Impact of e-cigarette aerosol on primary human alveolar epithelial type 2 cells.

Electronic cigarettes (e-cigarettes) are designed to simulate combustible cigarette smoking and to aid in smoking cessation. Although the number of e-cigarette users has been increasing, the potential health impacts and biological effects of e-cigarettes are still not fully understood. Previous research has focused on the biological effects of e-cigarettes on lung cancer cell lines and distal airway epithelial cells; however, there have been few published studies on the effect of e-cigarettes on primary lung alveolar epithelial cells. The primary purpose of this study was to investigate the direct effect of e-cigarette aerosol on primary human lung alveolar epithelial type 2 (AT2) cells, both alone and in the presence of viral infection. The Melo-3 atomizer caused direct AT2 cell toxicity, whereas the more popular Juul pod's aerosol did not have a detectable cytotoxic effect on AT2 cells. Juul nicotine aerosol also did not increase short-term susceptibility to viral infection. However, 3 days of exposure upregulated genes central to the generation of reactive oxygen species, lipid peroxidation, and carcinogen metabolism and downregulated key innate immune system genes related to cytokine and chemokine signaling. These findings have implications for the potentially injurious impact of long-term use of popular low-power e-cigarette pods on the human alveolar epithelium. Gene expression data might be an important endpoint for evaluating the potential harmful effects of vaping devices that do not cause overt toxicity.

Mesenchymal stem cell extracellular vesicles mitigate vascular permeability and injury in the small intestine and lung in a mouse model of hemorrhagic shock and trauma.

BACKGROUND: Hemorrhagic shock and trauma (HS/T)-induced gut injury may play a critical role in the development of multi-organ failure. Novel therapies that target gut injury and vascular permeability early after HS/T could have substantial impacts on trauma patients. In this study, we investigate the therapeutic potential of human mesenchymal stem cells (MSCs) and MSC-derived extracellular vesicles (MSC EVs) in vivo in HS/T in mice and in vitro in Caco-2 human intestinal epithelial cells. METHODS: In vivo, using a mouse model of HS/T, vascular permeability to a 10-kDa dextran dye and histopathologic injury in the small intestine and lungs were measured among mice. Groups were (1) sham, (2) HS/T + lactated Ringer's (LR), (3) HS/T + MSCs, and (4) HS/T + MSC EVs. In vitro, Caco-2 cell monolayer integrity was evaluated by an epithelial cell impedance assay. Caco-2 cells were pretreated with control media, MSC conditioned media (CM), or MSC EVs, then challenged with hydrogen peroxide (H2O2). RESULTS: In vivo, both MSCs and MSC EVs significantly reduced vascular permeability in the small intestine (fluorescence units: sham, 456 ± 88; LR, 1067 ± 295; MSC, 765 ± 258; MSC EV, 715 ± 200) and lung (sham, 297 ± 155; LR, 791 ± 331; MSC, 331 ± 172; MSC EV, 303 ± 88). Histopathologic injury in the small intestine and lung was also attenuated by MSCs and MSC EVs. In vitro, MSC CM but not MSC EVs attenuated the increased permeability among Caco-2 cell monolayers challenged with H2O2. CONCLUSION: Mesenchymal stem cell EVs recapitulate the effects of MSCs in reducing vascular permeability and injury in the small intestine and lungs in vivo, suggesting MSC EVs may be a potential cell-free therapy targeting multi-organ dysfunction in HS/T. This is the first study to demonstrate that MSC EVs improve both gut and lung injury in an animal model of HS/T.

Cryoprecipitate attenuates the endotheliopathy of trauma in mice subjected to hemorrhagic shock and trauma.

BACKGROUND: Plasma has been shown to mitigate the endotheliopathy of trauma. Protection of the endothelium may be due in part to fibrinogen and other plasma-derived proteins found in cryoprecipitate; however, the exact mechanisms remain unknown. Clinical trials are underway investigating early cryoprecipitate administration in trauma. In this study, we hypothesize that cryoprecipitate will inhibit endothelial cell (EC) permeability in vitro and will replicate the ability of plasma to attenuate pulmonary vascular permeability and inflammation induced by hemorrhagic shock and trauma (HS/T) in mice. METHODS: In vitro, barrier permeability of ECs subjected to thrombin challenge was measured by transendothelial electrical resistance. In vivo, using an established mouse model of HS/T, we compared pulmonary vascular permeability among mice resuscitated with (1) lactated Ringer's solution (LR), (2) fresh frozen plasma (FFP), or (3) cryoprecipitate. Lung tissue from the mice in all groups was analyzed for markers of vascular integrity, inflammation, and inflammatory gene expression via NanoString messenger RNA quantification. RESULTS: Cryoprecipitate attenuates EC permeability and EC junctional compromise induced by thrombin in vitro in a dose-dependent fashion. In vivo, resuscitation of HS/T mice with either FFP or cryoprecipitate attenuates pulmonary vascular permeability (sham, 297 ± 155; LR, 848 ± 331; FFP, 379 ± 275; cryoprecipitate, 405 ± 207; p < 0.01, sham vs. LR; p < 0.01, LR vs. FFP; and p < 0.05, LR vs. cryoprecipitate). Lungs from cryoprecipitate- and FFP-treated mice demonstrate decreased lung injury, decreased infiltration of neutrophils and activation of macrophages, and preserved pericyte-endothelial interaction compared with LR-treated mice. Gene analysis of lung tissue from cryoprecipitate- and FFP-treated mice demonstrates decreased inflammatory gene expression, in particular, IL-1ß and NLRP3, compared with LR-treated mice. CONCLUSION: Our data suggest that cryoprecipitate attenuates the endotheliopathy of trauma in HS/T similar to FFP. Further investigation is warranted on active components and their mechanisms of action.

Freeze-dried platelets promote clot formation, attenuate endothelial cell permeability, and decrease pulmonary vascular leak in a murine model of hemorrhagic shock.

BACKGROUND: Hemorrhagic shock (HS) and trauma induce endothelial barrier compromise, inflammation, and aberrant clotting. We have shown that fresh human platelets (Plts) and Plt extracellular vesicles mitigate vascular leak in murine models of injury. Here, we investigate the potential of freeze-dried platelets (FDPlts) to attenuate pulmonary vascular permeability, decrease inflammation, and promote clotting in a murine model of HS. METHODS: Human FDPlts were characterized using in vitro assays of Plt marker expression, aggregation, coagulation, and endothelial cell permeability. An intravital model of vascular injury in the mouse cremaster muscle was used to assess the ability of FDPlts to incorporate into clots. Mouse groups subjected to controlled hemorrhage for 90 minutes were (1) lactated Ringer solution (LR), (2) FDPlts, (3) fresh human Plts, (4) murine whole blood (WB), and (5) shams (only instrumented). Hemorrhagic shock mouse endpoints included coagulation, pulmonary vascular permeability, and lung injury. RESULTS: Freeze-dried Plts expressed Plt-specific markers and retained functionality similar to fresh Plts. In in vitro assays of Plt aggregation, differences were noted. In vivo, FDPlts and Plts were found to incorporate into clots in postcapillary venules in the mouse cremaster muscle. Hemorrhagic shock mice resuscitated with LR displayed increased pulmonary vascular permeability compared with sham (sham, 686.6 ± 359.7; shock-LR, 2,637 ± 954.7; p = 0.001), and treatment with FDPlts or WB attenuated permeability compared with shock: shock-FDPlts, 1,328 ± 462.6 (p = 0.05), and shock-WB, 1,024 ± 370.5 (p = 0.0108). However, human Plts (Days 1-3) did not attenuate vascular leak in HS mice compared with shock-LR (shock-Plts, 3,601 ± 1,581; p = 0.33). CONCLUSION: FDPlts contribute to clot formation similar to fresh human Plts. FDPlts also attenuated vascular permeability in vitro and in vivo. Mouse WB resuscitation but not fresh human Plts attenuated vascular permeability after HS. These data suggest that the effect of FDPlts may be a suitable alternative to fresh Plts in modulating hemostasis and the endotheliopathy associated with injury.