2-Mercaptoethanol enhances the yield of exosomes showing therapeutic potency in alleviating spinal cord injury mice.

BACKGROUND

Limited passage numbers of mesenchymal stem cells (MSCs) present challenges in producing sufficient exosomes for spinal cord injury (SCI) treatment.

OBJECTIVES

This study investigates whether β-mercaptoethanol (BME) preconditioning of MSCs can increase exosome yield for SCI therapy.

METHODS

Exosomal content was analyzed using silver staining and SYBR Gold staining. Cell viability was assessed via CCK-8 and EdU assays. IL-1β, IL-6, TNF-α, and MCP-1 levels were measured by enzyme-linked immunosorbent assay (ELISA). Neuronal differentiation influenced by astrocytes was evaluated through neurite outgrowth and migration assays. Neuronal survival and motor function recovery in SCI mice were assessed using TUNEL staining, the Basso Mouse Scale (BMS), muscle strength tests, and motor evoked potential (MEP) measurements.

RESULTS

BME treatment significantly increased exosome quantity, including proteins and microRNAs, without drastic changes in exosomal content spectrum. Exosomes from BME-treated MSCs more effectively suppressed IL-1β, IL-6, TNF-α, and MCP-1 secretion by astrocytes, reducing neuronal inflammation. Yap1 activation reduced the exosomes' inhibitory effects on inflammatory cytokines. Mice treated with exosomes from BME-treated MSCs showed better outcomes: lower GFAP and C3 expression, reduced inflammation, increased NF-H levels, higher BMS scores, and greater MEP peaks. Exosome treatment also reduced bladder volume, residual urine, and the time to regain spontaneous urination after uroschesis.

CONCLUSION

BME preconditioning enhances exosome yield from hUC-MSCs, offering improved therapeutic potential for SCI.