The extracellular matrix (ECM) is a natural microenvironment pivotal for stem cell survival, as well as proliferation, differentiation and metastasis, composed of a variety of biological molecular complexes secreted by resident cells in tissues and organs. Heparan sulfate proteoglycan (HSPG) is a type of ECM protein that contains one or more covalently attached heparan sulfate chains. Heparan sulphate chains have high affinity with growth factors, chemokines and morphogens, acting as cytokine-binding domains of great importance in development and normal physiology. Herein, we constructed endogenous HSPG2 overexpression in mouse embryonic fibroblasts based on the CRISPR/Cas9 synergistic activation mediator system and then fabricated a cell-derived HSPG2 functional ECM (ECMHSPG2). The ECMHSPG2 is capable of enriching basic fibroblast growth factor (bFGF), which binds more strongly than the negative control ECM. With a growing bFGF concentration, ECMHSPG2 could better maintain neural stem cell (NSCs) stemness and promote NSC proliferation and differentiation in culture. These findings provide a precise design strategy for producing a specific cell-derived ECM for biomaterials in research and regenerative medicine.