Journal: PLoS ONE

Article Title: DIOL Triterpenes Block Profibrotic Effects of Angiotensin II and Protect from Cardiac Hypertrophy

doi: 10.1371/journal.pone.0041545

Figure Lengend Snippet: Time course of angiotensin II (Ang II; 1 µM)-stimulated collagen I protein production (A). Effect of erythrodiol (ERY; 5 µM) or uvaol (UVA; 5 µM) on CTGF (B), collagen I (C) and galectin 3 (D) protein expression in angiotensin II-treated cardiac myofibroblasts for 12 hours. Representative immunoblots of 4 experiments. Values are mean ± SEM of four assays. *p<0.05 vs vehicle. †p<0.05 vs angiotensin II. Quantification of band intensities was measured by densitometry and normalized to respective α-tubulin.

Article Snippet: Membranes were probed with primary antibody for α-SMA (Biocare Medical, CA, USA), PPAR-γ (Santa Cruz, Inc, USA), collagen I (AbD Serotec, Oxford, UK), connective tissue growth factor (CTGF; Torrey Pines Biolabs Inc, East Orange, NJ) and galectin 3 (Thermo Scientific, Rockford, IL) followed by incubation with an HRP-linked secondary antibody.

Techniques: Expressing, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Galectin-3 Mediates NETosis and Acts as an Autoantigen in Systemic Lupus Erythematosus-Associated Diffuse Alveolar Haemorrhage

doi: 10.3390/ijms24119493

Figure Lengend Snippet: Expression of galectin-3 in patients with systemic lupus erythematosus (SLE) and mice with pristane-induced lupus. ( A ) Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) of galectin-3 in PBMCs of SLE patients (SLE) and in the same cohort with (LN-Yes) or without (LN-Nil) lupus nephritis and normal donors (Normal). ( B ) The significance of correlation between the expression levels of galectin-3 and SLE disease activity index 2000 (SLEDAI-2K) was calculated using Pearson’s correlation coefficient. ( C ) Anti-galectin-3 antibody levels in sera of SLE patients and normal donors. ( D ) Female mice aged 8 weeks were injected with 500 μL of pristane or phosphate-buffered saline (PBS) via the intra-peritoneal routes. Mice died from diffuse alveolar haemorrhage (DAH) around week 4 after pristane injection, as demonstrated by H&E staining. Immunohistochemical stainings of galectin-3 and moue IgG Fc in lung tissue sections from PBS and pristane-treated mice. Boxed areas are shown at higher magnification in the panels next to them. ( E ) Expression of galectin-3, citrullinated histone 3 (citH3), and histone 3 (H3) in lung tissue extracts from PBS and pristane-treated mice (n = 4). ( F ) Immunofluorescent stainings of myeloperoxidase (MPO, Texas red), and Ly6G (FITC) in lung tissue sections from PBS and pristane-treated WT and galectin-3 knockout (Gal-3 KO) mice. Boxed areas indicate MPO and Ly6G double-positive signals (×400 magnification). DAPI indicates nuclear staining. Values were presented as mean and SEM. Bars shown on the photomicrographs at ×40, ×100, and ×400 magnifications correspond to 500, 200, and 50 μm, respectively.

Article Snippet: NETotic cells were fixed with 4% paraformaldehyde and stained with DAPI for 10 min or with anti-human galectin-3 or anti-human MPO (Cat.#ab208670, Abcam) antibody and followed by Texas-red conjugated secondary antibody (Cat.#112-075-003, Jackson ImmunoResearch Laboratories) for immunofluorescence analysis.

Techniques: Expressing, Reverse Transcription, Polymerase Chain Reaction, Quantitative RT-PCR, Activity Assay, Injection, Saline, Staining, Immunohistochemical staining, Knock-Out

Journal: International Journal of Molecular Sciences

Article Title: Galectin-3 Mediates NETosis and Acts as an Autoantigen in Systemic Lupus Erythematosus-Associated Diffuse Alveolar Haemorrhage

doi: 10.3390/ijms24119493

Figure Lengend Snippet: Associations of galectin-3 and lupus manifestations in pristane-treated galectin-3 knockout (Gal-3 KO) mice. Female wild-type (WT) and Gal-3 KO mice aged 8 weeks were injected with 500 μL of pristane via the intra-peritoneal routes. ( A ) Representative images of DAH and inflammation from pristane-treated Gal-3 KO mice compared with WT counterparts, as demonstrated by H&E staining (n = 3). Boxed areas are shown at higher magnification in the panels beneath them ( B ) Percent survival in pristane-induced WT and Gal-3 KO mice. ( C ) Representative images of renal sections from pristane-treated Gal-3 KO mice compared with their WT counterparts, as demonstrated by H&E staining (×400 magnification). ( D ) Levels of proteinuria in pristane-induced WT and Gal-3 KO mice. ( E ) Levels of anti-RNP antibodies in sera from pristane-induced WT and Gal-3 KO mice, as determined by ELISA. ( F ) Expression of galectin-3, citH3, and H3 in lung tissue extracts from pristane-induced WT and Gal-3 KO mice, as determined by immunoblotting (n = 4). Values were presented as mean and SEM. Bars shown on the photomicrographs at ×40, ×100, and ×400 magnifications correspond to 500, 200, and 50 μm, respectively.

Article Snippet: NETotic cells were fixed with 4% paraformaldehyde and stained with DAPI for 10 min or with anti-human galectin-3 or anti-human MPO (Cat.#ab208670, Abcam) antibody and followed by Texas-red conjugated secondary antibody (Cat.#112-075-003, Jackson ImmunoResearch Laboratories) for immunofluorescence analysis.

Techniques: Knock-Out, Injection, Staining, Enzyme-linked Immunosorbent Assay, Expressing, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Galectin-3 Mediates NETosis and Acts as an Autoantigen in Systemic Lupus Erythematosus-Associated Diffuse Alveolar Haemorrhage

doi: 10.3390/ijms24119493

Figure Lengend Snippet: Effects of galectin-3 on NETosis. ( A ) Representative images of neutrophils isolated from normal donors and analyzed at baseline (T0) or after stimulation with phorbol myristate acetate (PMA, 100 μM) for 4 h (T4). Cells were stained with DAPI for 10 min or with anti-human galectin-3 or with anti-human MPO antibody and followed by Texas-red conjugated secondary antibodies for immunofluorescence analysis. ( B ) Representative images and quantification of neutrophils isolated from normal donors and treated with lactose or glucose (20 and 50 mM) at T4. Cells were stained with DAPI for 10 min. Values were presented as mean and SEM (n = 3). ( C ) NETosis and ( D ) Galectin-3, citH3, and H3 expression in lipopolysaccharide (LPS, 20 μM)-treated neutrophils from WT and Gal-3 KO mice. ( E ) Representative images of PBMCs isolated from SLE patients or normal donors and analyzed at T0. Cells were stained with DAPI for 10 min or with rat anti-human galectin-3 antibody or sera from indicated donors and followed by Texas-red conjugated anti-rat or FITC-conjugated anti-human IgG secondary antibody for immunofluorescence analysis. Higher-magnification views of the merged images are shown on the far right. The yellow arrows indicate immune complex deposition. Bars shown on the photomicrographs at ×100 and ×400 magnifications correspond to 200 and 50 μm, respectively. * p < 0.05, *** p < 0.001.

Article Snippet: NETotic cells were fixed with 4% paraformaldehyde and stained with DAPI for 10 min or with anti-human galectin-3 or anti-human MPO (Cat.#ab208670, Abcam) antibody and followed by Texas-red conjugated secondary antibody (Cat.#112-075-003, Jackson ImmunoResearch Laboratories) for immunofluorescence analysis.

Techniques: Isolation, Staining, Immunofluorescence, Expressing