Absence of SPARC results in increased cardiac rupture and dysfunction after acute myocardial infarction

The matricellular protein SPARC (secreted protein, acidic and rich in cysteine, also known as osteonectin) mediates cell–matrix interactions during wound healing and regulates the production and/or assembly of the extracellular matrix (ECM). This study investigated whether SPARC functions in infarct healing and ECM maturation after myocardial infarction (MI). In comparison with wild-type (WT) mice, animals with a targeted inactivation of SPARC exhibited a fourfold increase in mortality that resulted from an increased incidence of cardiac rupture and failure after MI. SPARC-null infarcts had a disorganized granulation tissue and immature collagenous ECM. In contrast, adenoviral overexpression of SPARC in WT mice improved the collagen maturation and prevented cardiac dilatation and dysfunction after MI. In cardiac fibroblasts in vitro, reduction of SPARC by short hairpin RNA attenuated transforming growth factor β (TGF)–mediated increase of Smad2 phosphorylation, whereas addition of recombinant SPARC increased Smad2 phosphorylation concordant with increased Smad2 phosphorylation in SPARC-treated mice. Importantly, infusion of TGF-β rescued cardiac rupture in SPARC-null mice but did not significantly alter infarct healing in WT mice. These findings indicate that local production of SPARC is essential for maintenance of the integrity of cardiac ECM after MI. The protective effects of SPARC emphasize the potential therapeutic applications of this protein to prevent cardiac dilatation and dysfunction after MI

Cartilage intermediate layer protein 1 (CILP1): a novel mediator of cardiac extracellular matrix remodelling

Heart failure is accompanied by extracellular matrix (ECM) remodelling, often leading to cardiac fibrosis. In the present study we explored the significance of cartilage intermediate layer protein 1 (CILP1) as a novel mediator of cardiac ECM remodelling. Whole genome transcriptional analysis of human cardiac tissue samples revealed a strong association of CILP1 with many structural (e.g. COL1A2 r2¿=¿0.83) and non-structural (e.g. TGFB3 r2¿=¿0.75) ECM proteins. Gene enrichment analysis further underscored the involvement of CILP1 in human cardiac ECM remodelling and TGFß signalling. Myocardial CILP1 protein levels were significantly elevated in human infarct tissue and in aortic valve stenosis patients. CILP1 mRNA levels markedly increased in mouse heart after myocardial infarction, transverse aortic constriction, and angiotensin II treatment. Cardiac fibroblasts were found to be the primary source of cardiac CILP1 expression. Recombinant CILP1 inhibited TGFß-induced ¿SMA gene and protein expression in cardiac fibroblasts. In addition, CILP1 overexpression in HEK293 cells strongly (5-fold p¿<¿0.05) inhibited TGFß signalling activity. In conclusion, our study identifies CILP1 as a new cardiac matricellular protein interfering with pro-fibrotic TGFß signalling, and as a novel sensitive marker for cardiac fibrosis

Hendrik Conscience.

De auteur heeft het over Conscience, over de vele, bij wijlen intensieve contacten die hij onderhield met Gent en over zijn vriendschapsbanden met tal van Gentse figuren.</jats:p

Recombinant staphylokinase variants with reduced antigenicity due to elimination of B-lymphocyte epitopes

Abstract Site directed mutagenesis (350 variants) of recombinant staphylokinase (SakSTAR), a potent fibrin-selective thrombolytic agent, was undertaken in order to reduce its antigenicity while maintaining its potency. Variants with K35A, (ie, Lys[K] in position 35 substituted with Ala[A]), E65D or E65Q, K74R or K74Q, E80A+D82A, K130T, and K135R displayed increased enzymatic activity or reduced binding of human staphylokinase-specific antibodies. Additive mutagenesis identified 8 variants with intact thrombolytic potencies, which absorbed down to less than a third of SakSTAR-specific antibodies. Intra-arterial administration in 61 patients with peripheral arterial occlusion caused no significant allergic reactions. Median neutralizing antibody titers (with 15 to 85 percentiles), expressed as microgram (μg) compound neutralized per milliliter plasma, were 4.4 (0.3 to 49) for the variants, compared with 12 (4 to 100) in 70 patients given wild-type SakSTAR (P = .002 by Mann-Whitney rank sum test). Overt neutralizing antibody induction (more than 5 μg compound neutralized per milliliter plasma) was observed in 57 of 70 patients (81%) given wild-type SakSTAR, but only in 28 of 60 patients (47%) treated with variants (P &amp;lt; .0001 by Fisher exact test). On the basis of this study, the variant SakSTAR (K35A, E65Q, K74R, D82A, S84A, T90A, E99D, T101S, E108A, K109A, K130T, K135R) (code SY155) has been selected for further clinical development.</jats:p

Recombinant staphylokinase variants with reduced antigenicity due to elimination of B-lymphocyte epitopes

Site directed mutagenesis (350 variants) of recombinant staphylokinase (SakSTAR), a potent fibrin-selective thrombolytic agent, was undertaken in order to reduce its antigenicity while maintaining its potency. Variants with K35A, (ie, Lys[K] in position 35 substituted with Ala[A]), E65D or E65Q, K74R or K74Q, E80A+D82A, K130T, and K135R displayed increased enzymatic activity or reduced binding of human staphylokinase-specific antibodies. Additive mutagenesis identified 8 variants with intact thrombolytic potencies, which absorbed down to less than a third of SakSTAR-specific antibodies. Intra-arterial administration in 61 patients with peripheral arterial occlusion caused no significant allergic reactions. Median neutralizing antibody titers (with 15 to 85 percentiles), expressed as microgram (μg) compound neutralized per milliliter plasma, were 4.4 (0.3 to 49) for the variants, compared with 12 (4 to 100) in 70 patients given wild-type SakSTAR (P = .002 by Mann-Whitney rank sum test). Overt neutralizing antibody induction (more than 5 μg compound neutralized per milliliter plasma) was observed in 57 of 70 patients (81%) given wild-type SakSTAR, but only in 28 of 60 patients (47%) treated with variants (P &lt; .0001 by Fisher exact test). On the basis of this study, the variant SakSTAR (K35A, E65Q, K74R, D82A, S84A, T90A, E99D, T101S, E108A, K109A, K130T, K135R) (code SY155) has been selected for further clinical development.</jats:p

Zondaars en sterren; De mooie zomer van 40 /

Met bijlage

Cartilage intermediate layer protein 1 (CILP1):A novel mediator of cardiac extracellular matrix remodelling

Abstract Heart failure is accompanied by extracellular matrix (ECM) remodelling, often leading to cardiac fibrosis. In the present study we explored the significance of cartilage intermediate layer protein 1 (CILP1) as a novel mediator of cardiac ECM remodelling. Whole genome transcriptional analysis of human cardiac tissue samples revealed a strong association of CILP1 with many structural (e.g. COL1A2 r2 = 0.83) and non-structural (e.g. TGFB3 r2 = 0.75) ECM proteins. Gene enrichment analysis further underscored the involvement of CILP1 in human cardiac ECM remodelling and TGFβ signalling. Myocardial CILP1 protein levels were significantly elevated in human infarct tissue and in aortic valve stenosis patients. CILP1 mRNA levels markedly increased in mouse heart after myocardial infarction, transverse aortic constriction, and angiotensin II treatment. Cardiac fibroblasts were found to be the primary source of cardiac CILP1 expression. Recombinant CILP1 inhibited TGFβ-induced αSMA gene and protein expression in cardiac fibroblasts. In addition, CILP1 overexpression in HEK293 cells strongly (5-fold p < 0.05) inhibited TGFβ signalling activity. In conclusion, our study identifies CILP1 as a new cardiac matricellular protein interfering with pro-fibrotic TGFβ signalling, and as a novel sensitive marker for cardiac fibrosis