Cytokine 73 (2015) 277–282

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IL-18 polymorphisms in hepatitis B virus related liver disease Vijay Kumar Karra a, Phani Kumar Gumma a, Soumya Jyoti Chowdhury a, Rajesh Ruttala a, Sunil Kumar Polipalli a, Anita Chakravarti b, Premashis Kar a,⇑ a b

PCR Hepatitis Lab, Dept. of Medicine, Maulana Azad Medical College, University of Delhi, New Delhi 110002, India Department of Medical Microbiology, Maulana Azad Medical College, University of Delhi, New Delhi 110002, India

a r t i c l e

i n f o

Article history: Received 19 November 2014 Received in revised form 13 February 2015 Accepted 14 February 2015

Keywords: Hepatitis B Interleukine-18 (IL-18) Single-nucleotide polymorphism (SNP)

a b s t r a c t Interleukine-18 (IL-18) was originally called interferon (INF-c) inducing factor and plays a critical dual role in Th1 polarization and viral clearance. We aimed to explore whether single-nucleotide promoter polymorphisms (SNPs) are associated with the outcome of hepatitis B virus (HBV) infection. 271 HBV infected patients were recruited in this study out of these 109 were spontaneously recovered and 162 were diagnosed to be having persistent HBV infection which includes 48 chronic hepatitis, 84 liver cirrhosis, 30 HCC cases and were compared with 280 healthy controls. IL-18 promoter genotyping was performed with sequence-specific primers. The results demonstrated the significant involvement of genotype AA at position -607 in healthy controls (38.6%) when compared to cases (26.0%) (OR = 0.54 (0.385–0.797)) and also associated with spontaneous clearance (37.6%) compared to persistent HBV infections (17.9%) (OR = 2.76 (1.582–4.832)). Whereas, genotype CC at position -607 in cases (18.0%) when compared to healthy controls (6.7%) (OR = 3.03 (1.734–5.303)) also associated with persistent HBV infections (24.1%) compared to spontaneous clearance (9.2%) (OR = 0.31 (0.151–0.67)). And genotype GC at position -137 in cases (49.5%) compared to healthy controls (38.5%) (OR = 1.55 (1.11–2.18)). Whereas, genotype GG at position -137 in healthy controls (56.8%) compared to cases (45.4%) (OR = 0.63 (0.451–0.885)). No significant difference at position -137 was observed between spontaneous clearance and persistent HBV infections. These polymorphisms of the IL-18 gene promoter region at position -607 and -137 could be associated with different outcomes of HBV infection. The people with allele A at position -607 may be protected against HBV infection; moreover AA genotype is associated with spontaneous clearance. Ó 2015 Elsevier Ltd. All rights reserved.

1. Introduction Hepatitis B virus (HBV) infection is a serious public health problem with more than 400 million HBV carriers around the world. Host factors determine the variable natural course of HBV infection, but much information regarding these factors are lacking [1,2]. The clinical outcome of HBV infection varies and possibly includes spontaneous recovery, hepatitis B surface antigen (HBsAg) carriers, chronic hepatitis (CHB), liver cirrhosis (LC) or hepatocellular carcinoma (HCC). Parenterally/sexually transmission is the major mode of HBV infection in endemic zones, including India [3,4]. In addition, the age at which the infection was Abbreviations: HBV, hepatitis B virus; CHB, chronic hepatitis; LC, liver cirrhosis; HCC, hepatocellular carcinoma; SNP, single nucleotide polymorphisms; IFN-c, interferon-c; PCR, polymerase chain reaction; SSP, sequence specific primer; OR, odds ratio; CI, confidence interval. ⇑ Corresponding author. Tel./fax: +91 11 23230132. E-mail address: [email protected] (P. Kar). http://dx.doi.org/10.1016/j.cyto.2015.02.015 1043-4666/Ó 2015 Elsevier Ltd. All rights reserved.

contracted effects the natural history of HBV infection [5] However, there are many other factors affecting the persistence of HBV infection after acute infection. There is an increasing evidence that host genetic factors influence the natural history of HBV infection [6]. Previous studies have shown that in various cytokine genes, the capacity of cytokine production differs among individuals and correlates with (SNPs) single nucleotide polymorphisms [7]. And also, SNPs in the cytokine gene were associated with the severity of the liver disease in HBV infections [8]. These may provide some new clues to clarify the mechanism of natural history of HBV infection. Cytokines were closely involved in both innate and adaptive immune response and defence against viral infection. Animal studies in transgenic mice have shown evidence that cytokine expression can inhibit HBV replication [9]. Interleukine-18 (IL-18) is a novel pro-inflammatory cytokine and known as immune activator involved in host defence mechanism. IL-18 was initially known as interferon-c (IFN-c) inducing factor which acts on Th 1 cells,

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natural killer cells (NK), nonpolarized T cells, dendritic cells, and B cells to produce interferon-c in the presence of IL-12 [10]. IL-18 has ability to induce IFN-c production in cytotoxic T lymphocytes and natural killer cells and plays a major role in the T cell helper type 1 (Th1) response [11]. IL-18 and IL-12 collectively induce IFN-c production at various different levels [12]. Inhibition of HBV replication is directly linked to interferon-c production by adoptively transfer CD8+ T cells [13]. After acute viral infection mature IL-18 is secreted from macrophages and HBV-specific T cells display a shift to type 1 T cells, and may contribute to effective clearance of hepatitis viruses [14]. Recent studied have shown IL18 gene promoter region regulates gene expression of this cytokine. Two SNPs at position -607 A/C and -137 C/G in the IL-18 gene promoter region were suggested to alter promoter activity (Fig. 1). Considering this evidence, we investigated the possible role of the SNPs of IL-18 gene promoter region in the natural history of HBV infections.

considered as spontaneously recovered, who had acute hepatitis B during the follow up and has spontaneously cleared the HBsAg over a period of 6 months, had evidence of HBV DNA (deoxyribonucleic acid) negativity, but were positive for antibody to hepatitis B surface antigen (anti-HBs) and antibody for hepatitis B core antigen (anti-HBc). In addition, 162 of the 271 patients were considered as persistent HBV infections such as chronic hepatitis B, liver cirrhosis and hepatocellular carcinoma, manifested by elevated liver function profile and by clinical or histological findings during the follow-up period. This group comprises of 48 chronic hepatitis B (38 male, 10 female; age mean ± SD, 28.65 ± 12.27 years), 84 liver cirrhosis (69 male, 15 female; age mean ± SD, 40.8 ± 14.6), and 30 HCC (27 male, 3 female; age mean ± SD, 55.2 ± 11.7) (Table 1). The study protocol was approved by institutional ethical committee of Maulana Azad Medical College, New Delhi, and informed consent was taken from all the patients. 2.2. DNA extraction

2. Material and methods 2.1. Patients and controls A total of 271 patients, with hepatitis B related liver disease were being followed up in the medical OPD of department of Medicine, Lok Nayak Hospital, New Delhi between February 2010, and February 2013, were enrolled in this study. Two hundred eighty healthy volunteers (157 male and 123 female) were included as control group who were HBV markers negative and were not exposed to HBV infection, we have excluded subjects who were HBsAg-negative and had anti-HBs, but not anti-HBc in the control. The patients were regularly followed with measurements of AST (aspartate aminotransferase), ALT (alanine aminotransferase), ALP (alkaline phosphatise) and HBV serological markers, like HBsAg, anti-HBs, anti-HBc IgM, anti-HBc IgG, HBeAg and anti-HBe, using commercially available ELISA kits every 3– 6 months. Ultrasonography or computed tomography, serum HBV-DNA was measured every 3–6 months. Of 271 patients, 109 (82 male, 27 female; age mean ± SD, 29.8 ± 11.7 years) were

Fig. 1. The schematic representation of IL-18 gene with promoter regions analyzed.

2 ml of peripheral venous blood was collected from all the patients after an overnight fasting in EDTA tube. Genomic DNA was extracted from whole blood by using QIAamp Blood kit (Qiagen, Germantown, USA) according to the manufacturer’s instructions and frozen at 20°. 2.3. Determination of IL-18 (-607/-137) genotypes Polymorphism at position -607 and -137 in the promoter region of IL-18 gene was analyzed by polymerase chain reaction with sequence specific primers (PCR-SSP) method [15]. For the position -607 A/C specific PCR, a common reverse primer and 2 sequencespecific primers were used. An amplified product of 196 bp was detected. A control forward primer was used to detect an amplified product of 301 bp fragment and acts as an internal positive amplification control (Table 1). PCR reaction was carried out in a final volume of 20 lL consisting of 2.0 lL 10 PCR buffer, 10 mmol/L dNTP, 100 ng/lL genomic DNA and 1 lL of Taq polymerase (5 U/lL). One sequence specific primer (for allele A or allele C) and common reverse primer with a concentration of 0.5 lmol/L was included in every reaction mixture. In addition, internal positive control primer was added to the reaction mixture. Therefore, for every individual two PCR reactions were performed. PCR reactions were carried out in Biometra T-Gradient thermal cycler (Goettingen Germany). The PCR conditions were as follows; denaturation at 94 °C for 2 min, followed by seven cycles for 20 s at 94 °C, 40 s at 64 °C and 40 s at 72 °C and 25 cycles for 20 s at 94 °C, 40 s at 57 °C, 40 s at 72 °C and 72 °C for 5 min. PCR products were examined by 2% agarose gel electrophoresis stained with ethidium bromide and documented. For the position -137 C/G specific PCR, a common reverse primer and 2 sequence specific forward primers were used. An amplified product of 261 bp was

Table 1 Primers for IL-18 gene promoter polymorphism. SNP

Primer

Sequence

Product size

-607 A/C-specific PCR

Forward primer Reverse primer Control forward primer Common reverse primer

50 -GTTGCAGAAAGTGTAAAAATTATTAC-30 50 -GTTGCAGAAAGTGTAAAAATTATTAA-30 50 -CTTTGCTATCATTCCAGGAA-30 50 -TAACCTCATTCAGGACTTCC-30

196-bp

Forward primer Reverse primer Control forward primer Common reverse primer

50 -CCCCAACTTTTACGGAAGAAAAG-30 50 -CCCCAACTTTTACGGAAGAAAAC-30 50 -CCAATAGGACTGATTAT TCCGCA-30 50 -AGGAGGGCAAAATG CACTGG-30

261-bp

-137 C/G-specific PCR

301-bp

446-bp

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detected. A control forward primer was used to detect an amplified product of 446 bp fragment and acts as an internal positive amplification control (Table 1). PCR reaction was carried out in a final volume of 20 lL consisting of 2.0 lL 10 PCR buffer, 10 mmol/L dNTP, 100 ng/lL genomic DNA and 1 lL of Taq polymerase (5 U/lL). One sequence specific primer (for allele C or allele G) and common reverse primer with a concentration of 0.5 lmol/L was included in every reaction mixture. In addition, internal positive control primer was added to the reaction mixture. The PCR conditions were as follows; denaturation at 94 °C for 2 min, followed by five cycles for 20 s at 94 °C, 1 min at 68 °C and 25 cycles for 20 s at 94 °C, 40 s at 62 °C, 40 s at 72 °C and 72 °C for 5 min. 2.4. Statistical analysis Comparison of genotypic and allelic frequencies in the IL-18 gene promoter region at position -607 and -137 were performed using chi-square or the Fisher exact test. Data were analyzed with SPSS 20.0 software. Clinical and biochemical results are expressed as mean ± SD. The Hardy–Weinberg equilibrium was determined by chi-square test. Odds ratio (OR) was calculated by logistic regression with 95% confidence interval (CI) level. P values less than 0.05 was considered statistically significant.

3. Results 3.1. Demographic profile The baseline clinical and biochemical characteristics of the cases and healthy controls in the study groups were summarized in Table 2. We have investigated the distribution of IL-18 gene promoter polymorphism at -607 (A/C) and -137 (C/G) in 271 north Indian HBV infected patients and 280 healthy controls.

3.2. Correlation of IL-18 promoter polymorphism at position -607 Genotypic and allelic frequencies at position -607 were compared between HBV cases and healthy controls and summarized in Table 3. Of 271 patients, 70 had the AA type (26%), 152 had the AC type (56%) and 49 the CC (18.0%). Of the 280 healthy controls, 102 had the AA type (36.4%), 144 had the AC type (51.4%) and 34 the CC (12.1%). There was a significant difference in the genotype and allele frequency between cases and the healthy controls. In genotypes, the AA type was present at a significantly higher frequency in the healthy controls compared to those in the cases (36.4% vs. 26.0%, v2 = 7.20, P = 0.007, OR = 0.607 (0.421– 0.875), and the CC type was present at significantly higher frequency in cases compared to those in the healthy control (18.0% vs. 12.1%, v2 = 3.79, P = 0.05, OR = 1.597 (0.994–2.564). But in phenotypes, the allele A at -607 was of significantly higher frequency in healthy controls than that in the cases (62.1% vs. 53.8%, v2 = 7.73, P = 0.005, OR = 0.711 (0.559–0.904). 3.3. Correlation of IL-18 promoter polymorphism at position -137 Genotypic and allelic frequencies at position -137 were compared between HBV cases and healthy controls and summarized in Table 3. Of 271 patients, 14 had the CC type (5.1%), 134 had the GC type (49.5%) and 123 the GG (45.4%). Of the 280 healthy controls, 13 had the CC type (4.7%), 108 had the GC type (38.5%) and 159 the GG (56.8%). There was a significant difference in the genotype and allele frequency between cases and the healthy controls. In genotypes, the GC type was present at a significantly higher frequency in the cases compared to those in the healthy controls (49.5% vs. 38.5%, v2 = 6.613, P = 0.01, OR = 1.558 (1.11– 2.185), and the GG type was present at significantly higher frequency in healthy control compared to those in the cases (56.8% vs. 45.4%, v2 = 7.161, P = 0.007, OR = 0.632 (0.451–0.885). But in

Table 2 Clinical and biochemical characteristics of HBV cases and healthy controls.

Gender Male/female Age (years) Mean ± SD AST (IU/L) ALT (IU/L) ALP (IU/L)

Spontaneously recovered (n = 109)

Chronic hepatitis B (n = 48)

Liver cirrhosis (n = 84)

HCC (n = 30)

Healthy control (n = 280)

82/27

38/10

69/15

27/3

157/123

29.8 ± 11.7 35.6 ± 16.4 46.8 ± 27.0 160.4 ± 69.0

28.65 ± 12.27 98.7 ± 244.5 120.4 ± 302.2 218.9 ± 100.1

40.8 ± 14.6 81.9 ± 51.2 65.0 ± 41.8 169.4 ± 72.3

55.2 ± 11.7 73.3 ± 47.5 109 ± 105.2 228.7 ± 183.2

30.9 ± 9.1 – – –

AST, aspartate aminotransferase; ALT, alanine aminotransferase; ALP, alkaline phosphatise.

Table 3 Comparison of IL-18 gene promoter polymorphism between cases and healthy controls. Polymorphism 607 position Genotypes AA AC CC Alleles A C 137 position Genotypes CC GC GG Alleles C G

Cases n = 271 (%)

Healthy controls n = 280 (%)

v2

P

OR (95%) CI

70 (26.0) 152 (56.0) 49 (18.0)

102 (36.4) 144 (51.4) 34 (12.1)

7.20 1.20 3.79

0.007 0.273 0.051

0.607 (0.421–0.875) 1.206 (0.862–1.687) 1.597 (0.994–2.564)

292 (53.8) 250 (46.2)

348 (62.1) 212 (37.8)

7.73

0.005

0.711 (0.559–0.904)

14 (5.1) 134 (49.5) 123 (45.4)

13 (4.7) 108 (38.5) 159 (56.8)

0.080 6.613 7.161

0.776 0.010 0.007

1.119 (0.515–2.426) 1.558 (1.11–2.185) 0.632 (0.451–0.885)

162 (29.8) 380 (70.2)

134 (24.0) 426 (76.0)

4.981

0.025

1.355 (1.037–1.771)

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phenotypes, the allele C at -607 was of significantly higher frequency in cases than that in the healthy control (29.8% vs. 24.0%, v2 = 4.981, P = 0.025, OR = 1.355 (1.037–1.771). 3.4. Distribution of IL-18 promoter polymorphisms (-607 and -137) in HBV infections The genotype frequencies at position -607 and -137 in each group of HBV infections were summarized in Table 4. With regard to position -607, 41 (37.6%) spontaneously recovered patients had AA genotype, 58 (53.2%) had the AC genotype, and 10 (9.2%) had the CC genotype. Of the 48 chronic hepatitis B patients 10 (20.8%) had the AA genotype, 36 (75.0%) had the AC genotype, and 2 (4.2%) had the CC genotype. Of the 84 liver cirrhosis patients 12 (14.3%) had the AA genotype, 38 (45.3%) had the AC genotype, and 34 (40.4%) had the CC genotype. And of the 30 HCC patients 7 (23.3%) had the AA genotype, 20 (66.7%) had the AC genotype, and 3 (10.0%) had the CC genotype. With regard to the position 137, 5 (4.5%) spontaneously recovered patients had CC genotype, 47 (43.2%) had the GC genotype, and 57 (52.3%) had the GG genotype. Of the 48 chronic hepatitis B patients 2 (4.2%) had the CC genotype, 27 (56.3%) had the GC genotype, and 19 (39.5%) had the GG genotype. Of the 84 liver cirrhosis patients 6 (7.1%) had the CC genotype, 42 (50.0%) had the GC genotype, and 36 (42.9%) had the GG genotype. And of the 30 HCC patients 1 (3.3%) had the CC genotype, 18 (60.0%) had the GC genotype, and 11 (36.7%) had the GG genotype (Table 4). 3.5. Correlation of IL-18 promoter polymorphisms (-607 and -137) in spontaneously recovered and persistent HBV infections These 271 HBV infected patients were divided into 2 groups; 109 patients were considered as spontaneously recovered and 162 were found to have persistent HBV infection (Table 5). With regard to position -607, there was significant difference in the

Table 4 Distribution of genotypic frequency in patients with HBV infections. Spontaneously recovered n = 109 (%)

Chronic hepatitis B n = 48 (%)

Liver cirrhosis n = 84 (%)

HCC n = 30 (%)

Locus-607 AA AC CC

41 (37.6) 58 (53.2) 10 (9.2)

10 (20.8) 36 (75.0) 2 (4.2)

12 (14.3) 38 (45.3) 34 (40.4)

7 (23.3) 20 (66.7) 3 (10.0)

Locus-137 CC GC GG

5 (4.5) 47 (43.2) 57 (52.3)

2 (4.2) 27 (56.3) 19 (39.5)

6 (7.1) 42 (50.0) 36 (42.9)

1 (3.3) 18 (60.0) 11 (36.7)

Genotypes

genotype frequency between spontaneously recovered cases and the persistent HBV infections. The AA genotype was present at a significantly higher frequency in the spontaneously recovered patients compared to those in the persistent HBV infection (37.6% vs. 17.9%, v2 = 13.22, P = 0.0002, OR = 2.765 (1.582–4.832). and the CC genotype was present at significantly higher frequency in the persistent HBV infection compared to those in the spontaneously recovered patients (24.1% vs. 9.2%, v2 = 9.766, P = 0.001, OR = 0.318 (0.151–0.67). With regard to position -137, no significant difference was found between the spontaneously recovered patients and persistent HBV infections (Table 5).

4. Discussion There are several factors governing the susceptibility to persistent HBV infection, which includes age at the time of infection. When acquired during the early neonatal period from HBV infected mother, only 10% of the children will resolve the virus. Whereas, when acquired during childhood or later, up to 90% will resolve the virus spontaneously [16]. In addition, people with same ethnic groups, age and sex exposed to the same HBV strain, shows broad spectrum of HBV infections with different clinical outcomes. These information including with twin studies suggest that host genetic background influences the clinical outcome of HBV infection [17]. A coordinated innate and adaptive humoral, cell-mediated immune response is required to clear the HBV infection [18]. Previous studies have demonstrated that cytokine production varies among individuals and correlates with the SNPs in the promoter region of cytokine gene [7]. Moreover, Disease progression in HBV infections have been seen to be associated with cytokine gene promoter polymorphism [19]. These factors may reveal some clues to understand the mechanism between spontaneous clearance in some people, whereas development of end stage complications in some others such as liver cirrhosis and hepatocellular carcinoma. IL-18 activates effecter cells which are involved in the inter cellular interactions that occur during the time of inflammation, and leads to activities against pathogens, part of the stages of acute and chronic viral hepatitis, and induces targeted cells apoptosis [20,21]. In a previous study, it was reported that in the liver of transgenic mice, after injection of a single dosage of 10 lg recombinant murine IL-18 rapidly, reversibly and non-cytopathically inhibited HBV replication. IL-18 mediates anti-viral effect by its ability to activate intra hepatic natural killer cells and T cells to produce IFN-c and its ability to induce IFN-a/b production in the liver [22]. These findings suggest that IL-18 potentially contributes to the control of HBV replication during self limited infection and it may have therapeutic value for the treatment. In this present study, we have compared the distribution of IL-18 gene promoter polymorphisms among Indian patients with

Table 5 Distribution of IL-18 genotype in spontaneously recovered and persistent HBV infections. Spontaneously recovered n = 109 (%)

Persistent HBV infections n = 162 (%)

v2

P

OR (95%) CI

607 position Genotypes AA AC CC

41 (37.6) 58 (53.2) 10 (9.2)

29 (17.9) 94 (58.0) 39 (24.1)

13.22 0.613 9.766

0.0002 0.433 0.001

2.765 (1.582–4.832) 0.822 (0.504–1.341) 0.318 (0.151–0.67)

137 position Genotypes CC GC GG

5 (4.5) 47 (43.2) 57 (52.3)

9 (5.6) 87 (53.7) 66 (40.7)

0.124 2.92 3.508

0.724 0.087 0.061

0.817 (0.266–2.508) 0.653 (0.400–1.066) 1.594 (0.977–2.601)

Polymorphism

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different clinical outcome of HBV infection. Our results demonstrated that the frequencies of the -607 AA genotype and the 607 A allele were significantly higher in the healthy controls compared with those in the patients. Whereas, the frequencies of -607 CC genotype were significantly higher in the patients compared to healthy controls. These results suggest that the -607 AA genotype and the -607 A allele have a protective effect on HBV infections. Moreover, when we compared the spontaneously recovered with the persistent HBV infection group we have found interesting results, the frequencies of the -607 AA genotype were significantly higher in spontaneously recovered compared with those in persistent HBV infection suggesting a protective effect of this genotype on the diseases progression and the frequencies of -607 CC genotype were significantly increased in the persistent HBV infection than the spontaneously recovered. These results suggest that 607 AA genotype was associated with a reduced risk of disease progression in HBV infection (OR, 2.76), the -607 CC genotype was associated with an increased risk of disease progression in HBV infection (OR, 0.31). And the frequencies of -137 GC genotype and the -137 C allele were significantly higher in the patients compared with those in the healthy control. Whereas, the frequencies of -137 GG genotype were significantly higher in the healthy controls compared to patients. These results suggest that the -137 GC genotype and the -137 C allele have a susceptible effect on HBV infections. And the -137 GG genotype have protective effect. There was no significant difference seen between spontaneously recovered and persistent HBV infection at position -137. The mechanism beyond this differential effect of these genotypes with the development of HBV infection was not clear in this study. It was reported that these two polymorphisms -607 A/C and -137 C/G were in strong linkage disequilibrium [23]. The C allele at position -607 (CC + C/A) has been seen to be associated with increased risk of cirrhosis and hepatocellular in HBV patients [24]. We have found similar results with high frequency of -607 CC genotype with 34 (40.4%) in liver cirrhosis and 3 (10%) in HCC patients. These findings address the question as how IL-18 polymorphisms are related to HBV infections. Giedraitis et al. demonstrated experimentally that the allele C at -137 disrupt the confirmed H4TF-binding site, while nucleotide substitution at -607 (C/A) may disrupt the potential cyclic-adenosine-monophosphate-responsive element-binding site [41]. Moreover, transcription activity assay at IL-18 promoter site demonstrated that the presence of both A and C allele at positions -607 and -137 in the same haplotype is associated with low promoter activity. The fundamental mechanism for the relationship between the IL-18 gene promoter polymorphism and the outcome of the HBV infection were not clearly understood in our study. Zhang et al. reported that the AA genotype at position -607 may be associated with HBV-DNA replication, and the presence of allele C at position -137 may play protective role in the development of HBV infection [15]. Our results may not be in concordance with the previous findings of IL-18 promoter polymorphism. These polymorphisms may vary with the ethnicity of the individual. In various populations, IL-18 gene promoter polymorphism at either -607 or -137 position, or both has been shown to be associated in many diseases. For example Systemic lupus erythematosus [25], Rheumatoid arthritis [26], Hepatocellular carcinoma [31], Human immunodeficiency virus (HIV) [38], Gastrointestinal cancers [39], and viral infections like Hepatitis C (HCV) [36,37] (Table 6). In conclusion, we have attempted to elucidate the role of IL-18 gene promoter polymorphism in the outcome of HBV infection. Our data suggest that the carriage of allele A at position -607 in the promoter may play a protective role in the development of HBV infection and AA genotype at position -607 may be associated with spontaneous clearance of HBV infection. And the allele C at position -137 is associated with HBV infection and GC genotype

Table 6 Association of IL-18 gene promoter polymorphism with different diseases and disease severity in various populations. Disease

Population

Gene position

Disease outcome

Systemic lupus erythematosus Rheumatoid arthritis Behcet’s disease Breast cancer Primary immune thrombocytopenia Multiple sclerosis Hepatocellular carcinoma Cervical squamous cell carcinoma Diabetes mellitus Bladder cancer Chronic obstructive pulmonary disease Hepatitis C (HCV) Human immunodeficiency virus (HIV) Gastrointestinal cancers Ovarian cancer

Egyptian [25]

-607, -137

Associated

Indian [26] Egyptian [27] Brazilian [28] Chinese [29]

-607, -137 -137 -607, -137 -607, -137

Associated Associated Associated No association

Turkish [30] Brazilian [31] Taiwanese [32]

-137 -607, -137 -607, -137

Associated Associated Associated

Brazilian [33] Indian [34] Chinese [35]

-607, -137 -137 -607

Associated Associated Associated

Tunisian [36,37] Brazilian [38]

-607, -137 -607

Associated Associated

Iranian [39] Iranian [40]

-137 -607, -137

Associated No association

at position -137 is associated with persistent HBV infections. However, some limitations exist in this study. Male to female ratio in cases and controls was not matched. To establish the real role of IL-18 gene promoter polymorphism and its relation in the outcome of HBV infections, more large-scale population-based studies are required.

Acknowledgments This study was funded by Indian Council of Medical Research, Government of India. The author thanks Dr. Anita chakravarthi HOD, Department of Medical Microbiology, Faculty of Medical sciences, University of Delhi, Maulana Azad Medical College, New Delhi, India, for critical input. And the research staff worked under this project.

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