Donor age effects on in vitro chondrogenic and osteogenic differentiation performance of equine bone marrow- and adipose tissue-derived mesenchymal stromal cells

Abstract

Background Bone marrow (BM)- and adipose tissue (AT)-derived mesenchymal stromal cells (MSCs) have shown potential as cell-based therapies for cartilage and bone injuries and are used increasingly in human and veterinary practice to facilitate the treatment of orthopedic conditions. However, human and rodent studies have documented a sharp decline in chondrogenic and osteogenic differentiation potential with increasing donor age, which may be problematic for the important demographic of older orthopedic patients. The aim of this study was to identify the effect of donor age on the chondrogenic and osteogenic differentiation performance of equine BM- and AT-MSCs in vitro. BM- and AT-MSCs and dermal fibroblasts (biological negative control) were harvested from horses in five different age groups (n = 4, N = 60); newborn (0 days), yearling (15–17 months), adult (5–8 years), middle-aged (12–18 years), and geriatric (≥ 22 years). Chondrogenic differentiation performance was assessed quantitatively by measuring pellet size, matrix proteoglycan levels, and gene expression of articular cartilage biomarkers. Osteogenic differentiation performance was assessed quantitatively by measuring alkaline phosphatase activity, calcium deposition, and gene expression of bone biomarkers. Results Chondrogenic and osteogenic differentiation performance of equine BM- and AT-MSCs declined with increasing donor age. BM-MSCs had a higher chondrogenic differentiation performance. AT-MSCs showed minimal chondrogenic differentiation performance in all age groups. For osteogenesis, alkaline phosphatase activity was also higher in BM-MSCs, but BM-MSCs calcium deposition was affected by donor age earlier than AT-MSCs. Chondrogenic and osteogenic differentiation performance of BM-MSCs exhibited a decline as early as between the newborn and yearling samples. Steady state levels of mRNA encoding growth factors, chondrogenic, and osteogenic biomarkers were lower with increasing donor age in both MSC types. Conclusions The data showed that chondrogenic and osteogenic differentiation performance of equine BM-MSCs declined already in yearlings, and that AT-MSCs showed minimal chondrogenic potential, but were affected later by donor age with regards to osteogenesis (calcium deposition). The results highlight the importance of donor age considerations and MSC selection for cell-based treatment of orthopedic injuries and will help inform clinicians on when to implement or potentially cryopreserve cells. Moreover, the study provides molecular targets affected by donor age.

Successful isolation of viable stem cells from cryopreserved microfragmented human adipose tissue from patients with knee osteoarthritis – a comparative study of isolation by tissue explant culture and enzymatic digestion

“…Aging is positively correlated with the number of senescent articular chondrocytes and osteoarthritis progression [33]. It is therefore important to consider cellular senescence as a contributing factor in the pathophysiology of osteoarthritis and in therapeutic stem cell potential [13, 34]. SA‐BGAL activity is considered a gold‐standard biomarker of cellular senescence associated with increased size and activity of the lysosomal compartment, which releases lipofuscin [26, 35, 36].…”

“…Cryopreservation and biobanking of stem cells are important for research purposes and for treatment of aged patients, as aging has demonstrated negative effects on the potential of AT-derived stem cells when treating osteoarthritis [13][14][15][16]. Cryopreservation potentially opens the possibility to store functional stem cells from an early age for use later in life.…”

Immunosenescence and inflammaging in the aged horse

“…Studies of equine stem cells derived from bone marrow, adipose, and synovial fluid have demonstrated progressive reductions in cell proliferation and differentiation potential, as well as increased prevalence of age-related binucleate or tetraploid cells. Additionally, stem cells derived from geriatric horses are more likely to show morphological features correlated with aging such as endoplasmic reticulum stress, autophagy, and mitophagy [55][56][57][58].…”

Use of Brain-Derived Stem/Progenitor Cells and Derived Extracellular Vesicles to Repair Damaged Neural Tissues: Lessons Learned from Connective Tissue Repair Regarding Variables Limiting Progress and Approaches to Overcome Limitations

“…There are at least two interesting points associated with this perspective; (1) there could be specific loss of PNRC numbers or functioning with aging [ 106 ] and life-span transitions, which would put the health and functioning of the other cells in a brain centre at risk; (2) it would be better to attempt to restore function earlier rather than later in a disease process when the number of residual cells is still prominent. Regarding Point 1, it is known that the numbers and function of PMRC/MSC decline with age [ 107 , 108 , 109 ] and the brain-associated NSC as defined by Reynolds et al [ 1 , 2 ] act as a reservoir of pluripotent cells for replenishment of small numbers of NSC that are required in a subclinical manner for, perhaps, maintenance of tissue integrity. Regarding Point 2, the medical model of relying on drugs during the early stages of disease and only entertaining serious, potentially more risky interventions until late in the disease process actually works against the concept of restoration of function when there are sufficient residual cell numbers in the affected tissues to make the restoration of function with NSC more achievable.…”