Abstract <p>Immunocompatibility and oxidative stress challenge the functional performance of biomaterials post-implantation hurdling the success of tissue engineering. Hence, a thorough in vitro assessment of immunocompatibility and the ability of biomaterials to withstand oxidative stress is critical for successful tissue engineering applications. Considering alginate-based hybrid hydrogels and in vitro challenged RAW264.7 macrophages as models, this study aims to assess their antioxidant potential and immunocompatibility. Alginate-CMC-PVA (ACPV) and Alginate-starch-PEG (ASPG) hydrogels were prepared in two batches ACPV1 and ACPV2 and ASPG1 and ASPG2 by varying alginate content. Ascorbic acid loaded ACPV and ASPG hydrogels displayed appreciable scavenging effects against DPPH and NO compared to untreated control. Additionally, both the hydrogels effectively shielded oxidative stress in H₂O₂ challenged RAW264.7 macrophages and retained their normal morphology suggesting the absence of immune activation. Both ACPV and ASPG hydrogels displayed the downregulation of anti-inflammatory TGF-β and IL-10 with a concomitant upregulation of the proinflammatory mediators IL6, NFκB, TNF-α, and IKB compared to LPS-treated controls. This observation confirms the immunocompatibility of ACPV and AGPG hydrogels. Moreover, network analysis revealed the pathways of inflammatory resolution and repair correlating with the level expression of IL6, NFκB, IL-10, TGF-β, TNF-α, and IKB revealing the possible gene interactions in determining the success of ACPV and ASPG hydrogels. Overall, both ACPVs and ASPGs exhibited appreciable antioxidant response and immunocompatibility; however, further optimizations are warranted for the translational applications in tissue engineering.</p> Lay Summary <p>Tissue engineering is an emerging field that utilizes hydrogels to reconstruct the organs to treat various diseases. The hydrogels serve primarily as scaffolds for the regeneration of the organ parts. However, as a foreign entity, the hydrogel scaffolds could elicit adverse effects (immune response) within the body. The present study focuses on assessing the immunological effects and protective mechanisms of tissue engineering hydrogels synthesized from polymers including alginate, starch, carboxy methyl cellulose, and poly vinyl alcohol. The hydrogels were rigorously tested using laboratory grown immune cells. The findings revealed that the hydrogels exhibited appreciable protective effects and immunocompatibility; however, further optimizations are warranted for the translational applications in tissue engineering.</p>