments with different serotypes of HBsAg. The viral S gene was PCR-amplified and mutation analysis was conducted. Plasmids expressing HBsAgs were constructed by inserting these PCR products into an eukaryotic expression vector and were then transfected into HepG2 cells. The cell culture supernatant and cellular extracts were detected for HBsAg respectively. Neutralization experiments were carried out in the cell culture supernatant from HBsAg plasmids transfected HepG2 cells and serum samples from these patients and others who had been confirmed to be positive for HBsAb.</p><p><b>RESULTS</b>Multiple tests using various diagnostic kits showed that the 2 patients were negative for HBsAg and the three different serotypes of HBsAg (adr, adw, ay) could neutralize 82.1%-100% of HBsAb existed in the 2 patients. Sequence analysis of S gene cloned from these patients revealed that the homology to reference strain were 95.13%-97.79% and 92.04%-95.58% respectively at the nucleotide and amino acid levels. Quantitation of HBsAg showed that the expression levels of HBsAg from the two patients were 41.1% and 22.6% respectively of that of control HBsAg in cell culture supernatant and 48.1% and 59.3% respectively in cellular extract, and the supernatant/cell lysate ratios were 0.85 and 0.38 respectively. In neutralization experiments, HBsAg could be totally absorbed by control serum, whereas could only be partially neutralized by HBsAbs from the two patients (F = 353.6 and 645.2, P is less than 0.01).</p><p><b>CONCLUSION</b>Both the antigenicity and the ability of HBsAg secreted outside of the cells are decreased in these HBsAb-positive patients with occult HBV infection. The HBsAbs are mainly specific for common epitopes among different serotypes of HBsAg and are probably different as compared with those produced by vaccine inoculation.</p>
Subject(s)
Adult , Humans , Male , Middle Aged , Hepatitis B , Blood , Virology , Hepatitis B Antibodies , Blood , Hepatitis B Surface Antigens , Blood , Hepatitis B virus , Serologic TestsABSTRACT
<p><b>OBJECTIVE</b>To determine the relationship between IgG antibody against the C-terminal region of the preS1 protein of hepatitis B virus and the early response to interferon therapy in chronic hepatitis B.</p><p><b>METHODS</b>69 patients with chronic hepatitis B virus (genotype B) infection were recruited in this study. 42 patients were treated with interferon-a-1b or a-2b, and 27 patients were treated with PEG interferon (a-2a). Peptide mimicking the C-terminal region of the preS1 protein (94-117aa) of genotype B HBV were synthesised, and the IgG antibody against this peptide was measured by ELISA, and the early response to IFN-alpha therapy was judged by the effect on the viral kinetics, transaminase and the status of HBeAg at 12th week after the treatment.</p><p><b>RESULTS</b>21 patients were positive for anti-preS1 antibody, and 48 patients were negative for anti-preS1 antibody. After 12 weeks of the treatment, the average decrease in viral levels was 3.37log10 copies/ml and 0.33log10 copies/ml in anti-PreS1 positive patients and anti-preS1 negative patients, respectively, the difference between the two groups was significant (Z = -3.658, P = 0.000); the average decrease in ALT levels was 92 U/L and 30.5 U/L in these two groups, respectively (Z = -2.132, P = 0.033). The rate of hepatitis B e antigen (HBeAg) loss was 41.2% (7/17) and the rate of anti-HBe seroconversion was 5.9% (1/17) in anti-PreS1 positive group, however, the rate of hepatitis B e antigen loss was only 12.8% (5/39), and none of the patients in anti-PreS1 negative group showed anti-HBe seroconversion, the difference between the two groups was significant (Z = -5.110, P = 0.000). The rates of response were 71.4% (15/21) and 16.7% (8/48), respectively, in anti-PreS1 positive group and anti-PreS1 negative group. The rates of complete response were 23.8% (5/21) and 6.25% (3/48), respectively, in these two groups. The positive predictive value (PPV) of anti-C-terminal region of preS1 (94-117aa) antibody in predicting early response was 71.6% and the negative predictive value (NPV) was 83.3%. CONCLUCIONS: Detection of anti-C-terminal region of preS1 (94-117aa) antibody may help to improve the efficacy of INF-alpha therapy for chronic hepatitis B (CHB).</p>
Subject(s)
Adolescent , Adult , Female , Humans , Male , Young Adult , Alanine Transaminase , Blood , Antiviral Agents , Therapeutic Uses , DNA, Viral , Blood , Hepatitis B Surface Antigens , Blood , Allergy and Immunology , Hepatitis B e Antigens , Blood , Hepatitis B virus , Genetics , Hepatitis B, Chronic , Blood , Drug Therapy , Allergy and Immunology , Immunoglobulin G , Blood , Allergy and Immunology , Interferon-alpha , Therapeutic Uses , Polymerase Chain Reaction , Methods , Predictive Value of Tests , Prospective Studies , Protein Precursors , Blood , Allergy and Immunology , Viral LoadABSTRACT
<p><b>OBJECTIVES</b>To investigate S gene mutations in HBsAg/HBsAb double positive chronic hepatitis B patients.</p><p><b>METHODS</b>HBV S gene from 8 patients (Group A) with HBsAg (+)/HBsAb (+) and 9 patients (Group B) with HBsAg (+)/HBsAb (-)was amplified by polymerase chain reaction (PCR) and sequencing. Both the distribution of genotype and serotype and the rate of MHR region were compared by Fisher's exact test. The mutation rate of both the DNA level and amino acid level was compared by t test.</p><p><b>RESULTS</b>No significant difference in distribution of genotypes was found between the two groups (P=0.153). In group A, 2 were genotype B, 6 were genotype C; In group B, 6 were genotype B, 3 were genotype C. No significant difference in distribution of serotypes was found between the two groups, either (P=0.218). In group A, 2 were adw, 5 were adr, 1 was ayr; In group B, 6 were adw, 3 were adr. The mutation rate of Pre-S1 region at both the DNA level (2.29% vs 1.80%, t=2.66, P more than 0.05) and the amino acid level (2.66% vs 1.59%, t=1.39, P>0.05) was not significantly different between these two groups; the mutation rate of Pre-S2 region in group A patients was significantly higher than that in group B at the DNA level (1.74% vs 0.91%, t=4.68, P<0.01), but not higher at the amino acid level (3.18% vs 2.05%, t=1.85, P>0.05), the mutation rate of S region in group A patients was significantly higher than that in group B at both the DNA level (2.13% vs 0.81%, t=6.00, P<0.01) and the amino acid level (4.37% vs 1.52%, t=5.32, P<0.01). Amino acid substitutions were found both within and beyond the MHR region. The rate of "a" determinant mutations in these two groups was also found to be significantly different (P<0.05).</p><p><b>CONCLUSION</b>Higher HBV S gene mutation rate exists in HBsAg/HBsAb double positive patients than that in HBsAg (+)/HBsAb (-) patients.</p>
Subject(s)
Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , Base Sequence , DNA, Viral , Blood , Genetics , Genes, Viral , Genetic Variation , Genotype , Hepatitis B Antibodies , Blood , Hepatitis B Surface Antigens , Blood , Allergy and Immunology , Hepatitis B virus , Genetics , Allergy and Immunology , Hepatitis B, Chronic , Allergy and Immunology , Virology , Polymerase Chain Reaction , Sequence Analysis, DNAABSTRACT
<p><b>OBJECTIVE</b>To establish the reference sequences of genotype B and C of hepatitis B virus in China.</p><p><b>METHODS</b>Genome sequences of Hepatitis B virus isolated from different area in China were retrieved from GenBank. These genome sequences were alignmented and the most common sequences were regarded as reference sequences. The amino acid sequences were also alignmented.</p><p><b>RESULTS</b>The homology was 99.32% between Bc genome and Ba genome, and it was 95.52% between Bc genome and Bj genome. The S gene sequence homology was 99.71% between Bc and Ba, and it was 98.68% between Bc and Bj. The homology was 98.44% between Cc genome and C genome, and it was 93.97% between Cc genome and Caus genome. The S gene sequence homology was 99.27% between Cc and C, and it was 95.01% between Cc and Caus. There was significant difference at the sites of 1762, 1764, 1858 between genotype B and genotype C (P < 0.05). The amino acid sequences were also different between genotype B and genotype C.</p><p><b>CONCLUSION</b>Bc and Cc sequences of Hepatitis B virus can be regarded as the reference sequences of genotype B and C.</p>
Subject(s)
Humans , Amino Acid Sequence , Base Sequence , China , DNA Primers , Genetic Variation , Genome, Viral , Genotype , Hepatitis B virus , Classification , Genetics , Hepatitis B, Chronic , Virology , Molecular Sequence Data , Reference Standards , Sequence Alignment , Sequence HomologyABSTRACT
<p><b>OBJECTIVE</b>To construct a vector that is competent to support the replication of hepatitis B virus (HBV) of genotype B.</p><p><b>METHODS</b>The HBV genome of genotype B was amplified by PCR and ligated into pBlueskript II KS(+) vector, the resulting plasmid was verified by enzyme digestion and DNA sequencing. After transfection of this plasmid into Huh7 cells, the HBsAg and HBeAg antigens in culture medium were quantified by ELISA, the transcripts and replication intermediates of HBV were detected by northern blot and southern blot respectively. On the other hand, the plasmid was hydrodynamically injected into BALB/cJ mice via tail vein. Then the HBV DNA in serum was quantified by real-time PCR, and HBcAg expression in liver tissue was detected by immunohistochemistry.</p><p><b>RESULTS</b>After transfection of the plasmid into Huh7 cells, the HBsAg and HBeAg antigens were detected in the culture medium, the transcripts and replication intermediates of HBV were detected in the cells. High titer of HBV DNA was detected in the serum of hydrodynamic-injected mice. Immunostaining indicated that HBcAg was expressed in hepatocytes of injected mice.</p><p><b>CONCLUSION</b>This construct is competent to support the replication of hepatitis B virus of genotype B.</p>
Subject(s)
Animals , Mice , Cell Line, Tumor , Cloning, Molecular , Disease Models, Animal , Genetic Vectors , Genotype , Hepatitis B , Metabolism , Virology , Hepatitis B Antigens , Genetics , Metabolism , Hepatitis B virus , Genetics , Metabolism , Liver , Metabolism , Virology , Mice, Inbred BALB C , Plasmids , Polymerase Chain Reaction , Transfection , Virus ReplicationABSTRACT
<p><b>OBJECTIVE</b>To develop a standard duck hepatitis B virus (DHBV) animal model using a local Hubei species of duck, Ma Ya, and use it as an in vivo experimental system to study antiviral strategies against hepatitis B.</p><p><b>METHODS</b>Two-day-old Ma Ya ducklings were experimentally infected via intraperitoneal injection with the DHBV inocula which was collected from the transfected culture supernatant of 1.5-fold-overlength genome recombinant plasmid. Blood samples were taken twice or thrice a week during post-inoculation for 50 days. Viremia was quantified by serum real-time PCR to show the peak. Antiviral treatment of the DHBV-infected ducklings was started 3 d post-inoculation. The animals received oral administration of lamivudine (3TC) at a dose of 25 mg/kg/d for 5 d, followed by a maintenance therapy thrice weekly for 3 more weeks. Serum was quantified to show the viremia peak and liver biopsy specimens were analysed by Southern blotting and in-situ hybridization at the end of antiviral drug treatment.</p><p><b>RESULTS</b>The experimental infection rate of 2-day-old ducklings was 87.5%. Viremia started to be detectable on day 7 and reached a peak on day 11 post-inoculation, followed by a decrease and fluctuations. Four weeks of oral administration of 3TC led to a significant decrease in viremia peak during. This effect was not sustained, as a rebound in viremia was observed after drug withdrawal. Similarly, the analysis of liver biopsies at the end of 3TC treatment showed a marked decrease in DHBV DNA. However, after drug withdrawal a rebound of intrahepatic DHBV DNA was observed in duck livers.</p><p><b>CONCLUSION</b>The Hubei duck model with experimental DHBV infection of transfected supernatant is more suitable for the hepadnavirus biologic research due to its stability and practicability.</p>
Subject(s)
Animals , Animals, Newborn , DNA, Viral , Genetics , Metabolism , Disease Models, Animal , Ducks , Hepadnaviridae Infections , Blood , Drug Therapy , Virology , Hepatitis B Virus, Duck , Genetics , Hepatitis, Viral, Animal , Blood , Drug Therapy , Virology , Lamivudine , Pharmacology , Liver , Pathology , Virology , Reverse Transcriptase Inhibitors , Pharmacology , Viremia , BloodABSTRACT
<p><b>OBJECTIVE</b>To search for and verify some common B cell epitopes in the core proteins of woodchuck hepatitis virus and human hepatitis B virus.</p><p><b>METHODS</b>Monoclonal antibodies against both core proteins of woodchuck hepatitis virus (WHV) and human hepatitis B virus (HBV) were prepared by inoculating Balb/c mice with denatured recombination WHV and HBV core proteins. ELISA and immunoblotting assays for WHcAg and HBcAg were carried out by using these antibodies. Immunohistochemistry was carried out with liver tissue sections of both WHV-infected woodchucks and chronic HBV-infected patients. The epitopes were mapped with the mouse mAbs (6D1 and 1H4) by using a panel of 24 16mer overlapping peptides covering the entire WHcAg. The amino acid sequences of WHcAg and HBcAg were compared.</p><p><b>RESULTS</b>Cross-reactions were observed between mAbs (6D1 and 1H4) and WHcAg and between Mabs and HBcAg/HBcAg in ELISA and immunoblotting assay. Liver tissue sections of both WHV-infected woodchucks and chronic HBV-infected patients could be stained specifically by mAbs. The epitopes were mapped at aa1-8 (6D1) and aa125-140 (1H4) of the core proteins of both WHV and HBV by using ELISA assay. WHcAg and HBcAg share similar amino acids sequences at aa1-8 and aa125-140 respectively.</p><p><b>CONCLUSION</b>The core proteins of woodchuck hepatitis virus and human hepatitis B virus share common linear B cell epitopes which span aa1-8 and aa125-140 respectively.</p>
Subject(s)
Animals , Humans , Mice , Antibodies, Monoclonal , B-Lymphocytes , Allergy and Immunology , Cell Line, Tumor , Cross Reactions , Epitopes, B-Lymphocyte , Allergy and Immunology , Hepatitis B Core Antigens , Allergy and Immunology , Hepatitis B Virus, Woodchuck , Genetics , Allergy and Immunology , Hepatitis B virus , Genetics , Allergy and Immunology , Marmota , Viral Core Proteins , Allergy and ImmunologyABSTRACT
<p><b>OBJECTIVES</b>To construct a prokaryotic plasmid expressing truncated human cervical cancer oncogene (HCCR-1(167-360)), to express and purify the recombinant protein, and to develop the polyclonal antibody against HCCR.</p><p><b>METHODS</b>HCCR-1(167-360) was amplified by RT-PCR from HepG2 cells and cloned into vector pRSET-B, then expressed in E.coli BL21(DE3) pLysS, which was induced by IPTG. The recombinant protein was purified using Ni-NTA spin column and acrylamide gel electrophoresis. A polyclonal antibody was developed by immunizing BALB/c mice with the purified recombinant protein, and their sensitivity and specificity were tested using enzyme-linked immunosorbent assay, immunohistochemical staining and Western blot analysis.</p><p><b>RESULTS</b>Recombinant plasmid expressing truncated HCCR-1167-360 was constructed. A protein of 2.70 x 10(4) was successfully expressed and purified. High titer polyclonal antibody with a high specificity was obtained by immunizing BALB/c mice with the purified recombinant protein.</p><p><b>CONCLUSIONS</b>The truncated recombinant HCCR-1(167-360) developed in this study is highly purified and shows strong antigenecity; the polyclonal antibody against this HCCR protein was generated by regular immunization method, showing both high sensitivity and specificity. The protein and the antibody can be used for further clinical examination and research of HCCR.</p>
Subject(s)
Animals , Humans , Mice , Antibodies, Monoclonal , Genetics , Biomarkers, Tumor , Genetics , Carcinoma, Hepatocellular , Pathology , Cloning, Molecular , Escherichia coli , Metabolism , Liver Neoplasms , Pathology , Mice, Inbred BALB C , Prokaryotic Cells , Metabolism , Proto-Oncogene Proteins , Genetics , Allergy and Immunology , Reverse Transcriptase Polymerase Chain Reaction , Tumor Cells, CulturedABSTRACT
Objective:To study the change of immunological response and cell proliferation in breast tissues augmented by polyacrylamide hydrogel injection(PHI).Methods:The expression of CD68,CD25 and PCNA in 20 breast tissues with indurations,12 without indurations after breast augmentation by PHI,and 10 normal breast tissues was examined by immunohistochemistry P-V6000; analysis was also done by H-E staining.Results:Hyperplasia of fibrous tissue and infiltration of inflammatory cells and macrophages were found in the breast and adjacent tissues 3-8 years after PHI.Positive cells of CD68,CD25 and PCNA hardly existed in the normal tissues,but the breast tissues around the polyacrylamide hydrogel had many positive cells of CD68 and PCNA,especially in cases with indurations;there were significant differences between the 3 groups(P