Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Differential expression of WISP1 in normal and tumor tissue samples. (A) Expression of WISP1 in pan-cancer tissues and adjacent normal tissues via TIMER2.0. (B) Expression levels of WISP1 in ESCA from GEPIA2 (|Log2FC|> 1, P < 0.01, log scale: log2 (TPM + 1), Jitter Size: 0.4; T: Tumor, N: Normal). (C) Expression of WISP1 mRNA in the ESCC dataset. (D–G) Results of IHC and WB analyses using 12 paired ESCC tissues and adjacent control samples (T: Tumor, N: Normal; scale bars = 100μm). (H) ROC curves predicting the prognostic ability of high WISP1 expression for 1-year, 3-year, and 5-year patient survival. *p < 0.05; **p < 0.01; ***p < 0.001,;****P < 0.0001.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Quantitative Proteomics, Expressing, Control

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: The impact of WISP1 expression on the survival of ESCC patients. (A–C) Kaplan-Meier analysis of overall survival for high-expression versus low-expression groups in the datasets GSE53624 , GSE53625 , and TCGA. (D–F) Time-dependent ROC analysis for patients in the datasets.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Identification of DEGs in ESCC, functional enrichment analysis, and functional annotation of WISP1. (A) Volcano plot shows the DEGs from four datasets. (B) A Venn diagram shows genes that are differentially expressed across all four datasets. (C) GO and KEGG analyses reveal the potential biological mechanisms of DEGs in the four datasets. (D, E) GeneMANIA and STRING databases identify target proteins and genes associated with WISP1, followed by enrichment analysis of the functions of these proteins and genes. (F) Visualization of enrichment analysis results.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Functional Assay

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Tumor microenvironment and immune cell infiltration analysis in the GSE53624 dataset. (A) Differences in the abundance of infiltrating immune cells between high-expression and low-expression groups. (B–D) Differences in stromal scores, immune scores, and estimate scores between high-expression and low-expression groups. (E) Evaluation of the expression of immune checkpoint molecules (CD274, PDCD1, TIGIT, CD276, CTLA4, LAG3) between high-risk and low-risk groups. (F) Correlation between WISP1 expression and immune checkpoint molecules (CD274, PDCD1, TIGIT, CD276, CTLA4, LAG3). Blue indicates positive correlation, red indicates negative correlation, and the numbers inside the boxes represent the magnitude of the correlation. *p < 0.05; **p < 0.01; ***p < 0.001; ****P < 0.0001.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Gene set enrichment analysis (GSEA) of hallmark pathways for esophageal squamous cell carcinoma (ESCC) patients stratified by WISP1 expression levels. (A–C) Enrichment patterns of associated pathways in the GSE53624 , GSE53625 , and TCGA datasets. The analyses were evaluated using a significance threshold corrected for the False Discovery Rate (FDR).

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Computational prediction of drug sensitivity for ESCC patients stratified by WISP1 expression levels. Using WISP1 expression grouping derived from the GSE53624 dataset, the “oncoPredict” R package was employed to evaluate the drug response spectrum. **** indicates statistical significance at p < 0.0001.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Expressing, Derivative Assay

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: Functional characterization of WISP1 in a subpopulation of fibroblasts. (A) Gene Set Enrichment Analysis (GSEA) shows enrichment results of differentially expressed genes between WISP1_Fib_Negative and WISP1_Fib_Positive clusters within the c5.all.v2024.1.Hs.symbols gene set; (B, C) display the enrichment results of DEGs in the h.all.v7.1.symbols gene set. (D) Proportional distribution of WISP1_Fib_Negative and WISP1_Fib_Positive clusters in normal esophageal tissues and ESCC samples. (E, F) Comparative analysis of intercellular communication signal intensity among fibroblast subpopulations. (G, H) Molecular stratification of fibroblasts using lineage-specific markers (DCN/Decorin, IGFBP6/Insulin Like Growth Factor Binding Protein 6, MFAP5/Microfibril Associated Protein 5, ACTA2/Actin Alpha 2 Smooth Muscle, TAGLN/Transgelin, CTHRC1/Collagen Triple Helix Repeat Containing 1)into NFs and CAFs subtypes. (I) Validation of CAF-specific markers (FAP, COL1A1, COL3A1, COL4A1, COL10A1, MMP1, MMP11, MMP14) and expression patterns of WISP1 in CAFs via single-cell RNA sequencing.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Functional Assay, Binding Assay, Biomarker Discovery, Expressing, RNA Sequencing

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: WISP1 regulates cancer-associated fibroblast function and extracellular matrix remodeling. (A, B) Immunofluorescence multiplex staining of α-smooth muscle actin (αSMA) and fibroblast activation protein (FAP) in paired cancer-associated fibroblasts (CAFs, tumor-derived) and normal fibroblasts (NFs, adjacent non-tumor tissues) (scale bars = 50μm). (C) Western blot (WB) analysis of WISP1, αSMA, and FAP expression in CAFs versus NFs. (D, E) Lentiviral short hairpin RNA (shRNA)-mediated WISP1 knockdown in CAFs, validated by quantitative reverse transcription PCR (qRT-PCR) and WB. (F–K) Functional characterization of CAF proliferation (CCK-8/EdU), migration, and invasion (Transwell) post-WISP1 silencing (scale bars = 50μm). (M) ELISA quantification of secreted WISP1 in supernatants from untransfected CAFs, CAFs-shVector, and CAFs-shWISP1 at 0 h, 24 h, and 48 h (N) Schematic of indirect co-culture system for CAF-ESCC interaction analysis. (L, P) WB assessment of extracellular matrix (ECM)-remodeling markers (COL1A1, MMP14) in WISP1-depleted CAFs and rescue via recombinant human WISP1 (rhWISP1). (O, Q) Transwell assay comparing the migration and invasion capacities of KYSE150 (left) and Eca109 (right) cells co-cultured with untransfected CAFs, CAFs-shVector, or CAFs-shWISP1(scale bars = 50μm). (R, S) Transwell assay assessing the migration and invasion capacities of KYSE150 (left) and Eca109 (right) cells in the CAFs-shWISP1 co-culture system following rescue experiments with rhWISP1 supplementation (scale bar = 50μm).Data are presented as mean ± standard deviation from three independent experiments. Statistical significance was analyzed by two-tailed Student’s t-tests. ns = not significant, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Immunofluorescence, Multiplex Assay, Staining, Activation Assay, Derivative Assay, Western Blot, Expressing, shRNA, Knockdown, Reverse Transcription, Quantitative RT-PCR, Functional Assay, CCK-8 Assay, Migration, Enzyme-linked Immunosorbent Assay, Co-Culture Assay, Recombinant, Transwell Assay, Cell Culture, Standard Deviation, Two Tailed Test

Journal: Frontiers in Immunology

Article Title: WISP1 drives esophageal squamous cell carcinoma progression via modulation of cancer-associated fibroblasts and immune microenvironment

doi: 10.3389/fimmu.2025.1586790

Figure Lengend Snippet: WISP1 regulates ECM remodeling in CAFs through STAT3 signaling. (A, B) Representative phospho-kinase antibody array membrane comparing CAFs-shVector and CAFs-shWISP1. Red boxes highlight phosphorylated STAT3 (Y705) signals. (C) Dose-response curve of STAT3 inhibitor Stattic in CAFs. (D) Western blot analysis of phosphorylated STAT3 (Y705), total STAT3, and GAPDH (loading control) in: CAFs-shVector, CAFs-shWISP1, CAFs-shWISP1 + rhWISP1 (0.8 μg/mL), and CAFs-shWISP1 + rhWISP1 (0.8 μg/mL) + Stattic (7 μM). (E) Western blot analysis of COL1A1, MMP14, and GAPDH in CAFs-shWISP1 + rhWISP1 (0.8 μg/mL) and CAFs-shWISP1 + rhWISP1 (0.8 μg/mL) + Stattic (7 μM).

Article Snippet: Primary fibroblasts were infected with shWISP1 lentivirus or treated with recombinant human WISP1 (rhWISP1) protein( NP_003873.1 , Sino Biological, China; endotoxin level <0.1 EU/μg, as determined by Limulus Amebocyte Lysate assay), followed by three washes with PBS.

Techniques: Ab Array, Membrane, Western Blot, Control

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Alterations in (A) EBA permeability, (B) wet:dry ratio of lungs, and (C) levels of total protein, (D) TNF-α and (E) IL-6, as well as (F) PMNs numbers in BALF at 4 h after poly(I:C) and MTV treatment. Data are presented as means ± SEM (n = 15) and compared by one-way ANOVA and Student-Newman-Keuls test. For comparing the poly(I:C) group with the untreated control group #P < 0.05, ##P < 0.01, ###P < 0.001. For comparing the poly(I:C) + MTV group with the poly(I:C) group **P < 0.01, ***P < 0.001.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Permeability

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Morphologic alterations of the lungs as determined by photomicro graphy. Control: photomicrograph of a pulmonary section from a control mouse. MTV: photomicrograph of a lung section from a normal mouse treated with MTV. Poly(I:C): photomicrograph of a lung section from a poly(I:C) challenged mouse. Poly(I:C) + MTV: photomicrograph of a lung section from a poly(I:C)-challenged mouse treated with MTV. Original magnification: 400×. A total histopathologic score of lung injury was calculated for each animal. Data are presented as means ± SEM (n = 5) and compared by one-way ANOVA and Student-Newman-Keuls test. For comparing the poly(I:C) group with the untreated control group ###P < 0.001. For comparing the poly(I:C) + MTV group with the poly(I:C) group ***P < 0.01.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques:

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Alterations in pulmonary levels of wisp1 mRNA expressed as (A) the ratio to the housekeeping gene β-actin and protein and (B) the ratio to the housekeeping protein β-actin at 4 h after poly(I:C) and MTV treatment. Data are presented as means ± SEM (n = 15) and compared by one-way ANOVA and Student-Newman-Keuls test. For comparing the poly(I:C) group with the untreated control group ##P < 0.01. For comparing the poly(I:C) + MTV group with the poly(I:C) group *P < 0.05, **P < 0.01.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques:

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Effects of anti-WISP1 treatment on (A) EBA permeability, (B) wet:dry ratio of lungs, and (C) levels of total protein, (D) TNF-α and (E) IL-6, as well as (F) PMNs numbers in BALF at 4 h after poly(I:C) and MTV treatment. Animals received the anti-WISP1 blocking antibody or control IgG i.p. 1 h before mechanical ventilation. Data are presented as mean ± SEM (n = 10) and compared by paired t test. For comparing the anti-WISP1 group with the IgG group *P < 0.05, **P < 0.01.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Permeability, Blocking Assay

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Alterations in pulmonary levels of integrin b3 mRNA expressed as (A) the ratio to the housekeeping gene β-actin and protein and (B) the ratio to the housekeeping protein β-actin at 4 h after poly(I:C) and MTV treatment. Data are presented as means ± SEM (n = 15) and compared by one-way ANOVA and Student-Newman-Keuls test. For comparing the poly(I:C) group and MTV group with the untreated control group: ###P < 0.001. And for comparing the poly(I:C) + MTV group with the poly(I:C) and MTV groups: ***P < 0.001.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques:

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Effects of poly(I:C) + MTV on WISP1/integrin β3 interaction. Lung tissues were harvested from C57BL/6J mice in the control and poly(I:C) + MTV groups. Total lysates were immunoprecipitated with an anti-WISP1 antibody or an anti-β3 antibody before immunoblot was performed. (A) Glycosylated form of integrin β3 (110 kD) was detected with an anti-integrin β3 antibody (IB: integrin β3) either in the whole lung tissue lysates (Total Lysates) or after immunoprecipitation with an anti-WISP1 antibody (IP: WISP1). (B) WISP1 was detected with an anti-WISP1 antibody (IB: WISP1) either in the whole lung tissue lysates (Total Lysates) or after immunoprecipitation with an anti-integrin β3 antibody (IP: integrin β3). Antibody species matched serum (Goat IgG) was used as a negative control for coprecipitation experiments. WISP1 and integrin β3 interact was enhanced in the poly(I:C) + MTV group as compared with the control group.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Immunoprecipitation, Western Blot, Negative Control

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Effects of WISP1 on poly(I:C)-induced TNF-α production in macrophages. (A) Peritoneal macrophage isolated from C57BL/6J mice were treated with WISP1 (10 μg/mL) in the presence or absence of various concentrations (0–10 μg/mL) of poly(I:C) in serum-free medium for 16 h. TNF-α levels in the supernatant were measured by ELISA. (B) Peritoneal macrophage isolated from C57BL/6J mice were treated with WISP1(10 μg/mL) in the presence or absence of poly(I:C) (5 μg/mL) in serum-free medium for up to 24 h. TNF-α levels in the supernatant were measured by ELISA at 0 h, 4 h, 8 h, 12 h, 16 h, 20 h and 24 h after stimulation. Data are presented as mean ± SEM (n = 5) and compared by two-way ANOVA. For comparing the poly(I:C) + WISP1 group with the poly(I:C) alone group *P < 0.05, ***P < 0.001.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Isolation, Enzyme-linked Immunosorbent Assay

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Effects of integrin β3 knockout on WISP1 plus poly(I:C)-induced TNF-α production in macrophages. Peritoneal macrophages isolated from C57BL/6J and integrin β3 knockout mice were treated with WISP1 (10 μg/mL) and poly(I:C) (5 μg/mL) for 16 h in serum free medium. TNF-α levels in the supernatant were measured by ELISA. Data are presented as mean ± SEM (n = 5) and compared by two-way ANOVA. ***P < 0.001 C57BL/6J versus integrin β3 knockout after WISP1 plus poly(I:C) treatment.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Knock-Out, Isolation, Enzyme-linked Immunosorbent Assay

Journal: Molecular Medicine

Article Title: Mechanical Ventilation Augments Poly(I:C)-Induced Lung Injury via a WISP1-Integrin β3-Dependent Pathway in Mice

doi: 10.2119/molmed.2015.00233

Figure Lengend Snippet: Effects of WISP1 and poly(I:C) on ERK phosphorylation in macrophages (A) Peritoneal macrophages isolated from C57BL/6J mice were treated with WISP1 (10 μg/mL) or poly(I:C) in serum free medium for 15, 30 and 60 min. ERK phosphorylation was detected by Western blot. Effects of an ERK inhibitor, U0126, on WISP1 and poly(I:C)-induced TNF-α release in macrophages. (B) Peritoneal macrophage isolated from C57BL/6J mice were treated with WISP1 (10 μg/mL) and poly(I:C) (5 μg/mL) in the presence or absence of various concentrations (0–1.0 μmol/L) of U0126 in serum free medium for 16 h. TNF-α levels in the supernatant were measured by ELISA. Data are presented as means ± SEM (n = 5) and compared by two-way ANOVA. For comparing the poly(I:C) + WISP1 group pretreated with U0126 0.5 μmol/L with poly(I:C) + WISP1 untreated with U0126 group ***P < 0.001.

Article Snippet: The Synergetic Effect of WISP1 and Poly(I:C) on Macrophages Peritoneal macrophages isolated from C57 BL/6J mice were treated with poly(I:C) in the presence or absence of recombinant WISP1 (1680-WS-050, R&D Systems) in serum-free medium for up to 24 h. TNF-α levels in the medium were measured by ELISA.

Techniques: Isolation, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: a Representative images of antibody arrays incubated with p14 or 20wo mouse serum. Spots corresponding to Wisp1 are marked with an empty square box. b Serum levels of Wisp1 in p14 ( n = 7), p28 ( n = 4), 11wo ( n = 6) and 20wo ( n = 7) mice measured with ELISA. c Plasma levels of WISP1 in children ( n = 11) and adult ( n = 14) measured with ELISA. d Quantification by qPCR of the expression of the indicated CCN genes in p14 ( n = 7, green) and 20wo mouse islets ( n = 6 for Cyr61 , n = 7 for other genes, gray). Values are expressed relative to Tbp . e Quantification by qPCR of the expression of the indicated CCN genes in adult human islets ( n = 6 for CYR61 , n = 5 for other genes). Values are expressed relative to TBP . f Quantification by qPCR of Wisp1 gene expression in the indicated p14 (green) and 20wo (gray) mouse tissues. Wisp1 gene expression is shown relative to levels in p14 bone, given the value of 1 ( n = 9 for bone, n = 4 for all other tissues). WAT: white adipose tissue; Gastroc: gastrocnemius. All data shown represent mean ± SEM from the indicated n . Indicated comparisons were made using two-tailed Student’s t test ( c , d ), one-way ( b ) and two-way ( f ) ANOVA. * p < 0.05; ** p < 0.01; **** p < 0.0001; ns: not significant. In f , at p14, bone Wisp1 gene expression was significantly higher than in all other tissues tested with p < 0.01–0.001.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Incubation, Enzyme-linked Immunosorbent Assay, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: a – c Beta cell proliferation in fixed pancreases from p14 Wisp1 + / + or Wisp1 − / − mice. a Representative images of pancreases co-immunostained for insulin (purple)/ki67 (green) or insulin (purple) /pHH3 (green). Nuclei are marked with Hoechst in blue. b Quantification of the percentage of beta (insulin+) cells that are ki67+ in Wisp1 + / + ( n = 5, yellow) or Wisp1 − / − ( n = 6, orange) mice. c Quantification of the percentage of beta (insulin+) cells that are pHH3+ in Wisp1 + / + ( n = 4, yellow) or Wisp1 − / − ( n = 5, orange) mice. d – f Beta cell proliferation in fixed pancreases from p12 Wisp1 − / − mice treated with saline or with rmWisp1 protein for three days (from p9 to p11). d Representative images of pancreases co-immunostained for insulin (purple)/ki67 (green) or insulin (purple)/pHH3 (green). Nuclei are marked with Hoechst in blue. e Quantification of the percentage of beta (insulin+) cells that are ki67+ in mice injected with rmWisp1 ( n = 4, orange) o saline ( n = 4, brown). f Quantification of the percentage of beta (insulin+) cells that are pHH3+ in mice injected with rmWisp1 ( n = 3, orange) o saline ( n = 3, brown). g – i Beta cell proliferation of 20wo mouse islet grafts transplanted into the anterior chamber of the eye of p16 Wisp1 + / + or Wisp1 − / − mouse recipients. g Representative images of islet grafts co-immunostained for insulin (purple)/ki67 (green) or insulin (purple)/pHH3 (green). Nuclei are marked with Hoechst in blue. h Quantification of the percentage of beta (insulin+) cells that are ki67+ in p16 Wisp1 + / + ( n = 7, yellow) or Wisp1 − / − ( n = 7, orange) mice. i Quantification of the percentage of beta (insulin+) cells that are pHH3+ in p16 Wisp1 + / + ( n = 4, yellow) or Wisp1 − / − ( n = 4, orange) mice. All data values represent mean ± SEM for the indicated n . * p < 0.05; ** p < 0.01 using two-tailed Student’s t test. Scale bars are 25 μm. ND: not detectable.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Saline, Injection, Two Tailed Test

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: Adenoviruses encoding human WISP1 (Ad-WISP1) or beta-galactosidase (Ad-betaGal) were injected via the tail vein into 12wo C57BL6/J mice. a Quantification by qPCR of human WISP1 transcripts in the livers of mice seven days and fourteen days ( n = 4) post-injection. Levels are expressed relative to values in mice injected with Ad-betaGal, given the value of 1. b Quantification by qPCR of mouse Wisp1 mRNA ( n = 5 for Ad-betaGal, red; n = 7 for Ad-WISP1, purple) and human WISP1 transcripts ( n = 5 for Ad-betaGal, red; n = 4 for Ad-WISP1, purple) in the livers of mice fourteen days post-injection. Expression levels are expressed relative to Tbp . c Serum human WISP1 levels were measured by ELISA at days 7 and 14 post-injection of Ad-betaGal ( n = 7) or Ad-WISP1 ( n = 8 at day 7; n = 4 at day 14, purple). Human WISP1 was not detectable (ND) in serum from mice injected with Ad-betaGal. d , e Beta cell proliferation following injection of Ad-WISP1 and Ad-betaGal. d Representative images of in toto immunofluorescence staining against ki67 (green) and insulin (purple) in islets isolated at day 7 after injection of the indicated adenoviruses. Nuclei are labeled with Hoechst (blue). e Percentage of beta cells (insulin+) that are ki67+ at day 7 after injection of Ad-betaGal ( n = 4, red) or Ad-WISP1 ( n = 7, purple). f Beta cell mass at day 14 following injection of Ad-betaGal ( n = 5, red) or Ad-WISP1 ( n = 7, purple). All data shown represent mean ± SEM for the indicated n . * p < 0.05; ** p < 0.01 using two-tailed Student’s t test ( b , e , f ) or two-way ANOVA ( a , c ). Scale bars are 25 μm.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Injection, Expressing, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Isolation, Labeling, Two Tailed Test

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: a Schematic of experimental plan. b Quantification by qPCR of mouse Wisp1 mRNA ( n = 6 for Ad-betaGal, yellow; n = 10 for Ad-WISP1, green) and human WISP1 transcripts ( n = 8 for Ad-betaGal, yellow; n = 6 for Ad-WISP1, green) in the livers of mice fourteen days post-injection. Expression levels are expressed relative to Tbp . c Serum human WISP1 levels were measured by ELISA at days 9 and 14 post-injection with Ad-betaGal ( n = 5) or Ad-WISP1 ( n = 4, green). Human WISP1 was not detectable (ND) in mice injected with Ad-betaGal. d Blood glucose concentrations measured at the indicated days post-injection with Ad-betaGal ( n = 8, yellow) or Ad-WISP1 ( n = 7, green). e Serum insulin at day 14 following administration of Ad-betaGal ( n = 11, yellow) or Ad-WISP1 ( n = 13, green). f , g Beta cell proliferation following injection of Ad-WISP1 and Ad-betaGal. f Representative images of immunofluorescence staining against ki67 (green) and insulin (purple) in fixed pancreases at day 14 after injection of the indicated adenoviruses. Nuclei are labeled with Hoechst (blue). g Percentage of beta cells (insulin+) that are ki67+ at day 14 days after injection of the indicated adenoviruses ( n = 7; Ad-betaGal in yellow, Ad-WISP1 in green). h Beta cell fractional area (insulin+ area relative to total pancreatic area) and i total beta cell mass at day 14 after injection of Ad-betaGal ( n = 8, yellow) or Ad-WISP1 ( n = 7, green). All data shown represent mean ± SEM for the indicated n . * p < 0.05; ** p < 0.01 using two-tailed Student’s t test ( b , e) , one-tailed Student’s t test ( g – i ) and two-way ANOVA ( d ). Scale bars are 25 μm.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Injection, Expressing, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Staining, Labeling, Two Tailed Test, One-tailed Test

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: a , b Beta cell proliferation in mouse islets incubated with Wisp1 recombinant mouse protein (rmWisp1). a Representative images of in toto immunofluorescence showing ki67 (green) and insulin (purple) staining in mouse islets cultured for 48 h with increasing amounts of rmWisp1 protein or harmine. Nuclei are marked with Hoechst in blue. b Percentage of beta cells (insulin+) that are ki67+ in cultured islets under the indicated conditions (control, n = 49 islets, in gray; rmWisp1-250, n = 19 islets, in blue; rmWisp1-500, n = 45 islets, in green; harmine, n = 15 islets, in pink; from four independent experiments except for harmine that are from two independent experiments). c , d Beta cell proliferation in mouse islets co-cultured with NIH3T3 cells expressing WISP1. c Representative images of in toto immunofluorescence showing ki67 (green) and insulin (purple) staining in mouse islets co-cultured for 24 or 48 h with NIH3T3 cells infected with the indicated adenoviruses. Nuclei are marked with Hoechst in blue. d Percentage of beta cells (insulin+) that are ki67+ in mouse islets cultured under the indicated conditions for 24 h (control, in gray: n = 25 islets; 3T3/WISP1, in blue: n = 21 islets, from three independent experiments) and for 48 h (control, in gray: n = 100 islets; 3T3/WISP1, in blue: n = 88 islets, from three independent experiments). e , f Beta cell proliferation in human islets incubated with recombinant human WISP1 protein (rhWISP1). e Representative images of in toto immunofluorescence showing ki67 (green) and insulin (purple) staining in human islets incubated for 48 h with increasing amounts of rhWISP1 protein or with harmine. Nuclei are marked with Hoechst in blue. f Percentage of beta cells (insulin+) that are ki67+ in human islets cultured under the indicated conditions (control, in gray: n = 79 islets; rhWISP1-250, in blue: n = 60 islets; rhWISP1-500, in green: n = 98 islets; harmine, in pink: n = 17 islets; from four donors). All data shown represent mean ± SEM for the indicated n . Comparisons were made using one-way ANOVA. * p < 0.05; ** p < 0.01; **** p < 0.0001. Scale bars are 25 μm.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Incubation, Recombinant, Immunofluorescence, Staining, Cell Culture, Expressing, Infection

Journal: Nature Communications

Article Title: Wisp1 is a circulating factor that stimulates proliferation of adult mouse and human beta cells

doi: 10.1038/s41467-020-19657-1

Figure Lengend Snippet: a Determination of Akt activation ( Ser473 phosphorylation) by immunoblot analysis in mouse islets incubated with recombinant mouse Wisp1 protein (rmWisp1) at 500 ng/ml for 30 min. Top: representative immunoblot image. Molecular weight markers are shown on the right. Bottom, quantification of Akt activation, expressed relative to control islets (no rmWisp1), given the value of 1 (control, in gray: n = 10; rmWisp1, in blue: n = 11, from five independent experiments). b , c Beta cell proliferation in mouse islets incubated with rmWisp1 protein and Akt inhibitors. b Representative immunofluorescence images showing ki67 staining in green and insulin in purple. Nuclei are marked with Hoechst (blue). c Percentage of beta cells (insulin+) that are ki67+ in islets incubated with rmWisp1protein at 500 ng/ml for 48 h alone ( n = 41 islets, blue) or with the Akt inhibitors AZD5363 ( n = 21 islets, pink) or Akti ( n = 25 islets, yellow), or left untreated (control, gray: n = 39 islets) from three different isolation experiments. d Determination of AKT activation ( Ser473 phosphorylation) by immunoblot analysis in human islets incubated with recombinant human WISP1 protein (rhWISP1) at 500 ng/ml for 15 min. Top: representative immunoblot image. Molecular weight markers are shown on the right. Bottom, quantification of AKT activation, expressed relative to control islets (no WISP1), given the value of 1 (control in gray: n = 12; rhWISP1 in blue: n = 7, from 3 donors) e , f Beta cell proliferation in human islets incubated with Wisp1 protein and Akt inhibitors. e Representative immunofluorescence images showing ki67 staining in green and insulin in red. Nuclei are marked with Hoechst (blue). f Percentage of beta cells (ins+) that are ki67+ in human islets incubated with rhWISP1 protein at 500 ng/ml for 48 h alone ( n = 21 islets, blue) or with the Akt inhibitors AZD5363 ( n = 31 islets, pink) or Akti ( n = 15 islets, yellow), or left untreated (control, gray: n = 20 islets) from 3 donors. All data shown represent mean ± SEM for the indicated n . Comparisons were made using two-tailed Student’s t test ( a , d ) or one-way ANOVA ( c , f ). * p < 0.05; *** p < 0.001; **** p < 0.0001. Scale bars are 25 μm.

Article Snippet: Mouse Wisp1 levels were determined using a Mouse/Rat Wisp-1/CCN4 Immunoassay (R&D Systems) that detects both natural and full-length recombinant Wisp1 protein.

Techniques: Activation Assay, Western Blot, Incubation, Recombinant, Molecular Weight, Immunofluorescence, Staining, Isolation, Two Tailed Test

Journal: Journal of Cell Communication and Signaling

Article Title: The role of CCN family genes in haematological malignancies

doi: 10.1007/s12079-015-0296-4

Figure Lengend Snippet: Summary of CCN-associated haematological malignancies

Article Snippet: Although no mast cell or dendritic/histiocytic neoplasms have been reported to have an association with CCN family members, two CCN proteins have been associated with myeloid neoplasms, CCN1 with AML and CCN3 with CML (Table ). table ft1 table-wrap mode="anchored" t5 Table 1 caption a7 WHO classification CCN family member Haematological malignancy CCN-expressing cell Disease outcome Reference (first cited) Myeloid neoplasms CCN1 Acute myelogenous leukaemia (AML) Over-expressed in AML Benefits neoplasm Niu et al. 2014 CCN3 Chronic myelogenous leukaemia (CML) Under-expressed in BCR-ABL-positive CML Benefits neoplasm McCallum et al. 2006 Lymphoid neoplasms Precursor lymphoid neoplasms CCN2 B-cell precursor acute lymphoblastic leukaemia (ALL) Over-expressed in B-cell precursor ALL Benefits neoplasm Vorwerk et al. 2000 Mature B-cell neoplasms CCN1 Multiple myeloma Over-expressed in tumour-associated MSCs Inhibits neoplasm Santra et al. 2011 CCN2 Multiple myeloma CCN2 whole protein reduced and N-terminal fragment increased in serum Inhibits neoplasm Munemasa et al. 2007 Diffuse large B-cell lymphoma (DLBCL) Over-expressed in DLBCL sub-types with stromal infiltration Inhibits neoplasm Blenk et al. 2007 Hodgkin lymphoma Over-expressed in nodular sclerosis Hodgkin lymphoma Inhibits neoplasm Birgersdotter et al. 2010 Mantle cell lymphoma Under-expressed in tumour cells Unknown Rizzatti et al. 2005 CCN4 Multiple myeloma Under-expressed in tumour-associated MSCs Unknown Corre et al. 2007 Mature T-cell and NK-cell neoplasms CCN1 Peripheral T-cell lymphoma Over-expressed in tumour cells Unknown Mahadevan et al. 2005 Angioimmunoblastic lymphoma Over-expressed in tumour cells Unknown Piccaluga et al. 2007b CCN2 Peripheral T-cell lymphoma Over-expressed in tumour cells Unknown Mahadevan et al. 2005 Open in a separate window Summary of CCN-associated haematological malignancies CCN1 CCN1 was found to be over-expressed in bone marrow samples from AML patients, showing heterogenous levels of expression when compared by immunoblots with normal bone marrow samples (Niu et al. 2014 ).

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