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Three-dimensional (3D) printing was utilized for the fabrication of a composite scaffold of poly(ε-caprolactone) (PCL) and calcium magnesium phosphate (CMP) bioceramics for bone tissue engineering application. Four groups of scaffolds, that is, PMC-0, PMC-5, PMC-10, and PMC-15, were fabricated using a custom 3D printer. Rheology analysis, surface morphology, and wettability of the scaffolds were characterized. The PMC-0 scaffolds displayed a smoother surface texture and an increase in the ceramic content of the composite scaffolds exhibited a rougher structure. The hydrophilicity of the composite scaffold was significantly enhanced compared to the control PMC-0. The effect of ceramic content on the bioactivity of fibroblast NIH/3T3 cells in the composite scaffold was investigated. Cell viability and toxicity studies were evaluated by comparing results from lactate dehydrogenase (LDH) and Alamar Blue (AB) colorimetric assays, respectively. The live-dead cell assay illustrated the biocompatibility of the tested samples with more than 100% of live cells on day 3 compared to the control one. The LDH release indicated that the composite scaffolds improved cell attachment and proliferation. In this research, the fabrication of a customized composite 3D scaffold not only mimics the rough textured architecture, porosity, and chemical composition of natural bone tissue matrices but also serves as a source for soluble ions of calcium and magnesium that are favorable for bone cells to grow. These 3D-printed scaffolds thus provide a desirable microenvironment to facilitate biomineralization and could be a new effective approach for preparing constructs suitable for bone tissue engineering.
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Author Correction: Novel stereological method for estimation of cell counts in 3D collagen scaffolds
Current methods for assessing cell proliferation in 3D scaffolds rely on changes in metabolic activity or total DNA, however, direct quantification of cell number in 3D scaffolds remains a challenge. To address this issue, we developed an unbiased stereology approach that uses systematic-random sampling and thin focal-plane optical sectioning of the scaffolds followed by estimation of total cell number (StereoCount). This approach was validated against an indirect method for measuring the total DNA (DNA content); and the Bürker counting chamber, the current reference method for quantifying cell number. We assessed the total cell number for cell seeding density (cells per unit volume) across four values and compared the methods in terms of accuracy, ease-of-use and time demands. The accuracy of StereoCount markedly outperformed the DNA content for cases with ~ 10,000 and ~ 125,000 cells/scaffold. For cases with ~ 250,000 and ~ 375,000 cells/scaffold both StereoCount and DNA content showed lower accuracy than the Bürker but did not differ from each other. In terms of ease-of-use, there was a strong advantage for the StereoCount due to output in terms of absolute cell numbers along with the possibility for an overview of cell distribution and future use of automation for high throughput analysis. Taking together, the StereoCount method is an efficient approach for direct cell quantification in 3D collagen scaffolds. Its major benefit is that automated StereoCount could accelerate research using 3D scaffolds focused on drug discovery for a wide variety of human diseases.
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- Award ID(s):
- 1926990
- PAR ID:
- 10478141
- Publisher / Repository:
- Sci Rep
- Date Published:
- Journal Name:
- Scientific Reports
- Volume:
- 13
- Issue:
- 1
- ISSN:
- 2045-2322
- Subject(s) / Keyword(s):
- Stereology Cell Proliferation Collagen Tissue Engineering Tissue Scaffolds*
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1038/s41598-023-37265-z
