An Abundant Tissue Macrophage Population in the Adult Murine Heart with a Distinct Alternatively-Activated Macrophage Profile

Cardiac tissue macrophages (cTMs) are a previously uncharacterised cell type that we have identified and characterise here as an abundant GFP+ population within the adult Cx3cr1GFP/+ knock-in mouse heart. They comprise the predominant myeloid cell population in the myocardium, and are found throughout myocardial interstitial spaces interacting directly with capillary endothelial cells and cardiomyocytes. Flow cytometry-based immunophenotyping shows that cTMs exhibit canonical macrophage markers. Gene expression analysis shows that cTMs (CD45+CD11b+GFP+) are distinct from mononuclear CD45+CD11b+GFP+ cells sorted from the spleen and brain of adult Cx3cr1GFP/+ mice. Gene expression profiling reveals that cTMs closely resemble alternatively-activated anti-inflammatory M2 macrophages, expressing a number of M2 markers, including Mrc1, CD163, and Lyve-1. While cTMs perform normal tissue macrophage homeostatic functions, they also exhibit a distinct phenotype, involving secretion of salutary factors (including IGF-1) and immune modulation. In summary, the characterisation of cTMs at the cellular and molecular level defines a potentially important role for these cells in cardiac homeostasis

The protein kinases AtMAP3Kε1 and BnMAP3Kε1 are functional homologues of S. pombe cdc7p and may be involved in cell division

We identified an Arabidopsis thaliana gene, AtMAP3Kε1, and a Brassica napus cDNA, BnMAP3Kε1, encoding functional protein serine/threonine kinases closely related to cdc7p and Cdc15p from Schizosaccharomyces pombe and Saccharomyces cerevisiae, respectively. This is the first report of cdc7-related genes in non-fungal eukaryotes; no such genes have as yet been identified in Metazoans. The B. napus protein is able to partially complement a cdc7 loss of function mutation in S. pombe. RT–PCR and in situ hybridisation revealed that the A. thaliana and B. napus genes are expressed in both the sporophytic and the gametophytic tissues of the respective plant species and revealed further that expression is highest in dividing cells. Moreover, AtMAP3Kε1 gene expression is cell cycle-regulated, with higher expression in G2-M phases. Our results strongly suggest that the plant cdc7p-related protein kinases are involved in a signal transduction pathway similar to the SIN pathway, which positively regulates cytokinesis in S. pombe.This work was mainly supported by a EU grant (SIME project BIOTEC-RTD-CEE PL 960275). The authors also acknowledge the financial support of the MERS and CNRS to UMR 8618, and DGESIG PB98–0678

Variable outcomes of human heart attack recapitulated in genetically diverse mice.

Clinical variation in patient responses to myocardial infarction (MI) has been difficult to model in laboratory animals. To assess the genetic basis of variation in outcomes after heart attack, we characterized responses to acute MI in the Collaborative Cross (CC), a multi-parental panel of genetically diverse mouse strains. Striking differences in post-MI functional, morphological, and myocardial scar features were detected across 32 CC founder and recombinant inbred strains. Transcriptomic analyses revealed a plausible link between increased intrinsic cardiac oxidative phosphorylation levels and MI-induced heart failure. The emergence of significant quantitative trait loci for several post-MI traits indicates that utilizing CC strains is a valid approach for gene network discovery in cardiovascular disease, enabling more accurate clinical risk assessment and prediction

Variable outcomes of human heart attack recapitulated in genetically diverse mice

Clinical variation in patient responses to myocardial infarction (MI) has been difficult to model in laboratory animals. To assess the genetic basis of variation in outcomes after heart attack, we characterized responses to acute MI in the Collaborative Cross (CC), a multi-parental panel of genetically diverse mouse strains. Striking differences in post-MI functional, morphological, and myocardial scar features were detected across 32 CC founder and recombinant inbred strains. Transcriptomic analyses revealed a plausible link between increased intrinsic cardiac oxidative phosphorylation levels and MI-induced heart failure. The emergence of significant quantitative trait loci for several post-MI traits indicates that utilizing CC strains is a valid approach for gene network discovery in cardiovascular disease, enabling more accurate clinical risk assessment and prediction

Skeletal muscle NOX4 is required for adaptive responses that prevent insulin resistance

Reactive oxygen species (ROS) generated during exercise are considered integral for the health-promoting effects of exercise. However, the precise mechanisms by which exercise and ROS promote metabolic health remain unclear. Here, we demonstrate that skeletal muscle NADPH oxidase 4 (NOX4), which is induced after exercise, facilitates ROS-mediated adaptive responses that promote muscle function, maintain redox balance, and prevent the development of insulin resistance. Conversely, reductions in skeletal muscle NOX4 in aging and obesity contribute to the development of insulin resistance. NOX4 deletion in skeletal muscle compromised exercise capacity and antioxidant defense and promoted oxidative stress and insulin resistance in aging and obesity. The abrogated adaptive mechanisms, oxidative stress, and insulin resistance could be corrected by deleting the H2O2-detoxifying enzyme GPX-1 or by treating mice with an agonist of NFE2L2, the master regulator of antioxidant defense. These findings causally link NOX4-derived ROS in skeletal muscle with adaptive responses that promote muscle function and insulin sensitivity

E-peptides control bioavailability of IGF-1

Insulin-like growth factor 1 (IGF-1) is a potent cytoprotective growth factor that has attracted considerable attention as a promising therapeutic agent. Transgenic over-expression of IGF-1 propeptides facilitates protection and repair in a broad range of tissues, although transgenic mice over-expressing IGF-1 propeptides display little or no increase in IGF-1 serum levels, even with high levels of transgene expression. IGF-1 propeptides are encoded by multiple alternatively spliced transcripts including C-terminal extension (E) peptides, which are highly positively charged. In the present study, we use decellularized mouse tissue to show that the E-peptides facilitate in vitro binding of murine IGF-1 to the extracellular matrix (ECM) with varying affinities. This property is independent of IGF-1, since proteins consisting of the E-peptides fused to relaxin, a related member of the insulin superfamily, bound equally avidly to decellularized ECM. Thus, the E-peptides control IGF-1 bioavailability by preventing systemic circulation, offering a potentially powerful way to tether IGF-1 and other therapeutic proteins to the site of synthesis and/or administration

A superior extracellular matrix binding motif to enhance the regenerative activity and safety of therapeutic proteins

Abstract Among therapeutic proteins, cytokines and growth factors have great potential for regenerative medicine applications. However, these molecules have encountered limited clinical success due to low effectiveness and major safety concerns, highlighting the need to develop better approaches that increase efficacy and safety. Promising approaches leverage how the extracellular matrix (ECM) controls the activity of these molecules during tissue healing. Using a protein motif screening strategy, we discovered that amphiregulin possesses an exceptionally strong binding motif for ECM components. We used this motif to confer the pro-regenerative therapeutics platelet-derived growth factor-BB (PDGF-BB) and interleukin-1 receptor antagonist (IL-1Ra) a very high affinity to the ECM. In mouse models, the approach considerably extended tissue retention of the engineered therapeutics and reduced leakage in the circulation. Prolonged retention and minimal systemic diffusion of engineered PDGF-BB abolished the tumour growth-promoting adverse effect that was observed with wild-type PDGF-BB. Moreover, engineered PDGF-BB was substantially more effective at promoting diabetic wound healing and regeneration after volumetric muscle loss, compared to wild-type PDGF-BB. Finally, while local or systemic delivery of wild-type IL-1Ra showed minor effects, intramyocardial delivery of engineered IL-1Ra enhanced cardiac repair after myocardial infarction by limiting cardiomyocyte death and fibrosis. This engineering strategy highlights the key importance of exploiting interactions between ECM and therapeutic proteins for developing effective and safer regenerative therapies

E-peptides bind heparin-agarose.

<p>Binding of IGF-1 isoforms to heparin coated agarose beads (lanes 2–4) and control agarose beads (lanes 6–8). The control lane (9) is the growth medium from IGF-1EbCD transfected cells.</p

Length (amino acids), Isoelectric Point (IP), and calculated charge at pH7 of human (h) (rows 1–5) and murine (mu) (rows 6–10) IGF-1 related peptides.

<p>Rows 1 and 6: mature IGF-1; rows 4,5,9,10: propeptides; rows 2,3,7,8: E-peptides alone.</p

Structure of the rodent IGF-1 gene.

<p>Exons 1 and 2 are transcribed from different promoters. Differential splicing gives rise to two different signal peptides (SP1 and SP2), which include a common C-terminal sequence encoded by Exon 3. Exon 3 also encodes the N-terminal part of the mature IGF-1 B chain. Exon 4 encodes the remaining mature IGF-1 protein (B,C,A and D chains), and also encodes the common N-terminal sequence of the E-peptides. Differential splicing excluding Exon 5 gives rise to the IGF-1Ea propeptide, or a longer IGF-1Eb propeptide when Exon 5 is included. Protease cleavage (arrowheads) removes the E peptides to produce the mature IGF-1 protein.</p