Evaluation of pain markers expression, nerve fibers ingrowth and establishment of an in vitro model of neurites growth in degenerated intervertebral disc (IVD)

Abstract

Low back pain due to the degeneration of the intervertebral disc (IVD) is the most common cause of disability worldwide. Pain is the main debilitating symptom of degenerated IVD, mainly caused by the ingrowth of nociceptive nerve fibres into the deeper layer of the IVD. The production of pain-related neuropeptides by IVD cells is also responsible for the triggering of nociceptive stimuli involved in pain perception. Therefore, it is crucial to investigate the nerve ingrowth processes and the expression of these pain-related markers in degenerative disc disease to determine new strategies for pain treatment. Indeed, it has been shown how the cyclooxygenase-2 (COX-2) inhibitor, celecoxib (CXB) is able to halt degeneration and address IVD-related pain via intradiscal delivery, reducing systemic side effects. In the present study, the presence of nerve fibres and the expression of the neurotrophin NGF in the degenerated IVD were determined in rat and canine models. In addition, the pain-related markers expression was evaluated in vitro in human nucleus pulposus (NP) and annulus fibrosus (AF) cell cultures, upon stimulation with TNFα. Overall, pain marker expression was increased upon proinflammatory stimuli. However after CXB treatment, their expression was increased in a dose-dependent manner, in contrast to what is observed in vivo, suggesting the presence and influence of other factors which play a crucial role in the therapeutical effect of CXB in vivo. Moreover, an in vitro model was established using mouse dorsal root ganglion (DRG) explants cultured in conditioned media of human inflamed IVD cells to evaluate the effects of the latter on neurites growth. Surprisingly, inflamed conditioned media seems to not have any effect on neurites outgrowth, as well as for the CXB-treated one. However, both conditioned media seemed to have a protective effect against neural cell death, resulting in higher number of living neurons. Certainly, follow-up studies are necessary to determine the mechanisms behind pain in the degeneration of intravertebral discs, including neurites ingrowth and neuropeptide related to pain.