Speaker
Description
Introduction
Breast cancer continues to be one of the leading causes of cancer-related mortality among women worldwide [1,2]. Conventional 2D cultures and animal models fall short in accurately replicating the breast tumor microenvironment, often lacking translational relevance [3]. The development of three-dimensional (3D) in vitro models through hydrogel-based bioprinting offers a promising alternative to better mimic the mechanical, structural, and biological characteristics of tumor tissue [4]. This study aimed to design bioprintable alginate-gelatin (ALG-GEL) hydrogels and evaluate their suitability to serve as advanced 3D platforms for better mimicking the breast tumor microenvironment and enabling testing of cellular behaviors or therapeutic responses in a more physiologically relevant context compared to traditional 2D cultures.
Methods
ALG-GEL hydrogels were prepared mixing different concentrations of alginate and gelatin, to match the stiffness typical of breast tumor tissue (~10 kPa). Swelling behavior, degradation rate, and pH stability were monitored over a 21-day incubation at 37°C. Structural integrity was evaluated via scanning electron microscopy (SEM), and chemical composition was verified through Fourier-transform infrared spectroscopy (FTIR). Rheological analysis assessed the mechanical properties of the hydrogels, while pre-crosslinking printability studies helped determine suitable compositions for bioprinting. MDA-MB-231 breast cancer cells were embedded within bioprinted constructs. Cellular viability and metabolic activity were assessed using CCK8 assays over 21 days. Confocal laser scanning microscopy with live/dead staining was used to confirm cell viability and spatial distribution within the constructs.
Results
All hydrogel formulations exhibited high initial swelling within 2 hours, followed by a controlled degradation profile over 21 days. FTIR confirmed the successful incorporation of gelatin without compromising the alginate backbone. SEM analysis revealed a well-interconnected and homogeneous microstructure. Rheological measurements demonstrated a storage modulus (G′) close to 10 kPa, suitable for mimicking tumor tissue stiffness. Printability studies identified optimal ALG-GEL ratios that ensured structural fidelity and cell compatibility. Importantly, bioprinted constructs showed a marked increase in metabolic activity from day 1 to day 21, suggesting robust cell proliferation. Confocal microscopy confirmed high cell viability and a uniformly distributed cell population throughout the 3D constructs.
Discussion
The combination of structural, mechanical, and biochemical evaluations demonstrated that ALG-GEL hydrogels are highly suitable for biofabrication of tumor-mimetic scaffolds. The progressive increase in metabolic activity suggests that these hydrogels effectively support cell proliferation over prolonged periods. Moreover, the confocal microscopy findings reinforce the scaffold's ability to provide a uniform 3D niche for cell survival and growth. These features are critical for the development of reliable in vitro breast cancer models that may reduce reliance on animal testing and offer new insights into tumor progression and treatment response.
References
[1] M. Arnold, doi.org/10.1016/j.breast.2022.08.010
[2] K. Barzaman, doi.org/10.1016/j.intimp.2020.106535
[3] M. Kapałczyńska, doi.org/10.5114/aoms.2016.63743
[4] A. Guller, doi.org/10.3390/bioengineering10010017
Acknowledgments
The authors acknowledge the support of the Interuniversity Center for the Promotion of the 3Rs Principles and the Nanotechnology Lab at Istituti Clinici Scientifici Maugeri IRCCS and the PNRR program.
Disclosure Information
The authors declare no conflicts of interest.
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