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

Abstract

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.

The long noncoding RNA LUCAT1 promotes colorectal cancer cell proliferation by antagonizing Nucleolin to regulate MYC expression

“…After knocking out LUCAT1 , CRC cell proliferation was significantly inhibited. This is also consistent with the results of studies in cervical cancer, ovarian cancer, breast cancer, choroidal melanoma, clear cell renal cell carcinoma, non-small-cell lung cancer and esophageal squamous cell carcinoma, showing that loss of LUCAT1 hinders cell proliferation 15 , 17 , 20 , 39 , 41 – 43 . The results described above indicate the oncogenic role of LUCAT1 in CRC.…”

Guiding Lights: Tissue Bioprinting Using Photoactivated Materials

“…[383][384][385] Recently, thermal inkjet printing was also associated with upregulation of oncogenic pathways, increased phosphorylation of kinases and large-scale alternation in gene expression of breast cancer cells. 386 Piezoelectric inkjet printing can also lead to mechanical stress-induced cell death. In particular, printing at high power and/or high vibration frequencies can cause damage to the cell membranes.…”

3D Bioprinted cancer models: Revolutionizing personalized cancer therapy

“…This offers promise for developing and screening personalized tumor anti-lymphangiogenic therapies in the future. Recent investigation suggested that the gene alterations during bioprinting are evident demonstrated by changes in the expression of LUCAT1, IL6, CCL26, and NRN1L genes and phosphorylation of critical oncogenic drug resistance pathways in breast cancer cells in thermal bioprinted constructs [135] . Understanding such changes at the molecular level in bioprinted constructs would help to develop models that mimic human tumor tissues.…”