Publications by authors named "Alessandra Ciullo"

10 Publications

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Extracellular Vesicles Secreted by TDO2-Augmented Fibroblasts Regulate Pro-inflammatory Response in Macrophages.

Front Cell Dev Biol 2021 22;9:733354. Epub 2021 Oct 22.

Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, United States.

Extracellular vesicles (EVs) are secreted lipid bilayer vesicles that mediate cell to cell communication and are effectors of cell therapy. Previous work has shown that canonical Wnt signaling is necessary for cell and EV therapeutic potency. Tryptophan 2,3-dioxygenase (TDO2) is a target gene of canonical Wnt signaling. Augmenting TDO2 in therapeutically inert fibroblasts endows their EVs with immunomodulatory capacity including attenuating inflammatory signaling in macrophages. Transcriptomic analysis showed that macrophages treated with EVs from fibroblasts overexpressing TDO2 had blunted inflammatory response compared to control fibroblast EVs. , EVs from TDO2-overexpressing fibroblasts preserved cardiac function. Taken together, these results describe the role of a major canonical Wnt-target gene (TDO2) in driving the therapeutic potency of cells and their EVs.
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http://dx.doi.org/10.3389/fcell.2021.733354DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8571098PMC
October 2021

Engineered Fibroblast Extracellular Vesicles Attenuate Pulmonary Inflammation and Fibrosis in Bleomycin-Induced Lung Injury.

Front Cell Dev Biol 2021 23;9:733158. Epub 2021 Sep 23.

Smidt Heart Institute, Cedars Sinai Medical Center, Los Angeles, CA, United States.

Pulmonary fibrosis is a progressive disease for which no curative treatment exists. We have previously engineered dermal fibroblasts to produce extracellular vesicles with tissue reparative properties dubbed activated specialized tissue effector extracellular vesicles (ASTEX). Here, we investigate the therapeutic utility of ASTEX and in a mouse model of bleomycin-induced lung injury. RNA sequencing demonstrates that ASTEX are enriched in micro-RNAs (miRs) cargo compared with EVs from untransduced dermal fibroblast EVs (DF-EVs). Treating primary macrophages with ASTEX reduced interleukin (IL)6 expression and increased IL10 expression compared with DF-EV-exposed macrophages. Furthermore, exposure of human lung fibroblasts or vascular endothelial cells to ASTEX reduced expression of smooth muscle actin, a hallmark of myofibroblast differentiation (respectively). , intratracheal administration of ASTEX in naïve healthy mice demonstrated a favorable safety profile with no changes in body weight, lung weight to body weight, fibrotic burden, or histological score 3 weeks postexposure. In an acute phase (short-term) bleomycin model of lung injury, ASTEX reduced lung weight to body weight, IL6 expression, and circulating monocytes. In a long-term setting, ASTEX improved survival and reduced fibrotic content in lung tissue. These results suggest potential immunomodulatory and antifibrotic properties of ASTEX in lung injury.
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http://dx.doi.org/10.3389/fcell.2021.733158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8512699PMC
September 2021

Exosomally derived Y RNA fragment alleviates hypertrophic cardiomyopathy in transgenic mice.

Mol Ther Nucleic Acids 2021 Jun 20;24:951-960. Epub 2021 Apr 20.

Smidt Heart Institute, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, Los Angeles, CA 90048, USA.

Cardiosphere-derived cell exosomes (CDC) and YF1, a CDC-derived non-coding RNA, elicit therapeutic bioactivity in models of myocardial infarction and hypertensive hypertrophy. Here we tested the hypothesis that YF1, a 56-nucleotide Y RNA fragment, could alleviate cardiomyocyte hypertrophy, inflammation, and fibrosis associated with hypertrophic cardiomyopathy (HCM) in transgenic mice harboring a clinically relevant mutation in cardiac troponin I (cTnI). By quantitative PCR, YF1 was detectable in bone marrow, spleen, liver, and heart 30 min after intravenous (i.v.) infusion. For efficacy studies, mice were randomly allocated to receive i.v. YF1 or vehicle, monitored for ambulatory and cardiac function, and sacrificed at 4 weeks. YF1 (but not vehicle) improved ambulation and reduced cardiac hypertrophy and fibrosis. In parallel, peripheral mobilization of neutrophils and proinflammatory monocytes was decreased, and fewer macrophages infiltrated the heart. RNA-sequencing of macrophages revealed that YF1 confers substantive and broad changes in gene expression, modulating pathways associated with immunological disease and inflammatory responses. Together, these data demonstrate that YF1 can reverse hypertrophic and fibrotic signaling pathways associated with HCM, while improving function, raising the prospect that YF1 may be a viable novel therapeutic candidate for HCM.
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http://dx.doi.org/10.1016/j.omtn.2021.04.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8141670PMC
June 2021

Cardiosphere-derived cells, with and without a biological scaffold, stimulate myogenesis and recovery of muscle function in mice with volumetric muscle loss.

Biomaterials 2021 07 27;274:120852. Epub 2021 Apr 27.

Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, 90048, USA. Electronic address:

Extremity trauma to military personnel and civilians commonly results in volumetric muscle loss (VML), leaving patients suffering chronic physical disability. Biomaterial-based technologies such as extracellular matrices (ECMs) are currently in clinical testing for soft tissue repair, but, in preclinical models of VML, the efficacy of ECMs is equivocal. In a murine model of VML, we investigated the effects of ECM and/or cardiosphere-derived cell (CDC) therapy; the latter improves skeletal myogenesis and muscle function in mdx mice, so we reasoned that CDCs may exert disease-modifying bioactivity in VML. While ECM alone improves functional recovery, CDCs have no additive or synergistic benefits with ECM transplantation following VML injury. However, CDCs alone are sufficient to promote muscle recovery, leading to sustained increases in muscle function throughout the study period. Notably, CDCs stimulate satellite cell accumulation in the muscle defect area and hasten myogenic progression (as evidenced by qPCR gene expression profiling), leading to global increases in myofiber numbers and anterior muscle compartment volume. Together, these data implicate CDCs as a viable therapeutic candidate to regenerate skeletal muscle injured by VML.
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http://dx.doi.org/10.1016/j.biomaterials.2021.120852DOI Listing
July 2021

Small molecule inhibitors and culture conditions enhance therapeutic cell and EV potency via activation of beta-catenin and suppression of THY1.

Nanomedicine 2021 04 13;33:102347. Epub 2020 Dec 13.

Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, USA. Electronic address:

Primary cell therapy continues to face significant hurdles to therapeutic translation including the inherent variations that exist from donor to donor, batch to batch, and scale-up driven modifications to the manufacturing process. Cardiosphere-derived cells (CDCs) are stromal/progenitor cells with clinically demonstrated tissue reparative capabilities. Mechanistic investigations have identified canonical Wnt/β-catenin signaling as a therapeutic potency marker, and THY1 (CD90) expression as inversely correlated with potency. Here we demonstrate that the cardiosphere formation process increases β-catenin levels and enriches for therapeutic miR content in the extracellular vesicles of these cells, namely miR-146a and miR-22. We further find that loss of potency is correlated with impaired cardiosphere formation. Finally, our data show that small GSK3β inhibitors including CHIR, and BIO and "pro-canonical Wnt" culturing conditions can rescue β-catenin signaling and reduce CD90 expression. These findings identify strategies that could be used to maintain CDC potency and therapeutic consistency.
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http://dx.doi.org/10.1016/j.nano.2020.102347DOI Listing
April 2021

Extracellular Vesicles as Therapeutic Agents for Cardiac Fibrosis.

Front Physiol 2020 21;11:479. Epub 2020 May 21.

Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, United States.

Heart disease remains an increasing major public health challenge in the United States and worldwide. A common end-organ feature in diseased hearts is myocardial fibrosis, which stiffens the heart and interferes with normal pump function, leading to pump failure. The development of cells for regenerative therapy has been met with many pitfalls on its path to clinical translation. Recognizing that regenerative cells secrete therapeutically bioactive vesicles has paved the way to circumvent many failures of cell therapy. In this review, we provide an overview of extracellular vesicles (EVs), with a focus on their utility as therapeutic agents for cardiac regeneration. We also highlight the engineering potential of EVs to enhance their therapeutic application.
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http://dx.doi.org/10.3389/fphys.2020.00479DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7255103PMC
May 2020

Inflammatory extracellular vesicles prompt heart dysfunction via TRL4-dependent NF-κB activation.

Theranostics 2020 3;10(6):2773-2790. Epub 2020 Feb 3.

Laboratory for Cardiovascular Theranostics, Cardiocentro Ticino Foundation, Lugano, Switzerland.

: After myocardial infarction, necrotic cardiomyocytes release damage-associated proteins that stimulate innate immune pathways and macrophage tissue infiltration, which drives inflammation and myocardial remodeling. Circulating inflammatory extracellular vesicles play a crucial role in the acute and chronic phases of ischemia, in terms of inflammatory progression. In this study, we hypothesize that the paracrine effect mediated by these vesicles induces direct cytotoxicity in cardiomyocytes. Thus, we examined whether reducing the generation of inflammatory vesicles within the first few hours after the ischemic event ameliorates cardiac outcome at short and long time points. : Myocardial infarction was induced in rats that were previously injected intraperitoneally with a chemical inhibitor of extracellular-vesicle biogenesis. Heart global function was assessed by echocardiography performed at 7, 14 and 28 days after MI. Cardiac outcome was also evaluated by hemodynamic analysis at sacrifice. Cytotoxic effects of circulating EV were evaluated in a Langendorff, system by measuring the level of cardiac troponin I (cTnI) in the perfusate. Mechanisms undergoing cytotoxic effects of EV derived from pro-inflammatory macrophages (M1) were studied in primary rat neonatal cardiomyocytes. : Inflammatory response following myocardial infarction dramatically increased the number of circulating extracellular vesicles carrying alarmins such as IL-1α, IL-1β and Rantes. Reducing the boost in inflammatory vesicles during the acute phase of ischemia resulted in preserved left ventricular ejection fraction . Hemodynamic analysis confirmed functional recovery by displaying higher velocity of left ventricular relaxation and improved contractility. When added to the perfusate of isolated hearts, post-infarction circulating vesicles induced significantly more cell death in adult cardiomyocytes, as assessed by cTnI release, comparing to circulating vesicles isolated from healthy (non-infarcted) rats. inflammatory extracellular vesicles induce cell death by driving nuclear translocation of NF-κB into nuclei of cardiomyocytes. : Our data suggest that targeting circulating extracellular vesicles during the acute phase of myocardial infarction may offer an effective therapeutic approach to preserve function of ischemic heart.
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http://dx.doi.org/10.7150/thno.39072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7052909PMC
May 2021

Intravenous administration of cardiac progenitor cell-derived exosomes protects against doxorubicin/trastuzumab-induced cardiac toxicity.

Cardiovasc Res 2020 02;116(2):383-392

Laboratory of Cellular and Molecular Cardiology, Cardiocentro Ticino Foundation, 6900 Lugano, Switzerland.

Aims: Combined administration of anthracyclines (e.g. doxorubicin; Dox) and trastuzumab (Trz), a humanized anti-human epidermal growth factor receptor 2 (HER2; ErbB2), is an effective treatment for HER2-positive breast cancer. However, both agents are associated with cardiac toxicity. Human cardiac-resident mesenchymal progenitor cells (CPCs) secrete extracellular vesicles including nanosized exosomes which protect against myocardial ischaemia. Here, we investigated the effects of these exosomes using a novel model of Dox/Trz-mediated cardiotoxicity.

Methods And Results: CPCs were derived from cardiac atrial appendage specimens from patients who underwent heart surgery for heart valve disease and/or ischaemic heart disease, and exosomes were purified from CPC conditioned media. Proteomics analyses revealed that CPC exosomes contained multiple proteins involved in redox processes. Dox/Trz induced a significant increase in reactive oxygen species (ROS) in rat cardiomyocytes, which was prevented by CPC exosomes. In vivo, rats received six doses of Dox (Days 1-11), followed by six doses of Trz (Days 19-28). Three doses of either exosomes or exosome suspension vehicle were injected intravenously on Days 5, 11, and 19 in the treatment and control groups, respectively. Dox/Trz induced myocardial fibrosis, CD68+ inflammatory cell infiltrates, inducible nitric oxide synthase expression, and left ventricular dysfunction. CPC exosomes prevented these effects. These vesicles were highly enriched in miR-146a-5p compared with human dermal fibroblast exosomes. Dox upregulated Traf6 and Mpo, two known miR-146a-5p target genes (which encode signalling mediators of inflammatory and cell death axes) in myocytes. CPC exosomes suppressed miR-146a-5p target genes Traf6, Smad4, Irak1, Nox4, and Mpo in Dox-treated cells. Specific silencing of miR-146a-5p abrogated exosome-mediated suppression of those genes leading to an increase in Dox-induced cell death.

Conclusions: Human CPC exosomes attenuate Dox-/Trz-induced oxidative stress in cardiomyocytes. Systemic administration of these vesicles prevents Dox/Trz cardiotoxicity in vivo. miR-146a-5p mediates some of the benefits of exosomes in this setting.
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http://dx.doi.org/10.1093/cvr/cvz108DOI Listing
February 2020

Exosomal Expression of CXCR4 Targets Cardioprotective Vesicles to Myocardial Infarction and Improves Outcome after Systemic Administration.

Int J Mol Sci 2019 Jan 22;20(3). Epub 2019 Jan 22.

Cellular and Molecular Cardiology Laboratory, Cardiocentro Ticino Foundation, 6900 Lugano, Switzerland.

Cell therapy has been evaluated to enhance heart function after injury. Delivered cells mostly act via paracrine mechanisms, including secreted growth factors, cytokines, and vesicles, such as exosomes (Exo). Intramyocardial injection of cardiac-resident progenitor cells (CPC)-derived Exo reduced scarring and improved cardiac function after myocardial infarction in rats. Here, we explore a clinically relevant approach to enhance the homing process to cardiomyocytes (CM), which is crucial for therapeutic efficacy upon systemic delivery of Exo. By overexpressing exosomal CXCR4, we increased the efficacy of plasmatic injection of cardioprotective Exo-CPC by increasing their bioavailability to ischemic hearts. Intravenous injection of Exo significantly reduced infarct size and improved left ventricle ejection fraction at 4 weeks compared to Exo ( < 0.01). Hemodynamic measurements showed that Exo improved dp/dt min, as compared to Exo and PBS group. In vitro, Exo was more bioactive than Exo in preventing CM death. This in vitro effect was independent from SDF-1α, as shown by using AMD3100 as specific CXCR4 antagonist. We showed, for the first time, that systemic administration of Exo derived from CXCR4-overexpressing CPC improves heart function in a rat model of ischemia reperfusion injury These data represent a substantial step toward clinical application of Exo-based therapeutics in cardiovascular disease.
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http://dx.doi.org/10.3390/ijms20030468DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6386845PMC
January 2019

Cardioprotection by cardiac progenitor cell-secreted exosomes: role of pregnancy-associated plasma protein-A.

Cardiovasc Res 2018 06;114(7):992-1005

Cellular and Molecular Cardiology Laboratory, Cardiocentro Ticino Foundation and Swiss Institute for Regenerative Medicine (SIRM), via Tesserete 48, 6900 Lugano, Switzerland.

Aims: Cell therapy trials using cardiac-resident progenitor cells (CPCs) and bone marrow-derived mesenchymal stem/progenitor cells (BMCs) in patients after myocardial infarction have provided encouraging results. Exosomes, nanosized extracellular vesicles of endosomal origin, figure prominently in the bioactivities of these cells. However, a head-to-head comparison of exosomes from the two cell types has not been performed yet.

Methods And Results: CPCs and BMCs were derived from cardiac atrial appendage specimens and sternal bone marrow, respectively, from patients (n = 20; age, 69.9 ± 10.9) undergoing heart surgery for aortic valve disease and/or coronary artery disease. Vesicles were purified from cell conditioned media by centrifugation/filtration and ultracentrifugation. Vesicle preparations were predominantly composed of exosomes based on particle size and marker expression (CD9, CD63, CD81, Alix, and TSG-101). CPC-secreted exosomes prevented staurosporine-induced cardiomyocyte apoptosis more effectively than BMC-secreted exosomes. In vivo, CPC-secreted exosomes reduced scar size and improved ventricular function after permanent coronary occlusion in rats more efficiently than BMC-secreted exosomes. Both types of exosomes stimulated blood vessel formation. CPC-secreted exosomes, but not BMC-derived exosomes, enhanced ventricular function after ischaemia/reperfusion. Proteomics profiling identified pregnancy-associated plasma protein-A (PAPP-A) as one of the most highly enriched proteins in CPC vs. BMC exosomes. The active form of PAPP-A was detected on CPC exosome surfaces. These vesicles released insulin-like growth factor-1 (IGF-1) via proteolytic cleavage of IGF-binding protein-4 (IGFBP-4), resulting in IGF-1 receptor activation, intracellular Akt and ERK1/2 phosphorylation, decreased caspase activation, and reduced cardiomyocyte apoptosis. PAPP-A knockdown prevented CPC exosome-mediated cardioprotection both in vitro and in vivo.

Conclusion: These results suggest that CPC-secreted exosomes may be more cardioprotective than BMC-secreted exosomes, and that PAPP-A-mediated IGF-1 release may explain the benefit. They illustrate a general mechanism whereby exosomes may function via an active protease on their surface, which releases a ligand in proximity to the transmembrane receptor bound by the ligand.
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http://dx.doi.org/10.1093/cvr/cvy055DOI Listing
June 2018
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