Thermal Bioprinting Causes Ample Alterations of Expression of LUCAT1, IL6, CCL26, and NRN1L Genes and Massive Phosphorylation of Critical Oncogenic Drug Resistance Pathways in Breast Cancer Cells
Thermal Bioprinting Causes Ample Alterations of Expression of LUCAT1, IL6, CCL26, and NRN1L Genes and Massive Phosphorylation of Critical Oncogenic Drug Resistance Pathways in Breast Cancer Cells
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DOI：
10.3389/fbioe.2020.00082
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发表时间：
2020-02-21
影响因子：
5.7
通讯作者：
Boland, Thomas
中科院分区：
文献类型：
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作者：
Campbell, Aleli;Mohl, Jonathon E.;Boland, Thomas
生物打印技术将工程学和生物学领域结合在一起，它们具有巨大的转化潜力，可以极大地影响再生医学和药物发现的未来。然而，热喷墨生物打印在乳腺癌细胞中引发的分子效应仍然难以捉摸。先前的研究表明，生物打印可以用于药物发现和药理学的组织模型。我们报道了生物打印的MCF7乳腺癌细胞在生物打印后不同时间点的活力、凋亡、磷酸化和RNA序列分析。Annexin A5-FITC细胞凋亡染色联合流式细胞术于生物打印后2和24 h进行。生物打印24小时后，使用Human phospho-MAPK阵列试剂盒进行抗体阵列。对生物打印后2、7、24小时采集的样品进行RNA序列分析。打印后24 h和48 h细胞存活率平均值分别为77%和76%，打印后2 h和24 h细胞凋亡率分别为31%和64%。在生物打印的细胞中，共有21个激酶被磷酸化，而在人工播种的对照中，有9个激酶被磷酸化。对生物打印细胞进行RNA序列分析，共鉴定出12235个基因，其中9.7%存在显著差异表达。以+/- 2倍变化为截断点，在生物打印细胞中观察到266个上调基因和206个下调基因，其中以下5个基因唯一表达NRN1L、LUCAT1、IL6、CCL26和LOC401585。这表明，热喷墨生物打印正在刺激大规模的基因改变，可能用于药物发现。此外，生物打印激活了涉及耐药、细胞运动、增殖、存活和分化的关键途径。
Bioprinting technology merges engineering and biological fields and together, they possess a great translational potential, which can tremendously impact the future of regenerative medicine and drug discovery. However, the molecular effects elicited by thermal inkjet bioprinting in breast cancer cells remains elusive. Previous studies have suggested that bioprinting can be used to model tissues for drug discovery and pharmacology. We report viability, apoptosis, phosphorylation, and RNA sequence analysis of bioprinted MCF7 breast cancer cells at separate timepoints post-bioprinting. An Annexin A5-FITC apoptosis stain was used in combination with flow cytometry at 2 and 24 h post-bioprinting. Antibody arrays using a Human phospho-MAPK array kit was performed 24 h post-bioprinting. RNA sequence analysis was conducted in samples collected at 2, 7, and 24 h post-bioprinting. The post-bioprinting cell viability averages were 77 and 76% at 24 h and 48 h, with 31 and 64% apoptotic cells at 2 and 24 h after bioprinting. A total of 21 kinases were phosphorylated in the bioprinted cells and 9 were phosphorylated in the manually seeded controls. The RNA seq analysis in the bioprinted cells identified a total of 12,235 genes, of which 9.7% were significantly differentially expressed. Using a +/- 2-fold change as the cutoff, 266 upregulated and 206 downregulated genes were observed in the bioprinted cells, with the following 5 genes uniquely expressed NRN1L, LUCAT1, IL6, CCL26, and LOC401585. This suggests that thermal inkjet bioprinting is stimulating large scale gene alterations that could potentially be utilized for drug discovery. Moreover, bioprinting activates key pathways implicated in drug resistance, cell motility, proliferation, survival, and differentiation.