Aging is a major risk factor for cardiovascular disease, the leading cause of death worldwide, and numerous other diseases, but the mechanisms of these aging-related effects remain elusive. Recent evidence suggests that chronic changes in the microenvironment and local paracrine signaling are major drivers of these effects, but the precise effect of aging on these factors remains understudied. Here, for the first time, we directly compare extracellular vesicles obtained from young and aged patients to identify therapeutic or disease-associated agents, and directly compare vesicles isolated from heart tissue matrix (TEVs) or plasma (PEVs). While young TEVs and PEVs showed notable overlap of miRNA cargo, aged EVs differed substantially, indicating differential aging-related changes between TEVs and PEVs. TEVs overall were uniquely enriched in miRNAs which directly or indirectly demonstrate cardioprotective effects, with 45 potential therapeutic agents identified in our analysis. Both populations also showed increased predisposition to disease with aging, though through different mechanisms. Changes in PEV cargo were largely correlated with chronic systemic inflammation, while those in TEVs were more related to cardiac homeostasis and local inflammation. From this, 17 protein targets were identified which were unique to TEVs and highly correlated with aging and the onset of cardiovascular disease. Further analysis via machine learning techniques implicated several new miRNA and protein targets, independently suggesting several of the targets identified by non-machine learning analysis, which correlated with aging-related changes in TEVs. With further study, this biomarker set may serve as a powerful, potential indicator of cardiac health and age which can be measured from PEVs. Additionally, several proposed “young-enriched” therapeutic agents were validated and, when tested, could successfully prevent cell death and cardiac fibrosis in disease-like conditions using a microfluidic heart-on-a-chip to model of acute and chronic fibrosis, making this study the first in literature to test the efficacy of a miRNA-based therapeutic encapsulated in lipid nanoparticles in an organ-on-a-chip device.

Cardiovascular disease (CVD) is the leading cause of death worldwide. Chemotherapy-induced CVD is increasingly recognized as a contributor to long-term morbidity in cancer survivors. Doxorubicin (DOX) is a widely used chemotherapeutic to treat breast cancer, one of the most common cancers in the United States. However, over 10% of treated women experience acute cardiotoxicity immediately following treatment, and approximately 2% develop severe cardiotoxicity up to 10 years after treatment, yet the mechanisms driving this delayed onset remain unclear. Here, we show that DOX treatment of cardiac cells changes their function and paracrine signaling profile. Subsequent exposure of healthy cells to altered extracellular vesicles (EV) recapitulates the effects of direct DOX exposure in 2D/3D in vitro models, suggesting a mechanism for propagating initial injury. Plasma-EV miRNA profiling of blinded patient samples revealed distinct clustering by DOX-cardiotoxicity risk, with high-risk patients exhibiting miRNA signatures similar to those from DOX-treated models. Pathway analysis of miRNAs linked them to cardiac homeostasis and cardiotoxicity-related mechanisms, supporting the potential of plasma-EV miRNAs as noninvasive biomarkers for early risk stratification and personalized cardioprotective interventions in oncological care, and targeting of key miRNA clusters to enhance understanding of and intervention strategies for preventing the onset of DOX cardiotoxicity. STATEMENT OF SIGNIFICANCE: Doxorubicin is an effective chemotherapy drug, but its use is limited by cardiotoxicity that can appear during treatment or years later. The mechanisms driving this delayed injury remain poorly understood. In this study, we show for the first time that small extracellular vesicles released from doxorubicin-treated cardiac cells can propagate doxorubicin-like dysfunction to healthy cardiac cells even in the absence of the drug itself. We further identify broad miRNA cargo changes in these vesicles and show that related vesicle-associated miRNA shifts are also detectable in patient plasma. These findings provide new mechanistic insight into anthracycline cardiotoxicity and highlight extracellular vesicle-associated miRNAs as promising candidates for minimally invasive diagnosis and future therapeutic intervention.

3D printing technology can precisely simulate the complex structure of natural meat, but balancing the viscosity and mechanical properties of printing inks remains challenging. This study proposes a one-pot dual-crosslinking strategy. CaCO3/GDL-mediated ionic crosslinking provides immediate structural support for 3D printing, while temperature regulation delays the TG enzyme crosslinking rate. This enables the interpenetration of covalent and ionic networks to form a composite network structure, ensuring long-term product stability. The effects of crosslinking sequence and gelatin concentration were investigated. Separating the crosslinking sequence significantly enhanced hydrogel viscoelasticity and structural recovery, improving print fidelity. Synergistic ion and delayed enzymatic crosslinking formed a uniform, dense honeycomb structure. Increasing gelatin concentration (2.5%-10%) markedly elevated the hydrogel's storage modulus, compressive modulus, hardness, and chewiness. Furthermore, water holding capacity approached 100%, with thawing loss and cooking loss reduced to 2.83% and 7.27%, respectively. This study provides novel strategies for texture regulation in 3D-printed meat analogues.

Reproducing the spatiotemporally coordinated emergence of vascular, biliary, and parenchymal compartments of the human liver remains a persistent bottleneck in organoid engineering. We introduce biodegradable poly(lactic-co-glycolic acid) microspheres that display epithelial (E) cadherin and vascular endothelial (VE) cadherin fusion proteins and deliver morphogens, mimicking key developmental signals that guide liver organogenesis. When combined with mesenchymal stem cells (MSCs), the microspheres supply adhesion cues that drive aggregate compaction. E-cadherin engagement induces Yes-associated protein activation, priming MSCs for lineage induction. Complementarily, VE-cadherin and vascular endothelial growth factor jointly establish an endothelial niche that delivers inductive cues for hepatic lineage divergence. The resulting organoids display compartmentalized architecture comprising hepatocytes, cholangiocytes, mesenchymal cells, and endothelial networks and respond to hepatotoxic compounds with clinically relevant phenotypes. Unlike induced pluripotent stem cell-based or multiple cell cocultures, this single-source MSC system achieves self-organization through cadherin-defined adhesion and timed morphogen delivery. These elements define a scalable, cadherin-defined biomaterial platform for liver organoid engineering, enabling translational use in drug evaluation and regenerative medicine.

Ischemic stroke inflicts severe neurological damage by disrupting the neurovascular unit. While promising, mesenchymal stem cell (MSC) therapies are hampered by poor posttransplantation survival and nonspecific secretomes. Here, we introduce a bioengineering strategy that employs cadherin-functionalized interfaces to generate cohesive multicellular MSC aggregates (Cad-MAs). Priming MSCs with recombinant N-cadherin and VE-cadherin stimulated endogenous cadherin expression and facilitated the self-assembly of stable spheroids with reinforced intercellular adherens junctions. Cad-MAs exhibited increased resistance to inflammatory stress and anoikis, and secreted a reparative profile enriched in neurotrophic and angiogenic factors, as well as exosomes carrying therapeutic miRNAs such as miR-21-5p and miR-126-3p. The in vitro analyses indicate that cadherin-empowered assembly yields MSC aggregates in which structural stability is coupled with a pro-survival, pro-regenerative phenotype. Furthermore, in a mouse stroke model, systemically delivered Cad-MAs significantly outperformed conventional dissociated MSCs, promoting functional recovery, reducing infarct volume, and improving cerebral perfusion alongside evidence of enhanced angiogenesis and preservation of blood-brain barrier integrity markers. This approach, termed functional aggregation-induced emergence (F-AIE), provides a versatile framework for engineering integrated cellular therapeutics with tailored functional outputs for regenerative applications.

Dietary polyphenols, such as curcumin have excellent bioactivity such as anti-inflammatory and antioxidant activity. However, their application in treating ulcerative colitis (UC) is hampered by poor aqueous solubility, stability, and oral bioavailability. These limitations can be effectively overcome using advanced oral delivery systems. Composite nanoparticles enhance the stability and bioavailability of curcumin and modulate its metabolic profile. The metabolite profiles of curcumin nanoparticles (Cur-awNPs) and free curcumin in serum and fecal samples from UC-treated mice were analyzed using UPLC-Q-TOF/MS. Metabolomics revealed over a dozen differential metabolites across the control, model, Cur-awNPs, and free curcumin groups. Notably, metabolites such as tryptophan, linoleic acid, and arachidonic acid were significantly altered in the model group, and treatment with Cur-awNPs effectively regulated these metabolite levels in UC mice. Metabolic pathway analysis indicated that Cur-awNPs primarily influenced linoleic acid metabolism, riboflavin metabolism, tryptophan metabolism, and arachidonic acid metabolism. Correlation analysis revealed significant correlations between riboflavin, tryptophan, arachidonic acid, linoleic acid and inflammatory factors, which can be used as potential biomarkers. This study provides novel insights and a theoretical basis for the mechanisms underlying the targeted delivery of dietary polyphenols to ameliorate UC.

Extracellular vesicles (EVs) are an important carrier of cellular communication that contain cargo such as cytokines, RNAs, or microRNAs (miRNA) and have been proven to play an important role in breast cancer tumorigenesis, progression, and metastasis. Although the role of cancer associated fibroblasts (CAFs), and EVs originated from them have been studied extensively, there is a lack in knowledge on the contribution of normal fibroblasts surrounding the tumor and their roles with respect to their proximity to the tumor. Here we investigate how the proximity of the tumor affects the EV production of the normal fibroblasts. We created stromal models by 3D bioprinting two different fibroblasts, normal human mammary fibroblasts (hMFs) and normal tumor adjacent fibroblasts (NTAF), within a collagen gel. We isolated EVs from both the effluent media and the 3D stromal model, which were then characterized and we found that EVs from each group were of consistent exosome size and displayed traditional exosome markers, however, the EVs from different groups also displayed different cytokine profiles of their cargo, with the NTAF media group showing an upregulation of cytokines associated with breast cancer progression. After this, we used the EVs to treat breast cancer cells to investigate the effects of the EV groups on the breast cancer cell behavior. The breast cancer cells treated with the NTAF groups had increased migration. Finally, we utilized a 3D breast tumor model to investigate the effects of the EVs on a tumor spheroid. Tumor spheroids treated with either NTAF EV groups showed increased proliferation, tumor diameter, and local invasion. This study is the first to investigate the effect of proximity to a breast tumor on EV production and the first to utilize 3D bioprinting of stromal models specifically to obtain EVs. Overall, our results show that EVs from normal fibroblasts closer to a tumor produce EVs that promote breast cancer progression, regardless of the secretion location of the EVs. These cells have a distinct EV secretome different from normal human mammary fibroblasts, showing that the proximity to a tumor influences the normal fibroblasts surrounding the tumor.

Elevated ferritin levels in breast invasive carcinoma correlate strongly with unfavorable clinical outcomes. However, current ferritin-reducing strategies (e.g., CRISPR, PROTACs) are limited by inefficient large protein aggregate degradation and poor cancer cell specificity. Ferritinophagy-an intrinsic ferritin-lowering process that supplies intracellular iron via autophagy-can induce ferroptosis when dysregulated. Inspired by this mechanism, we engineered Ferritin NanoChaperone (FerriNano), which consists of a ferritin-homing peptidefunctionalized nanoceria, (HK-CeO2) coated with hyaluronic acid (HA). Upon administration, FerriNano selectively accumulated in 4T1 tumor tissues via the interaction between HA and CD44 on cancer cells. The nanochaperone, HK-CeO2, can selectively bind to overexpressed ferritin, and the formed complex can be sequestered in autophagosomes by in situ autophagy activation of nanoceria, which subsequently fuse with lysosomes to execute ferritin degradation. Consequently, FerriNano demonstrated potent downregulated ferritin levels and reduced tumor metastasis. Notably, approximately 3.7 % of the administered FerriNano traversed the intestinal epithelium via hepatic bile secretion. This systemic distribution enhanced gut microbiota diversity and metabolic activity, potentially amplifying therapeutic outcomes through microbiota-immune interactions. Overall, this study not only identifies candidate nanochaperones for targeted ferritin degradation but also highlights the significance of evaluating the metabolic byproducts for breast cancer treatment.

Aging is one of the most significant risk factors for breast cancer. With the growing interest in the alterations of the aging breast tissue microenvironment, it is identified that aging is related to tumorigenesis, invasion, and drug resistance. However, current pre-clinical disease models often neglect the impact of aging and sometimes result in worse clinical outcomes. In this study, aged animal-generated materials are utilized to create and validate a novel age-mimetic breast cancer model that generates an aging microenvironment for cells and alters cells toward a more invasive phenotype found in the aged environment. Furthermore, the age-mimetic models are utilized for 3D breast cancer invasion assessment and high-throughput screening of over 700 drugs in the FDA-approved drug library. 36 potential effective drug targets as well as 34 potential drug targets with different drug responses in different age groups are identified, demonstrating the potential of this age-mimetic breast cancer model for further in-depth breast cancer studies and drug development.

Umbilical cord-derived mesenchymal stromal cells (MSCs) transplantation is a promising therapy for systemic sclerosis (SSc) because of their distinctive antifibrotic and immunomodulatory properties. To enhance the effects of MSCs transplantation, novel engineered MSC aggregates preconditioned with human E/N-cadherin fusion protein (hE/N-cad-Fc) and interleukin-6 (IL-6) for SSc treatment are fabricated and named 3D-Cad/IL6-MSCs. These novel-engineered MSC aggregates possess tighter cellular cohesion and exhibit enhanced antiapoptotic, immunosuppressive, and proangiogenic capabilities according to transcriptomic analyses. Moreover, 3D-Cad/IL6-MSCs have improved immunoregulatory effects on peripheral blood mononuclear cells (PBMCs), CD4+ T cells, and CD8+ T cells in vitro because of the synergistic preconditioning from IL-6 and bioactive hE/N-cad-Fc. Upon intravenous injection, 3D-Cad/IL6-MSCs significantly mitigate skin and lung fibrosis and prolong the retention duration in bleomycin (BLM)-induced SSc mouse model. In detail, they suppress excessive infiltration of macrophages and T cells in the injured skin and lungs and reestablish the immune equilibrium of circulating CD4+ T-cell subsets in vivo. These results suggest that 3D-Cad/IL6-MSCs are ideal candidates for SSc therapy and optimize the clinical utilization of MSCs.

Aging is a key risk factor for breast cancer; however, the independent effects of the aged extracellular matrix (ECM) remain understudied. To address this, we developed a novel hybrid in vivo model that enables the independent investigation of age-related ECM influences on breast cancer development and progression. To examine the effects of genes known to be enriched in a cancerous and aged microenvironment, we first seeded normal or stable knockdown cells onto decellularized ECM (dECM) from aged murine mammary glands, and implanted them into young Rag1 −/− mice. We identified LOX as a principal driver of tumor progression, with knockdown reducing invasion and stress-related pathways. To further isolate the independent influence of ECM aging on tumor growth, normal MCF10A cells were seeded atop young or aged matrices and implanted. Aged tumors exhibited significantly greater volume and a larger tumorigenic region when compared to young, with single-cell RNA sequencing revealing enrichment of inflammatory and invasive genes. Together, these findings identify LOX as a driver of tumor progression and a potential therapeutic target and demonstrate that the aged ECM alone is sufficient to promote breast cancer progression.

The extracellular matrix (ECM) is recognized as a key regulator of cell behavior, with its stiffness playing a crucial role in the progression of pathological conditions such as cancer and cardiovascular diseases. While extracellular vesicles (EVs) are essential mediators of intercellular communication, the influence of matrix stiffness on EV secretion remains poorly understood. This study investigates how substrate stiffness affects EV size and composition in mouse mammary and cardiac fibroblasts, the key stromal cell types in breast cancer and cardiac microenvironments. Importantly, stiffness-tuned EV proteomic cargo is uncovered, providing new insights into how mechanical cues can reprogram the signaling functions of fibroblast-derived vesicles. The findings show that substrate stiffness significantly alters EV characteristics, with sizes increasing below stiffnesses of 20 kPa and decreasing on stiffer substrates. Mechanotransduction pathways involving p53 and thioredoxin are identified as regulators of these alterations, with thioredoxin dominate the modulation in mammary fibroblasts and p53 in cardiac fibroblasts. These results underscore the importance of ECM stiffness in modulating EV secretion and highlight candidate pathways influenced by ECM remodeling that may warrant further investigation for therapeutic relevance.

Inflammatory bowel disease (IBD) is an incurable condition driven by chronic inflammatory dysregulation. Human mesenchymal stem cells (hMSCs) offer therapeutic promise, but limited persistence and functional heterogeneity constrain their clinical efficacy. Here, we present a bio-inspired priming platform built on poly (lactic-co-glycolic acid)/chitosan-heparin microparticles, engineered to display E/N-cadherin motifs and to enable controlled interleukin-1(3 release through PLGA biodegradation. These functionalized microparticles (hE/ N-cad@P/C-h-IL-1(3) assemble with hMSCs into hybrid aggregates that enhance cell survival under stress and improve adaptability to microenvironment cues. The priming process upregulates endogenous cadherin expression and modulates AKT/STAT signaling, conferring stimulus-and dose-responsive immune activity. In vitro, hybrid aggregates polarize macrophages toward an anti-inflammatory phenotype, expand regulatory T cells, and suppress pathogenic lymphocyte activation. In a dextran sulfate sodium (DSS) colitis model, intraperitoneal delivery of hybrid aggregates restores intestinal immune homeostasis, accelerates epithelial repair, and reduces disease severity to near baseline within fourteen days. By integrating cell-adhesive and cytokine signals within a modular platform, this strategy overcomes critical translational barriers in hMSC therapy and provides a blueprint for precisely targeted treatment of IBD and related inflammatory disorders.

Aging is a significant cancer risk factor, yet its impact on the extracellular matrix (ECM) in tumor initiation and progression has been traditionally overlooked. While significant amounts of research focus on cellular and genetic links between aging and cancer, recent studies highlight how age-induced ECM changes create a tumor-permissive environment. Here we review this emerging research area, where age-related ECM alterations, such as age-induced increases in matrix stiffness, biochemical changes, and resultant dysregulated mechanosensitive pathways, are explored for their influence in cancer initiation and progression. Additionally, recent studies have showed how aged cells contribute to ECM alterations, further reinforcing tumor-permissive changes. This review examines both aspects of ECM aging, i.e. material-driven and cell-driven, and highlights current understandings of how ECM aging influences interactions within the tumor microenvironment in multiple cancer types, with a focus on biomechanical aspects. We also discuss emerging age-mimetic in vitro models facilitating studies of age-dependent cancer progression and therapeutic responses. Finally, we review therapeutic strategies that target aging-associated components or ECM changes to improve treatment efficacy.

Even with the promising therapeutic potential of engineered bacteria that target tumors, effective cancer treatment will still need to involve combination strategies. Herein, a model strain of Escherichia coli (BL21) is aimed to engineer to internally produce copper sulfide nanoparticles (CuS), which are subsequently armed with disulfiram-loaded gold nanorods, resulting in Au-DSF@CuS-BL21 for hypoxic-targeted prodrug delivery and enhanced antitumor effects. The bacterial membrane in this system serves as a barrier between the prodrugs of CuS and DSF, preventing the premature formation and release of the highly toxic CuET complex during circulation, thus ensuring bacterial survival and minimizing drug leakage. The key findings revealed that bacterial activity of Au-DSF@CuS-BL21 is effectively sustained, exhibiting enhanced capabilities for tumor targeting and penetration. Upon activation by 808 nm near-infrared laser irradiation, photothermal bacterial lysis is induced, resulting in the release of CuS and DSF. This process facilitate the formation of the highly toxic CuET complex, as well as augmented antitumor efficacy by stimulating the innate immune response. Collectively, these findings offer promising opportunities for enhancing the biosafety of modified bacteria, as well as for compartmentalized prodrug delivery and improved effectiveness of combination antitumor therapies.

Aging is a major risk factor for cardiovascular disease, the leading cause of death worldwide, and numerous other diseases, but the mechanisms of these aging-related effects remain elusive. Recent evidence suggests that chronic changes in the microenvironment and local paracrine signaling are major drivers of these effects, but the precise effect of aging on these factors remains understudied. Here, for the first time, we directly compare extracellular vesicles obtained from young and aged patients to identify therapeutic or disease-associated agents, and directly compare vesicles isolated from heart tissue matrix (TEVs) or plasma (PEVs). While young TEVs and PEVs showed notable overlap of miRNA cargo, aged EVs differed substantially, indicating differential aging-related changes between TEVs and PEVs. TEVs overall were uniquely enriched in miRNAs which directly or indirectly demonstrate cardioprotective effects, with 45 potential therapeutic agents identified in our analysis. Both populations also showed increased predisposition to disease with aging, though through different mechanisms. Changes in PEV cargo were largely correlated with chronic systemic inflammation, while those in TEVs were more related to cardiac homeostasis and local inflammation. From this, 17 protein targets were identified which were unique to TEVs and highly correlated with aging and the onset of cardiovascular disease. Further analysis via machine learning techniques implicated several new miRNA and protein targets, independently suggesting several of the targets identified by non-machine learning analysis, which correlated with aging-related changes in TEVs. With further study, this biomarker set may serve as a powerful, potential indicator of cardiac health and age which can be measured from PEVs. Additionally, several proposed "young-enriched" therapeutic agents were validated and, when tested, could successfully prevent cell death and cardiac fibrosis in disease-like conditions.

Extracellular vesicles (EVs) are an important carrier of cellular communication that are secreted from the cell. Different cells will produce EVs with different cargo such as cytokines, RNAs, or microRNAs (miRNA). EVs have been proven to play an important role in breast cancer tumorigenesis, progression, and metastasis. Although the role of cancer associated fibroblasts (CAFs), and EVs originated from them have been studied extensively, there is a lack in knowledge on the contribution of normal fibroblasts surrounding the tumor and their roles with respect to their proximity to the tumor. Here we investigate how the proximity of the tumor affects the EV production of the normal fibroblasts. We created stromal models by 3D bioprinting two different fibroblasts, normal human mammary fibroblasts (hMFs) and normal tumor adjacent fibroblasts (NTAF), within a collagen gel. After one week of culture, we isolated EVs from both the effluent media and the 3D stromal model, which were then characterized using nanoparticle tracking analysis (NTA), transmission electron microscopy (TEM), ELISA, zeta potential, and cytokine array analysis of the cargo. The EVs from each group were of consistent exosome size and displayed traditional exosome markers, however the EVs from different groups also displayed different cytokine profiles of their cargo, with the NTAF media group showing an upregulation of cytokines associated with breast cancer progression. After this, we used the EVs to treat breast cancer cells to investigate the effects the EVs from different tumor proximities have on the breast cancer cell behavior. The breast cancer cells treated with the NTAF groups had increased migration. Finally, we utilized a 3D breast tumor model to investigate the effects of the EVs on a tumor spheroid. Tumor spheroids treated with either NTAF EV groups showed increased proliferation, tumor radius, and local invasion. This study is the first to investigate the effect of proximity to a breast tumor on EV production and the first to utilize 3D bioprinting of stromal models specifically to obtain EVs. Overall, our results show that EVs from normal fibroblasts closer to a tumor produce EVs that promote breast cancer progression, regardless of the secretion location of the EVs. These cells have a distinct EV secretome different from normal human mammary fibroblasts, showing that the proximity to a tumor influences the normal fibroblasts surrounding the tumor. ### Competing Interest Statement The authors have declared no competing interest.

beta -galactosidase (lactase) is commercially important as a dietary supplement to alleviate the symptoms of lactose intolerance. This work investigated a unique activation of CMP (carboxymethylated (1 -> 3)- beta - D -glucan) on lactase and its mechanism by comparing it with carboxymethyl chitosan (CMCS), an inhibitor of lactase. The results illustrated that the secondary and tertiary structures of lactase were altered and its active sites exposed after complexation with CMP, and dissociation of lactase aggregates was also observed. These changes favored better accessibility of the substrate to the active sites of lactase, resulting in a maximum increase of 60.5 % in lactase activity. Furthermore, the hydrophobic and electrostatic interactions with lactase caused by the carboxymethyl group of CMP were shown to be crucial for its activation ability. Thus, the improvement of lactase activity and stability by CMP shown here is important for the development of new products in the food and pharmaceutical industries.

Aging is one of the inherent risk factors for breast cancer. Although the influence of age-related cellular alterations on breast cancer development has been extensively explored, little is known about the alterations in the aging breast tissue microenvironment, specifically the extracellular matrix (ECM). Here, for the first time in literature, we have identified tissue resident matrix bound vesicles (MBVs) within the healthy mouse breast ECM, investigated and compared their characteristics in young and aged healthy breast tissues, and studied the effects of these MBVs on normal (KTB21) and cancerous (MDA-MB-231) human mammary epithelial cells with respect to the tissue age that they are extracted from. Using vesicle labeling technology, we were able to visualize cellular uptake of the MBVs directly from the native decellularized tissue sections, showing that these MBVs have regulatory roles in the tissue microenvironment. We mimicked the ECM by embedding the MBVs in collagen gels, and showed that MBVs could be taken up by the cells. The miRNA and cytokine profiling showed that MBVs shifted towards a more tumorigenic and invasive phenotype with age, as evidenced by the more pronounced presence of cancer-associated cytokines, and higher expression levels of oncomiRs miR-10b, miR-30e, and miR-210 in MBVs isolated from aged mice. When treated with MBVs or these upregulated factors, KTB21 and MDA-MB-231 cells showed significantly higher motility and invasion compared to untreated controls. Treatment of cells with a cocktail of miRNAs (miR-10b, miR-30e, and miR-210) or with the agonist of adiponectin (AdipoRon), which both were enriched in the aged MBVs, recapitulated the effect of aged MBVs on cells. This study shows for the first time that the MBVs have a regulatory role in the tissue microenvironment and that the MBV contents change towards cancer-promoting upon aging. Studying the effects of MBVs and their cargos on cellular behavior could lead to a better understanding of the critical roles of MBVs played in breast cancer progression and metastasis.

Aging is a risk factor for cardiovascular disease, the leading cause of death worldwide. Cardiac fibrosis is a harmful result of repeated myocardial infarction that increases risk of morbidity and future injury. Interestingly, both rates and outcomes of cardiac fibrosis differ between young and aged individuals, as well as men and women. Here, for the first time, we identify and isolate matrix-bound extracellular vesicles from the left ventricles (LVs) of young or aged males and females in both human and murine models. These LV vesicles (LVVs) show differences in morphology and content between these four cohorts in both humans and mice. LVV effects on fibrosis were also investigated in vitro, and aged male LVVs were pro-fibrotic while other LVVs were anti-fibrotic. From these LVVs, we could identify therapeutic miRNAs to promote anti-fibrotic effects. Four miRNAs were identified and together, but not individually, demonstrated significant cardioprotective effects when transfected. This suggests that miRNA synergy can regulate cell response, not just individual miRNAs, and also indicates that biological agent-associated therapeutic effects may be recapitulated using non-immunologically active agents. Furthermore, that chronic changes in LVV miRNA content may be a major factor in sex- and age-dependent differences in clinical outcomes of cardiac fibrosis.