Journal: PLoS ONE

Article Title: Neutrophils remain detrimentally active in hydroxyurea-treated patients with sickle cell disease

doi: 10.1371/journal.pone.0226583

Figure Lengend Snippet: (A) Purified and rested neutrophils from healthy donors (H) and SCD patients in steady state (SS) were left untreated in RPMI as control or treated with 20 μM hemin. Median Fluorescence Intensity of F-Actin (upper panel) and CD63 (lower panel) in F-Actin/CD63 double positive neutrophils with normal multi-lobulated nuclei is shown. (Healthy N = 7; SCD N = 6). (B) Z-stack stills of SCD neutrophils left in RPMI for 30 minutes showing neutrophils with roughed cell membranes (yellow arrows, upper panel). Staining for DNA (blue), F-Actin (green) and CD63 (red) is presented (lower panel). (C) MPO release was assayed by ELISA following 30 minutes incubation with or without the hemin stimulus (H (Healthy) N = 4; SS (SCD) N = 4). Degranulation response was assayed in terms of MPO ng/ml plasma (upper panel), or fold increase over healthy neutrophils (lower panel). Data presented as dot plots±S.D., significance calculated with an unpaired Mann-Whitney test.

Article Snippet: The supernatants were then assayed with a Human Myeloperoxidase Quantikine ELISA Kit from R&D Systems according to the manufacturer’s instruction.

Techniques: Purification, Control, Fluorescence, Staining, Enzyme-linked Immunosorbent Assay, Incubation, Clinical Proteomics, MANN-WHITNEY

Journal: bioRxiv

Article Title: IL-11 neutralising therapies target hepatic stellate cell-induced liver inflammation and fibrosis in NASH

doi: 10.1101/470062

Figure Lengend Snippet: a , IL-11 is upregulated in hepatic stellate cells (HSCs) stimulated with TGFβ1 (n=3). b , IL-11 protein is secreted from HSCs stimulated with TGFβ1 (ELISA, n=3). c , Human precision cut liver slices were stimulated with TGFβ1 and IL-11 protein was measured in supernatant (ELISA, n=3). d , IL6R, gp130, and IL11RA expression in HSCs (TPM, transcripts per million). e, f , Representative fluorescence images (scale bars, 200 µm) of HSCs and automated fluorescence quantification for ( e ) ACTA2 +ve cells and ( f ) Collagen I immunostaining following incubation without stimulus (-), with TGFβ1, PDGF, or IL-11. g , Collagen secretion supernatants of HSC stimulated with TGFβ1, PDGF, or IL-11 (Sirius red assay. n≥7). h , Dose-dependent matrigel invasion of HSCs induced by IL-11(n=3). i , Hyper IL-11 induces IL-11 protein secretion from HSCs (ELISA, n=3). a-c, e-g, i , TGFβ1 (5 ng ml -1 ), Hyper IL-11 (0.2 ng ml -1 ), PDGF (20 ng ml -1 ), IL-11 (5 ng ml -1 ); 24 h stimulation; h , 48 h stimulation. j , Schematic of mice receiving daily subcutaneous injection of either saline (control) or rmIl-11 (100 µg kg -1 ). k , Relative liver hydroxyproline content, l , mRNA expression of pro-fibrotic markers, and m , serum ALT levels (k, l , control, n=7; rmIl-11, n=10; m , control n=8; rmIl-11, n=11). a, b, g, h, i, k, m Data are represented as mean ±s.d; c-f, l , Box-and-whisker plots show median (middle line), 25th–75th percentiles (box) and min-max percentiles (whiskers). a-d, i, k-m , Two-tailed Student’s t -test; e-h , two-tailed Dunnett’s test. FC: fold change; I/A: intensity/area.

Article Snippet: Total secreted collagen in the cell culture supernatant was quantified using Sirius red collagen detection kit (9062, Chondrex).

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Fluorescence, Immunostaining, Incubation, Injection, Whisker Assay, Two Tailed Test

Journal: Open Access Rheumatology : Research and Reviews

Article Title: A suppressive effect of prostaglandin E 2 on the expression of SERPINE1 /plasminogen activator inhibitor-1 in human articular chondrocytes: An in vitro pilot study

doi:

Figure Lengend Snippet: Suppressive effect of PGE 2 on PAI-1 expression. A ) Western blot. Left: Chondrocytes were stimulated with PGE 2 (0.1, 1, or 10 nM) for 24 h, and the expression of PAI-1 was analyzed. NS: not stimulated. Representative results are shown. Bottom: Calculation of the relative intensity of the bands among four OA samples tested. B ) ELISA. Levels of PAI-1 in culture supernatants of chondrocytes after PGE 2 stimulation (0.1–10 nM) were measured. Notes: OA chondrocytes, N = 3, duplicate. Abbreviations: ELISA, enzyme-linked immunosorbent assay; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; PAI-1, plasminogen activator inhibitor; PGE 2 , prostaglandin E 2 .

Article Snippet: The level of PAI-1 in the culture supernatant was measured by using an ELISA kit (AssayMax Human Plasminogen activator inhibitor-1 ELISA Kit™, AssayPro LLC, St. Charles, MO, USA).

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

Journal: Open Access Rheumatology : Research and Reviews

Article Title: A suppressive effect of prostaglandin E 2 on the expression of SERPINE1 /plasminogen activator inhibitor-1 in human articular chondrocytes: An in vitro pilot study

doi:

Figure Lengend Snippet: The PAI-1 suppression by PGE 2 is delivered through EP4 receptor. The summary of PAI-1 ELISA with receptor antagonists are shown. Chondrocytes were stimulated with PGE 2 (10 nM) with or without AH6809 (10 ng/ml) or GW627368X (5 μM), and the levels of PAI-1 in culture supernatants were measured. Notes: OA chondrocytes, N = 6. Abbreviations: ELISA, enzyme-linked immunosorbent assay; PAI-1, plasminogen activator inhibitor; PGE 2 , prostaglandin E 2 .

Article Snippet: The level of PAI-1 in the culture supernatant was measured by using an ELISA kit (AssayMax Human Plasminogen activator inhibitor-1 ELISA Kit™, AssayPro LLC, St. Charles, MO, USA).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Open Access Rheumatology : Research and Reviews

Article Title: A suppressive effect of prostaglandin E 2 on the expression of SERPINE1 /plasminogen activator inhibitor-1 in human articular chondrocytes: An in vitro pilot study

doi:

Figure Lengend Snippet: Inhibition of p38 and ERK MAPK did not abrogate the effect of PGE 2 on PAI-1 downregulation. The summary of PAI-1 ELISA with receptor antagonists are shown. Chondrocytes were stimulated with PGE 2 (10 nM) with or without SB203580 or PD98059, and the levels of PAI-1 in culture supernatants were measured. Notes: OA chondrocytes, N = 3. *p <0.05; **p <0.02. Abbreviations: MAPK, mitogen-activated protein kinases, PAI-1, plasminogen activator inhibitor; PGE 2 , prostaglandin E 2 .

Article Snippet: The level of PAI-1 in the culture supernatant was measured by using an ELISA kit (AssayMax Human Plasminogen activator inhibitor-1 ELISA Kit™, AssayPro LLC, St. Charles, MO, USA).

Techniques: Inhibition, Enzyme-linked Immunosorbent Assay