Hypothesis

Age‑related decline in tropoelastin synthesis in the abdominal infrarenal aorta increases matrix stiffness sensed by vascular smooth muscle cells, driving nuclear translocation of YAP/TAZ and promoting an osteogenic phenotypic switch that leads to medial calcification. This mechanism explains regional susceptibility to stiffness and calcification beyond passive collagen accumulation.

Mechanistic Basis

• It's known that elastin fragments normally engage integrin‑αvβ3 to suppress FAK‑Src signaling; loss of tropoelastin reduces this brake, elevating focal adhesion kinase activity.
• heightened FAK‑Src activates the Hippo pathway antagonists LATS1/2, allowing YAP/TAZ to accumulate in the nucleus.
• Nuclear YAP/TAZ complexes with TEAD transcription factors to upregulate Runx2, Osterix and BMP2, orchestrating VSMC‑derived hydroxyapatite deposition.
• Low nitric oxide in the infrarenal aorta amplifies ROS‑mediated inhibition of phosphatases, further sustaining FAK‑Src activity.
• Thus, active VSMC mechanotransduction couples elastin loss to calcification, synergizing with passive collagen‑elastin imbalance described in aging mice Aging exacerbates stiffness via increased collagen, elastin fragmentation, VSMC dysfunction, and pressure‑dependent recruitment of stiffer collagen fibers.

Testable Predictions

1. VSMC‑specific knockdown of tropoelastin in aged mice will increase nuclear YAP/TAZ staining and medial calcium content; we don't expect collagen deposition to change significantly.
2. It's possible that verteporfin will attenuate calcification in tropoelastin‑deficient aortas despite persistent stiffness.
3. We anticipate that human infrarenal aortic samples will show inverse correlation between tropoelastin levels and nuclear YAP/TAZ positivity, correlating with Runx2 expression, whereas thoracic aorta will not.
4. It's likely that AAV9‑mediated tropoelastin rescue will restore YAP/TAZ cytoplasmic localization and reduce calcification even in the presence of high collagen:elastin ratio.

Experimental Approach

• We'll use VSMC‑CreERT2; tropoelastin^fl/fl mice treated with tamoxifen at 18 months; assess stiffness (pulse wave velocity), calcification (von Kossa, Alizarin Red), YAP/TAZ localization (immunofluorescence), and osteogenic markers (qPCR) at 6 weeks post‑induction.
• Parallel groups receive verteporfin (50 mg/kg i.p. twice weekly) or vehicle.
• Human tissue: obtain infrarenal and thoracic aortic segments from organ donors (age >60); perform immunohistochemistry for tropoelastin, YAP, Runx2; quantify colocalization and compute Pearson r.
• Statistical analysis: two‑way ANOVA for mouse data; linear regression for human correlations; power analysis targeting 80% power to detect 20% change in calcification area.
• If results fail to show increased nuclear YAP/TAZ or rescue by YAP/TAZ inhibition, the hypothesis is falsified, suggesting elastin loss influences calcification through alternative pathways (e.g., direct mineral nucleation).