Journal: Nature Communications

Article Title: Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

doi: 10.1038/s41467-021-27622-9

Figure Lengend Snippet: a Quantification of BMP1.3 protein levels in blood of healthy individuals and patients affected by acute myocardial infarction (MI) within 12 h from hospitalization. b Quantification of the expression levels of BMP1.3 normalized over GAPDH in human myocardium of patients without overt cardiac disease (control) or affected by ischemic heart disease (ischemic). c Quantification of BMP1-3 protein levels in the blood of sham-operated or infarcted mice at 24 h after MI. d Quantification of the expression levels of Bmp1.3 normalized over 18 S in the myocardium of sham-operated or infarcted mice at 48 h after MI. e Quantification of the levels of Bmp1.3 mRNA normalized on Actb in different cardiac cell types at 48 h after MI. f Quantification of the ejection fraction in rats treated with isoproterenol (Ip), alone (control) or in combination with anti-BMP1.3 antibody (150 µg/kg). Values for sham animals are also shown for reference. g Quantification of the area covered by collagen in each section. h Representative images of heart sections stained with Sirius red to show collagen deposition upon administration of isoproterenol alone (control) or in combination with anti-BMP1.3 antibody. i Quantification of the area covered by collagen in sections of murine hearts sham-operated or injured by myocardial infarction (MI) and treated with the indicated dose of anti-BMP1.3 antibody (150 or 500 µg/kg). j Representative images of murine heart sections stained with Sirius red to show collagen deposition upon MI and treatment with the indicated dose of anti-BMP1.3 antibody. k Quantification of the ejection fraction at 7, 14, 30, and 60 days after MI. Data in a , b , c , d , e , f , g , i , and k are shown as mean ± s.e.m. Sample number is indicated inside or above each bar. Statistical significance was determined using unpaired, two-sided t -test in a – d , one-way ANOVA, followed by Tukey’s test in e , f , g , i , and two-way ANOVA in k followed by Tukey’s multiple comparison test. Scale bar in h , j is 200 µm and 1 mm, respectively. Source data are provided as a Source Data file.

Article Snippet: Plasma levels of BMP1.3 were measured using the ELISA Human BMP1 Matched Antibody Pair Kit (Cloud-Clone PSA653Hu01) and additional reagents supplied in Antibody Pairs Support Pack (Cloud-Clone IS077).

Techniques: Expressing, Staining

Journal: Nature Communications

Article Title: Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

doi: 10.1038/s41467-021-27622-9

Figure Lengend Snippet: a Representative images of heart sections from COLL-EGFP mice after MI (control) and after treatment with anti-BMP1.3 antibody (150 µg/kg). Nuclei were stained with DAPI. b Quantification of the EGFP + area normalized on DAPI + area. c Quantification of both EGFP + or RFP + pixels in cultured cardiac fibroblasts from COLL-EGFP/SMA-RFP mice untreated (control), treated with human recombinant BMP1.3 (rhBMP1.3, 0.1 µg/ml) or anti-BMP1.3 antibody (BMP1.3 Ab, 5 µg/ml). d Representative images of cultured cardiac fibroblast from COLL-EGFP/SMA-RFP mice treated as indicated. e Quantification of the expression levels of Col1a , Lox, Ctgf, Fn , and Tgf1b genes normalized to Gapdh in cultured cardiac fibroblasts treated as described. f Representative images from tissue section of infarct area of animals untreated (control) or treated with anti-BMP1.3 antibody (150 µg/kg) stained with anti-Tgfβ1, anti-phospho-Smad2 and anti-CTGF antibodies (brown) and hematoxylin (violet) to visualize nuclei. g Quantification of the expression levels of Col1a , Ctgf, Fn, Lox , and Tgf1b genes normalized to Gapdh in hearts, either untreated (control) or treated with anti-BMP1.3 antibody (150 µg/kg). h Representative images from tissue section of the heart of sham animals or animals subjected to MI, either in the absence of treament (control) or treated with anti-BMP1.3 antibody (150 µg/kg), stained with anti-Lox antibodies (brown) and hematoxylin (violet) to visualize nuclei. i Biochemical analysis of collagen crosslinking in scar tissue dissected from hearts of untreated rats (control) or rats treated with anti-BMP1.3 antibody (150 µg/kg). DHNL dihydroxynorleucine, DHLNL dihydroxylysinonorleucine, HLNL hydroxylysinonorleucine, Pyr pyridinoline, d-Pyr deoxypyridinoline, ALD total aldehyde, NON RED non reduced aldehyde. Data in b , c , e , g and i are shown as mean ± s.e.m. Sample number is indicated inside or above each bar. Statistical significance was determined using unpaired, two-sided t -test in b , e , g and i or one-way ANOVA followed by Tukey’s multiple comparison test in c . Scale bars in a , f and h is 100 µm, while in d is 500 µm. Source data are provided as a Source Data file.

Article Snippet: Plasma levels of BMP1.3 were measured using the ELISA Human BMP1 Matched Antibody Pair Kit (Cloud-Clone PSA653Hu01) and additional reagents supplied in Antibody Pairs Support Pack (Cloud-Clone IS077).

Techniques: Staining, Cell Culture, Recombinant, Expressing

Journal: Nature Communications

Article Title: Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

doi: 10.1038/s41467-021-27622-9

Figure Lengend Snippet: a Representative images of TUNEL and α-actinin staining on heart sections of mice untreated (sham) or injured by a MI in the absence of treatment (control) or after injection of anti-BMP1.3 antibody (150 µg/kg). Nuclei were stained with DAPI. b Quantification of dying cells in the infarcted area. c Quantification of TUNEL + nuclei in primary cardiomyocytes and cardiac fibroblasts. d Representative images of primary cardiac cells, either untreated (control) or treated with anti-BMP1.3 antibody (5 µg/ml), stained with TUNEL and anti-α-actinin antibody. Nuclei were stained with DAPI. e Representative images of primary cardiac fibroblasts and cardiomyocytes, cultured in hypoxic conditions, either in the absence (control) or in the presence of recombinant BMP1.3 (0.1 µg/ml), stained with TUNEL and anti-α-actinin antibody. Nuclei were stained with DAPI. f Quantification of TUNEL + nuclei in cardiomyocytes and cardiac fibroblasts. Data in b , c , and f are shown as mean ± s.e.m. Sample number is indicated inside or above each bar. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparison test in b , c and f . Scale bars in a , d and e indicate 100 µm. Source data are provided as a Source Data file.

Article Snippet: Plasma levels of BMP1.3 were measured using the ELISA Human BMP1 Matched Antibody Pair Kit (Cloud-Clone PSA653Hu01) and additional reagents supplied in Antibody Pairs Support Pack (Cloud-Clone IS077).

Techniques: TUNEL Assay, Staining, Injection, Cell Culture, Recombinant

Journal: Nature Communications

Article Title: Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

doi: 10.1038/s41467-021-27622-9

Figure Lengend Snippet: a Quantification of the expression levels of Bmp2 and Bmp5 in primary cardiac fibroblasts and cardiomyocytes cultured in hypoxic conditions in the absence of treatment (control) or treated with anti-BMP1.3 antibody (5 µg/ml), normalized to Gapdh . b Quantification of the expression levels of Bmp2 and Bmp5 in the heart of mice subjected to MI in the absence of treatment (control) or after injection of anti-BMP1.3 antibody (150 µg/kg). c Representative images from tissue section of infarct area of untreated rats (control) or treated with anti-BMP1.3 antibody (150 µg/kg) stained with either anti-BMP2 or anti-BMP5 antibody (brown) and hematoxylin (violet) to visualize nuclei. d Quantification of living cardiomyocytes cultured in hypoxic conditions expressed as number of cardiomyocytes per field and exposed to the indicated enriched supernatants collected from CHO transfected with control plasmid (pGI) or plasmid for the overexpression of BMP2 and/or BMP5. e Representative images of rat neonatal cardiomyocytes cultured in hypoxic conditions and exposed to the indicated treatments. Cardiomyocytes were stained with anti-α-actinin antibody and nuclei with DAPI. f Quantification of living cardiomyocytes cultured in hypoxic conditions and exposed to the indicated treatments. g Representative images of rat neonatal cardiomyocytes cultured in hypoxic conditions in the absence of treatment (−Ab) or treated with anti-BMP1.3 antibody (+Ab) and transfected with the indicated siRNAs (mock = scramble siBMP5). Data in a , b , d and f are shown as mean ± s.e.m. Sample number is indicated inside or above each bar. Statistical significance was determined using unpaired, two-sided t -test in a , b or one-way ANOVA followed by Dunnet’s in d or Tukey’s multiple comparison test in f . Scale bar in c , e and g is 100 µm. Source data are provided as a Source Data file.

Article Snippet: Plasma levels of BMP1.3 were measured using the ELISA Human BMP1 Matched Antibody Pair Kit (Cloud-Clone PSA653Hu01) and additional reagents supplied in Antibody Pairs Support Pack (Cloud-Clone IS077).

Techniques: Expressing, Cell Culture, Injection, Staining, Transfection, Plasmid Preparation, Over Expression

Journal: Nature Communications

Article Title: Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

doi: 10.1038/s41467-021-27622-9

Figure Lengend Snippet: The anti-BMP1.3 monoclonal antibody blocks the enzymatic activity of BMP1.3, exerting pleiotropic therapeutic effects after MI. On the one hand, BMP1.3 inhibition reduces collagen maturation. On the other hand, it results in TGFβ pathway inhibition, which in turns reduces myofibroblast activation, Lox expression and collagen cross-linking and increases the expression of cardioprotective BMPs.

Article Snippet: Plasma levels of BMP1.3 were measured using the ELISA Human BMP1 Matched Antibody Pair Kit (Cloud-Clone PSA653Hu01) and additional reagents supplied in Antibody Pairs Support Pack (Cloud-Clone IS077).

Techniques: Activity Assay, Inhibition, Activation Assay, Expressing