Injectable catechol-functionalized hyaluronic acid/layered double hydroxide nanocomposite hydrogels for regulated...

Although catechol-functionalized hyaluronic acid hydrogels and layered double hydroxide (LDH)-based osteogenic delivery systems have each been investigated, the functional integration of LDH nanosheets as both structural reinforcing phases and cargo-regulating reservoirs within an injectable polysaccharide network remains insufficiently established. Here, we developed an oxidatively stabilized catechol-functionalized hyaluronic acid (HA-Cat) hydrogel incorporating ternary Ca-Mg-Al LDH nanosheets and the Hedgehog agonist SAG. The LDH nanosheets provided secondary organic-inorganic interactions, mechanical reinforcement, and regulated local presentation of SAG while preserving injectability and self-recovery. Transmission electron microscopy confirmed the nanosheet morphology of exfoliated LDH, and the ζ-potential shift from approximately +25 mV for bare LDH to -16 mV after mixing with HA-Cat supported electrostatic association at the organic-inorganic interface. LDH incorporation reduced swelling, increased stiffness, and delayed degradation without compromising rheological recovery. HPLC analysis determined the SAG loading efficiency to be approximately 50%. LDH@SAG exhibited slower SAG release than free SAG and the free SAG/blank LDH physical mixture, with the Weibull model providing the best full-profile fit (R² = 0.9690). Compared with freely incorporated SAG, LDH-associated SAG maintained comparable extracellular matrix mineralization after 14 days. The nanocomposite hydrogels also promoted osteogenic differentiation of three-dimensionally encapsulated bone marrow stromal cells and exhibited in vitro antimicrobial and antioxidant activities. In a murine critical-sized calvarial defect model, the LDH@SAG formulation supported new bone formation, as assessed by bone volume, bone surface, histology, and osteogenic marker expression. These findings demonstrate the dual structural and delivery-regulating role of LDH within an injectable HA-Cat network.