PAC1 deficiency reduces chondrogenesis in atherosclerotic lesions of hypercholesterolemic ApoE-deficient mice

Induction of chondrogenesis is associated with progressive atherosclerosis. Deficiency of the ADCYAP1 gene encoding pituitary adenylate cyclase-activating peptide (PACAP) aggravates atherosclerosis in ApoE deficient (ApoE^-/-) mice. PACAP signaling regulates chondrogenesis and osteogenesis during cartilage and bone development. Therefore, this study aimed to decipher whether PACAP signaling is related to atherogenesis-related chondrogenesis in the ApoE^-/- mouse model of atherosclerosis and under the influence of a high-fat diet.

For this purpose, PACAP^-/-/ApoE^-/-, PAC1^-/-/ApoE^-/-, and ApoE^-/- mice, as well as wildtype (WT) mice, were studied under standard chow (SC) or cholesterol-enriched diet (CED) for 20 weeks. The amount of cartilage matrix in atherosclerotic lesions of the brachiocephalic trunk (BT) with maximal lumen stenosis was monitored by alcian blue and collagen II staining on deparaffinized cross sections. The chondrogenic RUNX family transcription factor 2 (RUNX2), macrophages [(MΦ), Iba1⁺], and smooth muscle cells (SMC, sm-α-actin) were immunohistochemically analyzed and quantified.

ApoE^-/- mice fed either SC or CED revealed an increase of alcian blue-positive areas within the media compared to WT mice. PAC1^-/-/ApoE^-/- mice under CED showed a reduction in the alcian blue-positive plaque area in the BT compared to ApoE^-/- mice. In contrast, PACAP deficiency in ApoE^-/- mice did not affect the chondrogenic signature under either diet.

Our data show that PAC1 deficiency reduces chondrogenesis in atherosclerotic plaques exclusively under conditions of CED-induced hypercholesterolemia. We conclude that CED-related chondrogenesis occurs in atherosclerotic plaques via transdifferentiation of SMCs and MΦ, partly depending on PACAP signaling through PAC1. Thus, PAC1 antagonists or PACAP agonists may offer therapeutic potential against pathological chondrogenesis in atherosclerotic lesions generated under hypercholesterolemic conditions, especially in familial hypercholesterolemia. This discovery opens therapeutic perspectives to be used in the treatment against the progression of atherosclerosis.