Developing a Clinically Relevant Hemorrhagic Shock Model in Rats.

Over the recent decades, the development of animal models allowed us to better understand various pathologies and identify new treatments. Hemorrhagic shock, i.e., organ failure due to rapid loss of a large volume of blood, is associated with a highly complex pathophysiology involving several pathways. Numerous existing animal models of hemorrhagic shock strive to replicate what happens in humans, but these models have limits in terms of clinical relevance, reproducibility, or standardization. The aim of this study was to refine these models to develop a new model of hemorrhagic shock. Briefly, hemorrhagic shock was induced in male Wistar Han rats (11-13 weeks old) by a controlled exsanguination responsible for a drop in the mean arterial pressure. The next phase of 75 min was to maintain a low mean arterial blood pressure, between 32 mmHg and 38 mmHg, to trigger the pathophysiological pathways of hemorrhagic shock. The final phase of the protocol mimicked patient care with an administration of intravenous fluids, Ringer Lactate solution, to elevate the blood pressure. Lactate and behavioral scores were assessed 16 h after the protocol started, while hemodynamics parameters and plasmatic markers were evaluated 24 h after injury. Twenty-four hours post-hemorrhagic shock induction, the mean arterial and diastolic blood pressure were decreased in the hemorrhagic shock group (p < 0.05). Heart rate and systolic blood pressure remained unchanged. All organ damage markers were increased with the hemorrhagic shock (p < 0.05). The lactatemia and behavioral scores were increased compared to the sham group (p < 0.05). In conclusion, we demonstrated that the protocol described here is a relevant model of hemorrhagic shock that can be used in subsequent studies, particularly to evaluate the therapeutic potential of new molecules.

Evaluation of the dystrophin carboxy-terminal domain for micro-dystrophin gene therapy in cardiac and skeletal muscles in the DMDmdx rat model.

Duchenne muscular dystrophy (DMD) is a muscle wasting disorder caused by mutations in the gene encoding dystrophin. Gene therapy using micro-dystrophin (MD) transgenes and recombinant adeno-associated virus (rAAV) vectors hold great promise. To overcome the limited packaging capacity of rAAV vectors, most MD do not include dystrophin carboxy-terminal (CT) domain. Yet, the CT domain is known to recruit α1- and ß1-syntrophins and α-dystrobrevin, a part of the dystrophin-associated protein complex (DAPC), which is a signaling and structural mediator of muscle cells. In this study, we explored the impact of inclusion of the dystrophin CT domain on ΔR4-23/ΔCT MD (MD1), in DMDmdx rats, which allows for relevant evaluations at muscular and cardiac levels. We showed by LC-MS/MS that MD1 expression is sufficient to restore the interactions at a physiological level of most DAPC partners in skeletal and cardiac muscles, and that inclusion of the CT domain increases the recruitment of some DAPC partners at supra-physiological levels. In parallel, we demonstrated that inclusion of the CT domain does not improve MD1 therapeutic efficacy on DMD muscle and cardiac pathologies. Our work highlights new evidences of the therapeutic potential of MD1 and strengthens the relevance of this candidate for gene therapy of DMD.

Human MuStem Cell Grafting into Infarcted Rat Heart Attenuates Adverse Tissue Remodeling and Preserves Cardiac Function.

Myocardial infarction is one of the leading causes of mortality and morbidity worldwide. Whereas transplantation of several cell types into the infarcted heart has produced promising preclinical results, clinical studies using analogous human cells have shown limited structural and functional benefits. In dogs and humans, we have described a type of muscle-derived stem cells termed MuStem cells that efficiently promoted repair of injured skeletal muscle. Enhanced survival rate, long-term engraftment, and participation in muscle fiber formation were reported, leading to persistent tissue remodeling and clinical benefits. With the consideration of these features that are restricted or absent in cells tested so far for myocardial infarction, we wanted to investigate the capacity of human MuStem cells to repair infarcted hearts. Their local administration in immunodeficient rats 1 week after induced infarction resulted in reduced fibrosis and increased angiogenesis 3 weeks post-transplantation. Importantly, foci of human fibers were detected in the infarct site. Treated rats also showed attenuated left-ventricle dilation and preservation of contractile function. Interestingly, no spontaneous arrhythmias were observed. Our findings support the potential of MuStem cells, which have already been proposed as therapeutic candidates for dystrophic patients, to treat myocardial infarction and position them as an attractive tool for muscle-regenerative medicine.

Gap-134, a Connexin43 activator, prevents age-related development of ventricular fibrosis in Scn5a+/- mice.

Down-regulation of Connexin43 (Cx43) has often been associated with the development of cardiac fibrosis. We showed previously that Scn5a heterozygous knockout mice (Scn5a+/-), which mimic familial progressive cardiac conduction defect, exhibit an age-dependent decrease of Cx43 expression and phosphorylation concomitantly with activation of TGF-ß pathway and fibrosis development in the myocardium between 45 and 60 weeks of age. The aim of this study was to investigate whether Gap-134 prevents Cx43 down-regulation with age and fibrosis development in Scn5a+/- mice. We observed in 60-week-old Scn5a+/- mouse heart a Cx43 expression and localization remodeling correlated with fibrosis. Chronic administration of a potent and selective gap junction modifier, Gap-134 (danegaptide), between 45 and 60 weeks, increased Cx43 expression and phosphorylation on serine 368 and prevented Cx43 delocalization. Furthermore, we found that Gap-134 prevented fibrosis despite the persistence of the conduction defects and the TGF-ß canonical pathway activation. In conclusion, the present study demonstrates that the age-dependent decrease of Cx43 expression is involved in the ventricular fibrotic process occurring in Scn5a+/- mice. Finally, our study suggests that gap junction modifier, such as Gap-134, could be an effective anti-fibrotic agent in the context of age-dependent fibrosis in progressive cardiac conduction disease.