Adipose-derived stromal cells increase the formation of collagens through paracrine and juxtacrine mechanisms in a fibroblast co-culture model utilizing macromolecular crowding

Abstract

Background Adipose-derived stromal cells (ASCs) possess a multitude of regenerative capabilities, which include immunomodulation, angiogenesis, and stimulation of extracellular matrix (ECM) remodeling. However, the underlying mechanisms leading to ECM remodeling remain largely elusive and highlight the need for functional in vitro models for mode of action studies. Therefore, the purpose of this study was to develop an in vitro co-culture model to investigate the capabilities of ASCs to modulate fibroblasts and ECM. Methods An ECM in vitro model with ASCs and normal human dermal fibroblasts (NHDFs) was established utilizing macromolecular crowding, ascorbic acid, and TGF-β stimulation. Paracrine and juxtacrine co-cultures were created using transwell inserts and cell cultures with direct cell–cell contacts. The cultures were screened using RT2 PCR Profiler Arrays; the protein levels of myofibroblast differentiation marker alpha smooth muscle actin (αSMA) and ECM remodeling enzymes were analyzed using western blot on cell lysates; the formation of collagen type I, III, VI, and fibronectin was investigated using ELISA on culture supernatants; and the deposition of collagens was analyzed using immunocytochemistry. Results TGF-β stimulation of NHDF monocultures increased the expression of 18 transcripts relevant for ECM formation and remodeling, the protein levels of αSMA and matrix metalloproteinase-2 (MMP-2), the formation of collagen type I, III, VI, and fibronectin, and the deposition of collagen type I and VI and decreased the protein levels of MMP-14. Inclusion of ASCs in the ECM co-culture model increased the formation of collagen type I and III through paracrine mechanisms and the formation of collagen type VI through juxtacrine mechanisms. Conclusions The co-culture model provides effective stimulation of NHDF monocultures by TGF-β for enhanced formation and deposition of ECM. In the model, ASCs induce changes in ECM by increasing formation of collagen type I, III and VI. The obtained results could guide further investigations of ASCs’ capabilities and underlying mechanisms related to ECM formation and remodeling.

Advancements in adipose-derived stem cell therapy for skin fibrosis

“…In another study, ASC-CM reduced the synthesis of collagen and the expression of connective tissue growth factor, fibronectin, and α-SMA[ 65 ]. However, in a coculture model of ASCs and normal human dermal fibroblasts, ASCs increased the formation of collagen types I, III, and VI in the ECM[ 66 ]. It appears that ASCs could target abnormal fibroblasts and reduce pathological deposition of ECM.…”

Unraveling White Adipose Tissue Heterogeneity and Obesity by Adipose Stem/Stromal Cell Biology and 3D Culture Models

“…Auxenfans et al [ 69 ] also demonstrated that a 3D-reconstructed skin model co-cultured with ASCs increased thickness of the dermis, specifically the papillary dermis. In direct juxtacrine co-culture and in indirect paracrine co-culture, ASCs improved collagen maturation and metalloproteinase secretion compared to monoculture [ 70 ].…”

Potential of Estradiol‐Functionalized Polyisocyanide Hydrogels for Stimulating Tissue Regeneration of the Pelvic Floor

“…[ 18 ] Next, we investigated the bioactivity of PIC‐E2 on adipose‐derived stem cells (ASCs), because they are often used for various tissue engineering applications including POP; [ 19 ] ASCs possess various regenerative capabilities including stimulation of ECM production and their angiogenic and immunomodulatory potential. [ 20 ] We hypothesize that the combination of PIC‐E2 and ASCs may have a synergistic therapeutic effect on the regeneration of the pelvic floor in vivo and, hence, may improve surgical outcomes of POP.…”