title: Anti-RANKL antibodies decrease CGRP expression in dorsal root ganglion neurons innervating injured lumbar intervertebral discs in rats.
abstract: OBJECTIVE
  Nuclear factor-κB (NF-κB), receptor activator of NF-κB (RANK), and RANK ligand (RANKL) are transcriptional regulators of inflammatory cytokines. RANKL expression in dorsal root ganglion (DRG) neurons is elevated in animal models of pain or intervertebral disc herniation. We sought to evaluate the effect of anti-RANKL antibodies on sensory nerves innervating injured intervertebral discs.
  
  
  METHODS
  We labeled DRG neurons innervating L5-6 discs with FluoroGold (FG). The L5-6 discs of 36 rats were punctured using a 23-gage needle and 18 rats underwent sham surgery without disc puncture. The puncture group was evenly subdivided into a group in which 10 μl saline was administered to the injured disc and a group in which 10 μl of anti-RANKL antibody was administered. Seven and 14 days postsurgery, DRGs at L2 level were harvested, sectioned, and immunostained for calcitonin gene-related peptide (CGRP). The proportion of CGRP-immunoreactive (IR) DRG neurons of all FG-positive neurons was determined. Amount of tumor necrosis factor (TNF)-α and interleukin(IL)-6 was measured within the intervertebral discs in each group at 7 and 14 days after surgery using an enzyme-linked immunosorbent assay (ELISA).
  
  
  RESULTS
  The proportion of CGRP-IR DRG neurons to total FG-labeled neurons innervating injured intervertebral discs and amount of TNF-α and IL-6 in the injured discs in the saline control group was significantly increased compared with that found in rats from the sham surgery group (P < 0.05). However, application of anti-RANKL antibody to the injured discs significantly decreased the proportion of CGRP-IR DRG neurons to total FG-labeled neurons and amount of TNF-α and IL-6 in the injured discs (P < 0.05).
  
  
  CONCLUSIONS
  TNF-α and IL-6 in the injured discs increased and CGRP expression increased in DRG neurons innervating injured discs, and antibodies to RANKL could suppress this increased TNF-α, IL-6, and CGRP expression. RANKL may be a therapeutic target for pain control in patients with lumbar disc degeneration.
authors.affiliation: Department of Orthopaedic Surgery, Graduate School of Medicine, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba, 260-8670, Japan.
authors.forename: Masashi
authors.initials: M
authors.lastname: Sato
authors.forename: Kazuhide
authors.initials: K
authors.lastname: Inage
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Sakuma
authors.forename: Jun
authors.initials: J
authors.lastname: Sato
authors.forename: Sumihisa
authors.initials: S
authors.lastname: Orita
authors.forename: Kazuyo
authors.initials: K
authors.lastname: Yamauchi
authors.forename: Yawara
authors.initials: Y
authors.lastname: Eguchi
authors.forename: Nobuyasu
authors.initials: N
authors.lastname: Ochiai
authors.forename: Kazuki
authors.initials: K
authors.lastname: Kuniyoshi
authors.forename: Yasuchika
authors.initials: Y
authors.lastname: Aoki
authors.forename: Junichi
authors.initials: J
authors.lastname: Nakamura
authors.forename: Masayuki
authors.initials: M
authors.lastname: Miyagi
authors.forename: Miyako
authors.initials: M
authors.lastname: Suzuki
authors.forename: Gou
authors.initials: G
authors.lastname: Kubota
authors.forename: Takeshi
authors.initials: T
authors.lastname: Sainoh
authors.forename: Kazuki
authors.initials: K
authors.lastname: Fujimoto
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Shiga
authors.forename: Koki
authors.initials: K
authors.lastname: Abe
authors.forename: Hiroto
authors.initials: H
authors.lastname: Kanamoto
authors.forename: Gen
authors.initials: G
authors.lastname: Inoue
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Takahashi
authors.forename: Seiji
authors.initials: S
authors.lastname: Ohtori
mesh_terms: D000818:Animals; D000906:Antibodies / Q000494:pharmacology*; D015740:Calcitonin Gene-Related Peptide / Q000187:drug effects* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005456:Fluorescent Dyes; D005727:Ganglia, Spinal / Q000187:drug effects* / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D007403:Intervertebral Disc / Q000187:drug effects* / Q000294:innervation / Q000378:metabolism; D055959:Intervertebral Disc Degeneration / Q000378:metabolism; D008159:Lumbar Vertebrae; D008297:Male; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010146:Pain / Q000378:metabolism; D053245:RANK Ligand / Q000276:immunology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013266:Stilbamidines; D014409:Tumor Necrosis Factor-alpha / Q000187:drug effects / Q000378:metabolism
chemical_list: C049774:2-hydroxy-4,4'-diamidinostilbene, methanesulfonate salt; D000906:Antibodies; D005456:Fluorescent Dyes; D015850:Interleukin-6; D053245:RANK Ligand; D013266:Stilbamidines; D014409:Tumor Necrosis Factor-alpha; D015740:Calcitonin Gene-Related Peptide