Human Mesenchymal Stem Cell Hydrogen Sulfide Production Critically Impacts the Release of Other Paracrine Mediators After Injury.

BACKGROUND

The use of mesenchymal stem cells (MSCs) for treatment during ischemia is novel. Hydrogen sulfide (HS) is an important paracrine mediator that is released from MSCs to facilitate angiogenesis and vasodilation. Three enzymes, cystathionine-beta-synthase (CBS), cystathionine-gamma-lyase (CSE), and 3-mercaptopyruvate-sulfurtransferase (MPST), are mainly responsible for HS production. However, it is unclear how these enzymes impact the production of other critical growth factors and chemokines. We hypothesized that the enzymes responsible for HS production in human MSCs would also critically regulate other growth factors and chemokines.

MATERIALS AND METHODS

Human MSCs were transfected with CBS, MPST, CSE, or negative control small interfering RNA. Knockdown of enzymes was confirmed by polymerase chain reaction. Cells were plated in 12-well plates at 100,000 cells per well and stimulated with tumor necrosis factor-α (TNF-α; 50 ng/mL), lipopolysaccharide (LPS; 200 ng/mL), or 5% hypoxia for 24 h. Supernatants were collected, and cytokines measured by multiplex beaded assay. Data were compared with the Mann-Whitney U-test, and P < 0.05 was significant.

RESULTS

TNF-α, LPS, and hypoxia effectively stimulated MSCs. Granulocyte colony-stimulating factor (GCSF), epidermal growth factor, fibroblast growth factor, granulocyte/monocyte colony-stimulating factor (GMCSF), vascular endothelial growth factor, and interferon gamma-inducible protein 10 were all significantly elevated when CSE was knocked down during TNF-α stimulation (P < 0.05). Knockdown of MPST during LPS stimulation more readily increased GCSF and epidermal growth factor but decreased GMCSF (P < 0.05). CBS knockdown decreased production of GCSF, fibroblast growth factor, GMCSF, and vascular endothelial growth factor (P < 0.05) after hypoxia.

CONCLUSIONS

The enzymes that produce HS in MSCs are also responsible for the production of other stem cell paracrine mediators under stressful stimuli. Therefore, reprogramming MSCs to endogenously produce more HS as a therapeutic intervention could also critically impact other paracrine mediators, which may alter the desired beneficial effects.