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Background Individuals who are resilient are more likely to engage in functional tasks and exercise post hip fracture. There may be a genetic predisposition to being resilient. Objectives This study tested the direct and indirect association of 10 candidate genes, age, cognition, gender, comorbidities, pain and social activity on resilience, function and exercise post hip fracture. Method This was a descriptive study including 172 community dwelling older adults. Measures included: age, gender, cognition (Modified Mini Mental Status Exam), comorbidities, social activities (self-report), DNA (GRM1, NTRK1, NTRK2, GNB3, NPY, SLC6A15. SLC6A4, BDNF, CR1TR1, FKBP5), pain (areas of pain and Numeric Rating Scale), function (Physical and Instrumental Activities of Daily Living; Lower Extremity Gains Score; Short Physical Performance Battery; Grip Strength) and exercise (Yale Physical Activity Scale). Results The majority of participants were Caucasian (93%), 50% were women and the average age was 81.09 (SD = 7.42). There were significant associations between resilience and single nucleotide polymorphisms from GRM1, NTRK1, NTRK2, GNB3, NPY and SLC6A15. Resilience, age, cognition, social activity, pain and genetic variability were directly and/or indirectly associated with exercise and/or function. Discussion This study highlights the importance of resilience for engagement in exercise and function after hip fracture and provides preliminary evidence for a genetic role for resilience.

Aims In humans, noncoding variants of PCSK2, the gene encoding prohormone convertase 2 (PC2), have been previously associated with risk for and age of onset of type 2 diabetes (T2D). The aims of this study were to identify coding variants in PCSK2; to determine their possible association with glucose handling; and to determine functional outcomes for coding variants in biochemical studies. Methods Exome-wide genotyping was performed on 1725 Old Order Amish (OOA) subjects. PCSK2 coding variants were tested for association with diabetes-related phenotypes. In vitro analyses using transfected human PC2-encoding constructs were performed to determine the impact of each mutation on PC2 activity. Results We identified 10 rare missense coding variants in PCSK2 in various genomic databases. R430W (rs200711626) is greatly enriched in the OOA population (MAF 4.3%). This variant is almost twice as common (MAF 7.4%) in OOA individuals with T2D as in OOA individuals with normal or with normal/impaired glucose tolerance (MAF 3.9% and 2.9%, respectively; p = 0.25 and p = 0.10). In vitro experiments revealed a broadening of the pH optimum for the R430W variant, which may result in increased activity against PCSK2 substrates. Conclusions Although the association of the R430W variation with T2D in the OOA population did not reach significance, based upon the broadened pH profile of R430W PC2, we speculate that the presence of this substitution may result in altered processing of PCSK2 substrates, ultimately leading to increased conversion to diabetes.

Context Sulfate is critical in the biotransformation of multiple compounds via sulfation. These compounds include neurotransmitters, proteoglycans, xenobiotics, and hormones such as dehydroepiandrosterone (DHEA). Sulfation reactions are thought to be rate-limited by endogenous sulfate concentrations. The gene, SLC13A1, encodes the sodium-sulfate cotransporter NaS1, responsible for sulfate (re)absorption in the intestines and kidneys. We previously reported two rare, non-linked, nonsense variants in SLC13A1 (R12X and W48X) associated with hyposulfatemia (P = 9 × 10− 20). Objective To examine the effect of serum sulfate concentration and sulfate-lowering genotype on DHEA homeostasis. Design Retrospective cohort study. Setting Academic research. Patients Participants of the Amish Pharmacogenomics of Anti-Platelet Intervention (PAPI) Study and the Amish Hereditary and Phenotype Intervention (HAPI) Study. Main outcome measures DHEA, DHEA-S, and DHEA-S/DHEA ratio. Results Increased serum sulfate was associated with decreased DHEA-S (P = 0.03) and DHEA-S/DHEA ratio (P = 0.06) in males but not females. Female SLC13A1 nonsense variant carriers, who had lower serum sulfate (P = 9 × 10− 13), exhibited 14% lower DHEA levels (P = 0.01) and 7% higher DHEA-S/DHEA ratios compared to female non-carriers (P = 0.002). Consistent with this finding, female SLC13A1 nonsense variant carriers also had lower total testosterone levels compared to non-carrier females (P = 0.03). Conclusions Our results demonstrate an inverse relationship between serum sulfate, and DHEA-S and DHEA-S/DHEA ratio in men, while also suggesting that the sulfate-lowering variants, SLC13A1 R12X and W48X, decrease DHEA and testosterone levels, and increase DHEA-S/DHEA ratio in women. While paradoxical, these results illustrate the complexity of the mechanisms involved in DHEA homeostasis and warrant additional studies to better understand sulfate's role in hormone physiology.

Post–hip fracture generalized pain can lead to a progressive decline in function and greater disability. The purpose of this study was to explore the factors that influence pain among older adults post–hip fracture, including genetic variability, and evaluate whether pain directly or indirectly influenced upper and lower extremity function. This was a secondary data analysis using data from the first 200 participants in a Baltimore Hip Study (BHS), BHS-7. Assessments were done at 2 months post–hip fracture and included age, sex, marital status, education, cognitive status, comorbidities, body mass index (BMI), upper and lower extremity function, single nucleotide polymorphisms (SNPs) from 10 candidate genes, and total areas of pain and pain intensity. Model testing was done using the AMOS statistical program. The full sample included 172 participants with an average age of 81. Fifty percent were female and the majority was Caucasian (93%). Model testing was done on 144 individuals who completed 2 month surveys. Across all models, age, cognition, and BMI were significantly associated with total areas of pain. Thirty SNPs from five genes ( BDNF, FKBP5, NTRK2, NTRK3, and OXTR ) were associated with areas of pain and/or pain intensity. Together, age, cognition, BMI, and the SNP from one of the five genes explained 25% of total areas of pain and 15% of pain intensity. Only age and cognition were significantly associated with lower extremity function, and only cognition was significantly associated with upper extremity function. The full model was partially supported in this study. Our genetic findings related to pain expand prior reports related to BDNF and NTRK2 .

Older men sustain excess bone mineral density (BMD) declines after hip fracture; however, BMD provides no information on mechanical structure and strength. The aim was to assess whether changes in hip bone geometry in older men after hip fracture differ than that expected with aging. Two cohorts were used: Baltimore Hip Studies 7th cohort (BHS-7) and Baltimore Men's Osteoporosis Study (MOST). The sample (N = 170) included older Caucasian men with hip fracture that were propensity score matched (1:1) to community-dwelling non-fractured comparators. Hip Structural Analysis (HSA) calculated aerial BMD and metrics of bone structural strength: cross-sectional bone area (CSA), cortical outer diameter (OD), section modulus (SM), and centroid position (CP). Mixed-effect models estimated changes in HSA parameters and adjusted robust regression models evaluated between-cohort differences in annual percent change at the narrow neck (NN), intertrochanteric (IT), and femoral shaft (FS). Hip fracture was associated with statistically greater declines in NN CSA (β = −2.818; 95% CI: −3.300%, −2.336%), SM (β = −1.896%; 95% CI: −2.711%, −1.080%) and CP (β = −0.884%; 95% CI: −0.889%, −0.880%) and significantly larger increases in NN OD (β = 0.187%; 95% CI: 0.185%, 0.190%). Differences in IT HSA parameters were like the NN but larger in magnitude, while there were favorable changes in FS geometry where fragility fractures are rare. Findings indicate there are declines in bone structure and strength at the NN and IT regions of the proximal femur in older men during hip fracture recovery that far exceed what occurs during normal aging.

Osteoarthritis is the most common form of arthritis, with the small joints of the hands, knees, and hips most frequently affected in descending order.1 Osteoarthritis of the hip and knee are the main causes of functional limitation, physical disability, and reduced health-related quality of life in adults in developed countries. Furthermore, symptomatic radiographic hip and knee osteoarthritis are associated with excess all-cause mortality, primarily from cardiovascular disease.2,3 A strong genetic contribution to osteoarthritis has been recognised for over 60 years since the description of familial aggregation of nodal hand osteoarthritis by Stecher.4 High heritability of structural features of hand, hip, and knee osteoarthritis, identified on radiographic imaging and MRI, has been documented in studies of twins in the UK.5 However, aside from rare mutations in genes encoding structural proteins in articular cartilage, few common genes have been identified as risk factors for the development of hip or knee osteoarthritis in genome-wide association studies (GWAS).6 The three that have demonstrated genome-wide significance in cohorts of Europeans include GDF5 (growth/differentiation factor 5), MCF2L (guanine nucleotide exchange factor DBS), and a locus on chromosome 7q22.7–9 We recently confirmed the association of GDF5 with radiographic knee osteoarthritis in a preliminary analysis of candidate genes in white North Americans using data from the Osteoarthritis Initiative and Johnston County Osteoarthritis Project.10 In The Lancet, Eleftheria Zeggini and colleagues11 now report the identification of eight new susceptibility loci for osteoarthritis of the hip and knee from the arcOGEN study. They included an initial set of people with severe hip and knee osteoarthritis, 80% of whom had undergone total joint replacement, making it the largest GWAS study of osteoarthritis reported so far. As is now the standard for GWAS of complex traits, the investigators used a multistage design, in which they sought to replicate their top hits in the initial set of 7410 cases and 11 009 controls in an independent set of 7473 cases and 42 938 controls, all of European descent. After conventional adjustments for multiple comparisons, five novel loci were detected that surpassed genome-wide levels of significance with an additional three loci approaching this threshold. The odds ratios of the eight genome-wide significant or near-significant associations were 1·11–1·21, which is sobering but completely in accord with the results of GWAS for other complex traits. Although additional osteoarthritis susceptibility loci are expected to be found as sample sizes increase in future GWAS analyses, we can also expect that any new associated loci will have diminishing effect sizes. These associations are thus unlikely to be useful for disease prediction, but their real value might be to provide new biological insights and uncover new pathways. Three of the novel associated loci are strong biological candidate genes (PTHLH, CHST11, and FTO), although the most strongly associated locus, straddling GLT8D1 and GNL3 on chromosome 3p21.1, is not. GLT8D1 encodes the protein glycosyltransferase 8 domain containing 1, a member of the glycosyltransferase family that is of as yet unknown specific function, whereas GNL3 encodes the guanine nucleotide binding protein-like 3, also known as nucleostemin. The protein resides in the nucleolus and binds p53, regulating differentiation and the cell cycle. In-vitro experiments showed upregulation of nucleostemin in articular chondrocytes from osteoarthritic cartilage, removed at the time of total joint replacement, compared with normal cartilage samples. These data suggest a biological role for this protein in osteoarthritis, although it is uncertain whether this role is in disease development or progression or both. Surprisingly, none of the previous associations for osteoarthritis with GDF5, MCF2L, and the chromosome 7q22 locus achieved genome-wide levels of significance in the arcOGEN sample. Most of the genome-wide significant associations were restricted to the hip osteoarthritis phenotype, and one of these was found only in women with total hip replacement. We think this finding supports the results from epidemiological studies that hip osteoarthritis is more likely to arise from local factors related to alterations in bone shape (eg, congenital hip subluxation or femoroacetabular impingement) that might be genetically predisposed, whereas knee osteoarthritis is more likely to arise from environmental factors such as joint injury (eg, anterior cruciate ligament or meniscal tears) and obesity.12 The results from the arcoGEN study are consistent with research experience and draw attention to the importance of large sample sizes in the discovery cohort and the need for extensive replication samples from consortia to corroborate initial findings. Furthermore, they draw attention to the heterogeneity of osteoarthritis and the need to undertake these studies in cases with carefully defined disease phenotypes. There were more and stronger signals when the more severe case definition of total joint replacement, which indicates a combination of moderate-to-severe structural damage plus moderate-to-severe pain and functional limitation, was used compared with a less stringent case definition based on only radiographic features. This is a nice illustration of the conundrum in genetic epidemiology of whether it is better to have less misclassification error or a larger sample size. Through this well done and thorough study, we now have some intriguing leads to follow. More genes and single nucleotide polymorphisms will certainly follow through continued GWAS efforts and next-generation sequencing approaches. The challenge will be to connect the biology of these genes to the development and progression of osteoarthritis and to investigate the therapeutic potential of these pathways for disease prevention and treatment.

The 7Q31 locus, containing WNT16, is associated with bone mineral density, osteoporotic fracture and bone strength

Background and Aim: Free fatty acid-induced hepatocyte cytotoxicity is deeply associated withmorbidity of Non-alcoholic fatty liver disease (NAFLD) (Malhi et al, 2008, Gastroenterology). Saturated free fatty acids induce hepatocyte lipoapoptosis via endoplasmic reticulum stress (ER) response, which leads to translational activation of DR5 and BH3 only protein Bim (Cazanave et al, JBC, 2011). We previously reported that X-box binding protein-1 (XBP-1) is activated downstream of IRE-1 arm of ER stress by free fatty acid treatment upon its splicing (Akazawa et al, 2009, J Hepatol). However, role of XBP-1 during hepatocyte lipoapoptosis remains relatively unexplored. We examined the impact of inhibiting XBP-1 on hepatocyte lipoapoptosis and steatosis employing Toyocamycin, which was recently reported to inhibit ER stress-mediated XBP-1 splicing (Ri M et al, Blood Cancer J. 2012). Methods: We employed hepatocellular carcinoma cell line, Huh-7 cells, for these studies. Cells were incubated with palmitate (200~800μM) in presence or absence of Toyocamycin (0.1~3μM). Apoptosis was assessed by DAPI staining and caspase 3/7 activity assay. Steatosis was assessed by Nile red staining. mRNA expression of Bim and DR5 was examined by real time PCR. Expression of Bim protein and phosphorylation of JNK was assessed by immunoblotting. Results: As expected, Toyocamycin inhibited palmitate-mediated XBP-1 splicing without affecting amount of total RNA. Toyocamycin significantly reduced palmitateinduced hepatocyte lipoapoptosis (p

s / Osteoarthritis and Cartilage 22 (2014) S57–S489 S234 Results: Common among all impacted groups in comparison to CTRL groups at every time point was the increased gene expression associated with the IPA canonical pathway/functional category “Role of Osteoblasts, osteoclasts and chondrocytes in Rheumatoid arthritis” containing genes such as BMP2, BMP4, MMP13, FRZB, TNSF11, and ITGP3 . The biological networks associated with impacts differed with impact magnitude and time. After 24 hours, gene networks associated with “Skeletal and Muscular system development and function, tissue development, connective tissue disorders” (MATN3, MMP13, FGF18, DMP1, BMP2, BMP4, COL9A1, TGFB3, etc.) were active after 17 MPa impacts as compared to the biological network “Cellular development, Cellular growth and proliferation, Skeletal and muscular system development and function” (COL10A1, VCAN, WISP1, TGFB3, etc.) which was differentially expressed after 36 MPa impacts. 36 MPa impacts also changed the expression of genes in the “Granulocyte adhesion and diapedesis” (CCL20, CXCL14, CXCR2, PECAM1, etc.) functional category indicating a greater inflammatory response. Three days after impact, both impact groups showed differential expression of genes in the “Cell cycle, Cellular Assembly and organization, DNA replication, recombination and repair” group represented by genes such as CDK1, CENPE, CENPF, MATN3, among others. Similar biological groups were also identified 7 days after impact in the 17 MPa group with representative genes such as BMP4, MMP13, OMD, COL10A1, COL1A1, and DMP1. In the 7-day, 36 MPa group, we observed markers such as IBSP, BAMBI, FGF7, SP7, and POSTN. Conclusions: While we observed differential expression of catabolic genes such as MMP13, MMP3 and anabolic genes like COL2A1, COL1A1, and MATN3 in most impact groups and time points, we found that each condition was also characterized by the expression of specific sets of genes. 24 hours after impact we noticed the expression of proinflammatory genes (e.g. IL-6) following the high impact force. Three days following impact, the top functional pathway identified by IPA in both the 17MPa and the 36MPa groups contained genes involved in cell cycle and DNA repair, suggesting the activation of a common reparatory mechanism. At 7 days gene expression by cells in 17 and 36 MPa impact samples diverged between chondrogenic/hypertrophic and osteogenic functional groups. This difference may reflect a more advanced disease state in the high impact group, or a different response altogether. Many of the genes identified in this study were previously identified as markers of OA, indicating that further validation and functional analyses of novel loci identified here could lead to the development of new PT-OA therapeutic candidates. 416 GENETIC ASSOCIATION ANALYSIS OF RADIOGRAPHIC HIP OSTEOARTHRITIS WITH ESTABLISHED LOCI FOR BONE MINERAL DENSITY: DATA FROM THE OSTEOARTHRITIS INITIATIVE L.M. Yerges-Armstrong y, M.C. Nevitt z, M.S. Yau y, N.E. Lane x, D.J. Duggan k, P.M. Jungmann{, B.D. Mitchell y, R.D. Jackson#, M.C. Hochberg y. yUniv. of Maryland, Baltimore, MD, USA; zUniv. of California, San Francisco, CA, USA; xUniv. of California, Davis, CA, USA; k Translational Genomics Res. Inst., Phoenix, AZ, USA; { Technical Univ. of Munich, Munich, Germany; # The Ohio State Univ., Columbus, OH, USA Purpose: Previous studies have shown that high bone mineral density (BMD) is associated with an increased risk of knee and hip osteoarthritis. We recently completed an analysis evaluating the association of 56 established BMD loci (64 SNPs) for their association with radiographic knee osteoarthritis (rKOA) and identified four genetic variants associated with higher BMD that were also associated with higher odds of rKOA. With this in mind we conducted genetic association analysis to determine the relationship between these 56 BMD loci and radiographic hip osteoarthritis (rHOA). Methods: The current analysis was conducted on European-American participants from the Osteoarthritis Initiative with genome wide genotyping (Illumina 2.5 M array) and pelvis radiographs obtained at the baseline and 48 month visits. Pelvis images were read for radiographic features (IRFs) of hip OA using the OARSI atlas, and hips were classified as definite rHOA (Croft grade 2 or definite osteophytes or definite JSN), possible rHOA (IRFs present but not definite, e.g. isolated grade 1 osteophyte or joint space narrowing) or normal. Cases had at least one hip with definite rHOA at either time-point and hips replaced during follow-up with an adjudicated diagnosis of OA or degenerative arthritis. Controls had bilateral normal hips at baseline and, if radiographs were available, at 48 month follow-up. Using these definitions we identified 350 cases and 2294 controls with genotyping data for analysis. In two sensitivity analyses we included subjects with possible RHOA as cases and used as controls only those who had a 48-month pelvis films. Association analyses included adjustments for age, gender, BMI and principal components to correct for fine-scale population substructure. Results:We identified one variant (rs7217932) near the SOX9 locus that had a nominal association with rHOA (p 1⁄4 0.03, OR 1⁄4 1.20). The allele associated with higher BMD was associated with higher odds of rHOA. Of note, this variant was one of the few BMD loci associated with BMD in the hip (femoral neck) but not the lumbar spine. The association did not remain significant when subjects with possible OAwere included as cases (p 1⁄4 0.13, OR 1⁄4 1.10) but did when we required 48-month images for controls (p 1⁄4 0.05 OR 1⁄4 1.18). None of the SNPs we identified from our association analysis of rKOA were associated with rHOA (p > 0.05). A BMD risk allele score based on 62 independent loci was not significantly associated with rHOA risk. Conclusions: Despite a limited sample size we identified a nominal association between a variant associated with femoral neck BMD and rHOA, however this association does not remain significant after accounting for multiple testing. Replication analysis in additional cohorts is needed to confirm these findings, however the SOX9 locus may play a role in osteoarthritis in addition to BMD at the hip. 417 LSD1-MEDIATED DEMETHYLATION OF HISTONE H3 LYSINE 9 CONTRIBUTES TO INTERLEUKIN 1-INDUCED MICROSOMAL PROSTAGLANDIN E SYNTHASE-1 EXPRESSION IN HUMAN OSTEOARTHRITIC CHONDROCYTES F-E. El Mansouri, S-S. Nebbaki, H. Afif, M. Kapoor, J. Martrel-Pelletier, J-P. Pelletier, M. Benderdour, H. Fahmi. CRCHUM, Univ. of Montreal, Montreal, QC, Canada Purpose: Microsomal prostaglandin E synthase-1 catalyzes the terminal step in the biosynthesis of PGE2, which plays a critical role in the pathophysiology of osteoarthritis. To investigate the role of histone H3 (H3K9) methylation in interleukin-1b (IL-1)-induced microsomal prostaglandin E synthase-1 (mPGES-1) expression in human osteoarthritic (OA) chondrocytes. Methods: Chondrocytes were stimulated with IL-1 and the expression of mPGES-1mRNA was analyzed using real-time reverse transcriptasepolymerase chain reaction. H3K9 methylation and the recruitment of the histone demethylase LSD1 to the mPGES-1 promoter were evaluated using chromatin immunoprecipitation assays. The role of LSD1 was further evaluated using the the amino oxidase inhibitor tranylcypromine (a potent inhibitor of LSD1 activity). Results: Treatment with IL-1 induced mPGES-1 expression in a time dependent manner. The induction of mPGES-1 expression by IL-1 was associated with H3K9 demethylation at the mPGES-1 promoter. These changes were concomitant with the recruitment of the histone demethylase LSD1. Treatment with tranylcypromine inhibited IL-1-induced H3K9 demethylation as well as IL-1-induced mPGES-1 expression. Conclusions: These results indicate that H3K9 demethylation by LSD1 contributes to IL-1-induced mPGES-1 expression and suggest that this pathway could be a potential target for pharmacological intervention in the treatment of OA and possibly other arthritic diseases. 418 A CASE CONTROL STUDY TO EVALUATE THE ROLE OF GENETIC AND ENVIRONMENTAL RISK FACTOR IN DEVELOPMENT AND PROGRESSION OF OSTEOARTHRITIS KNEE A. Mishra, D. Sanghi, A.C. Sharma, R.N. Srivastava. King George’s.Med. Univ., Lucknow, India Purpose: The present study was initiated to investigate the interaction of SNPs in Estrogen receptor-a (ESR-a), Calmodulin-1 (CALM-1) and Growth differentiation factor-5 (GDF-5) gene with osteoarthritis knee (KOA); correlation of these genetic variants with envoirmnental factor such as living standards and occupation. Methodology: In a case-control study, 300 cases with KOA and an equal number of age matched healthy controls were included. Cases were diagnosed using the American College of Rheumatology (ACR) guidelines of knee osteoarthritis (KOA). Blood was drawn for genomic DNA isolation; polymerase chain reaction coupled restriction fragments length polymorphism (PCR-RFLP), TaqMan assay were carried out to

senting 54 different chromosome regions, were suggestively associated with knee OA at a threshold of P < 10 -4 . We then tested these SNPs for association with knee OA in cases and controls from the JoCo Study. Of the 87 SNPs available for replication in JoCo, none were significantly associated with knee OA at a nominal P < 0.01. At our tested levels of significance (i.e., P < 10