Lamparelli Erwin Pavel, Lovecchio Joseph, Ciardulli Maria Camilla, Giudice Valentina, Dale Tina P, Selleri Carmine, Forsyth Nicholas, Giordano Emanuele, Maffulli Nicola, Della Porta Giovanna
Department of Medicine, Surgery and Dentistry, University of Salerno, via S. Allende, 84081 Baronissi, SA, Italy.
Department of Electrical, Electronic and Information Engineering "Guglielmo Marconi" (DEI), University of Bologna, via dell'Università 50, 47522 Cesena, FC, Italy.
Pharmaceutics. 2021 Mar 17;13(3):399. doi: 10.3390/pharmaceutics13030399.
Tissue engineering strategies can be relevant for cartilage repair and regeneration. A collagen matrix was functionalized with the addition of poly-lactic-co-glycolic acid microcarriers (PLGA-MCs) carrying a human Transforming Growth Factor β1 (hTFG-β1) payload, to provide a 3D biomimetic environment with the capacity to direct stem cell commitment towards a chondrogenic phenotype. PLGA-MCs (mean size 3 ± 0.9 μm) were prepared via supercritical emulsion extraction technology and tailored to sustain delivery of payload into the collagen hydrogel for 21 days. PLGA-MCs were coseeded with human Bone Marrow Mesenchymal Stem Cells (hBM-MSCs) in the collagen matrix. Chondrogenic induction was suggested when dynamic perfusion was applied as indicated by transcriptional upregulation of COL2A1 gene (5-fold; < 0.01) and downregulation of COL1A1 (0.07-fold; < 0.05) and COL3A1 (0.11-fold; < 0.05) genes, at day 16, as monitored by qRT-PCR. Histological and quantitative-immunofluorescence (qIF) analysis confirmed cell activity by remodeling the synthetic extracellular matrix when cultured in perfused conditions. Static constructs lacked evidence of chondrogenic specific gene overexpression, which was probably due to a reduced mass exchange, as determined by 3D system Finite Element Modelling (FEM) analysis. Proinflammatory (IL-6, TNF, IL-12A, IL-1β) and anti-inflammatory (IL-10, TGF-β1) cytokine gene expression by hBM-MSC was observed only in dynamic culture (TNF and IL-1β 10-fold, < 0.001; TGF-β1 4-fold, < 0.01 at Day 16) confirming the cells' immunomodulatory activity mainly in relation to their commitment and not due to the synthetic environment. This study supports the use of 3D hydrogel scaffolds, equipped for growth factor controlled delivery, as tissue engineered models for the study of in vitro chondrogenic differentiation and opens clinical perspectives for injectable collagen-based advanced therapy systems.
组织工程策略可能与软骨修复和再生相关。通过添加携带人转化生长因子β1（hTFG-β1）的聚乳酸-乙醇酸共聚物微载体（PLGA-MCs）对胶原基质进行功能化处理，以提供一个三维仿生环境，使其具有引导干细胞向软骨生成表型分化的能力。PLGA-MCs（平均尺寸为3±0.9μm）通过超临界乳液萃取技术制备，并经过调整以在21天内持续将负载物递送至胶原水凝胶中。将PLGA-MCs与人骨髓间充质干细胞（hBM-MSCs）共同接种到胶原基质中。如通过qRT-PCR监测，在第16天时，当进行动态灌注时，COL2A1基因转录上调（5倍；<0.01）以及COL1A1（0.07倍；<0.05）和COL3A1（0.11倍；<0.05）基因下调，提示有软骨生成诱导作用。组织学和定量免疫荧光（qIF）分析证实，在灌注条件下培养时，细胞通过重塑合成细胞外基质表现出细胞活性。静态构建物缺乏软骨生成特异性基因过表达的证据，这可能是由于通过三维系统有限元建模（FEM）分析确定的质量交换减少所致。仅在动态培养中观察到hBM-MSC的促炎（IL-6、TNF、IL-12A、IL-1β）和抗炎（IL-10、TGF-β1）细胞因子基因表达（第16天时，TNF和IL-1β为10倍，<0.001；TGF-β1为4倍，<0.01），证实细胞的免疫调节活性主要与其分化有关，而非由于合成环境。本研究支持使用配备生长因子可控递送功能的三维水凝胶支架作为体外软骨生成分化研究的组织工程模型，并为基于注射用胶原的先进治疗系统开辟了临床前景。
