Journal: bioRxiv

Article Title: Apical expansion of calvarial osteoblasts and suture patency is dependent on graded fibronectin cues

doi: 10.1101/2023.01.16.524278

Figure Lengend Snippet: (A) Immunofluorescence of unfolded collagen chains in E12.5 coronal sections. Immunofluorescence of collagen III (B), Laminin (C), RGB Trichrome staining for collagen (D) in E12.5 coronal sections. (E, F, G) Immunofluorescence of FN1 in coronal sections showing graded FN1 protein expression in cranial mesenchyme at E11.5-13.5. N≥3 controls; 3 mutants. (H) Schematic depicting the graded FN1 substrate towards the apex of the cranial mesenchyme. Nuclei were labeled with DAPI in blue. CHP, collagen hybridizing peptide; FN1, Fibronectin 1; fbp, frontal bone primordia. mn, meninges. Scale Bar= 100 μM.

Article Snippet: Following primary antibodies were used: Collagen hybridizing peptide (CHP) (Hwang et al., 2017; Jussila et al., 2021) (1: 250; 3Helix BIO60), rabbit anti-Fibronectin antibody (1: 250; Abcam ab2413; RRID:AB_2262874), rabbit anti-Fibulin 1 (1: 250; Abcam ab230994), rabbit anti-Collagen I (1: 250; Abcam ab270993; RRID:AB_2927551), Collagen III (Rockland Labs 1:100), rabbit anti-Laminin (1: 250; Millipore L9393), rabbit anti-Osx (1:2000 or 1:4000; Abcam ab209484; RRID:AB_2892207), rabbit anti-Caspase 3 (1: 250; Abcam ab13847; RRID:AB_443014), Goat anti-Runx 2 (1:100; R&D Systems AF2006-SP; RRID:AB_2184528) and mouse anti-GM130 (1:100; Thermo Fisher Scientific BDB610822; RRID:AB_398141).

Techniques: Immunofluorescence, Staining, Expressing, Labeling

Journal: Hypertension

Article Title: Calpain-1 Regulation of Matrix Metalloproteinase 2 Activity in Vascular Smooth Muscle Cells Facilitates Age-Associated Aortic Wall Calcification and Fibrosis

doi: 10.1161/hypertensionaha.112.196840

Figure Lengend Snippet: Figure 3. Overexpression of calpain-1 induces collagen type I, II, and III and reduces elastin expression levels in vascular smooth muscle cells (VSMC). A, Representative Western blots of collagen I and collagen III in VSMC after infection by GFP or CANP1 adenoviruses for 48 hours. B, Photomicrographs (×400) of collagen II protein staining (brown color) from young (8-month) and old (30-month) rat aortic walls. Nuclei are counterstained with hematoxylin (blue color). L indicates lumen; M, media. C and D, Representative Western blots and average protein levels of collagen II (C) and elastin (D) in young VSMC infected with GFP or CANP1 for 48 hours. *P<0.05 compared with GFP-infected 8-month VSMC, n=3.

Article Snippet: Primary antibodies used for immunostaining (IS) and Western blotting (WB) Antibody Species Titer for IS Titer for WB Source -actin Mouse 1:10000 Sigma-Aldrich Co. LLC, St. Louis, MO calpain-1 Rabbit 1:100 1:2500 Sigma-Aldrich Co. LLC, St. Louis, MO collagen I Rabbit 1:1000 Rockland Antibodies & Assays, Gilbertsville, PA collagen II Rabbit 1:50 1:1000 Abcam, Cambridge, MA collagen III Rabbit 1:1000 Rockland Antibodies & Assays, Gilbertsville, PA elastin Rabbit 1:1000 Millipore, Billerica, MA MMP2 Mouse 1:50 1:1000 Millipore, Billerica, MA MT1MMP Rabbit 1:1000 Millipore, Billerica, MA osteonectin Rabbit 1:1000 Millipore, Billerica, MA osteoponin Rabbit 1:1000 Millipore, Billerica, MA Smad 2/3 Mouse 1:1000 Cell Signaling Technology, Inc, Danvers, MA TGF-1 Rabbit 1:500 Santa Cruz Biotechnology, Inc, Santa Cruz, CA TIMP2 Goat 1:500 R&D Systems, Inc. Minneapolis, MN 9 Figure S1: Interactions between calpain-1 and MMP2.

Techniques: Over Expression, Expressing, Western Blot, Infection, Staining

Journal: Hypertension

Article Title: Calpain-1 Regulation of Matrix Metalloproteinase 2 Activity in Vascular Smooth Muscle Cells Facilitates Age-Associated Aortic Wall Calcification and Fibrosis

doi: 10.1161/hypertensionaha.112.196840

Figure Lengend Snippet: Figure 6. Overexpression of calpain-1 regulates extracellular matrix remodeling, particularly fibrosis and vascular calcification in cultured carotid artery rings. Representative Western blot of tissue inhibitor of metalloprotease 2 (TIMP2), calpain-1, elastin, transforming growth factor-β1 (TGFβ1), (A) Smad 2/3, collagen I, II, III, (B) and osteopontin (OPN) and osteonectin (ON) (C) in cultured carotid artery rings isolated from 8-month-old rat after infection by GFP or CANP1 or TIMP2 adenoviruses for 48 hours.

Article Snippet: Primary antibodies used for immunostaining (IS) and Western blotting (WB) Antibody Species Titer for IS Titer for WB Source -actin Mouse 1:10000 Sigma-Aldrich Co. LLC, St. Louis, MO calpain-1 Rabbit 1:100 1:2500 Sigma-Aldrich Co. LLC, St. Louis, MO collagen I Rabbit 1:1000 Rockland Antibodies & Assays, Gilbertsville, PA collagen II Rabbit 1:50 1:1000 Abcam, Cambridge, MA collagen III Rabbit 1:1000 Rockland Antibodies & Assays, Gilbertsville, PA elastin Rabbit 1:1000 Millipore, Billerica, MA MMP2 Mouse 1:50 1:1000 Millipore, Billerica, MA MT1MMP Rabbit 1:1000 Millipore, Billerica, MA osteonectin Rabbit 1:1000 Millipore, Billerica, MA osteoponin Rabbit 1:1000 Millipore, Billerica, MA Smad 2/3 Mouse 1:1000 Cell Signaling Technology, Inc, Danvers, MA TGF-1 Rabbit 1:500 Santa Cruz Biotechnology, Inc, Santa Cruz, CA TIMP2 Goat 1:500 R&D Systems, Inc. Minneapolis, MN 9 Figure S1: Interactions between calpain-1 and MMP2.

Techniques: Over Expression, Cell Culture, Western Blot, Isolation, Infection

Journal: Photonics

Article Title: In Situ Structural Characterization of Cardiomyocyte Microenvironment by Multimodal STED Microscopy

doi: 10.3390/photonics11060533

Figure Lengend Snippet: Reconstructed models show the endocardium (in AF), cardiomyocytes (in AF), and interstitial matrix (in B-SHG) within papillary muscle cross-sectioned areas. Collagen type III surrounds individual cardiomyocytes and the cardiomyocyte bundle (top row). β1 integrin distributes on cardiomyocytes and endothelial cells (bottom row). Arrowheads highlight nuclei of squamous endothelial cells within the endocardium. Z-step, 0.5 μm. Stack depth, 13 μm (top row); 11.5 μm (bottom row).

Article Snippet: Sections were blocked in 2% ( w / v ) bovine serum albumin with 0.3% Triton X-100 at room temperature, then incubated with anti-laminin (ab11575, Abcam, Cambridge, UK), β1 integrin (ab179471, Abcam), collagen type III (600-401-105, Rockland, Philadelphia, USA), or collagen type IV (ab179471, Abcam) at 4 °C overnight.

Techniques: