M2 macrophages regulate nucleus pulposus cell extracellular matrix synthesis through the OPN-CD44 axis in intervertebral disc degeneration.

Objective: Macrophages play a crucial role in various physiological processes. In intervertebral disc degeneration (IDD), macrophage infiltration has been observed in human intervertebral disc (IVD) specimens, but how macrophages influence IDD remains unclear.

Methods: According to the single-cell transcriptome expression profiles from GSE165722, we verified the infiltration of macrophages in IDD and the possible interaction between infiltrated macrophages and nucleus pulposus cells (NPCs). The expression of macrophage-associated markers was verified in specimens of human nucleus pulposus, lumbar spinal instability mice and annulus fibrosus puncture mice. By treating NPCs cocultured with M2 macrophages with OPN neutralization antibody and siCD44, we demonstrated that both in vitro and in vivo macrophages regulated IDD through the OPN-CD44 axis. Using TGFβ1 and siCD44 treatment, we verified that CD44 regulated the pSMAD2/3 pathway.

Results: IDD engaged macrophage infiltration, mainly gathered in the endplate, and induced macrophage M2 polarization. Infiltrated macrophages showed high level expression of OPN, and NPCs showed upregulated CD44. Depletion of macrophages significantly decreased the expression of OPN and CD44 in degenerative IVD, concurrently exacerbating IDD. The co-culture of macrophages and NPCs in vitro demonstrated that the conditioned media from NPCs induced macrophage M2polarization. Further, M2 macrophages rescued NPCs extracellular matrix (ECM) phenotype through the OPN-CD44 axis, by regulating pSMAD2/3 nuclear translocation.

Conclusions: Our findings suggest that macrophages regulate NPC ECM expression in IDD through the OPN-CD44 axis, emphasizing the therapeutic potential of targeting macrophages and the OPN-CD44 axis for IDD prevention and treatment.