Milk extracellular vesicles: A burgeoning new presence in nutraceuticals and drug delivery

Abstract

Mammalian milk, a multifaceted developmental biofluid, has attracted new attention due to its diverse constituents and their implications for health and disease. Among these constituents, extracellular vesicles (EVs) have emerged as focal points of investigation. EVs, including exosomes and small EVs, have demonstrated biological activity in preclinical studies-including reports of enhancement of cognition and neural complexity, promotion of gastrointestinal development, barrier function and microbiome richness, the bolstering of immune response, and facilitation of musculoskeletal maturation in neonates. The richness of milk as a source of EVs is noteworthy, with hundreds of milliliters (at >10¹² EVs/mL) of nanovesicles extractable from a single liter of milk (>10¹⁴ EVs/starting liter of milk). Techniques such as tangential flow filtration hold promise for scalable production, potentially extending to thousands of liters. Together with the scale and increasing sophistication of the dairy industry, the abundance of EVs in milk underscores their commercial potential in various nutraceutical applications. Beyond natural bioactivity, milk EVs (mEVs) present intriguing possibilities as orally deliverable, non-immunogenic pharmaceutical carriers, with burgeoning interest in their utilization for heart disease and cancer chemotherapy and as vectors for gene-editing modules such as CrispR. This review synthesizes current knowledge on mEV biogenesis, characterization, isolation methodologies, and cargo contents. Moreover, it delves into the therapeutic potential of mEVs, both as inherently bioactive nanovesicles and as versatile platforms for drug delivery. As efforts progress toward large-scale implementation, rigorous attention to safe, industrial-scale production and robust assay development will be pivotal in harnessing the translational promise of small EVs from milk.

Keywords: drug delivery; exosome; extracellular vesicle; infant development; milk; nutraceutical; pharmaceutical.

FIGURE 1

Overview of extracellular vesicle (EV) biogenesis (left), composition (center), and uptake (right). Created with Biorender.com.

FIGURE 2

Proteomic, lipidomic, and transcriptomic composition of small milk extracellular vesicles. Created with BioRender.com.

FIGURE 3

Representative transmission electron microscopy image of milk extracellular vesicles with high magnification inset (left) and Calcein‐AM‐stained milk extracellular vesicles imaged by confocal microscopy (right) repurposed with permission from authors.

FIGURE 4

Overview of loading methods leveraged to date for mEVs. Electroporation and co‐incubation, , have shown the greatest loading ceiling (40%–60%), with notable floors of 4%. Chemical transfection has shown a consistent result of 30%, while other approaches, including sonication and saponification, have shown low efficacy. Combination approaches such as extrusion + sonication have shown improvements over sonication alone, but with notable limits on efficacy. See Table 2 for an itemized breakdown of loading efficiencies and cargoes utilized, as well as the specific citations associated with the methods summarized.