Mesenchymal Stromal Cell-Seeded Biomimetic Scaffolds as a Factory of Soluble RANKL in Rankl-Deficient Osteopetrosis

Abstract

Biomimetic scaffolds are extremely versatile in terms of chemical composition and physical properties, which can be defined to accomplish specific applications. One property that can be added is the production/release of bioactive soluble factors, either directly from the biomaterial, or from cells embedded within the biomaterial. We reasoned that pursuing this strategy would be appropriate to setup a cell-based therapy for RANKL-deficient Autosomal Recessive Osteopetrosis, a very rare skeletal genetic disease in which lack of the essential osteoclastogenic factor RANKL impedes osteoclast formation. The exogenously administered RANKL cytokine is effective in achieving osteoclast formation and function in vitro and in vivo, thus, we produced murine Rankl MSCs overexpressing human soluble RANKL (hsRL) following lentiviral transduction (LVhsRL). Here, we described a three-dimensional (3D) culture system based on a Magnesium-doped hydroxyapatite/collagen I (MgHA/Col) biocompatible scaffold closely reproducing bone physicochemical properties. MgHA/Col-seeded murine MSCs showed improved properties, as compared to two-dimensional (2D) culture, in terms of proliferation and hsRL production, with respect to LVhsRL-transduced cells. When implanted subcutaneously in Rankl mice, these cell constructs were well tolerated, colonized by host cells, and intensely vascularized. Of note, in the bone of Rankl mice that carried scaffolds with either WT or LVhsRL-transduced Rankl MSCs, we specifically observed formation of TRAP cells, likely due to sRL released from the scaffolds into circulation. Thus, our strategy proved to have the potential to elicit an effect on the bone; further work is required to maximize these benefits and achieve improvements of the skeletal pathology in the treated Rankl mice. Stem Cells Translational Medicine 2018.

Absence of Dipeptidyl Peptidase 3 Increases Oxidative Stress and Causes Bone Loss

“…In vitro osteoclastogenesis was induced following standard protocols, as reported . Briefly, 4 × 10 5 splenocytes/well were cultured in 96‐well plates in α‐MEM medium (Sigma‐Aldrich) supplemented with 10% FBS, 1% penicillin/streptomycin (P/S), and 1% glutamine in the presence of 25 ng/mL murine M‐CSF, 5 ng/mL human TGFβ1 (both from Peprotech, London, UK), 1 μM dexamethasone (Sigma‐Aldrich), and with or without 100 ng/mL RANKL for 6 days.…”

Mesenchymal Stromal Cells Anno 2019: Dawn of the Therapeutic Era? Concise Review

“…The secretome of MSCs also includes extracellular vesicles, which contain proteins and μRNAs that control target cell function. There are multiple examples of therapeutic effects of MSCs in preclinical models that are attributed to the MSC secretome, such as for instance the inhibition of colitis via the release of tumor necrosis factor‐induced protein 6 by intraperitoneal administered MSCs or via the release of the osteoclastogenic factor Receptor Activator of Nuclear factor Kappa‐B Ligand by MSCs seeded on biomimetic scaffolds for the treatment of osteopetrosis . Other studies have demonstrated that the effects of MSCs can be mimicked by infusion of MSC conditioned culture medium .…”

Expanded circulating hematopoietic stem/progenitor cells as novel cell source for the treatment of TCIRG1 osteopetrosis

“…Osteoclasts differentiated on bone slices were stained using alendronate conjugated to Alexa Fluor 488 24 , TRITC-conjugated phalloidin (Sigma-Aldrich) and TO-PRO-3 (Thermo Fisher Scientific). To evaluate bone resorption, the same bone slices were used for toluidine blue staining as previously described 25 . TRAP and toluidine blue images were acquired on a Zeiss AxioImager…”