Background/Purpose: Previous work by our group and others indicated that an accumulation of lamin A (LMNA) is associated with the osteoarthritis (OA) chondrocyte phenotype. Mutations of this protein are linked to laminopathies and specifically to Hutchinson-Guilford Progeria Syndrome (HGPS), an accelerated aging disease. Some authors have proposed that a deregulation of LMNA affects the differentiation potential of stem cells. In the present study, we examined the effect of the over-expression of LMNA, on the mesoderm and, specifically, chondrocyte differentiation potential of Mesenchymal Stem Cells (MSCs).

Methods: MSCs from human umbilical cord (UC) stroma have previously been isolated, expanded and differentiated towards mesoderm cell lineages. For efficient gene delivery of wt LMNA and GFP (Green Fluorescence Protein) as control, we used a lentiviral expression system. Osteogenic potential was studied by with alizarin red staining and Real-Time PCR of ALP, OC and Runx2 to assess early and late osteogenic differentiation. Adipogenic potential was studied with Oil Red staining and Real-Time PCR of LPL, FABP and ADIPOQ, for early and late adipogenic differentiation. Chondrogenesis and hypertrophy were studied using immunohistochemestry and Real-Time PCR of Aggrecan, MMP-13, Type II Collagen, Type I Collagen and Type I Collagen. Reactive oxygen species (ROS) generation was measured by flow cytometry and oriented migration capacity was tested in a Modified Boyden´s chamber assay.

Results: We found that over-expression of LMNA by lentiviral gene delivery leads to alterations in differentiation potential. Adipogenic capacity is increased in LMNA-MSCs, accompanied by an accumulation of Fatty Acid Synthase, as revealed by Western-blotting. The chondrogenic potential is defective in LMNA-MSCs, showing a decreased COL2/COL1 ratio and an increase in hypertrophy markers. These cells present lower levels of manganese superoxide dismutase (MnSODM) and an increase of mitochondrial MnSODM-dependent reactive oxygen species (ROS). This stable extra ROS generation is accompanied by alterations in the oriented migratory capacity of these cells under the effect of pro-inflammatory cytokines. ROS synthesis was partially and totally reverted by incubation with the ROS scavenger N-Acetyl Cystein (NAC) for 1 hour in culture. In addition, defects in chondrogenesis detected by immunohistochemestry and Real Time-PCR are partially reversed by periodic incubations with NAC for 1 hour.

Conclusion: Overall, our results show that chondrogenic differentiation of UC-MSCs is compromised by Lamin A deregulation whereas adipogenic differenciation is enhanced. We also demonstrated that this deregulation alters the oxidative stress balance in MSCs, modifies their migratory properties and induces defects in their capacity to differentiate into chondrocytes in our in vitro model. Further experiments are necessary to determine the in vivo significance of those alterations and to explore putative key targets for stem cell therapies in OA and other aging syndromes.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/overexpression-of-lamin-a-in-mesenchymal-stem-cells-compromises-chondrogenesis-and-enhances-adipogenesis/