Comprehensive proteogenomic analysis of human embryonic and induced pluripotent stem cells

Scalise

et al. 2021

Cells

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Embryonic stem cells (ESCs) are pluripotent cells with indefinite self-renewal ability and differentiation properties. To function properly and maintain genomic stability, ESCs need to be endowed with an efficient repair system as well as effective redox homeostasis. In this study, we investigated different aspects involved in ESCs’ response to iron accumulation following stable knockdown of the ferritin heavy chain (FTH1) gene, which encodes for a major iron storage protein with ferroxidase activity. Experimental findings highlight unexpected and, to a certain extent, paradoxical results. If on one hand FTH1 silencing does not correlate with increased ROS production nor with changes in the redox status, strengthening the concept that hESCs are extremely resistant and, to a certain extent, even refractory to intracellular iron imbalance, on the other, the differentiation potential of hESCs seems to be affected and apoptosis is observed. Interestingly, we found that FTH1 silencing is accompanied by a significant activation of the nuclear factor (erythroid-derived-2)-like 2 (Nrf2) signaling pathway and pentose phosphate pathway (PPP), which crosstalk in driving hESCs antioxidant cascade events. These findings shed new light on how hESCs perform under oxidative stress, dissecting the molecular mechanisms through which Nrf2, in combination with PPP, counteracts oxidative injury triggered by FTH1 knockdown.

Section: Discussionmentioning

confidence: 99%

Uncovering the Metabolic and Stress Responses of Human Embryonic Stem Cells to FTH1 Gene Silencing

Scalise

et al. 2021

Cells

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“…The detailed description of iPSCs generation from ULD1 and ULD2 (lines UNIMGi003-A and UNIMGi004-A , respectively) is described in [ 31 ]. The generation of iPSCs from the healthy subject used as control in all our experiments is described in [ 32 ] (see hiPSCs-3 line). Briefly, we generated control iPS cell line through genetic reprogramming of skin fibroblasts of a healthy Caucasian male, having the same age of our ULD patients.…”

Section: Methodsmentioning

confidence: 99%

Insights into the Genetic Profile of Two Siblings Affected by Unverricht-Lundborg Disease Using Patient-Derived hiPSCs

Scalise

et al. 2022

Cells

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Unverricht-Lundborg disease (ULD), also known as progressive myoclonic epilepsy 1 (EPM1), is a rare autosomal recessive neurodegenerative disorder characterized by a complex symptomatology that includes action- and stimulus-sensitive myoclonus and tonic-clonic seizures. The main cause of the onset and development of ULD is a repeat expansion of a dodecamer sequence localized in the promoter region of the gene encoding cystatin B (CSTB), an inhibitor of lysosomal proteases. Although this is the predominant mutation found in most patients, the physio-pathological mechanisms underlying the disease complexity remain largely unknown. In this work, we used patient-specific iPSCs and their neuronal derivatives to gain insight into the molecular and genetic machinery responsible for the disease in two Italian siblings affected by different phenotypes of ULD. Specifically, fragment length analysis on amplified CSTB promoters found homozygous status for dodecamer expansion in both patients and showed that the number of dodecamer repeats is the same in both. Furthermore, the luciferase reporter assay showed that the CSTB promoter activity was similarly reduced in both lines compared to the control. This information allowed us to draw important conclusions: (1) the phenotypic differences of the patients do not seem to be strictly dependent on the genetic mutation around the CSTB gene, and (2) that some other molecular mechanisms, not yet clearly identified, might be taken into account. In line with the inhibitory role of cystatin B on cathepsins, molecular investigations performed on iPSCs-derived neurons showed an increased expression of lysosomal cathepsins (B, D, and L) and a reduced expression of CSTB protein. Intriguingly, the increase in cathepsin expression does not appear to be correlated with the residual amount of CSTB, suggesting that other mechanisms, in addition to the regulation of cathepsins, could be involved in the pathological complexity of the disease.

“…Comparative gene expression analyses of human iPSCs and ESCs have indeed revealed the presence of a small number of genes that are differentially expressed between these human pluripotent stem cell lines, suggesting that iPSCs might display significant differences of molecular profiles including genomic instability and epigenetic, non-coding and coding-RNA expression [ 16 ]. Additionally, a combined “multi-omics” study encompassing transcriptomic, proteomic, and phosphoproteomic profile analysis of reprogrammed iPSCs versus ESCs has demonstrated the presence of a small, but statistically significant, group of signaling pathways exclusively enriched in iPSCs, providing again evidence that reprogrammed cells may have a unique molecular signature, highlighting the complexity of human pluripotency [ 17 ]. Although concerns about the real equivalence between iPSCs and ESCs still exist, the effort of the scientific community to make these differences irrelevant as much as possible is undeniable.…”

Section: Human Pluripotent Stem Cells: Applications Power and LImentioning

confidence: 99%

Modeling Cardiac Disease Mechanisms Using Induced Pluripotent Stem Cell-Derived Cardiomyocytes: Progress, Promises and Challenges

Parrotta

et al. 2020

IJMS

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Cardiovascular diseases (CVDs) are a class of disorders affecting the heart or blood vessels. Despite progress in clinical research and therapy, CVDs still represent the leading cause of mortality and morbidity worldwide. The hallmarks of cardiac diseases include heart dysfunction and cardiomyocyte death, inflammation, fibrosis, scar tissue, hyperplasia, hypertrophy, and abnormal ventricular remodeling. The loss of cardiomyocytes is an irreversible process that leads to fibrosis and scar formation, which, in turn, induce heart failure with progressive and dramatic consequences. Both genetic and environmental factors pathologically contribute to the development of CVDs, but the precise causes that trigger cardiac diseases and their progression are still largely unknown. The lack of reliable human model systems for such diseases has hampered the unraveling of the underlying molecular mechanisms and cellular processes involved in heart diseases at their initial stage and during their progression. Over the past decade, significant scientific advances in the field of stem cell biology have literally revolutionized the study of human disease in vitro. Remarkably, the possibility to generate disease-relevant cell types from induced pluripotent stem cells (iPSCs) has developed into an unprecedented and powerful opportunity to achieve the long-standing ambition to investigate human diseases at a cellular level, uncovering their molecular mechanisms, and finally to translate bench discoveries into potential new therapeutic strategies. This review provides an update on previous and current research in the field of iPSC-driven cardiovascular disease modeling, with the aim of underlining the potential of stem-cell biology-based approaches in the elucidation of the pathophysiology of these life-threatening diseases.