Mesenchymal Stromal Cells Donate Mitochondria to Articular Chondrocytes Exposed to Mitochondrial, Environmental, and Mechanical Stress

Abstract

Avascular soft tissues of the skeletal system, including articular cartilage, have limited healing capacity, in part due to their low metabolic activity. No drugs are available that can prevent or slow the development of osteoarthritis (OA) after joint injury. Therefore, mesenchymal stromal cell (MSC)-based regenerative therapies are increasingly common in the treatment of OA, but questions regarding their clinical efficacy and mechanisms of action remain unanswered. Our group recently reported that mitochondrial dysfunction is one of the earliest responses of cartilage to injury, resulting in chondrocyte death, extracellular matrix degeneration, and ultimately OA. MSCs have been found to rescue injured cells and improve healing by donating healthy mitochondria in highly metabolic tissues, but mitochondrial transfer has not been investigated in cartilage. Here, we demonstrate that MSCs transfer mitochondria to stressed chondrocytes in cell culture and in injured cartilage tissue. Conditions known induce chondrocyte mitochondrial dysfunction, including stimulation with rotenone/antimycin and hyperoxia, increased transfer. Stressed chondrocytes increased expression of genes related to inflammation and senescence, further supporting the link between mitochondrial dysfunction and transfer. MSC-chondrocyte mitochondrial transfer was blocked by non-specific and specific (connexin-43) gap-junction inhibition. When MSCs were exposed to mechanically injured cartilage they localized to areas of matrix damage and extended cellular processes deep into microcracks, delivering mitochondria to chondrocytes. This work provides insights into the chemical, environmental, and mechanical conditions that can elicit MSC-chondrocyte mitochondrial transfer in vitro and in situ, and our findings suggest a new potential role for MSC-based therapeutics after cartilage injury.Significance StatementRecent evidence suggests that although articular cartilage is avascular and relatively metabolically quiescent, acute injury induces chondrocyte mitochondrial dysfunction, driving cartilage degradation and OA. We present the first evidence that MSCs donate mitochondria to articular chondrocytes undergoing mitochondrial dysfunction in vitro and in situ. These findings support a new role for MSCs in the context of cartilage injury and OA, and intercellular mitochondrial transfer may represent a new biological approach to augment mitochondrial capacity in injured chondrocytes. This work establishes multiple experimental models to study MSC mitochondrial donation for the treatment of OA and related degenerative diseases of avascular orthopedic tissues.

Human mesenchymal stromal cells release functional mitochondria in extracellular vesicles

“…However, it should be noted that several factors, including culture time and MV numbers were not optimized for these experiments. In similar work, where chondrocytes and MSCs were directly co-cultured, the frequency of MT transfer was higher than reported here for mitoEV-mediated transfer; between 1-15% depending on experimental parameters, including culture time, cell ratios, and environmental conditions (Bennett et al, 2019;Fahey et al, 2022). Therefore, we anticipate that future work to optimize experimental parameters and identify mechanisms mediating MSC-chondrocyte mitoEV transfer will allow us to manipulate the rate of mitoEV release by MSCs and uptake by chondrocytes.…”

“…However, MSCderived mitoEV transfer has not been demonstrated to chondrocytes. Intriguingly, our group has reported MSCchondrocyte MT transfer in direct coculture and observed MSCs apparently shedding MT into the extracellular space during co-culture with injured chondrocytes (Bennett et al, 2019;Fahey et al, 2022). However, it was unclear if these MT were packaged within EVs (mitoEVs).…”