黏膜佐剂大肠埃希菌不耐热肠毒素B亚单位优化的淋病奈瑟菌孔洞蛋白B核酸疫苗的构建及其诱导小鼠的免疫应答

Objective To investigate the specific humoral immune response and cellular immune response induced by DNA vaccine with Neisseria gonorrhoeae porin B (PorB) fused with B subunit of Escherichia coli heat-labile enterotoxin B (LTB) in mice. Methods Target genes of porB, ltB and ltB-porB were amplified by polymerase chain reaction (PCR) and cloned into eukaryotic vector pcDNA3.1(-). The recombinants were identified by PCR, enzyme digestion and DNA sequencing.The vectors were transfected into Hela cells, and expressed proteins were checked by cytoimmunofluorescence. Female BALB/c mice were intranasally immunized with recombination vectors. The humoral immune response and cellular immune response were detected by enzyme linked immunosorbent assay (ELISA) and methyl thiazolyl tetrazolium (MTT) colorimetric assay. The expressions of recombination vectors in intranasal mucosal tissues of the immunized mice were detected by immunohistochemistry. The means between groups were compared by analysis of variance. Results All the three recombinants were expressed in Hela cells and intranasal mucosal tissues. The PorB specific IgG in serum and sIgA in vaginal secretions in DNA vaccine immunized mice were significantly higher than those in controls (P＜0.01 ; P＜0.05). Moreover, the sIgA level in pcDNA3.1 (-)/ltB-porB group was higher than that in peDNA3, 1(-)/porB group (P＝0. 002). The levels of interferon-gamma (IFN-γ) and interleukin-4 (IL-4) in the supernatants and stimulation index (SI) of spleen lymphocyte culture in pcDNA3, 1(-)/porB group were (170.04±23.89) pg/mL, (114.68±14.27) pg/mL and 1. 68±0.19, respectively; and those in pcDNA3, 1(-)/ltB-porB group were (161.42±27.50) pg/mL, (124.16±19.04) pg/mL and 1.73±0.28, respectively; which were both higher than those in pcDNA3.1(-)/ phosphate buffered saliae (PBS) group (P＜0. 01; P＜0.05) and pcDNA3.1 (-)/ltB group (all P＜0.05), while there was no significant difference between pcDNA3.1 (-)/ltB-porB group and pcDNA3. 1 (-)/porB group (0. 998, 0. 696, 0. 994; all P＞0.05). Conclusions The constructed DNA vaccines are all successfully expressed in Hela cells and murine intranasal mucosal tissues. The mucosal immunization of the vaccines [pcDNA3. 1 (- )/porB and pcDNA3.1 ( -)/ltBporB] could induce humoral immune response and cellular immune response, especially mucosal immune response. It is confirmed that mucosal adjuvant LTB could promote PorB to induce higher level of mucosal immune response in mice.     

黏膜佐剂大肠埃希菌不耐热肠毒素B亚单位优化的淋病奈瑟菌孔洞蛋白B核酸疫苗的构建及其诱导小鼠的免疫应答

目的 分析黏膜佐剂大肠埃希菌不耐热肠毒素B亚单位(LTB)辅佐的淋病奈瑟菌孔洞蛋白B(PorB)核酸疫苗诱导小鼠的免疫应答水平.方法 PCR法扩增目的 基因porB、ltB、ltB-porB,分别构建3种相应的peDNA3.1(-)真核重组表达载体,经PCR、双酶切及基因测序鉴定后转染Hela细胞,用细胞免疫荧光法鉴定质粒的蛋白表达.将核酸疫苗经鼻饲免疫雌性BALB/c小鼠,检测体液免疫及细胞免疫应答水平,同时用免疫组织化学法检测porB、ltB、ltB-porB基因在小鼠鼻黏膜内的表达.组间均数比较采用方差分析.结果 构建的真核重组质粒均能在Hela细胞内及小鼠鼻黏膜组织内表达.核酸疫苗免疫组小鼠的生殖道灌洗液PorB特异性sIgA及血清porB特异性IgG水平明显高于对照组(P＜0.01;P＜0.05),且ltB-porB融合基因组特异性sIgA明显高于porB组(P＜0.05);pcDNA3.1(-)/porB组小鼠脾淋巴细胞培养上清液中IFN-γ、IL-4和脾淋巴细胞刺激指数(SI)分别为(170.04±23.89)pg/mL、(114.68±14.27)pg/mL和1.68±0.19,pcDNA3.1(-)/ltB-porB组分别为(161.42±27.50)pg/mL、(124.16±19.04)pg/mL和1.73±0.28,均高于对照组pcDNA3.1(-)和PBS,差异均有统计学意义(P＜0.01;P＜0.05),高于对照组pcDNA3.1(-)/ltB,差异均有统计学意义(均P＜0.05),但两核酸疫苗免疫组间差异无统计学意义(均P＞0.05).结论 所构建的核酸疫苗分别在Hela细胞及小鼠鼻黏膜组织内获得了表达,经黏膜途径免疫能诱导小鼠产生较高水平的特异性体液免疫和细胞免疫应答,尤其是黏膜免疫应答;证实黏膜佐剂LTB可辅佐PorB诱导小鼠产生高水平的生殖道黏膜免疫.

Abstract：

Objective To investigate the specific humoral immune response and cellular immune response induced by DNA vaccine with Neisseria gonorrhoeae porin B (PorB) fused with B subunit of Escherichia coli heat-labile enterotoxin B (LTB) in mice. Methods Target genes of porB, ltB and ltB-porB were amplified by polymerase chain reaction (PCR) and cloned into eukaryotic vector pcDNA3.1(-). The recombinants were identified by PCR, enzyme digestion and DNA sequencing.The vectors were transfected into Hela cells, and expressed proteins were checked by cytoimmunofluorescence. Female BALB/c mice were intranasally immunized with recombination vectors. The humoral immune response and cellular immune response were detected by enzyme linked immunosorbent assay (ELISA) and methyl thiazolyl tetrazolium (MTT) colorimetric assay. The expressions of recombination vectors in intranasal mucosal tissues of the immunized mice were detected by immunohistochemistry. The means between groups were compared by analysis of variance. Results All the three recombinants were expressed in Hela cells and intranasal mucosal tissues. The PorB specific IgG in serum and sIgA in vaginal secretions in DNA vaccine immunized mice were significantly higher than those in controls (P＜0.01 ; P＜0.05). Moreover, the sIgA level in pcDNA3.1 (-)/ltB-porB group was higher than that in peDNA3, 1(-)/porB group (P＝0. 002). The levels of interferon-gamma (IFN-γ) and interleukin-4 (IL-4) in the supernatants and stimulation index (SI) of spleen lymphocyte culture in pcDNA3, 1(-)/porB group were (170.04±23.89) pg/mL, (114.68±14.27) pg/mL and 1. 68±0.19, respectively; and those in pcDNA3, 1(-)/ltB-porB group were (161.42±27.50) pg/mL, (124.16±19.04) pg/mL and 1.73±0.28, respectively; which were both higher than those in pcDNA3.1(-)/ phosphate buffered saliae (PBS) group (P＜0. 01; P＜0.05) and pcDNA3.1 (-)/ltB group (all P＜0.05), while there was no significant difference between pcDNA3.1 (-)/ltB-porB group and pcDNA3. 1 (-)/porB group (0. 998, 0. 696, 0. 994; all P＞0.05). Conclusions The constructed DNA vaccines are all successfully expressed in Hela cells and murine intranasal mucosal tissues. The mucosal immunization of the vaccines [pcDNA3. 1 (- )/porB and pcDNA3.1 ( -)/ltBporB] could induce humoral immune response and cellular immune response, especially mucosal immune response. It is confirmed that mucosal adjuvant LTB could promote PorB to induce higher level of mucosal immune response in mice.     

Enhancement of humoral immune responses to HBsAg by heat shock protein gp96 and its N-terminal fragment in mice

AIM: Most studies on the immune effect of gp96 were focused on its enhancement of CTLs. It is interesting to know whether gp96 could influence the humoral immune response, and whether the recombinant N-terminal fragment of gp96 could substitute native gp96 to stimulate the immune system. METHODS: gp96 isolated from livers of normal mice and its N-terminal fragment (amino acid 22-355) expressed in E coli were used for immunization of BALb/c mice. Eight groups of mice received one of the following regiments subcutaneously in 100 μL phosphate buffered saline (PBS) at an interval of 3 wk. Group 1: PBS only; group 2: gp96 only; group 3: N-terminal fragment only; group 4: HBsAg only; group 5: HBsAg+gp96; group 6: HBsAg+N-terminal fragment; group 7: HBsAg+incomplete Freud's adjuvant; group 8: HBsAg+N-terminal fragment (95℃ heated for 30 min). Serum anti-HBsAg antibody levels were assayed by ELISA. CTL responses in splenocytes were analyzed by ELISPOT after the last vaccination. RESULTS: The average titer of serum anti-HBsAg antibody in the mice immunized with HBsAg together with gp96 or its N-terminal fragment were much higher than those immunized with HBsAg alone detected by ELISA. The cellular immune response of the mice immunized with HBsAg together with gp96 or its N-terminal fragment was not different with those immunized with HBsAg alone measured by ELISPOT assay. CONCLUSION: gp96 or its N-terminal fragment greatly improved humoral immune response induced by HBsAg, but failed to enhance the CTL response, which demonstrated the potential of using gp96 or its N-terminal fragment as a possible adjuvant to augment humoral immune response against HBV infection.     

Hepatitis E virus chimeric DNA vaccine elicits immunologic response in mice

AIM: To construct the plasmid pcHEV23 containing fragments of HEV ORF2 and ORF3 chimeric gene and to assess its ability to elicit specific immunologic response in mice. METHODS: The gene encoding the structural protein of HEV ORF2 fragment and full-length ORF3 was amplified by PCR. The PCR products were cloned into an eucaryotic expression plasmid pcDNAS. The resulting plasmid pcHEV23 was used as a DNA vaccine to inoculate BALB/c mice intramuscularly thrice at a dose of 100 or 200 μg. Mice injected with empty pcDNA3 DNA or saline served as control and then specific immune responses in the mice were detected. RESULTS: After 2-3 times of inoculation, all mice injected with pcHEV23 had anti-HEV IgG seroconversion and specific T lymphocyte proliferation. The lymphocyte stimulation index in the group immunized with pcHEV23 (3.1±0.49) was higher than that in the control group (0.787±0.12, P<0.01). None in the control group had a detectable level of anti-HEV IgG. CONCLUSION: DNA vaccine containing HEV ORF2 and ORF3 chimeric gene can successfully induce specific humoral and cellular immune response in mice.     

Induction of hepatitis C virus-specific cytotoxic T and B cell responses by dendritic cells expressing a modified antigen targeting receptor

AIM: To find a novel antigen (Ag) presentation strategy to improve the immune responses induced by dendritic cell (DC) vaccine expressing hepatitis C virus (HCV) core antigen (pcDNA3HCV C-Fc) in Balb/c mice (H-2d). METHODS: pcDNA3HCV C-Fc plasmid and eukaryotic expression vector pcDNA3 were injected into mice sc. Immune responses to pcDNA3HCV C-Fc were studied. Meanwhile the effect of pcDNA3HCV C-Fc on anti-translated subcutaneous tumor of SP2/0 cells stably expressing HCV C Ag (SP2/0-HCV C-FC) was also studied. Anti-HCV C in serum was detected by enzyme-linked immunoadsordent assay (ELISA) and HCV specific cytotoxic T lymphocyte (CTL) activity was measured by LDH release assay. After 3 wk of DNA immunization, the cells of SP2/0-HCV C-FC were inoculated into mice subcutaneously and tumor growth was measured every 5 d. The survival rate and living time of mice were also calculated. RESULTS: After 4 wk of DC immunization, the A450 nm values of sera in mice immunized with pcDNA3HCV C-Fc-DC and pcDNA3-DC were 0.56±0.17 and 0.12±0.03 respectively. The antibody titres in mice codeliveried with pcDNA3HCV C-Fc with DC were significantly higher than those of mice injected with pcDNA3-DC. The HCV specific CTL activities in mice coinjected with DC and pcDNA3HCV C-Fc or empty expression vectors were(73.2±3.1) % and (24.4±8.8) % , which were significantly higher than those of mice injected with water. The DC vaccine could evidently inhibit tumor growth, prolong the survival time of mice and improve the survival rate of mice and these effects could be improved by HCV C-Fc (pcDNA3HCV C-Fc) gene codelivered. CONCLUSION: DC vaccine has a strong antigenicity in humoral and cellular immunities, which can be promoted by transduced pcDNA3HCV C-Fc expressing HCV C or Fc. Thus, pcDNA3HCV C-Fc-transduced DCs may be a promising candidate for a CTL-based vaccine against HCV.     

Antitumor immunopreventive effect in mice induced by DNA vaccine encoding a fusion protein of α-fetoprotein and CTLA4

AIM:To develop a tumor DNA vaccine encoding a fusion protein of murine AFP and CTLA4,and to study its ability to induce specific CTL response and its protective effect against AFP-producing tumor.METHODS:Murine α-fetoprotein (mAFP) gene was cloned from total RNA of Hepal-6 cells by RT-PCR.A DNA vaccine was constructed by fusion murine α-fetoprotein gene and extramembrane domain of murine CTLA4 gene.The DNA vaccine was identified by restriction enzyme analysis,sequencing and expression.EL-4 (mAFP) was developed by stable transfection of EL-4 cells with pmAFP.The frequency of cells producing IFN-γ in splenocytes harvested from the immunized mice was measured by ELISPOT.Mice immunized with DNA vaccine were inoculated with EL-4(mAFP) cells in back to observe the protective effect of immunization on tumor. On the other hand,blood samples were collected from the immunized mice to check the functions of liver and kidney.RESULTS:1.8kb mAFP cDNA was cloned from total RNA of Hepal-6 cells by RT-PCR.The DNA vaccine encoding a fusion protein of mAFP-CTLA4 was constructed and confirmed by restriction enzyme analysis,sequencing and expression.The expression of mAFP mRNA in EL-4 (mAFP) was confirmed by RT-PCR.The ELISPOT results showedthat the number of IFN-γ-producing cells in pmAFP-CTLA4 group was significantly higher than that in pmAFP,pcDNA3.1 and PBS group.The tumor volume in pmAFP-CTLA4 group was significantly smaller than that in pmAFP,pcDNA3.1 and PBS group, respectively. The hepatic and kidney functions in each group were not altered.CONCLUSION: AFP-CTLA4 DNA vaccine can stimulate potent specific CTL responses and has distinctive antitumor effect on AFP-producing tumor.The vaccine has no impact on the function of mouse liver and kidney.     

Establishment of mice model with human viral hepatitis B

AIM:To establish a mice model of hepatitis B by using HBV-transgenic mice, and to transfer HBV-specific cytotoxic T lymphoo/tes (CTL) induced from syngeneic BALB/c mice immunized by a eukaryotic expression vector containing HBV complete genome DNA.METHODS: HBV DNA was obtained from digested pBR3222HBV and ligated with the vector pcDNA3. Recombinant pcDNA3-HBV was identified by restriction endonuclease assay and transfected into human hepatoma cell line HepG2 with lipofectin. ELISA was used to detect the expression of HBsAg in culture supernatant, and RT-PCR to determine the existence of HBV PreS1 mRNA.BALB/c mice were immunized with pcDNA3-HBV or pcDNA3 by intramuscular injection.ELISA was used to detect the expression of HBsAb in serum. MTT assay was used to measure non-specific or specific proliferation ability and specific killing activity of spleen lymphocytes. Lymphocytes from immunized mice were transferred into HBV-transgenic mice (2.5&#215;10^7 per mouse).Forty-eight hours later,the level of serum protein and transaminase was detected with biochemical method,liver and kidney were sectioned and stained by HE to observe the pathological changes.RESULTS: By enzyme digestion with Eco RI, Xho I and Hind Ⅲ,the recombinant pcDNA3-HBV was verified to contain a single copy of HBV genome,which was inserted in the positive direction.HepG2 cells transfected with the recombinant could stably express PreS1 mRNA and HBsAg.After immunized by pcDNA3-HBV for 4 weeks,HBsAb was detected in the serum of BALB/c mice. The potential of spleen lymphoo/tes for both non-specific and specific proliferation and the specific killing activity against target cells were enhanced. The transgenic mice in model group had no significant changes in the level of serum protein but had an obvious increase of ALT and AST. The liver had obvious pathological changes, while the kidney had no evident damage.CONCLUSION:A eukaryotic expression vector pcDNA3-HBV containing HBV complete genome is constructed successfully. HepG2 cells transfected with the recombinant can express PreS1 mRNA and HBsAg stably.Specific cellular immune response can be induced in mice immunized by pcDNA3-HBV.A mice model of acute hepatitis with HBV has been established.     

Antitumor immunopreventive effect in mice induced by DNA vaccine encoding a fusion protein of α-fetoprotein and CTLA4

AIM:To develop a tumor DNA vaccine encoding a fusionprotein of murine AFP and CTLA4,and to study its ability toinduce specific CTL response and its protective effect againstAFP-producing tumor.METHODS:Murine a-fetoprotein (mAFP) gene was clonedfrom total RNA of Hepa1-6 cells by RT-PCR.A DNA vaccinewas constructed by fusion murine α-fetoprotein gene andextramembrane domain of murine CTLA4 gene.The DNAvaccine was identified by restriction enzyme analysis,sequencing and expression.EL-4 (mAFP) was developedby stable transfection of EL-4 cells with pmAFR Thefrequency of cells producing IFN-γ in splenocytes harvestedfrom the immunized mice was measured by ELISPOT.Miceimmunized with DNA vaccine were inoculated with EL-4(mAFP) cells in back to observe the protective effect ofimmunization on tumor.On the other hand,blood sampleswere collected from the immunized mice to check thefunctions of liver and kidney.RESULTS:1.8 kb mAFP cDNA was cloned from total RNAof Hepa1-6 cells by RT-PCR.The DNA vaccine encoding afusion protein of mAFP-CTLA4 was constructed andconfirmed by restriction enzyme analysis,sequencing andexpression.The expression of mAFP mRNA in EL-4 (mAFP)was confirmed by RT-PCR.The ELISPOT results showedthat the number of IFN-γ-producing cells in pmAFP-CTLA4group was significantly higher than that in pmAFP,pcDNA3.1and PBS group.The tumor volume in pmAFP-CTLA4 groupwas significantly smaller than that in pmAFP,pcDNA3.1 andPBS group,respectively.The hepatic and kidney functionsin each group were not altered.CONCLUSION:AFP-CTLA4 DNA vaccine can stimulatepotent specific CTL responses and has distinctive antitumoreffect on AFP-producing tumor.The vaccine has no impacton the function of mouse liver and kidney.     

A DNA vaccine against extracellular domains 1-3 of flk-1 and its immune preventive and therapeutic effects against H22 tumor cell in vivo

AIM:To construct a DNA vaccine against extracellulardomains 1-3 of fetal liver kinase-1 (ilk-1),and to investigateits preventive and therapeutic effect against H22 cell in vivo.METHODS:FIk-1 DNA vaccine was produced by cloningextracellular domains 1-3 of flk-1 and by inserting the clonedgene into pcDNA3.1 ( ).Fifteen mice were divided into3 groups and inoculated by vaccine,plasmid and salinerespectively to detect specific T lymphocyte response.ThirtyMice were equally divided into preventive group andtherapeutic group.Preventive group was further dividedinto V,P,and S subgroups,namely immunized by vaccine,pcDNA3.1 ( ) and saline,respectively,and attacked byH22 cell.Therapeutical group was divided into 3 subgroupsof V,P and S,and attacked by H22,then treated withvaccine,pcDNA3.1 ( ) and saline,respectively.The tumorsize,tumor weight,mice survival time and tumor latencyperiod were compared within these groups.Furthermore,intratumoral microvessel density (MVD) was assessed byimmunohistochemistry.RESULTS:DNA vaccine pcDNA3.1 ( )flk-1-domains 1-3was successfully constructed and could raise specific CTLactivity.In the preventive group and therapeutic group,tumor latency period and survival time were significantlylonger in vaccine subgroup than that in P and S subgroups(P<0.05);the tumor size,weight and MVD were significantlyless in vaccine subgroup than that in P and S subgroups(P<0.05).The survival time of therapeutic vaccine subgroupwas significantly shorter than that of preventive vaccinesubgroup (P<0.05);the tumor size,and MVD of therapeuticvaccine subgroup were significantly greater than that ofpreventive vaccine subgroup (P<0.05).CONCLUSION:DNA vaccine against flk-1 domains 1-3 canstimulate potent specific CTL activity;and has distinctiveprophylactic effect on tumor H22;and also can inhibit thetumor growth in vivo,This vaccine may be used as an adjuvanttherapy because it is less effective on detectable tumor.     

Establishment of mice model with human viral hepatitis B

AIM:To establish a mice model of hepatitis B by usingHBV-transgenic mice,and to transfer HBV-specific cytotoxicT lymphocytes (CTL) induced from syngeneic BALB/c miceimmunized by a eukaryotic expression vector containing HBVcomplete genome DNA.METHODS:HBV DNA was obtained from digested pBR322-2HBV and ligated with the vector pcDNA3.RecombinantpcDNA3-HBV was identified by restriction endonucleaseassay and transfected into human hepatoma cell line HepG2with lipofectin.ELISA was used to detect the expression ofHBsAg in culture supernatant,and RT-PCR to determinethe existence of HBV PreS1 mRNA.BALB/c mice wereimmunized with pcDNA3-HBV or pcDNA3 by intramuscularinjection.ELISA was used to detect the expression of HBsAbin serum.MTT assay was used to measure non-specific orspecific proliferation ability and specific killing activity ofspleen lymphocytes.Lymphocytes from immunized micewere transferred into HBV-transgenic mice (2.5×10~7 permouse).Forty-eight hours later,the level of serum proteinand transaminase was detected with biochemical method,liver and kidney were sectioned and stained by HE to observethe pathological changes.RESULTS:By enzyme digestion with Eco RI,Xho Ⅰ andHind Ⅲ,the recombinant pcDNA3-HBV was verified tocontain a single copy of HBV genome,which was insertedin the positive direction.HepG2 cells transfected with therecombinant could stably express PreS1 mRNA and HBsAg.After immunized by pcDNA3-HBV for 4 weeks,HBsAb wasdetected in the serum of BALB/c mice.The potential of spleenlymphocytes for both non-specific and specific proliferationand the specific killing activity against target cells wereenhanced.The transgenic mice in model group had nosignificant changes in the level of serum protein but had anobvious increase of ALT and AST.The liver had obviouspathological changes,while the kidney had no evidentdamage.CONCLUSION:A eukaryotic expression vector pcDNA3-HBV containing HBV complete genome is constructedsuccessfully.HepG2 cells transfected with the recombinantcan express PreS1 mRNA and HBsAg stably.Specific cellular immune response can be induced in mice immunized bypcDNA3-HBV.A mice model of acute hepatitis with HBV hasbeen established.     

Prevalence of hepatitis B virus genotype B in Vietnamese patients with chronic hepatitis B

Purpose  Hepatitis B virus (HBV) genotypes can affect treatment response to interferon-based therapy and disease outcomes in patients with chronic hepatitis B (CHB). Little data exist to characterize HBV genotypes in Vietnamese, one of the largest minority groups in the United States and also one with one of the highest CHB and liver cancer disease burdens. The goal of this study was to compare the distribution of HBV genotypes in Vietnamese and Chinese patients. Methods  We performed a cross-sectional study of 567 consecutive patients of Vietnamese (n = 478) or Chinese (n = 89) descent, with HBV genotype mutation analysis performed between 7/2,005 and 6/2,008 at a community gastroenterology clinic and a university-affiliated liver clinic in the United States. Results  There were no significant differences between the Vietnamese and Chinese groups in mean age (45 and 44 years), gender (58% and 61% male), HBeAg status (64% and 65% negative), median alanine aminotransferase (33 and 41 U/L), and log₁₀ HBV DNA (4.9 and 5.0 log₁₀ IU/ml), or the prevalence of precore/basic core promoter mutations (72% and 71%), respectively. Vietnamese patients had a much higher prevalence of HBV genotype B and a lower prevalence of genotype C than Chinese patients: 74% and 25% vs. 55% and 43% (P = 0.001). Conclusions  Chinese patients with CHB often carry either B or C genotype. Vietnamese patients with CHB mostly have HBV genotype B. Additional studies are needed to further characterize the clinical significance of HBV genotype in the natural history and treatment outcomes of CHB in Vietnamese patients.     

Hepatitis B virus genotype B is associated with earlier HBeAg seroconversion compared with hepatitis B virus genotype C

BACKGROUND & AIMS: Recent studies suggest that hepatitis B virus (HBV) genotype B is associated with less active liver disease than HBV genotype C. The aim of our study was to determine if HBV genotype B is associated with higher rates of spontaneous hepatitis B e antigen (HBeAg) seroconversion compared with genotype C. METHODS: A retrospective study using stored sera from 332 Chinese patients with chronic HBV infection followed for a mean of 48 months (range, 12-98) were tested for HBV genotype using a line-probe assay. RESULTS: HBV DNA was detected in 273 patients, 122 had HBV genotype B and 147 genotype C. Patients with genotype B had a significantly lower prevalence of HBeAg at presentation and significantly higher rates of spontaneous HBeAg seroconversion during follow-up. HBV genotype B patients who were HBeAg positive were significantly younger, and spontaneous HBeAg seroconversion occurred approximately 1 decade earlier compared with HBV genotype C patients. Multivariate analyses identified high alanine aminotransferase (baseline and follow-up), age >30 years, and genotype B as independent factors associated with spontaneous HBeAg seroconversion. CONCLUSIONS: HBV genotype B is associated with earlier HBeAg seroconversion than genotype C. This finding may explain the less active/progressive liver disease in patients with genotype B.     

Hepatitis B genotypes correlate with clinical outcomes in patients with chronic hepatitis B

BACKGROUND & AIMS: Six genotypes (A-F) of hepatitis B virus (HBV) have been identified; however, the genotype-related differences in the pathogenicity of HBV remain unknown. Therefore, we investigated the prevalence of HBV genotypes in Taiwan and the association between distinct genotypes and severity of liver disease in a cross-sectional study. METHODS: Using a molecular method, HBV genotypes were determined in 100 asymptomatic carriers and in 170 patients with histologically verified chronic liver disease and hepatocellular carcinoma (HCC). RESULTS: All genotypes except genotype E were identified in Taiwan, and genotypes B and C were predominant. Genotype C was prevalent in patients with cirrhosis and in those with HCC who were older than 50 years compared with age-matched asymptomatic carriers (60% vs. 23%, P < 0.001, and 41% vs. 15%, P = 0.005, respectively). Genotype B was significantly more common in patients with HCC aged less than 50 years compared with age-matched asymptomatic carriers (80% vs. 52%, P = 0.03). This predominance was more marked in younger patients with HCC (90% in those aged 相似文献   

Medication Nonadherence with Long-Term Management of Patients with Hepatitis B e antigen-Negative Chronic Hepatitis B

Background and Aims  

Antiviral treatment responses for patients with hepatitis B e antigen (HBeAg)-negative chronic hepatitis B (CHB) are well-defined by data from registration trials but may differ from patients seen in community settings where medical adherence is usually not as strictly monitored. The goal of this study was to examine the long-term outcomes of HBeAg-negative patients in a community clinical setting.     

慢性乙型肝炎患者HBV基因耐药突变分析

目的 探讨慢性乙型肝炎患者HBV基因耐药突变情况。方法 收集2010年12月至2014年12月我院感染病科诊治的慢性乙型肝炎患者60例,均为使用拉米夫定或阿德福韦酯后出现病毒学突变的患者,采用荧光定量PCR法检测HBV DNA,采用ELISA法检测HBeAg和HBeAb定量,对血清HBV DNA直接进行基因测序。结果 在60例患者,发现rtl 180M阳性17%,rtm 204V阳性20%, rtn 236T阳性12%,rtA 181V/T阳性10%;rtl 180M阳性和rtm 204V阳性患者HBV DNA拷贝数较高,rtn 236T阳性患者较rtn 236T阴性患者ALT和HBV DNA拷贝数较低;rtA 181V/T阳性患者较rtA 181V/T阴性患者ALT水平高。结论 rtl 180M、rtm 204V、rtn 236T和rtA 181V/T变异多见于年龄大、病程长的患者,HBV DNA拷贝数升高先于ALT升高者,预后较差,其中rtl 180M和rtm 204V变异患者病情容易复发。     

乙型肝炎病毒携带孕妇乳汁和血清 HBV DNA载量与婴儿感染率的关系分析

目的 了解HBV携带孕妇所生婴儿接受母乳喂养后HBV感染情况。方法 2013年6月~2016年8月在我院分娩的128例HBV携带孕妇,其中HBeAg阳性74例, HBeAg阴性54例。采用PCR法检测血清和乳汁HBV DNA水平。结果 HBeAg阳性组血清HBV DNA水平为（4.27±1.20） lg copies/ml, 乳汁HBV DNA水平为（3.29±1.02） lg copies/ml, 均显著高于HBeAg阴性组【分别为（1.05±0.23）lg copies/ml和（0.97±0.15）lg copies/ml,P均<0.05】;在婴儿6个月和12个月时,84例HBeAg阳性孕妇所生婴儿HBV感染18例（21.95%）和24例（29.27%）,均显著高于64例HBeAg阴性组婴儿【分别为3例（4.69%）和5例（7.81%）,P均<0.05】。结论 HBeAg阳性孕妇在分娩前后应接受积极的抗病毒治疗,以降低血清和乳汁HBV DNA水平,才能考虑给予婴儿母乳喂养。     

Severe illness with influenza B

Fifteen patients with recent influenza B infection were admitted to three Dallas hospitals in the first 11 weeks of 1977. The patients' ages ranged from five to 73 years, with a median of 18 years. Most had no significant underlying disease.The spectrum of clinical illness included postinfluenzal bacterial pneumonia (three cases), other severe chest disease (two cases), hyperpyrexia and possible rhabdomyolysis in the elderly (two cases), onset of toxemia of pregnancy, thyroid dysfunction, Stevens-Johnson syndrome, neurologic disorders (two cases), and Reye's syndrome (three cases). Two patients died. Two elderly men with high fever and weakness entered the hospital within three days of illness and two of three patients with Reye's syndrome were admitted four days after the onset of influenza, but the 11 other patients gave a history of seven to 31 days of symptoms before being hospitalized. Morbidity and mortality with influenza B are neither trivial nor restricted to debilitated hosts.