Fig. 5.

Hydrogel loaded with EV-NID1 significantly promotes angiogenesis at bone defects sites. (A) Program diagram of the construction and application of EVs-hydrogel. (B) The microstructure and EVs distribution inside hydrogels were characterized by SEM. (C) At 4 weeks post operation, HE staining was used to observe the cell infiltration and the hydrogel degradation at the defect site. The green dashed box represents the integration of the interface between the regenerative tissue and the host bone, and the blue dashed box represents the status of tissue filling and material degradation in the defect center. (D) Sirius red staining showed collagen coverage in the defect area. Under polarized light, red/orange represents typeⅠcollagen, and green represents type Ⅲ collagen. Scale bar,200 μm. (E) The relative coverage of type Ⅲ collagen in the defect area, and the proportion of type Ⅲ collagen in total typeⅠand Ⅲ collagen. (F) Immunofluorescence staining of CD31 and NID1, the white arrow represents the double-positive tubular structure of CD31 and NID1. Scale bar,100 μm. (G) Comparison of the relative number of CD31⁺ vessels, and count the CD31/NID1 double positive cells. Data were represented as the mean ± SD. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not significant from Student's t-test.