3D printed fibroblast-laden alginate-cellulose scaffolds support extracellular matrix formation and angiogenic growth factor secretion

Effective microvascular tissue engineering requires fibroblasts that remain phenotypically stable and secrete extracellular matrix (ECM) proteins and growth factors relevant for vascularization. This study evaluated 3D printed hydrogels based on sodium alginate (ALG) and carboxymethyl cellulose (CMC) to assess their ability to sustain fibroblast phenotype, ECM deposition, and angiogenic growth factor secretion during long-term culture. Seven formulations – including one with nanofibrillated cellulose – were compared by encapsulating fibroblasts and crosslinking with CaCl₂ or SrCl₂. All scaffolds were printable and exhibited comparable degradation profiles. Mechanical testing indicated stable compressive response, with Sr 2+ -crosslinked hydrogels generally showing higher apparent compressive modulus, while Ca 2+ -crosslinked scaffolds supported slightly higher cell viability. Encapsulated fibroblasts retained their phenotype for 30 days, evidenced by steadily increasing collagen I/III and fibronectin deposition, alongside sustained expression of specific fibroblast markers. After 30 days, all groups produced comparable levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF-2), with one formulation yielding a significantly higher FGF-2 output. This multiparametric study demonstrates that scaffold composition and crosslinker chemistry influence fibroblast phenotype maintenance, ECM deposition, and growth factor secretion. To our knowledge, this is the first systematic, 30-day screening of ALG-CMC hydrogels – tuned by polymer content, NFC addition, and Ca 2+ /Sr 2+ crosslinking – specifically for their ability to sustain fibroblast phenotype, extracellular matrix deposition, and growth factor secretion, providing design considerations to guide bioink development for microvascular models.