Efficacy of monoclonal antibodies against calcitonin gene-related peptide (CGRP) or its receptor (calcitonin receptor-like receptor/receptor activity modifying protein-1, CLR/RAMP1) implicates peripherally-released CGRP in migraine pain. However, the site and mechanism of CGRP-evoked peripheral pain remain unclear. By cell-selective RAMP1 gene deletion, we reveal that CGRP released from mouse cutaneous trigeminal fibers targets CLR/RAMP1 on surrounding Schwann cells to evoke periorbital mechanical allodynia. CLR/RAMP1 activation in human and mouse Schwann cells generates long-lasting signals from endosomes that evoke cAMP-dependent formation of NO. NO, by gating Schwann cell transient receptor potential ankyrin 1 (TRPA1), releases ROS, which in a feed-forward manner sustain allodynia via nociceptor TRPA1. When encapsulated into nanoparticles that release cargo in acidified endosomes, a CLR/RAMP1 antagonist provides superior inhibition of CGRP signaling and allodynia in mice. Our data suggest that the CGRP-mediated neuronal/Schwann cell pathway mediates allodynia associated with neurogenic inflammation, contributing to the algesic action of CGRP in mice.

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Although macrophages (MΦ) are known to play a central role in neuropathic pain, their contribution to cancer pain has not been established. Here we report that depletion of sciatic nerve resident MΦs (rMΦ) in mice attenuates mechanical/cold hypersensitivity and spontaneous pain evoked by intraplantar injection of melanoma or lung carcinoma cells. MΦcolony stimulating factor (M-CSF) was upregulated in the sciatic nerve trunk and mediated cancer-evoked pain via rMΦ expansion, transient receptor potential ankyrin 1 (TRPA1) activation, and oxidative stress. Targeted deletion of Trpa1 revealed a key role for Schwann cell TRPA1 in sciatic nerve rMΦ expansion and pain-like behaviors. Depletion of rMΦs in a medial portion of the sciatic nerve prevented pain-like behaviors. Collectively, we identified a feed-forward pathway involving M-CSF, rMΦ, oxidative stress and Schwann cell TRPA1 that operates throughout the nerve trunk to signal cancer-evoked pain. SignificanceSchwann cell TRPA1 sustains cancer pain through release of M-CSF and oxidative stress, which promote the expansion and the proalgesic actions of intraneural macrophages.

Background The mechanism underlying the pain symptoms associated with chemotherapeutic-induced peripheral neuropathy (CIPN) is poorly understood. Transient receptor potential ankyrin 1 (TRPA1), TRP vanilloid 4 (TRPV4), TRPV1, and oxidative stress have been implicated in several rodent models of CIPN-evoked allodynia. Thalidomide causes a painful CIPN in patients via an unknown mechanism. Surprisingly, the pathway responsible for such proalgesic response has not yet been investigated in animal models. Results Here, we reveal that a single systemic administration of thalidomide and its derivatives, lenalidomide and pomalidomide, elicits prolonged (~ 35 days) mechanical and cold hypersensitivity in C57BL/6J mouse hind paw. Pharmacological antagonism or genetic deletion studies indicated that both TRPA1 and TRPV4, but not TRPV1, contribute to mechanical allodynia, whereas cold hypersensitivity was entirely due to TRPA1. Thalidomide per se did not stimulate recombinant and constitutive TRPA1 and TRPV4 channels in vitro, which, however, were activated by the oxidative stress byproduct, hydrogen peroxide. Systemic treatment with an antioxidant attenuated mechanical and cold hypersensitivity, and the increase in oxidative stress in hind paw, sciatic nerve, and lumbar spinal cord produced by thalidomide. Notably, central (intrathecal) or peripheral (intraplantar) treatments with channel antagonists or an antioxidant revealed that oxidative stress-dependent activation of peripheral TRPA1 mediates cold allodynia and part of mechanical allodynia. However, oxidative stress-induced activation of central TRPV4 mediated the residual TRPA1-resistant component of mechanical allodynia. Conclusions Targeting of peripheral TRPA1 and central TRPV4 may be required to attenuate pain associated with CIPN elicited by thalidomide and related drugs.

Efficacy of monoclonal antibodies against calcitonin gene-related peptide (CGRP) or its receptor (calcitonin receptor-like receptor/receptor activity modifying protein-1, CLR/RAMP1) implicates peripherally-released CGRP in migraine pain. However, the site and mechanism of CGRP-evoked migraine pain remain unknown. By cell-selective RAMP1 gene deletion, we reveal that CGRP released from mouse cutaneous trigeminal fibers targets CLR/RAMP1 on surrounding Schwann cells to evoke periorbital mechanical allodynia. CLR/RAMP1 activation in human and mouse Schwann cells generates long-lasting signals from endosomes that evoke cAMP-dependent formation of NO. NO, by gating Schwann cell transient receptor potential ankyrin 1 (TRPA1), releases ROS, which in a feed-forward manner sustain allodynia via nociceptor TRPA1. When encapsulated into nanoparticles that release cargo in acidified endosomes, a CLR/RAMP1 antagonist provides superior inhibition of CGRP signaling and allodynia in mice. The CGRP-mediated neuronal/Schwann cell pathway is critical to mediate allodynia associated with neurogenic inflammation, thus contributing to the pro-migraine action of CGRP.

Background and Purpose The pro‐algesic transient receptor potential ankyrin 1 (TRPA1) channel, expressed by a subpopulation of primary sensory neurons, has been implicated in various pain models in mice. However, evidence in rats indicates that TRPA1 conveys nociceptive signals elicited by channel activators, but not those associated with tissue inflammation or nerve injury. Here, in rats, we explored the TRPA1 role in mechanical allodynia associated with stimulation of peptidergic primary sensory neurons (neurogenic inflammation) and moderate (partial sciatic nerve ligation, pSNL) or severe (chronic constriction injury, CCI) sciatic nerve injury. Experimental Approach Acute nociception and mechanical hypersensitivity associated with neurogenic inflammation and sciatic nerve injury (pSNL and CCI) were investigated in rats with TRPA1 pharmacological antagonism or genetic silencing. TRPA1 presence and function were analysed in cultured rat Schwann cells. Key Results Hind paw mechanical allodynia (HPMA), but not acute nociception, evoked by local injection of capsaicin or allyl isothiocyanate, the TRP vanilloid 1 (TRPV1) or the TRPA1 activators was mediated by CGRP released from peripheral sensory nerve terminals. CGRP‐evoked HPMA was sustained by a ROS‐dependent TRPA1 activation, probably in Schwann cells. HPMA evoked by pSNL, but not that evoked by CCI, was mediated by ROS and TRPA1 without the involvement of CGRP. Conclusions and Implications As found in mice, TRPA1 mediates mechanical allodynia associated with neurogenic inflammation and moderate nerve injury in rats. The channel contribution to mechanical hypersensitivity is a common feature in rodents and might be explored in humans.

Aim: The objective of this study was to investigate the possible synergy between doxycycline and photodynamic therapy against Helicobacter pylori and to evaluate the possible side effects on adenocarcinoma gastric cells with and without protoporphyrin IX. Materials & methods: Three H. pylori strains (ATCC 700392, 43504 and 49503) were grown on solid medium either with, or without, doxycycline at subinhibitory concentrations, and irradiated for 10, 20 and 30 minutes with a 400 nm-peaked light source. The phototoxicity tests on AGS cells were evaluated by MTT assay. Results: The photodynamic therapy and doxycycline combination showed an antibacterial synergistic effect with no significant toxicities. Conclusion: The synergistic treatment could be considered as an interesting therapeutic option.

Abstract. The aim of the present study was to determine the expression of vascular endothelial growth factor (VEGF) family receptors (VEGFR) in placentas from pregnancies complicated by hypertensive disorders of different clinical severity. Placental tissue from women with gestational hypertension, pre-eclampsia, pre-eclampsia with haemolysis, elevated liver enzymes and low platelets (HELLP syndrome) and normotensive women, as a control group, was examined. Immunohistochemical techniques, reverse transcription-polymerase chain reaction and western blot were used to evaluate receptor expression. In cases with gestational hypertension, as well as in control cases, VEGFR-1 and VEGFR-3 immunoreactivity was detected in all placental components, whereas in placentas from the pre-eclampsia and pre-eclampsia with HELLP syndrome groups, VEGFR-1 and VEGFR-3 immunoreactivity was detected only in some portions of trophoblast and/or some vessels and/or clusters of stromal cells. In the control group, VEGFR-2 immunoreactivity was observed only in the vessels, whereas the hypertensive groups showed VEGF-2 immunoreactivity also in trophoblast and stromal cells. The mRNA levels of the three receptors in the group with gestational hypertension were higher with respect to those in the control group. Placentas from pregnancies with pre-eclampsia showed lowest mRNA expression levels, whereas placentas from women with pre-eclampsia plus HELLP syndrome showed higher mRNA expression levels with respect to the three other groups. Receptor protein levels were lower in pathological cases compared with levles in the control group. These findings demonstrate a dysregulation of placental expression of VEGF family receptors related to the degree of clinical severity of the hypertensive disorder.