Apolipoprotein E Gene Revisited: Contribution of Rare Variants to Alzheimer's Disease Susceptibility in Southern Chinese.

BACKGROUND: APOE ε4 is the best-known risk factor for late-onset alzheimer's disease (AD). Population studies have demonstrated a relatively low prevalence of APOE ε4 among Chinese population, implying additional risk factors that are Chinese-specific may exist. Apart from - alleles, genetic variation profile along the full-length APOE has rarely been investigated. OBJECTIVE: In this study, we filled this gap by comprehensively determining all genetic variations in APOE and investigated their potential associations with late-onset AD and mild cognitive impairment (MCI) in southern Chinese. METHODS: Two hundred and fifty-seven southern Chinese participants were recruited, of whom 69 were AD patients, 83 had MCI, and 105 were normal controls. Full-length APOE from promoter to 3'UTR regions were sequenced. Genetic variants were identified and compared among the three groups. RESULTS: While APOE ε4 was more significantly found in AD patients, the prevalence of APOE ε4 in southern Chinese AD patients was the lowest when compared to other areas of China and nearby regions, as well as other countries worldwide. We further identified 13 rare non-singleton variants in APOE. Significantly more AD patients carried any of the rare non-singleton variants than MCI and normal subjects. Such difference was observed in the non-carriers of ε4-allele only. Among the identified rare variants, the potential functional impact was predicted for rs532314089, rs553874843, rs533904656 and rs370594287. CONCLUSION: Our study suggests an ethnic difference in genetic risk composition of AD in southern Chinese. Rare variants on APOE are a potential candidate for AD risk stratification biomarker in addition to APOE-ε4.

DIPPER, a spatiotemporal proteomics atlas of human intervertebral discs for exploring ageing and degeneration dynamics.

The spatiotemporal proteome of the intervertebral disc (IVD) underpins its integrity and function. We present DIPPER, a deep and comprehensive IVD proteomic resource comprising 94 genome-wide profiles from 17 individuals. To begin with, protein modules defining key directional trends spanning the lateral and anteroposterior axes were derived from high-resolution spatial proteomes of intact young cadaveric lumbar IVDs. They revealed novel region-specific profiles of regulatory activities and displayed potential paths of deconstruction in the level- and location-matched aged cadaveric discs. Machine learning methods predicted a 'hydration matrisome' that connects extracellular matrix with MRI intensity. Importantly, the static proteome used as point-references can be integrated with dynamic proteome (SILAC/degradome) and transcriptome data from multiple clinical samples, enhancing robustness and clinical relevance. The data, findings, and methodology, available on a web interface (http://www.sbms.hku.hk/dclab/DIPPER/), will be valuable references in the field of IVD biology and proteomic analytics.

The backbone of vertebrate animals consists of a series of bones called vertebrae that are joined together by disc-like structures that allow the back to move and distribute forces to protect it during daily activities. It is common for these intervertebral discs to degenerate with age, resulting in back pain and severely reducing quality of life. The mechanical features of intervertebral discs are the result of their proteins. These include extracellular matrix proteins, which form the external scaffolding that binds cells together in a tissue, and signaling proteins, which allow cells to communicate. However, how the levels of different proteins in each region of the disc vary with time has not been fully examined. To establish how protein composition changes with age, Tam, Chen et al. quantified the protein levels and gene activity (which leads to protein production) of intervertebral discs from young and old deceased individuals. They found that the position of different mixtures of proteins in the intervertebral disc changes with age, and that young people have high levels of extracellular matrix proteins and signaling proteins. Levels of these proteins decreased as people got older, as did the amount of proteins produced. To determine which region of the intervertebral disc different proteins were in, Tam, Chen et al. also performed magnetic resonance imaging (MRI) of the samples to correlate image intensity (which represents water content) with the corresponding protein signature. The data obtained provides a high-quality map of how the location of different proteins changes with age, and is available online under the name DIPPER. This database is an informative resource for research into skeletal biology, and it will likely advance the understanding of intervertebral disc degeneration in humans and animals, potentially leading to the development of new treatment strategies for this condition.

Population-Wide Genetic Risk Prediction of Complex Diseases: A Pilot Feasibility Study in Macau Population for Precision Public Healthcare Planning.

The genetic bases of many common diseases have been identified through genome-wide association studies in the past decade. However, the application of this approach on public healthcare planning has not been well established. Using Macau with population of around 650,000 as a basis, we conducted a pilot study to evaluate the feasibility of population genomic research and its potential on public health decisions. By performing genome-wide SNP genotyping of over a thousand Macau individuals, we evaluated the population genetic risk profiles of 47 non-communicable diseases and traits, as well as two traits associated with influenza infection. We found that for most of the diseases, the genetic risks of Macau population were different from those of Caucasian, but with similar profile with mainland Chinese. We also identified a panel of diseases that Macau population may have a high or elevated genetic risks. This pilot study showed that (1) population genomic study is feasible in Asian regions like Macau; (2) Macau may have different profile of population-based genetic risks than Caucasians, (3) the different prevalence of genetic risk profile indicates the importance of Asian-specific studies for Asian populations; and (4) the results generated may have an impact for going forward healthcare planning.

Prevalence, Patterns, and Genetic Association Analysis of Modic Vertebral Endplate Changes.

STUDY DESIGN: A prospective genetic association study. PURPOSE: The etiology of Modic changes (MCs) is unclear. Recently, the role of genetic factors in the etiology of MCs has been evaluated. However, studies with a larger patient subset are lacking, and candidate genes involved in other disc degeneration phenotypes have not been evaluated. We studied the prevalence of MCs and genetic association of 41 candidate genes in a large Indian cohort. OVERVIEW OF LITERATURE: MCs are vertebral endplate signal changes predominantly observed in the lumbar spine. A significant association between MCs and lumbar disc degeneration and nonspecific low back pain has been described, with the etiopathogenesis implicating various mechanical, infective, and biochemical factors. METHODS: We studied 809 patients using 1.5-T magnetic resonance imaging to determine the prevalence, patterns, distribution, and type of lumbar MCs. Genetic association analysis of 71 single nucleotide polymorphisms (SNPs) of 41 candidate genes was performed based on the presence or absence of MCs. SNPs were genotyped using the Sequenome platform, and an association test was performed using PLINK software. RESULTS: The mean age of the study population (n=809) was 36.7±10.8 years. Based on the presence of MCs, the cohort was divided into 702 controls and 107 cases (prevalence, 13%). MCs were more commonly present in the lower (149/251, 59.4%) than in the upper (102/251, 40.6%) endplates. L4-5 endplates were the most commonly affected levels (30.7%). Type 2 MCs were the most commonly observed pattern (n=206, 82%). The rs2228570 SNP of VDR (p=0.02) and rs17099008 SNP of MMP20 (p=0.03) were significantly associated with MCs. CONCLUSIONS: Genetic polymorphisms of SNPs of VDR and MMP20 were significantly associated with MCs. Understanding the etiopathogenetic mechanisms of MCs is important for planning preventive and therapeutic strategies.

Proteomic Analysis of Amyloid Corneal Aggregates from TGFBI-H626R Lattice Corneal Dystrophy Patient Implicates Serine-Protease HTRA1 in Mutation-Specific Pathogenesis of TGFBIp.

TGFBI-associated corneal dystrophies are inherited disorders caused by TGFBI gene variants that promote deposition of mutant protein (TGFBIp) as insoluble aggregates in the cornea. Depending on the type and position of amino acid substitution, the aggregates may be amyloid fibrillar, amorphous globular or both, but the molecular mechanisms that drive these different patterns of aggregation are not fully understood. In the current study, we report the protein composition of amyloid corneal aggregates from lattice corneal dystrophy patients of Asian origin with H626R and R124C mutation and compared it with healthy corneal tissues via LC-MS/MS. We identified several amyloidogenic, nonfibrillar amyloid associated proteins and TGFBIp as the major components of the deposits. Our data indicates that apolipoprotein A-IV, apolipoprotein E, and serine protease HTRA1 were significantly enriched in patient deposits compared to healthy controls. HTRA1 was also found to be 7-fold enriched in the amyloid deposits of patients compared to the controls. Peptides sequences (G511DNRFSMLVAAIQSAGLTETLNR533 and Y571HIGDEILVSGGIGALVR588) derived from the fourth FAS-1 domain of TGFBIp were enriched in the corneal aggregates in a mutation-specific manner. Biophysical studies of these two enriched sequences revealed high propensity to form amyloid fibrils under physiological conditions. Our data suggests a possible proteolytic processing mechanism of mutant TGFBIp by HTRA1 and peptides generated by mutant protein may form the ß-amyloid core of corneal aggregates in dystrophic patients.

How Reliable Are the Reported Genetic Associations in Disc Degeneration?: The Influence of Phenotypes, Age, Population Size, and Inclusion Sequence in 809 Patients.

STUDY DESIGN: Prospective genetic association study. OBJECTIVE: The aim of this study was to document the variations in the genetic associations, when different magnetic resonance imaging (MRI) phenotypes, age stratification, cohort size, and sequence of cohort inclusion are varied in the same study population. SUMMARY OF BACKGROUND DATA: Genetic associations with disc degeneration have shown high inconsistency, generally attributed to hereditary factors and ethnic variations. However, the effect of different phenotypes, size of the study population, age of the cohort, etc have not been documented clearly. METHODS: Seventy-one single-nucleotide polymorphisms (SNPs) of 41 candidate genes were correlated to six MRI markers of disc degeneration (annular tears, Pfirmann grading, Schmorl nodes, Modic changes, Total Endplate Damage score, and disc bulge) in 809 patients with back pain and/or sciatica. In the same study group, the correlations were then retested for different age groups, different sample, size and sequence of subject inclusion (first 404 and the second 405) and the differences documented. RESULTS: The mean age of population (M: 455, F: 354) was 36.7 ±â€Š10.8 years. Different genetic associations were found with different phenotypes: disc bulge with three SNPs of CILP; annular tears with rs2249350 of ADAMTS5 and rs11247361 IGF1R; modic changes with VDR and MMP20; Pfirmann grading with three SNPs of MMP20 and Schmorl node with SNPs of CALM1 and FN1 and none with Total End Plate Score.Subgroup analysis based on three age groups and dividing the total population into two groups also completely changed the associations for all the six radiographic parameters. CONCLUSION: In the same study population, SNP associations completely change with different phenotypes. Variations in age, inclusion sequence, and sample size resulted in change of genetic associations. Our study questions the validity of previous studies and necessitates the need for standardizing the description of disc degeneration, phenotype selection, study sample size, age, and other variables in future studies. LEVEL OF EVIDENCE: 4.

Genetic susceptibility of lumbar degenerative disc disease in young Indian adults.

PURPOSE: Although the exact mechanisms that lead to degenerative disc disease (DDD) are not well understood, a significant genetic influence has been found. Focusing on DDD that occurs in young adults can be valuable in determining the exact role of genetic predisposition to DDD. METHODS: Patients (<40 years) with lumbar disc degeneration were evaluated with MRI imaging (1.5 Tesla) and genetic association analysis for 58 single nucleotide polymorphism (SNP) of 35 candidate genes was performed. Disc degeneration of individual discs of lumbar spine from L1 to S1 was graded by Pfirrmann's grading. The subjects were stratified into two groups based on Total Disc Degenerative Score (TDDS). Based on TDDS, the severity of DDD was classified as mild (Group A: TDDS <10) and severe (Group B: TDDS >10). RESULTS: 695 Indian subjects including 308 with mild TDDS and 387 with severe TDDS were studied. The mean age of the patients was 29.6 ± 6.9 years in group A and 31.7 ± 6.1 in group B (p < 0.05). Five of the 35 candidate genes viz., rs1337185 of COL11A (p = 0.02), rs5275 (p = 0.03) and rs5277 (p = 0.05) of COX2, rs7575934 of IL1F5 (p = 0.04), rs3213718 of CALM1 (p = 0.04) and rs162509 of ADAMTS5 (p = 0.04) were found to be significantly associated with severe TDDS. CONCLUSION: The study identifies specific SNP associations of five genes in young adults with severe lumbar disc degeneration. These five genes (COL11A1, ADAMTS5, CALM1, IL1F5 and COX2) have different functions in the matrix metabolism, intracellular signalling and inflammatory cascade. This shows that disc degeneration is a complex disease with an intricate interplay of multiple genetic polymorphisms.

Detection of extracellular matrix degradation in intervertebral disc degeneration by diffusion magnetic resonance spectroscopy.

PURPOSE: To investigate whether diffusion magnetic resonance spectroscopy (MRS) can detect the extracellular matrix (ECM) degradation during intervertebral disc degeneration (IVDD) by the increased mobility of ECM macromolecules such as proteoglycans and collagens. METHODS: Fresh bovine intervertebral discs were injected with papain solution to induce ECM degradation. The apparent diffusion coefficients (ADCs), T2 values, and contents of ECM macromolecules and water resonances were measured longitudinally in the nucleus pulposus. RESULTS: The macromolecule ADCs increased drastically at day 1 after papain injection, and continued increasing for 5 days. In contrast, the proteoglycan content exhibited a small and slow decrease after injection while the macromolecule T2 values, water T2, ADC, and content showed slight increase or no change. The protein gel electrophoresis analysis confirmed the gradually increased ECM fragmentation in accordance with the observed macromolecule ADC increases. CONCLUSION: Diffusion MRS provides a new method to characterize the ECM degradation processes directly and sensitively. Macromolecule ADCs offer a potentially more sensitive and earlier marker for ECM degradation than the proteoglycan content and T2, and water MR properties during early IVDD. Such diffusion approach offers the possibility to directly monitor ECM integrity and degradation processes in vivo at molecular and microstructural levels in both preclinical and clinical settings.

Congenital respiratory epithelial cysts of the orbit: a rare cause of major orbital impairment.

Congenital respiratory epithelial cysts of the orbit are rare lesions with few reported cases. Extensive disease may cause an orbital apex syndrome, resulting in significant visual loss. Two such cases were described, the risk factors associated with surgical morbidity were identified, and the embryological origins of these cysts were reviewed.

Understanding the Basis of Genetic Studies: Adolescent Idiopathic Scoliosis as an Example.

STUDY DESIGN: A review of the general concepts of genetics studies with specific reference to adolescent idiopathic scoliosis (AIS). OBJECTIVES: To equip the average spine surgeon with the vocabulary and understanding needed to understand the genetics of scoliosis and the approaches used to identify risk genes. SUMMARY OF BACKGROUND DATA: Adolescent idiopathic scoliosis is a multifactorial disease. Increasing evidence from families and monozygotic twins suggests the involvement of genetic factors. An estimation of heritability also indicates a strong influence of genetics on the disease. Increasing focus has been placed on identifying genes and genetic variants associated with AIS. REVIEW: This is a review of genes and genetic variations, the phenotype definition of AIS in genetics studies, concepts and approaches to identifying associated genes, and the evaluation of results. Different types of genetic variations are present in the genome. These variations may modulate the expression or function of protein products, which in turn alter individuals' susceptibility to disease. Identifying the variants related to AIS requires an objective and clearly defined phenotype, among which the Cobb angle is commonly used. The phenotype helps classify subjects into cases and controls. By selecting candidate genes of growth factors and hormonal receptors, which are speculated to be involved in the mechanism of disease, the variants within these genes were compared between cases and controls to identify any differences. Another approach was to use large families and inspect the co-segregation of variants and phenotypes. Recently, arrays covering the variants of the whole genome were developed and assist in high-throughput screening for associated genes. CONCLUSIONS: Genetic factors have an important role in AIS. Deciphering the genes and genetic variants associated with AIS can improve our understanding of the mechanisms of the disease, as well as assist in designing treatment methods and preventive measures.

Lumbar disc degeneration is linked to a carbohydrate sulfotransferase 3 variant.

Lumbar disc degeneration (LDD) is associated with both genetic and environmental factors and affects many people worldwide. A hallmark of LDD is loss of proteoglycan and water content in the nucleus pulposus of intervertebral discs. While some genetic determinants have been reported, the etiology of LDD is largely unknown. Here we report the findings from linkage and association studies on a total of 32,642 subjects consisting of 4,043 LDD cases and 28,599 control subjects. We identified carbohydrate sulfotransferase 3 (CHST3), an enzyme that catalyzes proteoglycan sulfation, as a susceptibility gene for LDD. The strongest genome-wide linkage peak encompassed CHST3 from a Southern Chinese family­based data set, while a genome-wide association was observed at rs4148941 in the gene in a meta-analysis using multiethnic population cohorts. rs4148941 lies within a potential microRNA-513a-5p (miR-513a-5p) binding site. Interaction between miR-513a-5p and mRNA transcribed from the susceptibility allele (A allele) of rs4148941 was enhanced in vitro compared with transcripts from other alleles. Additionally, expression of CHST3 mRNA was significantly reduced in the intervertebral disc cells of human subjects carrying the A allele of rs4148941. Together, our data provide new insights into the etiology of LDD, implicating an interplay between genetic risk factors and miRNA.

Association of the asporin D14 allele with lumbar-disc degeneration in Asians.

Lumbar-disc degeneration (LDD) is a polygenic disease. Susceptibility genes reported so far are mainly extracellular matrix proteins. D14 allele of asporin (ASPN) is associated with osteoarthritis (OA). Candidate-gene association studies showed that the D14 allele is also significantly associated with LDD in Chinese and Japanese individuals. Meta-analysis showed that individuals harboring a D14 allele had higher risk with a summary odds ratio of 1.70 (p = 0.000013). ASPN expression in vertebral discs increased with age and degeneration. Our results indicate ASPN is a LDD gene in Asians, and common risk factors may be considered for OA and LDD.

Expression of the Trp2 allele of COL9A2 is associated with alterations in the mechanical properties of human intervertebral discs.

STUDY DESIGN: Biomechanical study into the association between genetic polymorphism in COL9A2 and mechanical properties of human nucleus pulposus. OBJECTIVE: To examine whether there is an association between genetic polymorphism in a structural gene, and alterations in the mechanical properties of the intervertebral discs that may predispose to disc degeneration. SUMMARY OF BACKGROUND DATA: Genetic studies have demonstrated that a polymorphism (Trp2 allele) in COL9A2 coding for alpha2 chain of collagen IX predisposes the individual to disc degeneration. The mechanism of this predisposition is not known. METHODS: Blood and whole disc samples were retrieved from adolescents and young adults during scoliosis surgery, degenerated discs were retrieved from patients with back pain during anterior spinal fusion. Anulus fibrosus and nucleus pulposus from a set of the scoliosis discs were used to perform immunohistochemistry to demonstrate the presence of collagen IX in the scoliosis discs. For the remaining samples, DNA was extracted from blood to determine the Trp2 status by sequencing. Nondegenerated (Trp2-), nondegenerated (Trp2+), and degenerated (Trp2-) nucleus pulposus samples were tested in confined compression. Swelling pressure and compressive modulus were measured and compared between groups. RESULTS: Positive staining of collagen IX was detected in both anulus fibrosus and nucleus pulposus sections confirming its presence in the scoliosis discs. The mean swelling pressure and compressive modulus values of 6 nondegenerated (Trp2+) samples (swelling pressure = 0.0019 MPa, compressive modulus = 0.97 MPa) were significantly lower (P < 0.05) than those of the 6 nondegenerated (Trp2-) samples (swelling pressure = 0.015 MPa; compressive modulus = 1.89 MPa). CONCLUSION: This is the first study to demonstrate an association between the Trp2 allele and disc mechanics, thus relating genetic variations and debilitating mechanical alterations that may ultimately result in intervertebral disc degeneration.