A recent article published in Applied Surface Science Advances describes the bioprinting of nanostructured patches composed of sodium alginate (NaAlg), polyvinyl alcohol (PVA), and indocyanine green (ICG), either freely dispersed or encapsulated in liposomes.
Three formulations were evaluated using scanning electron microscopy (SEM) and atomic force microscopy (AFM) to investigate their surface morphology and structural characteristics.
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Background
NaAlg and PVA composites are commonly used in tissue regeneration due to their biocompatibility and complementary properties. When combined, they form stable hydrogels suitable for biomedical use. Incorporating ICG enhances these systems by adding optical functionality. Encapsulation of ICG in liposomes further improves stability, enabling controlled release and real-time monitoring of material degradation via near-infrared fluorescence.
The interface between the patch and host tissue plays a critical role in regenerative outcomes. Therefore, tuning the microstructure through material processing is essential. To study these effects, three patch types were fabricated: NaAlg/PVA (PS0), NaAlg/PVA with free ICG (PS1), and NaAlg/PVA with liposome-encapsulated ICG (PS2).
Methods
Liposomes were prepared using L-α-phosphatidylcholine via the standard reverse-phase method. Particle size and size distribution were characterized using dynamic light scattering.
Three gel formulations were prepared for bioprinting: NaAlg/PVA (S0), NaAlg/PVA with free ICG (S1), and NaAlg/PVA with liposome-encapsulated ICG (S2). These were used to produce the corresponding patches: PS0, PS1, and PS2.
Patches containing ICG (PS1 and PS2) were visibly colored due to the dye. For S0, aqueous solutions of NaAlg and PVA were prepared separately in ultra-pure water and then combined. In S1 and S2, free ICG solution or liposomal ICG dispersions were added to the S0 mixture.
Patch designs were created using Autodesk Inventor Pro, modeled as square-based parallelepipeds with 10 mm sides and a height of 0.4 mm. The patches were printed using a BIOX Bioprinter and then air-dried for 48 hours. To protect the dye from light degradation, dyed samples were kept in the dark during this period. After drying, all patches were immersed in a 500 μM CaCl2 aqueous solution for one minute to complete crosslinking.
Surface morphology was characterized using SEM and AFM. Optical properties, including reflectance and transmittance, were evaluated with a fiber-based setup using a broadband light source covering the 500–2500 nm wavelength range.
Results and Discussion
The differing surface tensions of PVA and NaAlg contributed to anisotropic contraction of the patch following ion exposure. When additional chemical species were incorporated during fabrication, the bipolymeric composite underwent morphological restructuring. As a result, the patches exhibited distinct physicochemical characteristics and variable responses to crosslinking with CaCl2.
SEM analysis showed that crosslinking with CaCl2 affected surface topography, leading to the formation of aggregates, pores, and regular asperities. These surface features were quantified using AFM, which revealed differences in surface roughness. PS1 showed fewer and smaller pores than PS0, while PS2 exhibited the lowest overall pore density and area.
Fine-scale scans also captured liposome aggregation on the patch surface. Although liposomes formed clusters between 140 and 230 nm, individual vesicles remained intact at approximately 120 nm in diameter, consistent with their size in aqueous dispersion. The inclusion of liposomes increased surface roughness by approximately 5.8 %.
In contrast, patches containing either free or encapsulated ICG showed reduced surface roughness. This was attributed to interactions between ICG and the polymer matrix, which appeared to moderate the contraction induced by crosslinking.
Differences in the optical properties of PS1 and PS2 were relevant for tuning the patches for specific diagnostic or therapeutic use. Reflectance spectra indicated dye aggregation, which altered how the patches interacted with light.
In PS1, aggregation caused a red shift of approximately 20 nm in the absorption spectrum. This spectral broadening and shift may reduce precision in light absorption, which could influence the patch’s performance in photodynamic therapy or imaging applications.
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Conclusion
The study demonstrated the successful bioprinting of NaAlg/PVA-based patches containing either free or liposome-encapsulated ICG. Morphological analysis highlighted how both crosslinking and dye encapsulation influence surface roughness and structural organization. Optical measurements confirmed that liposomes preserve the photophysical properties of ICG and reduce aggregation.
These findings support the use of liposomal encapsulation to improve the functional stability of ICG-containing biomaterials. The ability to tune surface morphology and optical behavior positions these patches as promising candidates for applications in diagnostics, drug delivery, and regenerative medicine.
Journal Reference
de Nigris, A., Quero, G., Vanoli, G. P., & Ambrosone, L. (2025). Surface properties of nanostructured alginate-based bioprinted patches. Applied Surface Science Advances, 27, 100739. DOI: 10.1016/j.apsadv.2025.100739, https://www.sciencedirect.com/science/article/pii/S2666523925000479
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