title: Matrix metalloprotease inhibitors suppress initiation and progression of chondrogenic differentiation of mesenchymal stromal cells in vitro.
abstract: Mesenchymal stromal cells (MSC) are an attractive source for cell therapy and tissue engineering of joint cartilage. Common chondrogenic in vitro protocols, however, induce hypertrophic markers like COL10A1, matrix metalloproteinase 13 (MMP13), and alkaline phosphatase (ALP) reminiscent of endochondral bone formation. To direct MSC toward articular chondrocytes more specifically, a better understanding of the regulatory steps is desirable. Proteases are important players in matrix remodeling, display inhibitory effects on growth plate development and MMP13 inhibition prevented hypertrophy of bovine chondrocytes. The aim of this study was to evaluate whether the activity of proteases and MMPs, especially MMP13, is crucial for the transition of MSC toward mature chondrocytes and could allow to selectively influence aspects of early and late chondrogenic differentiation. Protease inhibitors were added during MSC chondrogenesis and stage-specific markers were assessed by histology, qPCR, and ALP quantification. Chondrogenesis was little affected by leupeptin, pepstatin, or aprotinin. In contrast, broad spectrum pan-MMP inhibitors dose dependently suppressed proteoglycan deposition, collagen type II and type X staining, ALP activity, and reduced SOX9 and COL2A1 expression. A selective MMP13 inhibitor allowed chondrogenesis and showed only weak effects on ALP activity. In conclusion, transition of MSC toward mature chondrocytes in vitro depended on molecules suppressed by pan-MMP inhibitors identifying chondrogenic differentiation of MSC as a sophistically regulated process in which catabolic enzymes are capable to directly influence cellular fate. In future therapeutic applications of diseased joints, the tested MMP13-specific inhibitor promises suppression of collagen type II degradation without imposing a risk to impair MSC-driven regeneration processes.
authors.affiliation: Division of Experimental Orthopaedics, Orthopaedic University Clinic Heidelberg, Heidelberg, Germany.
authors.forename: Helge
authors.initials: H
authors.lastname: Bertram
authors.forename: Stephane
authors.initials: S
authors.lastname: Boeuf
authors.forename: Jasper
authors.initials: J
authors.lastname: Wachters
authors.forename: Sandra
authors.initials: S
authors.lastname: Boehmer
authors.forename: Christian
authors.initials: C
authors.lastname: Heisel
authors.forename: Michael W
authors.initials: MW
authors.lastname: Hofmann
authors.forename: Dorothea
authors.initials: D
authors.lastname: Piecha
authors.forename: Wiltrud
authors.initials: W
authors.lastname: Richter