Abstract
Transient receptor potential (TRP) channels have emerged as potential sensors and transducers of inflammatory pain. The aims of this study were to investigate (1) the expression of TRP channels in intervertebral disc (IVD) cells in normal and inflammatory conditions and (2) the function of Transient receptor potential ankyrin 1 (TRPA1) and Transient receptor potential vanilloid 1 (TRPV1) in IVD inflammation and matrix homeostasis. RT-qPCR was used to analyze human fetal, healthy, and degenerated IVD tissues for the gene expression of TRPA1 and TRPV1. The primary IVD cell cultures were stimulated with either interleukin-1 beta (IL-1β) or tumor necrosis factor alpha (TNF-α) alone or in combination with TRPA1/V1 agonist allyl isothiocyanate (AITC, 3 and 10 µM), followed by analysis of calcium flux and the expression of inflammation mediators (RT-qPCR/ELISA) and matrix constituents (RT-qPCR). The matrix structure and composition in caudal motion segments from TRPA1 and TRPV1 wild-type (WT) and knock-out (KO) mice was visualized by FAST staining. Gene expression of other TRP channels (A1, C1, C3, C6, V1, V2, V4, V6, M2, M7, M8) was also tested in cytokine-treated cells. TRPA1 was expressed in fetal IVD cells, 20% of degenerated IVDs, but not in healthy mature IVDs. TRPA1 expression was not detectable in untreated cells and it increased upon cytokine treatment, while TRPV1 was expressed and concomitantly reduced. In inflamed IVD cells, 10 µM AITC activated calcium flux, induced gene expression of IL-8, and reduced disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and collagen 1A1, possibly via upregulated TRPA1. TRPA1 KO in mice was associated with signs of degeneration in the nucleus pulposus and the vertebral growth plate, whereas TRPV1 KO did not show profound changes. Cytokine treatment also affected the gene expression of TRPV2 (increase), TRPV4 (increase), and TRPC6 (decrease). TRPA1 might be expressed in developing IVD, downregulated during its maturation, and upregulated again in degenerative disc disease, participating in matrix homeostasis. However, follow-up studies with larger sample sizes are needed to fully elucidate the role of TRPA1 and other TRP channels in degenerative disc disease.
Highlights
Low back pain (LBP) is the leading cause of disability, activity limitation and lost productivity throughout the world today, with approximately 80% of all people suffering from back pain at least once in their life [1]
It is known that Transient receptor potential ankyrin 1 (TRPA1) can associate with Transient receptor potential vanilloid 1 (TRPV1), thereby regulating its intrinsic properties independently of intracellular calcium
Our study provided evidence that IL-1β itself can regulate gene expression of TRPV4 in intervertebral disc (IVD) cells., the Gene expression of TRPC6 was shown to be reduced in IVD cells under microgravity [18], but elevated in IVDs with increasing degeneration grade [19]
Summary
Low back pain (LBP) is the leading cause of disability, activity limitation and lost productivity throughout the world today, with approximately 80% of all people suffering from back pain at least once in their life [1]. Pro-inflammatory cytokines IL-1β and TNF-α directly act as nociceptive triggers, and activate the expression of other potentially nociceptive molecules including neuropeptides, interleukin-6 (IL-6), and interleukin-8 (IL-8) [4,5]. TRPV1 is a non-selective calcium channel, the expression and activity of which increases after inflammatory stimulation in dorsal root ganglions (DRGs), possibly causing chronic hyperalgesia. TRPA1 is a calcium permeable non-selective cation channel that is widely expressed in sensory neurons and in non-neuronal cells, including chondrocytes [11]. The aims of this study were to investigate the (1) expression of TRP channels in IVD cells in normal and inflammatory conditions and (2) the function of TRPA1 and TRPV1 in disc inflammation and matrix homeostasis
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