anti cd44 neutralizing antibody  (R&D Systems)

Journal: Stem Cell Research & Therapy

Article Title: Hyaluronan supports the limbal stem cell phenotype during ex vivo culture

doi: 10.1186/s13287-022-03084-8

Figure Lengend Snippet: Effects of HA on TKE2 cells cultured on differently coated dishes or in the presence of HA as a supplement to the media. TKE2 cells were cultured on differently coated dishes ( A ) or on uncoated dishes with HA, HA + anti-CD44 or 4MU + Hyalase supplemented to the media ( B ). The number of ∆Np63 positive cells and the total number of cells (DAPI) were quantified using QuPath and the percentage of ∆Np63 + cells calculated ( C ). *Represents p ≤ 0.05, scale bar represents 50 μm

Article Snippet: In order to verify the mechanism by which HA supports LESCs, anti-CD44 neutralizing antibody (R&D systems, Minneapolis, USA#AF6127) was added to the culture media in order to block the binding of HA to CD44.

Techniques: Cell Culture

Journal: Stem Cell Research & Therapy

Article Title: Hyaluronan supports the limbal stem cell phenotype during ex vivo culture

doi: 10.1186/s13287-022-03084-8

Figure Lengend Snippet: Effects of HA on the differentiation of TKE2 cells cultured on differently coated dishes or in the presence of HA as a supplement to the media. TKE2 cells were cultured on differently coated dishes ( A ) or on uncoated dishes with HA of different molecular weights, HA + anti-CD44 or 4MU + Hyalase supplemented to the media ( B ) under differentiating conditions. The number of ∆Np63 positive cells and the total number of cells (DAPI) were quantified using Qupath and the percentage of ∆Np63 + cells calculated for TKE cells maintained on differently coated dishes ( C ) and with HA, HA + anti-CD44 and 4MU + Hyalase supplemented to the media ( D ). *Represents p ≤ 0.05, scale bar represents 50 μm

Article Snippet: In order to verify the mechanism by which HA supports LESCs, anti-CD44 neutralizing antibody (R&D systems, Minneapolis, USA#AF6127) was added to the culture media in order to block the binding of HA to CD44.

Techniques: Cell Culture

Journal: bioRxiv

Article Title: Natural killer cells associate with epithelial cells in the pancreatic ductal adenocarcinoma tumor microenvironment

doi: 10.1101/2024.05.23.593868

Figure Lengend Snippet: A. Incoming and outgoing interaction strength for each cell population grouped by AdjNorm and PDAC disease states. B. Communication probabilities of signaling pathways targeting NK cells between AdjNorm and PDAC disease states as measured by information flow (pathway legend: pink = AdjNorm-specific, teal = PDAC-specific, black = equally specific). C. Significant ligand-receptor pairs from collagen, FN1, and laminin signaling pathways targeting NK cells upregulated in PDAC compared to AdjNorm. Violin plots showing differential expression of CD44 between PDAC (teal) and AdjNorm (pink) disease states in D. normal epithelial cells, E. malignant epithelial cells, F. fibroblasts, and G. NK cells (NS = not significant, ** P < 0.01, **** P < 0.0001, Bonferroni correction).

Article Snippet: Donor NK cells were added to Transwells with or without an α-CD44 neutralizing antibody (clone IM7; Bio X Cell; C f = 10 µg/ml) and incubated for 4 hours (37°C).

Techniques: Expressing

Journal: bioRxiv

Article Title: Natural killer cells associate with epithelial cells in the pancreatic ductal adenocarcinoma tumor microenvironment

doi: 10.1101/2024.05.23.593868

Figure Lengend Snippet: A. Schematic for Transwell invasion assays. Average relative invasion + SEM of B. Donor NK #1 (n = 9), C. Donor NK #2 (n = 12), D. Donor NK #3 (n = 6), and E. Donor NK #4 (n = 10) upon CD44 neutralization. * P < 0.05 as determined by Wilcoxon matched-pairs signed rank test. F. Schematic of spheroid invasion assay. G. Representative 20X immunofluorescence images of outlined (yellow) PANC-1 spheroids (pan-cytokeratin; red) embedded with NK cells (CD56; green), pointed out with white arrows, treated with or without α-CD44. Scale bar = 250 µm. H. Zoomed inset of NK cells in the α-CD44 treatment group in G (white box). Average number of NK cells per PANC-1 spheroid + SEM from I. Donor NK #1 (- α-CD44, n = 42; + α-CD44, n = 53), J. Donor NK #2 (- α-CD44, n = 43; + α-CD44, n = 29), and K. Donor NK #3 (- α-CD44, n = 47; + α-CD44, n = 27); n = # of spheroids analyzed; * P < 0.05 as determined by Wilcoxon matched-pairs signed rank test.

Article Snippet: Donor NK cells were added to Transwells with or without an α-CD44 neutralizing antibody (clone IM7; Bio X Cell; C f = 10 µg/ml) and incubated for 4 hours (37°C).

Techniques: Neutralization, Invasion Assay, Immunofluorescence

Journal: Cell Communication and Signaling : CCS

Article Title: All-trans retinoic acid pretreatment of mesenchymal stem cells enhances the therapeutic effect on acute kidney injury

doi: 10.1186/s12964-024-01671-1

Figure Lengend Snippet: HA activates the downstream PI3K/AKT signaling pathway by binding to CD44. a, b Representative images and quantification of immunofluorescence staining of HK-2 cells with CD44 (red) and DAPI (blue) ( n = 3). Scale bars, 20 μm. c, d Representative immunofluorescence staining of CD44 in renal tissue ( n = 6). Scale bars, 20 μm. e, f Representative immunohistochemical staining of CD44 in renal tissue ( n = 6). Scale bars, 40 μm. g-j Representative western blot images and quantitative analysis of p-AKT and total AKT levels in kidney tissues ( n = 3). k-n Representative western blot images and quantitative analysis of p-AKT and total AKT levels in HK-2 cells ( n = 3). Data are presented as the means ± SD. ** P < 0.01, * P < 0.05

Article Snippet: To block HA interactions to CD44, anti-CD44 neutralizing antibody (10 μg/mL, 9400-01, Southern Biotech, USA)) was added to HK-2 cells culture medium before hypoxia treatment.

Techniques: Binding Assay, Immunofluorescence, Staining, Immunohistochemical staining, Western Blot

Journal: Cell Communication and Signaling : CCS

Article Title: All-trans retinoic acid pretreatment of mesenchymal stem cells enhances the therapeutic effect on acute kidney injury

doi: 10.1186/s12964-024-01671-1

Figure Lengend Snippet: Inhibition of the HA/CD44 axis reverses the anti-inflammatory, anti-apoptotic and pro-proliferative effects of ATRA-MSCs. a, c-i Western blot images and quantitative densitometry analysis of TNF-a, IL-6, Bax, Bcl-2 and cleaved caspase-3 in HK-2 cells ( n = 3). b, j-l Western blot images and quantitative densitometry analysis of p-AKT and total AKT in HK-2 cells ( n = 3). Data are presented as the means ± SD. **** P < 0.0001, *** P < 0.001, ** P < 0.01, * P < 0.05

Article Snippet: To block HA interactions to CD44, anti-CD44 neutralizing antibody (10 μg/mL, 9400-01, Southern Biotech, USA)) was added to HK-2 cells culture medium before hypoxia treatment.

Techniques: Inhibition, Western Blot

Journal: Cell Communication and Signaling : CCS

Article Title: All-trans retinoic acid pretreatment of mesenchymal stem cells enhances the therapeutic effect on acute kidney injury

doi: 10.1186/s12964-024-01671-1

Figure Lengend Snippet: Schematic illustration of ATRA promoted HA production by MSCs and activated the PI3K/AKT pathway by binding to CD44 in renal tubular epithelial cells, thereby improving renal repair after AKI

Article Snippet: To block HA interactions to CD44, anti-CD44 neutralizing antibody (10 μg/mL, 9400-01, Southern Biotech, USA)) was added to HK-2 cells culture medium before hypoxia treatment.

Techniques: Binding Assay

Journal: Scientific Reports

Article Title: Melanocortin-4 receptor in macrophages attenuated angiotensin II-induced abdominal aortic aneurysm in mice

doi: 10.1038/s41598-023-46831-4

Figure Lengend Snippet: An osteopontin receptor CD44 mediates vascular vulnerability of MC4R TB/TB mice. ( A ) Leptin (100 nM)-induced Spp1 gene expression in primary vascular smooth muscle cells from WT male mice pretreated with or without the PI3K inhibitor LY294002 (LY, 10 nM; n = 4). ( B ) Representative pictures and ( C ) quantification of osteopontin (OPN) immunostaining in the aorta of WD-fed MC4R + / + and MC4R TB/TB mice (n = 6, scale bar is 100 μm). ( D ) Representative pictures of DAPI, α-smooth muscle cells, and merged images in the aorta of WD-fed MC4R TB/TB mice (scale bars represent 20 μm). ( E ) Changes in the sBP of 14–18-weeks-old MC4R TB/TB mice during Ang II (500 ng/kg/min) infusion concomitantly administered with anti-CD44 or control antibodies (Ab; n = 6–12). ( F ) Representative pictures of the aorta and ( G ) AAA incidence and ( H ) diameter after Ang II infusion into WD-fed MC4R + / + and MC4R TB/TB mice administered with anti-CD44 or control Abs (n = 6–12). * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: In the experiment of α-MSH (M4135, Sigma-Aldrich, 1 mg/kg/day) or anti-CD44 neutralizing antibody (BE0039, BioXCell, 200 μg/day) treatment, they were injected intraperitoneally into ApoE KO or MC4R TB/TB mice for 4 weeks starting on the day the minipumps were implanted.

Techniques: Expressing, Immunostaining

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Single-Cell RNA Sequencing of Coronary Perivascular Adipose Tissue From End-Stage Heart Failure Patients Identifies SPP1 + Macrophage Subpopulation as a Target for Alleviating Fibrosis

doi: 10.1161/ATVBAHA.123.319828

Figure Lengend Snippet: Conditioned medium from perivascular adipose tissue (PVAT) of patients with atherosclerosis (AS) significantly promotes the fibrogenesis of human fibroadipogenic progenitors (FAPs) through CD44/integrin axis. A , Schematic representation of the experimental procedure. B , Representative images of transwell migration assay ( top ) and EdU (5-ethynyl-2’-deoxyuridine) proliferation assay ( bottom ) of human FAP cells. Scale bar, 100 µm. C and D , Quantitative analysis of transwell migration assay ( C ) and EdU proliferation assay ( D ) of human FAP cells. Four points in each group in C and D represent 4 biological replicates. Comparisons of the cell migration or proliferation between each 2 groups were performed using the Student t test; P values were adjusted for multiple hypothesis testing using the Benjamini-Hochberg method.

Article Snippet: FAP cells were pretreated with vehicle or anti-CD44 (cluster of differentiation 44) neutralizing monoclonal antibody (10 μg/mL, 15675-1-AP; Proteintech), RGD (arginylglycylaspartic acid) peptide to inhibit integrin receptors (10 μM, A8052; Sigma Aldrich), or both in combination for 30 minutes as described previously.

Techniques: Transwell Migration Assay, Proliferation Assay, Migration