Maintaining the pluripotency of mouse embryonic stem cells on gold nanoparticle layers with nanoscale but not microscale surface roughness

Abstract

Efficient control of the self-renewal and pluripotency maintenance of embryonic stem cell (ESC) is a prerequisite for translating stem cell technologies to clinical applications. Surface topography is one of the most important factors that regulates cell behaviors. In the present study, micro/nano topographical structures composed of a gold nanoparticle layer (GNPL) with nano-, sub-micro-, and microscale surface roughnesses were used to study the roles of these structures in regulating the behaviors of mouse ESCs (mESCs) under feeder-free conditions. The distinctive results from Oct-4 immunofluorescence staining and quantitative real-time polymerase chain reaction (qPCR) demonstrate that nanoscale and low sub-microscale surface roughnesses (Rq less than 392 nm) are conducive to the long-term maintenance of mESC pluripotency, while high sub-microscale and microscale surface roughnesses (Rq greater than 573 nm) result in a significant loss of mESC pluripotency and a faster undirectional differentiation, particularly in long-term culture. Moreover, the likely signalling cascades engaged in the topological sensing of mESCs were investigated and their role in affecting the maintenance of the long-term cell pluripotency was discussed by analyzing the expression of proteins related to E-cadherin mediated cell-cell adhesions and integrin-mediated focal adhesions (FAs). Additionally, the conclusions from MTT, cell morphology staining and alkaline phosphatase (ALP) activity assays show that the surface roughness can provide a potent regulatory signal for various mESC behaviors, including cell attachment, proliferation and osteoinduction.

Iron overload induced death of osteoblasts in vitro: involvement of the mitochondrial apoptotic pathway

“…Next, ALP activity in the samples was measured by p-nitrophenyl phosphate method using a Alkaline Phosphatase Assay Kit (Beyotime Institute of Biotechnology, Jiangsu, China) as previous report (Lyu et al, 2014). …”

Nanostructured Brownian Surfaces Prepared through Two-Photon Polymerization: Investigation of Stem Cell Response

“…This finding differs from results obtained by applying other physical cues (i.e., roughness and stiffness) that are instead well-known to affect cell stemness. 38,39 Concerning MSC cytoskeleton organization, we noticed a gradual change from nominally flat conditions to highly fractal surfaces (H = 0.01), involving vinculin, f-actin and g-actin expression. In particular, the increase of the surface fractal dimension is accompanied by a gradual disorganization of the actin stress fibers and by a reduction of the vinculin cluster size.…”

Structurally Tunable Reduced Graphene Oxide Substrate Maintains Mouse Embryonic Stem Cell Pluripotency

“…[13,14,16] Here, we examine the protein expression levels of β-catenin and E-cadherin in ESCs and find that the expression levels of β-catenin and E-cadherin in ESCs cultured on RGO-50, RGO-30, and RGO-15 substrates are similar to that of the feeder group (Figure 7a,b; Figure S3a Long-term cultivation of ESC on RGO substrate. [13,14,16] Here, we examine the protein expression levels of β-catenin and E-cadherin in ESCs and find that the expression levels of β-catenin and E-cadherin in ESCs cultured on RGO-50, RGO-30, and RGO-15 substrates are similar to that of the feeder group (Figure 7a,b; Figure S3a Long-term cultivation of ESC on RGO substrate.…”

“…[10,11] Therefore, xenogeneic-free and chemically well-defined synthetic substrates provide unique advantages in culturing ESCs. [14][15][16][17] However, the substrates mentioned above have common disadvantages, such as they have poor chemical properties, are unable to arbitrarily adjust the content of oxygen-containing groups, are difficult to be modified by proteins, cannot be mass-produced, exhibit changes in properties after folding or bending, and cannot be transplanted into the body as a scaffold. [12] Human ESCs maintain pluripotency on polystyrene nanopillar substrates with a dia meter of 120-170 nm, mainly due to cell-nanotopography interactions that regulate focal adhesion formation and cytoskeleton reorganization, and enhance E-cadherin-mediated cell-cell adhesion in clones.…”

“…The ECM and the extracellular domain of cadherin, a type of transmembrane glycoprotein, play an important role in cell adhesion. [13,14,16] 3D elastin-like protein hydrogels are able to maintain neural progenitor cell (NPC) stemness via the modulation of cadherin-mediated β-catenin signaling. [26] The cytoplasmic domains of E-cadherin and β-catenin are closely related in mediating cell adhesion and stem maintenance.…”