title: Inflammation induces irreversible biophysical changes in isolated nucleus pulposus cells.
abstract: Intervertebral disc degeneration is accompanied by elevated levels of inflammatory cytokines that have been implicated in disease etiology and matrix degradation. While the effects of inflammatory stimulation on disc cell metabolism have been well-studied, their effects on cell biophysical properties have not been investigated. The hypothesis of this study is that inflammatory stimulation alters the biomechanical properties of isolated disc cells and volume responses to step osmotic loading. Cells from the nucleus pulposus (NP) of bovine discs were isolated and treated with either lipopolysaccharide (LPS), an inflammatory ligand, or with the recombinant cytokine TNF-α for 24 hours. We measured cellular volume regulation responses to osmotic loading either immediately after stimulation or after a 1 week recovery period from the inflammatory stimuli. Cells from each group were tested under step osmotic loading and the transient volume-response was captured via time-lapse microscopy. Volume-responses were analyzed using mixture theory framework to investigate two biomechanical properties of the cell, the intracellular water content and the hydraulic permeability. Intracellular water content did not vary between treatment groups, but hydraulic permeability increased significantly with inflammatory treatment. In the 1 week recovery group, hydraulic permeability remained elevated relative to the untreated recovery control. Cell radius was also significantly increased both after 24 hours of treatment and after 1 week recovery. A significant linear correlation was observed between hydraulic permeability and cell radius in untreated cells at 24 hours and at 1-week recovery, though not in the inflammatory stimulated groups at either time point. This loss of correlation between cell size and hydraulic permeability suggests that regulation of volume change is disrupted irreversibly due to inflammatory stimulation. Inflammatory treated cells exhibited altered F-actin cytoskeleton expression relative to untreated cells. We also found a significant decrease in the expression of aquaporin-1, the predominant water channel in disc NP cells, with inflammatory stimulation. To our knowledge, this is the first study providing evidence that inflammatory stimulation directly alters the mechanobiology of NP cells. The cellular biophysical changes observed in this study are coincident with documented changes in the extracellular matrix induced by inflammation, and may be important in disease etiology.
authors.affiliation: Center for Autoimmune and Musculoskeletal Diseases, The Feinstein Institute for Medical Research, North Shore-LIJ Health System, Manhasset, New York, United States of America.
authors.forename: Robert
authors.initials: R
authors.lastname: Maidhof
authors.affiliation: Center for Autoimmune and Musculoskeletal Diseases, The Feinstein Institute for Medical Research, North Shore-LIJ Health System, Manhasset, New York, United States of America.
authors.forename: Timothy
authors.initials: T
authors.lastname: Jacobsen
authors.affiliation: Center for Autoimmune and Musculoskeletal Diseases, The Feinstein Institute for Medical Research, North Shore-LIJ Health System, Manhasset, New York, United States of America.
authors.forename: Angelos
authors.initials: A
authors.lastname: Papatheodorou
authors.affiliation: Center for Autoimmune and Musculoskeletal Diseases, The Feinstein Institute for Medical Research, North Shore-LIJ Health System, Manhasset, New York, United States of America; Hofstra-North Shore LIJ School of Medicine, Hempstead, New York, United States of America.
authors.forename: Nadeen O
authors.initials: NO
authors.lastname: Chahine
mesh_terms: D000818:Animals; D051398:Aquaporin 1 / Q000378:metabolism; D002417:Cattle; D002463:Cell Membrane Permeability; D048429:Cell Size; D002478:Cells, Cultured; D003239:Connective Tissue Cells / Q000276:immunology* / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D007249:Inflammation / Q000378:metabolism; D007403:Intervertebral Disc / Q000276:immunology / Q000473:pathology; D055959:Intervertebral Disc Degeneration / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D009997:Osmotic Pressure; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology
chemical_list: D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; D051398:Aquaporin 1