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What Kind of Signaling Maintains Pluripotency and Viability in Human-Induced Pluripotent Stem Cells Cultured on Laminin-511 with Serum-Free Medium?

Nakashima Y, Omasa T - Biores Open Access (2016)

Bottom Line: In hiPSCs, the interaction of laminin-511/α6β1 integrin with the cell-cell adhesion molecule E-cadherin confers protection against apoptosis through the Ras homolog gene family member A (RhoA)/Rho kinase (ROCK) signaling pathway (the major pathways for cell death) and the proto-oncogene tyrosine-protein kinase Fyn (Fyn)-RhoA-ROCK signaling pathway.A combination of growth factors, medium constituents, cell membrane-located E-cadherin, and α6β1 integrin-induced signaling is required for pluripotent cell proliferation and for optimal cell survival on a laminin-511 scaffold.In this review, we discuss and explore the influence of growth factors on the cadherin and integrin signaling pathways in serum-free and xeno-free cultures of hiPSCs during the preparation of products for regenerative medicinal therapies.

View Article: PubMed Central - PubMed

Affiliation: Department of Material and Life Science, Graduate School of Engineering, Osaka University , Osaka, Japan .

ABSTRACT
Xeno-free medium contains no animal-derived components, but is composed of minimal growth factors and is serum free; the medium may be supplemented with insulin, transferrin, and selenium (ITS medium). Serum-free and xeno-free culture of human-induced pluripotent stem cells (hiPSCs) uses a variety of components based on ITS medium and Dulbecco's modified Eagle's medium/Ham's nutrient mixture F12 (DMEM/F12) that contain high levels of iron salt and glucose. Culture of hiPSCs also requires scaffolding materials, such as extracellular matrix, collagen, fibronectin, laminin, proteoglycan, and vitronectin. The scaffolding component laminin-511, which is composed of α5, β1, and γ1 chains, binds to α3β1, α6β1, and α6β4 integrins on the cell membrane to induce activation of the PI3K/AKT- and Ras/MAPK-dependent signaling pathways. In hiPSCs, the interaction of laminin-511/α6β1 integrin with the cell-cell adhesion molecule E-cadherin confers protection against apoptosis through the Ras homolog gene family member A (RhoA)/Rho kinase (ROCK) signaling pathway (the major pathways for cell death) and the proto-oncogene tyrosine-protein kinase Fyn (Fyn)-RhoA-ROCK signaling pathway. The expression levels of α6β1 integrin and E-cadherin on cell membranes are controlled through the activation of insulin receptor/insulin, FGF receptor/FGF2, or activin-like kinase 5 (ALK5)-dependent TGF-β signaling. A combination of growth factors, medium constituents, cell membrane-located E-cadherin, and α6β1 integrin-induced signaling is required for pluripotent cell proliferation and for optimal cell survival on a laminin-511 scaffold. In this review, we discuss and explore the influence of growth factors on the cadherin and integrin signaling pathways in serum-free and xeno-free cultures of hiPSCs during the preparation of products for regenerative medicinal therapies. In addition, we suggest the optimum serum-free medium components for use with laminin-511, a new scaffold for hiPSC culture.

No MeSH data available.


Related in: MedlinePlus

Signaling pathways in serum-free on feeder culture systems. BSA acts as a nutrient for MEFs. MEFs secrete heparan sulfate proteoglycans (HSPG), fibronectin, TGF-β, IGF1, and IGF2. HSA acts as a nutrient for hiPSCs. Albumin in the culture medium promotes expression of collagen I, collagen IV, and HSPG that detoxify β-mercaptoethanol. Collagen I binds to α2β1 integrin and transduces Ras function to activate PI3K binding to AKT; activation of the latter pathway promotes self-renewal in hiPSCs. Collagen IV binds to α2β1 integrin and transduces a RhoGAP function to activate PI3K binding to AKT; activation of the latter pathway promotes cell growth in hiPSCs. With regard to downstream functions in the IGF1R and ALK5 pathways of hiPSCs, activation induces E-cadherin transcription that recruits the PI3K/AKT and ERK/MAPK. This suggests that both PI3K/AKT and MAPK/ERK signaling are required for insulin- and TGF-β-induced E-cadherin upregulation on the cell membrane. Glucose promotes fibronectin and TGF-β manifestation through c-Jun signaling. TGF-β is activated by αVβ6 integrin. The pathways that IGF1R and ALK5 activate through unidentified growth factors, which are present in the culture medium for hiPSCs, are sufficient to maintain pluripotency in hiPSCs. Transferrin regulates cellular iron uptake and, thus, is an essential culture medium ingredient. Selenium is a component of some glutathione peroxidases; these enzymes prevent oxidative damage in cells by inducing the reduction of lipid hydroperoxides. With regard to downstream functions in the FGFR and ALK4 pathways of hiPSCs, the induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by ALK4-activin A signal pathway. Collagen I and HSPG concentrate on FGF2 in the vicinity of FGFR to activate laminin-332, -511, -521, and collagen IV transcription through the PI3K/AKT signaling pathway. Fibronectin is secreted by hiPSCs and binds with α8β1 integrin to promote cell survival of hiPSCs through PI3K/AKT signaling. In addition, fibronectin can also bind to αVβ1 integrin and α5β1 integrin to promote secretion of laminin-332, -511, -521, and collagen IV from hiPSCs. The pathways activated by fibronectin-integrin signaling are responsible for the secretion of ECM, which maintains pluripotency of hiPSCs. ALK4, activin-like kinase receptor-4; BSA, bovine serum albumin; ECM, extracellular matrix; ERK, extracellular signal-regulated kinase; hiPSCs, human-induced pluripotent stem cells; FGFR, fibroblast growth factor receptor; HSA, human serum albumin; IGF, insulin-like growth factor; IGF1R, insulin-like growth factor-1 receptor; MAPK, mitogen-activated protein kinase; MEF, mouse embryonic fibroblast; PI3K, phosphatidylinositol 3-kinase; TGF-β, transforming growth factor β.
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f1: Signaling pathways in serum-free on feeder culture systems. BSA acts as a nutrient for MEFs. MEFs secrete heparan sulfate proteoglycans (HSPG), fibronectin, TGF-β, IGF1, and IGF2. HSA acts as a nutrient for hiPSCs. Albumin in the culture medium promotes expression of collagen I, collagen IV, and HSPG that detoxify β-mercaptoethanol. Collagen I binds to α2β1 integrin and transduces Ras function to activate PI3K binding to AKT; activation of the latter pathway promotes self-renewal in hiPSCs. Collagen IV binds to α2β1 integrin and transduces a RhoGAP function to activate PI3K binding to AKT; activation of the latter pathway promotes cell growth in hiPSCs. With regard to downstream functions in the IGF1R and ALK5 pathways of hiPSCs, activation induces E-cadherin transcription that recruits the PI3K/AKT and ERK/MAPK. This suggests that both PI3K/AKT and MAPK/ERK signaling are required for insulin- and TGF-β-induced E-cadherin upregulation on the cell membrane. Glucose promotes fibronectin and TGF-β manifestation through c-Jun signaling. TGF-β is activated by αVβ6 integrin. The pathways that IGF1R and ALK5 activate through unidentified growth factors, which are present in the culture medium for hiPSCs, are sufficient to maintain pluripotency in hiPSCs. Transferrin regulates cellular iron uptake and, thus, is an essential culture medium ingredient. Selenium is a component of some glutathione peroxidases; these enzymes prevent oxidative damage in cells by inducing the reduction of lipid hydroperoxides. With regard to downstream functions in the FGFR and ALK4 pathways of hiPSCs, the induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by ALK4-activin A signal pathway. Collagen I and HSPG concentrate on FGF2 in the vicinity of FGFR to activate laminin-332, -511, -521, and collagen IV transcription through the PI3K/AKT signaling pathway. Fibronectin is secreted by hiPSCs and binds with α8β1 integrin to promote cell survival of hiPSCs through PI3K/AKT signaling. In addition, fibronectin can also bind to αVβ1 integrin and α5β1 integrin to promote secretion of laminin-332, -511, -521, and collagen IV from hiPSCs. The pathways activated by fibronectin-integrin signaling are responsible for the secretion of ECM, which maintains pluripotency of hiPSCs. ALK4, activin-like kinase receptor-4; BSA, bovine serum albumin; ECM, extracellular matrix; ERK, extracellular signal-regulated kinase; hiPSCs, human-induced pluripotent stem cells; FGFR, fibroblast growth factor receptor; HSA, human serum albumin; IGF, insulin-like growth factor; IGF1R, insulin-like growth factor-1 receptor; MAPK, mitogen-activated protein kinase; MEF, mouse embryonic fibroblast; PI3K, phosphatidylinositol 3-kinase; TGF-β, transforming growth factor β.

Mentions: FGFR1 (fibroblast growth factor receptor 1) is the predominant FGFR in hiPSCs. Binding of FGF2 activates FGFR and major signal transduction pathways, such as all three branches of the PI3K and AKT pathways, the mitogen-activated protein kinase (MAPK) pathway, and the extracellular signal-regulated kinase (ERK)/MAPK pathway. FGF2 activates the MAP kinases ERK1/ERK2, and also PI3K/AKT and c-Jun, and stimulates signaling cascades by activation of activin A.22,23 Induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by the ALK4-activin A signaling pathway. FGF2 signaling may act through the PI3K pathway to regulate the synthesis of pericellular laminin-332, -511, -521, and collagen IV isotypes and provide a framework for the basement membrane of hPSCs.24 Blocking the FGF2 pathway in hPSCs can lead to differentiation.6,8,25 Extracellular matrix (ECM) modulated signals may converge with FGF2 signaling and be involved in the maintenance of pluripotency (Fig. 1).


What Kind of Signaling Maintains Pluripotency and Viability in Human-Induced Pluripotent Stem Cells Cultured on Laminin-511 with Serum-Free Medium?

Nakashima Y, Omasa T - Biores Open Access (2016)

Signaling pathways in serum-free on feeder culture systems. BSA acts as a nutrient for MEFs. MEFs secrete heparan sulfate proteoglycans (HSPG), fibronectin, TGF-β, IGF1, and IGF2. HSA acts as a nutrient for hiPSCs. Albumin in the culture medium promotes expression of collagen I, collagen IV, and HSPG that detoxify β-mercaptoethanol. Collagen I binds to α2β1 integrin and transduces Ras function to activate PI3K binding to AKT; activation of the latter pathway promotes self-renewal in hiPSCs. Collagen IV binds to α2β1 integrin and transduces a RhoGAP function to activate PI3K binding to AKT; activation of the latter pathway promotes cell growth in hiPSCs. With regard to downstream functions in the IGF1R and ALK5 pathways of hiPSCs, activation induces E-cadherin transcription that recruits the PI3K/AKT and ERK/MAPK. This suggests that both PI3K/AKT and MAPK/ERK signaling are required for insulin- and TGF-β-induced E-cadherin upregulation on the cell membrane. Glucose promotes fibronectin and TGF-β manifestation through c-Jun signaling. TGF-β is activated by αVβ6 integrin. The pathways that IGF1R and ALK5 activate through unidentified growth factors, which are present in the culture medium for hiPSCs, are sufficient to maintain pluripotency in hiPSCs. Transferrin regulates cellular iron uptake and, thus, is an essential culture medium ingredient. Selenium is a component of some glutathione peroxidases; these enzymes prevent oxidative damage in cells by inducing the reduction of lipid hydroperoxides. With regard to downstream functions in the FGFR and ALK4 pathways of hiPSCs, the induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by ALK4-activin A signal pathway. Collagen I and HSPG concentrate on FGF2 in the vicinity of FGFR to activate laminin-332, -511, -521, and collagen IV transcription through the PI3K/AKT signaling pathway. Fibronectin is secreted by hiPSCs and binds with α8β1 integrin to promote cell survival of hiPSCs through PI3K/AKT signaling. In addition, fibronectin can also bind to αVβ1 integrin and α5β1 integrin to promote secretion of laminin-332, -511, -521, and collagen IV from hiPSCs. The pathways activated by fibronectin-integrin signaling are responsible for the secretion of ECM, which maintains pluripotency of hiPSCs. ALK4, activin-like kinase receptor-4; BSA, bovine serum albumin; ECM, extracellular matrix; ERK, extracellular signal-regulated kinase; hiPSCs, human-induced pluripotent stem cells; FGFR, fibroblast growth factor receptor; HSA, human serum albumin; IGF, insulin-like growth factor; IGF1R, insulin-like growth factor-1 receptor; MAPK, mitogen-activated protein kinase; MEF, mouse embryonic fibroblast; PI3K, phosphatidylinositol 3-kinase; TGF-β, transforming growth factor β.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4834485&req=5

f1: Signaling pathways in serum-free on feeder culture systems. BSA acts as a nutrient for MEFs. MEFs secrete heparan sulfate proteoglycans (HSPG), fibronectin, TGF-β, IGF1, and IGF2. HSA acts as a nutrient for hiPSCs. Albumin in the culture medium promotes expression of collagen I, collagen IV, and HSPG that detoxify β-mercaptoethanol. Collagen I binds to α2β1 integrin and transduces Ras function to activate PI3K binding to AKT; activation of the latter pathway promotes self-renewal in hiPSCs. Collagen IV binds to α2β1 integrin and transduces a RhoGAP function to activate PI3K binding to AKT; activation of the latter pathway promotes cell growth in hiPSCs. With regard to downstream functions in the IGF1R and ALK5 pathways of hiPSCs, activation induces E-cadherin transcription that recruits the PI3K/AKT and ERK/MAPK. This suggests that both PI3K/AKT and MAPK/ERK signaling are required for insulin- and TGF-β-induced E-cadherin upregulation on the cell membrane. Glucose promotes fibronectin and TGF-β manifestation through c-Jun signaling. TGF-β is activated by αVβ6 integrin. The pathways that IGF1R and ALK5 activate through unidentified growth factors, which are present in the culture medium for hiPSCs, are sufficient to maintain pluripotency in hiPSCs. Transferrin regulates cellular iron uptake and, thus, is an essential culture medium ingredient. Selenium is a component of some glutathione peroxidases; these enzymes prevent oxidative damage in cells by inducing the reduction of lipid hydroperoxides. With regard to downstream functions in the FGFR and ALK4 pathways of hiPSCs, the induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by ALK4-activin A signal pathway. Collagen I and HSPG concentrate on FGF2 in the vicinity of FGFR to activate laminin-332, -511, -521, and collagen IV transcription through the PI3K/AKT signaling pathway. Fibronectin is secreted by hiPSCs and binds with α8β1 integrin to promote cell survival of hiPSCs through PI3K/AKT signaling. In addition, fibronectin can also bind to αVβ1 integrin and α5β1 integrin to promote secretion of laminin-332, -511, -521, and collagen IV from hiPSCs. The pathways activated by fibronectin-integrin signaling are responsible for the secretion of ECM, which maintains pluripotency of hiPSCs. ALK4, activin-like kinase receptor-4; BSA, bovine serum albumin; ECM, extracellular matrix; ERK, extracellular signal-regulated kinase; hiPSCs, human-induced pluripotent stem cells; FGFR, fibroblast growth factor receptor; HSA, human serum albumin; IGF, insulin-like growth factor; IGF1R, insulin-like growth factor-1 receptor; MAPK, mitogen-activated protein kinase; MEF, mouse embryonic fibroblast; PI3K, phosphatidylinositol 3-kinase; TGF-β, transforming growth factor β.
Mentions: FGFR1 (fibroblast growth factor receptor 1) is the predominant FGFR in hiPSCs. Binding of FGF2 activates FGFR and major signal transduction pathways, such as all three branches of the PI3K and AKT pathways, the mitogen-activated protein kinase (MAPK) pathway, and the extracellular signal-regulated kinase (ERK)/MAPK pathway. FGF2 activates the MAP kinases ERK1/ERK2, and also PI3K/AKT and c-Jun, and stimulates signaling cascades by activation of activin A.22,23 Induction of α2β1, α3β1, α6β1, and α6β4 integrin transcription requires the FGF2-FGFR signal pathway and is reinforced by the ALK4-activin A signaling pathway. FGF2 signaling may act through the PI3K pathway to regulate the synthesis of pericellular laminin-332, -511, -521, and collagen IV isotypes and provide a framework for the basement membrane of hPSCs.24 Blocking the FGF2 pathway in hPSCs can lead to differentiation.6,8,25 Extracellular matrix (ECM) modulated signals may converge with FGF2 signaling and be involved in the maintenance of pluripotency (Fig. 1).

Bottom Line: In hiPSCs, the interaction of laminin-511/α6β1 integrin with the cell-cell adhesion molecule E-cadherin confers protection against apoptosis through the Ras homolog gene family member A (RhoA)/Rho kinase (ROCK) signaling pathway (the major pathways for cell death) and the proto-oncogene tyrosine-protein kinase Fyn (Fyn)-RhoA-ROCK signaling pathway.A combination of growth factors, medium constituents, cell membrane-located E-cadherin, and α6β1 integrin-induced signaling is required for pluripotent cell proliferation and for optimal cell survival on a laminin-511 scaffold.In this review, we discuss and explore the influence of growth factors on the cadherin and integrin signaling pathways in serum-free and xeno-free cultures of hiPSCs during the preparation of products for regenerative medicinal therapies.

View Article: PubMed Central - PubMed

Affiliation: Department of Material and Life Science, Graduate School of Engineering, Osaka University , Osaka, Japan .

ABSTRACT
Xeno-free medium contains no animal-derived components, but is composed of minimal growth factors and is serum free; the medium may be supplemented with insulin, transferrin, and selenium (ITS medium). Serum-free and xeno-free culture of human-induced pluripotent stem cells (hiPSCs) uses a variety of components based on ITS medium and Dulbecco's modified Eagle's medium/Ham's nutrient mixture F12 (DMEM/F12) that contain high levels of iron salt and glucose. Culture of hiPSCs also requires scaffolding materials, such as extracellular matrix, collagen, fibronectin, laminin, proteoglycan, and vitronectin. The scaffolding component laminin-511, which is composed of α5, β1, and γ1 chains, binds to α3β1, α6β1, and α6β4 integrins on the cell membrane to induce activation of the PI3K/AKT- and Ras/MAPK-dependent signaling pathways. In hiPSCs, the interaction of laminin-511/α6β1 integrin with the cell-cell adhesion molecule E-cadherin confers protection against apoptosis through the Ras homolog gene family member A (RhoA)/Rho kinase (ROCK) signaling pathway (the major pathways for cell death) and the proto-oncogene tyrosine-protein kinase Fyn (Fyn)-RhoA-ROCK signaling pathway. The expression levels of α6β1 integrin and E-cadherin on cell membranes are controlled through the activation of insulin receptor/insulin, FGF receptor/FGF2, or activin-like kinase 5 (ALK5)-dependent TGF-β signaling. A combination of growth factors, medium constituents, cell membrane-located E-cadherin, and α6β1 integrin-induced signaling is required for pluripotent cell proliferation and for optimal cell survival on a laminin-511 scaffold. In this review, we discuss and explore the influence of growth factors on the cadherin and integrin signaling pathways in serum-free and xeno-free cultures of hiPSCs during the preparation of products for regenerative medicinal therapies. In addition, we suggest the optimum serum-free medium components for use with laminin-511, a new scaffold for hiPSC culture.

No MeSH data available.


Related in: MedlinePlus