Role of Pdk4 in BAPN-induecd aortic dissection in mouse models

Thoracic aortic dissection (TAD) is a life-threatening acute vascular condition with extremely high morbidity and mortality. Endothelial cells (ECs) are critical for maintaining vascular homeostasis, yet the role of Endothelial-to-Mesenchymal Transition (EndoMT)—a key cell fate process in vascular development and disease—in TAD remains poorly defined. Furthermore, the functional significance of EndoMT-gene-program activation mediated by pyruvate dehydrogenase kinase 4 (PDK4) in TAD has not been elucidated. To explore the mechanistic involvement of EndoMT in TAD, a murine TAD model was established through systemic administration of β-aminopropionitrile (BAPN). RNA sequencing was performed to profile the activation landscape of the EndoMT-gene-program in ECs for identifying key regulatory targets. Concurrently, clinical samples from TAD patients and healthy controls were analyzed to quantify mRNA and protein levels of EndoMT-related markers in aortic tissues. Pyruvate dehydrogenase kinase isoenzyme 4 (PDK4) was identified as a critical regulator and validated as a downstream target of EndoMT signaling using an integrated approach combining RNA-seq, luciferase reporter assays, and immunofluorescent staining. The functional role of Pdk4 in TAD was further validated by using conditional knockout (CKO) mouse model. To facilitate better clinical translation, EC-specific Pdk4 deletion or overexpression mouse models were generated via adeno-associated virus (AAV)-mediated delivery and subjected to TAD model. Serum samples from these mice were collected, and enzyme-linked immunosorbent assay (ELISA) was used to measure levels of endothelial injury markers (sICAM1, sVCAM1, sE-Selectin) for assessing endothelial dysfunction. Additionally, the therapeutic potential of dichloroacetate (DCA), a specific PDK4 inhibitor, was evaluated by administering it to BAPN-fed mice to assess its efficacy in ameliorating TAD progression. A significant upregulation of the EndoMT-gene-program was observed in BAPN-fed murine models of TAD, aortic tissues from TAD patients, and HAECs treated with angiotensin II (Ang II). Administration of AAVEndoPdk4 directly induced TAD formation, which was exacerbated by BAPN co-treatment. Conversely, EC-specific knockdown of Pdk4 reduced TAD formation by inhibiting EndoMT signaling. The same results were also obtained in EC-specific conditional Pdk4 knockout mice (CKO, Pdk4flox/flox; Cdh5-CreERT2). PDK4 was identified as a critical factor in both TAD development and EndoMT process, with SNAI1 transcriptionally activating PDK4; vice versa, PDK4 further enhanced the activation of EndoMT signaling in both in vivo and vitro. PDK4 inhibitor, DCA, significantly slowed TAD progression in BAPN-fed mice. Mechanistically, a PDK4-EndoMT feedback loop was identified in TAD, where interruption of this self-reinforcing pathological cycle markedly attenuates the progression of TAD.