Restoration of Hydrogen Sulfide Production in Diabetic Mice Improves Reparative Function of Bone Marrow Cells

Abstract

Background Bone marrow cell-based treatment for critical limb ischemia (CLI) in diabetic patients yielded a modest therapeutic effect due to cell dysfunction. Therefore, approaches that improve diabetic stem/progenitor cell functions may provide therapeutic benefits. Here, we tested the hypotheses that restoration of hydrogen sulfide (H2S) production in diabetic bone marrow cells (BMCs) improves their reparative capacities. Methods Mouse BMCs were isolated by density-gradient centrifugation. Unilateral hind limb ischemia (HLI) was conducted in 12- to 14-week old db/+ and db/db mice by ligation of left femoral artery. H2S level was measured by either gas chromatography or staining with florescent dye sulfidefluor 7AM. Results Both H2S production and cystathionine γ-lyase (CSE), an H2S enzyme, levels were significantly decreased in BMCs from diabetic db/db mice. Administration of H2S donor diallyl trisulfide (DATS) or overexpression of CSE restored H2S production and enhanced cell survival and migratory capacity in high glucose (HG)-treated BMCs. Immediately after HLI surgery, the db/+ and db/db mice were administrated with DATS orally and/or local intramuscular injection of GFP-labeled BMCs or RFP-CSE-overexpressing BMCs (CSE-BMCs). Mice with HLI were divided into six groups: 1) db/+; 2) db/db; 3) db/db+BMCs; 4) db/db+DATS; 5) db/db+DATS+BMCs; 6) db/db+CSE-BMCs. DATS and CSE overexpression greatly enhanced diabetic BMCs retention in ischemic hind limbs (IHL) followed by improved blood perfusion, capillary/arteriole density, skeletal muscle architecture and cell survival, and decreased perivascular CD68+ cell infiltration in IHL of diabetic mice. Interestingly, DATS or CSE overexpression rescued HG-impaired migration, tube formation and survival of BMCs or mature human cardiac microvascular endothelial cells (HCMVECs). Mechanistically, DATS restored nitric oxide production and decreased eNOS-pT495 levels in HCMVECs, and improved BMC angiogenic activity under HG condition. Finally, silencing CSE by siRNA significantly increased eNOS-pT495 levels in HCMVECs. Conclusions Decreased CSE-mediated H2S bioavailability is an underlying source of BMC dysfunction in diabetes. Our data indicate that H2S and overexpression of CSE in diabetic BMCs may rescue their dysfunction and open novel avenues for cell-based therapeutics of CLI in diabetic patients.

Glutathione–Allylsulfur Conjugates as Mesenchymal Stem Cells Stimulating Agents for Potential Applications in Tissue Repair

“…These results are in agreement with the recent studies that demonstrate that H 2 S is able to promote migration and wound healing. The underlying mechanism involves increased levels of p-Akt, p-ERK1/2 and phosphorylated glycogen synthase kinase-3b [81][82][83]. Overall these results demonstrate that prolonged cell GSGa pre-conditioning could represent a powerful tool for selecting adult stem-cell lines that maintain an increased proliferative and migration capability and resistance to oxidative stress, suggesting a potential successful approach for future in vivo/therapeutic applications.…”

Wnt3a involved in the mechanical loading on improvement of bone remodeling and angiogenesis in a postmenopausal osteoporosis mouse model

“…We flushed the iliac and femur with Dulbecco's Modified Eagle's Medium (DMEM) containing 2% FBS. Bone marrow-derived cells were separated by Ficoll density gradient centrifugation and seeded in 6-well dishes (36). Cells were cultured in minimum essential medium alpha (MEM-a) with 30 ng/ ml murine macrophage-colony stimulating factor (M-CSF) and 20 ng/ml RANKL.…”

Protein S‐sulfhydration by hydrogen sulfide in cardiovascular system

“…However, recent publications have revealed that H 2 S is synthesized in mammalian tissues, and freely travels through cell membranes. H 2 S acts independently of any specific transporters, and it has a number of biological effects on various systems (Meng et al, ; Cao and Bian, ; Feliers et al, ; Ianaro et al, ; Katsouda et al, ; Cheng et al, ). Nowadays, H 2 S is regarded as the third endogenous gasotransmitter followed by NO and CO (Hine et al, ).…”