Molecular Changes Induced in Melanoma by Cell Culturing in 3D Alginate Hydrogels

Abstract

Alginate hydrogels have been used as a biomaterial for 3D culturing for several years. Here, gene expression patterns in melanoma cells cultivated in 3D alginate are compared to 2D cultures. It is well-known that 2D cell culture is not resembling the complex in vivo situation well. However, the use of very intricate 3D models does not allow performing high-throughput screening and analysis is highly complex. 3D cell culture strategies in hydrogels will better mimic the in vivo situation while they maintain feasibility for large-scale analysis. As alginate is an easy-to-use material and due to its favorable properties, it is commonly applied as a bioink component in the growing field of cell encapsulation and biofabrication. Yet, only a little information about the transcriptome in 3D cultures in hydrogels like alginate is available. In this study, changes in the transcriptome based on RNA-Seq data by cultivating melanoma cells in 3D alginate are analyzed and reveal marked changes compared to cells cultured on usual 2D tissue culture plastic. Deregulated genes represent valuable cues to signaling pathways and molecules affected by the culture method. Using this as a model system for tumor cell plasticity and heterogeneity, EGR1 is determined to play an important role in melanoma progression.

Direct early growth response-1 knockdown decreases melanoma viability independent of mitogen-activated extracellular signal-related kinase inhibition

“…From a clinical perspective, Kappelmann-Fenzl et al investigated EGR1-stained tissue sections, finding that EGR1 could be detected in most melanoma cells [31]. The authors further noted an absence of EGR1 protein expression in benign melanocytes, weak cytoplasmic staining in melanoma cells from primary tumor, and intense nuclear staining in melanoma metastasis, suggesting an association between increasing EGR1 expression and melanoma progression [31]. Kanemaru et al similarly found increased EGR1 expression in tumor tissues from patients with invasive melanoma compared to tissues of noninvasive melanoma [32].…”

The influence of viscosity of hydrogels on the spreading and migration of cells in 3D bioprinted skin cancer models

“…The gene early growth response 1 (EGR1) was the most notable upregulated gene in melanoma, with a 3-fold increase in mRNA expression compared to primary melanocytes. This highlighted the importance of EGR1 in melanoma progression ( Kappelmann-Fenzl et al, 2021 ).…”

Transcription factor activating enhancer-binding protein 2ε (AP2ε) modulates phenotypic plasticity and progression of malignant melanoma

“…To confirm that the isolated cells are from melanocytic origin, GRM1 expression was verified by RT-PCR, as previously described [ 23 ] (Supplementary Figure S1 ). Human melanoma cell lines Mel Juso, 501Mel, and Mel Im (FUCCI) were cultivated as indicated elsewhere [ 24 , 25 ]. Mycoplasma contamination was regularly excluded for all primary cells and cell lines.…”

“…The FUCCI (fluorescence ubiquitination-based cell cycle indicator)-labelled Mel Im cells were generated and quantified as described previously [ 24 ].…”