T. Fang, et al. Chemical Engineering Journal 370(2019)573-586 Blank BMP-2 BMP-2/HD NF-Ms B a BMP-2/HD NF-Ms 7Days 14 Days 14 Days BMP-2/HD NF-\s >⌒ 21 Days E BMP-2/HD NF-Ms □BMP2/HDNF-Ms sss 3 14 Days Fig. 5. The osteogenesis effects of 100 ng/ml soluble rhBMP-2(BMP-2 group)and rhBMP-2 released from Hep-Dopa NF-Ms(BMP-2/HD NF-Ms)on BMSCs in vitro. (A) The ALP staining and(B)quantitative analysis of relative alP expression on 7 days and 14 days.(C)Alizarin Red s(ARS)staining and (D)qu the 21st day.(E) Real-time PCR analysis of osteogenic-related gene ALP, Runx2 and Coll expression of BMSCs in different groups after induced with rhBMP-2 at 14 days. Statistically analysis by using one-way analysis of variance(ANOVA)(P 0.05, P 0.01).( For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) DAPI GFP-SHED Merge 6. The survival of sheds in the constructs fter implantation in vivo at different time B cryosections collected at week 1 (A-C), week 4 7Days plantation. The GFP-expressing SHED(green; B were observed in ectopic tissue grafts at l and 4 weeks, but undetectable at 8 weeks (H). Scale bars (A-D): 100 um. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this D F 4 Week G 8 Weeks21 Days C A 7 Days 14 Days Blank BMP-2 BMP-2/HD NF-Ms B D E Fig. 5. The osteogenesis effects of 100 ng/ml soluble rhBMP-2 (BMP-2 group) and rhBMP-2 released from Hep-Dopa NF-Ms (BMP-2/HD NF-Ms) on BMSCs in vitro. (A) The ALP staining and (B) quantitative analysis of relative ALP expression on 7 days and 14 days. (C) Alizarin Red S (ARS) staining and (D) quantitative results at the 21st day. (E) Real-time PCR analysis of osteogenic-related gene ALP, Runx2 and Col1 expression of BMSCs in different groups after induced with rhBMP-2 at 14 days. Statistically analysis by using one-way analysis of variance (ANOVA) (* P < 0.05, **P < 0.01). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.) DAPI GFP-SHED Merge 7 Days 4 Weeks 8 Weeks ABC D E F GHI 100ȝm Fig. 6. The survival of SHEDs in the constructs after implantation in vivo at different time points. Fluorescence detection was performed in cryo-sections collected at week 1 (A-C), week 4 (D-F), and week 8 (G-I) after subcutaneous im￾plantation. The GFP-expressing SHED (green; B, E) were observed in ectopic tissue grafts at 1 and 4 weeks, but undetectable at 8 weeks (H). Scale bars (A-I): 100 μm. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this ar￾ticle.) T. Fang, et al. Chemical Engineering Journal 370 (2019) 573–586 582