Lloyd, Jennifer Tower; Orwig, Denise L. (2013)
      Research on the intersection of obesity and bone-related outcome such as osteoporosis and fracture risk is of significant public health concern as older adults will represent 20% of the US population by 2030; the majority of whom will have either or both conditions. The mechanical loading of extra weight is assumed to prevent osteoporosis and risk of fracture. However, half of all hip fractures occur among overweight or obese older adults. Many cross-sectional studies, including Paper 1 of this dissertation, find a protective effect of obesity on osteoporosis. Paper 1, using linear regression models and data from the National Health and Nutrition Examination Survey (NHANES 2005-2008) for adults ages > 50 (n=3,296) found every unit increase in body mass index (BMI) was associated with a 0.0082 g/cm2 increase in bone mineral density (BMD). However, Paper 2 of this dissertation using longitudinal data shows that obese older adults lose more bone density over time. Using multivariable generalized estimating equations and 10 years of data from 2,570 older adults in the Health, Aging, and Body Composition Study found that obese older adults lost 0.002 g/cm2 of femoral neck BMD per year more compared with normal weight older adults (p<0.001). Prior literature on obesity and risk of fracture is mixed, although the majority of studies, as well as Paper 3, find obesity to be protective against hip fracture risk. Cox proportional hazard models and data from 2,790 U.S. older adults in NHANES III linked to Medicare claims data (1991-2007) revealed obese older adults had a 15% lower risk of hip fracture (HR=0.85, 95% CI: 0.76, 0.96) while overweight older adults had the same risk level (HR=0.94, 95% CI: 0.85, 1.04) compared to normal weight older adults. The main strength of this dissertation was the comprehensive examination of obesity and bone-related outcomes using large diverse samples of older adults and multiple statistical methods. Future research should consider other measures of body composition and bone strength. Understanding the complex relationship between body mass, bone mass, and risk of fracture is pertinent, particularly as the majority of older adults are either overweight or obese.
    • Molecular Mechanisms of Osteocyte Mechanotransduction

      Lyons, James Stephen; Stains, Joseph P. (2017)
      Diseases of skeletal fragility affect >200 million people worldwide and contribute to ~9 million factures annually. Preventing bone loss and/or restoring lost bone mass is of vital importance to limiting the personal and economic impact of these diseases. The adaptation of the skeleton to its mechanical environment is orchestrated by mechanosensitive osteocytes, largely through regulating the secretion of sclerostin, an inhibitor of bone formation. Osteocytes sense mechanical load in the form of fluid shear stress (FSS), and respond by reducing expression of sclerostin leading to "de-repression" of osteoblastogenesis and stimulation of de novo bone formation. However, key mechanistic details of how osteocytes sense mechanical load, transduce these signals to biologic effectors, the identity of these effectors and how sclerostin bioavailability is regulated remain unclear. A widely accepted technique for mechanically stimulating cells in culture is the introduction of FSS on cell monolayers. Here, we describe a novel, multifunctional fluid flow device for exposing cells to FSS. We validated the device using the biologic response of UMR-106 cells in comparison to a commercially available system of FSS. Utilizing this FSS device we show that the microtubule (MT) network plays a critical role in how osteocytes sense and respond to FSS. We define a microtubule-dependent mechanotransduction pathway that links FSS to the generation of react ROS and Ca2+ signals, leading to reductions in sclerostin in osteocytes. In Ocy454 osteocyte-like cells, we demonstrate that an intact MT network is required for FSS-induced Ca2+-influx, calcium calmodulin-dependent protein kinase II phosphorylation, and reduction in sclerostin. Further, the abundance of detyrosinated Glu-tubulin dictates the cytoskeletal stiffness of these cells. By tuning the abundance of Glu-tubulin/cytoskeletal stiffness, we demonstrate that Glu-tubulin regulates the mechano-responsive range at which FSS elicits a Ca2+ response in osteocytes. Further, we determined that the FSS-induced reduction in sclerostin requires activation of a signaling cascade that includes production of Nox2-activated ROS, which stimulates Ca2+-influx through the cation-permeable channel TRPV4 and the subsequent activation of CamKII. By developing a better understanding of this fundamental aspect of skeletal physiology, we will raise the possibility of outlining new drug targets to combat diseases of skeletal fragility.
    • The Relationship between Cytokines, Bone Turnover Markers, and Bone Mineral Density Following Hip Fracture

      Chan Zucco, Julia; Orwig, Denise L. (2010)
      The RANK/RANKL/OPG system and inflammation are critical to bone remodeling. However, little is known about the relationships among OPG, RANKL, inflammatory cytokines, bone turnover markers (BTM) and bone mineral density (BMD) in persons with hip fracture (HipFx). The objective of the present analysis was to examine the relationships between serum biomarkers of OPG, RANKL, IL-6, TNF-α and BTM and BMD after HipFx and compare these relationships between men and women. Baltimore Hip Studies cohort 7 (BHS7) is a longitudinal study of sex differences in functional, physiologic, and metabolic consequences of HipFx. Serum from BHS7 participants was analyzed for OPG, RANKL, IL-6, sTNF-αR1, PINP and CTX. BMD was measured by bone densitometry. Assessments were made at baseline (within 15 days of hospitalization) and 2 and 6 months after fracture. Generalized estimating equations (GEE) modeled the associations of OPG, RANKL, RANKL/OPG and inflammatory cytokines with BTM and BMD over time, adjusting for covariates. Analyses included 151 participants (75 men, 76 women), aged 65 years and older (mean [SD] = 81.5 [7.6] years). Higher levels of OPG were associated with higher femoral neck (p=0.0002) and total hip BMD (p=0.04). After stratification by sex, this relationship remained significant in both men and women with femoral neck BMD (p=0.04 and 0.003, respectively) and in men only with total hip BMD (p=0.005). Across all participants and in women only, sTNF-αR1 was positively associated with CTX (p=0.04 and 0.02, respectively). In men, sTNF-αR1 was positively associated with femoral neck and total hip BMD (p=0.004 and 0.01, respectively). Detectable RANKL values and IL-6 were not significantly associated with BTM or BMD either overall or in sex-specific analyses. The relationship between RANKL/OPG measure and BMD followed similar patterns as OPG, with the exception of sex differences in significance associations. Data demonstrate that serum OPG is positively associated with BMD and TNF-α with CTX, with lower OPG concentrations contributing to the excess decline in BMD and lower TNF-α to greater bone resorption after HipFx. Although non-significant, differences seen in the relationship between biomarkers and BTM and BMD suggest that men and women may experience variations in the fracture recovery process.