Mitochondrial Quality Control in Acute Lymphoblastic Leukemia Bone Marrow Stem Cells

Abstract:

The bone marrow niche plays a crucial role in maintaining healthy hematopoiesis. Mesenchymal stem cells (MSCs), a fundamental component of the bone marrow niche, support healthy hematopoiesis and undergo cellular and molecular changes during leukemogenesis, providing metabolic and antioxidant support to leukemic cells and contributing to drug resistance.

Alterations in energy production pathways impact the destiny of the cell, its capacity to differentiate, and its stemness. At this stage, the importance of the quality and functioning of mitochondria, regulated by fission, fusion, and mitophagy, becomes critical. Our study aimed to evaluate changes in mitochondrial quality in MSCs of pediatric ALL patients and investigate the relationship between mitochondrial quality in the niche and clinical outcomes.

Our study investigated mitochondrial quality in MSCs from children newly diagnosed with ALL or relapsed ALL, compared to healthy bone marrow donors. RT-PCR was performed to analyze the expression of genes related to mitochondrial fission, fusion, and mitophagy in MSCs. Q-PCR was used to assess the number of mitochondrial DNA copies. Confocal microscopy was employed to examine mitochondrial morphology, and mitochondrial volume was measured with flow cytometry.

The morphology of MSCs from relapse patients is similar to that of healthy and newly diagnosed ALL cells; however, the time required to reach 70-80% confluence was longer for relapse samples. Studies have shown that MSCs obtained from ALL patients have lower proliferation and support capacity than healthy MSCs; this finding may suggest relapse MSCs have a reduced self-renewal potential of the ALL microniche.

Flow cytometry analysis revealed that mitochondrial volume was significantly higher in patients than healthy individuals (p: 0.001). Patients exhibited higher mitochondrial Drp1, Fis1, Parkin, Opa, and mtDNA expression levels than healthy individuals. Conversely, Mfn1 and Mfn2 were more highly expressed in healthy donors than in patients. Drp1, Fis1, Mfn1, Opa, and mitochondrial volume were higher in relapsed patients than in newly diagnosed ALL patients. We suggest that higher fission in leukemic MSCs compared to the healthy population potentially supports cancer cells.

In newly diagnosed and relapsed ALL patients, mitochondrial volume was high, but the mtDNA amount was low. Patient MSCs contained less mtDNA and had a larger mitochondrial volume than healthy MSCs. When comparing relapse and newly diagnosed ALL, relapse ALLs had higher mitochondrial volume and lower mtDNA amount than newly diagnosed ALL.

Patients with high Drp-1 and Opa expression had an increased frequency of CNS involvement (p: 0.009 – 0.044). Additionally, increased mitochondrial volume was associated with CNS involvement (p: 0.020). We suggest that increased fission gene expression in MSCs could indicate damage and facilitate mitochondrial transfer to cancer cells, potentially making leukemic cells more invasive.

When we evaluated the bone marrow aspiration samples on days 15 and 33, we found that on day 15, Drp1 and Fis1, and on day 33, only Fis1 was associated with higher MRD percentages in patients with high gene expression. The elevated MRD rates found in leukemic cells on days 15 and 33 could potentially be linked to resistance to chemotherapy. Furthermore, it is possible that leukemic MSCs initially possess a high capacity for oxidative phosphorylation.

We found significant differences in mitochondrial quality parameters between healthy donors and patients. The altered mitochondrial dynamics, particularly the increased mitochondrial fission, are thought to be associated with excessive stress in the bone marrow niche and mitochondrial transfer. This study is the first to examine the relationship between mitochondrial quality parameters of pediatric ALL patients MSC’s. As the effects of the bone marrow niche and MSCs on cancer become better understood, they will provide guidance for developing new treatment methods.