Activated platelets present high mobility group box 1 to neutrophils, inducing autophagy and promoting the extrusion of neutrophil extracellular traps.

BACKGROUND: Increasing evidence implicates both platelets and neutrophils in the formation, stabilization, and growth of peripheral and coronary thrombi. Neutrophil extracellular traps (NETs) play a key role. The early events in the deregulated cross-talk between platelets and neutrophils are poorly characterized. OBJECTIVES: To identify at the molecular level the mechanism through which platelets induce the generation of NETs in sterile conditions. PATIENTS/METHODS: The presence of NETs was determined in 26 thrombi from patients with acute myocardial infarction by immunohistochemistry and immunofluorescence and markers of NETs assessed in the plasma. In vitro NET generation was studied in static and in physiological flow conditions. RESULTS: Coronary thrombi mainly consist of activated platelets, neutrophils, and NETs in close proximity of platelets. Activated platelets commit neutrophils to NET generation. The event abates in the presence of competitive antagonists of the high mobility group box 1 (HMGB1) protein. Hmgb1(-/-) platelets fail to elicit NETs, whereas the HMGB1 alone commits neutrophils to NET generation. Integrity of the HMGB1 receptor, Receptor for Advanced Glycation End products (RAGE), is required for NET formation, as assessed using pharmacologic and genetic tools. Exposure to HMGB1 prevents depletion of mitochondrial potential, induces autophagosome formation, and prolongs neutrophil survival. These metabolic effects are caused by the activation of autophagy. Blockade of the autophagic flux reverts platelet HMGB1-elicited NET generation. CONCLUSIONS: Activated platelets present HMGB1 to neutrophils and commit them to autophagy and NET generation. This chain of events may be responsible for some types of thromboinflammatory lesions and indicates novel paths for molecular intervention.

Autologous transplantation of muscle-derived CD133+ stem cells in Duchenne muscle patients.

Duchenne muscular dystrophy (DMD) is a lethal X-linked recessive muscle disease due to defect on the gene encoding dystrophin. The lack of a functional dystrophin in muscles results in the fragility of the muscle fiber membrane with progressive muscle weakness and premature death. There is no cure for DMD and current treatment options focus primarily on respiratory assistance, comfort care, and delaying the loss of ambulation. Recent works support the idea that stem cells can contribute to muscle repair as well as to replenishment of the satellite cell pool. Here we tested the safety of autologous transplantation of muscle-derived CD133+ cells in eight boys with Duchenne muscular dystrophy in a 7-month, double-blind phase I clinical trial. Stem cell safety was tested by measuring muscle strength and evaluating muscle structures with MRI and histological analysis. Timed cardiac and pulmonary function tests were secondary outcome measures. No local or systemic side effects were observed in all treated DMD patients. Treated patients had an increased ratio of capillary per muscle fibers with a switch from slow to fast myosin-positive myofibers.

T and B lymphocyte depletion has a marked effect on the fibrosis of dystrophic skeletal muscles in the scid/mdx mouse.

Abnormal connective tissue proliferation following muscle degeneration is a major pathological feature of Duchenne muscular dystrophy (DMD), a genetic myopathy due to lack of the sarcolemmal dystrophin protein. Since this fibrotic proliferation is likely to be a major obstacle to the efficacy of future therapies, research is needed to understand and prevent the fibrotic process in order to develop an effective treatment. Murine muscular dystrophy (mdx) is genetically homologous to DMD, and histopatological alterations are comparable to those of the muscles of patients with DMD. To investigate the development of fibrosis, we bred the mdx mouse with the scid immunodepressed mouse and analysed fibrosis histologically; we used ELISA analysis to determine TGF-beta1 expression. Significant reduction of fibrosis and TGF-beta1 expression was found in the muscles of the scid/mdx mice. However, we observed similar centrally located nuclei, necrosis, muscle degeneration and muscle force compared to the mdx animals. These data demonstrate a correlation between the absence of B and T lymphocytes and loss of fibrosis accompanied by reduction of TGF-beta1, suggesting the importance of modulation of the immune system in DMD.

Skin-derived stem cells transplanted into resorbable guides provide functional nerve regeneration after sciatic nerve resection.

The regeneration in the peripheral nervous system is often incomplete and the treatment of severe lesions with nerve tissue loss is primarily aimed at recreating nerve continuity. Guide tubes of various types, filled with Schwann cells, stem cells, or nerve growth factors are attractive as an alternative therapy to nerve grafts. In this study, we evaluated whether skin-derived stem cells (SDSCs) can improve peripheral nerve regeneration after transplantation into nerve guides. We compared peripheral nerve regeneration in adult rats with sciatic nerve gaps of 16 mm after autologous transplantation of GFP-labeled SDSCs into two different types of guides: a synthetic guide, obtained by dip coating with a L-lactide and trimethylene carbonate (PLA-TMC) copolymer and a collagen-based guide. The sciatic function index and the recovery rates of the compound muscle action potential were significantly higher in the animals that received SDSCs transplantation, in particular, into the collagen guide, compared to the control guides filled only with PBS. For these guides the morphological and immunohistochemical analysis demonstrated an increased number of myelinated axons expressing S100 and Neurofilament 70, suggesting the presence of regenerating nerve fibers along the gap. GFP positive cells were found around regenerating nerve fibers and few of them were positive for the expression of glial markers as S-100 and glial fibrillary acidic protein. RT-PCR analysis confirmed the expression of S100 and myelin basic protein in the animals treated with the collagen guide filled with SDSCs. These data support the hypothesis that SDSCs could represent a tool for future cell therapy applications in peripheral nerve regeneration.

High-resolution X-ray microtomography for three-dimensional visualization of human stem cell muscle homing.

In the perspective of clinical translation of stem cell research, it would be advantageous to develop new techniques to detect donor cells after transplantation to track their fate and thus better understand their role in regeneration of damaged and diseased tissues. In this study we use X-ray computed microtomography for three-dimensional visualization of stem cells that were labeled with magnetic nanoparticles and transplanted via intra-arterial infusion. We show that X-ray computed microtomography offers the possibility to detect with high definition and resolution human cells after transplantation, and opens new possibilities for both experimental stem cell research.