PhD Student The Chinese University of Hong Kong (CUHK), Hong Kong
Abstract: The meniscus is a crucial cartilage tissue bearing the majority of the mechanical load in the knee joint. Adult human meniscus stem/progenitor cells (hMeSPCs) are the tissue resident progenitor cells of meniscus tissue, and are shown to be sensitive to mechanical stimulations. Our previous study has revealed how the biomimetic mild cyclic tensile loading (0.5 Hz, 10% strain, 1h/day) activates hMeSPCs in 3D hydrogel cultures in the perspective of stem cell activation, ECM accumulation and remodelling. However, the influence of fast and intense tensile loading on hMeSPCs is yet to be revealed. In previous study, 20% of tensile stretch was applied to mimic intense mechanical stress on chondrocytes which shown an overload effect of the cells. By introducing servomotor and conveyor pulleys to in-house built bioreactor, we are able to implement considerable frequent stretch up to 4 Hz with a maximum 34% of hydrogel extension in length. We then seeded the hMeSPCs (n=8 biological donors) into GelMA hydrogel and introduce a fixed frequency of 1 Hz and 20% of stretching to the 3D cultured hMeSPCs, which is a biomimetic situation of joint meniscus load in fast walk scenario (120 step/min). The hMeSPCs were loaded (1 Hz, 1h/day, 10% strain and 20 strain, respectively) for 14 days. The cell-hydrogel cultures were then collected and analysed by histology of Safarinin O staining. We quantified the positive staining area of Safarinin O staining in hydrogel of each group (Ctrl: no loading; 10%: 1 Hz, 10% Strain; 20%: 1 Hz, 20% strain; n≥6 cell-hydrogel cultures/group), and observed increased cartilage-like ECM deposition in the hydrogel after frequent stretching in both 10% (15.52±9.41%, P < 0.05) and 20% stretch (13.89±7.99%, P < 0.01) compared with no stretching control (6.96±1.81%). Meanwhile, the porosity is not significantly changed by different loading parameters (Negative control: 40.33±8.18%, 10%: 43.62±11.17%, 20%: 44.24±9.04%). Next, we will apply flow cytometry and transcriptome analysis to further investigate the influence of intense stretch on hMeSPCs in the aspect of cell viability and underlying mechanisms. In summary, the in vitro findings provided better understanding of the mechanobiology of hMeSPCs, consolidating the knowledge for further cell-based therapy in meniscus-related diseases.
Funding Source: National Key R&D Program [2019YFA0111900, to YJ], MOST, China; General Research Fund (GRF, 14104022, to YJ) RGC, UGC of Hong Kong; the Center for Neuromusculoskeletal Restorative Medicine [CNRM at InnoHK, to RST, YJ]
