Journal: Frontiers in Immunology

Article Title: lncRNA-HEIM Facilitated Liver Fibrosis by Up-Regulating TGF- β Expression in Long-Term Outcome of Chronic Hepatitis B

doi: 10.3389/fimmu.2021.666370

Figure Lengend Snippet: Identification of differentially expressed lncRNAs correlated with TGF- β signaling pathway in long-term CHB patients. (A) Hierarchical clustering analysis of 4,718 differentially expressed lncRNA transcripts between CHB with long-term antiviral treatment and ASC samples (fold change >2, up- or down-regulated). Columns represent each gene, and rows represent each sample. The relative expression levels of mRNAs are depicted into color scale ranging from green (down-regulated) to red (up-regulated). (B) Alluvial diagram of candidate differentially expressed lncRNAs with up-regulated mRNAs from enriched KEGG pathways. (C) Pairwise correlations between lncRNAs and mRNAs (upper), with a color gradient denoting Pearson correlation coefficients. Pairwise comparisons between lncRNAs and mRNAs (lower). Edge width corresponds to the Mantel’s r statistic, and edge color denotes the statistical significance. (D) Real-time PCR validation of lncRNA ENST00000519726 in ASC or CHB samples. ∗ p < 0.05.

Article Snippet: Each labeled cRNA was fragmented and hybridized on Human LncRNA Microarray V4.0 (Arraystar, containing 40,173 annotated lncRNAs or lncRNAs of high confidence, as well as an entire collection of 20,730 protein coding mRNAs).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Biomarker Discovery

Journal: Frontiers in Immunology

Article Title: lncRNA-HEIM Facilitated Liver Fibrosis by Up-Regulating TGF- β Expression in Long-Term Outcome of Chronic Hepatitis B

doi: 10.3389/fimmu.2021.666370

Figure Lengend Snippet: lncRNA-HEIM ENST00000519726 was highly expressed in monocytes. (A) Relative expression levels of SMAD4 and TGF- β in CD14 + monocytes against those in CD14 − cells from CHB patients. (B) Relative expression levels of lncRNA ENST00000519726 in CD14 + monocytes against those in CD14 - cells from either healthy donor (HD) or CHB patients. (C) Relative expression levels of lncRNA-HEIM, SMAD4, and TGF- β in monocytes at 2, 8, 16, and 24 h upon HBV infection. Expression levels of lncRNA-HEIM, SMAD4, and TGF- β were normalized with negative control at each time point. Comparisons over groups were analyzed using two-way ANOVA followed by Tukey’s multiple comparisons test. ∗ p < 0.05. Blue line is for lncRNA-HEIM, while green and red lines are for SMAD4 and TGF- β , respectively. (D) Relative expression levels of lncRNA-HEIM, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and U6 in cytoplasm or nuclear of monocytes. GADPDH was used as cytoplasmic control transcripts, and U6 as nuclear control. (E) Relative expression levels of lncRNA-HEIM transfected with negative control or siRNAs targeting lncRNA-HEIM (silncRNAs). **p < 0.01, ***p < 0.001.

Article Snippet: Each labeled cRNA was fragmented and hybridized on Human LncRNA Microarray V4.0 (Arraystar, containing 40,173 annotated lncRNAs or lncRNAs of high confidence, as well as an entire collection of 20,730 protein coding mRNAs).

Techniques: Expressing, Infection, Negative Control, Control, Transfection

Journal: Frontiers in Immunology

Article Title: lncRNA-HEIM Facilitated Liver Fibrosis by Up-Regulating TGF- β Expression in Long-Term Outcome of Chronic Hepatitis B

doi: 10.3389/fimmu.2021.666370

Figure Lengend Snippet: Over-expression of lncRNA-HEIM remarkably promoted the expression of TGF- β and SMAD4. (A) Relative expression of SMAD4 or TGF- β in monocytes transfected with siRNAs for lncRNA-HEIM or negative control (NC). ∗∗ p < 0.01; ∗∗∗∗ p < 0.0001. (B) Western blotting assays of endogenous protein levels of TGF- β , SAMD4, phosphorylated SMAD2, 3, and total SMAD2, 3 in monocytes transfected with siRNAs for lncRNA-HEIM or NC. GAPDH protein levels were used as internal controls. Chemiluminescent densities were normalized with GAPDH, and fold changes were calculated based on NC. (C) ELISA based TGF- β levels in culture supernatant of monocytes transfected with siRNAs for lncRNA-HEIM or NC. ∗ p < 0.05. (D) Relative expression of lncRNA-HEIM in monocytes infected with lncRNA-HEIM lentivirus (LV-HEIM) or negative control (NC). (E) Relative expression of TGF- β or SMAD4 in monocytes stably over-expressing lncRNA-HEIM or NC. ∗ p < 0.05; ∗∗∗ p < 0.001. (F) ELISA based TGF- β levels in culture supernatant of monocytes with LV-HEIM or NC. ∗∗ p < 0.01. (G) Western blotting assays of endogenous protein levels of TGF- β , SAMD4 in monocytes with LV-HEIM or NC. Fold changes, same as above.

Article Snippet: Each labeled cRNA was fragmented and hybridized on Human LncRNA Microarray V4.0 (Arraystar, containing 40,173 annotated lncRNAs or lncRNAs of high confidence, as well as an entire collection of 20,730 protein coding mRNAs).

Techniques: Over Expression, Expressing, Transfection, Negative Control, Western Blot, Enzyme-linked Immunosorbent Assay, Infection, Stable Transfection

Journal: Frontiers in Immunology

Article Title: lncRNA-HEIM Facilitated Liver Fibrosis by Up-Regulating TGF- β Expression in Long-Term Outcome of Chronic Hepatitis B

doi: 10.3389/fimmu.2021.666370

Figure Lengend Snippet: lncRNA-HEIM dramatically facilitated the fibrosis of hepatic stellate cells. (A, B) Relative mRNA expression and western blotting assays of collagen I and α -smooth muscle actin ( α -SMA) in hepatic stellate cells (HSCs) co-cultured with monocytes transfected with siRNAs for lncRNA-HEIM or NC. Chemiluminescent densities in (B) were normalized with GAPDH, and fold changes were calculated based on NC. ∗ p < 0.05. (C, D) Relative mRNA expression and western blotting assays of collagen I and α -smooth muscle actin ( α -SMA) in HSCs co-cultured with LV-HEIM stable monocytes or NC. Fold changes in (D) , same as above. ∗∗ p < 0.01. (E) Confocal imaging of collagen-I (red), α -SMA (green), and DAPI (blue) in HSCs co-cultured with LV-HEIM stable monocytes or NC. Scale bar: 20 μm. Relative intensity of density normalized by DAPI was depicted on the right. ∗∗∗ p < 0.001.

Article Snippet: Each labeled cRNA was fragmented and hybridized on Human LncRNA Microarray V4.0 (Arraystar, containing 40,173 annotated lncRNAs or lncRNAs of high confidence, as well as an entire collection of 20,730 protein coding mRNAs).

Techniques: Expressing, Western Blot, Cell Culture, Transfection, Imaging