TRPC6-Mediated Ca2+ Influx Activates MAPK and NF κB Signaling and Elicits Pro-Inflammatory and Catabolic Responses in Human Intervertebral Disc Cells

Intervertebral disc degeneration is characterized by inflammation, extracellular matrix breakdown, and neurovascular ingrowth, processes that contribute to discogenic, chronic back pain. The transient receptor potential canonical 6 (TRPC6) channel is a calcium-permeable ion channel implicated in inflammation and pain signaling in multiple tissues; however, its functional role in human disc cells remain unknown. Here, we investigated the expression, activation, and downstream consequences of TRPC6 activation using Hyp9, a pharmacological activator of TRPC6. TRPC6 transcripts were consistently detected across all donors examined (n = 17). Functional TRPC6 activation induced a rapid, dose-dependent calcium (Ca2+) influx across 0.5-100 {micro}M Hyp9. TRPC6 activation did not reduce metabolic activity or increase cytotoxicity at concentrations commonly used for in vitro TRPC6 activation. Mechanistically, TRPC6 activation induced mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-{kappa}B) pathways, as demonstrated by increased phosphorylation of p38 and extracellular signal-regulated kinase (ERK), degradation of the inhibitor of {kappa}B-alpha (I{kappa}B-), and increased nuclear translocation of the NF-{kappa}B p65 subunit. Downstream of these early signaling events, TRPC6 activation elicited a robust inflammatory and catabolic response with upregulation of IL-6, IL-8, COX-2, MMP-1, MMP-3, NGF, and VEGF, with corresponding increases in protein secretion. These findings identify TRPC6 as an important signaling node linking calcium influx to inflammatory, catabolic, and neuro- and angiogenesis-associated pathways in disc cells, highlighting TRPC6 as a potential therapeutic target in degenerative disc disease. HighlightsO_ST_ABSWhat are the main findings?C_ST_ABSO_LITRPC6 is endogenously expressed in human intervertebral disc cells, and its activation induces rapid calcium influx that initiates MAPK and NF-{kappa}B signaling pathways. C_LIO_LITRPC6 activation initiates a broad inflammatory and degenerative program, elevating the expression of IL-6, IL-8, COX-2, MMP-1, MMP-3, NGF, and VEGF. C_LI What are the implications of the main findings?O_LITRPC6 functions as a key upstream regulator linking calcium influx with inflammatory, matrix-degrading, and neuro-angiogenic processes central to disc degeneration and discogenic back pain. C_LIO_LIPharmacological targeting of TRPC6 may offer a novel therapeutic approach to suppress early inflammatory signaling, limit extracellular matrix breakdown, and reduce neurovascular ingrowth in degenerative disc disease. C_LI