Optimization of photocrosslinkable resin components and 3D printing process parameters.

The role of 3D printing in the biomedical field is growing. In this context, photocrosslink-based 3D printing procedures for resorbable polymers stand out. Despite much work, more studies are needed on photocuring stereochemistry, new resin additives, new polymers and resin components. As part of these studies it is vital to present the logic used to optimize the amount of each resin constituent and how that effects printing process parameters. The present manuscript aims to analyze the effects of poly(propylene fumarate) (PPF) resin components and their effect on 3D printing process parameters. Diethyl fumarate (DEF), bisacylphosphine oxide (BAPO), Irgacure 784, 2-hydroxy-4-methoxybenzophenone (HMB) and, for the first time, in biomedical 3D printing, ethyl acetate (EA), were the resin components under investigation in this study. Regarding printing process parameters, Exposure Time, Voxel Depth, and Overcuring Depth were the parameters studied. Taguchi Design of Experiments was used to search for the effect of varying these resin constituent concentrations and 3D printing parameters on the curing behavior of 3D printable PPF resins. Our results indicate that resins with higher polymer cross-link density, especially those with a higher content of PPF, are able to be printed at higher voxel depth and with greater success (i.e., high yield). High voxel depth, as long as it does not sacrifice required resolution, is desirable as it speeds printing. Nevertheless, the overall process is governed by the correct setup of the voxel depth in relation to overcuring depth. In regards to resin biocompatibility, it was observed that EA is more effective than DEF, the material we had previously relied on. Our preliminary in vitro cytotoxicity tests indicate that the use of EA does not reduce scaffold biocompatibility as measured by standard cytotoxicity testing (i.e., ISO 10993-5). We demonstrate a workpath for resin constituent concentration optimization through thin film tests and photocrosslinkable process optimization. STATEMENT OF SIGNIFICANCE: We report here the results of a study of photo-crosslinkable polymer resin component optimization for the 3D printing of resorbable poly(propylene fumarate) (PPF) scaffolds. Resin additives are initially optimized for PPF thin film printing. Once those parameters have been optimized the 3D printing process parameters for PPF objects with complex, porous shapes can be optimized. The design of experiments to optimize both polymer thin films and complex porous resorbable polymer scaffolds is important as a guess and check, or in some cases a systematic method, are very likely to be too time consuming to accomplish. Previously unstudied resin components and process parameters are reported.

Screening of Additive Manufactured Scaffolds Designs for Triple Negative Breast Cancer 3D Cell Culture and Stem-Like Expansion.

Breast cancer stem cells (BCSCs) are tumor-initiating cells responsible for metastasis and tumor reappearance, but their research is limited by the impossibility to cultivate them in a monolayer culture. Scaffolds are three-dimensional (3D) cell culture systems which avoid problems related with culturing BCSC. However, a standardized scaffold for enhancing a BCSC population is still an open issue. The main aim of this study is to establish a suitable poly (lactic acid) (PLA) scaffold which will produce BCSC enrichment, thus allowing them to be studied. Different 3D printing parameters were analyzed using Taguchi experimental design methods. Several PLA scaffold architectures were manufactured using a Fused Filament Fabrication (FFF) 3D printer. They were then evaluated by cell proliferation assay and the configurations with the highest growth rates were subjected to BCSC quantification by ALDH activity. The design SS1 (0.2 mm layer height, 70% infill density, Zigzag infill pattern, 45° infill direction, and 100% flow) obtained the highest proliferation rate and was capable of enhancing a ALDH+ cell population compared to 2D cell culture. In conclusion, the data obtained endorse the PLA porous scaffold as useful for culturing breast cancer cells in a microenvironment similar to in vivo and increasing the numbers of BCSCs.

Design of a Scaffold Parameter Selection System with Additive Manufacturing for a Biomedical Cell Culture.

Open-source 3D printers mean objects can be quickly and efficiently produced. However, design and fabrication parameters need to be optimized to set up the correct printing procedure; a procedure in which the characteristics of the printing materials selected for use can also influence the process. This work focuses on optimizing the printing process of the open-source 3D extruder machine RepRap, which is used to manufacture poly(ε-caprolactone) (PCL) scaffolds for cell culture applications. PCL is a biocompatible polymer that is free of toxic dye and has been used to fabricate scaffolds, i.e., solid structures suitable for 3D cancer cell cultures. Scaffold cell culture has been described as enhancing cancer stem cell (CSC) populations related to tumor chemoresistance and/or their recurrence after chemotherapy. A RepRap BCN3D+ printer and 3 mm PCL wire were used to fabricate circular scaffolds. Design and fabrication parameters were first determined with SolidWorks and Slic3r software and subsequently optimized following a novel sequential flowchart. In the flowchart described here, the parameters were gradually optimized step by step, by taking several measurable variables of the resulting scaffolds into consideration to guarantee high-quality printing. Three deposition angles (45°, 60° and 90°) were fabricated and tested. MCF-7 breast carcinoma cells and NIH/3T3 murine fibroblasts were used to assess scaffold adequacy for 3D cell cultures. The 60° scaffolds were found to be suitable for the purpose. Therefore, PCL scaffolds fabricated via the flowchart optimization with a RepRap 3D printer could be used for 3D cell cultures and may boost CSCs to study new therapeutic treatments for this malignant population. Moreover, the flowchart defined here could represent a standard procedure for non-engineers (i.e., mainly physicians) when manufacturing new culture systems is required.