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<p>The use of synthetic scaffolds as an alternative strategy for repairing bone defects has garnered significant attention in bone tissue engineering. Human mesenchymal stem cells (MSCs) play a crucial role in the body’s regeneration processes, making the identification of suitable inducers for treating osteogenesis disorders essential. This study is the first report in the world on the induction of differentiation of human adipose-derived mesenchymal stem cells (hAD-MSCs) into osteoblast-like cells on an innovative scaffold of carboxymethylcellulose, polyglycerol, and sebacic acid, containing vitamin D2 and chondroitin-4 sulfate. Initially, the scaffolds were created using molding and salt washing techniques. Their morphology, structure, and mechanical characteristics were examined through techniques such as scanning electron microscopy (SEM), X-ray diffraction (XRD), Thermogravimetric Analysis (TGA), and Fourier-transform infrared spectroscopy (FTIR). The optimal concentrations of vitamin D2 and chondroitin sulfate (CS) in the differentiation medium and scaffold structure were determined using the MTT assay. Viability and morphology were assessed through acridine orange-ethidium bromide (AO/EB) staining. Differentiation experiments were conducted with various concentrations of inducers both in the medium and on the scaffolds. Cell differentiation into osteoblasts was quantified through alkaline phosphatase (ALP) enzyme activity assays and calcium content deposition (C.C). Qualitative assessments included staining the mineralized matrix with Alizarin Red and Kossa staining. Finally, the expression of bone marker genes, including <i>RUNX-2</i>, <i>collagen I</i>, <i>osteocalcin</i>, <i>osteonectin</i>, and <i>ALP</i>, was quantitatively evaluated using Real-Time PCR. The results of the osteogenic differentiation experiments demonstrated that scaffolds containing vitamin D2 and chondroitin-4-sulfate effectively induced osteogenic differentiation in mesenchymal stem cells. These scaffolds significantly increased alkaline phosphatase (ALP) activity and calcium deposition, as confirmed by specialized staining methods such as Alizarin Red and von Kossa. Furthermore, the expression levels of key genes involved in osteogenic differentiation were significantly upregulated, confirming the efficacy of these scaffolds in promoting osteogenesis.</p>

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Design and Fabrication of an Innovative Scaffold Made of Polyglycerol, Carboxymethyl Cellulose, and Sebacic Acid Containing Vitamin D2 and Chondroitin 4 Sulfate: Creating A Suitable Environment for Osteogenic Differentiation

  • 期刊论文

  • 发表日期：

    2025年10月25日

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  • Kiana Tabari,
  • Marzieh Ghollasi,
  • Raheleh Halabian,
  • Mohsen Ghiasi,
  • Hanieh Ahmadi

摘要

The use of synthetic scaffolds as an alternative strategy for repairing bone defects has garnered significant attention in bone tissue engineering. Human mesenchymal stem cells (MSCs) play a crucial role in the body’s regeneration processes, making the identification of suitable inducers for treating osteogenesis disorders essential. This study is the first report in the world on the induction of differentiation of human adipose-derived mesenchymal stem cells (hAD-MSCs) into osteoblast-like cells on an innovative scaffold of carboxymethylcellulose, polyglycerol, and sebacic acid, containing vitamin D2 and chondroitin-4 sulfate. Initially, the scaffolds were created using molding and salt washing techniques. Their morphology, structure, and mechanical characteristics were examined through techniques such as scanning electron microscopy (SEM), X-ray diffraction (XRD), Thermogravimetric Analysis (TGA), and Fourier-transform infrared spectroscopy (FTIR). The optimal concentrations of vitamin D2 and chondroitin sulfate (CS) in the differentiation medium and scaffold structure were determined using the MTT assay. Viability and morphology were assessed through acridine orange-ethidium bromide (AO/EB) staining. Differentiation experiments were conducted with various concentrations of inducers both in the medium and on the scaffolds. Cell differentiation into osteoblasts was quantified through alkaline phosphatase (ALP) enzyme activity assays and calcium content deposition (C.C). Qualitative assessments included staining the mineralized matrix with Alizarin Red and Kossa staining. Finally, the expression of bone marker genes, including RUNX-2, collagen I, osteocalcin, osteonectin, and ALP, was quantitatively evaluated using Real-Time PCR. The results of the osteogenic differentiation experiments demonstrated that scaffolds containing vitamin D2 and chondroitin-4-sulfate effectively induced osteogenic differentiation in mesenchymal stem cells. These scaffolds significantly increased alkaline phosphatase (ALP) activity and calcium deposition, as confirmed by specialized staining methods such as Alizarin Red and von Kossa. Furthermore, the expression levels of key genes involved in osteogenic differentiation were significantly upregulated, confirming the efficacy of these scaffolds in promoting osteogenesis.