DNA Hydroxymethylation by Ten-eleven Translocation Methylcytosine Dioxygenase 1 and 3 Regulates Nociceptive Sensitization in a Chronic Inflammatory Pain Model

Background: Ten-eleven translocation methylcytosine dioxygenase converts 5-methylcytosine in DNA to 5-hydroxymethylcytosine, which plays an important role in gene transcription. Although 5-hydroxymethylcytosine is enriched in mammalian neurons, its regulatory function in nociceptive information processing is unknown. Methods: The global levels of 5-hydroxymethylcytosine and ten-eleven translocation methylcytosine dioxygenase were measured in spinal cords in mice treated with complete Freund's adjuvant. Immunoblotting, immunohistochemistry, and behavioral tests were used to explore the downstream ten-eleven translocation methylcytosine dioxygenase-dependent signaling pathway. Results: Complete Freund's adjuvant-induced nociception increased the mean levels (+/- SD) of spinal 5-hydroxymethylcytosine (178 +/- 34 vs. 100 +/- 21; P = 0.0019), ten-eleven translocation methylcytosine dioxygenase-1 (0.52 +/- 0.11 vs. 0.36 +/- 0.064; P = 0.0088), and ten-eleven translocation methylcytosine dioxygenase-3 (0.61 +/- 0.13 vs. 0.39 +/- 0.08; P = 0.0083) compared with levels in control mice (n = 6/group). The knockdown of ten-eleven translocation methylcytosine dioxygenase-1 or ten-eleven translocation methylcytosine dioxygenase-3 alleviated thermal hyperalgesia and mechanical allodynia, whereas overexpression cytosinethem in naive mice (n = 6/ group). Down-regulation of spinal ten-eleven translocation methylcytosine dioxygenase-1 and ten-eleven translocation methylcytosine dioxygenase-3 also reversed the increases in Fos expression (123 +/- 26 vs. 294 +/- 6; P = 0.0031; and 140 +/- 21 vs. 294 +/- 60; P = 0.0043, respectively; n = 6/ group), 5-hydroxymethylcytosine levels in the Stat3 promoter (75 +/- 16.1 vs. 156 +/- 28.9; P = 0.0043; and 91 +/- 19.1 vs. 156 +/- 28.9; P = 0.0066, respectively; n = 5/group), and consequent Stat3 expression (93 +/- 19.6 vs. 137 +/- 27.5; P = 0.035; and 72 +/- 15.2 vs. 137 +/- 27.5; P = 0.0028, respectively; n = 5/group) in complete Freund's adjuvant-treated mice. Conclusions: This study reveals a novel epigenetic mechanism for ten-eleven translocation methylcytosine dioxygenase-1 and ten-eleven translocation methylcytosine dioxygenase-3 in the modulation of spinal nociceptive information via targeting of Stat3.