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3D Bioprinted Senescent Caco2 Modeling Using a Low-Cost DIY 3D Bioprinter: Challenges and Limitations
Sofian Al Shboul1, Yazan Al Dweiri2,3, Mai F AlSakhen4,5
1Department of Pharmacology and Public Health, Faculty of Medicine, The Hashemite University, Zarqa 13133, Jordan.
Background:
Therapy-induced senescence (TIS) is a major component of tumor cell response to a variety of anticancer treatments in preclinical models. Most TIS-related literature is derived from 2D-based studies with limited evidence in 3D models. This study aimed to investigate the challenges of developing an in-house (DIY) 3D bioprinted TIS cell model.
Materials And Methods:
A conventional thermoplastic extrusion-based 3D printer (Anet A8 kit) was modified to function as a bioprinter, enabling the fabrication of a Caco2 cell model via extrusion-based bioprinting. The printed 3D models were exposed to doxorubicin (DOX, at 1 and 10 μM), a well-established TIS trigger, and then assessed for senescence-associated β-galactosidase (SA-β-gal) activity and gene expression of several senescence regulators and components of the senescence-associated secretory phenotype (SASP) via real-time PCR.
Results:
Treated cells exhibited increased SA-β-gal staining and increased expression of CDKN2A and CDKN2B (and modestly TP53). Furthermore, the expression of SASP-related genes, including MMP3, MMP9, CXCL8, and TGF-β1 (but not IL-1A), was increased, albeit non-significantly, suggesting a potential opportunity to detect transcriptional SASP changes in 3D bioprinted tumor cell models.
Discussion:
This study provides an early attempt to develop a TIS 3D bioprinted tumor cell model, highlighting several challenges, including infrequent nozzle clogging, difficulty of image-based quantification of SA-β-gal staining, and inter-print variation in gene expression profiles.
Conclusion:
In-house DIY 3D bioprinting for investigating tumor cell senescence is feasible but requires further improvement prior to use in senolytic screening.
Insights
This study explored creating a 3D bioprinted model for therapy-induced senescence (TIS) in cancer cells. While feasible, challenges in DIY 3D bioprinting require refinement for senolytic drug screening.
Area of Science:
- Biomedical Engineering
- Cancer Research
- 3D Bioprinting Technology
Background:
- Therapy-induced senescence (TIS) is crucial in cancer treatment response, yet most studies use 2D models.
- Limited 3D models exist for TIS, hindering translation of preclinical findings.
- This research addresses the need for advanced 3D TIS models.
Purpose of the Study:
- To investigate the feasibility and challenges of developing an in-house, doxorubicin-induced TIS 3D bioprinted cell model.
- To assess key senescence markers and gene expression in a 3D bioprinted context.
- To evaluate the potential of 3D bioprinting for future senolytic drug screening.
Main Methods:
- A modified extrusion-based 3D printer was used to fabricate a Caco2 cell model.
- The 3D bioprinted models were treated with doxorubicin (DOX) to induce TIS.
- Senescence was assessed via SA-β-gal staining and real-time PCR for senescence regulators and SASP components.
Main Results:
- Doxorubicin treatment increased SA-β-gal activity and expression of senescence markers (CDKN2A, CDKN2B, TP53).
- Expression of SASP-related genes (MMP3, MMP9, CXCL8, TGF-β1) showed an increasing trend.
- The study identified potential for detecting transcriptional SASP changes in 3D bioprinted models.
Conclusions:
- Developing an in-house DIY 3D bioprinted TIS model presents challenges, including nozzle clogging and quantification difficulties.
- Inter-print variation in gene expression necessitates further optimization.
- DIY 3D bioprinting is a feasible approach for TIS research but needs improvement for senolytic screening.
