Electroacupuncture intervention of visceral hypersensitivity is involved in PAR-2-activation and CGRP-release in the spinal cord

Abstract

Electroacupuncture (EA) relieves visceral hypersensitivity (VH) with underlying inflammatory bowel diseases. However, the mechanism by which EA treats ileitis-induced VH is not clearly known. To assess the effects of EA on ileitis-induced VH and confirm whether EA attenuates VH through spinal PAR-2 activation and CGRP release, goats received an injection of 2,4,6-trinitro-benzenesulfonic-acid (TNBS) solution into the ileal wall. TNBS-injected goats were allocated into VH, Sham acupuncture (Sham-A) and EA groups, while goats treated with saline instead of TNBS solution were used as the control. Goats in EA group received EA at bilateral Hou-San-Li acupoints for 0.5 h at 7 days and thereafter repeated every 3 days for 6 times. Goats in the Sham-A group were inserted with needles for 0.5 h at the aforementioned acupoints without any hand manipulation and electric stimulation. Visceromotor responses to colorectal distension, an indicator of VH, were recorded by electromyography. The terminal ileum and thoracic spinal cord (T 11) were sampled for evaluating ileitis at days 7 and 22, and distribution and expression-levels of PAR-2, CGRP and c-Fos on day 22. TNBStreated-goats exhibited apparent transmural-ileitis on day 7, microscopically low-grade ileitis on day 22 and VH at days 7-22. Goats of Sham-A, VH or EA group showed higher (P < 0.01) VH at days 7-22 than the Control-goats. EA-treated goats exhibited lower (P < 0.01) VH as compared with Sham-A or VH group. Immunoreactive-cells and expression-levels of spinal PAR-2, CGRP and c-Fos in the EA group were greater (P < 0.01) than those in the Control group, but less (P < 0.01) than those in Sham-A and VH groups on day 22. Downregulation of spinal PAR-2 and CGRP levels by EA attenuates the ileitis and resultant VH. Inflammatory bowel diseases (IBDs) are chronic, immune-mediated and remittent-relapsing gastrointestinal inflammation. Crohn's disease (CD) and ulcerative colitis (UC) are the predominant clinical types of IBDs and their resultant symptoms include diarrhea, nausea, weight loss, abdominal pain and visceral hypersensitivity (VH) 1,2. There are two-thirds of IBD patients suffering from VH during their acute flares and remission 3,4. The persistent nature of VH impairs the patient's quality of life and consumes huge medical resources throughout the world. The environmental stress, inflammation and traumatic injuries trigger the development of VH. However, its underlying mechanism is not completely understood. A G-protein coupled receptor, PAR-2, is activated due to proteolytic cleavage of trypsin, tryptase and cathepsin-S 5-7. Over the years, studies have reported that the PAR-2 activation on resident mast cells, neutrophils or macrophages initiates for the release of inflammatory mediators, i.e., cytokines, chemokines and prostaglandins, which triggers the production of neurogenic mediators, i.e., substance-P (SP) and calcitonin gene-related peptide (CGRP) from the enteric neurons as well as afferent nerves to elicit eventually the VH 8-10. Cenac et al. 11 infu...

Mast Cells Mediate Inflammatory Injury and Aggravate Neurological Impairment in Experimental Subarachnoid Hemorrhage Through Microglial PAR-2 Pathway

“…PAR-2, also known as GPR11 or F2RL1, belongs to the 7-transmembrane receptors that couple to guanosine-nucleotide-binding proteins family that are encoded by F2RL1 gene ( Kawaguchi et al, 2020 ). PAR-2 is widely expressed in mammals, especially in epidermal keratinocytes and immune cells, including neutrophils, dendritic cells, T cells, and macrophages ( Sutherland et al, 2019 ; Shah et al, 2020 ). Upon its extracellular amino terminus between arginine and serine was cleaved by protease, PAR-2 is activated and further modulate multiple biological processes such as metabolism, tumorigenesis, and inflammatory response ( Kim et al, 2020 ; Wang et al, 2020 ; Bang et al, 2021 ; Jiang et al, 2021 ).…”

Peripheral and Spinal Mechanisms Involved in Electro-Acupuncture Therapy for Visceral Hypersensitivity

“…Different cytokines like tumor necrosis factor-alpha (TNF-α), interlukin-1β (IL-1β), and interlukin-6 (IL-6) and disease activity index, histopathological changes scour, and myeloperoxidase (MPO) enzyme expression increase in the inflamed intestine and have an important role in VH ( Adam et al, 2013 ; Shah et al, 2016 ; Tahir et al, 2016 ). After acupuncture treatment in the 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis or ileitis model, the level of TNF-α, IL-1β, and IL-6 and disease activity index, microscopic changes scour, and MPO level were decreased and caused to attenuate VH, and weight gain was improved ( Tian et al, 2003 ; Kim et al, 2017 ; Shah et al, 2020 ).…”

“…A previous study showed that protease-activated receptor-2 (PAR-2) activation in different cells like granulocytes and certain agranulocytic white blood cells could induce the release of different inflammatory mediators, i.e., chemokines and prostaglandins, which promoted the production of neurogenic mediators, i.e., substance P and calcitonin gene-related peptide (CGRP), from the enteric nerve plexus and afferent nerves to ultimately trigger the VH ( Cenac et al, 2003 ; Suckow and Caudle, 2009 ). Fecal supernatant of IBS patients was injected into the colon of healthy mice that resulted in higher mucosal inflammation and increased intestinal permeability, hence contributing to VH through a PAR-2 activation mechanism ( Cenac et al, 2007 ; Annaházi et al, 2009 ; Shah et al, 2020 ). The respective findings have shown that activated PAR-2 in the intestine enhances VH.…”

“…Therefore, c-Fos is providing the active site for evaluating the active substances or treatment in the CNS. Researchers have also established that EA reduced peripheral inflammation and c-Fos expression in the spinal cord ( Xu et al, 2010 ; Qi and Li, 2012 ; Shah et al, 2020 ). The c-Fos expression level increased in the SCDH superficial lamina during VH with TNBS-induced ileitis but decreased after EA therapy ( Shah et al, 2020 ).…”

Possible implications of animal models for the assessment of visceral pain

“…Wan et al 11 used an ileitis‐provoked visceral hypersensitivity (VH) model to confirm the effects of electroacupuncture on VH through Janus kinase 2 (JAK2)/signal transducers and activators of the transcription 3 (STAT3) signaling pathway in the periaqueductal gray‐rostral ventromedial medulla‐SDH axis. Very recently, Shah et al 12 used TNBS‐induced ileitis goats to study VH and reported that electroacupuncture attenuates VH through down‐regulation of spinal PAR‐2 and CGRP. The persistent pancreatitis pain induced by injection of dibutyltin dichloride in the tail vein, along with 10% alcohol in drinking water, has also been investigated in rats 13 .…”