MSP analysis.

<p>Representative examples of MSP analysis for MLH1, MSH2, PMS2 and p16 methylation in HCC and non-tumour adjacent tissues were shown. Bisulfite-modified DNA was amplified using MSP primers specific to a CpG-rich region of each gene promoter. PCR-amplified products were resolved by 2% agarose gel electrophoresis. (U) Lanes represent amplification of unmethylated alleles, and (M) lanes contain only methylated alleles.</p

Frequency of promoter methylation.

<p>Frequency of promoter methylation.</p

Relationship between promoter methylation and clinicopathologic parameters of HCC patients (N = 61).

<p>Relationship between promoter methylation and clinicopathologic parameters of HCC patients (N = 61).</p

Frequency of promoter methylation.

<p>Promoter methylation of MLH1, PMS2, MSH2 and p16 was determined in HCC of different origins. As presented in bar graph, the frequency of methylation in the tested promoters and tissues was 19.8% in HCV infected, 16.3% in HBV infected and 25% in patients with alcoholic liver disease.</p

Primer sequences and annealing temperatures used in MSP.

<p>Primer sequences and annealing temperatures used in MSP.</p

Clinical feature of the studied patients.

<p>Clinical feature of the studied patients.</p

MSI analysis.

<p>Analysis of commonly used mononucleotide MSI marker loci BAT25 and BAT26 was performed in all patients showing promoter methylation in one of the tested MMR genes. Three examples are shown here. (T) HCC tissue, (NT) non-tumour adjacent tissue.</p

The expression of A20 in HCV-associated BCL.

<p><b>A</b>: Expression levels of A20 in the spleen from RzCD19Cre mice with or without BCL. Whole-tissue extracts prepared from the spleen in CD19Cre mice (control, n = 3; lanes 1–3 217–2, 2 224–2, 224–3), RzCD19Cre mice without BCL (HCV/Tumour(-), n = 5; lanes 4–8 217–3, 224–4, 232–3, 254–4, 240–2) and RzCD19Cre mice with BCL (HCV/Tumour(+), n = 5; lanes 9–13 24–1, 56–5, 69–5, 59–1, 43–4) were subjected to SDS-PAGE and were analysed by immunoblotting using anti-N terminal (A20N), anti-C terminal A20 (A20C), and anti-GAPDH antibodies. GAPDH was used as protein loading control. <b>B</b>: Quantitation of A20 (N and C), the average is indicated and statistical analysis was performed. Vertical bars indicate S.D.</p

The expression of genes involved in oncogenic pathways associated with BCL.

<p><b>A</b>: Highly modified gene signals in B cell lymphoma in RzCD19Cre mice BCL vs. B cells in RzCD19Cre male (Pair 1) or female (Pair 3) mice (left), and the genes modified by HCV expression in B cells in male (Pair 2) or female (Pair 4) (right). Red indicates the relative enhancement of the expression ratio of the processed signal (Test/Control, 532/635), and green indicates the relative reduction of expression. <b>B</b>: Quantification of Fos mRNA in HCV-, HCV+ B cells and HCV-Tg BCL in mice (numbers of individual mice were indicated) by quantitative RT-PCR. Fos mRNA was normalised against 18S rRNA, and the relative ratio was calculated. Vertical bars indicate S.D. <b>C</b>: Quantification of C3 mRNA in HCV-, HCV+ B cells and HCV-Tg BCL in mice. C3 mRNA was normalised against 18S rRNA, and relative ratio was calculated. Vertical bars indicate S.D. <b>D</b>: Quantification of LT βR mRNA in HCV-, HCV+ B cells and HCV-Tg BCL in mice by quantitative RT-PCR. RNA copies per total RNA (μg) were indicated and vertical bars indicate S.D.</p

Data from array performed once with mixed RNA samples (<b>Table 1</b>) were analysed using MetaCore software.

<p>Signals were analysed in the disease network (<b>A</b>) and in the process network (<b>B</b>) the values for the microarray data [(Feature number; yellow), (Process Signal (635); blue), (Process signal (532); red), Test/Control (532/635); green], (Process Signal (635); orange), (Process signal (532); purple)] are indicated by coloured bars. Abbreviations: BCL = B cell lymphoma. Refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0091373#pone-0091373-t001" target="_blank">Table 1</a> for construction of pairings.</p