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Involvement of neuropeptide Y and its Y1 and Y5 receptors in maintaining self-renewal and proliferation of human embryonic stem cells.

ABSTRACT: Neuropeptide Y (NPY) and NPY receptors are widely expressed in various organs and cell types and have been shown to have pleiotropic functions. However, their presence or role in human embryonic stem cells (hESCs) remains unknown. We now show that undifferentiated hESCs primarily express NPY and its Y1 and Y5 receptors. Inhibition of NPY signalling using either the selective NPY Y1 or Y5 receptor antagonist reduces the maintenance of self-renewal and proliferation of undifferentiated hESCs. We also provide compelling evidence that exogenous NPY supports the long-term growth of undifferentiated hESCs in the absence of feeder cell factors using only knockout serum replacement media. Further, NPY facilitates the use of chemically defined medium made up of N2/B27 supplement and basic fibroblast growth factor (bFGF) for hESC feeder-free culture. Our results indicate that both Y1 and Y5 receptors appear to be involved in the NPY-mediated activation of AKT/protein kinase B and extracellular signal-regulated kinase 1/2 (ERK1/2) in hESCs. Notably, only Y1 receptor, but not Y5 receptor, is responsible for the NPY-induced activation of cAMP-response element binding (CREB) in hESCs. These results provide the first evidence that NPY and its Y1 and Y5 receptors have potential role in maintaining hESC self-renewal and pluripotency. We demonstrate the underlying importance of NPY signalling and its usefulness in the development of a defined and xeno-free culture condition for the large-scale propagation of undifferentiated hESCs.

SUBMITTER: Son MY

PROVIDER: S-EPMC3822502 | biostudies-literature | 2011 Jan

REPOSITORIES: biostudies-literature

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Involvement of neuropeptide Y and its Y1 and Y5 receptors in maintaining self-renewal and proliferation of human embryonic stem cells.

Son Mi-Young MY Kim Min-Jeong MJ Yu Kweon K Koo Deog-Bon DB Cho Yee Sook YS

Journal of cellular and molecular medicine 20110101 1

Neuropeptide Y (NPY) and NPY receptors are widely expressed in various organs and cell types and have been shown to have pleiotropic functions. However, their presence or role in human embryonic stem cells (hESCs) remains unknown. We now show that undifferentiated hESCs primarily express NPY and its Y1 and Y5 receptors. Inhibition of NPY signalling using either the selective NPY Y1 or Y5 receptor antagonist reduces the maintenance of self-renewal and proliferation of undifferentiated hESCs. We a ...[more]

PMID: 19874423

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Project description:The increased proliferation and migration of vascular smooth muscle cells (VSMCs) play important roles in pathophysiological remodeling of arteries during hypertension in pregnancy. However, the mechanisms involved in this process remain unclear. We hypothesized that Neuropeptide Y (NPY), which is a potent mitogenic peptide, participates in modulating proliferation and migration of VSMCs during hypertension in pregnancy. Using pregnant hypertensive rats, induced by intraperitoneal injection of L-nitro-arginine methylester (L-NAME), the plasma concentration of NPY was detected. Open angle, which reflects the non-uniform remodeling with high sensitivity, was used to detect the pathophysiological vascular remodeling in vivo. The results revealed that NPY concentration and artery open angle were both significantly increased in rats with hypertension in pregnant. The underlying mechanism of elevated NPY on vascular remodeling were further analyzed by using cultured VSMCs in vitro. In cultured VSMCs, NPY most effectively stimulated the migration and proliferation of VSMCs at 10-6 mol/L, similar to the plasma concentration in L-NAME hypertension in pregnant rats. NPY up-regulated the expressions of both Y1 and Y5 receptors, increased the phosphorylations of STAT3 on Tyr705 and Ser727 residues, and induced the expression of c-Fos. The NPY-induced VSMCs proliferation was reduced by Y5 receptor antagonist, and fully blocked by combinations with other antagonist, such as Y2+Y5, Y1+Y5, and Y1+Y2+Y5. In contrast, the NPY-induced VSMC migration was blocked by either Y receptor antagonist or any combination of Y receptor antagonists. These results suggest that the elevated plasma concentration of NPY during hypertension in pregnancy may induce VSMC proliferation mainly via Y5 receptor, which subsequently modulate STAT3 and c-Fos signaling pathways to result in the vascular remodeling. These results also suggest that NPY mainly acts on VSMCs in vitro via Y1, Y5 receptors and in vascular tissues in vivo via Y5 receptor.

Project description:Successful realization of the enormous potential of pluripotent stem cells in regenerative medicine demands the development of well-defined culture conditions. Maintenance of embryonic stem cells (ESCs) typically requires co-culture with feeder layer cells, generally mouse embryonic fibroblasts (MEFs). Concerns about xenogeneic pathogen contamination and immune reaction to feeder cells underlie the need for ensuring the safety and efficacy of future stem cell-based products through the development of a controlled culture environment. To gain insight into the effectiveness of MEF layers, here we have developed a biomimetic synthetic feeder layer (BSFL) that is acellular and replicates the stiffness and topography of MEFs. The mechanical properties of MEFs were measured using atomic force microscopy. The average Young's modulus of the MEF monolayers was replicated using tunable polyacrylamide (PA) gels. BSFLs replicated topographical features of the MEFs, including cellular, subcellular, and cytoskeletal features. On BSFLs, mouse ESCs adhered and formed compact round colonies; similar to on MEF controls but not on Flat PA. ESCs on BSFLs maintained their pluripotency and self-renewal across passages, formed embryoid bodies and differentiated into progenitors of the three germ layers. This acellular biomimetic synthetic feeder layer supports stem cell culture without requiring co-culture of live xenogeneic feeder cells, and provides a versatile, tailorable platform for investigating stem cell growth.Embryonic stem cells have enormous potential to aid therapeutics, because they can renew themselves and become different cell types. This study addresses a key challenge for ESC use - growing them safely for human patients. ESCs typically grow with a feeder layer of mouse fibroblasts. Since patients have a risk of immune response to feeder layer cells, we have developed a material to mimic the feeder layer and eliminate this risk. We investigated the influence of feeder layer topography and stiffness on mouse ESCs. While the biomimetic synthetic feeder layer contains no live cells, it replicates the stiffness and topography of feeder layer cells. Significantly, ESCs grown on BSFLs retain their abilities to grow and become multiple cell types.

Dataset Information

Involvement of neuropeptide Y and its Y1 and Y5 receptors in maintaining self-renewal and proliferation of human embryonic stem cells.

Publications

Involvement of neuropeptide Y and its Y1 and Y5 receptors in maintaining self-renewal and proliferation of human embryonic stem cells.

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