Ruxolitinib: A new hope for ventilator-induced diaphragm dysfunction.

AIM: Mechanical ventilation (MV) results in diminished diaphragm size and strength, termed ventilator-induced diaphragm dysfunction (VIDD). VID increases dependence, prolongs weaning, and increases discharge mortality rates. The Janus kinase (JAK)/Signal Transducer and Activator of Transcription (STAT) pathway is implicated in VIDD, upregulated following MV. JAK/STAT inhibition alleviates chronic muscle wasting conditions. This study aimed to explore the therapeutic potential of Ruxolitinib, an FDA approved JAK1/2 inhibitor (JI) for the treatment of VIDD. METHODS: Rats were subjected to 5 days controlled MV (CMV) with and without daily Ruxolitinib gavage. Muscle fiber size and function were assessed. RNAseq, mitochondrial morphology, respirometry, and mass spectrometry were determined. RESULTS: CMV significantly reduced diaphragm size and specific force by 45% (p < 0.01), associated with a two-fold P-STAT3 upregulation (p < 0.001). CMV disrupted mitochondrial content and reduced the oxygen consumption rate (p < 0.01). Expression of the motor protein myosin was unaffected, however CMV alters myosin function via post-translational modifications (PTMs). Daily administration of JI increased animal survival (40% vs. 87%; p < 0.05), restricted P-STAT3 (p < 0.001), and preserved diaphragm size and specific force. JI was associated with preserved mitochondrial content and respiratory function (p < 0.01), and the reversal or augmentation of myosin deamidation PTMs of the rod and head region. CONCLUSION: JI preserved diaphragm function, leading to increased survival in an experimental model of VIDD. Functional enhancement was associated with maintenance of mitochondrial content and respiration and the reversal of ventilator-induced PTMs of myosin. These results demonstrate the potential of repurposing Ruxolitinib for treatment of VIDD.

JAK/STAT inhibition augments soleus muscle function in a rat model of critical illness myopathy via regulation of complement C3/3R.

KEY POINTS: Critical illness myopathy (CIM) is a frequently observed negative consequence of modern critical care. Chronic Janus kinase (JAK)/signal transducer and activator of transcription activation impairs muscle size and function and is prominent following mechanical ventilation. We identify pSTAT-3 activation in tibialis anterior of CIM patients, before examining the potential benefits of JAK1/2 inhibition in an experimental model of CIM, where muscle mass and function are impaired. CIM activates complement cascade and increased monocyte infiltration in the soleus muscle, which was ameliorated by JAK1/2 inhibition, leading to reduced muscle degeneration and improved muscle force. Here, we demonstrate that JAK1/2 inhibition augments CIM muscle function through regulation of the complement cascade. ABSTRACT: Critical illness myopathy (CIM) is frequently observed in response to modern critical care with negative consequences for patient quality of life, morbidity, mortality and healthcare costs. Janus kinase (JAK)/signal transducer and activator of transcription (STAT) activation is observed in limb muscles following controlled mechanical ventilation. Chronic JAK/STAT activation promotes loss of muscle mass and function. Thus, we hypothesized that JAK1/2 inhibition would improve muscle outcomes for CIM. Following 12 days of intensive care unit conditions, pSTAT-3 levels increased in tibialis anterior muscle of CIM patients (P = 0.0489). The potential of JAK1/2 inhibition was assessed in an experimental model of CIM, where soleus muscle size and force are impaired. JAK1/2 inhibition restores soleus force (P < 0.0001). CIM activated muscle complement cascade, which was ameliorated by JAK1/2 inhibition (P < 0.05, respectively). Soleus macrophage number corresponded with complement activity, leading to reduced muscle degeneration and augmented muscle function (P < 0.05). Thus, JAK/STAT inhibition improves soleus function by modulating the complement cascade and muscle monocyte infiltration. Collectively, we demonstrate that JAK/STAT inhibition augments muscle function in CIM.

Chaperone co-inducer BGP-15 mitigates early contractile dysfunction of the soleus muscle in a rat ICU model.

AIM: Critical illness myopathy (CIM) represents a common consequence of modern intensive care, negatively impacting patient health and significantly increasing health care costs; however, there is no treatment available apart from symptomatic and supportive interventions. The chaperone co-inducer BGP-15 has previously been shown to have a positive effect on the diaphragm in rats exposed to the intensive care unit (ICU) condition. In this study, we aim to explore the effects of BGP-15 on a limb muscle (soleus muscle) in response to the ICU condition. METHODS: Sprague-Dawley rats were subjected to the ICU condition for 5, 8 and 10 days and compared with untreated sham-operated controls. RESULTS: BGP-15 significantly improved soleus muscle fibre force after 5 days exposure to the ICU condition. This improvement was associated with the protection of myosin from post-translational myosin modifications, improved mitochondrial structure/biogenesis and reduced the expression of MuRF1 and Fbxo31 E3 ligases. At longer durations (8 and 10 days), BGP-15 had no protective effect when the hallmark of CIM had become manifest, that is, preferential loss of myosin. Unrelated to the effects on skeletal muscle, BGP-15 had a strong positive effect on survival compared with untreated animals. CONCLUSIONS: BGP-15 treatment improved soleus muscle fibre and motor protein function after 5 days exposure to the ICU condition, but not at longer durations (8 and 10 days) when the preferential loss of myosin was manifest. Thus, long-term CIM interventions targeting limb muscle fibre/myosin force generation capacity need to consider both the post-translational modifications and the loss of myosin.

Vamorolone treatment improves skeletal muscle outcome in a critical illness myopathy rat model.

AIM: Critical illness myopathy (CIM) is a consequence of modern critical care, leading to skeletal muscle atrophy/paralysis with negative consequences for mortality/morbidity and health care costs. Glucocorticoids (GCs) have been proposed to trigger CIM. Here, we compare outcomes of two GCs, the commonly used prednisolone and the newly developed dissociative vamorolone in response to the intensive care unit (ICU) condition for 5 days, ie, sedation, immobilization, and mechanical ventilation. METHODS: Rats were divided into a 0-day sham-operated control group, and three groups exposed to 5 days ICU condition during treatment with prednisolone (PRED) or vamorolone (VAM) or none of these GCs (ICU-group). Survival, body and muscle weights, cytokine concentrations, regulation of muscle contraction in single fast- and slow-twitch muscle fibres, myofibrillar protein expression and protein degradation pathways were studied. RESULTS: Critical illness myopathy geno- and pheno-types were confirmed in the ICU group. However, VAM and PRED groups showed reduced atrophy/weakness than the ICU group, and muscle specific differences with more severe negative effects on fast-twitch muscle fibres in the PRED than the other groups. CONCLUSION: These results show that vamorolone provides a GC intervention superior to typical GCs in improving CIM outcomes. Further, the findings do not support the notion that moderate-dose GC treatment represents a factor triggering CIM.

Targeting Heat Shock Proteins Mitigates Ventilator Induced Diaphragm Muscle Dysfunction in an Age-Dependent Manner.

Intensive care unit (ICU) patients are often overtly subjected to mechanical ventilation and immobilization, which leads to impaired limb and respiratory muscle function. The latter, termed ventilator-induced diaphragm dysfunction (VIDD) has recently been related to compromised heat shock protein (Hsp) activation. The administration of a pharmacological drug BGP-15 acting as a Hsp chaperone co-inducer has been found to partially alleviate VIDD in young rats. Considering that the mean age in the ICU is increasing, we aimed to explore whether the beneficial functional effects are also present in old rats. For that, we exposed young (7-8 months) and old (28-32 months) rats to 5-day controlled mechanical ventilation and immobilization with or without systemic BGP-15 administration. We then dissected diaphragm muscles, membrane-permeabilized bundles and evaluated the contractile function at single fiber level. Results confirmed that administration of BGP-15 restored the force-generating capacity of isolated muscle cells from young rats in conjunction with an increased expression of Hsp72. On the other hand, our results highlighted that old rats did not positively respond to the BGP-15 treatment. Therefore, it is of crucial importance to comprehend in more depth the effect of VIDD on diaphragm function and ascertain any further age-related differences.

The chaperone co-inducer BGP-15 alleviates ventilation-induced diaphragm dysfunction.

Ventilation-induced diaphragm dysfunction (VIDD) is a marked decline in diaphragm function in response to mechanical ventilation, which has negative consequences for individual patients' quality of life and for the health care system, but specific treatment strategies are still lacking. We used an experimental intensive care unit (ICU) model, allowing time-resolved studies of diaphragm structure and function in response to long-term mechanical ventilation and the effects of a pharmacological intervention (the chaperone co-inducer BGP-15). The marked loss of diaphragm muscle fiber function in response to mechanical ventilation was caused by posttranslational modifications (PTMs) of myosin. In a rat model, 10 days of BGP-15 treatment greatly improved diaphragm muscle fiber function (by about 100%), although it did not reverse diaphragm atrophy. The treatment also provided protection from myosin PTMs associated with HSP72 induction and PARP-1 inhibition, resulting in improvement of mitochondrial function and content. Thus, BGP-15 may offer an intervention strategy for reducing VIDD in mechanically ventilated ICU patients.

Masseter muscle myofibrillar protein synthesis and degradation in an experimental critical illness myopathy model.

Critical illness myopathy (CIM) is a debilitating common consequence of modern intensive care, characterized by severe muscle wasting, weakness and a decreased myosin/actin (M/A) ratio. Limb/trunk muscles are primarily affected by this myopathy while cranial nerve innervated muscles are spared or less affected, but the mechanisms underlying these muscle-specific differences remain unknown. In this time-resolved study, the cranial nerve innervated masseter muscle was studied in a unique experimental rat intensive care unit (ICU) model, where animals were exposed to sedation, neuromuscular blockade (NMB), mechanical ventilation, and immobilization for durations varying between 6 h and 14d. Gel electrophoresis, immunoblotting, RT-PCR and morphological staining techniques were used to analyze M/A ratios, myofiber size, synthesis and degradation of myofibrillar proteins, and levels of heat shock proteins (HSPs). Results obtained in the masseter muscle were compared with previous observations in experimental and clinical studies of limb muscles. Significant muscle-specific differences were observed, i.e., in the masseter, the decline in M/A ratio and muscle fiber size was small and delayed. Furthermore, transcriptional regulation of myosin and actin synthesis was maintained, and Akt phosphorylation was only briefly reduced. In studied degradation pathways, only mRNA, but not protein levels of MuRF1, atrogin-1 and the autophagy marker LC3b were activated by the ICU condition. The matrix metalloproteinase MMP-2 was inhibited and protective HSPs were up-regulated early. These results confirm that the cranial nerve innervated masticatory muscles is less affected by the ICU-stress response than limb muscles, in accordance with clinical observation in ICU patients with CIM, supporting the model' credibility as a valid CIM model.

Effects of corticosteroids in the development of limb muscle weakness in a porcine intensive care unit model.

Severe muscle wasting is a debilitating condition in critically ill intensive care unit (ICU) patients, characterized by general muscle weakness and dysfunction, resulting in a prolonged mobilization, delayed weaning from the ventilator, and a decreased quality of life post-ICU. The mechanisms underlying limb muscle weakness in ICU patients are complex and involve the impact of primary disease, but also factors common to critically ill ICU patients such as sepsis, mechanical ventilation (MV), immobilization, and systemic administration of corticosteroids (CS). These factors may have additive negative effects on skeletal muscle structure and function, but their respective role alone remain unknown. The primary aim of this study was to examine how CS administration potentiates ventilator and immobilization-related limb muscle dysfunction at the gene level. Comparing biceps femoris gene expression in pigs exposed to MV and CS for 5 days with only MV pigs for the same duration of time showed a distinct deregulation of 186 genes according to microarray. Surprisingly, the decreased force-generation capacity at the single muscle fiber reported in response to the addition of CS administration in mechanically ventilated and immobilized pigs was not associated with an additional upregulation of proteolytic pathways. On the other hand, an altered expression of genes regulating kinase activity, cell cycle, transcription, channel regulation, oxidative stress response, cytoskeletal, sarcomeric, and heat shock protein, as well as protein synthesis at the translational level, appears to play an additive deleterious role for the limb muscle weakness in immobilized ICU patients.