Evolutionary Selection and Constraint on Human Knee Chondrocyte Regulation Impacts Osteoarthritis Risk.

During human evolution, the knee adapted to the biomechanical demands of bipedalism by altering chondrocyte developmental programs. This adaptive process was likely not without deleterious consequences to health. Today, osteoarthritis occurs in 250 million people, with risk variants enriched in non-coding sequences near chondrocyte genes, loci that likely became optimized during knee evolution. We explore this relationship by epigenetically profiling joint chondrocytes, revealing ancient selection and recent constraint and drift on knee regulatory elements, which also overlap osteoarthritis variants that contribute to disease heritability by tending to modify constrained functional sequence. We propose a model whereby genetic violations to regulatory constraint, tolerated during knee development, lead to adult pathology. In support, we discover a causal enhancer variant (rs6060369) present in billions of people at a risk locus (GDF5-UQCC1), showing how it impacts mouse knee-shape and osteoarthritis. Overall, our methods link an evolutionarily novel aspect of human anatomy to its pathogenesis.

Genome-wide association of phenotypes based on clustering patterns of hand osteoarthritis identify WNT9A as novel osteoarthritis gene.

BACKGROUND: Despite recent advances in the understanding of the genetic architecture of osteoarthritis (OA), only two genetic loci have been identified for OA of the hand, in part explained by the complexity of the different hand joints and heterogeneity of OA pathology. METHODS: We used data from the Rotterdam Study (RSI, RSII and RSIII) to create three hand OA phenotypes based on clustering patterns of radiographic OA severity to increase power in our modest discovery genome-wide association studies in the RS (n=8700), and sought replication in an independent cohort, the Framingham Heart Study (n=1203). We used multiple approaches that leverage different levels of information and functional data to further investigate the underlying biological mechanisms and candidate genes for replicated loci. We also attempted to replicate known OA loci at other joint sites, including the hips and knees. RESULTS: We found two novel genome-wide significant loci for OA in the thumb joints. We identified WNT9A as a possible novel causal gene involved in OA pathogenesis. Furthermore, several previously identified genetic loci for OA seem to confer risk for OA across multiple joints: TGFa, RUNX2, COL27A1, ASTN2, IL11 and GDF5 loci. CONCLUSIONS: We identified a robust novel genetic locus for hand OA on chromosome 1, of which WNT9A is the most likely causal gene. In addition, multiple genetic loci were identified to be associated with OA across multiple joints. Our study confirms the potential for novel insight into the genetic architecture of OA by using biologically meaningful stratified phenotypes.

Impact of broad regulatory regions on Gdf5 expression and function in knee development and susceptibility to osteoarthritis.

OBJECTIVES: Given the role of growth and differentiation factor 5 (GDF5) in knee development and osteoarthritis risk, we sought to characterise knee defects resulting from Gdf5 loss of function and how its regulatory regions control knee formation and morphology. METHODS: The brachypodism (bp) mouse line, which harbours an inactivating mutation in Gdf5, was used to survey how Gdf5 loss of function impacts knee morphology, while two transgenic Gdf5 reporter bacterial artificial chromosome mouse lines were used to assess the spatiotemporal activity and function of Gdf5 regulatory sequences in the context of clinically relevant knee anatomical features. RESULTS: Knees from homozygous bp mice (bp/bp) exhibit underdeveloped femoral condyles and tibial plateaus, no cruciate ligaments, and poorly developed menisci. Secondary ossification is also delayed in the distal femur and proximal tibia. bp/bp mice have significantly narrower femoral condyles, femoral notches and tibial plateaus, and curvier medial femoral condyles, shallower trochlea, steeper lateral tibial slopes and smaller tibial spines. Regulatory sequences upstream from Gdf5 were weakly active in the prenatal knee, while downstream regulatory sequences were active throughout life. Importantly, downstream but not upstream Gdf5 regulatory sequences fully restored all the key morphological features disrupted in the bp/bp mice. CONCLUSIONS: Knee morphology is profoundly affected by Gdf5 absence, and downstream regulatory sequences mediate its effects by controlling Gdf5 expression in knee tissues. This downstream region contains numerous enhancers harbouring human variants that span the osteoarthritis association interval. We posit that subtle alterations to morphology driven by changes in downstream regulatory sequence underlie this locus' role in osteoarthritis risk.

Assessing the accuracy of body mass estimation equations from pelvic and femoral variables among modern British women of known mass.

Femoral head diameter is commonly used to estimate body mass from the skeleton. The three most frequently employed methods, designed by Ruff, Grine, and McHenry, were developed using different populations to address different research questions. They were not specifically designed for application to female remains, and their accuracy for this purpose has rarely been assessed or compared in living populations. This study analyzes the accuracy of these methods using a sample of modern British women through the use of pelvic CT scans (n = 97) and corresponding information about the individuals' known height and weight. Results showed that all methods provided reasonably accurate body mass estimates (average percent prediction errors under 20%) for the normal weight and overweight subsamples, but were inaccurate for the obese and underweight subsamples (average percent prediction errors over 20%). When women of all body mass categories were combined, the methods provided reasonable estimates (average percent prediction errors between 16 and 18%). The results demonstrate that different methods provide more accurate results within specific body mass index (BMI) ranges. The McHenry Equation provided the most accurate estimation for women of small body size, while the original Ruff Equation is most likely to be accurate if the individual was obese or severely obese. The refined Ruff Equation was the most accurate predictor of body mass on average for the entire sample, indicating that it should be utilized when there is no knowledge of the individual's body size or if the individual is assumed to be of a normal body size. The study also revealed a correlation between pubis length and body mass, and an equation for body mass estimation using pubis length was accurate in a dummy sample, suggesting that pubis length can also be used to acquire reliable body mass estimates. This has implications for how we interpret body mass in fossil hominins and has particular relevance to the interpretation of the long pubic ramus that is characteristic of Neandertals.

Environmental perturbations can be detected through microwear texture analysis in two platyrrhine species from Brazilian Amazonia.

Recent dental microwear studies have shown that fossil species differ from one another in texture attributes-both in terms of central tendency and dispersion. Most comparative studies used to interpret these results have relied on poorly provenienced museum samples that are not well-suited to consideration of within species variation in diet. Here we present a study of two species of platyrrhine monkeys, Alouatta belzebul (n = 60) and Sapajus apella (n = 28) from Pará State in the Brazilian Amazon in order to assess effects of habitat variation on microwear (each species was sampled from forests that differ in the degree of disturbance from highly disturbed to minimally disturbed). Results indicate that microwear texture values vary between habitats-more for the capuchins than the howler monkeys. This is consistent with the notion that diets of the more folivorous A. belzebul are less affected by habitat disturbance than those of the more frugivorous S. apella. It also suggests that microwear holds the potential to reflect comparatively subtle differences in within-species variation in fossil taxa if sample size and control over paleohabitat allow.

The developmental impacts of natural selection on human pelvic morphology.

Evolutionary responses to selection for bipedalism and childbirth have shaped the human pelvis, a structure that differs substantially from that in apes. Morphology related to these factors is present by birth, yet the developmental-genetic mechanisms governing pelvic shape remain largely unknown. Here, we pinpoint and characterize a key gestational window when human-specific pelvic morphology becomes recognizable, as the ilium and the entire pelvis acquire traits essential for human walking and birth. We next use functional genomics to molecularly characterize chondrocytes from different pelvic subelements during this window to reveal their developmental-genetic architectures. We then find notable evidence of ancient selection and genetic constraint on regulatory sequences involved in ilium expansion and growth, findings complemented by our phenotypic analyses showing that variation in iliac traits is reduced in humans compared to African apes. Our datasets provide important resources for musculoskeletal biology and begin to elucidate developmental mechanisms that shape human-specific morphology.

Joint disease-specificity at the regulatory base-pair level.

Given the pleiotropic nature of coding sequences and that many loci exhibit multiple disease associations, it is within non-coding sequence that disease-specificity likely exists. Here, we focus on joint disorders, finding among replicated loci, that GDF5 exhibits over twenty distinct associations, and we identify causal variants for two of its strongest associations, hip dysplasia and knee osteoarthritis. By mapping regulatory regions in joint chondrocytes, we pinpoint two variants (rs4911178; rs6060369), on the same risk haplotype, which reside in anatomical site-specific enhancers. We show that both variants have clinical relevance, impacting disease by altering morphology. By modeling each variant in humanized mice, we observe joint-specific response, correlating with GDF5 expression. Thus, we uncouple separate regulatory variants on a common risk haplotype that cause joint-specific disease. By broadening our perspective, we finally find that patterns of modularity at GDF5 are also found at over three-quarters of loci with multiple GWAS disease associations.

Genetics of scapula and pelvis development: An evolutionary perspective.

In tetrapods, the scapular and pelvic girdles perform the important function of anchoring the limbs to the trunk of the body and facilitating the movement of each appendage. This shared function, however, is one of relatively few similarities between the scapula and pelvis, which have significantly different morphologies, evolutionary histories, embryonic origins, and underlying genetic pathways. The scapula evolved in jawless fish prior to the pelvis, and its embryonic development is unique among bones in that it is derived from multiple progenitor cell populations, including the dermomyotome, somatopleure, and neural crest. Conversely, the pelvis evolved several million years later in jawed fish, and it develops from an embryonic somatopleuric cell population. The genetic networks controlling the formation of the pelvis and scapula also share similarities and differences, with a number of genes shaping only one or the other, while other gene products such as PBX transcription factors act as hierarchical developmental regulators of both girdle structures. Here, we provide a detailed review of the cellular processes and genetic networks underlying pelvis and scapula formation in tetrapods, while also highlighting unanswered questions about girdle evolution and development.

Meta-Analysis of Genomewide Association Studies Reveals Genetic Variants for Hip Bone Geometry.

Hip geometry is an important predictor of fracture. We performed a meta-analysis of GWAS studies in adults to identify genetic variants that are associated with proximal femur geometry phenotypes. We analyzed four phenotypes: (i) femoral neck length; (ii) neck-shaft angle; (iii) femoral neck width, and (iv) femoral neck section modulus, estimated from DXA scans using algorithms of hip structure analysis. In the Discovery stage, 10 cohort studies were included in the fixed-effect meta-analysis, with up to 18,719 men and women ages 16 to 93 years. Association analyses were performed with ∼2.5 million polymorphisms under an additive model adjusted for age, body mass index, and height. Replication analyses of meta-GWAS significant loci (at adjusted genomewide significance [GWS], threshold p ≤ 2.6 × 10-8 ) were performed in seven additional cohorts in silico. We looked up SNPs associated in our analysis, for association with height, bone mineral density (BMD), and fracture. In meta-analysis (combined Discovery and Replication stages), GWS associations were found at 5p15 (IRX1 and ADAMTS16); 5q35 near FGFR4; at 12p11 (in CCDC91); 11q13 (near LRP5 and PPP6R3 (rs7102273)). Several hip geometry signals overlapped with BMD, including LRP5 (chr. 11). Chr. 11 SNP rs7102273 was associated with any-type fracture (p = 7.5 × 10-5 ). We used bone transcriptome data and discovered several significant eQTLs, including rs7102273 and PPP6R3 expression (p = 0.0007), and rs6556301 (intergenic, chr.5 near FGFR4) and PDLIM7 expression (p = 0.005). In conclusion, we found associations between several genes and hip geometry measures that explained 12% to 22% of heritability at different sites. The results provide a defined set of genes related to biological pathways relevant to BMD and etiology of bone fragility. © 2019 American Society for Bone and Mineral Research.

The role of Gdf5 regulatory regions in development of hip morphology.

Given GDF5 involvement in hip development, and osteoarthritis (OA) and developmental hip dysplasia (DDH) risk, here we sought to assess the role(s) of GDF5 and its regulatory sequence on the development of hip morphology linked to injury risk. The brachypodism (bp) mouse, which harbors a Gdf5 inactivating mutation, was used to survey how Gdf5 loss of function impacts the development of hip morphology. Two transgenic Gdf5 reporter BAC lines were used to assess the spatiotemporal expression of Gdf5 regulatory sequences. Each BAC line was also used to assess the functional roles of upstream and downstream sequence on hip morphology. bp/bp mice had shorter femora with smaller femoral heads and necks as well as larger alpha angles, smaller anterior offsets, and smaller acetabula, compared to bp/+ mice (p<0.04). Regulatory sequences downstream of Gdf5 drove strong prenatal (E17) expression and low postnatal (6 months) expression across regions of femoral head and acetabulum. Conversely, upstream regulatory sequences drove very low expression at E17 and no detectable expression at 6 months. Importantly, downstream, but not upstream Gdf5 regulatory sequences fully restored all the key morphologic features disrupted in bp/bp mice. Hip morphology is profoundly affected by Gdf5 absence, and downstream regulatory sequences mediate its effects by controlling Gdf5 expression during development. This downstream region contains numerous enhancers harboring risk variants related to hip OA, DDH, and dislocation. We posit that subtle alterations to morphology driven by changes in downstream regulatory sequence underlie this locus' role in hip injury risk.