Chondrocyte Response to High Regimens of Cyclic Hydrostatic Pressure in 3-Dimensional Engineered Constructs

Abstract

Purpose Despite widespread use of 3-dimensional (3D) micro-porous scaffolds to promote their potential application in cartilage tissue engineering, only a few studies have examined the response to hydrostatic pressure of engineered constructs. A high cyclic pressurization, currently believed to be the predominant mechanical signal perceived by cells in articular cartilage, was used here to stimulate bovine articular chondrocytes cultured in a synthetic 3D porous scaffold (DegraPol). Methods Construct cultivation lasted 3 days with applied pressurization cycles of amplitude 10 MPa, frequency 0.33 Hz, and stimulation sessions of 4 hours/day. Results At 3 days of culture, with respect to pre-culture conditions, the viability of the pressurized constructs did not vary, whereas it underwent a 16% drop in the unpressurized controls. Synthesis of α-actin was 34% lower in all cultured constructs. Synthesis of collagen II/collagen I did not vary in pressurized constructs, was 76% lower in unpressurized controls, and was around 230% higher in pressurized constructs with respect to unpressurized controls. Chondrocytes showed a phenotypic spherical morphology at time zero and at 3 days of pressurized culture. Conclusions Although the passage from 2D expansion to 3D geometry was effective to guide cell differentiation, only mechanical conditioning enabled the maintenance and further cell differentiation toward a mature chondrocytic phenotype.

Bioreactor mechanically guided 3D mesenchymal stem cell chondrogenesis using a biocompatible novel thermo-reversible methylcellulose-based hydrogel

“…Finally, COL 10 was detected in loaded specimens (C, lower panel, in green) but in a lower amount in comparison with COL 2, thus confirming that cells were not adopting a hypertrophic phenotype due to the continuous mechanical stimulation provided by daily bioreactor operating. These data are in accordance to literature as compressive stress combined with shear flow allows a higher MSC commitment11333442. Hence, IF staining confirmed the PCR data, providing further important indication of the successful bioreactor-guided chondrogenesis.…”

“…Thus, although cells are metabolically active (as confirmed by PCR findings and increased GAG synthesis), they seem to lack the appropriate environment or proper stimuli to produce a structured extracellular matrix within the timeframe of this experiment9. Moreover, by using fibrin glue as cells matrix in a newly developing construct, the bulk of the newly synthesized GAG was released into the culture medium as the pericellular matrix was not sufficiently developed to retain the newly produced GAG323334. Therefore, the MC hydrogel seems to act similarly to fibrin according to obtained and previous results.…”

Comparative chondrogenesis of human cells in a 3D integrated experimental–computational mechanobiology model

“…These stimulation levels were chosen because they were demonstrated to be able to promote expression of the chondrocyte phenotype in tissue engineering applications (Raimondi and Pietrabissa 2005;Raimondi et al 2008;Candiani et al 2008). Furthermore, the intermittent daily regime of the pressurisation cycles was based on previous experiments conducted with the same bioreactor system , in which was found evidence that this regime induces chondrogenesis.…”

Physical Stimulation of Chondrogenic Cells In Vitro: A Review

“…Others have found that in Type I collagen gel, under 40 KPa at 0.0125 Hz, Type II collagen and aggrecan were upregulated at the gene expression level but not at the protein level for human chondrocytes cultured for 14 days [28]. Finally, the phenotypic and proteomic expression pattern of chondrocytes seeded in a polyurethane scaffold (Degrapol1, Abmedica, Italy) with HP of 10 MPa at 0.33 Hz 4 hours per day was similar to those from nonloaded cells, although the maintenance of cell viability was improved after 3 days [13].…”