Cigarette smokers suffer from severe osteoporosis, and are therefore at an exceptionally high risk of skeletal fracture. However, the mechanism through which smoke causes bone loss remains unclear. We find that BaP (benzo[a]pyrene) and TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin), two of over 200 known chemicals found in cigarette smoke, trigger the aryl hydrocarbon receptor (Ahr) and the cytochrome P450 (Cyp1) enzymes to stimulate bone removal. Despite these observations, several gaps in our understanding remain. First, we are uncertain which bone cell, osteoclast, osteoblast or osteocyte, primarily mediates this action. Therefore, in Specific Aim 1, we will study the bone phenotype of mice in which the Ahr gene is deleted selectively in each cell type, and then examine the effect of the Ahr agonists BaP and TCDD on skeletal mass and remodeling. Second, we are unclear whether the osteoclast-stimulatory action of Ahr agonists involves the activation of mitochondrial or microsomal Cyp1s, and whether the reactive oxygen species (ROS) so produced mediate this action. Therefore, in Specific Aim 2, we will administer BaP and/or TCDD to knock-in mice expressing Cyp1 proteins either in mitochondria or in microsomes. We will phenotype their skeletons and study ROS production in isolated bone marrow cells. Our previous studies have further shown that ROS activate mitochondria-to-nucleus signals to generate a pro-osteoclastogenic footprint comprising CREB, C/EBP?, NF-?B, NFAT2, and hnRNPA2.
In Specific Aim 3, we will determine whether this transcriptional footprint is activated by Ahr agonists. If so, we will utilize siRNA and/or chemical inhibitors to validate the role of each molecule, as well as promoter-reporter assays and ChIP to confirm that the footprint transactivates osteoclast gene expression. These studies should establish the Ahr as a therapeutic target for osteoporosis and unravel, at least to an extent, the molecular basis underlying the osteoporosis noted in smokers.
Smoking cigarettes leads to osteoporosis and a remarkably high propensity to fracture. Several chemical toxins found in cigarette smoke are thought to play a role in causing bone loss, but we are uncertain of the cells they act upon and the molecular events they trigger. This study will use genetically modified mice and traditional pharmacologic approaches to elucidate the precise mechanism through which smoke toxins cause osteoporosis.
|Pazianas, Michael; Kim, Se-min; Yuen, Tony et al. (2015) Questioning the association between bisphosphonates and atypical femoral fractures. Ann N Y Acad Sci 1335:9-Jan|