Background Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) are emerging as a promising cell-free strategy for intervertebral disc degeneration (IDD) treatment. This study aimed to evaluate the anabolic effect of Wharton's Jelly MSC (WJ-MSC)-derived EVs on degenerative human nucleus pulposus cells (hNPCs) under in vitro inflammation using a 3D culture model.Methods Following isolation, hNPCs (n = 10) were encapsulated in alginate beads and treated with 10, 50, and 100 mu g/mL of WJ-MSC-EVs after preincubation with 10 ng/mL interleukin (IL)-1 beta. Cell proliferation, viability, nitrite production, and glycosaminoglycan (GAG) content were assessed. Histological analyses evaluated extracellular matrix (ECM) production. Phenotypic (SOX9, KRT19), catabolic (MMP1, MMP13, ADAMTS5, IL6, NOS2), and anabolic (ACAN) ECM markers were analyzed by RT-qPCR.Results WJ-MSC-EVs significantly promoted hNPC proliferation at all concentrations, with 10 mu g/mL effectively counteracting IL-1 beta catabolic effects. Live/dead staining showed reduced cell death in EV-treated hNPCs compared to the IL-1 beta-only group. Nitrite production decreased after 7 days with 10 mu g/mL WJ-EVs, supported by reduced NOS2 expression. GAG content increased dose-dependently, as confirmed by Alcian blue staining. WJ-EVs positively modulated anabolic (ACAN, KRT19, SOX9), catabolic (ADAMTS5, MMP1, MMP13), and inflammatory (IL6) gene expression levels.Conclusion WJ-MSC-derived EVs demonstrate potential as a cell-free therapeutic approach for IDD by enhancing hNPC growth, mitigating ECM degradation, and reducing oxidative stress-related IDD progression. These findings warrant further investigation into the use of WJ-EVs for IDD treatment.

Wharton's Jelly Mesenchymal Stromal Cell-Derived Extracellular Vesicles Attenuate Intervertebral Disc Degeneration Under Inflammatory Stress in an In Vitro 3D Culture System

Tilotta, Veronica;Vadalà, Gianluca;Russo, Fabrizio;Papalia, Rocco;Denaro, Vincenzo
2025-01-01

Abstract

Background Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) are emerging as a promising cell-free strategy for intervertebral disc degeneration (IDD) treatment. This study aimed to evaluate the anabolic effect of Wharton's Jelly MSC (WJ-MSC)-derived EVs on degenerative human nucleus pulposus cells (hNPCs) under in vitro inflammation using a 3D culture model.Methods Following isolation, hNPCs (n = 10) were encapsulated in alginate beads and treated with 10, 50, and 100 mu g/mL of WJ-MSC-EVs after preincubation with 10 ng/mL interleukin (IL)-1 beta. Cell proliferation, viability, nitrite production, and glycosaminoglycan (GAG) content were assessed. Histological analyses evaluated extracellular matrix (ECM) production. Phenotypic (SOX9, KRT19), catabolic (MMP1, MMP13, ADAMTS5, IL6, NOS2), and anabolic (ACAN) ECM markers were analyzed by RT-qPCR.Results WJ-MSC-EVs significantly promoted hNPC proliferation at all concentrations, with 10 mu g/mL effectively counteracting IL-1 beta catabolic effects. Live/dead staining showed reduced cell death in EV-treated hNPCs compared to the IL-1 beta-only group. Nitrite production decreased after 7 days with 10 mu g/mL WJ-EVs, supported by reduced NOS2 expression. GAG content increased dose-dependently, as confirmed by Alcian blue staining. WJ-EVs positively modulated anabolic (ACAN, KRT19, SOX9), catabolic (ADAMTS5, MMP1, MMP13), and inflammatory (IL6) gene expression levels.Conclusion WJ-MSC-derived EVs demonstrate potential as a cell-free therapeutic approach for IDD by enhancing hNPC growth, mitigating ECM degradation, and reducing oxidative stress-related IDD progression. These findings warrant further investigation into the use of WJ-EVs for IDD treatment.
2025
exosome; extracellular vesicles; intervertebral disc; intervertebral disc degeneration; intervertebral disc regeneration; low back pain; mesenchymal stromal cells
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Descrizione: Wharton's Jelly Mesenchymal Stromal Cell-Derived Extracellular Vesicles Attenuate Intervertebral Disc Degeneration Under Inflammatory Stress in an In Vitro 3D Culture System
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/20.500.12610/89605
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