Page 459 - IJB-10-6

International Journal of Bioprinting 3D-printed scaffold for biomolecule delivery

Previous studies have demonstrated how multilayer that the adherent stem cells on the GS scaffolds enhanced
coatings reduce burst release and offer better control of the expansion and formation of filopodia compared to
GF release kinetics. Multilayered poly(lactic-co-glycolic the cells on pristine PCL struts. The number of cells that
acid) (PLGA) microparticle systems have been reported to initially adhered to the GS-coated scaffolds is presented
significantly reduce the burst release of BMP-2, leading to in Figure 5b. The following in vitro experiments were
better-controlled delivery profiles. Similarly, electrospun performed using bilayer scaffolds divided into four groups:
28
nanofibrous scaffolds have been used to achieve biphasic no GFs (control), bFGF only (w/o/bFGF), BMP2 only
release profiles, with an initial burst followed by sustained (BMP2/w/o), and both bFGF and BMP2 (BMP2/bFGF).
release, effectively maintaining the bioactivity of GFs. 44,45 The stem cells were cultured for 24 h, and mitochondrial
In comparison to single-layer systems, the double-layer metabolism was assessed using the CCK-8 assay. The
coating in our study provides enhanced control over the optical density was converted into a standard curve, which
sequential release of bFGF and BMP-2, which is crucial was assessed using the cell number. A significant difference
for orchestrating different stages of tissue regeneration. in the number of adherent cells was observed among the
Soaking the hybrid containing GFs onto the hydrolyzed four groups. The groups containing bFGF reported higher
scaffolds resulted in a coating layer, and the use of the cell adherence than the control and BMP2-only groups.
scaffold facilitated the construction of multiple layers. The Figure 5c indicates that cell proliferation significantly
highly viscous hybrid was well-fixed on the hydrolyzed increased in cells on the scaffolds containing bFGF in the
polymer surface, and the loading efficiency of the GFs was second layer after 14 days compared to that in the control
enhanced. In contrast to previous studies, coating with or BMP-2-only samples. The osteogenic differentiation
GS has the potential for GF delivery due to the decrease of stem cells induced on the scaffolds for seven days was
in initial burst release, increase in loading capacity, the established based on ALP activity. The ALP activity of
possibility of dual-or multipronged delivery, and sustained the stem cells was higher in the bilayer containing both
release of GFs. BMP-2 and bFGF (Figure 5d). Additionally, ALP activity
was not significantly different in the BMP2-only groups,
As demonstrated in our study, both the PCL scaffold but a significant difference was observed in the other
and GS composite layers critically influence the release groups. The osteogenic capacity of the bilayer containing
kinetics of BMP-2 and bFGF. The dual-layer coating both BMP-2 and bFGF was observed through Alizarin
enables sequential release, where BMP-2 is released from Red S staining and an assay for the detection of mineral
the inner layer and bFGF from the outer layer, with the deposition (Figure 5e). Mineralization was significantly
release profile governed by diffusion barriers dependent on higher than in the other groups.
the thickness and density of the layers. This observation
aligns with previous studies, such as Lee et al., which Biomimetic or functional matrices on the scaffold
36
reported that thicker composite layers slow down have been engineered to stimulate and modulate stem cell
release kinetics. Therefore, the thickness of the coating behaviors, such as cell adhesion, proliferation, migration,
materials, in particular, plays a vital role in controlling and differentiation. 2,17,26 In particular, the attachment
the spatiotemporal release of biomolecules. Additionally, of stem cells to the artificial surface is related to various
while the release profiles of bFGF and BMP-2 were stable biological processes associated with subsequent stages
over the 42-day period, any potential uneven release and influences intracellular signaling pathways, including
could be attributed to factors such as variations in coating phenotype, stemness, growth, and differentiation.
thickness or localized differences in scaffold porosity. Therefore, research on the initial adhesion and mechanisms
To mitigate these issues, the fabrication process was of stem cells is of continuous interest in tissue engineering.
optimized to ensure uniform coating and consistent GF Generally, physical and chemical strategies are employed
loading. Similar studies have demonstrated that adjusting for the surface modification of biomaterials. Notably,
the ratio of gelatin to silica can fine-tune the release rates. geometrical engineering strategies at the nano- or micro-
46
In future research, we plan to investigate further the effects scale, including specific patterns, height and width of
of coating thickness, bonding, and variability to address pillars, size or shape of deposited particles, and roughness,
potential challenges associated with uneven release. have been induced to modulate stem cell behaviors.
Mimicking extracellular matrices, such as fibronectin,
3.4. Cellular responses of rat bone marrow-derived laminin, poly-D, L-lysine, and collagen, has also been
mesenchymal cells on the scaffold used for chemical surface modification. In a previous
To observe the morphology of the initially adherent cells, study, engineered bFGF was reportedly involved in cell
fluorescent images of stem cells cultured for 2 h were adhesion, i.e., heparan sulfate proteoglycan-mediated
captured using CLSM. The results in Figure 5a indicate adhesion and phosphorylation of focal adhesion kinase.

Volume 10 Issue 6 (2024) 451 doi: 10.36922/ijb.4638