Mesenchymal stem cells (MSC) have potent immunomodulatory and regenerative effects via soluble factors. One approach to improve stem cell-based therapies is encapsulation of MSC in hydrogels based on natural proteins such as collagen and fibrin, which play critical roles in bone healing. In this work, we comparatively studied the influence of collagen and fibrin hydrogels of varying stiffness on the paracrine interactions established by MSC with macrophages and osteoblasts. Type I collagen and fibrin hydrogels in a similar stiffness range loaded with MSC from donants were prepared by modifying the protein concentration. Viability and morphology of MSC in hydrogels as well as cell migration rate from the matrices were determined. Paracrine actions of MSC in hydrogels were evaluated in co-cultures with human macrophages from healthy blood donors or with osteoblasts from bone explants of patients with osteonecrosis of the femoral head. Lower matrix stiffness resulted in higher MSC viability and migration. Cell migration rate from collagen hydrogels was higher than from fibrin matrices. The secretion of the immunomodulatory factors interleukin-6 (IL-6) and prostaglandin E2 (PGE2) by MSC in both collagen and fibrin hydrogels increased with increasing matrix stiffness. Tumor necrosis factor-α (TNF-α) secretion by macrophages cultured on collagen hydrogels was lower than on fibrin matrices. Interestingly, higher collagen matrix stiffness resulted in lower secreted TNF-α while the trend was opposite on fibrin hydrogels. In all cases, TNF-α levels were lower when macrophages were cultured on hydrogels containing MSC than on empty gels, an effect partially mediated by PGE2. Finally, mineralization capacity of osteoblasts co-cultured with MSC in hydrogels increased with increasing matrix stiffness, although this effect was more notably for collagen hydrogels. Paracrine interactions established by MSC in hydrogels with macrophages and osteoblasts are regulated by matrix composition and stiffness.
Mesenchymal stem cells (MSC) are attractive candidates for bone regeneration approaches. Benefits of MSC therapy are mainly attributed to paracrine effects via soluble factors, exerting both immunoregulatory and regenerative actions. Encapsulation of MSC in hydrogels prepared with extracellular matrix (ECM) proteins has been proposed as a strategy to enhance their survival and potentiate their function after implantation. Functional activity of MSC can be regulated by the physical and mechanical properties of their microenvironment. In this work, we investigated whether matrix stiffness can modulate the crosstalk between MSC encapsulated in collagen hydrogels with macrophages and osteoblasts. Collagen hydrogels with a final collagen concentration of 1.5, 3 and 6 mg/mL loaded with human MSC were prepared. Viscoelastic properties of hydrogels were measured in a controlled stress rheometer. Cell distribution into the hydrogels was examined using confocal microscopy and the levels of the immunomodulatory factors interleukin-6 (IL-6) and prostaglandin E2 (PGE2) released by MSC were quantified by immunoassays. To determine the effect of matrix stiffness on the immunomodulatory potential of MSC, human macrophages obtained from healthy blood were cultured in media conditioned by MSC in hydrogels. The involvement of IL-6 and PGE2 in MSC-mediated immunomodulation was investigated employing neutralizing antibodies. Finally, the influence of soluble factors released by MSC in hydrogels on bone-forming cells was studied using osteoblasts obtained from trabecular bone explants from patients with osteonecrosis of the femoral head during total hip arthroplasty.Introduction and Objective
Materials and Method
Osteonecrosis of the femoral head (ONFH) is an evolving and disabling condition that often leads to subchondral collapse in late stages. It is the underlying diagnosis for approximately 3%–12% of total hip arthroplasties (THAs) and the most frequent aetiology for young patients undergoing THA. To date, the pathophysiological mechanisms underlying ONFH remain poorly understood. In this study, we investigated whether ONFH without an obvious etiological factor is related to impaired osteoblast activities, as compared to age-matched patients with primary OA. We cultured osteoblasts isolated from trabecular bone explants taken from the femoral head of patients with ONFH and from intertrochanteric region of patients with ONFH or with OA and compared their in vitro mineralisation capacity and secretion of paracrine factors.Introduction and Objective
Materials and Methods
