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Astrocytes play a crucial role in the onset and progression of amyotrophic lateral sclerosis (ALS), a fatal disorder marked by the degeneration of motor neurons (MNs) in the central nervous system. Although astrocytes in ALS are known to be toxic to MNs, the pathological changes leading to their neurotoxic phenotype remain poorly understood. In this study, we generated human astrocytes from induced pluripotent stem cells (iPSCs) carrying the ALS-associated A4V mutation in superoxide dismutase 1 (SOD1) to examine early cellular pathways and network changes. Proteomic analysis revealed that ALS astrocytes are both dysfunctional and reactive compared to control astrocytes. We identified significant alterations in the levels of proteins linked to ALS pathology and the innate immune cGAS-STING pathway. Furthermore, we found that ALS astrocyte reactivity differs from that of control astrocytes treated with tumor necrosis factor alpha (TNFα), a key cytokine in inflammatory reactions. We then evaluated the potential of fibroblast growth factor (FGF) 2, 4, 16, and 18 to reverse ALS astrocyte phenotype. Among these, FGF4 successfully reversed ALS astrocyte dysfunction and reactivity in vitro. When delivered to the spinal cord of the SOD1 G93A mouse model of ALS, FGF4 lowered astrocyte reactivity. However, this was not sufficient to protect MNs from cell death. Further analysis indicated that TNFα abrogated the reactivity reduction achieved by FGF4, suggesting that complete rescue of the ALS phenotype by FGF4 is hindered by ongoing complex neuroinflammatory processes in vivo. In summary, our data demonstrate that astrocytes generated from ALS iPSCs are inherently dysfunctional and exhibit an immune reactive phenotype. Effectively targeting astrocyte dysfunction and reactivity in vivo may help mitigate ALS and prevent MN death., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

(1) Background: the use of Mesenchymal Stromal Cells (MSC) in emerging therapies for spinal cord injury (SCI) hold the potential to improve functional recovery. However, the development of cell-based medicines is challenging and preclinical studies addressing quality, safety and efficacy must be conducted prior to clinical testing; (2) Methods: herein we present (i) the characterization of the quality attributes of MSC from the Wharton's jelly (WJ) of the umbilical cord, (ii) safety of intrathecal infusion in a 3-month subchronic toxicity assessment study, and (iii) efficacy in a rat SCI model by controlled impaction (100 kdynes) after single (day 7 post-injury) and repeated dose of 1 × 10 6 MSC,WJ (days 7 and 14 post-injury) with 70-day monitoring by electrophysiological testing, motor function assessment and histology evaluation; (3) Results: no toxicity associated to MSC,WJ infusion was observed. Regarding efficacy, recovery of locomotion was promoted at early time points. Persistence of MSC,WJ was detected early after administration (day 2 post-injection) but not at days 14 and 63 post-injection. (4) Conclusions: the safety profile and signs of efficacy substantiate the suitability of the presented data for inclusion in the Investigational Medicinal Product Dossier for further consideration by the competent Regulatory Authority to proceed with clinical trials.

Postmitotic cells, like neurons, must live through a lifetime. For this reason, organisms/cells have evolved with self-repair mechanisms that allow them to have a long life. The discovery workflow of neuroprotectors during the last years has focused on blocking the pathophysiological mechanisms that lead to neuronal loss in neurodegeneration. Unfortunately, only a few strategies from these studies were able to slow down or prevent neurodegeneration. There is compelling evidence demonstrating that endorsing the self-healing mechanisms that organisms/cells endogenously have, commonly referred to as cellular resilience, can arm neurons and promote their self-healing. Although enhancing these mechanisms has not yet received sufficient attention, these pathways open up new therapeutic avenues to prevent neuronal death and ameliorate neurodegeneration. Here, we highlight the main endogenous mechanisms of protection and describe their role in promoting neuron survival during neurodegeneration., Competing Interests: The authors declare no conflict of interest.

Musculoskeletal injuries represent a challenging medical problem. Although the skeletal muscle is able to regenerate and recover after injury, the process engaged with conservative therapy can be inefficient, leading to a high re-injury rate. In addition, the formation of scar tissue implies an alteration of mechanical properties in muscle. There is still a need for new treatments of the injured muscle. NeuroHeal may be one option. Published studies demonstrated that it reduces muscle atrophy due to denervation and disuse. The main objective of the present work was to assess the potential of NeuroHeal to improve muscle regeneration after traumatic injury. Secondary objectives included characterizing the effect of NeuroHeal treatment on satellite cell biology. We used a rat model of sport-induced injury in the gastrocnemius and analyzed the effects of NeuroHeal on functional recovery by means of electrophysiology and tetanic force analysis. These studies were accompanied by immunohistochemistry of the injured muscle to analyze fibrosis, satellite cell state, and fiber type. In addition, we used an in vitro model to determine the effect of NeuroHeal on myoblast biology and partially decipher its mechanism of action. The results showed that NeuroHeal treatment advanced muscle fiber recovery after injury in a preclinical model of muscle injury, and significantly reduced the formation of scar tissue. In vitro, we observed that NeuroHeal accelerated the formation of myotubes. The results pave the way for novel therapeutic avenues for muscle/tendinous disorders.

Cells and organisms are intrinsically prepared to effectively deal with damage caused by insults and heal themselves by triggering a plethora of stress responses including macroautophagy/autophagy. However, autophagy may become malfunctional during aging, neurodegeneration, and neurotrauma. We aimed to overcome autophagy dysfunction by refining therapeutics using multi-target approaches. Thus, we have demonstrated that modulation of autophagy with the multitarget drug NeuroHeal is neuroprotective in several neurodegeneration models in which previous autophagy modulators have failed. The key element of success is the coordinated activation of opposing forces that modulate autophagy with NeuroHeal, probably leading to the autophagy-dependent degradation of death executors such as PARP1. The precise tuning of autophagy thus allows the neuron to adapt to insults, survive and repair itself. These findings support the advent a new era of neuroprotectants that counteract neuronal damage by targeting in unison different pathways of the self-repair process, including autophagy.

Muscle wasting is an unmet medical need which leads to a reduction of myofiber diameter and a negative impact on the functional performance of daily activities. We previously found that a new neuroprotective drug called NeuroHeal reduced muscle atrophy produced by transient denervation. Aiming to decipher whether NeuroHeal has a direct role in muscle biology, we used herein different models of muscle atrophy: one caused by chronic denervation, another caused by hindlimb immobilization, and lastly, an in vitro model of myotube atrophy with Tumor Necrosis Factor-α (TNFα). In all these models, we observed that NeuroHeal reduced muscle atrophy and that SIRT1 activation seems to be required for that. The treatment downregulated some critical markers of protein degradation: Muscle Ring Finger 1 (MuRF1), K48 poly-Ub chains, and p62/SQSTM1. Moreover, it seems to restore the autophagy flux associated with denervation. Hence, we envisage a prospective use of NeuroHeal at clinics for different myopathies., Competing Interests: The authors declare no conflict of interest. NeuroHeal use for muscle disorders is under patent review.

Rationale : Protective mechanisms allow healthy neurons to cope with diverse stresses. Excessive damage as well as aging can lead to defective functioning of these mechanisms. We recently designed NeuroHeal using artificial intelligence with the goal of bolstering endogenous neuroprotective mechanisms. Understanding the key nodes involved in neuroprotection will allow us to identify even more effective strategies for treatment of neurodegenerative diseases. Methods : We used a model of peripheral nerve axotomy in rat pups, that induces retrograde apoptotic death of motoneurons. Nourishing mothers received treatment with vehicle, NeuroHeal or NeuroHeal plus nicotinamide, an inhibitor of sirtuins, and analysis of the pups were performed by immunohistochemistry, electron microscopy, and immunoblotting. In vitro , the post-translational status of proteins of interest was detailed using organotypic spinal cord cultures and genetic modifications in cell lines to unravel the neuroprotective mechanisms involved. Results : We found that the concomitant activation of the NAD + -dependent deacetylase SIRT1 and the PI3K/AKT signaling pathway converge to increase the presence of deacetylated and phosphorylated FOXO3a, a transcription factor, in the nucleus. This favors the activation of autophagy, a pro-survival process, and prevents pro-apoptotic PARP1/2 cleavage. Major conclusion : NeuroHeal is a neuroprotective agent for neonatal motoneurons that fine-tunes autophagy on by converging SIRT1/AKT/FOXO3a axis. NeuroHeal is a combo of repurposed drugs that allow its readiness for prospective pediatric use., Competing Interests: Competing Interests: The authors have declared that no competing interest exists. NeuroHeal is currently under patent protection., (© The author(s).)

Injured neurons should engage endogenous mechanisms of self-protection to limit neurodegeneration. Enhancing efficacy of these mechanisms or correcting dysfunctional pathways may be a successful strategy for inducing neuroprotection. Spinal motoneurons retrogradely degenerate after proximal axotomy due to mechanical detachment (avulsion) of the nerve roots, and this limits recovery of nervous system function in patients after this type of trauma. In a previously reported proteomic analysis, we demonstrated that autophagy is a key endogenous mechanism that may allow motoneuron survival and regeneration after distal axotomy and suture of the nerve. Herein, we show that autophagy flux is dysfunctional or blocked in degenerated motoneurons after root avulsion. We also found that there were abnormalities in anterograde/retrograde motor proteins, key secretory pathway factors, and lysosome function. Further, LAMP1 protein was missorted and underglycosylated as well as the proton pump v-ATPase. In vitro modeling revealed how sequential disruptions in these systems likely lead to neurodegeneration. In vivo, we observed that cytoskeletal alterations, induced by a single injection of nocodazole, were sufficient to promote neurodegeneration of avulsed motoneurons. Besides, only pre-treatment with rapamycin, but not post-treatment, neuroprotected after nerve root avulsion. In agreement, overexpressing ATG5 in injured motoneurons led to neuroprotection and attenuation of cytoskeletal and trafficking-related abnormalities. These discoveries serve as proof of concept for autophagy-target therapy to halting the progression of neurodegenerative processes.