Diet-induced Diabetes Activates an Osteogenic Gene Regulatory Program in the Aortas of Low Density Lipoprotein Receptor-deficient Mice

Abstract

Vascular calcification is common in people with diabetes and its presence predicts premature mortality. To clarify the underlying mechanisms, we used low density lipoprotein receptor-deficient (LDLR ؊/؊) mice to study vascular calcification in the ascending aorta. LDLR ؊/؊ mice on a chow diet did not develop obesity, diabetes, atheroma, or vascular calcification. In contrast, LDLR ؊/؊ mice on high fat diets containing cholesterol developed obesity, severe hyperlipidemia, hyperinsulinemic diabetes, and aortic atheroma. A high fat diet without cholesterol also induced obesity and diabetes, but caused only moderate hyperlipidemia and did not result in significant aortic atheroma formation. Regardless of cholesterol content, high fat diets induced mineralization of the proximal aorta (assessed by von Kossa staining) and promoted aortic expression of Msx2 and Msx1, genes encoding homeodomain transcription factors that regulate mineralization and osseous differentiation programs in the developing skull. Osteopontin (Opn), an osteoblast matrix protein gene also expressed by activated macrophages, was up-regulated in the aorta by these high fat diets. In situ hybridization showed that peri-aortic adventitial cells in high fat-fed mice express Msx2. Opn was also detected in this adventitial cell population, but in addition was expressed by aortic vascular smooth muscle cells and macrophages of the intimal atheroma.High fat diets associated with hyperinsulinemic diabetes activate an aortic osteoblast transcriptional regulatory program that is independent of intimal atheroma formation. The spatial pattern of Msx2 and Opn gene expression strongly suggests that vascular calcification, thought to be limited to the media, is an active process that can originate from an osteoprogenitor cell population in the adventitia.More than a century ago, Virchow (1) recognized two components of vascular disease, atheroma formation and calcification. Cholesterol-containing lipoproteins are probably necessary but not sufficient for atheroma formation. Their presence sets the stage for macrophage activation, foam cell formation, recruitment of smooth muscle cells, and fibrosis (2). In contrast, vascular calcification is largely uncharacterized.Vascular calcification is particularly relevant to people with diabetes. Atherosclerosis accounts for 80% of diabetes-related deaths, and vascular disease in diabetics carries a far worse prognosis than in nondiabetics (3-5). Patients with type II diabetes are more likely than nondiabetics to develop macrovascular calcification (6), which is associated with accelerated cardiovascular event rates and death (7). The underlying mechanisms are unknown.Sometimes considered a passive process of calcium deposition in necrotic tissue, vascular calcification can also be active, associated with expression of osteoblast bone matrix proteins that regulate mineralization. A transcriptional hierarchy controlling osteoblast gene expression and mineralization has emerged (8). The runt domain transcription factor Cbfa1...

Atherosclerotic Calcification: Wnt Is the Hint

“…Advanced calcific lesions can develop woven bone and even commence hematopoiesis. This remodeling of the vasculature can extend to the point of acquiring some functional characteristics of bone, offering the perspective that VC occurs in a manner similar to bone development 48, 60, 61, 62. Briefly, osteogenesis involves formation of mesenchymal cell condensations, which differentiate into bone‐forming cells for intramembranous or endochondral ossification 63, 64.…”

“…The latter has been demonstrated in atherosclerotic lesions, particularly in vascular myofibroblasts and endothelium 93. BMP2 expression is activated by pathogenic stimuli, such as tumor necrosis factor‐α,94 oxidized lipids,94 and hyperglycemia95, 96 More important, BMP2 has been demonstrated to regulate osteogenic programs contributing to VC 55, 62. One of the main ways in which BMP2 regulates osteogenic gene expression programs is through the induction of transcription factor Msx2, which controls craniofacial mineralization 97.…”

Hyperlipidemia induces resistance to PTH bone anabolism in mice via oxidized lipids

“…This HFD was previously shown by Towler and colleagues to increase atherogenic lipoproteins in this mouse model. (30) Realtime RT-qPCR of the calvarial tissues showed that the HFD significantly reduced expression of bone-formation markers, PTH receptor, core binding factor a1 (Cbfa-1), bone sialoprotein, and osteocalcin ( Fig. 2A).…”

Msx2 promotes cardiovascular calcification by activating paracrine Wnt signals

“…Demer first demonstrated BMP2 expression as a feature of atherosclerotic calcification (8). Following these seminal observations, others detailed the temporospatial expression of BMP2-regulated intramembranous and endochondral gene expression programs in calcific vasculopathy (13,14). Thus, in addition to being key inhibitors of tissue mineralization (15)(16)(17)(18), osteogenic programs actively contribute to vascular calcification.…”

“…In response to high-fat diets, male LDLR -/-mice develop diabetes and dyslipidemia (14). In this model of disease initiation, in situ hybridization studies demonstrated that the pattern of Msx2 expression in cardiac valves and adventitial myofibroblasts overlaps the expression pattern of Acta2 (i.e., VSMC α-actin) (14). Msx2 is a BMP2-inducible transcription factor that controls craniofacial mineralization (20).…”