Page 213 - IJB-10-5

International Journal of Bioprinting Biomimetic osteochondral scaffold

cartilage layers) were tightly bonded together and exhibited intensity of COL II in the group treated with 50 ng/mL
their original morphology. FGF-18 was significantly higher than that in groups treated
with 0 and 10 ng/mL FGF-18. In contrast, no significant
To investigate the mechanical compatibility between
the scaffold and native osteochondral tissues, we conducted difference was observed among the groups treated with
50, 100, and 150 ng/mL BMP-2, as well as the positive
compression tests on the 3D-printed osteochondral control group (Figure 3D). The optimal concentrations for
scaffolds. The stress–strain curve demonstrated that osteogenic and chondrogenic differentiation of rBMSCs
the compressive strengths of different layers of the were determined to be 150 ng/mL of BMP-2 and 50 ng/mL
osteochondral scaffold were of a considerable difference of FGF-18, respectively.
(cartilage layer < subchondral layer < interface layer),
and the overall compressive strength of the osteochondral Next, we fabricated subchondral scaffolds containing
scaffold was between the subchondral and interface different amounts of BMP-2 (1, 2, 3, and 4 μg BMP-2 per
layers (Figure 2C). The Young’s modulus of the cartilage g scaffold) and cartilage scaffolds containing different
layer (2.01 ± 0.22 MPa), the interface layer (53.43 ± 2.75 amounts of FGF-18 (1, 2, 3, and 4 μg BMP-2 per g scaffold)
MPa), the subchondral layer (19.20 ± 1.13 MPa), and to determine the precise GF loading amount required to
the integrated osteochondral scaffold (30.33 ± 2.80 MPa) maintain the optimal concentration mentioned above.
were comparable to that of human osteochondral tissues Figure 3E–G features the release behavior of BMP-2 from
(cartilage tissue: approximately 1.0 MPa; subchondral subchondral scaffolds in SBF over a 30-day period (measured
tissue: 20–500 MPa) (Figure 2D). 37–40 Next, the degradation using ELISA). The subchondral scaffolds loaded with BMP-
behavior of scaffolds was investigated over an 8-week 2 exhibited decreased release rates when the initial loading
period. As displayed in Figure 2E, both the subchondral amount was decreased (Figure 3E). The percentage of BMP-
and interface layers demonstrated a slow degradation 2 release increased and reached 84.81–86.50% within 30
rate (<12% at 8 weeks), and no significant difference was days (Figure 3F). The BMP-2 released every 3 days revealed
observed. The slow but continuous degradation was due to that subchondral scaffolds loaded with 3 and 4 μg/g BMP-
the hydrolysis of PLGA and the decomposition of β-TCP. 2 could maintain a concentration of 150 ng/mL for up to
The cartilage layer reported a much faster degradation 14 days, whereas subchondral scaffolds loaded with 1 and 2
rate (>47% at 8 weeks), mainly owing to the relatively fast μg/g BMP-2 could not achieve this concentration retention
degradation of GelMA hydrogel and slow hydrolysis of (Figure 3G). Figure 3H–J presents the release behavior of
the P(DLLA-TMC) matrix. Conversely, the osteochondral FGF-18 from cartilage scaffolds in SBF measured using
ELISA for 30 days. For cartilage scaffolds loaded with
scaffold exhibited a moderate degradation rate between different amounts of FGF-18, the total FGF-18 release was
the other layers. These data indicated that a tri-phasic also positively correlated with the loading amount (Figure
osteochondral scaffold that structurally and mechanically 3H). The percentage release of FGF-18 reached 64.11–
matched the native osteochondral tissues was successfully 66.87% in 30 days (Figure 3I). The FGF-18 released every 3
constructed via cryogenic 3D printing.
days demonstrated that cartilage scaffolds loaded with 2, 3,
3.3. The spatiotemporally controlled release of and 4 μg/g of FGF-18 could maintain a concentration of 50
BMP-2 and FGF-18 ng/mL for up to 21 days, whereas cartilage scaffolds loaded
To investigate the optimal concentration of BMP-2 and with 1 μg/g of FGF-18 could not achieve this concentration
FGF-18 for rBMSCs differentiation, ALP staining and retention (Figure 3J). These results demonstrated that BMP-
immunofluorescence staining of COL II were conducted, 2 could sustain osteogenic effects for 14 days, while FGF-
respectively. ALP staining revealed that the addition 18 could sustain chondrogenic effects for 21 days, thus
of BMP-2 could significantly enhance the osteogenic validating the time-controlled release function of BMP-2
differentiation of rBMSCs (Figure 3A). Specifically, after and FGF-18. Therefore, the optimal loading amount of
14 days of culture, the ALP staining area in the group BMP-2 and FGF-18 in subchondral scaffolds and cartilage
treated with 150 ng/mL BMP-2 was significantly larger scaffolds for improved osteochondral regeneration was set
than that treated with 0, 50, and 100 ng/mL BMP-2. In as 3 and 2 μg/g, respectively.
contrast, no significant difference was observed among Furthermore, to investigate the spatially controlled
the groups treated with 150 and 200 ng/mL BMP-2 and release of BMP-2 and FGF-18 in integrated osteochondral
the positive control group (Figure 3B). Furthermore, the scaffolds, the osteochondral scaffolds were encapsulated
immunofluorescence staining of COL II demonstrated with 5% GelMA gel to mimic the in vivo matrix
that the addition of FGF-18 could significantly enhance environment, which would contribute to the restricted
the chondrogenic differentiation of rBMSCs (Figure 3C). diffusion of GFs (Figure 4A). In the cartilage layer of the
Specifically, after 21 days of cultivation, the fluorescence osteochondral scaffold, the percentage release of FGF-18

Volume 10 Issue 5 (2024) 205 doi: 10.36922/ijb.3229