Multivariate genome-wide association analysis identifies novel and relevant variants associated with anterior cruciate ligament rupture risk in the dog model.

BACKGROUND: Anterior cruciate ligament rupture (ACLR) is a debilitating and potentially life-changing condition in humans, as there is a high prevalence of early-onset osteoarthritis after injury. Identification of high-risk individuals before they become patients is important, as post-treatment lifetime burden of ACLR in the USA ranges from $7.6 to $17.7 billion annually. ACLR is a complex disease with multiple risk factors including genetic predisposition. Naturally occurring ACLR in the dog is an excellent model for human ACLR, as risk factors and disease characteristics in humans and dogs are similar. In a univariate genome-wide association study (GWAS) of 237 Labrador Retrievers, we identified 99 ACLR candidate loci. It is likely that additional variants remain to be identified. Joint analysis of multiple correlated phenotypes is an underutilized technique that increases statistical power, even when only one phenotype is associated with the trait. Proximal tibial morphology has been shown to affect ACLR risk in both humans and dogs. In the present study, tibial plateau angle (TPA) and relative tibial tuberosity width (rTTW) were measured on bilateral radiographs from purebred Labrador Retrievers that were recruited to our initial GWAS. We performed a multivariate genome wide association analysis of ACLR status, TPA, and rTTW. RESULTS: Our analysis identified 3 loci with moderate evidence of association that were not previously associated with ACLR. A locus on Chr1 associated with both ACLR and rTTW is located within ROR2, a gene important for cartilage and bone development. A locus on Chr4 associated with both ACLR and TPA resides within DOCK2, a gene that has been shown to promote immune cell migration and invasion in synovitis, an important predictor of ACLR. A third locus on Chr23 associated with only ACLR is located near a long non-coding RNA (lncRNA). LncRNA's are important for regulation of gene transcription and translation. CONCLUSIONS: These results did not overlap with our previous GWAS, which is reflective of the different methods used, and supports the need for further work. The results of the present study are highly relevant to ACLR pathogenesis, and identify potential drug targets for medical treatment.

Craniectomies for Dogs With Skull Multilobular Osteochondrosarcoma Using the Misonix Bone Scalpel: Cadaveric Evaluation and Retrospective Case Series.

To evaluate the Misonix bone scalpel (MBS) for craniotomies in dogs and describe clinical findings and surgical experience in 3 dogs with large multilobular osteochondrosarcoma (MLO) of the skull. Cadaver evaluation and retrospective case series. One canine cadaver; 3 client-owned dogs. Craniotomies of different sizes and at different locations were performed with MBS. Dural tear and bone discoloration were recorded. Clinical, imaging, and surgical findings of dogs diagnosed with MLO and where MBS was used for craniectomies were retrospectively included. Cadaveric evaluation identified MBS as an efficient tool for rapid craniectomies (>5minutes) albeit dural tears and some small foci of bone discoloration were observed. Craniectomies could be performed without complications in 3 dogs with MLO without dural tear or bone discoloration. .Excision was in complete in all cases. The short-term outcome was good, and the long-term outcome was fair to good. Piezoelectric bone surgery with the Misonix bone scalpel is an alternative technology to perform craniectomies in dogs. It was not associated with complications in 3 dogs diagnosed and surgically treated for MLO. Dural tears and suspected bone necrosis can occur. Great care should be taken when using CT to establish disease free surgical osteotomy.

Analysis of copy number variation in dogs implicates genomic structural variation in the development of anterior cruciate ligament rupture.

Anterior cruciate ligament (ACL) rupture is an important condition of the human knee. Second ruptures are common and societal costs are substantial. Canine cranial cruciate ligament (CCL) rupture closely models the human disease. CCL rupture is common in the Labrador Retriever (5.79% prevalence), ~100-fold more prevalent than in humans. Labrador Retriever CCL rupture is a polygenic complex disease, based on genome-wide association study (GWAS) of single nucleotide polymorphism (SNP) markers. Dissection of genetic variation in complex traits can be enhanced by studying structural variation, including copy number variants (CNVs). Dogs are an ideal model for CNV research because of reduced genetic variability within breeds and extensive phenotypic diversity across breeds. We studied the genetic etiology of CCL rupture by association analysis of CNV regions (CNVRs) using 110 case and 164 control Labrador Retrievers. CNVs were called from SNPs using three different programs (PennCNV, CNVPartition, and QuantiSNP). After quality control, CNV calls were combined to create CNVRs using ParseCNV and an association analysis was performed. We found no strong effect CNVRs but found 46 small effect (max(T) permutation P<0.05) CCL rupture associated CNVRs in 22 autosomes; 25 were deletions and 21 were duplications. Of the 46 CCL rupture associated CNVRs, we identified 39 unique regions. Thirty four were identified by a single calling algorithm, 3 were identified by two calling algorithms, and 2 were identified by all three algorithms. For 42 of the associated CNVRs, frequency in the population was <10% while 4 occurred at a frequency in the population ranging from 10-25%. Average CNVR length was 198,872bp and CNVRs covered 0.11 to 0.15% of the genome. All CNVRs were associated with case status. CNVRs did not overlap previous canine CCL rupture risk loci identified by GWAS. Associated CNVRs contained 152 annotated genes; 12 CNVRs did not have genes mapped to CanFam3.1. Using pathway analysis, a cluster of 19 homeobox domain transcript regulator genes was associated with CCL rupture (P = 6.6E-13). This gene cluster influences cranial-caudal body pattern formation during embryonic limb development. Clustered genes were found in 3 CNVRs on chromosome 14 (HoxA), 28 (NKX6-2), and 36 (HoxD). When analysis was limited to deletion CNVRs, the association was strengthened (P = 8.7E-16). This study suggests a component of the polygenic risk of CCL rupture in Labrador Retrievers is associated with small effect CNVs and may include aspects of stifle morphology regulated by homeobox domain transcript regulator genes.

Influence of trial repetition on lameness during force platform gait analysis in a heterogeneous population of clinically lame dogs each trotting at its preferred velocity.

OBJECTIVE To determine variance effects influencing ground reaction forces (GRFs) in a heterogeneous population of lame dogs during trotting. ANIMALS 30 client-owned dogs with thoracic limb lameness and 31 dogs with pelvic limb lameness. PROCEDURES GRFs, velocity, height at the dorsal aspect of the scapulae (ie, withers), and shoulder height were obtained. Each dog was trotted across a force platform at its preferred velocity. Variance effects for 12 velocity and associated relative velocity (V*) ranges were examined. RESULTS Individual dog, velocity, V*, and limb significantly influenced GRFs. Withers height V* ranges were associated with small variance in GRFs, but all absolute and V* ranges were associated with significant effects for all 4 limbs and both types of lameness. Significant changes in lame limb GRFs and velocity in ipsilateral trials in dogs with thoracic limb and pelvic limb lameness were evident with trial repetition. Withers height V* range of 0.55 to 0.93 captured a large proportion of trials (> 90%) in dogs with thoracic limb or pelvic limb lameness, with limited effects on peak vertical force and vertical impulse. CONCLUSIONS AND CLINICAL RELEVANCE Trial repetition caused alterations to GRFs and subject velocity that may have confounded assessment of lameness, which supported the concept that a priori selection of a velocity or V* range for force platform gait analysis should use a range that captures valid trials efficiently while minimizing GRF variance. These ranges typically would span the preferred velocity of subject dogs, such as withers height V* of 0.55 to 0.93.

Variance associated with walking velocity during force platform gait analysis of a heterogeneous sample of clinically normal dogs.

OBJECTIVE To determine whether walking at specific ranges of absolute and relative (V*) velocity would aid efficient capture of gait trial data with low ground reaction force (GRF) variance in a heterogeneous sample of dogs. ANIMALS 17 clinically normal dogs of various breeds, ages, and sexes. PROCEDURES Each dog was walked across a force platform at its preferred velocity, with controlled acceleration within 0.5 m/s2. Ranges in V* were created for height at the highest point of the shoulders (withers; WHV*). Variance effects from 8 walking absolute velocity ranges and associated WHV* ranges were examined by means of repeated-measures ANCOVA. RESULTS The individual dog effect provided the greatest contribution to variance. Narrow velocity ranges typically resulted in capture of a smaller percentage of valid trials and were not consistently associated with lower variance. The WHV* range of 0.33 to 0.46 allowed capture of valid trials efficiently, with no significant effects on peak vertical force and vertical impulse. CONCLUSIONS AND CLINICAL RELEVANCE Dogs with severe lameness may be unable to trot or may have a decline in mobility with gait trial repetition. Gait analysis involving evaluation of individual dogs at their preferred absolute velocity, such that dogs are evaluated at a similar V*, may facilitate efficient capture of valid trials without significant effects on GRF. Use of individual velocity ranges derived from a WHV* range of 0.33 to 0.46 can account for heterogeneity and appears suitable for use in clinical trials involving dogs at a walking gait.

Genome-wide association analysis in dogs implicates 99 loci as risk variants for anterior cruciate ligament rupture.

Anterior cruciate ligament (ACL) rupture is a common condition that can be devastating and life changing, particularly in young adults. A non-contact mechanism is typical. Second ACL ruptures through rupture of the contralateral ACL or rupture of a graft repair is also common. Risk of rupture is increased in females. ACL rupture is also common in dogs. Disease prevalence exceeds 5% in several dog breeds, ~100 fold higher than human beings. We provide insight into the genetic etiology of ACL rupture by genome-wide association study (GWAS) in a high-risk breed using 98 case and 139 control Labrador Retrievers. We identified 129 single nucleotide polymorphisms (SNPs) within 99 risk loci. Associated loci (P<5E-04) explained approximately half of phenotypic variance in the ACL rupture trait. Two of these loci were located in uncharacterized or non-coding regions of the genome. A chromosome 24 locus containing nine genes with diverse functions met genome-wide significance (P = 3.63E-0.6). GWAS pathways were enriched for c-type lectins, a gene set that includes aggrecan, a gene set encoding antimicrobial proteins, and a gene set encoding membrane transport proteins with a variety of physiological functions. Genotypic risk estimated for each dog based on the risk contributed by each GWAS locus showed clear separation of ACL rupture cases and controls. Power analysis of the GWAS data set estimated that ~172 loci explain the genetic contribution to ACL rupture in the Labrador Retriever. Heritability was estimated at 0.48. We conclude ACL rupture is a moderately heritable highly polygenic complex trait. Our results implicate c-type lectin pathways in ACL homeostasis.

Color-encoded distance visualization of cranial nerve-vessel contacts.

PURPOSE: Visualization of pathological contact between cranial nerves and vascular structures at the surface of the brainstem is important for diagnosis and treatment of neurovascular compression (NVC) syndromes. We developed a method for improved visualization of this abnormality. METHODS: Distance fields were computed using preoperative MRI scans of individuals with NVC syndromes to support the topological representation of brainstem surface structures with quantitative information. Polygonal models of arteries, cranial nerves and the brainstem were generated using segmented T2 weighted MR data. After color-coding the polygonal models with the respective distances, enhanced color visualization of vessel-nerve locations with possible contacts was achieved. RESULTS: The proposed method was implemented and applied to surgical planning in a dozen cases of NVC syndrome. Two selected cases were chosen to demonstrate the feasibility and subjective improvement provided by our visualization technique. Expert neurosurgeons found the improvement valuable and useful for these cases. CONCLUSION: Color-encoded distance information significantly improves the perceptibility of potential nerve-vessel contacts. This method contributes to a better understanding of the complex anatomical situation at the surface of the brainstem and assists in planning of surgery.