Rescue of the senescence phenotype of AD MSCs by autophagy activation in 3D spheroids
Human MSCs (hMSCs) are cells capable of self-renewal and multi-lineage differentiation into various tissues of mesodermal origin. These cells can be easily isolated and expanded from the stroma of virtually all organs, although the preferred sources are bone marrow and subcutaneous fat. MSCs have been broadly applied in the treatment of various diseases, including graft-versus-host disease (GVHD), Crohn's disease (CD), diabetes mellitus (DM), multiple sclerosis (MS), myocardial infarction (MI), liver failure, and rejection after liver transplant. Upon isolation, hMSCs are characterized by their capability to develop as fibroblast colony-forming-units, and differentiate into osteocytes, chondrocytes, and adipocytes. hMSCs are positive for CD73, CD90, CD105, СD106, CD29, CD166, and negative for CD11b, CD14, CD34, CD45, HLA-DR, CD79α and CD19. Cultured primary cells do not grown infinitely, but undergo only a limited number of cell division, in a process called cellular senescence. Cell therapy protocols generally require hundreds of million hMSCs per treatment and, consequently, these cells need to be expanded in vitro for about 10 weeks before implantation. Notably, patient’s clinical history, age, and genetic makeup strongly influence the length of this expansion period and the quality of the obtained cells. Aged MSCs generally perform less well than their younger counterparts in various disease models and mounting evidence strongly suggests that cellular senescence contribute to aging and age-related diseases. It would, thus, be of great significance to monitor the occurrence of a senescent phenotype in hMSCs addressed to clinical uses and to evaluate the functional consequences of senescence in hMSCs which could affect their clinical therapeutic potential, taking into account their paracrine effects, immunomodulatory activity, differentiation potential, and cell migration ability. The function of MSCs is known to decline with age, a process that may be implicated in the loss of maintenance of tissue homeostasis leading to organ failure and diseases of aging The term senescence was applied to cells that ceased to divide in culture, based on the speculation that their behaviour recapitulated organismal ageing. Consequently, cellular senescence is sometimes termed cellular ageing or replicative senescence Telomere shortening provided the first molecular explanation for why many cells cease to divide in culture. Dysfunctional telomeres trigger senescence through the p53 pathway. This response is often termed telomere-initiated cellular senescence. Some cells undergo replicative senescence independently of telomere shortening. Resistance to apoptosis might partly explain why senescent cells are so stable in culture. This attribute might also explain why the number of senescent cells increases with age.
Changes in cell cycle inhibitors: p21Cip1 and p16InK4a. These CDKIs are components of tumour suppressor pathways that are governed by the p53 and retinoblastoma pRB proteins. New markers in Oncogene induced senescence: DEC1 (differentiated embryo chondrocyte expressed1), p15 (a CDKI) and DCR2 (decoy death receptor2). The specificity and significance of these proteins for senescent cells are not yet clear, but they are promising additional markers.
Dramatic structural changes of chromatin in senescent cells- Lamin B1. Presence of certain heterochromatin associated histone modifications (H3 lys9 methylation) and heterochromatin protein1 (Hp1)). In some cases - global heterochromatin loss, characterized by markers H3K9me3 and H3K27me3. Predominantly during OIS in vitro, heterochromatin is redistributed into 30–50 punctate DNA-dense senescence-associated heterochromatin foci (SAHF). SAHF are silent domains that co-localize with H3K9me3 and heterochromatin protein 1 (HP1) and may lock cells in a senescent state by transcriptionally repressing genes involved in cell proliferation.
MARKERS of SENESCENCE
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