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Objective:To study the clinical characteristics of Lynch syndrome-associated endometrial cancer in elderly patients and the relationship of the disease with MSH2 gene mutations in patients' families.Methods:Clinical data of 473 elderly patients with endometrial cancer admitted to our hospital between January 2016 and January 2021 were retrospectively analyzed.MSH2 gene mutations were detected and verified by DNA sequence analysis, real-time quantitative PCR and reverse transcription PCR.Patients were divided into a Lynch syndrome group and a non-Lynch syndrome group, and the clinicopathological features of the two groups were compared.Results:Of 473 endometrial carcinoma patients, 46(9.7%)had embryogenic mutations of the MMR gene and were diagnosed with Lynch syndrome, with 18, 6, 24 and 10 patients carrying MLH1, PMS2, MSH2 and MSH6 mutations, respectively.There were 3 mutations in the MSH2 gene, including exon 7 1380C>A, exon 12 2011A>G and exon 13 2756A→AC.The proportions of patients with G3 grade endometrioid adenocarcinoma, lower uterine segment involvement and a history of Lynch syndrome-associated malignant tumors in the Lynch syndrome group were significantly higher than those in the non-Lynch syndrome group( χ2=8.935, 8.303, 9.770, all P<0.05). Conclusions:Poorly differentiated endometrioid adenocarcinoma, predisposition to lower uterine segment involvement and familial inheritance are the main clinical manifestations of Lynch syndrome-associated endometrial carcinoma in elderly patients, with the most common mutations seen in the MSH2 gene.
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Objective To explore the infection status of hepatitis B virus (HBV) in families of hepatitis B surface antigen (HbsAg) positive students and the mutations of HBV related to hepatocellular carcinoma, so as to provide theoretic evidence for the prevention and control of HBV infection and hepatocellular carcinoma. Methods A total of 1 611 students were investigated; they were from 60 classes of 15 schools and kindergartens in Pudong New Area, Shanghai, China, and 8 HBsAg positive students were found. These 8 students and their 18 first-degree relatives were enrolled in this study. Venous blood samples were collected to test the 5 markers of hepatitis B using enzyme linked immunosorbent assay. HBV DNA was detected by fluorescent PCR. HBV genome, basic core promoter (BCP) region and PreS region were detected using multiplex-PCR and nested PCR combined with cloning and sequencing. Results The positive rates of HBsAg and HBcAb in the first-degree relatives were 33.3% (6/18) and 38.9% (7/18), respectively. Both HBsAg and HBcAb positive rates in the mothers were 71.4% (5/7), which were significantly higher than those of the other first-degree relatives (P0.05). Seven of 8 families (87.5%) had 2 or more members infected or had ever infected with HBV. Fourteen of 26 members in 8 families had positive HBsAg, with a positive rate of 53.8%. A total of 4 groups of mothers and children received gene detection. Three groups of them had type C HBV gene, and 1 group had type C in mother and type B in child. Among the hepatocellular carcinoma-related HBV mutations, the mutation frequency of hot spots in BCP region was lower in the children than that in the mothers. Eight HBV mutation sites of type C in PreS region were found in both the mothers and children, and none of the remaining key sites were found in the children. Conclusion There is obvious family clustering of HBV infection, suggesting that HBV infection of students is more likely to be transmitted through mother-to-child transmission, but there are other ways of infection, such as acquired blood. The evolution degree of HBV gene in children is lower than that in mothers, which conforms to the rule of HBV evolution.
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HBV A1762T/G1764A double nucleotide substitution (also called TA mutation) is relatively common in liver diseases. Hepatocyte nuclear factor 4 (HNF4) is one of liver-enriched transcription factors, and TA mutation is located in the binding region of HNF4 and HBV and plays an important regulatory role in HBV gene transcription and replication. Several studies have pointed out that TA mutation may aggravate liver diseases after HBV infection and increase the risk of chronic liver failure and liver cancer; however, it is still unclear how TA mutation can aggravate liver disease after HBV infection, and more studies are needed for clarification in the future. This article reviews the association between HBV A1762T/G1764A double nucleotide substitution and liver diseases.
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Objective To systematically assess the relation between angiotensin-I converting enzyme(ACE) gene insertion/deletion (I/D) variation and type 2 diabetic nephropathy (T2DN) onset risk among Chinese population.Methods The related literatures were retrieved from the China National Knowledge Infrastructure (CNKI) and Wanfang Data until June 1st,2016.The RevMan 5.0 was used to conduct the statistical analysis.The merge OR value and corresponding 95% confidence interval(95%CI) were used to assess ACE gene I/D polymorphism and T2DN onset risk.Results Totally 29 papers with 4 357 subjects were included according to the inclusion and exclusion standard,including 2 208 cases of DN and 2 149 cases of T2DM without DN.Meta analysis showed that compared with ACE gene I/D polymorphism I allele,D allele could significantly increase the risk of T2DM patients suffering from DN,the OR value and corresponding 95%CI were 1.44(1.25,1.66);the gene analysis showed that ACE gene I/D polymorphism loci were significantly correlated with DN onset risk in the Asian population.The corresponding relative onset risk OR and 95%CI were 1.42(1.15,1.76) and 1.75(1.46,2.10) in the dominant and recessive genetic model.The Begg′s test showed that the included data had no obvious publication bias existence.Conclusion ACE gene I/D polymorphism is closely correlated with the onset risk of T2DN,and D allele might be a risk genetic factor for DN occurrence in the patients with T2DM.
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OBJECTIVE To study the association between polymorphism of DNA repair gene XRCC3Thr 241 Met and the risks of developing laryngeal carcinoma. METHODS Sixty patients with laryngeal carcinoma and 120 cancer-free controls were genotyped for the polymorphism by Multiplex SNaPshot.The odds ratios (ORs) and 95% confidence intervals (Cls) were calculated using unconditional logistic regression model.RESULTS The XRCC3 241 Met allele increased the risks of developing laryngeal carcinoma.Comparing with subjects having the XRCC3241 Thr/Thr genotype,the subjects at least having one XRCC3 241 Met allele had OR of 4.27 (95%CI 1.49-12.18). CONCLUSION XRCC3 Thr 241 Met plays an important role in the development of laryngeal carcinoma.
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Objective To study the phylogenetic evolution and genetic variations of gag gene among the prevalent human immunodeficiency virus (HIV )‐1 strains in Guangxi Zhuang Autonomous Region . Methods Plasma samples of 158 HIV‐1 infected patients in Guangxi area were collected during October 2011 to March 2012 .The gag gene fragments of HIV‐1 were amplified by reverse transcription/nested‐polymerase chain reaction and then sequenced .MEGA 5 .03 was utilized to construct phylogenetic tree and to calculate the genetic distances and selection pressures (globle ω) of gag gene and its coding regions . The comparisons between two groups were tested by Student′s t test ,and the comparisons of multiple groups were tested by one‐way ANOVA .Results A total of 140 amplification products of gag gene were obtained from 158 samples .Four subtypes of HIV‐1 were found ,including CRF01_AE (80 ,57 .1% ) , CRF08_BC (46 ,32 .9% ) ,CRF07_BC (10 ,7 .1% ) ,and subtype B (B′) (4 ,2 .9% ) .The genetic distances of gag gene of the above subtypes were 0 .036 ± 0 .001 ,0 .031 ± 0 .002 ,0 .043 ± 0 .003 and 0 .102 ± 0 .006 ,respectively ,with statistical significance (F=220 .62 ,P<0 .01) .The p17 and p24 coding regions suffered negative selection pressure (globleω<1) .Neither the globle ω in p17 region nor that in p24 region had significant differences among different subtypes (F=0 .761 ,P=0 .469 and F=0 .037 ,P=0 .964 , respectively ) . Conclusion CRF01_AE is the major subtypes of HIV‐1 in Guangxi Zhuang Autonomous Region .The coding regions of gag gene are relatively conserved during evolution .Changes of HIV‐1 prevalence ,however ,may affect the genetic variation of gag gene ,which should be continuously monitored .
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Objective To investigate the relationship between genetic variants in the Toll-like receptor (TLR) pathway genes and susceptibility of gastric cancer (GC) and esophageal squamous cell carcinoma (ESCC).Methods The data of whole genome association studies of the high-risk population of GC and ESCC in China were analyzed by adaptive rank-truncated product (ARTP) method in pathway and gene level.The associations between single nucleotide polymorphism (SNP) and susceptibility of GC and ESCC were analyzed with additive model of unconditional Logistic regressions.PLINK 1.07 and SPSS 19.0 software were performed for statistical analyses,and ARTP package in R3.0.2 was used for pathway and gene level analysis.Results In gene-level analyses,eight genes were found to be associated with susceptibility of GC (P ＜0.05) and six genes were associated with susceptibility of ESCC (P ＜ 0.05).In single SNP-level analyses,21 SNPs were statistically correlated with susceptibility of GC (P ＜ 0.01),and 11 SNPs were statistically correlated with susceptibility of ESCC (P ＜0.01).Conclusions Some genetic variants in TLR pathway are associated with risk of GC and ESCC.The potential molecular mechanisms need further investigation.
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Objective To explore the mutations of hepatocellular carcinoma (HCC)-related hepatitis B virus (HBV) during mother-to-child transmission. so as to provide theoretic evidence for prophylaxis of HCC from the very beginning. Methods A total of 413 HBsAg-positive mothers and their newborns were enrolled in this study. Serum HBV DNA levels in maternal peripheral blood and cord blood of the newborns were measured using real-time quantitative PCR. Nested PCR together with cloning and sequencing methods were applied to examine the HCC-related HBV mutations in the preS and basal core promoter regions of HBV genome. All the newborns received standard HBV vaccination. Of the 413 newborns. 104 were successfully followed-up 7 months after birth. and the HBV mutations were examined if their circulating HBV DNA was detectable. Results Of the 413 newborns. 41 (9. 9%) had HBV DNA level >103 copies/mL in their cord blood. Four (3. 8%) of the 104 newborns who were successfully followed up had circulating HBV DNA level >103 copies/mL 7 months after birth. Compared to mothers without HBV trans-placental transmission. those with HBV trans-placental transmission had no increase in HBV mutations in the basal core promoter region. However. the viral mutations containing T2898G/C. C3000T. C3116T. T31C • and T52C in the preS region of HBV subgenotype C2 significantly increased the risk of HBV trans-placental transmission 8<0. 05). The frequencies of the HCC-related mutations in the preS and basal core promoter regions of HBV genome were not significantly different between maternal peripheral blood and the cord blood of the newborns. Importantly. the HCC-related mutations were rarely found in the HBV-positive infants at 7 months after birth. Conclusion The HBV mutations in the preS region of HBV subgenotype C2 may affect the trans-placental transmission of HBV. However, the quasispecies of HCC-related HBV mutants have no advantage in causing chronic HBV infection in infants. The HBV mutants which can promote HCC are selected during the long term chronic infection.
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Objective To explore the relationship between the Ebola virus genome variations and its epidemiological characteristics by analyzing the 102 whole genome sequences of Ebola viruses in 2014 outbreak. Methods Whole genome sequences of Ebola viruses (EBOVs) were obtained from the NCBI database, and the variations in genome sequences were analyzed by Mummer3. 0. The evolutionary analysis was carried out through MEGA5; and the 3D modeling of the protein was performed using CPHmodels and PyMOL software. Results It was found that there were 606 single nucleotide variants (SNVs) in the genome of 2014 EBOVs, of which 49 nonsynonymous SNVs were unique. The amino acids of NP-182, GP-82 and L-1951, which were highly conserved not only among all the Zaire EBOVs before 2014, but also among different EBOV species, were altered in 2014 EBOVs. Conclusion The unique mutation of 2014 EBOVs resulting in alterations of NP, GP and L protein, especially the alteration of aa82 of GP (Ala→Val), might weaken the stability of α-helix where the amino acid is located, which might be associated with the weakened lethality and enhanced transmission of the virus. Further studies are needed to confirm whether genomic variations in 2014 EBOVs is responsible for change of the epidemiological characteristics.
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China is among the middle-high endemic regions of HBV infection.The pathological outcomes of chronic HBV infection have been shown to be greatly influenced by several important factors,including HBV genotype,sub-genotype and gene viability mutation.HBV genome mutation,on the one hand,could alter its replication and secretion and thus change viral pathogenicity.In addition,host immune microenvironment and host-virus interaction,disease progression and the effect of antiviral therapy could be adapted at the same time.The detection of HBV genotypes,genetic subtypes and the key hotspot mutation is helpful to clinical risk assessment and prognosis prediction of HBV-related end-stage liver diseases (cirrhosis and hepatocellular carcinoma),it is also helpful to auxiliary predict the liver diseases recurrence and metastasis after treatment.Thus persistent care should be taken on the HBV mutation and its clinical translation so as to provide solid evidences for the personalized,standardized and fine management of HBV-related liver diseases.
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The development and progression of colorectal cancer (CRC) is a process that accumulates the driver somatic mutations elicited by chronic inflammation. Epigenetic modifications and genetic mutations play key roles in the whole evolutionary process of chronic inflammation-induced CRC. Adenoma gradually progresses into adenocarcinoma via accumulating genetic mutations stimulated by cancer promoting stimulations. The next generation sequencing technology provides an effective way to identify the “driver” mutations and fusion genes in the carcinogenesis, providing molecular evidences for cancer evolution. The most significant genetic changes during malignant transformation from adenoma to adenocarcinoma are the significant increases in microsatellite instability and chromosome instability. The main purpose of investigating the evolutionary process of CRC, especially somatic mutations, is to identify the related signaling pathways, which are the key steps to explore early effective intervention strategies and targeted therapies for CRC.
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Objective To investigate the binding capacities of two variants of quinolone resistance-determining region in the DNA gyrase subunit A to substrates in Klebsiella pneumonia (K.pneumonia).Methods Tertiary structures of two variants (type Ⅰ and type FH) of quinolone resistance-determining region in the DNA gyrase subunit A in K.pneumonia were predicted by homology modeling referring to that of wild type.Then,DOCK module in ArgusLab 4.1 software was used to perform molecular docking of two variants and wild type to seven kinds of quinolones substrates,and calculate binding free energies (△G).Moreover,numbers and distances of interaction between amino acid residues of DNA gyrase subunit A and ciprofloxacin were calculated.Results Molecular docking showed that binding free energies of type Ⅰ and type FH to pipemidic acid,ciprofloxacin,gatifloxacin were-26.607 50,-29.530 39,-29.493 09 kJ/mol and-26.696 44,-28.972 83,-29.590 50 kJ/mol,respectively,which declined greater than those of wild type (-27.188 82,-30.872 00 and-30.244 04 kJ/mol,respectively) and showed drug resistance.While binding free energies of type Ⅰ and type FH to levofloxacin were-29.013 81 and-29.497 57kJ/mol,respectively,and that of wild type was-28.016 20 kJ/mol.The binding free energies of type Ⅰ and type FH to nalidixic acid,norfloxacin,ofloxacin increased or declined.Moreover,if distance was less than 5 angstroms,atom pairs formed between wild type of DNA gyrase subunit A and ciprofloxacin had 16 pairs,while type Ⅰ and type FH had 2 pairs and 4 pairs,respectively.If distance was less than 4 angstroms,atom pairs formed between wild type and ciprofloxacin had 8 pairs,while type Ⅰ and type FH had no atom pairs.Conclusion Decline of binding capacities of two variants of DNA gyrase subunit A in K.pneumonia to ciprofloxacin played a role in drug resistance.
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Objective To analyze the VP1 and VP4 genetic region of enterovirus 71 (EV71)isolated from severe cases and mild cases with hanD-foot-mouth disease (HFMD) in Shanghai in 2011.Methods Five EV71 strains isolated from severe cases and five EV71 strains from mild cases in 2011 were included.Reverse transcription-polymerase chain reaction (RT-PCR) method was used to amplify and sequence the VP1 and VP4 genes of EV71,and then the sequencing results were compared with those of A,B,C genotype reference EV71 strains from GenBank by nucleotide alignment,amino acid alignment and phylogenetic tree analyses.Results The homogeneity between EV71 strains from severe cases and mild cases was 96.0％-98.1％ and 93.7％-99.5％ for VP1 and VP4 nucleotide sequences,respectively.The VP1 nucleotide sequences of 5 strains isolated from severe cases and 5 strains from mild cases in Shanghai shared 86.9％-98.2％ and 87.4％ 98.5％ identity with genotype C,respectively,while the homogeneity of VP4 nucleotide sequence was 85.5％-100.0％and 84.5％-99.5％,respectively.In addition,compared with the Fuyang EV71 strains (representative of C4 subtype),the strains from mild and severe cases shared homogeneity of 97.0％-98.2％ and 97.9％-98.5％ for VP1 gene,respectively,96.1％-100.0％ and 97.1％-99.5％ for VP4 gene,respectively.Among 3 strains isolated from severe cases,mutations at the residue 282 in the VP1 protein (N→S) and residue 7 in the VP4 protein (T→A) were discovered simultaneously.Conclusions The 10 EV71 strains isolated from severe and mild cases in Shanghai belong to subgenogroup C4.Among 3 strains isolated from severe cases,mutations at the residue 282 in the VP1 protein (N→S) and residue 7 in the VP4 protein(T→A) are discovered simultaneously.
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Objective To investigate the evolutionary characteristics and important amino acid sites of the neuraminidase (NA) gene of the novel influenza virus A/H7N9 in 2013 epidemic. Methods The NA gene sequences of influenza virus A N9 subtype of different times and areaswere downloaded from the database of The National Center for Biotechnology Information (NCBI) and The Global Initiative on Sharing All Influenza Data (GISAID). MEGA 5. 05 and BioEdit softwares were used for phylogenetic tree construction and nucleotide/protein sequence analysis. Results The NA gene sequence of the novel influenza virus A/H7N9 in 2013 shared a 96% similarity with that of the H11N9 avian strain found in Czech Republic in 2010. There were 5 amino acid deletions in this novel influenza virus,and one of the new strains had a variation in potential enzymatic active sites. Conclusion The NA gene of this 2013 novel influenza virus might originate from the avian H11N9 strains detected in Czech Republic. The deletion of amino acid might result in human infection and high fatality rate.
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Objective To investigate the evolution and variations in coding amino acids of hemagglutinin (HA) gene of the novel avian influenza virus H7N9 in 2013 epidemic. Methods The HA gene sequences of influenza virus H7N9, H7N2, H7N3 and H7N7 subtypeswere downloaded from the database of The National Center for Biotechnology Information (NCBI) and The Global Initiative on Sharing All Influenza Data (GISAID). MEGA 5. 05 software was used for sequence analysis and N- J method was used for constructing the phylogenetic trees. The amino acid sequences at the receptor binding sites, glycosylation sites, and cleavage siteswas aligned and analyzed. Results The HA genes this novel A/H7N9 virus in 2013 shared a 95. 3%- 95. 6% similarity with JQ906573. 1| Zhejiang (H7N3 virus) isolated in 2011. The most important variation in this novel H7N9 isolates was found at the receptor binding site: Q226L. The 5 glycosylation sites were highly conservative. One basic amino acid (R) at the HA cleavage sites, located between aa339 and aa340, was also found in this novel isolate. Conclusion The HA gene of this novel H7N9 isolate might originate from H7 subtypes carried by birds in China. Thebinding site change caused by Q226L variation might be responsible for human infection of this novel H7N9 isolate.
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Hepatitis B virus has a high degree of genetic variation. Multiple selection pressure,exerted by vaccine,antiviral drugs and diagnostic reagents,might favor certain mutations of HBV. HBV variation under the selective pressure may affect the pathogenesis of HBV-related liver disease,immunization effect,antiviral efficacy and laboratory tests.In this paper,we summarize the molecular characteristics and its significance of HBV variants,which will help us to manage hepatitis B.
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The DNA copy-number variant (CNV) is a kind of segments of DNA ranging from 1 kb to 3 Mb that is present in a variable number of copies.CNVs widely distribute across the human genome,and dramatically increases genetic diversity.In recent years,researches have found that most CNVs are closely related to complex diseases.If a cancer gene is directly encompassed or overlapped by a CNV,it may lead to activation of oncogenes or inactivation of tumor suppressor genes,and finally results in tumorigenesis.CNVs can affect gene expression,phenotype differences and phenotypic adaptations by changing gene dosages and gene activities,and then sequentially lead to tumor or any other genetic dieases.Investigating CNVs is apparently helpful for studing chromosome recombination,genomic evolution,gene expression and the pathogenesis of multiple complex diseases especially tumor.
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Objective To evaluate the efficacy and drug resistance profiles of nucleosides (NA) retreatment in NA experienced chronic hepatitis B (CHB) patients. Methods Totally 104 NA experienced CHB subjects were enrolled in this study.All these subjects had received at least 3 months NA monotherapy and stopped the treatment,and then received NA retreatment for at least one year.The subjects were divided into three groups according to the following criteria:reached the therapy endpoint of China guideline when they stopped NA-naive treatment (group A,n =39); did not reach the therapy endpoint when they stopped NA-naive treatment but hepatitis B virus (HBV) DNA＜1.0× 103 copy/mL (group B,n=33); and with HBV DNA＞1.0× 103 copy/mL (group C,n=32).The efficacy and drug resistance profiles of retreatment were compared among three groups. The effects of baseline alanine aminotransferase (ALT) levels,HBV DNA levels and HBeAg titers on the retreatment efficacies were analyzed. The mutations of HBV P gene were detected by nested polymerase chain reaction (PCR) and direct sequencing.The data were analyzd by Wilcoxon test and x2 test.Results The time to ALT normalization in patients with baseline ALT＜ 1.3 × upper limit normal (ULN) was shorter than that in patients with ALT≥1.3×ULN (2 months vs 4 months; Z=2.281,P=0.023).The time to virological response in patients with baseline HBV DNA＜5 lg copy/mL was shorter than that in patients with HBV DNA≥5 lg copy/mL (1 month vs 2 months; Z=2.054,P =0.040). The time to virological response and ALT normalization in baseline HBeAg negative were both shorter than those in patients with baseline HBeAg positive patents ( 1 month vs 3 months and 2 months vs 4.5 months,respectively; Z=2.580 and 2.304,respectively; both P＜0.05). The subjects in group A achieved virological response and HBeAg seroconversion after retreatment earlier compared to previous NA-naive therapy ([1.61 ± 1.76] months vs [3.48±4.066]months and [3.38 ± 3.34] months vs [9.92-11.22] months,respectively; Z=-2.854 and-1.094,respectively; both P＜0.05).The cumulative HBeAg seroconversion rate in group A was higher compared to those in group B and group C (80.0％ vs 36.8％ and 37.5％,respectively; x2 =4.368 and 5.100,respectively; both P＜0.05).Thirteen patients developed clinical resistance and four of them developed genotypic resistance proved by PCR direct sequencing.Among the patients retreated with the same regimen as previous in the C group,the cumulative resistance rate was highest compared to group A and B (44％ vs 9％ and 0,respectively; x2 =5.019 and 6.588,respectively;both P＜0.05).No resistance was detected in the 14 patients retreated with combined NA treatment without cross resistance.Conclusions For NA experienced CHB patients who fulfill the indication of antiviral therapy,the retreatment should be started as earlier as possible. Retreatment with NA combination without cross resistance can prevent (reduce) the risk of developing drug resistance.
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Objective To compare the efficacy and safety of Iamivudine-interferon sequential therapy and lamivudine monotherapy in HBeAg-positive chronic hepatitis B (CHB) patients.MethodsA total of 172 patients with HBeAg-positive CHB were randomized to sequential group (n=83) or lamivudine group (n=89).Sequential group were administrated with lamivudine 100 mg/d and 5 million units interferon alpha 2b subcutaneous injection every other day for 24 weeks were added since week 25 of treatment.Lamivudine group were administrated with lamivudine 100 mg/d for 48 weeks.All subjects were followed up for 24 weeks after drug withdrawal.Measurement data with homogeneity of variance were analyzed by using t test and data with heterogeneity of variance were analyzed by using rank sum test.The comparison of rates was done by chi square test or Fisher exact test.ResultsThe baseline hepatitis B virus (HBV) DNA levels of patients in sequential group and lamivudine group were (7.8±1.0) and (7.9±1.1) lg copy/mL,respectively (P＞0.05),and the baseline alanine aminotransferase (ALT) levels were (210.5 ± 150.1 ) and (211.9 ± 160.9) U/L,respectively (P＞0.05).At the end of treatment,higher ALT levels [(78.4±146.1) vs (36.1±32.4) U/L,P＜0.05)] and HBV DNA levels [(4.5±1.5) vs (3.8±1.3) lg copy/mL,P＜0.05)] levels,lower response rates (65.8％ vs 83.5％,P＜0.05),and similar HBeAg loss rates (31.6％ vs 22.2％,P＞0.05) and HBeAg seroconversion rates (27.6％ vs 16.0％,P＞0.05) were found in sequential group compared with lamivudine group.At the end of follow-up,higher ALT levels [(126.0±143.1) vs (82.7±83.0) U/L,P＜0.05)],similar HBV DNA levels [(5.3±1.5) vs (5.0±1.5) lg copy/mL,P＞0.05)],similar HBeAg loss rates (25.0％ vs 32.3％,P＞0.05) and HBeAg seroconversion rates (25.0 ％ vs 26.2 ％,P＞0.05) were found in sequential group compared with lamivudine group.YMDD motif mutation rate in sequential group was lower than lamivudine group at week 48 of treatment (10.5％ vs 26.9％,P＜0.05).ConclusionsLamivudine-interferon sequential therapy and lamivudine monotherapy are both effective in HBeAg-positive CHB patients,while HBV mutations are reduced in patients with sequential therapy.
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Objective To detect nucleos(t)ide-resistance mutations in hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) isolated from hepatocytes of patients with chronic HBV infection and to analyze the correlation between the mutations found in cccDNA and relaxed circular DNA (rcDNA). MethodsForty patients with chronic HBV infection were investigated. Preoperation serum samples and non-tumor liver tissues were collected.Intrahepatic HBV cccDNA and rcDNA were selectively extracted by co-precipitation of sodium dodecyl sulphate-protein and QIAamp DNA Mini Kit, and further purification with plasmid-safe ATP-dependent DNase (PSAD).Thereafter,cccDNA were amplified by selective polymerase chain reaction (PCR) or nested PCR using the primers spanning both the two gaps in HBV genome and covering the common mutations associated with nucleoside analogues resistance (rt169- rt250).Intrahepatic HBV rcDNA and pre-operation serum HBV rcDNA were also extracted and then amplified by PCR.The PCR products were then purified and sequenced.Results Among the 40 patients,intrahepatic HBV cccDNA were detected in 31 patients,and HBV rcDNA were detected in liver samples of 35 patients and pre-operation serum samples of 21 patients. The PCR products amplified from these samples were all successfully sequenced.rtM204Ⅰ mutation was detected in intracellular HBV cccDNA,rcDNA and serum HBV rcDNA in 2 patients.Both rtM204Ⅰ and rtQ215H were detected in intrahepatic HBV cccDNA and rcDNA in 2 patients,while no corresponding mutation was observed in serum HBV rcDNA of these two patients.Three variants including rtM204V,rtM204V and rtV173L-rtL180M-rtM204V were detected in serum HBV rcDNA in 3 patients,while corresponding mutants were not detected in intracellular HBV cccDNA or rcDNA of these patients.Condsions The results suggest that antiviral nucleos(t) ide resistance mutations can be found in HBV cccDNA in chronic hepatitis B patients. The dominant resistant mutation found in intrahepatic HBV cccDNA/rcDNA may be different from that in serum HBV rcDNA.