SARS‐CoV‐2 Spike Protein Exacerbates Endothelial Dysfunction and Promotes Lung Injury in Aging Type 2 Diabetes Mellitus Cell Culture Model

ABSTRACT Advanced age and diabetes mellitus (DM) are major risk factors for severe COVID‐19, associated with acute respiratory distress syndrome (ARDS), cytokine storm, and increased mortality. SARS‐CoV‐2 spike protein (SP) binding to angiotensin‐converting enzyme 2 (ACE2) potentially disrupts the renin–angiotensin system (RAS), promoting angiotensin II (Ang II) accumulation and vascular dysfunction. In individuals with preexisting endothelial dysfunction (ED), such as older adults with type 2 DM (T2DM), COVID‐19 may exacerbate inflammation, vascular permeability, and tissue damage. This study aimed to elucidate the mechanisms by which SP exacerbates diabetes‐associated ED using an in vitro aortic endothelial–bronchial epithelial co‐culture model. Serum from healthy adults and T2DM patients was used to stimulate human aortic endothelial cells (HAoECs), with or without SP. ED was assessed via reactive oxygen species (ROS), intercellular adhesion molecule‐1 (ICAM‐1) and vascular cell adhesion molecule‐1 (VCAM‐1), and endothelial permeability. ACE2 expression and Ang II levels were measured in ACE2‐transfected human bronchial epithelial cells (BEAS‐2B ACE2‐eGFP ). Combined effects on lung injury and cytokines (CXCL8, IL‐1β, and IL‐6) were evaluated using a co‐culture system. T2DM serum significantly increased ROS (609.33 ± 108.23%; p = 0.0016), ICAM‐1 and VCAM‐1 expression (68.55 ± 5.47 copies/μL; p = 0.0011 and 84.77 ± 3.81 copies/μL; p < 0.0001, respectively), and permeability (3.86 × 10 7 ± 4.8 × 10 6 FU; p = 0.0119) in HAoECs. In BEAS‐2B ACE2‐eGFP , T2DM serum significantly increased ACE2 expression (5.45 ± 1.99 copies/μL; p = 0.0089), whereas co‐exposure to T2DM + SP markedly reduced ACE2 (0.97 ± 0.29 copies/μL; p = 0.0086) and elevated Ang II (111.60 ± 15.81 pg./mL; p = 0.1343). Co‐treatment with T2DM + SP further exacerbated ED, driving heightened inflammation, lung epithelial injury, and cytokine release. Diabetes‐associated ED synergizes with SP to aggravate vascular inflammation and lung injury, identifying adhesion molecules and pro‐inflammatory cytokines as potential biomarkers, and RAS dysregulation and endothelial activation as therapeutic targets for severe COVID‐19 in T2DM.