Oral cancer induced TRPV1 sensitization is mediated by PAR2 signaling in primary afferent neurons innervating the cancer microenvironment

Abstract

Oral cancer patients report sensitivity to spicy foods and liquids. The mechanism responsible for chemosensitivity induced by oral cancer is not known. We simulate oral cancer-induced chemosensitivity in a xenograft oral cancer mouse model using two-bottle choice drinking and conditioned place aversion assays. An anatomic basis of chemosensitivity is shown in increased expression of TRPV1 in anatomically relevant trigeminal ganglion (TG) neurons in both the xenograft and a carcinogen (4-nitroquinoline 1-oxide)-induced oral cancer mouse models. The percent of retrograde labeled TG neurons that respond to TRPV1 agonist, capsaicin, is increased along with the magnitude of response as measured by calcium influx, in neurons from the cancer models. To address the possible mechanism of TRPV1 sensitivity in tongue afferents, we study the role of PAR2, which can sensitize the TRPV1 channel. We show co-expression of TRPV1 and PAR2 on tongue afferents and using a conditioned place aversion assay, demonstrate that PAR2 mediates oral cancer-induced, TRPV1-evoked sensitivity in an oral cancer mouse model. The findings provide insight into oral cancer-mediated chemosensitivity.

The impact of tumor immunogenicity on cancer pain phenotype using syngeneic oral cancer mouse models

“…For investigation into the syngeneic allograft transplant model, we chose to focus on three nociception-related proteins expressed on primary afferent sensory neurons, TRPV1, CGRP, and ATF3 and their associated genes, Trpv1, Calca, Atf3 . Cancer-induced increases in TRPV1 and CGRP protein expression in the TG neurons have been previously reported in association of nociception in both rat ( 47 ) and mouse ( 19 , 22 ) oral cancer models. We found, within the MOC1-and MOC2- models, that Calca gene expression and CGRP protein in tongue-innervating neurons are increased in primarily female mice, an effect which occurs along with the sex differences in nociceptive behavior.…”

“…However, given the low immunogenic quality of MOC2, the tumor growth and corresponding evoked nociceptive behavior and weight loss had a rapid onset, with tumor growth detectable at PID1 and significant changes in nociceptive behavior at PID8. Sex differences in oral cancer-evoked nociceptive behavior using mouse models ( 22 , 23 , 30 ) have also been previously reported but are still debated in the clinical literature ( 12 , 23 , 42 , 43 ). In the MOC1 model, despite similar tumor size over time, only tumor-bearing female mice demonstrated significant tumor-evoked nociceptive behavior over time compared to sex-matched shams.…”

“…Therefore, we also assessed relative gene expression in specifically tongue-innervating primary afferent neurons using retrograde labeling and single cell PCR. We have previously identified a role for the sensory neuropeptide, calcitonin gene-related peptide (CGRP), and chemosensitivity receptor, transient receptor vanilloid channel 1 (TRPV1), in oral cancer pain ( 19 , 22 ). Additionally, given the differences in immunogenicity and tumor growth in vivo , we sought to understand the potential contribution of nerve injury and subsequent neuropathic pain ( 9 ) for each model.…”

“…While our results are consistent with previous findings using the dolognawmeter assay, our report is limited by using only one behavioral assay. Additional nociceptive behaviors have been validated previously on animal models of oral cancer pain such as conditioned place preference ( 22 , 30 , 44 ) (CPP), facial von Frey ( 45 ), and meal analysis ( 44 , 46 ); however, sex differences in orofacial nociception have only been previously documented using the CPP ( 22 , 30 ).…”

“…Reverse transcription was performed with Quantitect Reverse Transcription Kit (Qiagen Inc.) according to the manufacturer's instructions. For single cell analysis, DiI-positive single neurons were dissociated as previously described ( 19 , 21 , 22 ), identified using fluorescence microscopy, and limited to ≤ 25 μm in diameter. Cells were then picked up using glass capillaries (World Precision Instruments) which were held by micromanipulator (Sutter Instruments), headstage (EPC10 HEKA) and electrode holder under brightfield optics.…”