The researchers tested whether the scaffold’s geometry could tune mechanical performance without changing the hydrogel formulation or photocrosslinking conditions. Using melt electrowriting, they produced PLGA fiber lattices with a porosity above 85%, then compared square grids spaced 1,000 and 500 micrometers apart and a 500-micrometer diamond lattice. The finer square grid produced more continuous load-transfer pathways and the highest maximum compressive force during 10 loading cycles. In tensile dynamic mechanical analysis, the square lattice reached a storage modulus above 100 megapascals, approximately one order of magnitude above the diamond design. By comparison, pure GelMA had a storage modulus of about 6 kilopascals. Notably, the PLGA framework represented only 1% by weight of the entire composite, yet it increased stiffness by several orders of magnitude relative to the unfilled gel.

Bioactive glass added a second, biologically relevant form of reinforcement. Particles measuring 20–50 micrometers were mixed into the GelMA precursor at 1, 5, 10 or 20% by weight, and scanning electron microscopy showed that they remained distributed throughout the hydrogel without obvious aggregation. The 1% formulation produced the most favorable balance. It had the lowest swelling, retained its shape after immersion in phosphate-buffered saline and reduced weight loss after 28 days from about 60% for the glass-free composite to approximately 25%. Dynamic mechanical analysis likewise showed an increase in storage modulus at 1% glass, followed by weakening at higher loading, which the authors attributed to particles disrupting continuity in the crosslinked GelMA network. The composite also released calcium, silicon and phosphorus over 21 days as the glass dissolved and the surrounding gel degraded.

Biocompatibility tests showed that the optimum was not simply the formulation with the most glass or the stiffest network. Rat bone-marrow mesenchymal stem cells maintained high viability on the fiber-reinforced composites containing 1 or 5% glass, with the 1% material producing the strongest proliferation signal and cell spreading. High glass contents of 10 and 20% suppressed proliferation during longer culture periods. All formulations produced hemolysis rates below the cited 5% threshold. These results identified P/G@1BG, containing 1% bioactive glass, as the formulation that combined mechanical stability, controlled swelling, sustained ion release and favorable cellular responses. Accelerated alkaline testing also showed that PLGA was more hydrolytically susceptible than polycaprolactone, supporting its selection as a reinforcement intended to lose structural contribution over time.

The optimized composite also accelerated osteogenic differentiation in rat bone-marrow stromal cells. After seven days, alkaline phosphatase activity in cells exposed to P/G@1BG was approximately 1.6 times that of the blank control, and by day 21 the material produced the greatest calcium-mineralization signal in Alizarin Red assays. Molecular measurements showed significant increases in early and late bone-associated genes, including alkaline phosphatase, BMP-2, Runx2, osteopontin, osteocalcin and collagen type I alpha 1. Immunofluorescence at day 14 placed Runx2 mainly in cell nuclei and revealed extensive osteocalcin expression in the cytoplasm. The authors linked these responses to the combined effects of a cell-adhesive GelMA matrix, sustained glass-derived ions and the PLGA scaffold’s guidance of cell organization. The evidence supports enhanced osteogenic differentiation in vitro; it does not by itself establish clinical effectiveness in people.

Pro-angiogenic effects were observed through a second route. P/G@1BG raised the expression of vascular endothelial growth factor A and hypoxia-inducible factor 1-alpha in the stem cells. When the culture medium from treated cells was transferred to human umbilical vein endothelial cells, the P/G@1BG-conditioned medium produced approximately 75.6% closure of a scratch after 24 hours, compared with 45.4% for medium from cells on the glass-free composite and 50.0% for the control medium. The same conditioned medium generated the most extensive capillary-like networks in a six-hour tube-formation assay, with more branching junctions and greater total tube length. These assays indicate that the hydrogel stimulated rat stem cells to release soluble signals that promoted endothelial migration and network formation in vitro.

The researchers next implanted the optimized material into bilateral, full-thickness, 5-millimeter calvarial defects in 18 adult Sprague-Dawley rats, assigning the two defects in each animal as paired sites for a blank control, the glass-free P/G scaffold or P/G@1BG. Specimens were examined at eight and 12 weeks by micro-computed tomography, histology and immunostaining. At 12 weeks, bone coverage reached approximately 60% in defects treated with P/G@1BG, compared with 40% for P/G and 25% for controls. Micro-CT analysis found a bone-volume-to-total-volume ratio of 59.85 ± 7.39%, a relative bone mineral density of 8.00 ± 0.40 times control, a trabecular-number increase of 2.87 ± 0.63 times and a trabecular-separation reduction to 0.56 ± 0.07 times control. Histology showed mature, collagen-rich bone closely associated with the implant in the optimized group, whereas controls contained mostly fibrous tissue and limited peripheral bone.

Markers of both tissue-building systems also rose within the treated defects. At 12 weeks, relative Runx2 and CD31 signals in P/G@1BG defects were 3.0 and 2.8 times those in blank controls, respectively. Double labeling showed persistent spatial association between Runx2-positive osteoblasts and CD31-positive vessels at both eight and 12 weeks, indicating coupled osteogenic and angiogenic activity during healing. The scaffolds progressively degraded while host tissue grew into them, and only mild inflammation was observed. Histological examination of major visceral organs at eight weeks showed no obvious pathological lesions, inflammatory infiltration or necrosis attributable to the material. These findings demonstrate enhanced bone repair and vascularization in a rat cranial-defect model, not yet in human patients or in defects comparable to every form of craniofacial trauma.

RNA sequencing offered a possible molecular explanation for the dual response. Compared with control cells, rat stem cells cultured on P/G@1BG had 784 significantly upregulated and 401 downregulated genes. Enrichment analysis associated the upregulated genes with cell proliferation, extracellular-matrix deposition and signaling pathways including Ras/MAPK. The pathway was also connected to the measured increases in Runx2, osteocalcin, HIF-1-alpha and VEGFA, suggesting that it may participate in osteogenic commitment and the paracrine activation of endothelial cells. The authors were careful not to treat enrichment as proof of causation: additional scaffold controls and experiments that directly disrupt Ras/MAPK signaling would be needed to establish causality. Taken together, the results show how a 3D-printed polymer skeleton, a soft bioactive matrix and glass-derived ions can be allocated different roles, producing a bioresorbable material that supports vascularized bone formation in preclinical testing.

Subject of Research: A mechanically reinforced bioactive hydrogel for osteogenic and angiogenic coupling in craniofacial bone repair

Article Title: Synergistically reinforced bioresorbable hydrogel for mechanical–biological decoupling and osteogenic–angiogenic coupling

Article References: Hong, Y., Lu, H., Luo, J., Li, Y., Ye, Y., Hua, Y., Xu, H., & Xu, T. (2026). Synergistically reinforced bioresorbable hydrogel for mechanical–biological decoupling and osteogenic–angiogenic coupling. Materials Today Bio, 41, Article 103736. https://doi.org/10.1016/j.mtbio.2026.103736

Image Credits: AI Generated

DOI: 10.1016/j.mtbio.2026.103736

Keywords: bioresorbable hydrogel, craniofacial bone repair, melt electrowriting, bioactive glass, PLGA scaffold, osteogenesis, angiogenesis, tissue engineering, rat calvarial defects, Synergistically, reinforced, bioresorbable