Differential Cytotoxicity of Corticosteroids on Human Mesenchymal Stem Cells

• Published in: Clinical orthopaedics and related research Vol. 473; no. 3; pp. 1155 - 1164
• Main Authors: Wyles, Cody C., Houdek, Matthew T., Wyles, Saranya P., Wagner, Eric R., Behfar, Atta, Sierra, Rafael J.
• Format: Journal Article
• Language: English
• Published: Boston Springer US 01.03.2015; Lippincott Williams & Wilkins Ovid Technologies
• Subjects: Adrenal Cortex Hormones - administration & dosage; Adrenal Cortex Hormones - toxicity; Adult; Anti-Inflammatory Agents - administration & dosage; Anti-Inflammatory Agents - toxicity; Basic Research; Cell Survival - drug effects; Chondrocytes - drug effects; Conservative Orthopedics; Dose-Response Relationship, Drug; Female; Humans; Male; Medicine; Medicine & Public Health; Mesenchymal Stromal Cells - drug effects; Middle Aged; Orthopedics; Sports Medicine; Surgery; Surgical Orthopedics; Average Optical Density; Avascular Necrosis; Triamcinolone; Betamethasone; Triamcinolone Acetonide
• ISSN: 0009-921X; 1528-1132
• DOI: 10.1007/s11999-014-3925-y

Background Corticosteroids are a common, short-term, local antiinflammatory and analgesic for treating patients with musculoskeletal disorders. Studies have shown the deleterious effects of corticosteroids on chondrocytes, suggesting a potentiation of degenerative joint disease. Mesenchymal stem cells (MSCs) are the direct progenitors of chondrocytes and other musculoskeletal tissue. Additionally, they serve an important antiinflammatory role, which can combat the chronic inflammatory state that mediates degenerative joint disease. Little is known about how corticosteroids interact with this regenerative and reparative cell population. Questions/purposes We asked: (1) Are corticosteroids cytotoxic to MSCs in a dose–response fashion? (2) Is there a differential effect in the level of cytotoxicity to MSCs between commercially available corticosteroid preparations? Methods Human MSCs were isolated and cultured from periarticular adipose tissue obtained from 20 patients undergoing primary THA. MSCs were exposed for 60 minutes to one of four commonly used corticosteroid preparations: betamethasone sodium phosphate-betamethasone acetate (6 mg/mL), dexamethasone sodium phosphate (4 mg/mL), methylprednisolone (40 mg/mL), or triamcinolone acetonide (40 mg/mL). Among the four preparations (treatment groups), cells were exposed to increasing concentrations of drugs according to the following titrations of the commercially available preparation: 0.0 (control solution of 1X phosphate buffered saline), 3.125, 6.25, 12.5, 25, 50, 75, and 100 % (undiluted commercial product). Cells were allowed to recover in standard culture media for 24 hours. After the recovery period, cell viability was measured using -(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) tetrazolium dye-based cellular viability assay and live-dead cell fluorescent staining. For the MTS assay, measurements were quantified in units of optical density (OD). ANOVA was performed at every experimental steroid concentration. When this global test was statistically significant, all pairwise comparisons were performed at that concentration with p values adjusted by the Tukey method to guard against Type I error. Results Exposure to corticosteroids decreased MSC viability in a curvilinear dose–response pattern. For betamethasone, the mean MTS OD at 0% steroid concentration was 1.03 (SD, 0.12) and decreased to 0.00 (SD, 0.00) at 25% steroid concentration. For dexamethasone, the mean MTS OD at 0% steroid concentration was 1.00 (SD, 0.07) and decreased to 0.00 (SD, 0.01) at 100% steroid concentration. For methylprednisolone, the mean MTS OD at 0% steroid concentration was 1.03 (SD, 0.09) and decreased to 0.00 (SD, 0.00) at 100% steroid concentration. For triamcinolone, the mean MTS OD at 0% steroid concentration was 1.02 (SD, 0.09) and decreased to 0.00 (SD, 0.00) at 75% steroid concentration. There were large differences among commercially available preparations, and these differences were present at every concentration. In general, dexamethasone was most gentle on MSCs (average OD by steroid concentration: 0% = 1.00; 3.125% = 0.86; 6.25% = 0.74; 12.5% = 0.53; 25% = 0.30; 50% = 0.20; 75% = 0.09; 100% = 0.00, triamcinolone and methylprednisolone were intermediate (triamcinolone average OD by steroid concentration: 0% = 1.02; 3.125% = 0.82; 6.25% = 0.64; 12.5% = 0.45; 25% = 0.18; 50% = 0.03; 75% = 0.00; 100% = 0.00; methylprednisolone average OD by steroid concentration: 0% = 1.03; 3.125% = 0.74; 6.25% = 0.54; 12.5% = 0.31; 25% = 0.12; 50% = 0.01; 75% = 0.00; 100% = 0.00), and betamethasone was most toxic (average OD by steroid concentration: 0% = 1.03; 3.125% = 0.74; 6.25% = 0.27; 12.5% = 0.02; 25% = 0.00; 50% = 0.00; 75% = 0.00; 100% = 0.00). ANOVA testing showed p values less than 0.0001 at every tested concentration (with the exception of the 0% control solution; p = 0.204) with subsequent pairwise comparisons supporting the relationships described above. The outcomes were maintained after stratifying by age, sex, or indication for THA (osteoarthritis versus avascular necrosis). Conclusions Commonly used intraarticular corticosteroids had a dose-dependent, profound, and differential effect on MSCs in this in vitro model, with betamethasone being the most toxic. Further studies are needed to assess if the in vitro effects of these agents translate into similar in vivo outcomes. Clinical Relevance Corticosteroids frequently are used by physicians to reduce inflammation in patients with musculoskeletal disorders, but these agents may hinder MSCs’ innate regenerative capacity in exchange for temporary analgesia. Our study suggests that choosing dexamethasone may result in less harmful effects when compared with other injectable steroids.