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Background Cardiac ageing is associated to heart failure with preserved ejection fraction (HF-pEF) and increased senescence, hypertrophy and diastolic dysfunction. No treatments have yet proved to reduce HF-pEF morbidity and mortality. Cardiosphere-derived cells (CDCs) and their secreted extracellular vesicles (CDC-EVs) have demonstrated efficacy in old animals with cardiac dysfunction, but variability of effect and lack of adequate potency tests remain as challenges. Purpose To explore in vitro predictors of cardiac protective potency of CDC-EVs, focusing on the chronological age of the CDC-donors, CDC-senescence, and their in vitro anti-senescent and pro-angiogenic effect. Methods CDCs derived from 34 patients (age range 0–81 years old, both sexes) were characterized in terms of senescence, proliferative and migration capacities, VEGF secretion, expression of specific surface markers and cardiosphere size. CDC-EVs were purified and their in vitro anti-senescent potential (at genetic, secretory and cellular level over cardiac stromal cells) and their pro-angiogenic potential (ability to induce tube formation over endothelial cells) quantified. According to the performance in each of these tests, potency was scored and CDC-EVs were classified as potent (P-EVs) and non-potent (NP-EVs). The effect of P-EVs and NP-EVs were then tested in vivo in rats with induced cardiac aging. SD-rats received 3-months of daily intraperitoneal injections (IP) of saline (healthy control) or D-galactose. Rats in the D-Gal group were randomly allocated to receive IP saline (sham control, n=12), P-EVs (n=7) or NP-EVs (n=6) and followed-up for one month. Results Chronological age of the donor or expression of surface markers did not relate to most CDC properties nor to their in vitro potency. CDC senescence did relate to other CDC bioactive properties, but this was insufficient to predict CDC-EV anti-senescence and pro-angiogenic in vitro potency. In vivo, EV classified as P-EVs, but not NP-EVs, prevented D-gal induced hypertrophy (2.4 vs. 2.9 mg/gr, p=0.05). This finding was in parallel to the levels of galactosidase-beta 1 expression in cardiac tissue, which were increased in sham vs. P-EVs (1.22 vs. 0.73, p=0.03) but levels in NP-EVs were not significantly different to the sham group (0.86, p=0.1). P-EVs tended to reduce TGFB1 expression, while NP-EVs significantly increased cardiac fibrosis and reduced cardiac perfusion. At systemic level, while P-EVs significantly improved glucose metabolism and tended to drive total antioxidant capacity and hair growth to a healthier profile, NP-EVs did not significantly improve any of the explored parameters and significantly increased total antioxidant capacity. Conclusions After further validation, the matrix potency assay proposed here, scoring the anti-senescent and pro-angiogenic in vitro effect of CDC-EVs could be used to predict EV suitability as an allogenic product in the treatment of cardiac ageing. Funding Acknowledgement Type of funding sources: Public Institution(s). Main funding source(s): Instituto de Salud Carlos III, Ministerio de Ciencia e Innovaciόn, Spain: PI16/01123; PI19/00161; Red de Terapia Celular, Tercel, (RD16.0011.0029) and CIBERCV (CB16.11.00292)

Background Cardiac ageing is characterized by hypertrophy and diastolic dysfunction and related with heart failure with preserved ejection fraction, an unmet medical need for treatments development. Cardiosphere-derived cells (CDCs) have demonstrated glimpses of efficacy in old animals with cardiac dysfunction, however variability of effect among different CDC-donors is observed. Purpose To identify markers of in vitro potency of CDCs and to test it in an in vivo model of cardiac ageing. Methods CDCs were derived from cardiac biopsies of forty patients (age range 0 - 81 years old, 16 females) who underwent cardiac surgery for other reasons. CDC-secreted extracellular vesicles (CDC-EVs) were purified and used in in vitro and in vivo experiments. Since anti-ageing effect was targeted in vivo, in vitro potency of CDC-EVs was checked in terms of their anti-senescence bioactivity. A panel of six parameters (genetic, secretory and cellular) related with cellular senescence were analysed in vitro for all 40-donors CDC-EVs and those with most favourable and unfavourable profiles were classified as potent (P-EVs) and non-potent (NP-EVs), respectively. P-EVs and NP-EVs were then tested in vivo in 25 rats with D-galactose-induced cardiac ageing and 7-healthy controls in a randomized blinded study. Results In vitro, despite variations in the extent of the observed effect, all CDC-EVs significantly reduced cellular senescence and increased IL-6 secretion of human cardiac stromal cells. Most CDC-EVs decreased the expression of senescence-related genes (p21, p16, p53, TGF-b). Based on the observed profiles as P-EVs were selected CDC-EVs from 75- and 14-years old donors and as NP-EVs from 73-years old. In vivo, 3-months old SD-rats (30% female) after 3-months of daily intraperitoneal injections of saline (healthy control) or D-Gal, were randomly allocated (D-Gal group) to receive intraperitoneal injection of saline (sham control, n=12), P-EVs (n=7), NP-EVs (n=6) and followed-up for one month. While sham control group developed myocardial hypertrophy compared to healthy control animals (heart/body weight 2.7 vs. 2.2 mg/gr, p=0.03), this effect was prevented by P-EVs but not by NP-EVs (2.5 vs. 2.9 mg/gr, p=0.02). This finding was in parallel to the levels of serum antioxidants which were increased in sham vs healthy (p=0.02) and in sham vs. P-EVs (p=0.01) but levels in NP-EVs were similar to the sham group. Conclusions Anti-senescence in vitro effect of CDC-EVs is correlated to their therapeutic efficacy in vivo model of cardiac ageing. The relevance of this study is double: identification of senescence-related markers as determinant of therapeutic potency of the tested product and confirming the potential utility of CDC-EVs as therapy for cardiac ageing-related pathologies. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Instituto de Salud Carlos III, Ministerio de Ciencia e Innovaciόn,CIBERCV, Spain

Percutaneous treatment of patent ductus arteriosus (PDA) in extreme premature infants is technically difficult, and therefore, often not consider as an alternative to surgery. The main objective of our work was to compare respiratory status prior and post ductal closure and morbi-mortality, in our series of preterm infants with percutaneous PDA closure versus surgical ligation in the same time-period. Retrospective review of all premature infants submitted to percutaneous and surgical PDA closure from January 2011 to December 2016. All the antenatal, perinatal, and postnatal characteristics were collected. The main outcome was the assessment of the pulmonary status before and after ductal closure using a pulmonary score. Secondary outcomes included moderate-severe disability in neurodevelopment, death before discharge, moderate-severe chronic lung disease, and morbidity at discharge. 25 patients with a mean weight of 1330 g (± 280) underwent percutaneous closure of PDA with ADO-II-AS, and a total of 53 underwent surgical ligation. 28/53 with similar gestational age, birth weight, and procedure weight to those in the percutaneous group, were selected to perform the comparative study. Ductal closure (percutaneous and surgical) resulted in improved respiratory status. However, percutaneous group achieved a fastest respiratory improvement, than surgical group. The surgical closure group associated higher morbidity among survivors (HIV, number of sepsis, need, and duration of inotropics post-interventionism). The incidence of recurrent laryngeal nerve palsy among the surgical group was 17%. Percutaneous closure of PDA in carefully selected low-weight preterm infants is a safe and reliable alternative to surgical ligation.