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Transcriptional signatures in iPSC-derived neurons are reproducible across labs when differentiation protocols are closely matched.

ABSTRACT: Reproducibility of expression patterns in iPSC-derived cells from different labs is an important first step in ensuring replication of biochemical or functional assays that are performed in different labs. Here we show that reproducible gene expression patterns from iPSCs and iPSC-derived neurons matured and collected at two separate laboratory locations can be achieved by closely matching protocols and reagents. While there are significant differences in gene expression between iPSCs and differentiated neurons, as well as between different donor lines of the same cell type, transcriptional changes that vary with laboratory sites are relatively small. These results suggest that making great efforts to match protocols, reagents and technical methods between labs may improve the reproducibility of iPSC-derived cell models.

SUBMITTER: Reed X

PROVIDER: S-EPMC8655646 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Transcriptional signatures in iPSC-derived neurons are reproducible across labs when differentiation protocols are closely matched

Project description:Reproducibility of expression patterns in iPSC-derived cells from different labs is an important first step in ensuring replication of biochemical or functional assays that are performed in different labs. Here we show that reproducible gene expression patterns from iPSCs and iPSC-derived neurons matured and collected at two separate laboratory locations can be achieved by closely matching protocols and reagents. While there are significant differences in gene expression between iPSCs and differentiated neurons, as well as between different donor lines of the same cell type, transcriptional changes that vary with laboratory sites are relatively small. These results suggest that making great efforts to match protocols, reagents and technical methods between labs may improve the reproducibility of iPSC-derived cell models.

2021-12-01 | GSE167051 | GEO

Transcriptional signatures in iPSC-derived neurons are reproducible across labs when differentiation protocols are closely matched

Project description:Transcriptional signatures in iPSC-derived neurons are reproducible across labs when differentiation protocols are closely matched

| PRJNA702793 | ENA

The Alzheimer Disease-Causing Presenilin-1 L435F Mutation Causes Increased Production of Soluble Aβ43 Species in Patient-Derived iPSC-Neurons, Closely Mimicking Matched Patient Brain Tissue.

Project description:Familial Alzheimer disease-causing mutations in Presenilin 1 (PSEN1) are generally thought to shift the processing of APP toward longer, more amyloidogenic Aβ fragments. However, certain PSEN1 mutations cause severe reduction in gamma secretase function when expressed in the homozygous state, thus challenging the amyloid hypothesis. We sought to evaluate the effects of one such mutation, PSEN1 L435F, in more physiologic conditions and genetic contexts by using human induced pluripotent stem cell (iPSC)-derived neurons from an individual with familial AD (fAD) linked to the PSEN1 L435F mutation, and compared the biochemical phenotype of the iPS-derived neurons with brain tissue obtained at autopsy from the same patient. Our results demonstrate that in the endogenous heterozygous state, the PSEN1 L435F mutation causes a large increase in soluble Aβ43 but does not change the overall levels of soluble Aβ40 or Aβ42 when compared with control iPSC-neurons. Increased pathologically phosphorylated tau species were also observed in PSEN1-mutant iPSC-neurons. Concordant changes in Aβ species were present in autopsy brain tissue from the same patient. Finally, the feasibility of using Aβ43 immunohistochemistry of brain tissue to identify fAD cases was evaluated in a limited autopsy case series with the finding that strong Aβ43 staining occurred only in fAD cases.

| S-EPMC7241938 | biostudies-literature

Modeling anorexia nervosa: transcriptional insights from human iPSC-derived neurons.

Project description:Anorexia nervosa (AN) is a complex and multifactorial disorder occurring predominantly in women. Despite having the highest mortality among psychiatric conditions, it still lacks robust and effective treatment. Disorders such as AN are most likely syndromes with multiple genetic contributions, however, genome-wide studies have been underpowered to reveal associations with this uncommon illness. Here, we generated induced pluripotent stem cells (iPSCs) from adolescent females with AN and unaffected controls. These iPSCs were differentiated into neural cultures and subjected to extensive transcriptome analysis. Within a small cohort of patients who presented for treatment, we identified a novel gene that appears to contribute to AN pathophysiology, TACR1 (tachykinin 1 receptor). The participation of tachykinins in a variety of biological processes and their interactions with other neurotransmitters suggest novel mechanisms for how a disrupted tachykinin system might contribute to AN symptoms. Although TACR1 has been associated with psychiatric conditions, especially anxiety disorders, we believe this report is its first association with AN. Moreover, our human iPSC approach is a proof-of-concept that AN can be modeled in vitro with a full human genetic complement, and represents a new tool for understanding the elusive molecular and cellular mechanisms underlying the disease.

| S-EPMC5416680 | biostudies-literature

Somatic mutations alter the differentiation outcomes of iPSC-derived neurons.

Project description:The use of induced pluripotent stem cells (iPSC) as models for development and human disease has enabled the study of otherwise inaccessible tissues. A remaining challenge in developing reliable models is our limited understanding of the factors driving irregular differentiation of iPSCs, particularly the impact of acquired somatic mutations. We leveraged data from a pooled dopaminergic neuron differentiation experiment of 238 iPSC lines profiled with single-cell RNA and whole-exome sequencing to study how somatic mutations affect differentiation outcomes. We found that deleterious somatic mutations in key developmental genes, notably the BCOR gene, are strongly associated with failure in dopaminergic neuron differentiation and a larger proliferation rate in culture. We further identified broad differences in cell type composition between incorrectly and successfully differentiating lines, as well as significant changes in gene expression contributing to the inhibition of neurogenesis. Our work calls for caution in interpreting differentiation-related phenotypes in disease-modeling experiments.

| S-EPMC10112289 | biostudies-literature

Improved cell therapy protocols for Parkinson's disease based on differentiation efficiency and safety of hESC-, hiPSC-, and non-human primate iPSC-derived dopaminergic neurons.

Project description:The main motor symptoms of Parkinson's disease are due to the loss of dopaminergic (DA) neurons in the ventral midbrain (VM). For the future treatment of Parkinson's disease with cell transplantation it is important to develop efficient differentiation methods for production of human iPSCs and hESCs-derived midbrain-type DA neurons. Here we describe an efficient differentiation and sorting strategy for DA neurons from both human ES/iPS cells and non-human primate iPSCs. The use of non-human primate iPSCs for neuronal differentiation and autologous transplantation is important for preclinical evaluation of safety and efficacy of stem cell-derived DA neurons. The aim of this study was to improve the safety of human- and non-human primate iPSC (PiPSC)-derived DA neurons. According to our results, NCAM(+) /CD29(low) sorting enriched VM DA neurons from pluripotent stem cell-derived neural cell populations. NCAM(+) /CD29(low) DA neurons were positive for FOXA2/TH and EN1/TH and this cell population had increased expression levels of FOXA2, LMX1A, TH, GIRK2, PITX3, EN1, NURR1 mRNA compared to unsorted neural cell populations. PiPSC-derived NCAM(+) /CD29(low) DA neurons were able to restore motor function of 6-hydroxydopamine (6-OHDA) lesioned rats 16 weeks after transplantation. The transplanted sorted cells also integrated in the rodent brain tissue, with robust TH+/hNCAM+ neuritic innervation of the host striatum. One year after autologous transplantation, the primate iPSC-derived neural cells survived in the striatum of one primate without any immunosuppression. These neural cell grafts contained FOXA2/TH-positive neurons in the graft site. This is an important proof of concept for the feasibility and safety of iPSC-derived cell transplantation therapies in the future.

| S-EPMC3775937 | biostudies-literature

Super-Obese Patient-Derived iPSC Hypothalamic Neurons Exhibit Obesogenic Signatures and Hormone Responses.

Project description:The hypothalamus contains neurons that integrate hunger and satiety endocrine signals from the periphery and are implicated in the pathophysiology of obesity. The limited availability of human hypothalamic neurons hampers our understanding of obesity disease mechanisms. To address this, we generated human induced pluripotent stem cells (hiPSCs) from multiple normal body mass index (BMI; BMI ≤ 25) subjects and super-obese (OBS) donors (BMI ≥ 50) with polygenic coding variants in obesity-associated genes. We developed a method to reliably differentiate hiPSCs into hypothalamic-like neurons (iHTNs) capable of secreting orexigenic and anorexigenic neuropeptides. Transcriptomic profiling revealed that, although iHTNs maintain a fetal identity, they respond appropriately to metabolic hormones ghrelin and leptin. Notably, OBS iHTNs retained disease signatures and phenotypes of high BMI, exhibiting dysregulated respiratory function, ghrelin-leptin signaling, axonal guidance, glutamate receptors, and endoplasmic reticulum (ER) stress pathways. Thus, human iHTNs provide a powerful platform to study obesity and gene-environment interactions.

| S-EPMC6398951 | biostudies-literature

Recapitulation of Human Neural Microenvironment Signatures in iPSC-Derived NPC 3D Differentiation.

Project description:Brain microenvironment plays an important role in neurodevelopment and pathology, where the extracellular matrix (ECM) and soluble factors modulate multiple cellular processes. Neural cell culture typically relies on heterologous matrices poorly resembling brain ECM. Here, we employed neurospheroids to address microenvironment remodeling during neural differentiation of human stem cells, without the confounding effects of exogenous matrices. Proteome and transcriptome dynamics revealed significant changes at cell membrane and ECM during 3D differentiation, diverging significantly from the 2D differentiation. Structural proteoglycans typical of brain ECM were enriched during 3D differentiation, in contrast to basement membrane constituents in 2D. Moreover, higher expression of synaptic and ion transport machinery was observed in 3D cultures, suggesting higher neuronal maturation in neurospheroids. This work demonstrates that 3D neural differentiation as neurospheroids promotes the expression of cellular and extracellular features found in neural tissue, highlighting its value to address molecular defects in cell-ECM interactions associated with neurological disorders.

| S-EPMC6094163 | biostudies-literature

A survey of protocols from 54 infant and toddler neuroimaging research labs.

Project description:Infant and toddler MRI enables unprecedented insight into the developing brain. However, consensus about optimal data collection practices is lacking, which slows growth of the field and impedes replication efforts. The goal of this study was to collect systematic data across a large number of infant/toddler research laboratories to better understand preferred practices. Survey data addressed MRI acquisition strategies, scan success rates, visit preparations, scanning protocols, accommodations for families, study design, and policies regarding incidental findings. Respondents had on average 8 years' experience in early life neuroimaging and represented more than fifty research laboratories. Areas of consensus across labs included higher success rates among newborns compared to older infants or toddlers, high rates of data loss across age groups, endorsement of multiple layers of hearing protection, and age-specific scan preparation and participant accommodation. Researchers remain divided on decisions in longitudinal study design and practices regarding incidental findings. This study summarizes practices honed over years of work by a large collection of scientists, which may serve as an important resource for those new to the field. The ability to reference data about best practices facilitates future harmonization, data sharing, and reproducibility, all of which advance this important frontier in developmental science.

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A review of protocols for human iPSC culture, cardiac differentiation, subtype-specification, maturation, and direct reprogramming.

Project description:The methods for the culture and cardiomyocyte differentiation of human embryonic stem cells, and later human induced pluripotent stem cells (hiPSC), have moved from a complex and uncontrolled systems to simplified and relatively robust protocols, using the knowledge and cues gathered at each step. HiPSC-derived cardiomyocytes have proven to be a useful tool in human disease modelling, drug discovery, developmental biology, and regenerative medicine. In this protocol review, we will highlight the evolution of protocols associated with hPSC culture, cardiomyocyte differentiation, sub-type specification, and cardiomyocyte maturation. We also discuss protocols for somatic cell direct reprogramming to cardiomyocyte-like cells.

| S-EPMC9405110 | biostudies-literature

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