IL‐6 counteracts the inhibitory effect of IL‐4 on osteogenic differentiation of human adipose stem cells

Abstract

Fracture repair is characterized by cytokine production and hypoxia. To better predict cytokine modulation of mesenchymal stem cell (MSC)‐aided bone healing, we investigated whether interleukin 4 (IL‐4), IL‐6, and their combination, affect osteogenic differentiation, vascular endothelial growth factor (VEGF) production, and/or mammalian target of rapamycin complex 1 (mTORC1) activation by MSCs under normoxia or hypoxia. Human adipose stem cells (hASCs) were cultured with IL‐4, IL‐6, or their combination for 3 days under normoxia (20% O 2) or hypoxia (1% O 2), followed by 11 days without cytokines under normoxia or hypoxia. Hypoxia did not alter IL‐4 or IL‐6‐modulated gene or protein expression by hASCs. IL‐4 alone decreased runt‐related transcription factor 2 (RUNX2) and collagen type 1 (COL1) gene expression, alkaline phosphatase (ALP) activity, and VEGF protein production by hASCs under normoxia and hypoxia, and decreased mineralization of hASCs under hypoxia. In contrast, IL‐6 increased mineralization of hASCs under normoxia, and enhanced RUNX2 gene expression under normoxia and hypoxia. Neither IL‐4 nor IL‐6 affected phosphorylation of the mTORC1 effector protein P70S6K. IL‐4 combined with IL‐6 diminished the inhibitory effect of IL‐4 on ALP activity, bone nodule formation, and VEGF production, and decreased RUNX2 and COL1 expression, similar to IL‐4 alone, under normoxia and hypoxia. In conclusion, IL‐4 alone, but not in combination with IL‐6, inhibits osteogenic differentiation and angiogenic stimulation potential of hASCs under normoxia and hypoxia, likely through pathways other than mTORC1. These results indicate that cytokines may differentially affect bone healing and regeneration when applied in isolation or in combination.

Interleukin‐4 overexpressing mesenchymal stem cells within gelatin‐based microribbon hydrogels enhance bone healing in a murine long bone critical‐size defect model

“…During maturation of mesenchymal‐osteoblast lineage cells, the cells express ALP, osterix, and collagen type I, important markers of osteogenesis (Wu, Scadden, & Kronenberg, 2009). IL‐4 has an inhibitory effect on ALP expression and retards bone mineralization (Bastidas‐Coral et al, 2016; Bastidas‐Coral et al, 2019; Riancho, Gonzalez‐Marcias, Amado, Olmos, & Fernandez‐Luna, 1995). Lewis et al reported that mice that overproduce IL‐4 had severe osteoporosis primarily due to a marked decrease in osteoblast and ALP activity (Lewis et al, 1993).…”

Glucocorticoids decreased Cx43 expression in osteonecrosis of femoral head: The effect on proliferation and osteogenic differentiation of rat BMSCs

“…The results showed that Cx43 expression was decreased significantly in the model group in a comparison with the control group, and also the expressions of Runx2, ALP and COLⅠ were decreased significantly in the model group. All these factors are known as the primary regulators or associated factors of the osteogenic differentiation process 43,44 . Osteocytes are mainly originated from BMSCs, where BMSCs firstly undergo lineage differentiation and become pre‐osteoblasts.…”

“…All these factors are known as the primary regulators or associated factors of the osteogenic differentiation process. 43,44 Osteocytes are mainly originated from BMSCs, where BMSCs firstly undergo lineage differentiation and become pre-osteoblasts. Glucocorticoid use could inhibit the differentiation and maturation of pre-osteoblasts through inhibiting Runx2 expression, which followed by a decrease of the expressions of ALP and COLⅠ.…”

Effect on Osteogenic Differentiation of Genetically Modified IL4 or PDGF-BB Over-Expressing and IL4-PDGF-BB Co-Over-Expressing Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro

“…We implanted IL-4 MSCs within μRB scaffolds on the third day after the initial surgical defect was created so as not to interfere with the acute inflammatory phase, which lasts about 3 days and is essential for fracture healing in mice [ 44 ]; this resulted in enhanced bone formation in a murine critical-size bone defect model [ 45 ]. There are also several reports that local delivery of IL4 acutely inhibits osteogenic differentiation of MSCs [ 18 , 46 ]. In addition, we have reported that the injection of IL4-MSCs significantly decreased the number of empty lacunae compared to MSCs in a model of corticosteroid induced osteonecrosis but resulted in less osteogenesis in vivo [ 20 ].…”