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Abstract Chinese hamster ovary (CHO) cells release and exchange large quantities of extracellular vesicles (EVs). EVs are highly enriched in microRNAs (miRs, or miRNAs), which are responsible for most of their biological effects. We have recently shown that the miR content of CHO EVs varies significantly under culture stress conditions. Here, we provide a novel stoichiometric (“per‐EV”) quantification of miR and protein levels in large CHO EVs produced under ammonia, lactate, osmotic, and age‐related stress. Each stress resulted in distinct EV miR levels, with selective miR loading by parent cells. Our data provide a proof of concept for the use of CHO EV cargo as a diagnostic tool for identifying culture stress. We also tested the impact of three select miRs (let‐7a, miR‐21, and miR‐92a) on CHO cell growth and viability. Let‐7a—abundant in CHO EVs from stressed cultures—reduced CHO cell viability, while miR‐92a—abundant in CHO EVs from unstressed cultures—promoted cell survival. Overexpression of miR‐21 had a slight detrimental impact on CHO cell growth and viability during late exponential‐phase culture, an unexpected result based on the reported antiapoptotic role of miR‐21 in other mammalian cell lines. These findings provide novel relationships between CHO EV cargo and cell phenotype, suggesting that CHO EVs may exert both pro‐ and antiapoptotic effects on target cells, depending on the conditions under which they were produced. 

Abstract Megakaryocytic extracellular vesicles (MkEVs) promote the growth and megakaryopoiesis of hematopoietic stem and progenitor cells (HSPCs) largely through endogenous miR‐486‐5p and miR‐22‐3p cargo. Here, we examine the impact of biomechanical force and culture age/differentiation on the formation, properties, and biological efficacy of MkEVs. We applied biomechanical force to Mks using two methods: shake flask cultures and a syringe pump system. Force increased MkEV production in a magnitude‐dependent manner, with similar trends emerging regardless of whether flow cytometry or nanoparticle tracking analysis was used for MkEV counting. Both methods produced MkEVs that were relatively depleted of miR‐486‐5p and miR‐22‐3p cargo. However, while the shake flask‐derived MkEVs were correspondingly less effective in promoting megakaryocytic differentiation of HSPCs, the syringe pump‐derived MkEVs weremoreeffective in doing so, suggesting the presence of unique, unidentified miRNA cargo components. Higher numbers of MkEVs were also produced by “older” Mk cultures, though miRNA cargo levels and MkEV bioactivity were unaffected by culture age. A reduction in MkEV production by Mks derived from late‐differentiating HSPCs was also noted. Taken together, our results demonstrate that biomechanical force has an underappreciated and deeply influential role in MkEV biology, though that role may vary significantly depending on the nature of the force. Given the ubiquity of biomechanical force in vivo and in biomanufacturing, this phenomenon must be grappled with before MkEVs can attain clinical relevance. 

Abstract. The IceCube Neutrino Observatory instruments about 1 km3 of deep, glacial ice at the geographic South Pole. It uses 5160 photomultipliers to detect Cherenkov light emitted by charged relativistic particles. An unexpected light propagation effect observed by the experiment is an anisotropic attenuation, which is aligned with the local flow direction of the ice. We examine birefringent light propagation through the polycrystalline ice microstructure as a possible explanation for this effect. The predictions of a first-principles model developed for this purpose, in particular curved light trajectories resulting from asymmetric diffusion, provide a qualitatively good match to the main features of the data. This in turn allows us to deduce ice crystal properties. Since the wavelength of the detected light is short compared to the crystal size, these crystal properties include not only the crystal orientation fabric, but also the average crystal size and shape, as a function of depth. By adding small empirical corrections to this first-principles model, a quantitatively accurate description of the optical properties of the IceCube glacial ice is obtained. In this paper, we present the experimental signature of ice optical anisotropy observed in IceCube light-emitting diode (LED) calibration data, the theory and parameterization of the birefringence effect, the fitting procedures of these parameterizations to experimental data, and the inferred crystal properties.