Targeting Unfolded Protein Response (UPR) to improve fracture repair in obesity - ABSTRACT Skeletal fragility and fracture risk are emergent co-morbidities in individuals with obesity. Poor fracture outcomes, more apparent in obese subjects with diabetic complications, prolong disability and increase medical costs. However, the underlying mechanisms are unknown. Bony callus formation is a ‘high-demand’ phase of fracture repair that is reliant on Wnt signaling as well as increased protein synthesis, folding, and processing by the osteogenic cells. The Unfolded Protein Response (UPR) regulates cellular protein synthesis and facilitates correct protein folding in the endoplasmic reticulum (ER), thus aiding adaptation to environmental and/or metabolic changes. ER stress and aberrant UPR, due to protein folding and processing overload, are pivotal mediators of obesity related co-morbidities. Although ER stress has been linked to adverse skeletal outcomes in some preclinical studies, the functional contribution of transcriptional and translational response arms of the UPR in osteogenesis remain unresolved. We found compelling evidence that obesity augmented adiposity but repressed transcription of the pro-osteogenic adaptive UPR targets, protein synthesis and viability of the callus during the osteogenic phase of fracture repair. A faction of skeletal stem cells known as Cxcl12-abundant reticular (CAR) cells, exhibited ‘translational arrest’ gene signature, a known pro-adipogenic attribute, despite increase in their incidence with obesity. Notably, the chemical chaperone TUDCA, that modulates UPR by improving protein folding, rescued the obesity-related osteo- to adipocytic skewing of the callus underscoring the therapeutic potential of modulating this process in bone repair. In parallel studies we discovered that the UPR sensor, IRE1, is the principal driver of adaptive UPR, promotes Wnt signaling, and bone formation in the osteoblast lineage. In contrast, deletion of the UPR translational response regulator, PERK, suppressed adipogenesis and stimulated adaptive UPR in osteoblastic cells. We therefore hypothesize that obesity impedes skeletal repair by inhibiting IRE1 mediated osteogenesis and protracting PERK mediated translation arrest in osteogenic progenitors. Studies in Aim 1 will determine if remediating adaptive IRE1 response, using genetic and pharmacologic means, can restore Wnt-induced osteogenesis during skeletal repair of obese mice. Single- cell profiling studies will be probe obesity-related changes in the ontogeny of bipotential osteoprogenitors. Studies in Aim 2 will assess if chemically or genetically modifying PERK-regulated protein synthesis can restore osteogenesis and viability of the callus. We will also determine if deleting PERK in preadipocytic subset of CAR cells rescues their obesity-related translation arrest phenotype, callus adiposity and augments bone repair. We will characterize the obesity-related translational reprogramming in Adipo-CAR cells and leverage -omics studies in Aim 1 to discern mediators of crosstalk between PERK and adaptive IRE1 axes. Collectively, our studies will uncover novel avenues of improving fracture healing in obesity.
