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Jeanne F Loring

Publications and source records attributed to Jeanne F Loring.

17 recordsLinked to original sources
PubMed2026

Genomic and transcriptomic quality control for an autologous iPSC-derived cell therapy for Parkinson's disease.

Toward development of an autologous, induced pluripotent stem cell (iPSC)-based cell therapy for Parkinson's disease (PD), we demonstrate successful, reproducible genomic and transcriptomic qualification of patient-derived dopaminergic neuron precursor cells (DANPCs) across multiple donors. Our analysis includes whole-genome sequencing data from fibroblasts, iPSCs, and DANPCs and the development of NeuriTest, an RNAseq-based bioinformatic analysis of DANPCs designed to predict cell quality based on empirical animal data. Autologous cell therapies are immune matched to the patient, potentially augmenting durability of benefit compared to allogeneic cells while negating the need for immunosuppression and accompanying side effects. Patient-specific iPSCs are an autologous cell source that can be differentiated to dopaminergic neurons, the cell type lost in PD. We report here our preclinical manufacturing strategy and results demonstrating efficacy in a PD rodent model and safety in a 9-month GLP toxicology study.

Parkinson’s disease↗
PubMed2025

Diversifying the reference iPSC line concept.

We discuss how to diversify the reference iPSC line concept. We highlight workflows for generating diverse iPSC lines. We ask whether reference lines can act as inclusive sources of human diversity for use in benchmarking controls for disease models or drug screens, or as clinical grade lines for cell therapies.

Induced Pluripotent Stem Cells↗
PubMed2006

Human embryonic stem cells have a unique epigenetic signature.

Human embryonic stem (hES) cells originate during an embryonic period of active epigenetic remodeling. DNA methylation patterns are likely to be critical for their self-renewal and pluripotence. We compared the DNA methylation status of 1536 CpG sites (from 371 genes) in 14 independently isolated hES cell lines with five other cell types: 24 cancer cell lines, four adult stem cell populations, four lymphoblastoid cell lines, five normal human tissues, and an embryonal carcinoma cell line. We found that the DNA methylation profile clearly distinguished the hES cells from all of the other cell types. A subset of 49 CpG sites from 40 genes contributed most to the differences among cell types. Another set of 25 sites from 23 genes distinguished hES cells from normal differentiated cells and can be used as biomarkers to monitor differentiation. Our results indicate that hES cells have a unique epigenetic signature that may contribute to their developmental potential.

Cell Differentiation↗
PubMed2006

Cryopreservation by slow cooling with DMSO diminished production of Oct-4 pluripotency marker in human embryonic stem cells.

We tested a "standard" cryopreservation protocol (slow cooling with 10% DMSO) on the human embryonic stem cell (hESC) line H9 containing an Oct-4 (POU5F1) promoter-driven, enhanced green fluorescent protein (EGFP) reporter to monitor maintenance of pluripotency. Cells were cooled to -80 degrees C in cryovials and then transferred to a -80 degrees C freezer. Cells were held at -80 degrees C for 3 days ("short-term storage") or 3 months ("long-term storage"). Vials were thawed in a +36 degrees C water bath and cells were cultured for 3, 7, or 14 days. Propidium iodide (PI) was used to assess cell viability by flow cytometry. Control cells were passaged on the same day that the frozen cells were thawed. The majority of cells in control hESC cultures were Oct-4 positive and almost 99% of EGFP+ cells were alive as determined by exclusion of PI. In contrast, the frozen cells, even after 3 days of culture, contained only 50% live cells, and only 10% were EGFP-positive. After 7 days in culture, the proportion of dead cells decreased and there was an increase in the Oct-4-positive population but microscopic examination revealed large patches of EGFP-negative cells within clusters of colonies even after 14 days of culturing. After 3 months of storage at -80 degrees C the deleterious effect of freezing was even more pronounced: the samples regained a quantifiable number of EGFP-positive cells only after 7 days of culturing following thawing. It is concluded that new protocols and media are required for freezing hESC and safe storage at -80 degrees C as well as studies of the mechanisms of stress-related events associated with cell cryopreservation.

Animals↗
PubMed2006

Genome wide profiling of human embryonic stem cells (hESCs), their derivatives and embryonal carcinoma cells to develop base profiles of U.S. Federal government approved hESC lines.

BACKGROUND: In order to compare the gene expression profiles of human embryonic stem cell (hESC) lines and their differentiated progeny and to monitor feeder contaminations, we have examined gene expression in seven hESC lines and human fibroblast feeder cells using Illumina bead arrays that contain probes for 24,131 transcript probes. RESULTS: A total of 48 different samples (including duplicates) grown in multiple laboratories under different conditions were analyzed and pairwise comparisons were performed in all groups. Hierarchical clustering showed that blinded duplicates were correctly identified as the closest related samples. hESC lines clustered together irrespective of the laboratory in which they were maintained. hESCs could be readily distinguished from embryoid bodies (EB) differentiated from them and the karyotypically abnormal hESC line BG01V. The embryonal carcinoma (EC) line NTera2 is a useful model for evaluating characteristics of hESCs. Expression of subsets of individual genes was validated by comparing with published databases, MPSS (Massively Parallel Signature Sequencing) libraries, and parallel analysis by microarray and RT-PCR. CONCLUSION: we show that Illumina's bead array platform is a reliable, reproducible and robust method for developing base global profiles of cells and identifying similarities and differences in large number of samples.

Carcinoma, Embryonal↗
PubMed2006

Transcriptome coexpression map of human embryonic stem cells.

BACKGROUND: Human embryonic stem (ES) cells hold great promise for medicine and science. The transcriptome of human ES cells has been studied in detail in recent years. However, no systematic analysis has yet addressed whether gene expression in human ES cells may be regulated in chromosomal domains, and no chromosomal domains of coexpression have been identified. RESULTS: We report the first transcriptome coexpression map of the human ES cell and the earliest stage of ES differentiation, the embryoid body (EB), for the analysis of how transcriptional regulation interacts with genomic structure during ES self-renewal and differentiation. We determined the gene expression profiles from multiple ES and EB samples and identified chromosomal domains showing coexpression of adjacent genes on the genome. The coexpression domains were not random, with significant enrichment in chromosomes 8, 11, 16, 17, 19, and Y in the ES state, and 6, 11, 17, 19 and 20 in the EB state. The domains were significantly associated with Giemsa-negative bands in EB, yet showed little correlation with known cytogenetic structures in ES cells. Different patterns of coexpression were revealed by comparative transcriptome mapping between ES and EB. CONCLUSION: The findings and methods reported in this investigation advance our understanding of how genome organization affects gene expression in human ES cells and help to identify new mechanisms and pathways controlling ES self-renewal or differentiation.

Animals↗
PubMed2006

Gene therapy: can neural stem cells deliver?

Neural stem cells are a self-renewing population that generates the neurons and glia of the developing brain. They can be isolated, proliferated, genetically manipulated and differentiated in vitro and reintroduced into a developing, adult or pathologically altered CNS. Neural stem cells have been considered for use in cell replacement therapies in various neurodegenerative diseases, and an unexpected and potentially valuable characteristic of these cells has recently been revealed--they are highly migratory and seem to be attracted to areas of brain pathology such as ischaemic and neoplastic lesions. Here, we speculate on the ways in which neural stem cells might be exploited as delivery vehicles for gene therapy in the CNS.

Animals↗
PubMed2006

Adhesive interactions between human neural stem cells and inflamed human vascular endothelium are mediated by integrins.

Understanding the mechanisms by which stem cells home precisely to regions of injury or degeneration is of importance to both basic and applied regenerative medicine. Optimizing regenerative processes may depend on identifying the range of molecules that subserve stem cell trafficking. The "rolling" of extravasating cells on endothelium under conditions of physiological flow is the first essential step in the homing cascade and determines cell adhesion and transmigration. Using a laminar flow chamber to simulate physiological shear stress, we explored an aspect of this process by using human neural stem cells (hNSCs). We observed that the interactions between hNSCs and tumor necrosis factor-alpha (TNF-alpha)-stimulated human endothelium (simulating an inflamed milieu) are mediated by a subclass of integrins--alpha2, alpha6, and beta1, but not alpha4, alphav, or the chemokine-mediated pathway CXCR4-stromal cell-derived factor-1alpha--suggesting not only that the mechanisms mediating hNSC homing via the vasculature differ from the mechanisms mediating homing through parenchyma, but also that each step invokes a distinct pathway mediating a specialized function in the hNSC homing cascade. (TNF-alpha stimulation also upregulates vascular cell adhesion molecule-1 expression on the hNSCs themselves and increases NSC-endothelial interactions.) The selective use of integrin subgroups to mediate homing of cells of neuroectodermal origin may also be used to ensure that cells within the systemic circulation are delivered to the pathological region of a given organ to the exclusion of other, perhaps undesired, organs.

Cell Adhesion↗
PubMed2005

Assessing self-renewal and differentiation in human embryonic stem cell lines.

Like other cell populations, undifferentiated human embryonic stem cells (hESCs) express a characteristic set of proteins and mRNA that is unique to the cells regardless of culture conditions, number of passages, and methods of propagation. We sought to identify a small set of markers that would serve as a reliable indicator of the balance of undifferentiated and differentiated cells in hESC populations. Markers of undifferentiated cells should be rapidly downregulated as the cells differentiate to form embryoid bodies (EBs), whereas markers that are absent or low during the undifferentiated state but that are induced as hESCs differentiate could be used to assess the presence of differentiated cells in the cultures. In this paper, we describe a list of markers that reliably distinguish undifferentiated and differentiated cells. An initial list of approximately 150 genes was generated by scanning published massively parallel signature sequencing, expressed sequence tag scan, and microarray datasets. From this list, a subset of 109 genes was selected that included 55 candidate markers of undifferentiated cells, 46 markers of hESC derivatives, four germ cell markers, and four trophoblast markers. Expression of these candidate marker genes was analyzed in undifferentiated hESCs and differentiating EB populations in four different lines by immunocytochemistry, reverse transcription-polymer-ase chain reaction (RT-PCR), microarray analysis, and quantitative RT-PCR (qPCR). We show that qPCR, with as few as 12 selected genes, can reliably distinguish differentiated cells from undifferentiated hESC populations.

Animals↗
PubMed2005

Establishing standards for the characterization of human embryonic stem cell lines.

As of August 2005, 22 human embryonic stem cell (hESC) lines listed on the National Institutes of Health (NIH) hESC Registry were being distributed to investigators. At a June 2005 meeting of NIH-supported hESC researchers, we proposed that a set of shared standards should be available in order to characterize the cells unambiguously in multiple laboratories. Here, we elaborate such a plan to identify a set of standard methods and to initiate collaborative efforts to validate the standards. The standard assays we propose should be comprehensive enough to ensure that hESC banks can provide a consistent and reliable product for NIH researchers, and inexpensive enough that individual laboratories can afford to use at least some of the methods routinely in their laboratories. We expect that as data accumulate and standards evolve, a core set of tests will become the norm for routine assessment of hESC cultures and that these tests will lay the groundwork for clinical applications of these cells.

Cell Differentiation↗
PubMed2005

Evolution of microarray analysis.

Reddy's very thorough review of the history of gene expression analysis of neurodegenerative disease made me think about the early days of microarrays, so I want to reminisce a little before commenting on the evolution of the technology and some issues that I think still continue to plague us.

Alzheimer Disease↗
PubMed2005

NTera2: a model system to study dopaminergic differentiation of human embryonic stem cells.

NTera2, a human embryonal carcinoma (EC) stem cell line, shares many characteristics with human embryonic stem cells (hESCs). To determine whether NTera2 can serve as a useful surrogate for hESCs, we compared global gene expression between undifferentiated NTera2, multiple undifferentiated hESC cell lines, and their differentiated derivatives, and we showed that NTera2 cells share multiple markers with hESCs. Similar to hESCs, NTera2 cells differentiated into TH-positive cells that express dopaminergic markers including AADC, DAT, Nurr1, TrkB, TrkC, and GFRA1 when co-cultured with PA6 cells. Flow cytometry analysis showed that tyrosine hydroxylase (TH) and neural cell adhesion molecule (NCAM) expression increased, whereas SSEA4 expression decreased as cells differentiated. Medium conditioned by PA6 cells stimulated differentiation of NTera2 cells to generate TH-positive cells that expressed dopaminergic markers. Flow cytometry selected polysialylated (PSA-NCAM) cells responded to medium conditioned by PA6 cells by differentiating into TH-positive cells and expressed dopaminergic markers. Sorted cells differentiated for 4 weeks in PA6 cell conditioned media included functional neurons that responded to neurotransmitters and exhibited electronic excitability. Therefore, NTera2 cell dopaminergic neuronal differentiation and PSA-NCAM enrichment provides a useful system for the future study of hESCs.

Animals↗
PubMed2005

A role for stem cell biology in the physiological and pathological aspects of aging.

The stem cell is an essential component of a developmental phenomenon-one of the key components of a program fundamental to organogenesis and the maintenance of homeostasis throughout life. The stem cell confers plasticity to the process. Because aging is part of the developmental arc, an understanding of the stem cell is essential to understanding aging and some of the degenerative conditions that accompany it. Because stem cells may be isolated and manipulated ex vivo and then reimplanted into organs, they may serve a variety of functions. They have the capability of migrating long distances and targeting pathological conditions, of expressing therapeutic genes and responding to cues that shift their differentiation toward deficient lineages. Therefore, stem cells may be used for cell replacement, for therapeutic interventions, and potentially to modify aging.

Aging↗
PubMed2003

Selectively reduced expression of synaptic plasticity-related genes in amyloid precursor protein + presenilin-1 transgenic mice.

A critical question in Alzheimer's disease (AD) research is the cause of memory loss that leads to dementia. The amyloid precursor protein + presenilin-1 (APP+PS1) transgenic mouse is a model for amyloid deposition, and like AD, the mice develop memory deficits as amyloid deposits accumulate. We profiled gene expression in these transgenic mice by microarray and quantitative RT-PCR (qRT-PCR). At the age when these animals developed cognitive dysfunction, they had reduced mRNA expression of several genes essential for long-term potentiation and memory formation (Arc, Zif268, NR2B, GluR1, Homer-1a, Nur77/TR3). These changes appeared to be related to amyloid deposition, because mRNA expression was unchanged in the regions that did not accumulate amyloid. Transgene expression was similar in both amyloid-containing and amyloid-free regions of the brain. Interestingly, these changes occurred without apparent changes in synaptic structure, because a number of presynaptic marker mRNAs (growth-associated protein-43, synapsin, synaptophysin, synaptopodin, synaptotagmin, syntaxin) remained stable. Additionally, a number of genes related to inflammation were elevated in transgenic mice, primarily in the regions containing amyloid. In AD cortical tissue, the same memory-associated genes were downregulated. However, all synaptic and neuronal transcripts were reduced, implying that the loss of neurons and synapses contributed to these changes. We conclude that reduced expression of selected genes associated with memory consolidation are linked to memory loss in both circumstances. This suggests that the memory loss in APP+PS1 transgenic mice may model the early memory dysfunction in AD before the degeneration of synapses and neurons.

Aged↗
PubMed2002

Complexity of inflammatory responses in endothelial cells and vascular smooth muscle cells determined by microarray analysis.

To better understand the molecular basis of vascular cell system behavior in inflammation, we used gene expression microarrays to analyze the expression of 7,075 genes and their response to IL-1beta and TNFalpha in cultures of coronary artery endothelium and smooth muscle derived from a single coronary artery. The most noticeable difference between the cell types was the considerably greater magnitude and complexity of the transcriptional response in the endothelial cells. Two hundred and nine genes were regulated in the endothelium and only 39 in vascular smooth muscle. Among the 209 regulated genes in the endothelium, 99 have not been previously associated with endothelial cell activation and many implicate the endothelium in unconventional roles. For example, the induced genes include several that have only been associated with leukocyte function (e.g., IL-7 receptor, EBI-3 receptor) and others related to antiviral and antibacterial defense (e.g., oligoadenylate synthetase, LMP7, toll-like receptor 4, complement component 3). In addition, 43 genes likely to participate in signal transduction (eg. IL-18 receptor, STK2 kinase, STAF50, ANP receptor, VIP receptor, RAC3, IFP35) were regulated providing evidence that a major effect of TNFalpha and IL-1beta is to alter the potential of the endothelial cell to respond to various other external stimuli.

Cells, Cultured↗