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The success of cell-based approaches for the treatment of cartilage or fibro-cartilaginous tissue defects requires an optimal cell source with chondrogenic differentiation ability that maintains its differentiated properties and stability following implantation. For this purpose, the aim of this study was to evaluate the use of endostatin (COL18A1), an anti-angiogenic factor, which is physiologically involved in cell differentiation during meniscus development. Swine neonatal meniscal cells not yet subjected to mechanical stimuli were extracted, cultured in fibrin hydrogel scaffolds, and treated at two different time points (T1 = 9 days and T2 = 21 days) with different concentrations of COL18A1 (10 ng/mL; 100 ng/mL; 200 ng/mL). At the end of the treatments, the scaffolds were examined through biochemical, molecular, and histochemical analyses. The results showed that the higher concentration of COL18A1 promotes a fibro-chondrogenic phenotype and improves cellularity index (DNA content, <i>p</i> < 0.001) and cell efficiency (GAGs/DNA ratio, <i>p</i> < 0.01) after 21 days. These data are supported by the molecular analysis of collagen type I (COL1A1, a marker of fibrous-like tissue, <i>p</i> < 0.001), collagen type II (COL2A1, a marker of cartilaginous-like tissue, <i>p</i> < 0.001) and SRY-Box Transcription Factor 9 (SOX9, an early marker of chondrogenicity, <i>p</i> < 0.001), as well as by histological analysis (Safranin-O staining), laying the foundations for future studies evaluating the involvement of 3D endostatin hydrogel scaffolds in the differentiation of avascular tissues.