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Mesenchymal stem cells proliferation and osteogenic differentiation on polymeric scaffolds and microspheres for bone tissue engineering

Walczak Kamila, Krok-Borkowicz Małgorzata, Pamuła Elżbieta

Engineering of Biomaterials

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2023

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vol. 26, no. 170

14--19

EN

In this study, we aimed to compare how the microstructure and architecture of polymer supports influence adhesion, growth and differentiation of human mesenchymal stem cells (hMSC) in the context of bone tissue engineering. We manufactured poly(L-lactide-co-glycolide) (PLGA) three-dimensional supports in the form of microspheres by emulsification and porous scaffolds by solvent casting/ porogen leaching. HMSC were seeded on both materials and on control tissue culture polystyrene (TCPS, bottom of the wells) and cultured in basal or osteogenic medium for 1, 3, 7 and 14 days. HMSC proliferation and osteogenic differentiation were studied using lactate dehydrogenase (LDH) and alkaline phosphatase (ALP) assays, respectively. Furthermore, cell morphology and viability were analyzed after live/dead fluorescence staining. The results show that the optimized emulsification conditions allowed the production of PLGA microspheres with a median size of 95 µm. The PLGA scaffolds had a porosity of 82.1% ± 4.2% and a pore size of 360 µm ± 74 µm. HMSC cultured on control TCPS in osteogenic medium were more spread and polygonal than those in basal medium. They were characterized with a lower proliferation rate, as shown by the LDH results, but higher ALP activity. This suggests that hMSC osteogenic differentiation was achieved. The same tendency was observed for cells cultured on microspheres and scaffolds. Cell proliferation was more efficient on both materials and control in growth medium as compared to differentiation medium. The amount of ALP, i.e. a marker of osteogenic differentiation, was elevated, as expected, in differentiation medium. However, on day 14 cells cultured on the scaffolds in basal medium exhibited the same osteogenic potential as those cultured in differentiation medium. In general, both microspheres and scaffolds promoted hMSC adhesion, proliferation, and osteogenic differentiation and may be used for bone tissue engineering.

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Osteoblasts response to novel chitosan/agarose/hydroxyapatite bone scaffold – studies on MC3T3-E1 and hFOB 1.19 cellular models

Kazimierczak Paulina, Vivcharenko Vladyslav, Truszkiewicz Wiesław, Wójcik Michał, Przekora Agata

Engineering of Biomaterials

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2019

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Vol. 22, no. 151

24--29

EN

Since it is known that various cell lines may ex-press different behaviours on the scaffolds surface, a comprehensive analysis using various cellular mo-dels is needed to evaluate the biomedical potential of developed biomaterials under in vitro conditions. Thus, the aim of this work was to fabricate bone scaffolds composed of a chitosan-agarose matrix reinforced with nanohydroxyapatite and compare the biological response of two cell lines, i.e. mouse calvarial preosteoblasts (MC3T3-E1 Subclone 4) and human foetal osteoblasts (hFOB 1.19). Within this study, the osteoblasts number on the scaffold surface and the osteogenic markers level produced by MC3T3-E1 and hFOB 1.19 cells were determined. Furthermore, changes in calcium and phosphorous ions concentrations in the culture media dedicated for MC3T3-E1 and hFOB 1.19 were estimated after the biomaterial incubation. The obtained results proved that the fabricated biomaterial is characterized by biocompatibility and osteoconductivity since it favours osteoblasts attachment and growth. It also supports the production of osteogenic markers (collagen, bALP, osteocalcin) by MC3T3-E1 and hFOB 1.19 cells. Interestingly, the developed biomaterial exhibits different ion reactivity values in the two culture media dedicated for the mentioned cell lines. It was also revealed that mouse and human osteoblasts differ in the cellular response to the fabricated scaffold. Thus, the use of at least two various cellular models is recommended to carry out a reliable biological characterization of the novel biomaterial. These results demonstrate that the tested bone scaffold is a promising biomaterial for bone regeneration applications, however further biological and physicochemical experiments are essential to fully assess its biomedical potential.

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Integrin αv signaling influences phenotype and maturation of primary human osteoblasts on alumina surface

Wróbel E., Witkowska-Zimny M., Mrówka P., Głodkowska-Mrówka E.

Engineering of Biomaterials

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2014

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Vol. 17, no. 127

33--39

EN

Due to the growing interest in stem cells application in tissue engineering the better understanding of primary human osteoblasts behavior in vitro, on biomaterial surface, is required. Among other molecules integrins may be taken into account as being involved in these phenomena. Integrins are a family of cell adhesion receptors, which may regulate many cellular functions e.g., adhesion, motility, phenotype and cell maturation. The aim of this study was to determine the effect of the biomaterial surfaces and αv integrin signaling pathway on the behavior, phenotype and maturation of human osteoblasts in vitro. Human bone derived cells (HBDCs) obtained from adult femoral bone fragments were cultured on both alumina disks and tissue culture polystyrene (TCPS) dishes. After 7, 14, and 21 days of culture, localization and mRNA expression level of αv integrin subunits and BGLAP (osteocalcin) on polystyrene were analyzed in addition, we treated the cell cultures with monoclonal antibodies against human αv integrin to block its ligand-binding activity, on both alumina and TCPS substrates. We found that the αv integrin was present in focal contacts and cell cytoplasm at subsequent stages of cell maturation and the level of αv integrin mRNA was the highest in mature osteoblasts. Blocking αv integrin transduction pathway caused changes in cell activity and morphology, decreased cells proliferation on TCPS and reduced expression of alkaline phosphatase (ALP) on both materials. The results suggest that αv integrin is involved as an important receptor facilitating osteogenic differentiation.