Influence of Matrigel on Single- and Multiple-Spheroid Cultures in Breast Cancer Research

Abstract

This study aimed to develop and compare single and multiple 3D models such as multicellular tumor spheroids and to investigate the influence of Matrigel on their morphological and functional behavior. MDA-MB-231 3D models were generated in the presence and absence of Matrigel and their key biological properties within 6 days of culture were monitored. Our results revealed the formation of well-defined 3D models in the presence of Matrigel, with a uniform morphology, increased diameter, good circularity, and increased expression of a proliferation marker (PCNA). In comparison, 3D models generated without Matrigel were characterized by an irregular border, reduced dimensions and circularity, and a decrease of PCNA expression. Similarities between the single and multiple 3D cultures were found in their viability, Nrf2 expression, and glutathione (GSH) content. The influence of Matrigel on MDA-MB-231 spheroids metabolism under hypoxic conditions was highlighted by released lactate dehydrogenase and nitric oxide, GSH levels and expression of Nrf2 and Hsp70 proteins. Based on the increased expression of PCNA and the development of the hypoxia process in the presence of extracellular matrix, our study showed that the addition of Matrigel improves the growing environment of tumor spheroids, making it closer to that of in vivo tumor conditions.

Fibroblasts Accelerate Formation and Improve Reproducibility of 3D Cellular Structures Printed with Magnetic Assistance

“…MDA-MB-231, a human epithelial triple-negative metastatic breast cancer cell line, is notorious for being difficult to grow in 3D [4]. Efforts to form 3D cellular structures with MDA-MB-231 often incorporate biologically based extracellular matrix (ECM) constituents, such as recombinant basement membrane (rBM) [1] or Matrigel [5,6].…”

“…These cells display stellate morphologies when grown in 3D in the presence of an extracellular matrix, indicating a malignant phenotype [7,8]. Matrigel, which promotes intercellular interactions for cell agglomeration [6], is derived from mouse Englebreth-Holm-Swarm tumor [9] and contains a mixture of ECM proteins and growth factors [6,9]. It is used in numerous scaffold-based 3D culture models [10][11][12][13][14].…”

“…In addition, it does not appropriately represent a human microenvironment due to its murine-derived origin [18]. Regulating the formation of these 3D cellular structures is critical for a drug discovery process [6,16,19,20].…”

“…The use of scaffold-free aggregations of tumor cells is an appropriate model for cancer research [2]. Adherent or anchorage-dependent cells cultured on an ultralow attachment (ULA) surface undergo spontaneous agglomeration, referred to as the forced floating [21] or liquid overlay technique [1,6]. On a flat ULA surface, this forced floating results in the formation of numerous 3D cellular structures of variable dimensions [2], limiting the reproducibility of the desired 3D model.…”

Bioengineering a novel 3D in-vitro model to recreate physiological oxygen levels and tumor-immune interactions

“…In order to overcome these limitations, relevant three-dimensional (3D) culture models have been studied. A wide array of matrices, including synthetic and natural, have been developed to recapitulate key features of the TME (36)(37)(38)(39)(40)(41). While biochemical and physical parameters, such as conduciveness to key biochemical signals, stiffness, degradability, permeability to nutrients, diffusibility to gases and swelling indices have been heavily studied (42)(43)(44)(45)(46)(47)(48), recreation of physiological oxygen levels and their influence on tumor-immune interactions remains fairly under investigated.…”