Human pluripotent stem cells have the potential to change the way in which human diseases are cured. Clinical-grade human embryonic stem cells and human induced pluripotent stem cells have to be created according to current good manufacturing practices and regulations. Quality and safety must be of the highest importance when humans' lives are at stake. With the rising number of clinical trials, there is a need for a consensus on hPSCs characterization. Here, we summarize mandatory and 'for information only' characterization methods with release criteria for the establishment of clinical-grade hPSC lines.

• Klíčová slova
• cGMP, cell therapy, characterization, clinical, hESC, hPSCs, hiPSC, human embryonic stem cells, human induced pluripotent stem cells, human pluripotent stem cells,
• MeSH
• Bacteria MeSH
• buněčná a tkáňová terapie metody   MeSH
• endotoxiny MeSH
• indukované pluripotentní kmenové buňky MeSH
• lidé MeSH
• lidské embryonální kmenové buňky MeSH
• Mycoplasma MeSH
• pluripotentní kmenové buňky * MeSH
• viry MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• endotoxiny MeSH

Neural differentiation of human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) can produce a valuable and robust source of human neural cell subtypes, holding great promise for the study of neurogenesis and development, and for treating neurological diseases. However, current hESCs and hiPSCs neural differentiation protocols require either animal factors or embryoid body formation, which decreases efficiency and yield, and strongly limits medical applications. Here we develop a simple, animal-free protocol for neural conversion of both hESCs and hiPSCs in adherent culture conditions. A simple medium formula including insulin induces the direct conversion of >98% of hESCs and hiPSCs into expandable, transplantable, and functional neural progenitors with neural rosette characteristics. Further differentiation of neural progenitors into dopaminergic and spinal motoneurons as well as astrocytes and oligodendrocytes indicates that these neural progenitors retain responsiveness to instructive cues revealing the robust applicability of the protocol in the treatment of different neurodegenerative diseases. The fact that this protocol includes animal-free medium and human extracellular matrix components avoiding embryoid bodies makes this protocol suitable for the use in clinic. Stem Cells Translational Medicine 2017;6:1217-1226.

• Klíčová slova
• Cellular therapy, Clinical translation, Differentiation, Embryonic stem cells, Induced pluripotent stem cells, Neural differentiation, Pluripotent stem cells,
• MeSH
• buněčná a tkáňová terapie MeSH
• buněčná diferenciace fyziologie   MeSH
• embryonální kmenové buňky fyziologie   MeSH
• indukované pluripotentní kmenové buňky cytologie   MeSH
• kultivované buňky MeSH
• lidé MeSH
• pluripotentní kmenové buňky cytologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Significance: Since their discovery, induced pluripotent stem cells (iPSCs) had generated considerable interest in the scientific community for their great potential in regenerative medicine, disease modeling, and cell-based therapeutic approach, due to their unique characteristics of self-renewal and pluripotency. Recent Advances: Technological advances in iPSC genome-wide epigenetic profiling led to the elucidation of the epigenetic control of cellular identity during nuclear reprogramming. Moreover, iPSC physiology and metabolism are tightly regulated by oxidation-reduction events that mainly occur during the respiratory chain. In theory, iPSC-derived differentiated cells would be ideal for stem cell transplantation as autologous cells from donors, as the risks of rejection are minimal. Critical Issues: However, iPSCs experience high oxidative stress that, in turn, confers a high risk of increased genomic instability, which is most often linked to DNA repair deficiencies. Genomic instability has to be assessed before iPSCs can be used in therapeutic designs. Future Directions: This review will particularly focus on the links between redox balance and epigenetic modifications-in particular based on the histone variant macroH2A1-that determine DNA damage response in iPSCs and derived differentiated cells, and that might be exploited to decrease the teratogenic potential on iPSC transplantation. Antioxid. Redox Signal. 34, 335-349.

• Klíčová slova
• DNA damage, histone variant macroH2A1, induced pluripotent stem cells (iPSCs), oxidative stress,
• MeSH
• buněčná diferenciace * genetika   MeSH
• buněčná sebeobnova MeSH
• epigeneze genetická * MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• lidé MeSH
• metylace DNA MeSH
• mitochondrie genetika   metabolismus   MeSH
• nádorová transformace buněk genetika   metabolismus   MeSH
• nestabilita genomu MeSH
• oxidace-redukce * MeSH
• oxidační stres MeSH
• oxidativní fosforylace MeSH
• pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• přeprogramování buněk genetika   MeSH
• regenerativní lékařství MeSH
• transplantace kmenových buněk MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Hereditary retinal dystrophies, specifically retinitis pigmentosa (RP) are clinically and genetically heterogeneous diseases affecting primarily retinal cells and retinal pigment epithelial cells with blindness as a final outcome. Understanding the pathogenicity behind these diseases has been largely precluded by the unavailability of affected tissue from patients, large genetic heterogeneity and animal models that do not faithfully represent some human diseases. A landmark discovery of human induced pluripotent stem cells (hiPSCs) permitted the derivation of patient-specific cells. These cells have unlimited self-renewing capacity and the ability to differentiate into RP-affected cell types, allowing the studies of disease mechanism, drug discovery, and cell replacement therapies, both as individual cell types and organoid cultures. Together with precise genome editing, the patient specific hiPSC technology offers novel strategies for targeting the pathogenic mutations and design therapies toward retinal dystrophies. This study summarizes current hiPSC-based RP models and highlights key achievements and challenges of these cellular models, as well as questions that still remain unanswered. Stem Cells 2018;36:474-481.

• Klíčová slova
• Differentiation, Gene targeting, Induced pluripotent stem, Induced pluripotent stem cells, Retina, Retinal photoreceptors, Retinal pigmented epithelium,
• MeSH
• autologní štěp MeSH
• buněčná diferenciace * MeSH
• editace genu * MeSH
• genom lidský * MeSH
• indukované pluripotentní kmenové buňky metabolismus   patologie   MeSH
• lidé MeSH
• modely nemocí na zvířatech MeSH
• retinopathia pigmentosa * genetika   metabolismus   patologie   terapie   MeSH
• transplantace kmenových buněk * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

Induced pluripotent stem (iPS) cells are derived from differentiated cells by different reprogramming techniques, by introducing specific transcription factors responsible for pluripotency. Induced pluripotent stem cells can serve as an excellent source for differentiated neural stem/progenitor cells (NSCs/NPs). Several methods and protocols are utilized to create a robust number of NSCs/NPs without jeopardizing the safety issues required for in vivo applications. A variety of disease-specific iPS cells have been used to study nervous system diseases. In this chapter, we will focus on some of the derivation and differentiation approaches and the application of iPS-NPs in the treatment of spinal cord injury and stroke.

• Klíčová slova
• Induced pluripotent stem cells, Neural stem cells, Neuronal differentiation, Spinal cord injury, Stroke,
• MeSH
• buněčná diferenciace * MeSH
• cévní mozková příhoda patologie   terapie   MeSH
• indukované pluripotentní kmenové buňky cytologie   MeSH
• lidé MeSH
• modely neurologické * MeSH
• nervové kmenové buňky cytologie   MeSH
• poranění míchy patologie   terapie   MeSH
• přeprogramování buněk MeSH
• transkripční faktory metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• transkripční faktory MeSH

New approaches in regenerative medicine and vasculogenesis have generated a demand for sufficient numbers of human endothelial cells (ECs). ECs and their progenitors reside on the interior surface of blood and lymphatic vessels or circulate in peripheral blood; however, their numbers are limited, and they are difficult to expand after isolation. Recent advances in human induced pluripotent stem cell (hiPSC) research have opened possible avenues to generate unlimited numbers of ECs from easily accessible cell sources, such as the peripheral blood. In this study, we reprogrammed peripheral blood mononuclear cells, human umbilical vein endothelial cells (HUVECs), and human saphenous vein endothelial cells (HSVECs) into hiPSCs and differentiated them into ECs. The phenotype profiles, functionality, and genome stability of all hiPSC-derived ECs were assessed and compared with HUVECs and HSVECs. hiPSC-derived ECs resembled their natural EC counterparts, as shown by the expression of the endothelial surface markers CD31 and CD144 and the results of the functional analysis. Higher expression of endothelial progenitor markers CD34 and kinase insert domain receptor (KDR) was measured in hiPSC-derived ECs. An analysis of phosphorylated histone H2AX (γH2AX) foci revealed that an increased number of DNA double-strand breaks upon reprogramming into pluripotent cells. However, differentiation into ECs restored a normal number of γH2AX foci. Our hiPSCs retained a normal karyotype, with the exception of the HSVEC-derived hiPSC line, which displayed mosaicism due to a gain of chromosome 1. Peripheral blood from adult donors is a suitable source for the unlimited production of patient-specific ECs through the hiPSC interstage. hiPSC-derived ECs are fully functional and comparable to natural ECs. The protocol is eligible for clinical applications in regenerative medicine, if the genomic stability of the pluripotent cell stage is closely monitored.

• Klíčová slova
• endothelial differentiation, induced pluripotent stem cells, peripheral blood mononuclear cells,
• MeSH
• biologické markery metabolismus   MeSH
• buněčná diferenciace fyziologie   MeSH
• endoteliální buňky pupečníkové žíly (lidské) cytologie   metabolismus   MeSH
• endoteliální buňky cytologie   metabolismus   MeSH
• fibroblasty cytologie   metabolismus   MeSH
• fyziologická neovaskularizace fyziologie   MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• kultivované buňky MeSH
• leukocyty mononukleární cytologie   metabolismus   MeSH
• lidé MeSH
• regenerativní lékařství metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biologické markery MeSH

The generation of haematopoietic progenitors from human pluripotent stem cells (hPSCs) presents great promise for cell-replacement therapies. However, current protocols for haematopoietic differentiation of hPSCs suffer from low efficiency and functional defects in the derived cells. The technology is also limited by variable ability of hPSC lines to generate blood cells in vitro. To address this issue, methodologies for haematopoietic differentiation in feeder-free conditions were applied to available human embryonic stem cell (hESC) and human induced pluripotent stem cell (hiPSC) lines in this study. It was found that these cell lines did not generate haematopoietic progenitors to such an extent as did H1 and H9 hESC lines that were used for this purpose in the vast majority of relevant studies. These results suggest that for clinical application of blood cells derived from hPSCs, possibly from autologous hiPSCs, it is necessary to overcome the variability in the haematopoietic developmental potential of individual hPSC lines.

• MeSH
• buněčná diferenciace MeSH
• buněčné linie MeSH
• hematopoetické kmenové buňky cytologie   MeSH
• indukované pluripotentní kmenové buňky cytologie   MeSH
• lidé MeSH
• pluripotentní kmenové buňky cytologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The generation of human pluripotent stem cells (hPSCs) of sufficient quantity and quality remains a major challenge for biomedical application. Here we present an efficient feeder-free, high-density monolayer system in which hPSCs become SSEA-3-high and gradually more viable than their feeder-dependent counterparts without changes attributed to culture adaptation. As a consequence, monolayer hPSCs possess advantages over their counterparts in embryoid body development, teratoma formation, freezing as a single-cell suspension, and colony-forming efficiency. Importantly, this monolayer culture system is reversible, preserving the competence of hPSCs to gradually reacquire features of colony growth, if necessary. Therefore, the monolayer culture system is highly suitable for long-term, large-scale propagation of hPSCs, which is necessary in drug development and pluripotent stem cell-based therapies.

• MeSH
• analýza kolonii tvořících jednotek MeSH
• biologické markery metabolismus   MeSH
• buněčné kultury metody   MeSH
• buněčné linie MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• lidé MeSH
• myši MeSH
• pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• podkladové buňky cytologie   MeSH
• proliferace buněk MeSH
• teratom patologie   MeSH
• viabilita buněk MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biologické markery MeSH

The successful development and characterization of human induced pluripotent stem cells (iPSCs) provides a powerful tool to study the molecular mechanisms that control cell fate decisions and differentiation toward distinct lineages. Here we focus on the ability of donors derived iPSCs to differentiate toward hematopoietic progenitor cells and on the analysis of their telomere length. The ability to screen telomere length in individual donors is important for defining cellular senescence, which correlates with their differentiation potential toward hematopoietic lineages. We have modified iPSC culture protocol and telomere length analysis to suit for high throughput screening of telomere length in large number of individual donors. This approach can be used to demonstrate the heterogeneity or changes of telomere length and its shortening as an exclusion criterion for selection of suitable donors for future stem cell therapies.

• Klíčová slova
• Cellular senescence, Hematopoiesis, Hematopoietic stem cells, Induced pluripotent stem cells, Telomere length,
• MeSH
• biologické markery * MeSH
• buněčné kultury * MeSH
• čipová analýza tkání metody   MeSH
• hematopoetické kmenové buňky cytologie   metabolismus   MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• lidé MeSH
• rychlé screeningové testy * MeSH
• stárnutí buněk * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biologické markery * MeSH

The genomic destabilization associated with the adaptation of human embryonic stem cells (hESCs) to culture conditions or the reprogramming of induced pluripotent stem cells (iPSCs) increases the risk of tumorigenesis upon the clinical use of these cells and decreases their value as a model for cell biology studies. Base excision repair (BER), a major genomic integrity maintenance mechanism, has been shown to fail during hESC adaptation. Here, we show that the increase in the mutation frequency (MF) caused by the inhibition of BER was similar to that caused by the hESC adaptation process. The increase in MF reflected the failure of DNA maintenance mechanisms and the subsequent increase in MF rather than being due solely to the accumulation of mutants over a prolonged period, as was previously suggested. The increase in the ionizing-radiation-induced MF in adapted hESCs exceeded the induced MF in nonadapted hESCs and differentiated cells. Unlike hESCs, the overall DNA maintenance in iPSCs, which was reflected by the MF, was similar to that in differentiated cells regardless of the time spent in culture and despite the upregulation of several genes responsible for genome maintenance during the reprogramming process. Taken together, our results suggest that the changes in BER activity during the long-term cultivation of hESCs increase the mutagenic burden, whereas neither reprogramming nor long-term propagation in culture changes the MF in iPSCs.

• MeSH
• buněčná diferenciace účinky záření   MeSH
• buněčné linie MeSH
• genetické lokusy * MeSH
• hypoxanthinfosforibosyltransferasa genetika   metabolismus   MeSH
• indukované pluripotentní kmenové buňky cytologie   metabolismus   MeSH
• lidé MeSH
• mutační rychlost * MeSH
• záření gama MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• hypoxanthinfosforibosyltransferasa MeSH