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Recreating the human bone marrow microenvironment in vitro remains a critical challenge in advancing our understanding of hematopoiesis and its disruption in disease. Here, we present a fully tunable bone marrow model based on silk fibroin scaffolds engineered through thermo-chemical processing to replicate the mechanical and structural features of native marrow. This 3D platform integrates mesenchymal stromal cells (MSCs) and supports the functional differentiation of hematopoietic stem and progenitor cells (HSPCs) into mature megakaryocytes and platelets. RNA sequencing of MSCs cultured on physiologically tuned scaffolds revealed transcriptional programs closely aligned with native stroma, validating the fidelity of the engineered niche. The model captures essential marrow dynamics, including matrix remodeling and perfusion flow, enabling direct assessment of thrombopoietic function. To simulate fibrotic remodeling, scaffolds were functionalized with TGF-β1, inducing MSC transition into myofibroblast-like cells and recreating pathological features of myeloproliferative neoplasms. In this context, patient-derived HSPCs exhibited impaired megakaryocyte maturation and aberrant calcium signaling, partially restored by interfering with calcium flux. To quantify microenvironment-driven dysfunction, we calculated changes in megakaryocyte size distribution using a Divergence Index. Combined with the engineered niche, this functional metric offers a powerful and quantifiable platform to dissect dysregulated hematopoiesis and evaluate therapeutic strategies in patient-derived systems.