Ficolin-2 Levels and FCN2 Haplotypes Influence Hepatitis B Infection Outcome in Vietnamese Patients

Human Ficolin-2 (L-ficolins) encoded by FCN2 gene is a soluble serum protein that plays an important role in innate immunity and is mainly expressed in the liver. Ficolin-2 serum levels and FCN2 single nucleotide polymorphisms were associated to several infectious diseases. We initially screened the complete FCN2 gene in 48 healthy individuals of Vietnamese ethnicity. We genotyped a Vietnamese cohort comprising of 423 clinically classified hepatitis B virus patients and 303 controls for functional single nucleotide polymorphisms in the promoter region (-986G>A, -602G>A, -4A>G) and in exon 8 (+6424G>T) by real-time PCR and investigated the contribution of FCN2 genotypes and haplotypes to serum Ficolin-2 levels, viral load and liver enzyme levels. Haplotypes differed significantly between patients and controls (P = 0.002) and the haplotype AGGG was found frequently in controls in comparison to patients with hepatitis B virus and hepatocellular carcinoma (P = 0.0002 and P<0.0001) conferring a protective effect. Ficolin-2 levels differed significantly between patients and controls (p<0.0001). Patients with acute hepatitis B had higher serum Ficolin-2 levels compared to other patient groups and controls.The viral load was observed to be significantly distributed among the haplotypes (P = 0.04) and the AAAG haplotype contributed to higher Ficolin-2 levels and to viral load. Four novel single nucleotide polymorphisms in introns (-941G>T, -310G>A, +2363G>A, +4882G>A) and one synonymous mutation in exon 8 (+6485G>T) was observed. Strong linkage was found between the variant -986G>A and -4A>G. The very first study on Vietnamese cohort associates both Ficolin-2 serum levels and FCN2 haplotypes to hepatitis B virus infection and subsequent disease progression

Primers and probes used for genotyping and sequencing.

<p>FL: Fluorescein; Cy5: Cyanine 5.18; PH: Phosphate group; <b>^(*)</b>: sequencing primers</p

Association of viral load and <i>FCN2</i> haplotypes in HBV patients.

<p>Box-plots illustrate medians with 25 and 75 percentiles with whiskers to 10 and 90 percentiles. The number in parenthesis indicates number of observed haplotypes. <i>P</i> values were calculated by Kruskal- Wallis test.</p

Distribution of <i>FCN2</i> haplotypes (-986/-602/-4/+6424) in hepatitis B patients and controls.

<p>AHB: acute hepatitis B; ASYM: asymptomatic hepatitis B carriers; CHB: chronic hepatitis B; HCC: hepatocellular carcinoma; LC: liver cirrhosis. n =  Number of chromosomes; NA: not applicable; NS: not significant.</p

Ficolin-2 serum levels in HBV patients and controls.

<p>Box-plots illustrate medians with 25 and 75 percentiles with whiskers to 10 and 90 percentiles; <i>P</i> value was calculated by t-test. Panel (A): Ficolin-2 serum levels were segregated based on patient groups including acute hepatitis B (AHB), asymptomatic HBV carriers (ASYM), chronic hepatitis B (CHB), hepatocellular carcinoma (HCC), liver cirrhosis (LC) and controls. Statistical analysis was performed using contingency table analysis with one-way ANOVA and P-values were illustrated. Panel (B): Ficolin-2 serum levels in HBV-infected patients with acute and chronic HBV (ASYM/CHB/HCC/LC). Panel (C): Ficolin-2 serum levels in HBV-infected patients with cancer (HCC) and without cancer (AHB/ASYM/CHB/LC).</p

Distribution of <i>FCN2</i> gene variants in Vietnamese individuals (n = 48).

<p>Distribution of <i>FCN2</i> gene variants in Vietnamese individuals (n = 48).</p

Characteristics of Vietnamese HBV patients and control individuals.

<p>AHB =  acute hepatitis B; CHB =  chronic hepatitis B; LC =  liver cirrhosis; HCC =  hepatocellular carcinoma;</p><p>AST and ALT =  aspartate and alanine amino transferase; IU =  international units; NA =  Not available. Values given are medians.</p><p>*P<0.001 significantly different among patient groups.</p