Environmental factors and their impact on chronic pain development and maintenance.

It is more than recognized and accepted that the environment affects the physiological responses of all living things, from bacteria to superior vertebrates, constituting an important factor in the evolution of all species. Environmental influences range from natural processes such as sunlight, seasons of the year, and rest to complex processes like stress and other mood disorders, infections, and air pollution, being all of them influenced by how each creature deals with them. In this chapter, it will be discussed how some of the environmental elements affect directly or indirectly neuropathic pain, a type of chronic pain caused by a lesion or disease of the somatosensory nervous system. For that, it was considered the edge of knowledge in translational research, thus including data from human and experimental animals as well as the applicability of such findings.

Pulmonary Low Malignant Potential Adenocarcinoma: A Validation of the Proposed Criteria for This Novel Subtype.

Adenocarcinoma (ADC) is the most common histologic type of lung cancer, including in situ (lepidic), minimally invasive, and invasive forms. While the former 2 types are associated with a favorable outcome, the latter includes tumors with variable behavior, often tumor stage-related. A recent study proposed strict morphologic criteria defining a new subgroup of resected stage I invasive ADC (16% of cases) with favorable outcomes (100% disease-specific survival), named "ADC of low malignant potential (LMP-ADC)." The following criteria were met: ≤3 cm size, nonmucinous histotype, ≥15% lepidic growth, and the absence of the following: high-grade patterns, >1 mitosis/2 mm 2 , necrosis, and vascular/pleural invasion. The aim of the present study was to validate the performance of such criteria to identify LMP-ADC in a series of 274 stage IA resected lung ADCs from a single institution. Thirty-four tumors (12.4%) met the proposed criteria for LMP-ADC, as confirmed by additional stains for mitotic figures, Ki67 index, and elastic fibers (helpful to assess alveolar wall invasion). Minor differences between the lepidic and invasive components were observed regarding cell atypia and proliferation. p53 was normally expressed by invasive tumor cells. Mutations occurred in known lung cancer genes (mostly KRAS and EGFR). Five patients (14.7%) developed disease progression and 2 of them (5.9%) died of the disease. In our series, the disease-specific survival was 94.1%. In conclusion, in resected invasive lung ADC, a subgroup presenting low-grade morphologic features and associated with favorable prognosis does exist. Morphologic criteria for LMP-ADC supported by ancillary techniques represent a valid tool to better define this novel subgroup and to refine the stratification of invasive lung ADC, possibly suggesting modified follow-up protocols, based on the observed indolent behavior in most cases.

4-Hydroxynonenal impairs miRNA maturation in heart failure via Dicer post-translational modification.

BACKGROUND AND AIMS: Developing novel therapies to battle the global public health burden of heart failure remains challenging. This study investigates the underlying mechanisms and potential treatment for 4-hydroxynonenal (4-HNE) deleterious effects in heart failure. METHODS: Biochemical, functional, and histochemical measurements were applied to identify 4-HNE adducts in rat and human failing hearts. In vitro studies were performed to validate 4-HNE targets. RESULTS: 4-HNE, a reactive aldehyde by-product of mitochondrial dysfunction in heart failure, covalently inhibits Dicer, an RNase III endonuclease essential for microRNA (miRNA) biogenesis. 4-HNE inhibition of Dicer impairs miRNA processing. Mechanistically, 4-HNE binds to recombinant human Dicer through an intermolecular interaction that disrupts both activity and stability of Dicer in a concentration- and time-dependent manner. Dithiothreitol neutralization of 4-HNE or replacing 4-HNE-targeted residues in Dicer prevents 4-HNE inhibition of Dicer in vitro. Interestingly, end-stage human failing hearts from three different heart failure aetiologies display defective 4-HNE clearance, decreased Dicer activity, and miRNA biogenesis impairment. Notably, boosting 4-HNE clearance through pharmacological re-activation of mitochondrial aldehyde dehydrogenase 2 (ALDH2) using Alda-1 or its improved orally bioavailable derivative AD-9308 restores Dicer activity. ALDH2 is a major enzyme responsible for 4-HNE removal. Importantly, this response is accompanied by improved miRNA maturation and cardiac function/remodelling in a pre-clinical model of heart failure. CONCLUSIONS: 4-HNE inhibition of Dicer directly impairs miRNA biogenesis in heart failure. Strikingly, decreasing cardiac 4-HNE levels through pharmacological ALDH2 activation is sufficient to re-establish Dicer activity and miRNA biogenesis; thereby representing potential treatment for patients with heart failure.

Wnt signaling is involved in crotalphine-induced analgesia in a rat model of neuropathic pain.

The aberrant activation of Wnt/ß-catenin and atypical Wnt/Ryk signaling pathways in the spinal cord is critical for the development and maintenance of neuropathic pain. Crotalphine is a structural analog to a peptide first identified in Crotalus durissus terrificus snake venom, which induces antinociception by activating kappa-opioid and CB2 cannabinoid receptors. Consistent with previous data, we showed that the protein levels of the canonical Wnt/ß-catenin and the atypical Wnt/Ryk signaling pathways are increased in neuropathic rats. Importantly, the administration of crotalphine downregulates these protein levels, including its downstream cascades, such as TCF4 from the canonical pathway and NR2B glutamatergic receptor and Ca2+-dependent signals, via the Ryk receptor. The CB2 receptor antagonist, AM630, abolished the crotalphine-induced atypical Wnt/Ryk signaling pathway activation. However, the selective CB2 agonist affects both canonical and non-canonical Wnt signaling in the spinal cord. Next, we showed that crotalphine blocked hypersensitivity and significantly decreased the concentration of IL-1ɑ, IL-1ß, IL-6, IL-10, IL-18, TNF-ɑ, MIP-1ɑ and MIP-2 induced by intrathecal injection of exogenous Wnt-3a agonist. Taken together, our findings show that crotalphine induces analgesia in a neuropathic pain model by down-regulating the canonical Wnt/ß-catenin and the atypical Wnt/Ryk signaling pathways and, consequently controlling neuroinflammation. This effect is, at least in part, mediated by CB2 receptor activation. These results open a perspective for new approaches that can be used to target Wnt signaling in the context of chronic pain. PERSPECTIVE: Our work identified that crotalphine-induced activation of CB2 receptors plays a critical role in the impairment of Wnt signaling during neuropathic pain. This work suggests that drugs with opioid/cannabinoid activity may be a useful strategy to target Wnt signaling in the context of chronic pain.

Exploring circular MET RNA as a potential biomarker in tumors exhibiting high MET activity.

BACKGROUND: MET-driven acquired resistance is emerging with unanticipated frequency in patients relapsing upon molecular therapy treatments. However, the determination of MET amplification remains challenging using both standard and next-generation sequencing-based methodologies. Liquid biopsy is an effective, non-invasive approach to define cancer genomic profiles, track tumor evolution over time, monitor treatment response and detect molecular resistance in advance. Circular RNAs (circRNAs), a family of RNA molecules that originate from a process of back-splicing, are attracting growing interest as potential novel biomarkers for their stability in body fluids. METHODS: We identified a circRNA encoded by the MET gene (circMET) and exploited blood-derived cell-free RNA (cfRNA) and matched tumor tissues to identify, stratify and monitor advanced cancer patients molecularly characterized by high MET activity, generally associated with genomic amplification. RESULTS: Using publicly available bioinformatic tools, we discovered that the MET locus transcribes several circRNA molecules, but only one candidate, circMET, was particularly abundant. Deeper molecular analysis revealed that circMET levels positively correlated with MET expression and activity, especially in MET-amplified cells. We developed a circMET-detection strategy and, in parallel, we performed standard FISH and IHC analyses in the same specimens to assess whether circMET quantification could identify patients displaying high MET activity. Longitudinal monitoring of circMET levels in the plasma of selected patients revealed the early emergence of MET amplification as a mechanism of acquired resistance to molecular therapies. CONCLUSIONS: We found that measurement of circMET levels allows identification and tracking of patients characterized by high MET activity. Circulating circMET (ccMET) detection and analysis could be a simple, cost-effective, non-invasive approach to better implement patient stratification based on MET expression, as well as to dynamically monitor over time both therapy response and clonal evolution during treatment.

Treatment with levosimendan in an experimental model of early ventilator-induced diaphragmatic dysfunction.

Introduction: Mechanical ventilation (MV) is a life-saving approach in critically ill patients. However, it may affect the diaphragmatic structure and function, beyond the lungs. Levosimendan is a calcium sensitizer widely used in clinics to improve cardiac contractility in acute heart failure patients. In vitro studies have demonstrated that levosimendan increased force-generating capacity of the diaphragm in chronic obstructive pulmonary disease patients. Thus the aim of this study was to evaluate the effects of levosimendan administration in an animal model of ventilator-induced diaphragmatic dysfunction (VIDD) on muscle contraction and diaphragm muscle cell viability. Methods: Sprague-Dawley rats underwent prolonged MV (5 hours). VIDD+Levo group received a starting bolus of levosimendan immediately after intratracheal intubation and then an intravenous infusion of levosimendan throughout the study. Diaphragms were collected for ex vivo contractility measurement (with electric stimulation), histological analysis and Western blot analysis. Healthy rats were used as the control. Results: Levosimendan treatment maintained an adequate mean arterial pressure during the entire experimental protocol, preserved levels of autophagy-related proteins (LC3BI and LC3BII) and the muscular cell diameter demonstrated by histological analysis. Levosimendan did not affect the diaphragmatic contraction or the levels of proteins involved in the protein degradation (atrogin). Conclusions: Our data suggest that levosimendan preserves muscular cell structure (cross-sectional area) and muscle autophagy after 5 hours of MV in a rat model of VIDD. However, levosimendan did not improve diaphragm contractile efficiency.

Formyl peptide receptors are involved in CTX-induced impairment of lymphocyte functions.

Crotoxin (CTX) is a neurotoxin that is isolated from the venom of Crotalus durissus terrificus, which displays immunomodulatory, anti-inflammatory, and anti-tumoral effects. Previous research has demonstrated that CTX promotes the adherence of leukocytes to the endothelial cells in blood microcirculation and the high endothelial venules of lymph nodes, which reduces the number of blood cells and lymphocytes. Studies have also shown that these effects are mediated by lipoxygenase-derived mediators. However, the exact lipoxygenase-derived eicosanoid involved in the CTX effect on lymphocytes is yet to be characterized. As CTX stimulates lipoxin-derived mediators from macrophages and lymphocyte effector functions could be modulated by activating formyl peptide receptors, we aimed to investigate whether these receptors were involved in CTX-induced redistribution and functions of lymphocytes in rats. We used male Wistar rats treated with CTX to demonstrate that Boc2 (butoxycarbonyl-Phe-Leu-Phe-Leu-Phe), an antagonist of formyl peptide receptors, prevented CTX-induced decrease in the number of circulating lymphocytes and increased the expression of the lymphocyte adhesion molecule LFA1. CTX reduced the T and B lymphocyte functions, such as lymphocyte proliferation in response to the mitogen Concanavalin A and antibody production in response to BSA immunization, respectively, which was prevented by the administration of Boc2. Importantly, mesenteric lymph node lymphocytes from CTX-treated rats showed an increased release of 15-epi-LXA4. These results indicate that formyl peptide receptors mediate CTX-induced redistribution of lymphocytes and that 15-epi-LXA4 is a key mediator of the immunosuppressive effects of CTX.

A human TRPV1 genetic variant within the channel gating domain regulates pain sensitivity in rodents.

Pain signals are relayed to the brain via a nociceptive system, and in rare cases, this nociceptive system contains genetic variants that can limit the pain response. Here, we questioned whether a human transient receptor potential vanilloid 1 (TRPV1) missense variant causes a resistance to noxious stimuli and, further, whether we could target this region with a cell-permeable peptide as a pain therapeutic. Initially using a computational approach, we identified a human K710N TRPV1 missense variant in an otherwise highly conserved region of mammalian TRPV1. After generating a TRPV1K710N-knockin mouse using CRISPR/Cas9, we discovered that the K710N variant reduced capsaicin-induced calcium influx in dorsal root ganglion neurons. The TRPV1K710N rodents also had less acute behavioral responses to noxious chemical stimuli and less hypersensitivity to nerve injury, while their response to noxious heat remained intact. Furthermore, blocking this K710 region in WT rodents using a cell-penetrating peptide limited acute behavioral responses to noxious stimuli and returned pain hypersensitivity induced by nerve injury to baseline levels. These findings identify K710 TRPV1 as a discrete site that is crucial for the control of nociception and provide insights into how to leverage rare genetic variants in humans to uncover fresh strategies for developing pain therapeutics.

Potential Applications for Growth Hormone Secretagogues Treatment of Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) arises from neuronal death due to complex interactions of genetic, molecular, and environmental factors. Currently, only two drugs, riluzole and edaravone, have been approved to slow the progression of this disease. However, ghrelin and other ligands of the GHS-R1a receptor have demonstrated interesting neuroprotective activities that could be exploited in this pathology. Ghrelin, a 28-amino acid hormone, primarily synthesized and secreted by oxyntic cells in the stomach wall, binds to the pituitary GHS-R1a and stimulates GH secretion; in addition, ghrelin is endowed with multiple extra endocrine bioactivities. Native ghrelin requires esterification with octanoic acid for binding to the GHS-R1a receptor; however, this esterified form is very labile and represents less than 10% of circulating ghrelin. A large number of synthetic compounds, the growth hormone secretagogues (GHS) encompassing short peptides, peptoids, and non-peptidic moieties, are capable of mimicking several biological activities of ghrelin, including stimulation of GH release, appetite, and elevation of blood IGF-I levels. GHS have demonstrated neuroprotective and anticonvulsant effects in experimental models of pathologies both in vitro and in vivo. To illustrate, some GHS, currently under evaluation by regulatory agencies for the treatment of human cachexia, have a good safety profile and are safe for human use. Collectively, evidence suggests that ghrelin and cognate GHS may constitute potential therapies for ALS.

TEGDMA-Functionalized Dicalcium Phosphate Dihydrate Resin-Based Composites Prevent Secondary Caries in an In Vitro Biofilm Model.

This study evaluated the efficacy of experimental TEGDMA-functionalized dicalcium phosphate dihydrate (T-DCPD) filler-based resin-based composites (RBC) in preventing caries lesions around the restoration margins (secondary caries, SC). Standardized Class-II cavities were made in sound molars with the cervical margin in dentin. Cavities were filled with a commercial resin-modified glass-ionomer cement (RMGIC) or experimental RBCs containing a bisGMA-TEGDMA resin blend and one of the following inorganic fractions: 60 wt.% Ba glass (RBC-0); 40 wt.% Ba glass and 20 wt.% T-DCPD (RBC-20); or 20 wt.% Ba glass and 40 wt.% T-DCPD (RBC-40). An open-system bioreactor produced Streptococcus mutans biofilm-driven SC. Specimens were scanned using micro-CT to evaluate demineralization depths. Scanning electron microscopy and energy-dispersive X-ray spectroscopy characterized the specimen surfaces, and antimicrobial activity, buffering effect, and ion uptake by the biofilms were also evaluated. ANOVA and Tukey's tests were applied at p < 0.05. RBC-0 and RBC-20 showed SC development in dentin, while RBC-40 and RMGIC significantly reduced the lesion depth at the restoration margin (p < 0.0001). Initial enamel demineralization could be observed only around the RBC-0 and RBC-20 restorations. Direct antibiofilm activity can explain SC reduction by RMGIC, whereas a buffering effect on the acidogenicity of biofilm can explain the behavior of RBC-40. Experimental RBC with CaP-releasing functionalized T-DCPD filler could prevent SC with the same efficacy as F-releasing materials.

Microbiological models for accelerated development of secondary caries in vitro.

OBJECTIVES: The current study aimed to compare the efficacy of two in vitro microbiological models based on open and closed systems designed to obtain secondary caries in an accelerated and reproducible way. METHODS: A conventional resin-based composite (RBC - Majesty ES-2; Kuraray, Japan) and a resin-modified glass-ionomer cement (RMGIC - Ionolux; VOCO, Germany) were used to restore standardized class II cavities (n = 4/tooth, cervical margin in dentin) in 16 human molars. The ability to produce secondary caries with Streptococcus mutans biofilms was tested using either an open-cycle or closed-cycle bioreactor (n = 8 specimens/model). Specimens were scanned before and after the biofilm exposure using micro-CT (Skyscan 1176, 9 µm resolution, 80 kV, 300 mA). Image reconstruction was performed, and demineralization depths (µm) were evaluated at the restoration margins and a distance of 1.0 mm. RESULTS: Dentin demineralization could be observed in all specimens, and enamel demineralization in 50% of the specimens. The open system bioreactor produced lesions with significantly higher overall demineralization depths (p < .001). However, demineralization depths at a 1.0 mm distance from the restoration margins showed no difference between open and closed systems or materials. In the open system, significantly lower demineralization depths were observed in proximity to RMGIC than RBC (p < .001), which was not significantly different in the closed system (p = .382). CONCLUSIONS: Both systems produced in vitro secondary caries in an accelerated way. However, the open-cycle bioreactor system confirmed the caries-protective activity exerted by the RMGIC material in contrast to the RBC, better simulating materials' clinical behavior. CLINICAL SIGNIFICANCE: The possibility of obtaining accelerated and reproducible secondary caries development in vitro is fundamental in testing the behavior of conventional and yet-to-come restorative dental materials. Such systems can provide faster outcomes regarding the performance of dental restorative materials compared to clinical studies, notwithstanding the importance of the latter.

Crotalphine Modulates Microglia M1/M2 Phenotypes and Induces Spinal Analgesia Mediated by Opioid-Cannabinoid Systems.

Pain is a worldwide public health problem and its treatment is still a challenge since clinically available drugs do not completely reverse chronic painful states or induce undesirable effects. Crotalphine is a 14 amino acids synthetic peptide that induces a potent and long-lasting analgesic effect on acute and chronic pain models, peripherally mediated by the endogenous release of dynorphin A and the desensitization of the transient receptor potential ankyrin 1 (TRPA1) receptor. However, the effects of crotalphine on the central nervous system (CNS) and the signaling pathway have not been investigated. Thus, the central effect of crotalphine was evaluated on the partial sciatic nerve ligation (PSNL)-induced chronic neuropathic pain model. Crotalphine (100 µg/kg, p.o.)-induced analgesia on the 14th day after surgery lasting up to 24 h after administration. This effect was prevented by intrathecal administration of CB1 (AM251) or CB2 (AM630) cannabinoid receptor antagonists. Besides that, crotalphine-induced analgesia was reversed by CTOP, nor-BNI, and naltrindole, antagonists of mu, kappa, and delta-opioid receptors, respectively, and also by the specific antibodies for ß-endorphin, dynorphin-A, and met-enkephalin. Likewise, the analgesic effect of crotalphine was blocked by the intrathecal administration of minocycline, an inhibitor of microglial activation and proliferation. Additionally, crotalphine decreased the PSNL-induced IL-6 release in the spinal cord. Importantly, in vitro, crotalphine inhibited LPS-induced CD86 expression and upregulated CD206 expression in BV-2 cells, demonstrating a polarization of microglial cells towards the M2 phenotype. These results demonstrated that crotalphine, besides activating opioid and cannabinoid analgesic systems, impairs central neuroinflammation, confirming the neuromodulatory mechanism involved in the crotalphine analgesic effect.

Caspase-1 and Cathepsin B Inhibitors from Marine Invertebrates, Aiming at a Reduction in Neuroinflammation.

Neuroinflammation is a condition associated with several types of dementia, such as Alzheimer's disease (AD), mainly caused by an inflammatory response to amyloid peptides that induce microglial activation, with subsequent cytokine release. Neuronal caspase-1 from inflammasome and cathepsin B are key enzymes mediating neuroinflammation in AD, therefore, revealing new molecules to modulate these enzymes may be an interesting approach to treat neurodegenerative diseases. In this study, we searched for new caspase-1 and cathepsin B inhibitors from five species of Brazilian marine invertebrates (four cnidarians and one echinoderm). The results show that the extract of the box jellyfish Chiropsalmus quadrumanus inhibits caspase-1. This extract was fractionated, and the products monitored for their inhibitory activity, until the obtention of a pure molecule, which was identified as trigonelline by mass spectrometry. Moreover, four extracts inhibit cathepsin B, and Exaiptasia diaphana was selected for subsequent fractionation and characterization, resulting in the identification of betaine as being responsible for the inhibitory action. Both molecules are already found in marine organisms, however, this is the first study showing a potent inhibitory effect on caspase-1 and cathepsin B activities. Therefore, these new prototypes can be considered for the enzyme inhibition and subsequent control of the neuroinflammation.

MicroRNA profiling predicts positive nodal status in papillary thyroid carcinoma in the preoperative setting.

BACKGROUND: The molecular characterization of thyroid nodules in cytological samples has so far been focused on discriminating between benign and malignant forms in a purely diagnostic setting. The evidence on the impact of molecular biomarkers to determine the risk of aggressiveness in cytologically "neoplastic" lesions is limited to genomic alterations (such as BRAF and TERT mutations). The aim of our study was to assess the preoperative role of microRNAs (miRNAs) in predicting the nodal status of patients with papillary thyroid cancer. METHODS: A pilot series of histological samples of papillary thyroid carcinoma with (6 cases) or without (6 cases) lymph node metastases, matched for other major clinical and pathological features, was analyzed for global miRNA expression in a screening phase. A set of miRNAs was then validated in a series of 63 consecutive cytological samples of papillary carcinomas: 48 pN-negative and 15 pN-positive at histology. RESULTS: Unsupervised cluster analysis segregated surgical pN-negative and pN-positive samples, except for 1 case. The 45 differentially expressed miRNAs in pN-positive versus pN-negative cases were predicted to regulate a wide range of cellular pathways, enriched for Wnt, gonadotropin-releasing hormone receptor, and cerulein/cholecystokinin receptor signaling. In agreement with their profiles in surgical samples, 4 miRNAs of the 10 selected for validation (miR-154-3p, miR-299-5p, miR-376a-3p, and miR-302E) had a significant differential expression in cytological samples of papillary carcinoma with lymph node metastases and predicted the positive nodal status with a relatively good performance. CONCLUSIONS: MiRNA profiling is a potential promising strategy to define papillary carcinoma aggressiveness in the preoperative setting.

Human Sensory Neuron-like Cells and Glycated Collagen Matrix as a Model for the Screening of Analgesic Compounds.

Increased collagen-derived advanced glycation end-products (AGEs) are consistently related to painful diseases, including osteoarthritis, diabetic neuropathy, and neurodegenerative disorders. We have recently developed a model combining a two-dimensional glycated extracellular matrix (ECM-GC) and primary dorsal root ganglion (DRG) that mimicked a pro-nociceptive microenvironment. However, culturing primary cells is still a challenge for large-scale screening studies. Here, we characterized a new model using ECM-GC as a stimulus for human sensory-like neurons differentiated from SH-SY5Y cell lines to screen for analgesic compounds. First, we confirmed that the differentiation process induces the expression of neuron markers (MAP2, RBFOX3 (NeuN), and TUBB3 (ß-III tubulin), as well as sensory neuron markers critical for pain sensation (TRPV1, SCN9A (Nav1.7), SCN10A (Nav1.8), and SCN11A (Nav1.9). Next, we showed that ECM-GC increased c-Fos expression in human sensory-like neurons, which is suggestive of neuronal activation. In addition, ECM-GC upregulated the expression of critical genes involved in pain, including SCN9A and TACR1. Of interest, ECM-GC induced substance P release, a neuropeptide widely involved in neuroinflammation and pain. Finally, morphine, the prototype opiate, decreased ECM-GC-induced substance P release. Together, our results suggest that we established a functional model that can be useful as a platform for screening candidates for the management of painful conditions.

PKCζ-Mitogen-Activated Protein Kinase Signaling Mediates Crotalphine-Induced Antinociception.

Crotalphine (CRP) is a structural analogue to a peptide that was first identified in the crude venom from the South American rattlesnake Crotalus durissus terrificus. This peptide induces a potent and long-lasting antinociceptive effect that is mediated by the activation of peripheral opioid receptors. The opioid receptor activation regulates a variety of intracellular signaling, including the mitogen-activated protein kinase (MAPK) pathway. Using primary cultures of sensory neurons, it was demonstrated that crotalphine increases the level of activated ERK1/2 and JNK-MAPKs and this increase is dependent on the activation of protein kinase Cζ (PKCζ). However, whether PKCζ-MAPK signaling is critical for crotalphine-induced antinociception is unknown. Here, we biochemically demonstrated that the systemic crotalphine activates ERK1/2 and JNK and decreases the phosphorylation of p38 in the lumbar spinal cord. The in vivo pharmacological inhibition of spinal ERK1/2 and JNK, but not of p38, blocks the antinociceptive effect of crotalphine. Of interest, the administration of a PKCζ pseudosubstrate (PKCζ inhibitor) prevents crotalphine-induced ERK activation in the spinal cord, followed by the abolishment of crotalphine-induced analgesia. Together, our results demonstrate that the PKCζ-ERK signaling pathway is involved in crotalphine-induced analgesia. Our study opens a perspective for the PKCζ-MAPK axis as a target for pain control.

Activation of PKCε-ALDH2 Axis Prevents 4-HNE-Induced Pain in Mice.

Protein kinase Cε (PKCε) is highly expressed in nociceptor neurons and its activation has been reported as pro-nociceptive. Intriguingly, we previously demonstrated that activation of the mitochondrial PKCε substrate aldehyde dehydrogenase-2 (ALDH2) results in anti-nociceptive effects. ALDH2 is a major enzyme responsible for the clearance of 4-hydroxy-2-nonenal (4-HNE), an oxidative stress byproduct accumulated in inflammatory conditions and sufficient to induce pain hypersensitivity in rodents. Here we determined the contribution of the PKCε-ALDH2 axis during 4-HNE-induced mechanical hypersensitivity. Using knockout mice, we demonstrated that PKCε is essential for the nociception recovery during 4-HNE-induced hypersensitivity. We also found that ALDH2 deficient knockin mice display increased 4-HNE-induced nociceptive behavior. As proof of concept, the use of a selective peptide activator of PKCε (ΨεHSP90), which favors PKCε translocation to mitochondria and activation of PKCε-ALDH2 axis, was sufficient to block 4-HNE-induced hypersensitivity in WT, but not in ALDH2-deficient mice. Similarly, ΨεHSP90 administration prevented mechanical hypersensitivity induced by endogenous production of 4-HNE after carrageenan injection. These findings provide evidence that selective activation of mitochondrial PKCε-ALDH2 axis is important to mitigate aldehyde-mediated pain in rodents, suggesting that ΨεHSP90 and small molecules that mimic it may be a potential treatment for patients with pain.