Modulation of inflammatory pain in intervertebral disc degeneration: A therapeutic biomaterials approach
Mohd Isa, Isma Liza
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Discogenic low back pain (LBP) is a very common health problem that imposes a socio-economic burden on society. Primarily, intervertebral disc degeneration is one of the causative factors of LBP. It is mediated by an inflammation and an imbalance of extracelullar matrix homeostasis that can induce hyper-innervation and sensory sensitization in the disc and results in discogenic pain. The use of hyaluronic acid (HA) as a therapeutic agents offers promising results in tissue regeneration as it can reduce inflammation and pain, but the effects of HA therapy on glycosylation and pain associated with disc degeneration have not previously been determined. The overall aim of this study was to develop an optimally cross-linked high molecular weight of HA hydrogel, and further informed as to its therapeutic efficacy in alleviating inflammatory pain in degenerative disc disease. In phase I, the hydrogel was optimally crosslinked at 75 mM PEG, exerted hydrolytic stability and resistance to enzymatic degradation. No cytotoxic effect of NP cells was marked after treatment with HA hydrogels for one, three and seven days. IL-1β signalling molecules of IL-1R1 and MyD88 were significantly suppressed in the presence of HA hydrogel. Additionally, NGF and BDNF mRNA were down-regulated after treatment with cross-linked HA hydrogel. A possible protective mechanism of HA is shown by the binding of the CD44 receptor of NP cells to HA and which prevents NP cells from undergoing further inflammation. These findings indicate that optimally stabilized cross-linked HA hydrogel has a therapeutic effect in response to neurogenic inflammatory insult and becomes an ideal matrices hydrogel for NP repair. Nonetheless, the efficacy of HA hydrogel in alleviating pain in degenerative disc disease is still unknown. Therefore, a novel pre-clinical model of pain was established in phase II of the study as an intervention platform. The surgically puncture-induced disc injury was adopted in the rat tail. Here, for the first time, surgically puncture-induced disc injury evoked robust pain phenotype by exhibiting thermal hyperalgesia and mechanical allodynia close to the site of injury and up- regulation of spinal nociception marker, c-Fos mRNA. Conversely, the tail flick test revealed hypoalgesia at a lower level of the rat tail, suggesting a phenomenon of persistent pain as seen in clinical chronic pain. Moreover, disc injury inducedstructural disc degeneration and differently regulated glycosignature in both AF and NP tissues. Thus, the inhibition of nociceptive behaviour by low dose morphine, a current reversible analgesic that validates this pain model. Conclusively, the results indicate the first demonstration of nociception in the rat tail, anatomical disc degeneration and distinct glycosylation in AF and NP, thereby provide a platform to study pathogenesis of disc degeneration and testing of potential therapeutic target for disc repair in treating discogenic pain. Using this model, the potential efficacy of HA hydrogel in alleviating inflammatory pain in disc degeneration was evaluated in phase III of the study. HA hydrogel has been seen to alleviate nociceptive behaviour by reducing thermal hyperalgesia and mechanical allodynia close to the site of injury in the rat tail. Using the tail flick test, HA hydrogel demonstrated an inhibition of hypoalgesia at a lower level of the rat tail. These results indicate an anti-nociceptive effect of HA hydrogel that is comparable to low dose morphine treatment. The HA hydrogel also down- regulated spinal nociception markers, c-Fos and Tac1 (precusor for substance P), and pro-nociceptor of TRPV1 and Trk-A in AF and NP. An inhibition of sensory hyper- innervation was indicated by GAP43 and CGRP protein. Furthermore, HA altered glycosylation pattern than in injured AF and NP tissues. The mechanism of action of HA was determined by modulating key inflammatory pathways of IL-6 and IL-1β, and regulatory signalling pathways of Smad3 via TGF-β1 to respectively attenuate inflammation and regulate matrix components. Therefore, these results represent inflammatory pain modulation by HA hydrogel through alteration in glycomic and protein regulatory pathways, and thus suggesting that HA hydrogel as a promising clinical candidate for treatment of back pain caused by degenerated discs.