Development of a Robust Consensus Modeling Approach for Identifying Cellular and Media Metabolites Predictive of Mesenchymal Stromal Cell Potency

Stem Cells, Год журнала: 2023, Номер 41(8), С. 792 - 808

Опубликована: Июнь 3, 2023

Abstract Mesenchymal stromal cells (MSCs) have shown promise in regenerative medicine applications due part to their ability modulate immune cells. However, MSCs demonstrate significant functional heterogeneity terms of immunomodulatory function because differences MSC donor/tissue source, as well non-standardized manufacturing approaches. As metabolism plays a critical role expand therapeutic numbers ex vivo, we comprehensively profiled intracellular and extracellular metabolites throughout the expansion process identify predictors (T-cell modulation indoleamine-2,3-dehydrogenase (IDO) activity). Here, media non-destructive manner through daily sampling nuclear magnetic resonance (NMR), at end using mass spectrometry (MS). Using robust consensus machine learning approach, were able panels predictive for 10 independent lines. This approach consisted identifying 2 or more models then building based on these metabolite panels. Consensus with high value included multiple lipid classes (such phosphatidylcholines, phosphatidylethanolamines, sphingomyelins) while proline, phenylalanine, pyruvate. Pathway enrichment identified metabolic pathways significantly associated such sphingolipid signaling metabolism, arginine proline autophagy. Overall, this work establishes generalizable framework that predict function, guiding future efforts identification high-potency lines engineering.

Язык: Английский

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Intra-individual variability in the neuroprotective and promyelinating properties of conditioned culture medium obtained from human adipose mesenchymal stromal cells

Stem Cell Research & Therapy, Год журнала: 2023, Номер 14(1)

Опубликована: Май 11, 2023

Abstract Background Greater knowledge of mesenchymal stromal cell (MSC)-based therapies is driving the research into their secretome, identified as main element responsible for therapeutic effects. The aim this study to characterize individual variability secretome adipose tissue-derived MSCs (adMSCs) with regard potential therapeutical applications in neurology. Methods adMSCs were isolated from intact tissue ten subjects undergoing abdominal plastic surgery or reduction mammoplasty. Two commercial lines also included. We analyzed expansion rate, production, and secretion growth factors interest neurological (VEGF-A, BDNF, PDGF-AA AA/BB, HGF, NGF, FGF-21, GDNF, IGF-I, IGF-II, EGF FGF-2). To correlate these characteristics biological effects on cellular targets, we used media conditioned various donors primary cultures neurons/astrocytes oligodendrocyte precursor cells (OPCs) exposed noxious stimuli (oxygen–glucose deprivation, OGD) evaluate protective promyelinating properties, using MSC medium a control group. Results showed significant within considered population PDGF-AA, PDGF-AB/BB, VEGF-A BDNF. None MSC-derived supernatants affected neuron viability normoxia, while substantial protection by high BDNF-containing was observed neuronal OGD conditions. In OPC cultures, protected OGD-induced death, increasing differentiation mature oligodendrocytes. Neuroprotection positive correlation VEGF-A, BDNF concentrations culture supernatants, an inverse following positively correlated concentration. Conclusions Despite limited number adMSC donors, properties under-researched aspect which may represent important step translation acellular products clinical practice. capability oligodendroglial lineages oxygen–glucose deprivation. These are directly concentration specific factors, indicate that remyelination should be included target MSC-based therapies.

Язык: Английский

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Chondrocyte Isolation from Loose Bodies—An Option for Reducing Donor Site Morbidity for Autologous Chondrocyte Implantation

International Journal of Molecular Sciences, Год журнала: 2023, Номер 24(2), С. 1484 - 1484

Опубликована: Янв. 12, 2023

Loose bodies (LBs) from patients with osteochondritis dissecans (OCD) are usually removed and discarded during surgical treatment of the defect. In this study, we address question whether these LBs contain sufficient viable functional chondrocytes that could serve as a source for autologous chondrocyte implantation (ACI) how required prolonged in vitro expansion affects their phenotype. Chondrocytes were isolated 18 compared control non-weight-bearing joint regions (n = 7) bone marrow mesenchymal stromal cells (BMSCs, n 6) obtained primary arthroplasty. No significant differences initial cell yield per isolation expression progenitor markers CD44 + /CD146+ found between populations (LB-CH) (Ctrl-CH). During long-term expansion, LB-CH exhibited comparable viability proliferation rates to no ultimate cycle arrest was observed within 12 passages respectively 15.3 ± 1.1 mean cumulative doublings (CPD). The chondrogenic differentiation potential Ctrl-CH, but both groups showed significantly higher ability form hyaline cartilage matrix than BMSC. Our data suggest promising obtaining qualitatively quantitatively suitable therapeutic applications, thereby circumventing donor site morbidity consequence biopsies current ACI procedure.

Язык: Английский

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Decreased CRISPLD2 expression impairs osteogenic differentiation of human mesenchymal stem cells during in vitro expansion

Journal of Cellular Physiology, Год журнала: 2023, Номер 238(6), С. 1368 - 1380

Опубликована: Апрель 6, 2023

Abstract Human mesenchymal stem cells (hMSCs) are the cornerstone of regenerative medicine; large quantities hMSCs required via in vitro expansion to meet therapeutic purposes. However, quickly lose their osteogenic differentiation potential during expansion, which is a major roadblock clinical applications. In this study, we found that human bone marrow (hBMSCs), dental pulp (hDPSCs), and adipose (hASCs) was severely impaired after expansion. To clarify molecular mechanism underlying expansion‐related loss capacity hMSCs, transcriptome changes following these were compared. Cysteine‐rich secretory protein LCCL domain‐containing 2 (CRISPLD2) identified as most downregulated gene shared by late passage hBMSCs, hDPSCs, hASCs. Both secreted non‐secreted CRISPLD2 proteins progressively declined when gradually lost potential. We thus hypothesized expression critical for maintain Our studies showed knockdown early hBMSCs inhibited cells' siRNA dose‐dependent manner. Transcriptome analysis immunoblotting indicated knockdown‐induced osteogenesis suppression might be attributed downregulation matrix metallopeptidase 1 (MMP1) forkhead box Q1 (FOXQ1). Furthermore, adeno‐associated virus (AAV)‐mediated overexpression could somewhat rescue These results revealed contributes findings shed light on understanding provide target bone‐related diseases.

Язык: Английский

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Development of serum substitute medium for bone tissue engineering

Journal of Biomedical Materials Research Part A, Год журнала: 2023, Номер 111(9), С. 1423 - 1440

Опубликована: Апрель 6, 2023

Abstract In tissue engineering, cells are grown often on scaffolds and subjected to chemical/mechanical stimuli. Most such cultures still use fetal bovine serum (FBS) despite its known disadvantages including ethical concerns, safety issues, variability in composition, which greatly influences the experimental outcomes. To overcome of using FBS, chemically defined substitute medium needs be developed. Development depends cell type application—which makes it impossible define one universal for all any application. Here, we developed a bone engineering (BTE) step‐by‐step process. Essential components were added while human marrow mesenchymal stromal (hBMSCs, osteoblast progenitor cells) cultured two‐dimensional three‐dimensional substrates. 3‐week culture, worked equally well as FBS containing term attachment substrate, survival, differentiation, deposition extracellular matrix. next step, was evaluated when culturing under mechanical loading form shear stress. The outcomes showed that application stress is essential improve matrix formation medium. could pave way replacing BTE studies eliminating controversial providing better‐defined chemical environment studies.

Язык: Английский

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Development of serum substitute medium for bone tissue engineering

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2022, Номер unknown

Опубликована: Окт. 7, 2022

Abstract In tissue engineering, cells are grown often on scaffolds and subjected to chemical/mechanical stimuli. Most such cultures still use fetal bovine serum (FBS) despite its known disadvantages including ethical concerns, safety issues, variability in composition, which greatly influences the experimental outcomes. To overcome of using FBS, chemically defined substitute medium needs be developed. Development depends cell type application - makes it impossible define one universal for all any application. Here, we developed a bone engineering (BTE) step-by-step process. Essential components were added while human marrow mesenchymal stromal (hBMSCs, osteoblast progenitor cells) cultured two-dimensional (2D) three-dimensional (3D) substrates. 3-week culture, worked equally well as FBS containing term attachment substrate, survival, differentiation, deposition extracellular matrix. next step, was evaluated when culturing under mechanical loading form shear stress. The outcomes showed that stress is essential improve matrix formation medium. could pave way replacing BTE studies eliminating controversial providing better-defined chemical environment studies. Graphical abstract

Язык: Английский

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Development of a Robust Consensus Modeling Approach for Identifying Cellular and Media Metabolites Predictive of Mesenchymal Stromal Cell Potency

bioRxiv (Cold Spring Harbor Laboratory), Год журнала: 2023, Номер unknown

Опубликована: Фев. 4, 2023

Abstract Mesenchymal stromal cells (MSCs) have shown promise in regenerative medicine applications due part to their ability modulate immune cells. However, MSCs demonstrate significant functional heterogeneity terms of immunomodulatory function because differences MSC donor/tissue source, as well non-standardized manufacturing approaches. As metabolism plays a critical role expand therapeutic numbers ex vivo , we comprehensively profiled intracellular and extracellular metabolites throughout the expansion process identify predictors (T cell modulation indoleamine-2,3-dehydrogenase (IDO) activity). Here, media non-destructive manner through daily sampling nuclear magnetic resonance (NMR), at end using mass spectrometry (MS). Using robust consensus machine learning approach, were able panels predictive for 10 independent lines. This approach consisted identifying 2 or more models then building based on these metabolite panels. Consensus with high value included multiple lipid classes (such phosphatidylcholines, phosphatidylethanolamines, sphingomyelins) while proline, phenylalanine, pyruvate. Pathway enrichment identified metabolic pathways significantly associated such sphingolipid signaling metabolism, arginine proline autophagy. Overall, this work establishes generalizable framework that predict function, guiding future efforts identification potency lines engineering.

Язык: Английский

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