Different Effects of Insulin-Like Growth Factor-1 and Insulin-Like Growth Factor-2 on Myogenic Differentiation of Human Mesenchymal Stem Cells

Abstract

Insulin-like growth factors (IGFs) are critical components of the stem cell niche, as they regulate proliferation and differentiation of stem cells into different lineages, including skeletal muscle. We have previously reported that insulin-like growth factor binding protein-6 (IGFBP-6), which has high affinity for IGF-2, alters the differentiation process of placental mesenchymal stem cells (PMSCs) into skeletal muscle. In this study, we determined the roles of IGF-1 and IGF-2 and their interactions with IGFBP-6. We showed that IGF-1 increased IGFBP-6 levels within 24 hours but decreased after 3 days, while IGF-2 maintained higher levels of IGFBP-6 throughout myogenesis. IGF-1 increased IGFBP-6 in the early phase as a requirement for muscle commitment. In contrast, IGF-2 enhanced muscle differentiation as shown by the expression of muscle differentiation markers MyoD, MyoG, and MHC. IGF-1 and IGF-2 had different effects on muscle differentiation with IGF-1 promoting early commitment to muscle and IGF-2 promoting complete muscle differentiation. We also showed that PMSCs acquired increasing capacity to synthesize IGF-2 during muscle differentiation, and the capacity increased as the differentiation progressed suggesting an autocrine and/or paracrine effect. Additionally, we demonstrated that IGFBP-6 could enhance the muscle differentiation process in the absence of IGF-2.

Insulin-Like Growth Factor Binding Protein-6 Promotes the Differentiation of Placental Mesenchymal Stem Cells into Skeletal Muscle Independent of Insulin-Like Growth Factor Receptor-1 and Insulin Receptor

“…IGFBP-6 is expressed in the developing cells [23, 26–28]. We have demonstrated that the balance between intracellular and extracellular IGFBP-6 levels is required for modulating muscle differentiation by PMSCs [26] and that the effects of IGFBP-6 on muscle differentiation are both IGF-dependent and IGF-independent [31]. These findings provided basic insight into the role of IGFBP-6 and IGFs on PMSC muscle differentiation.…”

“…For IGFBP-6 supplementation with the inhibitors, recombinant human IGFBP-6 (ProSpec) was added to the media (375 ng/mL) every 3 days at the time of media change. The dose of IGFBP-6 was based on our previous studies [26, 31].…”

“…After washing, HRP-conjugated streptavidin was added; then, a TMB substrate solution was used to develop a blue color in proportion to the amount of IGFBP-6 or IGF-2 bound. The stop solution changes color from blue to yellow, and the intensity was measured at 450 nm using the Multiskan Ascent plate reader and analysis software [26, 31].…”

“…Cells were washed and resuspended in 500 μ L of ice-cold Aldefluor assay buffer, and ALDH activity was measured using flow cytometry. As a negative control, Aldefluor™ DEAB reagent was used [26, 31].…”

Mesenchymal Stem Cell-Based Immunomodulation: Properties and Clinical Application

“…Mechanisms of interaction were shown to rely on cell–cell contact working in collaboration with secretion of soluble immune factors to induce MSC-regulated immunosuppression [ 21 ]. These specific modulators, including a multitude of immune-modulatory factors, cytokines, and growth factors, modulate inflammatory responses and balance immune profiles [ 22 ]. Namely, soluble immune secretomes, such as prostaglandin E2 (PGE-2), indoleamine 2,3-dioxygenase (IDO), or nitric oxide (NO), respond to immune cells to activate immunoregulation by MSCs [ 23 ].…”