Promoted Viability and Differentiated Phenotype of Cultured Chondrocytes With Low Level Laser Irradiation Potentiate Efficacious Cells for Therapeutics.

Xiaohong Yang,Shaojie Liu,Peihong Liang,Timon Chengyi Liu,Shuliang Cui,Weicong Zhu,Honglin Li,Suihui Ye

doi:10.3389/fbioe.2020.00468

Abstract

Effective clinical treatments of cartilage lesions in affected joints require large numbers of viable chondrogenic cells generated through in vivo stimulation or ex vivo expansion of chondrocytes isolated from small biopsy specimens. Conventional passaging of chondrocytes in culture provides sufficient cells for treatments but these cells usually lose their differentiated phenotype. This leads to the formation of fibrocartilaginous tissue due to a malfunctioning repair process. Biostimulation of passaging chondrocytes with low level laser irradiation (LLLI) may theoretically produce more functional chondrocytes for cell-based repair of cartilage defects. Molecular and cellular analyses, cytochemistry, cell cultivation, and microscopy showed that LLLI treatments were found to (1) increase chondrocyte viability, (2) promote secretion of matrix proteins, (3) upregulate expression of chondrogenic genes, and (4) downregulate gene expression of cell destructive proteases and genes coding for mediators involved in the extrinsic apoptosis signaling pathway. Furthermore, LLLI attenuated induction of genes associated with cell death and matrix breakdown induced by IL-1β, some of which was seen at the protein level, with verification of effects on gene expression in the C28/I2 human chondrocyte line. LLLI treatments during culture generated larger numbers of viable chondrocytes compared to untreated cultures. Moreover, LLLI-treated chondrocytes in culture also rectified and simultaneously maintained their differentiated phenotype. Cultured chondrocytes treated with LLLI are a promising cell source for repairing cartilage lesions in vivo and restoration of articular function using tissue engineering strategies.

Highlights

The ability of articular cartilage for remodeling and damage repair is limited by its avascular nature, and by the low cellularity (Oldershaw, 2012) and functionality (Archer and Francis-West, 2003) of chondrocytes in surface layers
The viability of cultured chondrocytes treated by level laser irradiation (LLLI) for 5 min (3.58 J/cm2) in the 1 day treatment group became significantly (P < 0.05) higher than that of untreated cells, peaked (P < 0.01) at 8 min (5.74 J/cm2), decreased to a significant level as treated for 11 min (7.87 J/cm2) before becoming insignificant when treated for 13 min (9.30 J/cm2) (Figure 1A)
Cultured chondrocytes responded to LLLI treatment for 3 days with a significant increment (P < 0.05) in cellular viability by 8 min (≈17 J/cm2)

Summary

Introduction

The ability of articular cartilage for remodeling and damage repair is limited by its avascular nature, and by the low cellularity (Oldershaw, 2012) and functionality (Archer and Francis-West, 2003) of chondrocytes in surface layers. A plethora of evidence showed that passaged chondrocytes alter their gene expression profiles (Lin et al, 2008) and become more fibroblastic (Stokes et al, 2001). This process of dedifferentiation typically shows decreased collagen type II (COL II) and aggrecan (ACAN) accompanied by increased collagen type I (COL I) (Hsu et al, 2002; Darling and Athanasiou, 2005; Frohlich et al, 2007). Dedifferentiated chondrocytes have failed to achieve long term repair and restoration of functional articular cartilage due to the formation of fibrocartilage as shown in ACI and MACI (Roberts et al, 2009), and microfracture (Gobbi et al, 2005). Effective numbers of expanded chondrocytes with enhanced differentiated phenotype could be achieved by modulation with various factors, including the approach of accessible laser irradiation

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Conclusion