Discussion
The lack of metformin-mediated suppression of inflammatory response in the nonstretch groups indicates that metformin may be enacting its effects through a stretch-dependent manner. These results suggest a foundation for pursuing further research into metformin's potential role as an anti-inflammatory agent for curtailing intervertebral disc degeneration.
Methods
Five Fischer 344 rats were sacrificed and their spines isolated. AF cells were cultured and plated in flexible silicone membrane-based six-well plates. Wells were split into eight groups and subjected to metformin, IL-1β, mechanical stretch, and combined treatments. Relative gene expressions of MMP-13, COX-2, iNOS, AGC, and Col1 were assessed with quantitative real-time polymerase chain reaction (qRT-PCR), and downstream prostaglandin E2 (PGE2) production was quantified with enzyme-linked immunosorbent assay (ELISA). NF-kB nuclear translocation was also quantified.
Results
Metformin in the presence of the combined stress treatments (M + IL/S) significantly increased Col1, COX-2, and MMP-13 gene expression, decreased PGE2 production compared to IL/S conditions alone. Metformin treatment of cultured rat annulus fibrosus cells significantly reduced the nuclear translocation of NF-κB after 4 h of IL-1β treatment from 43.1% in case of IL-1β treatment down to 26.2% in the case of metformin + IL-1β treatment.