Milk Exosomes Research Articles

Supplementation with a Whey Protein Concentrate Enriched in Bovine Milk Exosomes Improves Longitudinal Growth and Supports Bone Health During Catch-Up Growth in Rats

Supplementation with a Whey Protein Concentrate Enriched in Bovine Milk Exosomes Improves Longitudinal Growth and Supports Bone Health During Catch-Up Growth in Rats

Human Breast Milk Exosomes: Affecting Factors, Their Possible Health Outcomes, and Future Directions in Dietetics

Background: Human breast milk is a complex biological fluid containing multifaceted biological compounds that boost immune and metabolic system development that support the short- and long-term health of newborns. Recent literature suggests that human breast milk is a substantial source of nutrients, bioactive molecules, and exosomes. Objectives: This review examines the factors influencing exosomes noted in human milk and the impacts of exosomes on infant health. Furthermore, it discusses potential future prospects for exosome research in dietetics. Methods: Through a narrative review of the existing literature, we focused on exosomes in breast milk, exosome components and their potential impact on exosome health. Results: Exosomes are single-membrane extracellular vesicles of endosomal origin, with an approximate radius of 20–200 nm. They are natural messengers that cells secrete to transport a wide range of diverse cargoes, including deoxyribonucleic acid, ribonucleic acid, proteins, and lipids between various cells. Some studies have reported that the components noted in exosomes in human breast milk could be transferred to the infant and cause epigenetic changes. Thus, it can affect gene expression and cellular event regulation in several tissues. Conclusions: In this manner, exosomes are associated with several pathways, including the immune system, oxidative stress, and cell cycle, and they can affect the short- and long-term health of infants. However, there is still much to learn about the functions, effectiveness, and certain impacts on the health of human breast milk exosomes.

Association Between Dairy Production System and Milk Functionality Based on Analysis of miRNAs in Exosomes from Milk.

Dairy farming practices significantly affect the nutritional and functional properties of milk. This study compares miRNAs in milk exosomes from extensive and intensive dairy systems and explores their potential implications for human consumers. Extensive systems are believed to produce milk of higher quality with better animal welfare compared to intensive systems. Milk samples from eight extensive and nine intensive dairy farms were analysed. Milk-derived extracellular vesicles were isolated through sequential ultracentrifugation and characterised through Dynamic Light Scattering and Nanosight to determine the size and the concentration of the extracellular vesicles, in addition to immunoblotting to ensure the presence of exosome-specific proteins in their membrane. miRNA levels were quantified using RT-qPCR, and metabolic pathways associated with miRNAs showing significant differences between farm groups were analysed. EVs from extensive farms had higher concentrations. Notably, bta-miR-451 levels were significantly higher in milk from extensive farms (p = 0.021). Like human miRNA hsa-miR-451, it is linked to pathways related to Parkinson's disease and cancer. Our research suggests that milk production in extensive systems not only provides socioeconomic and environmental benefits but may also have positive effects on human health. Further research is warranted to explore the bioactive potential of these miRNAs and their implications for human health.

Goat Milk Exosomes Ameliorate Ulcerative Colitis in Mice through Modulation of the Intestinal Barrier, Gut Microbiota, and Metabolites.

Goat milk is rich in a variety of nutrients that are important for intestinal health and disease prevention. However, the role of exosomes in goat milk remains to be elucidated. This study investigated for the first time the therapeutic efficacy and molecular underlying mechanisms of mature milk exosomes (M-exo) and goat colostrum exosomes (C-exo) on dextran sodium sulfate-induced ulcerative colitis (UC) in mice. The findings demonstrate that M-exo and C-exo significantly improved physiological indices, suppressed the secretion of proinflammatory cytokines, and diminished oxidative stress and apoptosis in UC mice. Moreover, C-exo and M-exo restored the intestinal barrier function, remodeled the gut microbiota, and improved metabolite composition in the feces of colitis mice. In conclusion, goat milk exosomes ameliorate UC in mice, which provides a basis for the development of functional food applications for the prevention and treatment of inflammatory bowel disease.

Pig Milk Exosome Packaging ssc-miR-22-3p Alleviates Pig Intestinal Epithelial Cell Injury and Inflammatory Response by Targeting MAPK14

Inflammatory diseases of the intestinal tract in piglets severely impair the economic performance of pig farms. Pig milk exosomes can encapsulate miRNAs which can then enter the piglet intestine to play an immunomodulatory role. Previously, we comparatively analyzed and identified exosomal miRNAs in the colostrum and mature milk of Bamei and Landrace pigs, and we screened for ssc-miR-22-3p, which is associated with inflammation and immune response; however, the role played by ssc-miR-22-3p in the immune response in IPEC-J2 cells is not yet clear. In this study, we first constructed a pig intestinal inflammatory response model using Lipopolysaccharide (LPS) and Polyinosinic-polycytidylic acid (Poly (I:C)), and we investigated the role of ssc-miR-22-3p targeting MAPK14 in the regulation of LPS and Poly (I:C)-induced inflammatory injury in IPEC-J2 cells by RT-qPCR, cell counting kit-8 (CCK-8), EdU staining, lactate dehydrogenase (LDH) activity assay, and dual luciferase reporter gene assay. We successfully established LPS and Poly (I:C)-induced cell damage models in IPEC-J2 cells. The immune response of IPEC-J2 cells was stimulated by induction of IPEC-J2 cells at 10 μg/mL LPS and 20 μg/mL Poly (I:C) for 24 h. Overexpression of ssc-miR-22-3p decreased cytokine expression and promoted cell viability and proliferation. The functional enrichment analysis revealed that ssc-miR-22-3p targets genes enriched in the pathways of negative regulation of inflammatory response and bacterial invasion of epithelial cells. The validity of the binding site of ssc-miR-22-3p to MAPK14 was tested by a dual luciferase reporter gene. Pig milk exosome ssc-miR-22-3p promotes cell viability and proliferation by targeting MAPK14, and it alleviates LPS and Poly (I:C)-induced inflammatory responses in IPEC-J2 cells.

Anti-inflammatory and Restorative effects of milk exosomes and Dexamethasone-Loaded exosomes in a corneal alkali burn model

Corneal alkali burn is a common and challenging ocular trauma, necessitating the use of dexamethasone (DXMS) as a therapeutic agent. However, prolonged and frequent administration of this drug can lead to undesirable side effects, limiting its clinical application. This study aimed to investigate the role and mechanism of action of exosomes as drug carriers in corneal alkali burn repair. We employed centrifugation to isolate milk exosomes (EXO) as nanocarriers. We observed that EXO enhanced the activity and migration of corneal epithelial cells, expediting the repair process following corneal injury. Additionally, a nano-drug delivery model (DXMS@EXO) was designed using ultrasound to load DXMS into exosomes, thus enabling targeted delivery to inflammatory cells and enhancing drug efficacy. DXMS@EXO inhibited the inflammatory processes in the corneal alkali burn model by modulating the classical Wnt signaling pathway, thereby promoting corneal re-epithelialization and wound healing and accelerating the repair process of corneal alkali burn. Neither EXO nor DXMS@EXO exhibited significant side effects during the course of treatment. This study highlighted the substantial potential of EXO and DXMS@EXO in improving drug efficacy and facilitating the repair of corneal alkali burn.

Sheep (Ovis aries) Milk Exosomal miRNAs Attenuate Dextran Sulfate Sodium-Induced Colitis in Mice via TLR4 and TRAF-1 Inhibition.

Mammalian milk exosomal miRNAs play an important role in maintaining intestinal immune homeostasis and protecting epithelial barrier function, but the specific miRNAs and whether miRNA-mediated mechanisms are responsible for these benefits remain a matter of investigation. This study isolated sheep milk-derived exosomes (sheep MDEs), identifying the enriched miRNAs in sheep MDEs, oar-miR-148a, and oar-let-7b as key components targeting TLR4 and TRAF1, which was validated by a dual-luciferase reporter assay. In dextran sulfate sodium-induced colitis mice, administration of sheep MDEs alleviated colitis symptoms, reduced colonic inflammation, and systemic oxidative stress, as well as significantly increased colonic oar-miR-148a and oar-let-7b while reducing toll-like receptor 4 (TLR4) and TNF-receptor-associated factor 1 (TRAF1) level. Further characterization in TNF-α-challenged Caco-2 cells showed that overexpression of these miRNAs suppressed the TLR4/TRAF1-IκBα-p65 pathway and reduced IL-6 and IL-12 production. These findings indicate that sheep MDEs exert gastrointestinal anti-inflammatory effects through the miRNA-mediated modulation of TLR4 and TRAF1, highlighting their potential in managing colitis.

Exosomal Delivery Enhances the Antiproliferative Effects of Acid-Hydrolyzed Apiaceae Spice Extracts in Breast Cancer Cells.

Breast cancer remains a leading cause of death worldwide. The Apiaceae plant family includes many culinary spices that have been shown to have medicinal properties. Many phytochemicals exhibit potent bioactivities but often suffer from poor uptake and oral bioavailability. Bovine milk and colostrum exosomes are a compelling drug delivery platform that could address this issue; these natural nanoparticles can be loaded with hydrophilic and lipophilic small molecules and biologics, resulting in lower doses needed to inhibit cancer growth. Ethanolic extracts of eight Apiaceae spices were examined for phytochemical content and antiproliferative potential. Acid hydrolysis (AH) was employed to remove glycosides, asses its impacts on extract efficacy, and evaluate its effects on exosome loading and subsequent formulation efficacy. Antiproliferative activity was assessed through MTT assays on T-47D, MDA-MB-231, and BT-474 breast cancer cells; all extracts exhibited broad antiproliferative activity. AH enhanced the bioactivity of cumin, caraway, and fennel in T-47D cells. Celery, cumin, anise, and ajwain showed the highest activity and were assayed in exosomal formulations, which resulted in reduced doses required to inhibit cellular proliferation for all extracts except AH-cumin. Apiaceae spice extracts demonstrated antiproliferative activities that can be improved with AH and further enhanced with exosomal delivery.

Chemo-Enzymatic Functionalization of Bovine Milk Exosomes with an EGFR Nanobody for Target-specific Drug Delivery.

Bovine milk exosomes (BmExo) have been identified as versatile nanovesicles for anti-cancer drugs delivery due to their natural availability and biocompatibility. However, tumor-specific delivery based on BmExo often requires post-isolation modifications of the membrane surface with active-targeting ligands. In this study, we report an alternative approach to functionalize BmExo with nanobody combining facile chemical modification and Sortase A-mediated site-specific ligation, as demonstrated by the development of an epidermal growth factor receptor (EGFR)-targeted drug delivery system. The BmExo membrane was first coated with a diglycine-containing amphiphile molecule, NH2-GG-PEG2000-DSPE, by hydrophobic insertion. The diglycine as nucleophiles displayed on the membrane enabled the subsequent ligation of the EGFR nanobody (7D12) by Sortase A (SrtA)-mediated site-specific transpeptidation. The successful construction of BmExo-7D12 was confirmed by Western blotting analysis, electron microscopy, and dynamic light scattering (DLS). As a demonstration model, BmExo-7D12 loaded with the chemotherapeutic drug doxorubicin (Dox) was shown to be able to deliver Dox to cancer cells in response to the expression of EGFR as manifested by immunocytochemistry and flow cytometry analysis. Finally, the cytotoxicity assay showed that BmExo-7D12-Dox was more effective in killing tumor cells with high EGFR expression while significantly reduced the non-specific toxicity to EGFR negative cells. In conclusion, these results demonstrate that 7D12-functionalized BmExo can serve as a target-specific delivery system for Dox to selectively kill EGFR-expressing tumor cells. This approach should prove to be versatile and efficient for the generation of protein-ligands modified BmExo.

Milk Exosome-Glow Nanosystem for Cancer Cellular and Tissue Bioimaging.

Milk-derived exosomes are widely used for diagnosis, delivery, imaging, and theranostic applications. Near-Infrared (NIR) based fluorescence bioimaging is an attractive and safer technique that is used for clinical applications. However, almost all NIR imaging agents tend to have poor photostability, short half-life, nonspecific protein binding, and concentration-dependent aggregation(s). Therefore, there is an unmet clinical need to develop newer and safer modalities to package and deliver NIR imaging agents. Bovine milk exosomes are natural, biocompatible, safe, and efficient nanocarriers that facilitate the delivery of micro- and macromolecules. Herein, we developed an exosome-based NIR dye loaded nanoimaging formulation that offers improved solubility and photostability of NIR dye. Following the acetic acid based extracellular vesicle (EV) treatment method, we extracted the bovine milk exosomes from a variety of pasteurized grade milk. The EVs were screened for their physicochemical properties such as particle size and concentration and zeta potential. The stability of these exosomes was also determined under different conditions, including storage temperatures, pH, and salt concentrations. Next, indocyanine green, a model NIR dye was loaded into these exosomes (Exo-Glow) via a sonication method and further assessed for their improved fluorescence intensity and photostability using an IVIS imaging system. Initial screening suggested that size of the selected bovine milk exosomes was ∼100-135 nm with an average particle concentration of 5.8 × 102 particles/mL. Exo-Glow further demonstrated higher fluorescence intensity in cancer cells and tissues when compared to free dye. These results showed that Exo-Glow has the potential to serve as a safer NIR imaging tool for cancer cells/tissues.

Loading milk exosomes with urolithins boosts their delivery to the brain: Comparing the activity of encapsulated vs. free urolithins in SH-SY5Y neuroblastoma cells

The gut microbial-derived metabolites of ellagitannins and ellagic acid, urolithins (Uros) are well-established anti-cancer metabolites according to preclinical studies. However, their efficacy is limited in systemic tissues, including the brain, by phase-II metabolism. Exosomes (EXOs), extracellular vesicles involved in cell signaling with the ability to cross the blood-brain barrier (BBB), could protect polyphenols from metabolism. Therefore, we loaded milk EXOs with Uro-A, Uro-B or IsoUro-A to evaluate their brain delivery and anticancer effects compared to non-encapsulated Uros. In Sprague-Dawley rats, perfused brain tissue analyses by UPLC-ESI-QTOF-MS showed higher Uro levels (∼3−4-fold) at 5 min following intravenous administration of EXO-Uros compared to non-encapsulated Uros, except for Uro-B, using similar Uro concentrations (17−30 μM). Experiments on neuroblastoma SH-SY5Y cells revealed dose-dependent antiproliferative effects for all EXO-Uros (0.3−1.2 μM), but not with non-encapsulated Uros (10 μM). Flow cytometry analyses indicated that only EXO-Uros arrested the cell cycle and induced apoptosis. Finally, using fluorescent-labeled EXOs and selective inhibitors, the primary endocytic pathway was revealed to be clathrin-dependent. Overall, encapsulating Uros into EXOs is promising for enhancing brain delivery and anticancer activity.

An Investigation of In Vitro Anti-Cancer Efficacy of Dihydroartemisinin-Loaded Bovine Milk Exosomes Against Triple-Negative Breast Cancer.

Repurposing drugs offers several advantages, including reduced time and cost compared to developing new drugs from scratch. It leverages existing knowledge about drug safety, dosage, and pharmacokinetics, expediting the process of clinical trials and regulatory approval. Dihydroartemisinin (DHA) is a semi-synthetic and active metabolite of all artemisinin molecules and is FDA-approved for the treatment of malaria. Apart from having anti-malarial properties, DHA also possesses anticancer properties. However, its pharmacological actions are limited by toxicity and solubility problems. To overcome these challenges and enhance its anticancer effectiveness, we designed an exosomal formulation of DHA. We isolated exosomes from bovine milk using differential ultracentrifugation and loaded DHA using sonication. Scanning and transition electron microscopy revealed a size of roughly 100nm, with a spherical shape. Furthermore, in pH 7.4 and 5.5, the exosomes exhibited burst release followed by sustained release. Multiple in vitro cell culture tests demonstrated that Exo-DHA exhibited enhanced anticancer activity, including cytotoxicity, cellular uptake, generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential, and inhibition of colony formation. Additional evidence supporting Exo-DHA's anti-migration ability came from transwell migration and scratch assays. Based on these results, it was concluded that the anticancer efficacy of DHA was improved when loaded into bovine milk-derived exosomes. While the in vitro results are encouraging, more in vivo testing in suitable animal models and biochemical marker analysis are warranted.

Milk Exosome-Liposome Hybrid Vesicles with Self-Adapting Surface Properties Overcome the Sequential Absorption Barriers for Oral Delivery of Peptides.

Milk exosomes (mExos) have demonstrated significant promise as vehicles for the oral administration of protein and peptide drugs owing to their superior capacity to traverse epithelial barriers. Nevertheless, certain challenges persist due to their intrinsic characteristics, including suboptimal drug loading efficiency, inadequate mucus penetration capability, and susceptibility to membrane protein loss. Herein, a hybrid vesicle with self-adaptive surface properties (mExos@DSPE-Hyd-PMPC) was designed by fusing functionalized liposomes with natural mExos, aiming to overcome the limitations associated with mExos and unlock their full potential in oral peptide delivery. The surface property transformation of mExos@DSPE-Hyd-PMPC was achieved by introducing a pH-sensitive hydrazone bond between the highly hydrophilic zwitterionic polymer and the phospholipids, utilizing the pH microenvironment on the jejunum surface. In comparison to natural mExos, hybrid vesicles exhibited a 2.4-fold enhancement in the encapsulation efficiency of the semaglutide (SET). The hydrophilic and neutrally charged surfaces of mExos@DSPE-Hyd-PMPC in the jejunal lumen exhibited improved preservation of membrane proteins and efficient traversal of the mucus barrier. Upon reaching the surface of jejunal epithelial cells, the highly retained membrane proteins and positively charged surfaces of the hybrid vesicle efficiently overcame the apical barrier, the intracellular transport barrier, and the basolateral exocytosis barrier. The self-adaptive surface properties of the hybrid vesicle resulted in an oral bioavailability of 8.7% and notably enhanced the pharmacological therapeutic effects. This study successfully addresses some limitations of natural mExos and holds promise for overcoming the sequential absorption barriers associated with the oral delivery of peptides.

Chitosan nanoparticles, camel milk exosomes and/or Sorafenib induce apoptosis, inhibit tumor cells migration and angiogenesis and ameliorate the associated liver damage in Ehrlich ascites carcinoma-bearing mice

BackgroundIt is crucial to improve cancer patients' quality of life by developing medications that can treat cancer with minimum adverse effects. This study aimed to evaluate the therapeutic effect of chitosan nanoparticles (CNPs) and camel milk exosomes (CMEs) alone or in combination with Sorafenib (SOR) on Ehrlich ascites carcinoma (EAC)-bearing mice and to assess whether EAC-associated liver injury would be ameliorated due to this combination. Liver function and oxidant/antioxidant status were determined spectrophotometrically, while the levels of inflammatory cytokines were estimated by enzyme-linked immunosorbent assay. Gene expression was detected using real-time polymerase chain reaction.ResultsThe tumor burden in EAC-bearing mice was reduced after treatment with CNPs ± CMEs ± SOR as indicated by (1) reduced ascetic fluid volume and tumor-cell viability; (2) induction of apoptosis [high p53, BCL2 associated X (Bax), caspase 3, low B-cell leukemia/lymphoma 2 protein (Bcl2)]; (3) increased intracellular reactive oxygen species; (4) decreased migration [high matrix metalloproteinase 9 (MMP9) and low TIMP metallopeptidase inhibitor 1 (TIMP1)]; (5) declined angiogenesis [low vascular endothelial growth factor (VEGF). These treatments also reduced liver injury induced by EAC as noticed by (1) restored liver function indices [alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), and albumin]; (2) restored redox balance [low malondialdehyde (MDA) levels and high superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities]; (3) increased antioxidant gene expression [high nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1)]; (4) declined inflammation [low interleukin-1β (IL1β) and tumor necrosis factor alpha (TNFα) levels), and (5) enhanced structure of liver. SOR + CNPs-treated mice showed the most improvement, followed by SOR + CMEs-treated animals.ConclusionsBased on these findings, we determined that CNPs and CMEs enhanced SOR's anticancer efficacy and had an ameliorative role against EAC-induced liver injuries.Graphic abstract

Nebulized milk exosomes loaded with siTGF-β1 ameliorate pulmonary fibrosis by inhibiting EMT pathway and enhancing collagen permeability

Pulmonary Fibrosis (PF) is a fatal disease in the interstitial lung associated with high mortality, morbidity, and poor prognosis. Transforming growth factor-β1 (TGF-β1) is a fibroblast-activating protein that promotes fibrous diseases. Herein, an inhalable system was first developed using milk exosomes (M-Exos) encapsulating siRNA against TGF-β1 (MsiTGF-β1), and their therapeutic potential for bleomycin (BLM)-induced PF was investigated. M-siTGF-β1 was introduced into the lungs of mice with PF through nebulization. The collagen penetration effect and lysosomal escape ability were verified in vitro. Inhaled MsiTGF-β1 notably alleviated inflammatory infiltration, attenuated extracellular matrix (ECM) deposition, and increased the survival rate of PF mice by 4.7-fold. M-siTGF-β1 protected lung tissue from BLM toxicity by efficiently delivering specific siRNA to the lungs, leading to TGF-β1 mRNA silencing and epithelial mesenchymal transition pathway inhibition. Therefore, M-siTGF-β1 offers a promising avenue for therapeutic intervention in fibrosis-related disorders.

Milk exosomes: Harnessing nature's duality for cancer therapy

AbstractMilk exosomes, extracellular nanovesicles that are naturally present in milk, have gained significant attention in cancer research for their potential to revolutionize cancer treatment strategies. They possess a specific set of characteristics that make them promising nanoscale vehicles for targeted drug delivery systems. Their inherent biocompatibility, coupled with their ability to effectively encapsulate and transport therapeutic agents directly into tumor cells, suggests the possibility of developing novel cancer therapies, potentially minimizing side effects associated with conventional therapies. However, recent studies have shown that milk exosomes have a dual nature, which is not entirely positive. Although they show potential in delivering anticancer therapeutics, evidence suggests they may also, under certain conditions, contribute to cancer progression. This paradoxical nature necessitates a better understanding of how they interact and work in different stages of cancer. Further investigation is crucial to understand the factors influencing their behaviour and to develop strategies that can maximize their therapeutic prospects while mitigating potential risks associated with their use in cancer treatment.

Large-Scale Isolation of Milk Exosomes for Skincare.

Exosomes are small membrane vesicles in a cell culture. They are secreted by most cells and originate from the endosomal pathway. A variety of proteins, lipids, and genetic materials have been shown to be carried by exosomes. Once taken up by neighboring or distant cells, the bioactive compounds in exosomes can regulate the condition of recipient cells. Typically, producing exosomes in large quantities requires cell culture, resulting in high production costs. However, exosomes are abundant in milk and can be isolated on a large scale at a low cost. In our study, we found that milk exosomes can promote the synthesis and reconstruction of stratum corneum lipids, enhance skin barrier function, and provide greater protection for the skin. Furthermore, milk exosomes have anti-inflammatory properties that can reduce skin irritation, redness, and other symptoms, giving immediate relief. They also exhibit antioxidant activity, which helps neutralize free radicals and slows down the skin aging process. Additionally, milk exosomes inhibit melanin production, aiding in skin whitening. Ongoing research has uncovered the benefits of milk exosomes for skin improvement and their application in cosmetics, skin healthcare, and other fields, and these applications are continuing to expand.

Isolation and Characterization of Milk Exosomes for Use in Advanced Therapies.

Exosomes are cell-derived extracellular vesicles (EVs) with diameters between 30 and 120 nm. In recent years, several studies have evaluated the therapeutic potential of exosomes derived from different fluids due to their low immunogenicity and high biocompatibility. However, producing exosomes on a large scale is still challenging. One of the fluids from which they could be isolated in large quantities is milk. Moreover, regeneration is a well-known property of milk. The present work seeks to optimize a method for isolating exosomes from bovine and human milk, comparing different storage conditions and different extraction protocols. We found differences in the yield extraction associated with pre-storage milk conditions and observed some differences according to the processing agent. When we removed milk fat globules and added rennet before freezing, we obtained a cleaner final fraction. In summary, we attempted to optimize a rennet-based new milk-exosome isolation method and concluded that pre-treatment, followed by freezing of samples, yielded the best exosome population.

Targeting delivery of miR-146a via IMTP modified milk exosomes exerted cardioprotective effects by inhibiting NF-κB signaling pathway after myocardial ischemia-reperfusion injury

Reperfusion therapy is critical for saving heart muscle after myocardial infarction, but the process of restoring blood flow can itself exacerbate injury to the myocardium. This phenomenon is known as myocardial ischemia-reperfusion injury (MIRI), which includes oxidative stress, inflammation, and further cell death. microRNA-146a (miR-146a) is known to play a significant role in regulating the immune response and inflammation, and has been studied for its potential impact on the improvement of heart function after myocardial injury. However, the delivery of miR-146a to the heart in a specific and efficient manner remains a challenge as extracellular RNAs are unstable and rapidly degraded. Milk exosomes (MEs) have been proposed as ideal delivery platform for miRNA-based therapy as they can protect miRNAs from RNase degradation. In this study, the effects of miR-146a containing MEs (MEs-miR-146a) on improvement of cardiac function were examined in a rat model of MIRI. To enhance the targeting delivery of MEs-miR-146a to the site of myocardial injury, the ischemic myocardium-targeted peptide IMTP was modified onto the surfaces, and whether the modified MEs-miR-146a could exert a better therapeutic role was examined by echocardiography, myocardial injury indicators and the levels of inflammatory factors. Furthermore, the expressions of miR-146a mediated NF-κB signaling pathway-related proteins were detected by western blotting and qRT-PCR to further elucidate its mechanisms. MiR-146 mimics were successfully loaded into the MEs by electroporation at a square wave 1000 V voltage and 0.1 ms pulse duration. MEs-miR-146a can be up-taken by cardiomyocytes and protected the cells from oxygen glucose deprivation/reperfusion induced damage in vitro. Oral administration of MEs-miR-146a decreased myocardial tissue apoptosis and the expression of inflammatory factors and improved cardiac function after MIRI. The miR-146a level in myocardium tissues was significantly increased after the administration IMTP modified MEs-miR-146a, which was higher than that of the MEs-miR-146a group. In addition, intravenous injection of IMTP modified MEs-miR-146a enhanced the targeting to heart, improved cardiac function, reduced myocardial tissue apoptosis and suppressed inflammation after MIRI, which was more effective than the MEs-miR-146a treatment. Moreover, IMTP modified MEs-miR-146a reduced the protein levels of IRAK1, TRAF6 and p-p65. Therefore, IMTP modified MEs-miR-146a exerted their anti-inflammatory effect by inhibiting the IRAK1/TRAF6/NF-κB signaling pathway. Taken together, our findings suggested miR-146a containing MEs may be a promising strategy for the treatment of MIRI with better outcome after modification with ischemic myocardium-targeted peptide, which was expected to be applied in clinical practice in future.

Shape Engineering of Exosomes for Endoplasmic Reticulum-Targeted Delivery and Amplified Anticancer Efficacy

Exosomes are nanoscale extracellular vesicles that have become pivotal in advancing targeted drug delivery strategies for cancer therapy. In this study, we conducted a comparative analysis of the intracellular targeting capabilities of differently shaped exosomes, including milk exosome nanorods (MR), ginger exosome nanorods (GR), and cancer cell exosome nanorods (HR), compared to their spherical counterparts. Our observations revealed that exosome nanorods demonstrated effective and sustained targeting of the endoplasmic reticulum (ER) within cancer cells, while exosome nanospheres were captured within lysosomes. Building on this principle, we chose milk-derived exosomes (mExo) for the in vivo research, engineering the surface of MR with folate to enhance their tumor-targeting efficacy. We demonstrated the effective accumulation of these folate-modified MR (FMR) around the ER in cancer cells, as validated in both orthotopic colorectal cancer (CRC) tissues and human CRC biopsy samples. Furthermore, when loaded with curcumin (Cur), the FMR@Cur exhibited remarkable efficacy in suppressing tumors in orthotopic CRC mouse models. This effect is attributed to the targeted delivery of FMR@Cur to the ER, leading to enhanced ER-stress induced apoptosis. Overall, our study underscores the pivotal role of shape engineering in exosome-mediated drug delivery, offering novel insights and paving the way for innovative strategies to enhance the precision of intracellular drug targeting in cancer therapy.