An In Vitro Model to Test the Influence of Immune Cell Secretome on Mesenchymal Stromal Cell Osteogenic Differentiation
Khokhani, Paree; Belluomo, Ruggero; Croes, Michiel; Gawlitta, Debby; Kruyt, Moyo C; Weinans, Harrie
(2022) Tissue engineering. Part C, Methods, volume 28, issue 8, pp. 420 - 430
(Article)
Abstract
Immune cells and their soluble factors have an important role in the bone healing process. Modulation of the immune response, therefore, offers a potential strategy to enhance bone formation. To investigate the influence of the immune system on osteogenesis, we developed and applied an in vitro model that incorporates both
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innate and adaptive immune cells. Human peripheral blood mononuclear cells (PBMCs) were isolated and cultured for 24 h and subsequently stimulated with immune-modulatory agents; C-class CpG oligodeoxynucleotide (CpG ODN C), polyinosinic acid-polycytidylic acid [Poly(I:C)], and lipopolysaccharide (LPS); all pathogen recognition receptor agonists, that target Toll-like receptors (TLRs) 9, 3, and 4, respectively. The conditioned medium (CM) obtained from PBMCs after 24 h was used to investigate its effects on the metabolic activity and osteogenic differentiation capacity of human bone marrow-derived mesenchymal stromal cells (MSCs). Conditioned media from unstimulated PBMCs did not affect the metabolic activity and osteogenic differentiation capacity of MSCs. The CM from CpG ODN C and LPS-stimulated PBMCs increased alkaline phosphatase activity (ALP) of MSCs by approximately threefold as compared with the unstimulated control, whereas Poly(I:C) CM did not enhance ALP activity of MSCs. Moreover, direct stimulation of MSCs with the immune-modulatory stimuli did not result in increased ALP. These results demonstrate that soluble factors present in CM from PBMCs stimulated with immune-modulatory factors enhance osteogenesis of MSCs. This in vitro model can serve as a tool in screening immune-modulatory stimulants from a broad variety of immune cells for (indirect) effects on osteogenesis and also to identify soluble factors from multiple immune cell types that may modulate bone healing.
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Keywords: Cell Differentiation, Cells, Cultured, Culture Media, Conditioned/pharmacology, Humans, Leukocytes, Mononuclear, Lipopolysaccharides/metabolism, Mesenchymal Stem Cells, Osteogenesis, Secretome, bone healing, immune cells, inflammation, osteogenesis, osteoimmunology, pathogen recognition receptors, Bioengineering, Biomedical Engineering, Medicine (miscellaneous), Journal Article
ISSN: 1937-3384
Publisher: Mary Ann Liebert Inc.
Note: Funding Information: This article is supported by PPS allowance from the Health∼Holland LSH-TKI (grant no.: LSHM18011) and the EU's H2020 research and innovation program under Marie S. Curie Cofund RESCUE (grant agreement no. 801540). Publisher Copyright: © 2022 Mary Ann Liebert, Inc., publishers.
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