Exploiting the human amniotic fluid stem cell secretome for cardiomyocyte renewal
Creators: name: COSTA, AMBRA
Publisher: name: Universit� degli studi di Genova
Year: 2022

description: Functional replacement of cardiomyocytes following injury represents a limiting aspect in
cardiac regeneration. Given the extremely low intrinsic turnover rate of adult cardiomyocytes,
their therapeutic renewal represents an unmet clinical need. Against such scenario, progenitor
cell-based paracrine therapy has gained increasing attention in terms of candidate strategy to
stimulate endogenous mechanism of repair and regeneration within the injured heart.
Previous results from the group where I have carried out my PhD demonstrated that the
secretome (namely the whole of the paracrine soluble factors that cells can release in vitro in
their conditioned medium) of a specific subpopulation of human mesenchymal stromal cells
from the amniotic fluid (hAFS: human amniotic fluid-derived stem cells) can exert significant
cardioprotective effects and stimulate resident cardiomyocytes to engage into the DNA
duplication step of the cell cycle in a preclinical adult mouse model of acute myocardial
infarction. While such results advocate the hAFS relevant paracrine potential, further detailed
studies have been necessary to better characterize the cardio-active capacity of the hAFS
secretome. This has been pursued by (i) profiling the hAFS secretome formulations, (including
the particulate fraction of their secreted extracellular vesicles - EVs) more in details, and (ii) by
means of a more responsive experimental model represented by the neonatal mouse model of
myocardial infarction.
In this PhD project attention has been driven towards the extensive investigation of the cardiac
regenerative paracrine effects exerted by two different sources of human progenitor cells: II
trimester of gestation fetal hAFS, obtained from leftover samples of amniocentesis prenatal
screening and term III trimester perinatal hAFS, isolated from clinical waste amniotic fluid from
scheduled caesarean delivery. The hAFS secretome has been considered as either the total
cell-conditioned medium (hAFS-CM), as compared to the extracellular vesicles sub-fraction
within it (hAFS-EVs).
Gestational stage, underlying the comparison of II trimester fetal hAFS versus III trimester
perinatal hAFS, did not influence either the cell phenotype or the yield of the secretome
formulations. While fetal and perinatal hAFS-EVs presented similar size distribution with a
stable miRNA core in their cargo, proteomic and cytokine/chemokine profiling of the hAFS-CM
highlighted some discrepancies between the formulations. Fetal hAFS-EVs resulted the
secretome fraction endowed with the most stimulatory paracrine potential in sustaining
cardiomyocyte cell cycle progression from S- up to M-phase with evidence of cytokinesis
events. This was further supported by decreased expression of Cofilin-2 a pivotal player in
cardiomyocyte cytoskeleton re-arrangement as required for cell-cycle re-entry. Fetal hAFS-EVs
resulted particularly enriched in Agrin -a well-known mediator of cardiac regeneration in the
neonatal mouse heart- as putative functional role mediator of the paracrine cardiogenic effects
observed in vitro. Similar findings were obtained in vivo in a 4-days-old mouse model of
myocardial infarction where fetal hAFS-EVs supported to some extent resident cardiomyocyte
S-M phase transition in the remote myocardium within 3 days from injury and acute treatment.
These encouraging results suggest that EVs obtained by immature fetal hAFS may retain
promising cardio-active paracrine potential for myocardial renewal. Nonetheless, additional
analyses are required to pinpoint the detailed molecular mechanism(s) of function in order to
optimize such effect.