Alkaline phosphatase is an enzyme commonly expressed in almost all living organisms. In humans and other mammals, determinations of the expression and activity of alkaline phosphatase have frequently been used for cell determination in developmental studies and/or within clinical trials. Alkaline phosphatase also seems to be one of the key markers in the identification of pluripotent embryonic stem as well as related cells. However, alkaline phosphatases exist in some isoenzymes and isoforms, which have tissue specific expressions and functions. Here, the role of alkaline phosphatase as a stem cell marker is discussed in detail. First, we briefly summarize contemporary knowledge of mammalian alkaline phosphatases in general. Second, we focus on the known facts of its role in and potential significance for the identification of stem cells.

Argon plasma treatment was used to modify the surface of atelocollagen films using a plasmochemical reactor. To evaluate the effects of the treatment, the untreated and treated samples were characterized by Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (ATR-FTIR), Scanning Electron Microscopy (SEM) imaging, and X-ray Photoelectron Spectroscopy (XPS) techniques. Cell growth was carried out by culturing human immortalized keratinocyte (HaCaT) cells and proliferation was measured via MTT assay. It was observed that argon plasma treatment significantly enhanced the extent of cell proliferation, which was ascribed to the favourable role of plasma treatment in inducing surface oxygen-containing entities together with increasing surface roughness. This can be considered as a potentially promising approach for tissue regeneration purposes.

Reactive oxygen species (ROS) are important regulators of cellular functions. In embryonic stem cells, ROS are suggested to influence differentiation status. Regulated ROS formation is catalyzed primarily by NADPH‐dependent oxidases (NOXs). Apocynin and diphenyleneiodonium are frequently used inhibitors of NOXs; however, both exhibit uncharacterized effects not related to NOXs inhibition. Interestingly, in our model of mouse embryonic stem cells we demonstrate low expression of NOXs. Therefore we aimed to clarify potential side effects of these drugs. Both apocynin and diphenyleneiodonium impaired proliferation of cells. Surprisingly, we observed prooxidant activity of these drugs determined by hydroethidine. Further, we revealed that apocynin inhibits PI3K/Akt pathway with its downstream transcriptional factor Nanog. Opposite to this, apocynin augmented activity of canonical Wnt signaling. On the contrary, diphenyleneiodonium activated both PI3K/Akt and Erk signaling pathways without affecting Wnt. Our data indicates limits and possible unexpected interactions of NOXs inhibitors with intracellular signaling pathways.

Hypoxia is involved in the regulation of stem cell fate, and hypoxia‐inducible factor 1 (HIF‐1) is the master regulator of hypoxic response. Here, we focus on the effect of hypoxia on intracellular signaling pathways responsible for mouse embryonic stem (ES) cell maintenance. We employed wild‐type and HIF‐1α‐deficient ES cells to investigate hypoxic response in the ERK, Akt, and STAT3 pathways. Cultivation in 1% O₂ for 24 h resulted in the strong dephosphorylation of ERK and its upstream kinases and to a lesser extent of Akt in an HIF‐1‐independent manner, while STAT3 phosphorylation remained unaffected. Downregulation of ERK could not be mimicked either by pharmacologically induced hypoxia or by the overexpression. Dual‐specificity phosphatases (DUSP) 1, 5, and 6 are hypoxia‐sensitive MAPK‐specific phosphatases involved in ERK downregulation, and protein phosphatase 2A (PP2A) regulates both ERK and Akt. However, combining multiple approaches, we revealed the limited significance of DUSPs and PP2A in the hypoxia‐mediated attenuation of ERK signaling. Interestingly, we observed a decreased reactive oxygen species (ROS) level in hypoxia and a similar phosphorylation pattern for ERK when the cells were supplemented with glutathione. Therefore, we suggest a potential role for the ROS‐dependent attenuation of ERK signaling in hypoxia, without the involvement of HIF‐1.

Organic semiconductors are constantly gaining interest in regenerative medicine. Their tunable physico-chemical properties, including electrical conductivity, are very promising for the control of stem-cell differentiation. However, their use for combined material-based and electrical stimulation remains largely underexplored. Therefore, we carried out a study on whether a platform based on the conductive polymer poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS) can be beneficial to the differentiation of mouse embryonic stem cells (mESCs). The platform was prepared using the layout of a standard 24-well cell-culture plate. Polyethylene naphthalate foil served as the substrate for the preparation of interdigitated gold electrodes by physical vapor deposition. The PEDOT:PSS pattern was fabricated by precise screen printing over the gold electrodes. The PEDOT:PSS platform was able to produce higher electrical current with the pulsed-direct-current (DC) electrostimulation mode (1 Hz, 200 mV/mm, 100 ms pulse duration) compared to plain gold electrodes. There was a dominant capacitive component. In proof-of-concept experiments, mESCs were able to respond to such electrostimulation by membrane depolarization and elevation of cytosolic calcium. Further, the PEDOT:PSS platform was able to upregulate cardiomyogenesis and potentially inhibit early neurogenesis per se with minor contribution of electrostimulation. Hence, the present work highlights the large potential of PEDOT:PSS in regenerative medicine.