Andrew Laslett

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Dominic D'Agostino

University of South Florida

David Vago

Vanderbilt University

Sudhir K Shukla

Bhabha Atomic Research Centre

YIH-SHARNG CHEN

National Taiwan University

Alaa Al-Charrakh

University of Babylon

Phillip Tully

University of Adelaide

Andrea Chiba

University of California, San Diego

Ranil Jayawardena

University of Colombo, Sri Lanka

Pavel Rozsíval

Charles University, Prague

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University of Pretoria

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Papers by Andrew Laslett

Restricted ethnic diversity in human embryonic stem cell lines

by Eyitayo S Fakunle and Andrew Laslett

Nature Methods, 2010

Restricted ethnic diversity in human embryonic stem cell lines To the Editor: Human pluripotent s... more Restricted ethnic diversity in human embryonic stem cell lines To the Editor: Human pluripotent stem cells (hPSCs) have the capacity to self-renew indefinitely and to differentiate into a wide array of cell types, which make them a potential source of essentially unlimited quantities of differentiated cells for basic and clinical research. The tremendous self-renewal of hPSCs might lead one to conclude that a small number of cell lines would be sufficient to meet all needs. However, it is becoming increasingly clear that the genetic background of human cell lines can have significant effects on experimental results. Although hundreds of associations between individual alleles and specific traits or diseases are discovered each year, a full understanding of the relationships between genetic variation and cellular or organismal phenotype is still remote 1. The HapMap project 2 , along with many other studies, has made it clear that there are large differences in the allelic frequencies for many single-nucleotide polymorphisms (SNPs) among different ethnicities. Ethnicity can serve as a proxy for genetic variation to ensure diversity in genetic backgrounds in a study population. The availability of hPSCs from a variety of ethnic backgrounds would ensure the generalizability of results as well as increasing the likelihood that specific alleles or combinations of alleles of interest will be available for study. This is particularly important for the use of hPSCs and hPSC-derived cells for drug screening and toxicity studies. Several idiosyncratic drug effects have been attributed to specific genetic variations 3 , and the efficacy and toxicity of numerous drugs are presumed to be influenced by genetic factors. On the clinical side, hPSCs and their derivatives are potential sources for cell therapy, and recipients of cell transplants are more likely to find immunologic matches with donors who share their ethnic background. However, because most hPSC lines have been generated from de-identified material, the ethnic backgrounds of the donors are not known. We determined the ethnic origin of 47 human embryonic stem cell (hESC) lines (including 42 standard hESC lines and 5 parthenote hESC lines), 5 hiPSC lines and 58 non-pluripotent samples (19 blood and tissue samples and 39 primary cell lines). Although this study was not intended to be comprehensive, we included hESC lines derived in a variety of geographical locations in order to sample the spectrum of ethnic diversity present in hESC lines in general (Supplementary Table 1). Using genome-wide SNP genotyping and Bayesian analysis of population structure (BAPS; see Supplementary Methods), we determined that the ethnic origins of the hESC lines included in this study were quite restricted (Fig. 1, Supplementary Table 1). The large majority of hESC lines (43 of 47) were of European and East Asian ethnicity. The diversity of the hiPSC lines and nonpluripotent cells is also shown (Fig. 1, Supplementary Fig. 1). We note that the location of derivation of the hESC lines is not necessarily indicative of their ethnic origin. For instance, four of the five hESC lines described in one landmark publication 4 were derived in the United States (Wisconsin) but reportedly 5 came from blastocysts transported from Israel by collaborators. Our results on three of these lines indicate that they show a genetic profile similar to those of people from the region around Adygea (WA01), Tuscany (WA09) and Tuscany/Palestine (WA07), with indication of correspondence

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Reprogramming roadmap reveals route to human induced trophoblast stem cells

Nature

An integrated analysis of human myeloid cells identifies gaps in in vitro models of in vivo biology

Stem Cell Reports

The application of cell surface markers to demarcate distinct human pluripotent states

Experimental Cell Research

Tailored emulsion-templated porous polymer scaffolds for iPSC-derived human neural precursor cell culture

Polymer Chemistry

The work here describes the synthesis of tailor-made, porous, polymeric materials with elastic mo... more

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Single-Cell Profiling Identifies Key Pathways Expressed by iPSCs Cultured in Different Commercial Media

iScience, Jan 28, 2018

We assessed the pluripotency of human induced pluripotent stem cells (iPSCs) maintained on an aut... more We assessed the pluripotency of human induced pluripotent stem cells (iPSCs) maintained on an automated platform using StemFlex and TeSR-E8 media. Analysis of transcriptome of single cells revealed similar expression of core pluripotency genes, as well as genes associated with naive and primed states of pluripotency. Analysis of individual cells from four samples consisting of two different iPSC lines each grown in the two culture media revealed a shared subpopulation structure with three main subpopulations different in pluripotency states. By implementing a machine learning approach, we estimated that most cells within each subpopulation are very similar between all four samples. The single-cell RNA sequencing analysis of iPSC lines grown in both media reports the molecular signature in StemFlex medium and how it compares to that observed in the TeSR-E8 medium.

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Physiological oxygen culture reveals retention of metabolic memory in human induced pluripotent stem cells

PloS one, 2018

Reprogramming somatic cells to a pluripotent cell state (induced Pluripotent Stem (iPS) cells) re... more Reprogramming somatic cells to a pluripotent cell state (induced Pluripotent Stem (iPS) cells) requires reprogramming of metabolism to support cell proliferation and pluripotency, most notably changes in carbohydrate turnover that reflect a shift from oxidative to glycolytic metabolism. Some aspects of iPS cell metabolism differ from embryonic stem (ES) cells, which may reflect a parental cell memory, or be a consequence of the reprogramming process. In this study, we compared the metabolism of 3 human iPS cell lines to assess the fidelity of metabolic reprogramming. When challenged with reduced oxygen concentration, ES cells have been shown to modulate carbohydrate use in a predictably way. In the same model, 2 of 3 iPS cell lines failed to regulate carbohydrate metabolism. Oxygen is a well-characterized regulator of cell function and embryo viability, and an inability of iPS cells to modulate metabolism in response to oxygen may indicate poor metabolic fidelity. As metabolism is l...

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Comprehensive characterization of distinct states of human naive pluripotency generated by reprogramming

Nature Methods

Recent reports on the characteristics of naive human pluripotent stem cells (hPSCs) obtained usin... more Recent reports on the characteristics of naive human pluripotent stem cells (hPSCs) obtained using independent methods differ. Naive hPSCs have been mainly derived by conversion from primed hPSCs or by direct derivation from human embryos rather than by somatic cell reprogramming. To provide an unbiased molecular and functional reference, we derived genetically matched naive hPSCs by direct reprogramming of fibroblasts and by primed-to-naive conversion using different naive conditions (NHSM, RSeT, 5iLAF and t2iLGöY). Our results show that hPSCs obtained in these different conditions display a spectrum of naive characteristics. Furthermore, our characterization identifies KLF4 as sufficient for conversion of primed hPSCs into naive t2iLGöY hPSCs, underscoring the role that reprogramming factors can play for the derivation of bona fide naive hPSCs.

Scaffolds for 3D in vitro culture of neural lineage cells

Acta biomaterialia, May 1, 2017

Understanding how neurodegenerative disorders develop is not only a key challenge for researchers... more Understanding how neurodegenerative disorders develop is not only a key challenge for researchers but also for the wider society, given the rapidly aging populations in developed countries. Advances in this field require new tools with which to recreate neural tissue in vitro and produce realistic disease models. This in turn requires robust and reliable systems for performing 3D in vitro culture of neural lineage cells. This review provides a state of the art update on three-dimensional culture systems for in vitro development of neural tissue, employing a wide range of scaffold types including hydrogels, solid porous polymers, fibrous materials and decellularised tissues as well as microfluidic devices and lab-on-a-chip systems. To provide some context with in vivo development of the central nervous system (CNS), we also provide a brief overview of the neural stem cell niche, neural development and neural differentiation in vitro. We conclude with a discussion of future directions...

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New Monoclonal Antibodies to Defined Cell Surface Proteins on Human Pluripotent Stem Cells

Stem cells (Dayton, Ohio), Mar 23, 2016

The study and application of human pluripotent stem cells (hPSCs) will be enhanced by the availab... more The study and application of human pluripotent stem cells (hPSCs) will be enhanced by the availability of well-characterized monoclonal antibodies (mAbs) detecting cell-surface epitopes. Here, we report generation of seven new mAbs that detect cell surface proteins present on live and fixed human ES cells (hESCs) and human iPS cells (hiPSCs), confirming our previous prediction that these proteins were present on the cell surface of hPSCs. The mAbs all show a high correlation with POU5F1 (OCT4) expression and other hPSC surface markers (TRA-160 and SSEA-4) in hPSC cultures and detect rare OCT4 positive cells in differentiated cell cultures. These mAbs are immunoreactive to cell surface protein epitopes on both primed and naive state hPSCs, providing useful research tools to investigate the cellular mechanisms underlying human pluripotency and states of cellular reprogramming. In addition, we report that subsets of the seven new mAbs are also immunoreactive to human bone marrow-derive...

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Dynamic Changes in the Copy Number of Pluripotency and Cell Proliferation Genes in Human ESCs and iPSCs during Reprogramming and Time in Culture

by Jeanne Loring and Andrew Laslett

Cell Stem Cell, 2011

Genomic stability is critical for the clinical use of human embryonic and induced pluripotent ste... more Genomic stability is critical for the clinical use of human embryonic and induced pluripotent stem cells. We performed high resolution SNP (single nucleotide polymorphism) analysis on 186 pluripotent and 119 non-pluripotent samples. We report a higher frequency of subchromosomal copy number variations in pluripotent samples compared to non-pluripotent samples, with variations enriched in specific genomic regions. The distribution of these variations differed between hESCs and hiPSCs, characterized by large numbers of duplications found in a few hESC samples and moderate numbers of deletions distributed across many hiPSC samples. For hiPSCs, the reprogramming process was associated with deletions of tumor suppressor genes, while time in culture was associated with duplications of oncogenic genes. We also observed duplications that arose during a differentiation protocol. Our results illustrate the dynamic nature of genomic abnormalities in pluripotent stem cells and the need for frequent genomic monitoring to assure phenotypic stability and clinical safety.

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Flow Cytometric Analysis of Human Pluripotent Stem Cells

by Andrew Fryga and Andrew Laslett

Methods in Molecular Biology, 2011

Human pluripotent stem cells, human embryonic stem cells and induced pluripotent stem cells, repr... more Human pluripotent stem cells, human embryonic stem cells and induced pluripotent stem cells, represent an exciting new era in regenerative medicine and drug discovery. However, prior to their clinical translation, there is a need to gain an in-depth understanding of human pluripotent stem cell biology by characterizing these potentially heterogeneous populations of cells. Flow cytometry provides a rapid and efficient approach with which to isolate, purify, and study the functional properties of defined pluripotent stem cell types.

Flow Cytometric Analysis of Human Embryonic Stem Cells

by Andrew Fryga and Andrew Laslett

Human Stem Cell Manual, 2007

Gonocyte-Sertoli cell interactions during development of the neonatal rodent testis

Current topics in developmental biology, 2000

During neonatal testicular development in the rat, events critical for subsequent germ cell devel... more During neonatal testicular development in the rat, events critical for subsequent germ cell development occur that set the stage for fertility later in life. Some gonocytes resume mitotic activity and/or migrate to the surrounding basal lamina, and use of a carefully defined Sertoli cell-gonocyte coculture system indicates that these crucial events occur without added factors or hormones and are hence likely to depend on interaction with adjacent Sertoli cells. Coupling of the Kit receptor protein on gonocytes to stem cell factor from Sertoli cells is vital for successful migration by gonocytes, as antagonism of the former suppresses and addition of the latter stimulates gonocyte migration. During the neonatal period, intercellular adhesion is modified in a developmental manner such that neural cell adhesion molecule (NCAM) is the main adhesive molecule expressed and functioning at birth, with a progressive decline as development proceeds. This decline in NCAM expression is supporte...

Developmental Differences in Heparan Sulfate Proteoglycan-Mediated Fibroblast Growth Factor-2 Actions on Leydig Cell Steroidogenesis

DEVELOPMENTAL DIFFERENCES IN HEPARAN-SULFATE PROTEOGLYCAN MEDIATED FGF-2 ACTIONS ON LEYDIG-CELL STEROIDOGENESIS

5 Gonocyte-Sertoli Cell Interactions during Development of the Neonatal Rodent Testis-II. The Sertoli Cell-Gonocyte Coculture Model

5 Gonocyte-Sertoli Cell Interactions during Development of the Neonatal Rodent Testis-References

Effects of T-3 on diverse biological processes in neonatal stem cell-Sertoli cell cocultures: A gene expression profile

REQUIREMENT FOR HEPARAN-SULFATE PROTEOGLYCANS TO MEDIATE BASIC FIBROBLAST GROWTH-FACTOR (FGF-2)-INDUCED STIMULATION OF LEYDIG-CELL STEROIDOGENESIS

This study reports that, in contrast to previous findings, basic fibroblast growth factor (FGF-2)... more This study reports that, in contrast to previous findings, basic fibroblast growth factor (FGF-2) stimulates immature Leydig cell steroidogenesis in the absence of luteinizing hormone (LH). Heparan sulphate proteoglycans (HSPGs) are essential for this action of FGF-2 and the data suggest that HSPG/FGF-2 interactions have a significant role in the maintenance of immature Leydig cell steroidogenesis. Culture conditions were established

Restricted ethnic diversity in human embryonic stem cell lines

by Eyitayo S Fakunle and Andrew Laslett

Nature Methods, 2010

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Reprogramming roadmap reveals route to human induced trophoblast stem cells

Nature

An integrated analysis of human myeloid cells identifies gaps in in vitro models of in vivo biology

Stem Cell Reports

The application of cell surface markers to demarcate distinct human pluripotent states

Experimental Cell Research

Tailored emulsion-templated porous polymer scaffolds for iPSC-derived human neural precursor cell culture

Polymer Chemistry

The work here describes the synthesis of tailor-made, porous, polymeric materials with elastic mo... more

Download

Single-Cell Profiling Identifies Key Pathways Expressed by iPSCs Cultured in Different Commercial Media

iScience, Jan 28, 2018

Download

Physiological oxygen culture reveals retention of metabolic memory in human induced pluripotent stem cells

PloS one, 2018

Download

Comprehensive characterization of distinct states of human naive pluripotency generated by reprogramming

Nature Methods

Scaffolds for 3D in vitro culture of neural lineage cells

Acta biomaterialia, May 1, 2017

Download

New Monoclonal Antibodies to Defined Cell Surface Proteins on Human Pluripotent Stem Cells

Stem cells (Dayton, Ohio), Mar 23, 2016

Download

Dynamic Changes in the Copy Number of Pluripotency and Cell Proliferation Genes in Human ESCs and iPSCs during Reprogramming and Time in Culture

by Jeanne Loring and Andrew Laslett

Cell Stem Cell, 2011

Download

Flow Cytometric Analysis of Human Pluripotent Stem Cells

by Andrew Fryga and Andrew Laslett

Methods in Molecular Biology, 2011

Flow Cytometric Analysis of Human Embryonic Stem Cells

by Andrew Fryga and Andrew Laslett

Human Stem Cell Manual, 2007

Gonocyte-Sertoli cell interactions during development of the neonatal rodent testis

Current topics in developmental biology, 2000

Developmental Differences in Heparan Sulfate Proteoglycan-Mediated Fibroblast Growth Factor-2 Actions on Leydig Cell Steroidogenesis

DEVELOPMENTAL DIFFERENCES IN HEPARAN-SULFATE PROTEOGLYCAN MEDIATED FGF-2 ACTIONS ON LEYDIG-CELL STEROIDOGENESIS

5 Gonocyte-Sertoli Cell Interactions during Development of the Neonatal Rodent Testis-II. The Sertoli Cell-Gonocyte Coculture Model

5 Gonocyte-Sertoli Cell Interactions during Development of the Neonatal Rodent Testis-References

Effects of T-3 on diverse biological processes in neonatal stem cell-Sertoli cell cocultures: A gene expression profile

REQUIREMENT FOR HEPARAN-SULFATE PROTEOGLYCANS TO MEDIATE BASIC FIBROBLAST GROWTH-FACTOR (FGF-2)-INDUCED STIMULATION OF LEYDIG-CELL STEROIDOGENESIS

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