Improved neural differentiation of normal and abnormal induced pluripotent stem cell lines in the presence of valproic acid

Fernandes, Sophia; Vinnakota, Rajesh; Kumar, Janesh; Kale, Vaijayanti; Limaye, Lalita

doi:10.1002/term.2904

Cited by 4 publications

(4 citation statements)

References 42 publications

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“…VPA has been used to promote the differentiation of human stem cells into different types of somatic cells. It improved neural differentiation of human iPSCs [23], ESCs [24], mesenchymal stem cells [25] and adipose-derived stem cells [26], also hepatic and cardiomyocyte differentiation of human stem cells [27,28]. We showed that VPA robustly elevated the SSCLC induction efficiency in the presence of GDNF and b-FGF, and the combination of low concentrations of VPA and VC achieved the highest SSCLC induction efficiency.…”

Section: Introductionmentioning

confidence: 80%

Valproic acid promotes the in vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells

Wang

et al. 2021

Stem Cell Res Ther

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Background Studying human germ cell development and male infertility is heavily relied on mouse models. In vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells (SSCLCs) can be used as a model to study human germ cells and infertility. The current study aimed to develop the SSCLC induction protocol and assess the effects of the developed protocol on SSCLC induction. Methods We examined the effects of valproic acid (VPA), vitamin C (VC) and the combination of VPA and VC on the SSCLC induction efficiency and determined the expression of spermatogonial genes of differentiated cells. Haploid cells and cells expressed meiotic genes were also detected. RNA-seq analysis was performed to compare the transcriptome between cells at 0 and 12 days of differentiation and differently expressed genes were confirmed by RT-qPCR. We further evaluated the alteration in histone marks (H3K9ac and H3K27me3) at 12 days of differentiation. Moreover, the SSCLC induction efficiency of two hiPSC lines of non-obstructive azoospermia (NOA) patients was assessed using different induction protocols. Results The combination of low concentrations of VPA and VC in the induction medium was most effective to induce SSCLCs expressing several spermatogonial genes from human pluripotent stem cells at 12 days of differentiation. The high concentration of VPA was more effective to induce cells expressing meiotic genes and haploid cells. RNA-seq analysis revealed that the induction of SSCLC involved the upregulated genes in Wnt signaling pathway, and cells at 12 days of differentiation showed increased H3K9ac and decreased H3K27me3. Additionally, two hiPSC lines of NOA patients showed low SSCLC induction efficiency and decreased expression of genes in Wnt signaling pathway. Conclusions VPA robustly promoted the differentiation of human pluripotent stem cells into SSCLCs, which involved the upregulated genes in Wnt signaling pathway and epigenetic changes. hiPSCs from NOA patients showed decreased SSCLC induction efficiency and Wnt signaling pathway gene expression, suggesting that SSC depletion in azoospermia testes might be associated with inactivation of Wnt signaling pathway. Our developed SSCLC induction protocol provides a reliable tool and model to study human germ cell development and male infertility.

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Section: Introductionmentioning

confidence: 80%

Valproic acid promotes the in vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells

Wang

et al. 2021

Stem Cell Res Ther

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“…VPA was evaluated in this protocol for several reasons. The effect of VPA has been assessed in various studies of multipotent and pluripotent cells towards neuronal differentiation, however the biological mechanism and pathways have not been identified (Duan et al, 2019; Fernandes et al 2019). It has also been documented that VPA, as a histone deacetylase inhibitor, may create a neuroprotective niche (Dekker et al, 2021).…”

Section: Discussionmentioning

confidence: 99%

Robust and reproducible neuronal differentiation of human embryonic stem cells for neurotoxicology

Samara

Falck

Spildrejorde

et al. 2022

Preprint

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Neuronal differentiation from pluripotent cells is commonly used to recapitulate events in early brain development. Neuronal precursors and developing neurons cultured in vivo, rely on attachment, density and cell-to-cell communication, and cell-passaging steps should be minimised to avert disruption of network cross-connectivity. This is crucial both for viability and maturation, and the former also applies to human embryonic stem cells (hESCs). Neuronal differentiation protocols from hESCs for neurotoxicology assessment studies should therefore provide standardized cell density requirements, and optimised coating matrix conditions. The effect of drug treatments may be masked by compound instability and degradation in cell culture medium but performing daily media changes can circumvent these issues. Here, we describe a robust neuronal differentiation protocol using dual SMAD/WNT signalling inhibitors LDN193189, SB431542 and XAV939 (LSX) for neural induction of hESCs, followed by self-patterning and cell maturation stages, for the generation of ventral telencephalic progenitors and neurons. We provide critical information on the optimized cell culture parameters and standardized methods of stage-specific validation. Using cell counts, immunofluorescence, qRT-PCR, and a proof of principle treatment with valproic acid to showcase issues with drug-induced cell toxicity, we facilitate reproducibility of the protocol.

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“…Differentiation toward the astrocytic lineage is decreased under these conditions, which is usually observed during differentiation to the neuronal lineage. Length and connections established by neurites are also significantly increased by VPA addition [115]. Besides, the use of retinoic acid (NaB) increases the acetylation of H3K9 at the PAX6 promoter, which promotes the induction of neural progenitor cells [116].…”

Section: Role Of Histone Acetylation and Deacetylationmentioning

confidence: 99%

Chromatin Structure and Dynamics: Focus on Neuronal Differentiation and Pathological Implication

Nothof

Magdinier

Van-Gils

2022

Genes

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Chromatin structure is an essential regulator of gene expression. Its state of compaction contributes to the regulation of genetic programs, in particular during differentiation. Epigenetic processes, which include post-translational modifications of histones, DNA methylation and implication of non-coding RNA, are powerful regulators of gene expression. Neurogenesis and neuronal differentiation are spatio-temporally regulated events that allow the formation of the central nervous system components. Here, we review the chromatin structure and post-translational histone modifications associated with neuronal differentiation. Studying the impact of histone modifications on neuronal differentiation improves our understanding of the pathophysiological mechanisms of chromatinopathies and opens up new therapeutic avenues. In addition, we will discuss techniques for the analysis of histone modifications on a genome-wide scale and the pathologies associated with the dysregulation of the epigenetic machinery.

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Improved neural differentiation of normal and abnormal induced pluripotent stem cell lines in the presence of valproic acid

Cited by 4 publications

References 42 publications

Valproic acid promotes the in vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells

Valproic acid promotes the in vitro differentiation of human pluripotent stem cells into spermatogonial stem cell-like cells

Robust and reproducible neuronal differentiation of human embryonic stem cells for neurotoxicology

Chromatin Structure and Dynamics: Focus on Neuronal Differentiation and Pathological Implication

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