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. 2023 Aug 1;135(2):271-278.
doi: 10.1152/japplphysiol.00243.2023. Epub 2023 Jun 22.

Endothelial-derived extracellular vesicles associated with electronic cigarette use impair cerebral microvascular cell function

Hannah L Cardenas  1 , Nicholas G Evanoff  2 , Hannah K Fandl  1 , Auburn R Berry  1 , Kendra N Wegerson  1 , Emily I Ostrander  1 , Jared J Greiner  1 , Sheena R Dufresne  3 , Michael Kotlyar  3 , Donald R Dengel  2 , Christopher A DeSouza  1 , Vinicius P Garcia  1
Affiliations

Endothelial-derived extracellular vesicles associated with electronic cigarette use impair cerebral microvascular cell function

Hannah L Cardenas et al. J Appl Physiol (1985). .

Abstract

The aim of this study was to determine the effect of circulating endothelial cell-derived microvesicles (EMVs) isolated from e-cigarette users on human cerebral microvascular endothelial cells (hCMECs) nitric oxide (NO) and endothelin (ET)-1 production and tissue-type plasminogen activator (t-PA) release. Circulating EMVs (CD144-PE) were isolated (flow cytometry) from 27 young adults (19-25 yr): 10 nonsmokers (6 M/4 F), 10 e-cigarette users (6 M/4 F), and 7 tobacco cigarette smokers (4 M/3 F). hCMECs were cultured and treated with isolated EMVs for 24 h. EMVs from e-cigarette users and cigarette smokers induced significantly higher expression of p-eNOS (Thr495; 28.4 ± 4.6 vs. 29.1 ± 2.8 vs. 22.9 ± 3.8 AU), Big ET-1 (138.8 ± 19.0 vs. 141.7 ± 19.1 vs. 90.3 ± 18.8 AU) and endothelin converting enzyme (107.6 ± 10.1 and 113.5 ± 11.8 vs. 86.5 ± 13.2 AU), and significantly lower expression of p-eNOS (Ser1177; 7.4 ± 1.7 vs. 6.5 ± 0.5 vs. 9.7 ± 1.6 AU) in hCMECs than EMVs from nonsmokers. NO production was significantly lower and ET-1 production was significantly higher in hCMECs treated with EMVs from e-cigarette (5.7 ± 0.8 µmol/L; 33.1 ± 2.9 pg/mL) and cigarette smokers (6.3 ± 0.7 µmol/L; 32.1 ± 3.9 pg/mL) than EMVs from nonsmokers (7.6 ± 1.2 µmol/L; 27.9 ± 3.1 pg/mL). t-PA release in response to thrombin was significantly lower in hCMECs treated with EMVs from e-cigarette users (from 38.8 ± 6.3 to 37.4 ± 8.3 pg/mL) and cigarette smokers (31.5 ± 5.5 to 34.6 ± 8.4 pg/mL) than EMVs from nonsmokers (38.9 ± 4.3 to 48.4 ± 7.9 pg/mL). There were no significant differences in NO, ET-1, or t-PA protein expression or production in hCMECs treated with EMVs from e-cigarette users and smokers. Circulating EMVs associated with e-cigarette use adversely affects brain microvascular endothelial cells and may contribute to reported cerebrovascular dysfunction with e-cigarette use.NEW & NOTEWORTHY In the present study, we determined the effect of circulating endothelial cell-derived microvesicles (EMVs) isolated from e-cigarette users on human cerebral microvascular endothelial cells (hCMECs) nitric oxide (NO) and endothelin (ET)-1 production and tissue-type plasminogen activator (t-PA) release. EMVs from e-cigarette users reduced brain microvascular endothelial cell NO production, enhanced ET-1 production, and impaired endothelial t-PA release. EMVs are a potential mediating factor in the increased risk of stroke associated with e-cigarette use.

Keywords: e-cigarette; endothelial cells; endothelial microvesicles; smoking.

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Conflict of interest statement

No conflicts of interest, financial or otherwise, are declared by the authors.

Christopher DeSouza was an editor of Journal of Applied Physiology and was not involved and did not have access to information regarding the peer-review process or final disposition of this article. An alternate editor oversaw the peer-review and decision-making process for this article.

Figures

None
Graphical abstract
Figure 1.
Figure 1.
Expression of e-NOS (A), p-eNOS (Ser1177; B), and p-eNOS (Thr495; C) as well as NO production (D) in human cerebral microvascular endothelial cells treated with EMVs from nonsmokers, e-cigarette users, and cigarette smokers. Relation between p-eNOS (Thr495) expression and NO production (E) across samples. Mean value for each group is denoted. AU indicates arbitrary units. *P < 0.05. EMVs, endothelial cell-derived microvesicles; NO, nitric oxide.
Figure 2.
Figure 2.
Expression of big ET-1 (A) and ECE (B) as well as ET-1 production (C) in human cerebral microvascular endothelial cells treated with EMVs from nonsmokers, e-cigarette users, and cigarette smokers. Relation between big ET-1 (D) and ECE (E) expression and ET-1 production across samples. Mean value for each group is denoted. *P < 0.05. AU, arbitrary units; ECE, endothelin converting enzyme; EMVs, endothelial cell-derived microvesicles; ET, endothelin.
Figure 3.
Figure 3.
Representative protein histograms of immunodetection by capillary electrophoresis immunoassay for e-NOS (A), p-eNOS (Ser1177; B), p-eNOS (Thr495; C), big ET-1 (D), ECE (E), and t-PA (F). ECE, endothelin converting enzyme; e-NOS, endothelial nitric oxide synthase; ET, endothelin; t-PA, tissue-type plasminogen activator.
Figure 4.
Figure 4.
Expression of t-PA (A), basal t-PA release (B), and t-PA release in response to thrombin (C) in human cerebral microvascular endothelial cells treated with EMVs from nonsmokers, e-cigarette users, and cigarette smokers. Mean value for each group is denoted. *P < 0.05. AU, arbitrary units; EMVs, endothelial cell-derived microvesicles; t-PA, tissue-type plasminogen activator.
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