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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 Platelet transfusions are used to treat idiopathic or drug-induced thrombocytopenia. Platelets are an expensive product in limited supply, with limited storage and distribution capabilities because they cannot be frozen. We have demonstrated that, in vitro, human megakaryocytic microparticles (huMkMPs) target human CD34+ hematopoietic stem and progenitor cells (huHSPCs) and induce their Mk differentiation and platelet biogenesis in the absence of thrombopoietin. In this study, we showed that, in vitro, huMkMPs can also target murine HSPCs (muHSPCs) to induce them to differentiate into megakaryocytes in the absence of thrombopoietin. Based on that, using wild-type BALB/c mice, we demonstrated that intravenously administering 2 × 106 huMkMPs triggered de novo murine platelet biogenesis to increase platelet levels up to 49% 16 hours after administration. huMkMPs also largely rescued low platelet levels in mice with induced thrombocytopenia 16 hours after administration by increasing platelet counts by 51%, compared with platelet counts in thrombocytopenic mice. Normalized on a tissue-mass basis, biodistribution experiments show that MkMPs localized largely to the bone marrow, lungs, and liver 24 hours after huMkMP administration. Beyond the bone marrow, CD41+ (megakaryocytes and Mk-progenitor) cells were frequent in lungs, spleen, and especially, liver. In the liver, infused huMKMPs colocalized with Mk progenitors and muHSPCs, thus suggesting that huMkMPs interact with muHSPCs in vivo to induce platelet biogenesis. Our data demonstrate the potential of huMkMPs, which can be stored frozen, to treat thrombocytopenias and serve as effective carriers for in vivo, target-specific cargo delivery to HSPCs.