Abstract
Nuclear integrity and mechanical stability of the nuclear envelope (NE) are conferred by the nuclear lamina, a meshwork of intermediate filaments composed of A- and B-type lamins, supporting the inner nuclear membrane and playing a pivotal role in chromatin organization and epigenetic regulation. During cell senescence, nuclear alterations also involving NE architecture are widely described. In the present study, we utilized induced pluripotent stem cells (iPSCs) upon prolonged in vitro culture as a model to study aging and investigated the organization and expression pattern of NE major constituents. Confocal and four-dimensional imaging combined with molecular analyses, showed that aged iPSCs are characterized by nuclear dysmorphisms, nucleoskeletal components (lamin A/C-prelamin isoforms, lamin B1, emerin, and nesprin-2) imbalance, leading to impaired nucleo-cytoplasmic MKL1 shuttling, actin polymerization defects, mitochondrial dysfunctions, SIRT7 downregulation and NF-kBp65 hyperactivation. The observed age-related NE features of iPSCs closely resemble those reported for premature aging syndromes (e.g., Hutchinson-Gilford progeria syndrome) and for somatic cell senescence. These findings validate the use of aged iPSCs as a suitable cellular model to study senescence and for investigating therapeutic strategies aimed to treat premature aging.
Highlights
Nuclear envelope (NE) is a bilayer membrane enclosing the nucleus contents in eukaryotic cells; it is formed by the inner (INM) and the outer (ONM) nuclear membranes, composed of numerous integral membrane proteins playing pivotal roles in chromatin organization and gene expression
In this work we deepened the knowledge of induced pluripotent stem cells (iPSCs) biology, studying the behavior of nuclear envelope (NE) constituents both in pluripotent cells and in the same cells cultured for prolonged time, in order to investigate if NE dysfunctions are induced in pluripotent stem cells by aging
We used iPSCs obtained from fibroblasts of a healthy male adult and, in particular, we used three clones for all the experiments reported in this work
Summary
Nuclear envelope (NE) is a bilayer membrane enclosing the nucleus contents in eukaryotic cells; it is formed by the inner (INM) and the outer (ONM) nuclear membranes, composed of numerous integral membrane proteins playing pivotal roles in chromatin organization and gene expression. Several human diseases have been linked to mutations affecting LMNA or genes encoding B-type lamins, LAPs and other NE proteins. Among these disorders, referred to as “laminopathies”, the most severe phenotype is found in premature aging conditions, such as Hutchinson-Gilford progeria syndrome (HGPS), a rare autosomal dominant syndrome characterized by the appearance of aging hallmarks early in childhood [9]. Normal aging is a complex biological process characterized by several dysregulated pathways, some of which contributing to premature aging in HGPS, namely mitochondrial and telomere dysfunctions, heterocromatin loss and disorganization, reactive oxygen species (ROS) accumulation and alterations of NE components [12, 13]
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