The Effect of ICOS Polymorphism Interactions with HBV Mutations on HBV Subtype Infection Outcomes.

INTRODUCTION AND AIM: Hepatitis B virus (HBV) infection remains a public health problem worldwide. In addition, HBV infection results are influenced by various virological, immunological, and genetic factors. Inducible T-cell costimulator (ICOS) polymorphisms involving chronic HBV infection have been confirmed in previous studies. This study was to explore the effects of ICOS single nucleotide polymorphisms in HBV subtypes and their interactions with viral mutations on HBV infection outcomes. MATERIAL AND METHODS: A total of 1,636 Han Chinese individuals were recruited, including 47 asymptomatic HBV carriers (ASC), 353 chronic hepatitis B (CHB) patients, 327 HBV-related liver cirrhosis (LC) patients, 193 HBV-related hepatocellular carcinoma (HCC) patients, 464 patients with spontaneous recovery from HBV infection (SR), and 252 healthy controls (HC). DNA samples from these subjects were genotyped for four ICOS SNPs (rs11883722, rs10932029, rs1559931, and rs4675379). Direct sequencing was used to determine the HBV mutations in the enhancer II, basal core promoter, and pre-core regions. RESULTS: We found that the genotype "TC" of ICOS rs10932029 SNP was associated with decreased HBV-related LC risk in the genotype C group. Additionally, the A1762T, G1764A and A1762T/G1764A mutations were associated with an increased risk of LC in the genotype C group. Further study indicated that interactions between ICOS rs10932029 genotype "TC" and A1762T or A1762T/G1764A mutations significantly decreased the LC risk in the genotype C group. CONCLUSION: The rs10932029 genotype "TC" might be an LC-protective factor for HBV genotype C infection. The interactions between the rs10932029 genotype "TC" and A1762T or A1762T/G1764A mutations could decrease the risk of LC.

Retinoid X Receptor α-Dependent HBV Minichromosome Remodeling and Viral Replication.

BACKGROUND AND AIM: The HBV covalently closed circular DNA (cccDNA) is organized into a minichromosome in the nuclei of infected hepatocytes through interactions with histone and nonhistone proteins. Retinoid X receptor α (RXRα), a liver-enriched nuclear receptor, participates in regulation of HBV replication and transcription through modulation of HBV enhancer 1 and core promoter activity. MATERIAL AND METHODS: This study investigated RXRα involvement in HBV cccDNA epigenetic modifications. Quantitative cccDNA chromatin immunoprecipitation (ChIP) was applied to study the recruitment of RXRα, histones, and chromatin-modifying enzymes to HBV minichromosome in HepG2 cells after transfection of the linear HBV genome. RESULTS: RXRα Was found to directly bind to HBV cccDNA; recruitment of RXRα to HBV mini-chromosome paralleled HBV replication, histone recruitment, and histone acetylation in HBVcccDNA. Moreover, RXRα overexpression or knock-down significantly increased or impaired the recruitment of the p300 acetyltransferase to cccDNAminichromosome. CONCLUSIONS: Our results confirmed the regulation of RXRα on HBV replication in vitro and demonstrated the modulation of RXRα on HBV cccDNA epigenetics. These findings provide a profound theoretical and experimental basis for late-model antiviral treatment acting on the HBV cccDNA and minichromosome.

Association of HLA-DQ and IFNL4 polymorphisms with susceptibility to hepatitis B virus infection and clearance.

UNLABELLED:  Background and aim. Leukocyte antigen DQ (HLA-DQ) and interferon-λ4 (IFNL4) gene polymorphisms were associated with susceptibility to chronic hepatitis B and C virus infection. This study further confirmed that variants of these genes were associated with susceptibility and spontaneous clearance of HBV infection in a Chinese population. MATERIAL AND METHODS: A total of 1,069 subjects were recruited and divided into three groups i.e. 397 with CLD (HBV-related chronic liver disease), 434 with SC (spontaneous clearance), and 238 HC (healthy controls). HLA-DQrs9275319 and IFNL4rs368234815, rs12971396, rs12979860, and rs8099917SNPs were genotyped using the Sequenom MassARRAY MALDI-TOF system. RESULTS: HLA-DQ rs9275319 showed a significant association with HBV infection (allele model, OR, 0.514; 95% CI, 0.359-0.738, adjusted p = 0.0003) and with natural clearance (allele model, OR, 1.659; 95% CI, 1.197-2.300, adjusted. However, there was no association between IFNL4 polymorphism and HBV susceptibility or natural clearance (all p > 0.05). The multifactor dimensionality reduction (MDR) test with permutation correction showed that a three-way interaction between IFNL4 and HLA-DQ SNPs was identified for HBV susceptibility (permutation p = 0.009 for the best factor model) and clearance (permutation p = 0.014 for the best factor model). CONCLUSIONS: The data from the current study provided additional evidence for an SNP-SNP interaction between HLA-DQ and IFNL4 in regulation to HBV infection and natural clearance.

Effects of interaction between genetic variants in human leukocyte antigen DQ and granulysin genes in Chinese Han subjects infected with hepatitis B virus.

Single nucleotide polymorphisms (SNPs) of HLA-DQ and granulysin (GNLY) are reportedly associated with HBV infection. The aim of this study was to investigate the effects of interactions between SNPs in HLA-DQ and GNLY on the outcome of hepatitis B virus (HBV) infection in Chinese Han subjects. HLA-DQ (rs9275572) and GNLY (rs1866139 and rs11127) were genotyped in 310 subjects with HBV-related chronic liver disease, 295 in whom spontaneous clearance of HBV had occurred and 316 who had not been exposed to HBV. HLA-DQ rs9275572 was significantly correlated with HBV clearance (dominant genetic model: OR, 1.84; 95% CI, 1.30-2.61; adjusted P = 0.001). There was no statistical association of GNLY rs1866139 and rs11127with HBV infection outcomes. However, significant sex-specific associations with HBV susceptibility were observed in men who carried rs1866139 CG or rs11127 TC and in women who carried rs1866139 GG or rs11127 CC. The findings were the same in the validation cohort, which was composed of 829 subjects. Based on a multifactor dimensionality reduction test with permutation correction, a three-way interaction between SNPs in HLA-DQ and GNLY was identified in terms of HBV clearance. In conclusion, additional evidence for an association of HLA-DQ and GNLY SNPs with HBV infection outcomes has been identified and a SNP-SNP interaction between HLA-DQ and GNLY on HBV clearance observed.

Dynamics of lamivudine-resistant hepatitis B virus strains in patients with entecavir rescue therapy.

Entecavir rescue therapy is frequently used in patients with lamivudine-resistant hepatitis B virus (HBV) strains. The aim of this study was to investigate evolutionary patterns of HBV quasispecies during entecavir rescue therapy and evaluate their impacts on therapeutic efficacy. We enrolled 21 chronic hepatitis B patients who failed to respond to lamivudine therapy and were switched to entecavir treatment. Measurement of serum HBV DNA and sequence analysis of HBV reverse transcriptase were done up to 144 weeks. Four patients of this series showed a reversion to wild-type HBV after entecavir treatment and in three of them, a complete viral response (<2.6 log10 copies/ml) was achieved. An additional five patients developed entecavir genotypic resistance, with prior occurrence of lamivudine-resistant mutation (L180 M ± M204 V/I). A viral breakthrough was observed in four of the five patients with entecavir-resistant mutants. The remaining 12 patients of this series showed dominance of lamivudine-resistant mutants throughout the entecavir rescue therapy, and five of them achieved a complete viral response at the end of follow-up. The average HBV DNA level was significantly lower in patients with a reversion to wild-type HBV than in those without it (P < 0.05). In conclusion, reversion to wild-type HBV is a favorable indicator for response to entecavir rescue therapy in lamivudine-refractory patients with chronic hepatitis B. The presence of lamivudine-resistant mutations contributes to the development of entecavir resistance.

Phenotypic assay of a hepatitis B virus strain carrying an rtS246T variant using a new strategy.

Phenotypic assays of hepatitis B virus (HBV) play an important role in research related to the problem of drug resistance that emerges during long-term nucleot(s)ide therapy in patients with chronic hepatitis B. Most of the phenotypic assay systems that are available currently rely on the transfection of recombinant replication-competent HBV DNA into hepatoma cell lines. Cloning clinical HBV isolates using conventional digestion-and-ligation techniques to generate replication-competent recombinants can be very difficult because of the sequence heterogeneity and unique structure of the HBV genome. In this study, a new strategy for constructing an HBV 1.1× recombinant was developed. The core of this strategy is the "fragment substitution reaction" (FSR). FSR allows PCR fragments to be cloned without digestion or ligation, providing a new tool for cloning fragments or genomes amplified from serum HBV DNA, and therefore making the assay of HBV phenotypes more convenient. Using this strategy, a phenotypic assay was performed on an HBV strain carrying an rtS246T variant isolated from a patient with chronic hepatitis B that was only responsive partially to entecavir therapy. The results indicated that this strain is sensitive to entecavir in vitro.

Comparison of a novel real-time PCR assay with sequence analysis, reverse hybridization, and multiplex PCR for hepatitis B virus type B and C genotyping.

We compared a novel real-time genotyping and quantitative PCR (GQ-PCR) assay, direct sequence analysis, reverse hybridization, and multiplex PCR for genotyping hepatitis B virus (HBV) in 127 HBV-infected patients. We found that GQ-PCR had the highest concordance with sequence analysis and the highest detection rate for mixed genotype detecting.

[Evolution of hepatitis B virus quasispecies during antiviral therapy in one chronic hepatitis B patient].

OBJECTIVE: To investigate the evolution of hepatitis B virus (HBV) quasispecies in one patient during lamivudine (LAM) monotherapy and switching to entecavir (ETV) rescue treatment. METHODS: Serum samples were taken at seven different time points during antiviral therapy (0, 24, 48, 60, 72, 96, 152 weeks, respectively), the HBV DNA polymerase gene was amplified, cloned and sequenced to analyze the amino acid substitutions within HBV DNA polymerase gene and distribution of virus quasispecies. Quantitative detection of the HBV wild strains and total virus was performed by amplification refractory mutation system real-time PCR (ARMS-PCR). RESULTS: Three mutation patterns detected during antiviral therapy in the patient: rtM204V, rtM204V+rtL180M and rtM204I. The HBV quasispecies were found always in dynamic variation. The HBV populations were completely replaced with the LAM-resistant variants when the viral breakthrough was encountered during LAM monotherapy. Interestingly, the wild-type variants presented gradually dominant (79.3%) with the decline of HBV DNA load after switching to ETV rescue administration. ARMS-PCR results showed that the wild-type variants account ed for 68.55% of the HBV populations at baseline and this proportion declined to 0.21% when the viral breakthrough emerged under LAM therapy. The wild-type variants gradually increased from week 24 after switching to ETV rescue therapy and the proportion of HBV wild-type variants in the population fluctuated between 16.01% to 26.93%. CONCLUSIONS: The distribution of virus quasispecies were always in dynamic variation during sequential therapy with nucleotide analogs in chronic hepatitis B patients. Different patterns of dynamic HBV quasispecies may have different contribution in ETV resistance in LMV refractory patients with ETV administration.

Simultaneous genotyping and quantification of hepatitis B virus for genotypes B and C by real-time PCR assay.

Hepatitis B virus (HBV) is an important cause of human chronic liver diseases and is a major public health problem. Viral load and HBV genotype play critical roles in determining clinical outcomes and response to antiviral treatment in hepatitis B patients. Viral genotype detection and quantification assays are currently in use with different levels of effectiveness. In this study, the performance of a real-time genotyping and quantitative PCR (GQ-PCR)-based assay was evaluated. Through the use of genotype-specific primers and probes, this assay provides simultaneous identification and quantification of genotypes B and C in a single reaction. Our GQ-PCR correctly identified all predefined genotypes B and C, and no cross-reaction between genotypes B and C were observed. The GQ-PCR identified more cases of HBV infections with mixed genotypes B and C than direct sequencing did. Samples from 127 HBV-infected Chinese patients were genotyped with GQ-PCR, revealing 56.7% HBV as genotype B, 13.4% as genotype C, and 29.8% as mixed genotypes B and C. This assay provides a reliable, efficient, and cost-effective means for quantification of the B and C genotypes of HBV in single or mixed infections. This assay is suitable for sequential monitoring of viral load levels and for determining the relationship between the genotype viral load and stage of disease in Asians.

[Dissection of mechanism for the adefovir dipivoxil resistance in chronic hepatitis B patients].

OBJECTIVE: To explore the mechanism for adefovir dipivoxil (ADV) resistance occurred in chronic hepatitis B patients of a series of phase III clinical trails. METHODS: 30 resistant HBV strains were selected out from 177 cases of ADV treated chronic hepatitis B patients. HBV polymerase RT region were amplified by nested PCR and analyzed with the standard nucleotide sequence of HBV strains deposited in GeneBank. RESULTS: 21 out of 30 HBV strains were primary resistant strains, among them 5 HBV strains (23.8%, 5/21) had the polymorphism site of rtN118H. While the other 9 HBV strains showed secondary resistance, variations in conservative region C (rtM207V) and other non-conservative regions were found. The classic mutation sites such as rtN236T and rtA181V/T were not found. CONCLUSIONS: Polymorphism site of rtN118H might be responsible for HBV primary resistance to ADV therapy. rtM207V variation in HBV RT C domain and other variation sites might play a role in HBV secondary resistance to ADV treatment, and natural resistant quasispecies may be the basis for the ADV quick resistance. These conclusions await further confirmation by phenotype test.

Effect of Dry-Wet Ratio on Properties of Concrete Under Sulfate Attack.

In order to explore the drying-wetting cycle test method of concrete under sulfate accelerating erosion, the influence of dry-wet time ratio on concrete sulfate erosion was studied. Under the condition of 7 days for one cycle, five different dry-wet time ratios were designed: 1:3, 1:1, 3:1, 5:1, and 10:1. The basic properties such as compressive strength, splitting tensile strength and dynamic elastic modulus of concrete were tested. Scanning electron microscopy (SEM) was used to analyze the microstructure of concrete before and after erosion. The test results show that under the environment of sulfate drying-wetting cycle erosion, the change of mechanical properties of concrete are divided into three stages: ascending period, fluctuating period and rapid descending period. Concrete is subjected to periodic damage process of initial damage followed by filling compaction, cracking, further filling, and cracking again, in that order. Dry-wet ratio has a significant effect on concrete sulfate attack. Under the same drying-wetting cycle period, as the dry-wet ratio increases, the degree of deterioration of concrete by sulfate attack increases first and then decreases. When the dry-wet ratio is 5:1, the deterioration is the most serious.

Hepatitis B virus genotype-associated variability in antiviral response to adefovir dipivoxil therapy in Chinese Han population.

It is well known that different genotypes of hepatitis B virus (HBV) have a different sensitivity to interferon-alpha or lamivudine (nucleoside analogue) antiviral therapy. However, for adefovir dipivoxil (ADV, a nucleotide analogue), the antiviral response of the different genotypes remains to be clarified. In order to evaluate the response of HBV genotypes to ADV therapy and to identify factors that might affect initial virological response, we performed a retrospective analysis on patients with chronic hepatitis B (CHB) in Chinese Han population. The study included 183 patients, who had been tested positive for hepatitis B e antigen (HBeAg) and had been treated with ADV (10 mg/day) for 48 weeks. The numbers of patients infected with HBV genotype B and genotype C were 98 and 75 cases, respectively, and the remaining 10 patients were mixture infection of genotypes B plus C or genotypes B plus D. The mean HBV-DNA reduction and HBV-DNA seroclearance of genotypes B and C at 48 weeks were 3.6 log(10) and 3.1 log(10) copies/ml (p < 0.05) and 41.8% and 34.6% (p < 0.05), respectively. There were no statistically significant differences between genotypes B and C in terms of HBeAg loss, anti-HBe seroconversion and normalization of serum alanine aminotransferase (ALT). Multivariate analysis showed that young age, low pretreatment HBV-DNA and/or elevated ALT level might be independent predictive factors associated with initial virological response. Thus, in Han CHB patients who are HBeAg-positive, HBV genotype B shows a better virological response to ADV therapy than does genotype C.

[Genotyping 238 HBV strains using type-specific primer PCR combined with type-specific nucleotide analysis].

OBJECTIVE: To establish a set of suitable and reliable methods for HBV genotyping and to study the distribution of HBV genotypes. METHODS: Type-specific nucleotides were searched through alignment of S genes (more than 1000 sequences) listed in GenBank. Then, type-specific primers were designed and type-specific primer PCR was used to genotype the 238 HBV strains. S genes of the untyped strains were further amplified and sequenced to find out their genotypes with type-specific nucleotide analysis. RESULTS: All the 238 HBV strains were genotyped. 159 (66.8%) cases were genotype B, 69 (28.9%) were genotype C, 6 (2.5%) were mixtures of genotypes B and C and 4 (1.6%) were mixtures of genotypes B and D. No genotypes of A, E, F, G, and H were found. CONCLUSION: Genotypes B and C are the most common types for HBV strains. Mixtures of genotypes B and C or genotypes B and D coinfection rarely existed. There is no relationship between the gender of the patients and HBV genotypes (X2 = 0.794, P more than 0.05).

[Comparison of antiviral responses to adefovir dipivoxil therapy of genotype B and genotype C HBV infected patients].

OBJECTIVE: To investigate the role of HBV genotypes on their response to adefovir dipivoxil (ADV) antiviral therapy. METHODS: HBV genotypes from 177 HBeAg-positive chronic hepatitis B (CHB) patients were identified and the patients were treated with ADV 10 mg per day for 48 weeks. The clinical data in terms of serum HBV DNA seroclearance, mean HBV DNA reduction (log value), HBeAg loss, anti-HBe seroconversion and serum ALT of those patients were analyzed against their HBV genotypes. RESULTS: Genotype B and genotype C were found in 102 and 65 cases, respectively. The mean HBV DNA reduction in patients with genotype B and genotype C at their treatment times of 12, 24 and 48 weeks was 2.2 log10copies/ml, 2.1 log10copies/ml (P more than 0.05), 2.7 log10copies/ml, 2.4 log10copies/ml (P more than 0.05) and 3.6 log10copies/ml, 3.1 log10copies/ml (P less than 0.05), respectively. At the end of the therapy (48 weeks), 43 (42.2%) patients with genotype B HBV infection and 22 (33.8%) patients with genotype C HBV infection had achieved HBV DNA seroclearance (P less than 0.05). CONCLUSIONS: Our results suggest that genotype B HBV has a better virological response to ADV therapy in HBeAg-positive chronic hepatitis B patients than that of genotype C. Longer terms of ADV treatment are needed to confirm this conclusion.

[Effect of Fertilizers on Cadmium Uptake and Accumulation by Sunflowers].

A pot experiment was conducted to assess the phytoextraction potential for cadmium (Cd) of three types of sunflowers (edible, ornamental, and oil sunflowers) and the effects of the application of N, NP, and NPK fertilizers on Cd uptake of the three plants using Cd-contaminated soils collected from northern China. The results showed that fertilization could significantly increase the biomass, aboveground Cd concentrations and accumulation of ornamental and oil sunflowers, and the effect of NPK fertilizer was significantly better than those of other treatments. Cd concentrations of the aboveground parts of edible, ornamental, and oil sunflowers were 6.89, 8.92, 6.97 mg·kg-1, respectively. Fertilization treatment significantly improved the transport ability of Cd of the three types of sunflowers, and bioconcentration factors of edible, ornamental, and oil sunflowers were 2.63 (control) to 3.10 (NPK fertilizer), 2.80 (control) to 4.02 (NPK fertilizer), and 2.11 (control) to 3.14 (NPK fertilizer), respectively. The results further showed that the metal-enriched granules and cellular debris were the main enrichment sources in the subcellular fraction of the three types of sunflowers (more than 55%). In summary, sunflowers can be considered as plant material for remediation of Cd-contaminated soil. In addition, NPK fertilizers can effectively improve the efficiency of sunflowers.

Histone deacetylation is involved in activation of CXCL10 upon IFNgamma stimulation.

Histone deacetylase (HDAC) activity is commonly associated with transcriptional repression. However, there is also evidence for a function in transcriptional activation. Previous studies have demonstrated a fundamental role of deacetylase activity in IFNa-responsive gene transcription. In the case of type II IFN (IFNgamma) results are controversial: some genes require HDAC activity, while transcription of others is repressed by HDAC. To investigate the effect of HDAC on transcription of an IFNgamma-activated gene, real-time PCR was used to measure CXCL10 mRNA in Hela cells stimulated with IFNgamma in the presence or absence of the HDAC inhibitor TSA. Chromatin imunoprecipitation combined with real-time PCR was used to check acetylation of histone H4 and recruitment of the STAT1 complex to the ISRE locus of the CXCL10 gene. Activation of CXCL10 transcription in response to IFNgamma was paralleled by a decrease in histone H4 acetylation and an increase in recruitment of the STAT1 complex to the CXCL10 ISRE locus. The transcription of CXCL10 and histone H4 deacetylation were blocked by TSA, but the latter had no obvious affect on recruitment of the STAT1 complex. Our data indicate that IFNgamma and STAT-dependent gene transcription requires the participation of HDAC, as does the IFNalpha-STAT pathway.

[The relationship between histone acetylation modification and IFN-gamma responsive gene regulation].

OBJECTIVES: To study the role of histone modification in the regulation of IFN-gamma-activated gene using chromatin immunoprecipitation technique. METHODS: Real-time PCR was used to measure the mRNA level of interferon-gamma-inducible protein 10 (IP-10) in Hela cells. Chromatin immunoprecipitation combined with Real-time PCR was used to check the histone H4 acetylation level at IFN-stimulated response element (ISRE) locus of IP-10 gene. RESULTS: IP-10 was strongly activated by IFN-gamma. The histone H4 deacetylation happened at the ISRE locus when IP-10 was induced by IFN-gamma. The activation of IP-10 and the deacetylation of histone H4 at the ISRE site induced by IFN-gamma were inhibited or blocked by histone deacetylase (HDAC) inhibitor trichostatin A (TSA). CONCLUSION: The histone H4 deacetylation at the ISRE site is related with the activation of IP-10 by IFN-gamma.

PNPLA3 rs738409 Polymorphism Associated with Hepatic Steatosis and Advanced Fibrosis in Patients with Chronic Hepatitis C Virus: A Meta-Analysis.

BACKGROUND/AIMS: The recognition of a correlation between patatin-like phospholipase domain containing-protein 3 (PNPLA3) rs738409 (C>G) and the severity of liver steatosis or fibrosis in chronic hepatitis C (CHC) has not reached a consensus. This meta-analysis sought to investigate with accuracy the association between the PNPLA3 rs738409 (C>G) polymorphism and liver steatosis and advanced fibrosis in CHC patients. METHODS: We performed a comprehensive literature search from the PubMed, Embase, Web of Science, and Google Scholar databases up to December 31, 2014. Crude odds ratios (ORs) with 95% confidence intervals (CIs) were calculated. Statistical analyses were performed using Stata 12.0 software. RESULTS: The meta-analysis revealed the severity of liver fibrosis was significantly higher in CHC patients with PNPLA3 rs738409 GG in Caucasians (versus CC+CG OR, 2.29; 95% CI, 1.57 to 3.35; p<0.05) but not Asian populations. In Caucasians, liver steatosis was also more severe in CHC patients with rs738409 GG (versus CC+CG; OR, 4.33; 95% CI, 2.59 to 7.22; p<0.05). The sensitivity analysis indicated the results of this meta-analysis were stable and no publication bias was detected. CONCLUSIONS: PNPLA3 rs738409 (C>G) was associated with the risk of both advanced liver fibrosis and steatosis in patients with CHC, especially among Caucasian populations.

[A method of HPRE synthesis via transcription by T7 RNA polymerase in vitro].

OBJECTIVE: To synthesize highly pure HBV post-transcriptional regulatory element (HPRE) via transcription in vitro by T7 RNA polymerase. METHODS: HPRE gene was amplified by PCR from a template containing HBV complete genomic DNA and cloned into plasmid pGEM-11zf. The cloned DNA sequence was transcribed by T7 RNA polymerase. RESULTS: The construction of HPRE gene recombinant plasmid and production of HPRE via transcription in vitro was successful. CONCLUSION: In vitro transcription by T7 RNA polymerase can be used to synthesize highly pure HPRE.

Association of genetic polymorphisms in CD8+ T cell inhibitory genes and susceptibility to and progression of chronic HBV infection.

BACKGROUND: Previous studies have shown that multiple inhibitory genes play an important role in HBV-specific CD8+ T cell exhaustion and dysfunction in the setting of chronic HBV infection. Polymorphic variants of these genes are thought to be predisposing factors for HBV susceptibility, clearance, and disease progression. The aim of this retrospective study was to identify variants affecting chronic HBV infection in a Chinese Han population. METHODS: We chose 28 tgSNPs from HapMap data on 5 key genes. They were genotyped on a total of 858 chronic HBV patients, 429 patients who underwent spontaneous recovery, and 239 healthy controls. We evaluated the correlation between the polymorphisms and HBV susceptibility, spontaneous clearance, and disease progression. RESULTS: The association of rs3827537 of BIM genotype TA and allele A was significantly different (P=0.016, OR=2.049; P=0.031, OR=1.925) between HBV patients and healthy controls. The rs36084323 of PD-1, as well as rs3766377, rs485618, rs4656942 of CD244 showed significant associations with the risk for HBV-related cirrhosis and hepatocellular carcinoma (HCC) (P=0.009, OR=0.482; P=0.009, OR=4.573; P=0.015, OR=0.580; P=0.028, OR=2.855). MDR analysis revealed that the four SNPs (rs36084323, rs3766377, rs485618, rs4656942) modulated the predisposition to cirrhosis and HCC in patients with chronic HBV infection (P=0.006). Using a luciferase reporter assay, we demonstrated that various alleles of rs3766377 had differential effects, and rs3766377 and rs485618 might have interactive effects. CONCLUSIONS: The present study reveals genetic associations among PD-1 and CD244 variants that may be involved in the development of cirrhosis and HCC in patients with chronic HBV infection. The BIM variant was associated with HBV susceptibility.