Fibroblasts that are present in the stroma secrete glycoproteins and polysaccharides to form extracellular matrix. They maintain the structural integrity within the connective tissue and play a key role in cancer, chronic obstructive pulmonary diseases, idiopathic pulmonary fibrosis, scleroderma and several other diseases.
Fibroblasts grown in 3D environments express natural cell physiology closer to in-vivo conditions than their counterpart cultured in 2D environments. TrueGel3D™ hydrogel is a robust, chemically defined hydrogel made up of biologically inert, synthetic biopolymers that can be customized to match the native cell environment. The objective of this study is to investigate the fibroblast spreading within TrueGel3D™ hydrogel when crosslinked with CD cell-degradable and PEG non cell-degradable crosslinkers.
TrueGel3D™ hydrogel preparation:
All steps were performed in a sterile hood and the volume ratio of each component was added as indicated below.
Chemical fixation and confocal microscopy:
3T3 fibroblasts grown in hydrogels with PEG non cell-degradable crosslinker appeared round and in tightly packed aggregates, while cells grown in hydrogels with CD-cell degradable crossinker spread locally.
Figure 1.3T3 fibroblasts in TrueGel3D™ hydrogels made of FAST-PVA polymer modified with TrueGel3D™ RGD integrin adhesion peptide and crosslinked with either PEG non cell-degradable (left) or CD cell-degradable crosslinker (right). Pictures show collapsed stacks of confocal frames representing a height of 300 µm of the gel. Red: nuclei; green: actin cytoskeleton. Scale bar: 200 µm
Trugel3D™ hydrogels consist of small pores at a size of only several nanometers; 3T3 fibroblasts grown in hydrogel with CD cell-degradable crosslinker spread and migrated within the gel by breaking the matrix metalloproteases (MMP) cleavable peptides included in the CD degradable crosslinker.
Fibroblasts grown in a 3D environment showed an increased level of paracrine factors1, in addition TNF receptor-1 expression and NF-kB activation2 levels were closer to in-vivo conditions. The 3D culture systems like TrueGel3D™ hydrogels present a more sensitive and biologically relevant tool for investigating various diseases involving fibroblasts.
TrueGel3D™ hydrogels are also compatible with other cell types (immune cells, cancer cell lines, epithelial cell lines, stem cells) and can be customized to study cell migration/invasion, endothelial transmigration, cell adhesion and cell differentiation.