Journal: medRxiv

Article Title: In severe COVID-19, SARS-CoV-2 induces a chronic, TGF-β-dominated adaptive immune response

doi: 10.1101/2020.09.04.20188169

Figure Lengend Snippet: Peripheral blood activated B cells (CD27 high CD38 high ) were isolated and sorted by FACS for single cell sequencing. a) Percentage of CD27 high CD38 high cells among live B cells. Each sample is represented by a circle. The line indicates the median. Unpaired two sided Mann-Whitney U test. b) UMAP representation of 72277 CD27 high CD38 high sorted B cells from 9 COVID-19 ICU patients and 3 healthy controls. Between 2416 and 11229 cells were recovered per sample. Transcriptionally similar clusters were identified using shared nearest neighbor (SNN) modularity optimization (left). The combination of CD19, MS4A1, CD27, CD38, IFIT1, MIKI67, IRF4 and PRDM1 expression were used for annotation of the different activation/differentiation stages (right). c) Heatmap of genes that resemble markers for the six different clusters. Depicted are genes with a p-value < 0.01 (Wilcoxon rank sum test) after bonferroni correction, and an average absolute fold-change > log2(1.3). Shown are z-scores of the average expression. d) UMAP representation of the expression levels of selected signature genes for activated/differentiated B cells. e) UMAP representation of analyzed cells from one healthy control and two ICU patients, representing early (first week, patient #1) and late (>7days, patient #5) phase after ICU admission. f) Percentage of cells belonging to a defined cluster among all sequenced B cells per donor. Each dot represents one time point from one single donor. Donors were grouped as “HC” for healthy controls (n = 3), “1st week” for patients within 7 days after ICU admission (n = 6) and “Late” for patients who have been admitted to the ICU for more than a week at the time of analysis (n = 8). The line indicates the median. Significance was determined by using a two-sided analysis of varience (ANOVA) folowed by Tukey’s multiple comparison test with corrected p values as indicated in the figure.

Article Snippet: Fluorescence-coupled antibodies used: IgA2-PE (Clone REA995, Miltenyi, Cat. No.130-117-763, dilution 1:50); CD27-PE (Clone REA499, Miltenyi, Cat. No. 130-114-166, dilution 1:50); CD38-PE (Clone IB6, Miltenyi, Cat. No. 130-113-427, dilution 1:50) and IgA-PE (Clone IS11–8E10, Miltenyi, Cat. No. 130-114-002, dilution 1:50).

Techniques: Isolation, Sequencing, MANN-WHITNEY, Expressing, Activation Assay, Control, Comparison

Journal: medRxiv

Article Title: In severe COVID-19, SARS-CoV-2 induces a chronic, TGF-β-dominated adaptive immune response

doi: 10.1101/2020.09.04.20188169

Figure Lengend Snippet: a) 5-marker MELC panel of SARS-CoV2-positive and control lungs (SARS-CoV2-negative). Respective patient characteristics are given in supplementary Fig. 4a. Each image of each patient depicts the same field of view of the same section, sequentially stained with the fluorescence-labelled antibodies indicated and the nuclear stain DAPI. Magenta arrows indicate IgA2+IgA+CD27 + CD38 + cells (containing a nucleus). Images contain 2048 × 2048 pixels and are generated using an inverted wide-field fluorescence microscope with a 20x objective, a lateral resolution of 325 nm and an axial resolution above 5 µm. Scale bar: 100 µm. b) Absolute numbers of IgA2+IgA+CD27 + CD38 + cells per field of view in all MELC runs acquired (two runs per patient except for COVID-19_A with four runs and Control_B with one single run; see and supplementary Fig. 4b). Each field of view is represented by a circle. The line indicates the median. Unpaired two sided Mann-Whitney U test. c) Region of interest of one exemplary control and COVID-19 lung (as in a)), showing an overlay of the indicated markers. White arrows point out IgA2+CD27 + CD38 + cells. Scale bar: 20 µm. (d-e) Bronchoalveolar lavage (BAL) cells for single cell sequencing were enriched for CD45 + cells via MACS and live cells were further sorted using FACS (see supplementary Fig. 4c). d) UMAP of 5459 cells representing clusters containing T cells, B cells and CD14 -expressing cells from patient #1 on day 59 following ICU admission. B and T cell cluster identification based on BCR/TCR, CD19, CD3E, CD4 and CD8A expression (see supplementary Fig. 4d). UMAP representation of expression of TGFB1, IL21, CD40LG and IFNG . e) UMAP coordinates and clustering was computed for 433 and 2723 cells from patient #9 on days 31 and 46 following ICU admission, respectively. UMAP representation of expression of TGFB1, IL21, CD40LG and IFNG at the two different time points is shown side by side.

Article Snippet: Fluorescence-coupled antibodies used: IgA2-PE (Clone REA995, Miltenyi, Cat. No.130-117-763, dilution 1:50); CD27-PE (Clone REA499, Miltenyi, Cat. No. 130-114-166, dilution 1:50); CD38-PE (Clone IB6, Miltenyi, Cat. No. 130-113-427, dilution 1:50) and IgA-PE (Clone IS11–8E10, Miltenyi, Cat. No. 130-114-002, dilution 1:50).

Techniques: Marker, Control, Staining, Fluorescence, Generated, Microscopy, MANN-WHITNEY, Sequencing, Expressing

Journal: Stem Cell Research & Therapy

Article Title: Bone marrow mesenchymal stem cells alleviate liver fibrosis after rat liver transplantation through JAK1/STAT5 pathway

doi: 10.1186/s13287-025-04353-y

Figure Lengend Snippet: BMSCs may alleviate liver fibrosis after liver transplantation through the JAK1/STAT5 pathway. ( A ) Box plot for FPKM values. PCA analysis. Bar plot of the statistic of differentially expressed genes. Venn graph of the differentially expressed genes. Volcano plot of the differentially expressed genes. Heat map of RNA differential expression. GO analysis reveals the potential impact of the differentially expressed RNAs on functions. KEGG analysis reveals the potential impact of the differentially expressed RNA on pathways. KEGG map of the JAK/STAT pathway. ( B ) Western blot analyses of JAK1, p-STAT5, STAT5, p-STAT1, STAT1, p-STAT3, STAT3, IL7R, and IL7 in liver tissue. TUBA1A was used as the loading control except for p-STAT5, p-STAT1, and p-STAT3. STAT5, STAT1, and STAT3 were used as the loading control for p-STAT5, p-STAT1, and p-STAT3, respectively. Jak1 , Il7r , and Il7 mRNA levels in liver tissue. ( C ) Microscopic images of each group stained with HE, Masson, Sirius Red, and ACTA2 (magnification: ×100, n = 6). Area stained with Masson, Sirius Red, and ACTA2, as well as Metavir fibrosis staging. The levels of serum ALT, AST, and TBil in each group. ( D ) Western blot analyses of COL1A1, JAK1, p-STAT5, STAT5, ACTA2, and CASP3 in liver tissues. TUBA1A was used as the loading control except for p-STAT5. STAT5 was used as the loading control for p-STAT5. Full-length blots/gels are presented in Supplementary Fig. . The samples were derived from the same experiment and processed in parallel. Col1a1 , Jak1 , Acta2 , and Casp3 mRNA levels in liver tissues. ACTA2, actin alpha 2; ALT, alanine aminotransferase; AST, aspartate aminotransferase; BMSCs, bone mesenchymal stem cells; CASP3, caspase 3; COL1A1, collagen type I alpha 1 chain; GO, Gene Ontology; IL, interleukin; JAK, Janus-activated kinase; KEGG, Kyoto Encyclopedia of Genes and Genomes; LT, liver transplantation; STAT, signal transducer and activator of transcription; TBil, total bilirubin. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: The LT + anti-IL7 group received anti-IL7 antibody (M25; Cat# HY-P990210, MCE, Shanghai, China) (250 μg/kg of body weight, per ip, once a week) for 2 weeks after LT.

Techniques: Transplantation Assay, Quantitative Proteomics, Western Blot, Control, Staining, Derivative Assay

Journal: Stem Cell Research & Therapy

Article Title: Bone marrow mesenchymal stem cells alleviate liver fibrosis after rat liver transplantation through JAK1/STAT5 pathway

doi: 10.1186/s13287-025-04353-y

Figure Lengend Snippet: BMSCs may inhibit the activation of HSC-T6 cells after hypoxia–reoxygenation and reduce their apoptosis through the JAK1/STAT5 pathway. ( A ) Western blot analyses of JAK1, p-STAT5, STAT5, p-STAT1, STAT1, p-STAT3, STAT3, IL7R, and IL7 in HSC-T6 cells. TUBA1A was used as the loading control except for p-STAT5, p-STAT1, and p-STAT3. STAT5, STAT1, and STAT3 were used as the loading control for p-STAT5, p-STAT1, and p-STAT3, respectively. Jak1 and Il7r mRNA levels in HSC-T6 cells. ( B ) Western blot analyses of COL1A1, JAK1, p-STAT5, STAT5, and ACTA2 in HSC-T6 cells. TUBA1A was used as the loading control except for p-STAT5. STAT5 was used as the loading control for p-STAT5. Col1a1 , Jak1 and Acta2 mRNA levels in HSC- T6 cells. ( C ) Apoptosis results of flow cytometry in each group. Full-length blots/gels are presented in Supplementary Fig. . The samples were derived from the same experiment and processed in parallel. ACTA2, actin alpha 2; BMSCs, bone mesenchymal stem cells; COL1A1, collagen type I alpha 1 chain; IL, interleukin; JAK, Janus-activated kinase; STAT, signal transducer and activator of transcription. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: The LT + anti-IL7 group received anti-IL7 antibody (M25; Cat# HY-P990210, MCE, Shanghai, China) (250 μg/kg of body weight, per ip, once a week) for 2 weeks after LT.

Techniques: Activation Assay, Western Blot, Control, Flow Cytometry, Derivative Assay

Journal: Stem Cell Research & Therapy

Article Title: Bone marrow mesenchymal stem cells alleviate liver fibrosis after rat liver transplantation through JAK1/STAT5 pathway

doi: 10.1186/s13287-025-04353-y

Figure Lengend Snippet: BMSCs regulate the IL7R/JAK1/STAT5 pathway in HSCs by modulating the secretion of IL7 by hepatic cells. ( A ) Microscopic images of each group stained with HE, Masson, Sirius Red, ACTA2, p-STAT5 and STAT5 (magnification: ×100, n = 6). Area stained with Masson, Sirius Red, ACTA2, and p-STAT5/STAT5, as well as Metavir fibrosis staging. The levels of serum ALT, AST, and TBil in each group. ( B ) Western blot analyses of COL1A1, JAK1, p-STAT5, STAT5 and ACTA2 in liver tissue. TUBA1A was used as the loading control except for p-STAT5. STAT5 was used as the loading control for p-STAT5. Col1a1 , Jak1 , and Acta2 mRNA levels in liver tissue. ( C ) Western blot analyses of IL7 in IAR20 cells. TUBA1A was used as the loading control. Il7 mRNA levels in IAR20 cells. ( D ) Western blot analyses of COL1A1, JAK1, p-STAT5, STAT5 and ACTA2 in HSC-T6 cells. TUBA1A was used as the loading control except for p-STAT5. STAT5 was used as the loading control for p-STAT5. Col1a1 , Jak1 and Acta2 mRNA levels in HSC- T6 cells. Full-length blots/gels are presented in Supplementary Fig. . The samples were derived from the same experiment and processed in parallel. ACTA2, actin alpha 2; ALT, alanine aminotransferase; AST, aspartate aminotransferase; BMSCs, bone mesenchymal stem cells; COL1A1, collagen type I alpha 1 chain; IL, interleukin; JAK, Janus-activated kinase; LT, liver transplantation; STAT, signal transducer and activator of transcription; TBil, total bilirubin. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001

Article Snippet: The LT + anti-IL7 group received anti-IL7 antibody (M25; Cat# HY-P990210, MCE, Shanghai, China) (250 μg/kg of body weight, per ip, once a week) for 2 weeks after LT.

Techniques: Staining, Western Blot, Control, Derivative Assay, Transplantation Assay

Journal: World Journal of Gastroenterology

Article Title: Fas counterattack in cholangiocarcinoma: A mechanism for immune evasion in human hilar cholangiocarcinomas

doi: 10.3748/wjg.v7.i6.860

Figure Lengend Snippet: Western blotting of FasL protein with mAb from QBC939 cell cultures clone 33 from QBC939 cell cultures

Article Snippet: Immunohistochemistry for FasL and CD45 Detection of FasL expression and CD45 positive cells was performed using a rabbit polyclonal anti-human FasL specific IgG and a mouse anti-human monoclonal antibody on paraffin sections of human cholangiocarcinoma respectively (Boster Biological Technology Company, Wuhan, China).

Techniques: Western Blot

Journal: Bioscience Reports

Article Title: The final proteolytic step in transmembrane signaling of multiple RsgI anti-σ factors in Clostridium thermocellum

doi: 10.1042/BSR20253055

Figure Lengend Snippet: (A ) SDS-PAGE of the eight purified RsgI-NFs samples. The expected RsgI-NF bands are indicated by red arrows. ( B ) Western blot analysis of the eight purified RsgI-NFs using the anti-His6-tag antibodies.

Article Snippet: These membranes were blocked using TBST-blocking buffer (50 mM Tris, 150 mM NaCl, 0.07% Tween-20, 5% skim milk, pH 7.4) for a duration of 2 h. Then, the membranes were incubated with mouse anti-His tag monoclonal antibody (Bioss Antibodies, Beijing, China) overnight at 4°C.

Techniques: SDS Page, Purification, Western Blot