Oklahoma Medical Research Foundation Oklahoma City, OK, United States
Biji T Kurien1, Pharaoh Gavin2, Joshua Cavett1, Valerie Lewis3, Ambre Chambers1, Bhaskaran Shylesh4, Anjum Juvaria1, Brittany Karfonta1, Lida Radfar1, Astrid Rasmussen4, Christopher Lessard4, Darise Farris4, Kathy Sivils5, Kristi A Koelsch4, Holly Van Remmen4 and R. Hal Scofield1, 1University of Oklahoma Health Sciences Center, Oklahoma City, OK, 2University of Washington, Seattle, WA, 3Veterans Affairs Medical Center, Oklahoma City, OK, 4Oklahoma Medical Research Foundation, Oklahoma City, OK, 5Janssen Research and Development, LLC, Spring House, PA
Background/Purpose: Sjögren's disease (SjD) is a chronic inflammatory, autoimmune disorder with reduced lacrimal/salivary gland secretion resulting in keratoconjunctivitis sicca and xerostomia, respectively. Also, SjD patients have increased rates of metabolic syndrome and experience significant fatigue. Metabolic syndrome and fatigue are associated with chronic low-level inflammation. Our previous studies showed oxidative damage in systemic lupus erythematosus and oxidative modification of proteins in SjD. Oxidative damage, mitochondrial dysfunction, and metabolic syndrome have been studied only sporadically in SjD, and their associations have not been studied. We hypothesized that SjD subjects have mitochondrial dysfunction and that fatigue and oxidative damage will be present in a subset of subjects with SjD.
Methods: We enrolled 17 SjD subjects and seven age and sex-matched subjects at the Oklahoma Shared Clinical and Translational Resources in this study. Subjects underwent a fasting blood draw, lipid and glucose testing, BMI measurement, and completed a fatigue questionnaire. After isolating PBMCs from the blood, B cells were purified through positive selection, and T cells through negative selection using Miltenyi Biotech B and T cell isolation kit. We plated one million T cells/well on Cell Tak coated plate to make the cells adherent. The cells were analyzed for mitochondrial oxygen consumption rate (OCR) and extracellular acidification rate (glycolysis) using the Seahorse XF24 assay. SDS-PAGE analysis of plasma proteins followed by immunoblotting of 4-hydroxynonenal (HNE) modified proteins using anti-HNE antibody were also performed.
Results: Our previous studies confirmed oxidative damage in SjD, demonstrated by an HNE-modified plasma protein migrating at ~16 kD. In the present study, we did not find significant differences in non-mitochondrial respiration between controls and SjD [17.67±2.77 Standard Error (SE) Vs. 17.95±3.1 SE pmol/min/million cells]. However, basal OCR was significantly higher (p< 0.0001) in controls compared to SjD (114.94±16.13 SE Vs. 70±5.69 SE pmol/min/million cells). ATP-linked respiration was also significantly higher (p< 0.001) in controls than in SjD (116.07±16.79 SE Vs. 66.93±4.71 SE pmol/min/million cells). The biggest differences were in the maximal respiration (p< 0.0005) (controls- 421.3±88.35 SE; SjD- 173.97±17.21 SE pmol/min/million cells) and reserve capacity (p< 0.0002) (controls- 310.93±74.37 SE; SjD- 105.01±12.34 SE pmol/min/million cells). There was no significant difference in basal glycolysis or glycolytic stressed levels in controls and SjD. Our next steps will be to determine which of these SjD subjects have metabolic syndrome and oxidative damage and whether mitochondrial dysfunction, metabolic syndrome and oxidative damage occur in a sub-set of these SjD subjects.
Conclusion: Mitochondrial dysfunction appears to be a significant problem in SjD. We will analyze if it is associated with the metabolic syndrome and fatigue that frequently accompany SjD.
Disclosures: B. Kurien, None; P. Gavin, None; J. Cavett, None; V. Lewis, None; A. Chambers, None; B. Shylesh, None; A. Juvaria, None; B. Karfonta, None; L. Radfar, None; A. Rasmussen, None; C. Lessard, None; D. Farris, None; K. Sivils, Janssen Research & Development, LLC; K. Koelsch, None; H. Van Remmen, None; R. Scofield, None.
