Heme-induced ferroptosis promotes human lumbar disc degeneration analyzed by MALDI-TOF MS

Abstract BackgroundNeovasculogenesis is a characteristic of degenerative lumbar discs, which makes extruded tissues exposed to heme-iron cytotoxicity (increased oxidative stress by ferroptosis). However, the present analyses for neovascularization are very complicated, and its mechanism of action is rarely reported.MethodsMatrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed to analyze human degenerative lumbar discs. Then, clinical relevance was analyzed between MALDI-TOF MS results and Pfirrmann classification of degenerative discs. In order to explore the mechanism, a heme-induced ferroptosis effect was evaluated both at tissue and cell levels using high-resolution MALDI-TOF MS and molecular biology methods. ResultsThe spectra revealed that hemoglobin (Hb) and heme signals were highly increased, thus serving as biomarkers of vasculogenesis in degenerative tissues. Clinical relevance analysis demonstrated that the intensity of Hb and heme peaks was closely related to Pfirrmann classification of degenerative discs. Mechanically, increased heme catabolism and down-regulation of glutathione peroxidase 4 (GPX4) levels were detected in degenerative discs, reflecting a iron-dependent cell death or ferroptosis. Further, accuracy mass measurements confirmed that the levels of ferroptosis-related metabolites such as glutathione, arachidonic acid (AA), sphinganine, polyunsaturated fatty acid (PUFA), and tricarboxylic acid (TCA) cycle were significantly different between the degenerative and normal tissues, indicating the interior of degenerative tissues was a prooxidant environment. Moreover, the heme-induced ferroptosis was verified in human nucleus pulposus cells, and the underlying mechanism might be associated with the Notch pathway. ConclusionsThe neovascularization in degenerative discs may expose the tissues to high heme toxicity, which further induces the ferroptosis effect within the tissues and accelerates the degeneration progression of discs. This study is beneficial for the pathological mechanism in degenerative discs and facilitate the development of non-operative intervention for lumbar disc herniation (LDH).