In summary, we made the CS-GE hydrogels crosslinked with three different
concentration of gloxal (0.0025%, 0.005% and 0.01%). The rheology study showed that
CS-GE hydrogels crosslinked with 0.005% glyoxal and 0.01% glyoxal had better mechanical properties since the its elastic modulus (G’) was higher than the other
groups. Degradation behavior of hydrogels crosslinked with 0.01% glyoxal obviously
degrade slower than the other group. In the sustained release experiment, our result
suggested that CS-GE hydrogels crosslinked with different concentration of glyoxal had
similar release profile of HPL, 4 kDa FITC-dextran and 40 kDa FITC-dextran.
Nevertheless, in the 500 kDa FITC-dextran release experiment, CS-GE hydrogels
crosslinked with 0.01% glyoxal showed slower release rate than the other groups. In
addition, we also proved that HPL can not only stimulate the migration of HS68 cells
and HUVEC but promote the tube formation of HUVEC. Though we haven’t prove that
CS-GE hydrogels combined with HPL can stimulate angiogenesis in the CAM assay, we
still observed that eggs treated with hydrogels and HPL showed more complicated
vascular network in some cases. On the other hand, we also try to build a 3D skin
equivalent model to assess the angiogenic effect of HPL. Even there are some
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Appendix:
Fig. 4 Gelification samples(2% chitosan, 1% gelatin and 7.125% β-GP) with different concentrations of glyoxal.
Fig. 5 In vitro degradation test of CS-GE hydrogels. All the samples are composed of 2%
chitosan , 1% gelatin and 7.125% β-GP with different concentrations of glyoxal.
Fig. 6 Mechanical properties of different composition hydrogels. All the samples are composed of 2% chitosan , 1% gelatin and 7.125% β-GP with different concentrations of glyoxal.
Hydrogels are tested as frequency sweeps at 37℃.
Fig. 7 Human platelet lysate(A) & FITC-dextran (molecular weight: (B)4, (C)40 and (D)500 kDa) release from chitosan/gelatin/β-GP based hydrogel with different concentrations of glyoxal.(*: p value<0.05)
Fig. 9 Effect of HPL on cell activity tested by alamar blue assay (A) Different amount of human platelet lysate in EBM incubates with HUVEC. The group 2% FBS(Lonza) without growth factor and 10% FBS(Hyclone) are as control. (B) Different amount of HPL in DMEM incubates with HS68 cells. The 10% FBS(Hyclone) are as control. (*: p value<0.05, **: p value<0.01)
Fig. 11 HPL embedded in chitosan/gelatin/β-GP based hydrogel with 0.005% glyoxal to promote HUVEC migration.(A)Representative picture of HUVEC in transwell migration assay. The cells migrated in 24 hours are stained with crystal violet (B)Number of cells migrated in 24 hours are counted by image J (*: p value<0.05, **: p value<0.01 and scale bar=100μm)
Fig. 12 Tube formation assay of HUVEC on different amount of HPL.(A)Morphology of HUVEC on 2h, 4h, 6h and 24h (B)Number of
Fig. 13 Scheme of 3D skin equivalent model
Fig. 14 3D skin equivalent model photographed (A) 1.5 hr after seeding HUVEC (B) after taking oring out (C) after adding hydrogel+HPL (scale bar=100μm)
Fig. 15 Timeline of CAM assay