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. 2019 Sep 21;9(23):6949–6961. doi: 10.7150/thno.38061

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This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.

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Fabrication of GDF5-conjugated BMSC-laden scaffold for cartilage repair. A. 3D-bioprinted cartilage scaffold for implantation. (a) Schematic Illustration of constructed scaffolds for cartilage repair in rabbit knee. (b) 3D CAD of each layer of the cartilage scaffold and (c) the dispensing path (yellow box outlined in b) of (d) aligned PCL and hydrogel (green box outline in c). GDF5-conjugated BMSC-laden hydrogel was dispensed into the space between PCL fibers. (e) Gross appearance of GDF5-conjugated BMSC-laden hydrogel printed with OPUS. (f) Hydrogel was further observed under light microscopy. (g) Gross appearance of the cartilage scaffold with GDF5-conjugated BMSC-laden hydrogel and PCL as supporting structure. (h) SEM images of GDF5-conjugated PLGA μS. (i) Implantation process of the cartilage scaffold into the defect site in a rabbit knee. (j) Higher resolution image of the implanted scaffold in (i). (k) The alignment of PCL and hydrogel in the scaffold under microscopy. (l) Higher resolution image of the blue box area outlined in (k). (m to p) Minimal toxicity and distribution PLGA μS in BMSC-laden hydrogel in the scaffolds. (m) Fluorophore-conjugated rhodamine was encapsulated into PLGA μS and delivered to the hydrogel in the printed scaffolds. (n) At day 7, live BMSCs and (o) dead BMSCs in the PLGA-conjugated hydrogel printed between the PCL fibers were demonstrated with live/dead assay and observed under confocal microscope. (p) Merged image for (m to o). B. Mechanical spectra of different component (gel: gelatin; GF: gelatin + fibrinogen; GFHG: gelatin + fibrinogen + hyaluronic acid + glycerol) and the cross-linked hydrogel (GFHG hydrogel) measured at 17 °C. C. Dynamic thermal rheological observations of the cross-linkage of GFHG. D. Degradation rate of BMSC-laden hydrogel and E. PCL in vitro and in vivo. CH: BMSC cell-laden hydrogel: CHG: BMSC cell-laden hydrogel with conjugated GDF5. CHM and CHGM: CH and CHG assessed in nude mice. F. Released GDF5 concentration from PLGA μS was measured using enzyme-linked immunosorbent assay (ELISA) kits. Spheres showed controlled release of GDF5 that sustained over 60 days in vitro. G. Cell proliferation in the cartilage scaffolds. To determine cell proliferation in the scaffolds, we examined survival of BMSCs in the scaffolds compared to BMSCs cultured in fibrin through 21 days post printing with AlamarBlue assay kit. H. Biomechanical properties of the in vitro cartilage construct, including bulk tensile modulus and I) UTS after 12 weeks of culture. *P < 0.05 between the native or the GDF5-conjugated group and control group. All data are means ± SD (n = 6) and were analyzed by two-way ANOVA with Tukey's test.