Yohance M. Allette

The HMGB1-RAGE axis mediates traumatic brain injury–induced pulmonary dysfunction in lung transplantation

Traumatic brain injury induces acute lung injury that negatively impacts the physiology of the donor lung before and after lung transplantation. Sounding the Alarm for RAGE Only 20% of lungs are transplantable because traumatic brain injury, a major cause of death in organ doors, may induce acute lung injury. High-mobility group box-1 (HMGB1) release from the injured brain likely contributes to acute lung injury in donors by preferentially interacting with receptor for advanced glycation end products (RAGE) in the lung. Blocking the HMGB1-RAGE axis improves lung function in murine donors with traumatic brain injury and after transplant. In translational studies, lungs sourced from donors with high HMGB1 levels had worse pulmonary function after transplant. Targeting the HMGB1-RAGE axis may increase the number of lungs available for transplantation and improve patient outcomes. Traumatic brain injury (TBI) results in systemic inflammatory responses that affect the lung. This is especially critical in the setting of lung transplantation, where more than half of donor allografts are obtained postmortem from individuals with TBI. The mechanism by which TBI causes pulmonary dysfunction remains unclear but may involve the interaction of high-mobility group box-1 (HMGB1) protein with the receptor for advanced glycation end products (RAGE). To investigate the role of HMGB1 and RAGE in TBI-induced lung dysfunction, RAGE-sufficient (wild-type) or RAGE-deficient (RAGE−/−) C57BL/6 mice were subjected to TBI through controlled cortical impact and studied for cardiopulmonary injury. Compared to control animals, TBI induced systemic hypoxia, acute lung injury, pulmonary neutrophilia, and decreased compliance (a measure of the lungs’ ability to expand), all of which were attenuated in RAGE−/− mice. Neutralizing systemic HMGB1 induced by TBI reversed hypoxia and improved lung compliance. Compared to wild-type donors, lungs from RAGE−/− TBI donors did not develop acute lung injury after transplantation. In a study of clinical transplantation, elevated systemic HMGB1 in donors correlated with impaired systemic oxygenation of the donor lung before transplantation and predicted impaired oxygenation after transplantation. These data suggest that the HMGB1-RAGE axis plays a role in the mechanism by which TBI induces lung dysfunction and that targeting this pathway before transplant may improve recipient outcomes after lung transplantation.

Acrolein involvement in sensory and behavioral hypersensitivity following spinal cord injury in the rat

Growing evidence suggests that oxidative stress, as associated with spinal cord injury (SCI), may play a critical role in both neuroinflammation and neuropathic pain conditions. The production of the endogenous aldehyde acrolein, following lipid peroxidation during the inflammatory response, may contribute to peripheral sensitization and hyperreflexia following SCI via the TRPA1‐dependent mechanism. Here, we report that there are enhanced levels of acrolein and increased neuronal sensitivity to the aldehyde for at least 14 days after SCI. Concurrent with injury‐induced increases in acrolein concentration is an increased expression of TRPA1 in the lumbar (L3–L6) sensory ganglia. As proof of the potential pronociceptive role for acrolein, intrathecal injections of acrolein revealed enhanced sensitivity to both tactile and thermal stimuli for up to 10 days, supporting the compounds pro‐nociceptive functionality. Treatment of SCI animals with the acrolein scavenger hydralazine produced moderate improvement in tactile responses as well as robust changes in thermal sensitivity for up to 49 days. Taken together, these data suggest that acrolein directly modulates SCI‐associated pain behavior, making it a novel therapeutic target for preclinical and clinical SCI as an analgesic.

Suppression of pain-related behavior in two distinct rodent models of peripheral neuropathy by a homopolyarginine-conjugated CRMP2 peptide

The N‐type voltage‐gated calcium channel (CaV2.2) is a clinically endorsed target in chronic pain treatments. As directly targeting the channel can lead to multiple adverse side effects, targeting modulators of CaV2.2 may prove better. We previously identified ST1‐104, a short peptide from the collapsin response mediator protein 2 (CRMP2), which disrupted the CaV2.2–CRMP2 interaction and suppressed a model of HIV‐related neuropathy induced by anti‐retroviral therapy but not traumatic neuropathy. Here, we report ST2‐104 –a peptide wherein the cell‐penetrating TAT motif has been supplanted with a homopolyarginine motif, which dose‐dependently inhibits the CaV2.2–CRMP2 interaction and inhibits depolarization‐evoked Ca2+ influx in sensory neurons. Ca2+ influx via activation of vanilloid receptors is not affected by either peptide. Systemic administration of ST2‐104 does not affect thermal or tactile nociceptive behavioral changes. Importantly, ST2‐104 transiently reduces persistent mechanical hypersensitivity induced by systemic administration of the anti‐retroviral drug 2′,3′‐dideoxycytidine (ddC) and following tibial nerve injury (TNI). Possible mechanistic explanations for the broader efficacy of ST2‐104 are discussed.

Bone Pain Induced by Multiple Myeloma Is Reduced by Targeting V-ATPase and ASIC3

Multiple myeloma patients experience severe bone pain (MMBP) that is undertreated and poorly understood. In this study, we studied MMBP in an intratibial mouse xenograft model that employs JJN3 human multiple myeloma cells. In this model, mice develop MMBP associated in bone with increased sprouting of calcitonin gene-related peptide-positive (CGRP+) sensory nerves and in dorsal root ganglia (DRG) with upregulation of phosphorylated ERK1/2 (pERK1/2) and pCREB, two molecular indicators of neuron excitation. We found that JJN3 cells expressed a vacuolar proton pump (V-ATPase) that induced an acidic bone microenvironment. Inhibition of JJN3-colonized bone acidification by a single injection of the selective V-ATPase inhibitor, bafilomycin A1, decreased MMBP, CGRP+ sensory neuron sprouting, and pERK1/2 and pCREB expression in DRG. CGRP+ sensory nerves also expressed increased levels of the acid-sensing nociceptor ASIC3. Notably, a single injection of the selective ASIC3 antagonist APETx2 dramatically reduced MMBP in the model. Mechanistic investigations in primary DRG neurons cocultured with JJN3 cells showed increased neurite outgrowth and excitation inhibited by bafilomycin A1 or APETx2. Furthermore, combining APETx2 with bafilomycin A1 reduced MMBP to a greater extent than either agent alone. Finally, combining bafilomycin A1 with the osteoclast inhibitor zoledronic acid was sufficient to ameliorate MMBP, which was refractory to zoledronic acid. Overall, our results show that osteoclasts and multiple myeloma cooperate to induce an acidic bone microenvironment that evokes MMBP as a result of the excitation of ASIC3-activated sensory neurons. Furthermore, they present a mechanistic rationale for targeting ASIC3 on neurons along with the multiple myeloma-induced acidic bone microenvironment as a strategy to relieve MMBP in patients. Cancer Res; 77(6); 1283-95. ©2017 AACR.

Carbamazepine potentiates the effectiveness of morphine in a rodent model of neuropathic pain.

Approximately 60% of morphine is glucuronidated to morphine-3-glucuronide (M3G) which may aggravate preexisting pain conditions. Accumulating evidence indicates that M3G signaling through neuronal Toll-like receptor 4 (TLR4) may be central to this proalgesic signaling event. These events are known to include elevated neuronal excitability, increased voltage-gated sodium (NaV) current, tactile allodynia and decreased opioid analgesic efficacy. Using an in vitro ratiometric-based calcium influx analysis of acutely dissociated small and medium-diameter neurons derived from lumbar dorsal root ganglion (DRG), we observed that M3G-sensitive neurons responded to lipopolysaccharide (LPS) and over 35% of these M3G/LPS-responsive cells exhibited sensitivity to capsaicin. In addition, M3G-exposed sensory neurons significantly increased excitatory activity and potentiated NaV current as measured by current and voltage clamp, when compared to baseline level measurements. The M3G-dependent excitability and potentiation of NaV current in these sensory neurons could be reversed by the addition of carbamazepine (CBZ), a known inhibitor of several NaV currents. We then compared the efficacy between CBZ and morphine as independent agents, to the combined treatment of both drugs simultaneously, in the tibial nerve injury (TNI) model of neuropathic pain. The potent anti-nociceptive effects of morphine (5 mg/kg, i.p.) were observed in TNI rodents at post-injury day (PID) 7–14 and absent at PID21–28, while administration of CBZ (10 mg/kg, i.p.) alone failed to produce anti-nociceptive effects at any time following TNI (PID 7–28). In contrast to either drug alone at PID28, the combination of morphine and CBZ completely attenuated tactile hyperalgesia in the rodent TNI model. The basis for the potentiation of morphine in combination with CBZ may be due to the effects of a latent upregulation of NaV1.7 in the DRG following TNI. Taken together, our observations demonstrate a potential therapeutic use of morphine and CBZ as a combinational treatment for neuropathic pain.

Decoy peptide targeted to Toll-IL-1R domain inhibits LPS and TLR4-active metabolite morphine-3 glucuronide sensitization of sensory neurons.

Accumulating evidence indicates that Toll-like receptor (TLR) signaling adapter protein interactions with Toll/Interleukin-1 Receptor (TIR) domains present in sensory neurons may modulate neuropathic pain states. Following ligand interaction with TLRs, TIR serves to both initiate intracellular signaling and facilitate recruitment of signaling adapter proteins to the intracytoplasmic domain. Although TLR TIR is central to a number of TLR signaling cascades, its role in sensory neurons is poorly understood. In this study we investigated the degree to which TLR TIR decoy peptide modified to include a TAT sequence (Trans-Activator of Transcription gene in HIV; TAT-4BB) affected LPS-induced intracellular calcium flux and excitation in sensory neurons, and behavioral changes due to TLR4 active metabolite, morphine-3-glucuronide (M3G) exposure in vivo. TAT-4BB inhibited LPS-induced calcium changes in a majority of sensory neurons and decreased LPS-dependent neuronal excitability in small diameter neurons. Acute systemic administration of the TAT-4BB reversed M3G-induced tactile allodynia in a dose-dependent manner but did not affect motor activity, anxiety or responses to noxious thermal stimulus. These data suggest that targeting TLR TIR domains may provide novel pharmacological targets to reduce or reverse TLR4-dependent pain behavior in the rodent.