Lihuan Liang

Hydrogen peroxide is a novel mediator of inflammatory hyperalgesia, acting via transient receptor potential vanilloid 1-dependent and independent mechanisms

Abstract Inflammatory diseases associated with pain are often difficult to treat in the clinic due to insufficient understanding of the nociceptive pathways involved. Recently, there has been considerable interest in the role of reactive oxygen species (ROS) in inflammatory disease, but little is known of the role of hydrogen peroxide (H2O2) in hyperalgesia. In the present study, intraplantar injection of H2O2‐induced a significant dose‐ and time‐dependent mechanical and thermal hyperalgesia in the mouse hind paw, with increased c‐fos activity observed in the dorsal horn of the spinal cord. H2O2 also induced significant nociceptive behavior such as increased paw licking and decreased body liftings. H2O2 levels were significantly raised in the carrageenan‐induced hind paw inflammation model, showing that this ROS is produced endogenously in a model of inflammation. Moreover, superoxide dismutase and catalase significantly reduced carrageenan‐induced mechanical and thermal hyperalgesia, providing evidence of a functionally significant endogenous role. Thermal, but not mechanical, hyperalgesia in response to H2O2 (i.pl.) was longer lasting in TRPV1 wild type mice compared to TRPV1 knockouts. It is unlikely that downstream lipid peroxidation was increased by H2O2. In conclusion, we demonstrate a notable effect of H2O2 in mediating inflammatory hyperalgesia, thus highlighting H2O2 removal as a novel therapeutic target for anti‐hyperalgesic drugs in the clinic.

The transient receptor potential vanilloid 1 (TRPV1) receptor protects against the onset of sepsis after endotoxin

Transient potential vanilloid 1 (TRPV1) receptor is an ion channel receptor primarily localized on sensory nerves and activated by specific stimuli to initiate and amplify pain and inflammation, as typified by murine models of scald and arthritis. Little is known of the role of TRPV1 in sepsis, an infective disease associated with inflammation. Through use of a suble‐thal murine model of lipopolysaccharide‐induced peritoneal sepsis, we provide novel evidence that genetic deletion of TRPV1 leads to an enhanced onset of various pathological components of systemic endotox‐emia. Paired studies of TRPV1 knockout (KO) and wild‐type mice demonstrate significantly enhanced hypotension (56±2% vs. 38±6% decrease in blood pressure, n=12), hypothermia (13±3% vs. 7±1% decrease in core temperature, n=6), and peritoneal exudate mediator levels (TNF‐α, 0.78±0.2 vs. 0.38±0.1 ng/ml; nitrite, for NO, 35±10 vs. 15±3 μM; n=8) in TRPV1 KO mice, indicating loss of protective effect. Findings correlated with liver edema and raised plasma levels of aspartate aminotransferase in TRPV1 KO mice. These data suggest that TRPV1 may play an important regulatory role in sepsis independent of the major sensory neuropeptide substance P. The findings are relevant to developing strategies that increase the beneficial, and reduce the harmful, components of sepsis to prevent and treat this often fatal condition.—Clark, N., Keeble, J., Fernandes, E. S., Starr, A., Liang, L., Sugden, D., de Winter, P., Brain, S. D. The transient receptor potential vanilloid 1 (TRPV1) receptor protects against the onset of sepsis after endotoxin. FASEB J. 21, 3747–3755 (2007)

TRPV1 Deletion Enhances Local Inflammation and Accelerates the Onset of Systemic Inflammatory Response Syndrome

The transient receptor potential vanilloid 1 (TRPV1) is primarily localized to sensory nerve fibers and is associated with the stimulation of pain and inflammation. TRPV1 knockout (TRPV1KO) mice show enhanced LPS-induced sepsis compared with wild type (WT). This implies that TRPV1 may have a key modulatory role in increasing the beneficial and reducing the harmful components in sepsis. We investigated immune and inflammatory mechanisms in a cecal ligation and puncture (CLP) model of sepsis over 24 h. CLP TRPV1KO mice exhibited significant hypothermia, hypotension, and organ dysfunction compared with CLP WT mice. Analysis of the inflammatory responses at the site of initial infection (peritoneal cavity) revealed that CLP TRPV1KO mice exhibited: 1) decreased mononuclear cell integrity associated with apoptosis, 2) decreased macrophage tachykinin NK1-dependent phagocytosis, 3) substantially decreased levels of nitrite (indicative of NO) and reactive oxygen species, 4) increased cytokine levels, and 5) decreased bacteria clearance when compared with CLP WT mice. Therefore, TRPV1 deletion is associated with impaired macrophage-associated defense mechanisms. Thus, TRPV1 acts to protect against the damaging impact of sepsis and may influence the transition from local to a systemic inflammatory state.

A Role for TRPV1 in Influencing the Onset of Cardiovascular Disease in Obesity

Obesity induced by Western diets is associated with type 2 diabetes mellitus and cardiovascular diseases, although underlying mechanisms are unclear. We investigated a murine model of diet-induced obesity to determine the effect of transient potential receptor vanilloid 1 (TRPV1) deletion on hypertension and metabolic syndrome. Wild-type and TRPV1 knockout mice were fed normal or high-fat diet from 3 to 15 weeks. High-fat diet-fed mice from both genotypes became obese, with similar increases in body and adipose tissue weights. High-fat diet-fed TRPV1 knockout mice showed significantly improved handling of glucose compared with high-fat diet-fed wild-type mice. Hypertension, vascular hypertrophy, and altered nociception were observed in high-fat diet-fed wild-type but not high-fat diet-fed TRPV1 knockout mice. Wild-type, but not high-fat diet-fed TRPV1 knockout, mice demonstrated remodeling in terms of aortic vascular hypertrophy and increased heart and kidney weight, although resistance vessel responses were similar in each. Moreover, the wild-type mice had significantly increased plasma levels of leptin, interleukin 10 and interleukin 1&bgr;, whereas samples from TRPV1 knockout mice did not show significant increases. Our results do not support the concept that TRPV1 plays a major role in influencing weight gain. However, we identified a role of TRPV1 in the deleterious effects observed with high-fat feeding in terms of inducing hypertension, impairing thermal nociception sensitivity, and reducing glucose tolerance. The observation of raised levels of adipokines in wild-type but not TRPV1 knockout mice is in keeping with TRPV1 involvement in stimulating the proinflammatory network that is central to obesity-induced hypertension and sensory neuronal dysfunction.

Superoxide generation and leukocyte accumulation: key elements in the mediation of leukotriene B4-induced itch by transient receptor potential ankyrin 1 and transient receptor potential vanilloid 1

The underlying mechanisms of itch are poorly understood. We have investigated a model involving the chemoattractant leukotriene B4 (LTB4) that is up‐regulated in common skin diseases. Intradermal injection of LTB4 (0.1 nmol/site) into female CD1 mice induced significant scratching movements (used as an itch index) compared with vehicle‐injected (0.1% bovine serum albumin‐saline) mice. Intraperitoneal transient receptor potential (TRP) channel antagonist treatment significantly inhibited itch as follows: TRP vanilloid 1 (TRPV1) antagonist SB366791 (0.5 mg/kg, by 97%) and the TRP ankyrin 1 (TRPA1) antagonists TCS 5861528 (10 mg/kg; 82%) and HC‐030031 (100 mg/kg; 76%). Leukotriene B4 receptor 2 antagonism by LY255283 (5 mg/kg i.p.; 62%) reduced itch. Neither TRPV1‐knockout (TRPV1‐KO) nor TRPA1‐knockout (TRPA1‐KO mice exhibited LTB4‐induced itch compared with their wild‐type counterparts. The reactive oxygen species scavengers N‐acetylcysteine (NAC; 204 mg/kg i.p.; 86%) or superoxide dismutase (SOD; 10 mg/kg i.p.; 83%) also inhibited itch. LTB4‐induced superoxide release was attenuated by TCS 5861528 (56%) and HC‐030031 (66%), NAC (58%), SOD (50%), and LY255283 (59%) but not by the leukotriene B4 receptor 1 antagonist U‐75302 (9 nmol/site) or SB366791. Itch, superoxide, and myeloperoxidase generation were inhibited by the leukocyte migration inhibitor fucoidan (10 mg/kg i.v.) by 80, 61, and 34%, respectively. Myeloperoxidase activity was also reduced by SB366791 (35%) and SOD (28%). TRPV1‐KO mice showed impaired myeloperoxidase release, whereas TRPA1‐KO mice exhibited diminished production of superoxide. This result provides novel evidence that TRPA1 and TRPV1 contribute to itch via distinct mechanisms.—Fernandes, E. S., Vong, C. T., Quek, S., Cheong, J., Awal, S., Gentry, C., Aubdool, A. A., Liang, L., Bodkin, J.V., Bevan, S., Heads, R., Brain, S.D. Superoxide generation and leukocyte accumulation: key elements in the mediation of leukotriene B4‐induced itch by transient receptor potential ankyrin 1 and transient receptor potential vanilloid 1. FASEB J. 27, 1664–1673 (2013). www.fasebj.org

The sympathetic nervous system is controlled by transient receptor potential vanilloid 1 in the regulation of body temperature

Transient receptor potential vanilloid 1 (TRPV1) is involved in sensory nerve nociceptive signaling. Recently, it has been discovered that TRPV1 receptors also regulate basal body temperature in multiple species from mice to humans. In the present study, we investigated whether TRPV1 modulates basal sympathetic nervous system (SNS) activity. C57BL6/J wild‐type (WT) mice and TRPV1 knockout (KO) mice were implanted with radiotelemetry probes for measurement of core body temperature. AMG 9810∗∗∗ (50 mg/kg) or vehicle (2% DMSO/5% Tween 80/10 ml/kg saline) was injected intraperitoneally. Adrenoceptor antagonists or vehicle (5 ml/kg saline) was injected subcutaneously. In WT mice, the TRPV1 antagonist, AMG9810, caused significant hyperthermia, associated with increased noradrenaline concentrations in brown adipose tissue. The hyperthermia was significantly attenuated by the β‐adrenoceptor antagonist propranolol, the mixed α‐/β‐adrenoceptor antagonist labetalol, and the α1‐adrenoceptor antagonist prazosin. TRPV1 KO mice have a normal basal body temperature, indicative of developmental compensation. D‐Amphetamine (potent sympathomimetic) caused hyperthermia in WT mice, which was reduced in TRPV1 KO mice, suggesting a decreased sympathetic drive in KOs. This study provides new evidence that TRPV1 controls thermoregulation upstream of the SNS, providing a potential therapeutic target for sympathetic hyperactivity thermoregulatory disorders.—Alawi, K. M., Aubdool, A. A., Liang, L., Wilde, E., Vepa, A., Psefteli, M.‐P., Brain, S. D., Keeble, J. E. The sympathetic nervous system is controlled by transient receptor potential vanilloid 1 in the regulation of body temperature. FASEB J. 29, 4285‐4298 (2015). www.fasebj.org

Protection of Angiotensin II–Induced Vascular Hypertrophy in Vascular Smooth Muscle–Targeted Receptor Activity-Modifying Protein 2 Transgenic Mice

The vasodilator and vascular regulatory peptide adrenomedullin (AM), a member of the calcitonin gene-related peptide family of peptides, is predicted to play a pivotal protective role in cardiovascular dysfunction. The principle AM (AM1) receptor is composed of a G protein–linked calcitonin receptor-like receptor and a receptor activity-modifying protein (receptor activity-modifying protein 2). There is little knowledge of the receptors via which AM acts in diseases. Using smooth muscle-targeted receptor activity–modifying protein 2 transgenic mice with increased vascular density of functional AM1 receptors, we demonstrate that receptor activity-modifying protein 2 transgenic mice are not protected against angiotensin II–induced hypertension or cardiac hypertrophy. However, vascular hypertrophy, together with vascular cell adhesion molecule 1 and monocyte chemotactic protein 1 expression, is significantly reduced in the aortic walls of transgenic mice, as determined by histological techniques. This indicates that the AM1 vascular smooth muscle receptor can mediate local protection in vivo. This is supported by proliferation studies in cultured smooth muscle cells. By comparison, levels of hypotension and inflammation in a shock model were similar to those in wild-type mice. Thus, a role of the AM1 receptor in the vasoactive component could not be detected, and evidence is provided to show that the hypotensive response to AM is subject to desensitization in vivo. The finding that the vascular smooth muscle AM1 receptor acts at a local level to protect against hypertension-induced vascular hypertrophy and inflammation provides evidence that targeting this receptor may be a beneficial therapeutic approach.

Investigating the potential role of TRPA1 in locomotion and cardiovascular control during hypertension

Radiotelemetry was used to investigate the in vivo cardiovascular and activity phenotype of both TRPA1 (transient receptor potential ankyrin 1) wild‐type (WT) and TRPA1 knockout (KO) mice. After baseline recording, experimental hypertension was induced using angiotensin II infusion (1.1 mg−1 kg−1 a day, for 14 days). TRPA1 WT and KO mice showed similar morphological and functional cardiovascular parameters, including similar basal blood pressure (BP), heart rate, size, and function. Similar hypertension was also displayed in response to angiotensin II (156 ± 7 and 165 ± 11 mmHg, systolic BP ± SEM, n = 5–6). TRPA1 KO mice showed increased hypertensive hypertrophy (heart weight:tibia length: 7.3 ± 1.6 mg mm−1 vs. 8.8 ± 1.7 mg mm−1) and presented with blunted interleukin 6 (IL‐6) production compared with hypertensive WT mice (151 ± 24 vs. 89 ± 16 pg mL−1). TRPA1 expression in dorsal root ganglion (DRG) neurones was upregulated during hypertension (163% of baseline expression). Investigations utilizing the TRPA1 agonist cinnamaldehyde (CA) on mesenteric arterioles isolated from näive mice suggested a lack of TRPA1‐dependent vasoreactivity in this vascular bed; a site with notable ability to alter total peripheral resistance. However, mesenteric arterioles isolated from TRPA1 KO hypertensive mice displayed significantly reduced ability to relax in response to nitric oxide (NO) (P < 0.05). Unexpectedly, naïve TRPA1 KO mice also displayed physical hyperactivity traits at baseline, which was exacerbated during hypertension. In conclusion, our study provides a novel cardiovascular characterization of TRPA1 KO mice in a model of hypertension. Results suggest that TRPA1 has a limited role in global cardiovascular control, but we demonstrate an unexpected capacity for TRPA1 to regulate physical activity.

Vascular Actions of CGRP and Adrenomedullin: Mechanisms and Potential Contribution to Inflammation in the Cutaneous Microvasculature

Calcitonin gene-related peptide (CGRP) has a well known hypotensive effect on blood pressure primarily due to dilatation of small resistance vessels. This microvascular vasodilator effect can also lead to potentiation of inflammatory oedema formation and cell accumulation in the cutaneous microvasculature when applied or released locally. Moreover, an involvement of CGRP in hyperalgesia accompanying the inflammatory processes has been confirmed. Adrenomedullin (AM) is a member of calcitonin peptide family first isolated from pheochromocytoma cells. It is primarily produced by non-neural cell types. AM acts on CGRP receptors and also activates AM1 receptors, which CGRP does not. Both are G protein-linked receptors consisting of the same calcitonin receptor-like receptor in association with a different receptor activity modifying proteins. In this chapter we compare the actions of AM and CGRP on the microcirculation, on inflammatory oedema formation/cell accumulation and hyperalgesia.