Systemically administered wound-homing peptide accelerates wound healing by modulating syndecan-4 function.

CAR (CARSKNKDC) is a wound-homing peptide that recognises angiogenic neovessels. Here we discover that systemically administered CAR peptide has inherent ability to promote wound healing: wounds close and re-epithelialise faster in CAR-treated male mice. CAR promotes keratinocyte migration in vitro. The heparan sulfate proteoglycan syndecan-4 regulates cell migration and is crucial for wound healing. We report that syndecan-4 expression is restricted to epidermis and blood vessels in mice skin wounds. Syndecan-4 regulates binding and internalisation of CAR peptide and CAR-mediated cytoskeletal remodelling. CAR induces syndecan-4-dependent activation of the small GTPase ARF6, via the guanine nucleotide exchange factor cytohesin-2, and promotes syndecan-4-, ARF6- and Cytohesin-2-mediated keratinocyte migration. Finally, we show that genetic ablation of syndecan-4 in male mice eliminates CAR-induced wound re-epithelialisation following systemic administration. We propose that CAR peptide activates syndecan-4 functions to selectively promote re-epithelialisation. Thus, CAR peptide provides a therapeutic approach to enhance wound healing in mice; systemic, yet target organ- and cell-specific.

3D matrix adhesion feedback controls nuclear force coupling to drive invasive cell migration.

Cell invasion is a multi-step process, initiated by the acquisition of a migratory phenotype and the ability to move through complex 3D extracellular environments. We determine the composition of cell-matrix adhesion complexes of invasive breast cancer cells in 3D matrices and identify an interaction complex required for invasive migration. ßPix and myosin18A (Myo18A) drive polarized recruitment of non-muscle myosin 2A (NM2A) to adhesion complexes at the tips of protrusions. Actomyosin force engagement then displaces the Git1-ßPix complex from paxillin, establishing a feedback loop for adhesion maturation. We observe active force transmission to the nucleus during invasive migration that is needed to pull the nucleus forward. The recruitment of NM2A to adhesions creates a non-muscle myosin isoform gradient, which extends from the protrusion to the nucleus. We postulate that this gradient facilitates coupling of cell-matrix interactions at the protrusive cell front with nuclear movement, enabling effective invasive migration and front-rear cell polarity.

Hydrogen Sulfide Is a Novel Protector of the Retinal Glycocalyx and Endothelial Permeability Barrier.

Significantly reduced levels of the anti-inflammatory gaseous transmitter hydrogen sulfide (H2S) are observed in diabetic patients and correlate with microvascular dysfunction. H2S may protect the microvasculature by preventing loss of the endothelial glycocalyx. We tested the hypothesis that H2S could prevent or treat retinal microvascular endothelial dysfunction in diabetes. Bovine retinal endothelial cells (BRECs) were exposed to normal (NG, 5.5 mmol/L) or high glucose (HG, 25 mmol/L) ± the slow-release H2S donor NaGYY4137 in vitro. Glycocalyx coverage (stained with WGA-FITC) and calcein-labeled monocyte adherence were measured. In vivo, fundus fluorescein angiography (FFA) was performed in normal and streptozotocin-induced (STZ) diabetic rats. Animals received intraocular injection of NaGYY4137 (1 µM) or the mitochondrial-targeted H2S donor AP39 (100 nM) simultaneously with STZ (prevention) or on day 6 after STZ (treatment), and the ratio of interstitial to vascular fluorescence was used to estimate apparent permeability. NaGYY4137 prevented HG-induced loss of BREC glycocalyx, increased monocyte binding to BRECs (p ≤ 0.001), and increased overall glycocalyx coverage (p ≤ 0.001). In rats, the STZ-induced increase in apparent retinal vascular permeability (p ≤ 0.01) was significantly prevented by pre-treatment with NaGYY4137 and AP39 (p < 0.05) and stabilized by their post-STZ administration. NaGYY4137 also reduced the number of acellular capillaries (collagen IV + /IB4-) in the diabetic retina in both groups (p ≤ 0.05). We conclude that NaGYY4137 and AP39 protected the retinal glycocalyx and endothelial permeability barrier from diabetes-associated loss of integrity and reduced the progression of diabetic retinopathy (DR). Hydrogen sulfide donors that target the glycocalyx may therefore be a therapeutic candidate for DR.

PPFIA1 drives active α5ß1 integrin recycling and controls fibronectin fibrillogenesis and vascular morphogenesis.

Basolateral polymerization of cellular fibronectin (FN) into a meshwork drives endothelial cell (EC) polarity and vascular remodelling. However, mechanisms coordinating α5ß1 integrin-mediated extracellular FN endocytosis and exocytosis of newly synthesized FN remain elusive. Here we show that, on Rab21-elicited internalization, FN-bound/active α5ß1 is recycled to the EC surface. We identify a pathway, comprising the regulators of post-Golgi carrier formation PI4KB and AP-1A, the small GTPase Rab11B, the surface tyrosine phosphatase receptor PTPRF and its adaptor PPFIA1, which we propose acts as a funnel combining FN secretion and recycling of active α5ß1 integrin from the trans-Golgi network (TGN) to the EC surface, thus allowing FN fibrillogenesis. In this framework, PPFIA1 interacts with active α5ß1 integrin and localizes close to EC adhesions where post-Golgi carriers are targeted. We show that PPFIA1 is required for FN polymerization-dependent vascular morphogenesis, both in vitro and in the developing zebrafish embryo.

Fibronectin remodelling: cell-mediated regulation of the microenvironment.

The biophysical, mechanical and chemical characteristics of extracellular matrixes influence many cellular functions to control tissue homoeostasis and drive progression of cancer and inflammatory diseases. To maintain normal tissue function, fibronectin-rich matrixes are subject to dynamic cell-mediated structural and chemical modification. In this article, we discuss how localized application of mechanical force, heterodimer-specific integrin engagement and matrix proteolysis regulate fibronectin assembly and turnover. We also speculate that recently identified integrin trafficking, syndecan signalling and adhesion receptor-growth factor receptor cross-talk mechanisms might dynamically control the function, assembly and mechanical properties of a viable, and mechanoresponsive, fibronectin network.

Hydrogen sulfide in cell signaling, signal transduction, cellular bioenergetics and physiology in C. elegans.

Hydrogen sulfide (H2S), long viewed as a toxic gas and environmental hazard, is emerging as a biological mediator with remarkable physiological and pathophysiological relevance. H2S is now viewed as the third main gasotransmitter in the mammalian body. Its pharmacological characteristic possesses similarities to the other two gasotransmitters - nitric oxide (NO) and carbon monoxide (CO). Many of the biological effects of H2S follow a bell-shaped concentration-response; at low concentration or at lower release rates it has beneficial and cytoprotective effects, while at higher concentrations or fast release rates toxicity becomes apparent. Cellular bioenergetics is a prime example for this bell-shaped dose-response, where H2S, at lower concentrations/rates serves as an inorganic substrate and electron donor for mitochondrial ATP generation, while at high concentration it inhibits mitochondrial respiration by blocking the Complex IV in the mitochondrial electron transport chain. The current review is aimed to focus on the following aspects of H2S biology: 1) a general overview of the general pharmacological characteristics of H2S, 2) a summary of the key H2S-mediated signal transduction pathways, 3) an overview of role of H2S in regulation of cellular bioenergetics, 4) key aspects of H2S physiology in C. elegans (a model system) and, finally 5) the therapeutic potential of H2S donating molecules in various disease states.

Epithelial ovarian cancer-induced angiogenic phenotype of human omental microvascular endothelial cells may occur independently of VEGF signaling.

Epithelial ovarian cancer (EOC) metastasizes transcoelomically to the peritoneum and omentum, and despite surgery and chemotherapy, recurrent disease is likely. Metastasis requires the induction of proangiogenic changes in the omental microenvironment and EOC-induced omental angiogenesis is currently a key therapeutic target. In particular, antiangiogenic therapies targeting the vascular endothelial growth factor A (VEGFA) pathway are commonly used, although, with limited effects. Here, using human omental microvascular endothelial cells (HOMECs) and ovarian cancer cell lines as an in vitro model, we show that factors secreted from EOC cells increased proliferation, migration, and tube-like structure formation in HOMECs. However, EOC-induced angiogenic tube-like formation and migration were unaffected by inhibition of tyrosine kinase activity of VEGF receptors 1 and 2 (Semaxanib; SU5416) or neutralization of VEGFA (neutralizing anti-VEGFA antibody), although VEGFA165-induced HOMEC migration and tube-like structure formation were abolished. Proteomic investigation of the EOC secretome identified several alternative angiogenesis-related proteins. We screened these for their ability to induce an angiogenic phenotype in HOMECs, i.e., proliferation, migration, and tube-like structure formation. Hepatocyte growth factor (HGF) and insulin-like growth factor binding protein 7 (IGFBP-7) increased all three parameters, and cathepsin L (CL) increased migration and tubule formation. Further investigation confirmed expression of the HGF receptor c-Met in HOMECs. HGF- and EOC-induced proliferation and angiogenic tube structure formation were blocked by the c-Met inhibitor PF04217903. Our results highlight key alternative angiogenic mediators for metastatic EOC, namely, HGF, CL, and IGFBP-7, suggesting that effective antiangiogenic therapeutic strategies for this disease require inhibition of multiple angiogenic pathways.

Retinopathy in diabetes.

With the incidence, and prevalence of diabetes mellitus increasing worldwide, diabetic retinopathy is expected to reach epidemic proportions. The aim of this chapter is to introduce diabetic retinopathy, a leading cause of blindness in people of the working age. The clinical course of retinopathy, anatomical changes, its pathogenesis and current treatment are described, followed by an overview of the emerging drug therapies for the potential treatment of this sight-threatening complication of diabetes.

Synthesis and antiproliferative properties of ibuprofen-oligo(3-hydroxybutyrate) conjugates.

Synthesis of novel conjugates of the non-steroidal anti-inflammatory drug - ibuprofen with nontoxic oligo(3-hydroxybutyrate) (OHB) is described. Presented results indicate that anionic ring-opening polymerization of (R,S)-beta-butyrolactone initiated with an alkali metal salt of (S)-(+)-2-(4-isobutylphenyl)propionic acid (ibuprofen) may constitute a convenient method of conjugation of selected drugs with biodegradable OHB. Furthermore using the MTT cell proliferation assay we demonstrated that ibuprofen conjugated with OHB exhibited significantly increased, as compared to free ibuprofen, potential to inhibit proliferation of HT-29 and HCT 116 colon cancer cells. However, the conjugates of ibuprofen and OHB are less toxic as was shown in oral acute toxicity test in rats. Although the mechanism of antiproliferative activity of ibuprofen-OHB conjugates (Ibu-OHB) has to be established, we suggest that partially it can be related to more effective cellular uptake of the conjugate than the free drug. This assumption is based on the observation of much more efficient accumulation of a marker compound - OHB conjugated with fluorescein, in contrast to fluorescein sodium salt, which entered cells inefficiently. Further characterization of biological properties of the ibuprofen-OHB conjugates would provide insight into the mechanism of their antiproliferative effect and assess the potential relevance of their anticancer activity.