Journal: BMC Molecular and Cell Biology

Article Title: Minicircle DNA vector expressing interferon-lambda-3 inhibits hepatitis B virus replication and expression in hepatocyte-derived cell line

doi: 10.1186/s12860-020-00250-9

Figure Lengend Snippet: MC.IFNλ3 permits hepatocyte-specific expression of IFNλ3. HepG2.2.15, HEK293 and Hela cells were transfected with MC vectors. a Schematic illustration of the MC.IFNs. MC.IFNα is 1656-bp in length, MC.IFNλ3 is 1677-bp in length. attR represents a 36-bp attR recombinant site. ApoE indicates ApoE promoter. CDS represents coding sequence. bpA represents bovine growth hormone polyadenylation signal. b The expression of IFNα and IFNλ3 in cell lysate was determined by Western Blot at 3 days post-transfection. Lane 1–5 represents the untreated control (HepG2.2.15 cells without MC transfection), MC.IFNα transfected HepG2.2.15 cells, and MC.IFNλ3 transfected HepG2.2.15 cells, MC.IFNλ3 transfected HEK293 cells, MC.IFNλ3 transfected Hela cells, respectively

Article Snippet: After blocked the non-specific binding sites with 5% skim milk in TBST (Sigma, US), the membrane was subjected to immunoblotting using a primary antibody listed as below: the rabbit polyclonal antibody specific to IFN⍺ (ProteinTech, US; #18013–1-AP) and IFNλ3 (ProteinTech, US; #24199–1-AP); the rabbit monoclonal antibodies specific to JAK1 (Cell Signaling Technology, US; #3344) and phosphorylated JAK1 (p-JAK1) (Cell Signaling Technology, US; #3331); the rabbit polyclonal or monoclonal antibodies specific to STAT1 (Abcam, UK; #ab2415), STAT2 (Abcam, UK; #ab53149), phosphorylated STAT1 (p-STAT1) (Cell Signaling Technology, US; #9171) and phosphorylated STAT2 (p-STAT2) (Millipore, US; #07–224).

Techniques: Expressing, Transfection, Recombinant, Sequencing, Western Blot, Control

Journal: BMC Molecular and Cell Biology

Article Title: Minicircle DNA vector expressing interferon-lambda-3 inhibits hepatitis B virus replication and expression in hepatocyte-derived cell line

doi: 10.1186/s12860-020-00250-9

Figure Lengend Snippet: MC.IFNλ3 inhibits viral antigens expression and viral DNA replication in HepG2.2.15 cells. HepG2.2.15 cells were transfected with MC.IFNλ3 and MC.IFNα. While the untreated HepG2.2.15 cells served as a blank control (Blank). The levels of viral antigens, namely HBsAg ( a ) and HBeAg ( b ), and viral DNA in cell culture supernatant were determined by chemiluminiscence and qPCR, respectively, at the indicated time-points (3 or 6 days post-transfection). All data are shown as mean ± SD from three independent experiments. * indicates statistically significant ( P -value < 0.05), ns indicates not significant ( P -value > 0.05)

Article Snippet: After blocked the non-specific binding sites with 5% skim milk in TBST (Sigma, US), the membrane was subjected to immunoblotting using a primary antibody listed as below: the rabbit polyclonal antibody specific to IFN⍺ (ProteinTech, US; #18013–1-AP) and IFNλ3 (ProteinTech, US; #24199–1-AP); the rabbit monoclonal antibodies specific to JAK1 (Cell Signaling Technology, US; #3344) and phosphorylated JAK1 (p-JAK1) (Cell Signaling Technology, US; #3331); the rabbit polyclonal or monoclonal antibodies specific to STAT1 (Abcam, UK; #ab2415), STAT2 (Abcam, UK; #ab53149), phosphorylated STAT1 (p-STAT1) (Cell Signaling Technology, US; #9171) and phosphorylated STAT2 (p-STAT2) (Millipore, US; #07–224).

Techniques: Expressing, Transfection, Control, Cell Culture

Journal: BMC Molecular and Cell Biology

Article Title: Minicircle DNA vector expressing interferon-lambda-3 inhibits hepatitis B virus replication and expression in hepatocyte-derived cell line

doi: 10.1186/s12860-020-00250-9

Figure Lengend Snippet: Viral antigens and viral DNA in HepG2.2.15 cell culture supernatant after transfection

Article Snippet: After blocked the non-specific binding sites with 5% skim milk in TBST (Sigma, US), the membrane was subjected to immunoblotting using a primary antibody listed as below: the rabbit polyclonal antibody specific to IFN⍺ (ProteinTech, US; #18013–1-AP) and IFNλ3 (ProteinTech, US; #24199–1-AP); the rabbit monoclonal antibodies specific to JAK1 (Cell Signaling Technology, US; #3344) and phosphorylated JAK1 (p-JAK1) (Cell Signaling Technology, US; #3331); the rabbit polyclonal or monoclonal antibodies specific to STAT1 (Abcam, UK; #ab2415), STAT2 (Abcam, UK; #ab53149), phosphorylated STAT1 (p-STAT1) (Cell Signaling Technology, US; #9171) and phosphorylated STAT2 (p-STAT2) (Millipore, US; #07–224).

Techniques: Cell Culture, Transfection, Control

Journal: BMC Molecular and Cell Biology

Article Title: Minicircle DNA vector expressing interferon-lambda-3 inhibits hepatitis B virus replication and expression in hepatocyte-derived cell line

doi: 10.1186/s12860-020-00250-9

Figure Lengend Snippet: MC.IFNλ3 induce JAK1 and STAT1/STAT2 phosphorylation in HepG2.2.15 cells. HepG2.2.15 cells were transfected with MC vectors. The levels of a STAT1/STAT2 proteins and their phosphorylated form (p-STAT1/p-STAT2), b JAK1 and phosphorylated JAK1 (p-JAK1) in transfected HepG2.2.15 cells were determined by Western Blot at 6 days post-transfection. Lane 1, 2 and 3 represents untreated Control, MC.IFNα, and MC.IFNλ3 group, respectively

Article Snippet: After blocked the non-specific binding sites with 5% skim milk in TBST (Sigma, US), the membrane was subjected to immunoblotting using a primary antibody listed as below: the rabbit polyclonal antibody specific to IFN⍺ (ProteinTech, US; #18013–1-AP) and IFNλ3 (ProteinTech, US; #24199–1-AP); the rabbit monoclonal antibodies specific to JAK1 (Cell Signaling Technology, US; #3344) and phosphorylated JAK1 (p-JAK1) (Cell Signaling Technology, US; #3331); the rabbit polyclonal or monoclonal antibodies specific to STAT1 (Abcam, UK; #ab2415), STAT2 (Abcam, UK; #ab53149), phosphorylated STAT1 (p-STAT1) (Cell Signaling Technology, US; #9171) and phosphorylated STAT2 (p-STAT2) (Millipore, US; #07–224).

Techniques: Phospho-proteomics, Transfection, Western Blot, Control

Journal: BMC Molecular and Cell Biology

Article Title: Minicircle DNA vector expressing interferon-lambda-3 inhibits hepatitis B virus replication and expression in hepatocyte-derived cell line

doi: 10.1186/s12860-020-00250-9

Figure Lengend Snippet: MC.IFNλ3 up-regulates ISGs expression in HepG2.2.15 cells. MC.IFNλ3 up-regulates ISGs expression in HepG2.2.15 cells. The relative mRNA transcriptional levels of ten ISGs MC transfected HepG2.2.15 cells were quantified at 3 or 6 days post-transfection by qPCR. The ISGs mRNA levels in HepG2.2.15 cells after MC.IFNλ3 ( a ) and MC.IFNα ( b ) treatment were compared between 3 days and 6 days post-transfection groups. The ISGs mRNA levels in HepG2.2.15 cells between MC.IFNλ3 and MC.IFNα treatment groups were compared at 3 days ( c ) or 6 days ( d ) post-transfection. All data are shown as mean ± SD from three independent experiments

Article Snippet: After blocked the non-specific binding sites with 5% skim milk in TBST (Sigma, US), the membrane was subjected to immunoblotting using a primary antibody listed as below: the rabbit polyclonal antibody specific to IFN⍺ (ProteinTech, US; #18013–1-AP) and IFNλ3 (ProteinTech, US; #24199–1-AP); the rabbit monoclonal antibodies specific to JAK1 (Cell Signaling Technology, US; #3344) and phosphorylated JAK1 (p-JAK1) (Cell Signaling Technology, US; #3331); the rabbit polyclonal or monoclonal antibodies specific to STAT1 (Abcam, UK; #ab2415), STAT2 (Abcam, UK; #ab53149), phosphorylated STAT1 (p-STAT1) (Cell Signaling Technology, US; #9171) and phosphorylated STAT2 (p-STAT2) (Millipore, US; #07–224).

Techniques: Expressing, Transfection

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Frequencies of PD‐1 and IFNL4 genotypes in patients with HCV‐related diseases and in blood donors

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques:

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Genomic positions of IFNL4 genetic variants in samples homozygous at rs12979860. In addition to rs12979860, eight different SNPs were identified in 24 of 36 samples. Three SNPs (including one in exon 3) have not previously been reported. The table below the gene diagram indicates the genotype at each polymorphism for all 36 samples

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques:

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Frequency of IFNL4 genetic variants associated with IFNλ4 in patients and blood donors

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques: Expressing, Mutagenesis

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Boxplot describing the relationship of IFNL4 genotype related to a fully active IFNL4 secretion with a younger median age of HCC patients. HCC patients carrying a fully active (rs12979860‐T/T combined without P70S mutation, rs117648444 C/C) were diagnosed with tumour at younger age compared to those with a reduced expression (rs12979860‐T/T combined with P70S mutation, rs117648444 C/T or T/T), median age 61y and 72y, respectively, P = .02 ANOVA test. Boxes range from the 25th to the 75th percentile with a horizontal black line at the median and vertical lines extending to the 10th and 90th percentiles. These data suggest that IFNL4 levels may represent a risk factor for HCC development

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques: Mutagenesis, Expressing

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Pairwise linkage disequilibrium (LD) relationships between the PD‐1.3, PD‐1.5, PD‐1.7 and IFNL4 rs12979860 polymorphism. A–F, Results from linkage analysis conducted for the polymorphism of blood donors (A), CHC (B), cirrhosis (C), HCC (D), MC (E) and NHL (F) respectively

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques:

Journal: Liver International

Article Title: PDCD1 and IFNL4 genetic variants and risk of developing hepatitis C virus‐related diseases

doi: 10.1111/liv.14667

Figure Lengend Snippet: Epistatic interaction defined by PD‐1 and IFNL4 polymorphisms and their associations with HCV‐related diseases compared to patients with a chronic HCV infection

Article Snippet: Genotyping of IFNL4 rs12979860 and rs117648444 (C>T, P70S) was performed using custom TaqMan SNP genotyping assays (Applied Biosystems) on a 7900HT Fast Real‐Time PCR system (Applied Biosystems).

Techniques:

Journal: PLOS ONE

Article Title: Utility of a buccal swab point-of-care test for the IFNL4 genotype in the era of direct acting antivirals for hepatitis C virus

doi: 10.1371/journal.pone.0280551

Figure Lengend Snippet: Comparisons between Genedrive buccal swab and whole blood genotyping: CC, CT, and TT.

Article Snippet: The Genedrive® (previously Epistem) IFNL4 genotyping platform is a small, handheld ‘Point of Care’ (PoC) molecular diagnostic device developed by Genedrive PLC UK.

Techniques:

Journal: PLOS ONE

Article Title: Utility of a buccal swab point-of-care test for the IFNL4 genotype in the era of direct acting antivirals for hepatitis C virus

doi: 10.1371/journal.pone.0280551

Figure Lengend Snippet: Comparisons between Genedrive buccal swab and genome-wide genotyping: CC vs. non-CC.

Article Snippet: The Genedrive® (previously Epistem) IFNL4 genotyping platform is a small, handheld ‘Point of Care’ (PoC) molecular diagnostic device developed by Genedrive PLC UK.

Techniques: Genome Wide, Sequencing