Analysis of mechanosensing in human cardiac stem cellsElisa Mazzega
, Eliana Pomarè
, Angela Caragnano
, Sebastian Martewicz
, Nicola Elvassore
, Antonio Paolo Beltrami
, Ugo Livi
, 2015, objavljeni povzetek znanstvenega prispevka na konferenci
Opis: Objectives: We have shown that age and pathology impair the biological properties of stem cells isolated from human hearts (CSC) and functional assays showed differences between CSC isolated from normal (DCSC) and end-stage failing (ECSC) hearts. As alterations of mechanical properties of the myocardium, such as stiffening and increased wall stress, are crucial features of cardiac remodeling, this work addresses the biological effects exerted on CSC by mechanical stimuli. Materials and methods: DCSC and ECSC were cultured under deﬁned conditions to mimic speciﬁc features of the pathologic condition: increased mechanical loading (up to 15%, cyclic at 1 Hz), differential substrate stiffness (ranging from 1 to 231 kPa), differential cell densities. After 24, 48 and 72 h, cells were ﬁxed and stained for analysis of proliferation and subcellular localization of YAP or lysed for RT-PCR analysis.Results: Cyclic stretch was signiﬁcantly associated with both increased proliferation of DCSC (n = 6, p<0.0001) and ECSC (n = 4, p = 0.003), and with a signiﬁcant reduction of nuclear localized YAP (nYAP) as a function of time (p<0.05). However, while signiﬁcant correlation between cell density and decreased nYAP (p = 0.003, r2 = 0.37) characterized ECSC, this was not evident for unstretched DCSC, suggesting a less stringent regulation of contact inhibition in DCSC. These data were further conﬁrmed by seeding cells at differential density. As opposed to what previously shown for epithelial cell lines, DCSC did not reduce nYAP positivity as a function of cell density, when grown in serum containing medium, suggesting that soluble factors present in the serum could maintain the nuclear localization of YAP, independently from the cell density. In line, serum signiﬁcantly increased the nYAP expressing cells in DCSC, while a signiﬁcant positive correlation between cell density and nYAP positivity can be demonstrated in DCSC cultured in serum free medium. RT-PCR for YAP-regulated targets conﬁrmed immuno- ﬂuorescence data. Furthermore, independently from the pathologic status, cyclic stretch was signiﬁcantly associated with a persistent YAP signaling at high cell density. Besides, tension and assembly of cytoskeletal network, induced by increasing substrate stiffness, correlates with nYAP (p<0.05) and YAP transcriptional activation (p<0.05). Conclusions: D- and ECSC differ in their mechanosensing properties. However, in the ﬁrst cell type, nYAP localization is dictated by the combined action of paracrine factors and cytoskeletal tension, thus reducing the contact inhibition effect. This ﬁnding is in line with a more primitive phenotype of SC isolated from normal hearts.
Ključne besede: Stem cells, human cardiac stem cells, mechanosensing, mechanotransduction, heart failure, YAP
Objavljeno v RUNG: 21.03.2017; Ogledov: 3849; Prenosov: 0
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