Immunolipidomics Reveals a Globoside Network During the Resolution of Pro-Inflammatory Response in Human Macrophages.

Toll-like receptor 4 (TLR4)-mediated changes in macrophages reshape intracellular lipid pools to coordinate an effective innate immune response. Although this has been previously well-studied in different model systems, it remains incompletely understood in primary human macrophages. Here we report time-dependent lipidomic and transcriptomic responses to lipopolysaccharide (LPS) in primary human macrophages from healthy donors. We grouped the variation of ~200 individual lipid species measured by LC-MS/MS into eight temporal clusters. Among all other lipids, glycosphingolipids (glycoSP) and cholesteryl esters (CE) showed a sharp increase during the resolution phase (between 8h or 16h post LPS). GlycoSP, belonging to the globoside family (Gb3 and Gb4), showed the greatest inter-individual variability among all lipids quantified. Integrative network analysis between GlycoSP/CE levels and genome-wide transcripts, identified Gb4 d18:1/16:0 and CE 20:4 association with subnetworks enriched for T cell receptor signaling (PDCD1, CD86, PTPRC, CD247, IFNG) and DC-SIGN signaling (RAF1, CD209), respectively. Our findings reveal Gb3 and Gb4 globosides as sphingolipids associated with the resolution phase of inflammatory response in human macrophages.

MS-based lipidomics of human blood plasma: a community-initiated position paper to develop accepted guidelines.

Human blood is a self-regenerating lipid-rich biological fluid that is routinely collected in hospital settings. The inventory of lipid molecules found in blood plasma (plasma lipidome) offers insights into individual metabolism and physiology in health and disease. Disturbances in the plasma lipidome also occur in conditions that are not directly linked to lipid metabolism; therefore, plasma lipidomics based on MS is an emerging tool in an array of clinical diagnostics and disease management. However, challenges exist in the translation of such lipidomic data to clinical applications. These relate to the reproducibility, accuracy, and precision of lipid quantitation, study design, sample handling, and data sharing. This position paper emerged from a workshop that initiated a community-led process to elaborate and define a set of generally accepted guidelines for quantitative MS-based lipidomics of blood plasma or serum, with harmonization of data acquired on different instrumentation platforms across independent laboratories as an ultimate goal. We hope that other fields may benefit from and follow such a precedent.

Defining neurodegeneration on Guam by targeted genomic sequencing.

OBJECTIVE: Amyotrophic lateral sclerosis/parkinsonism-dementia complex has been described in Guam, Western Papua, and the Kii Peninsula of Japan. The etiology and pathogenesis of this complex neurodegenerative disease remains enigmatic. METHODS: In this study, we have used targeted genomic sequencing to evaluate the contribution of genetic variability in the pathogenesis of amyotrophic lateral sclerosis, parkinsonism, and dementia in Guamanian Chamorros. RESULTS: Genes previously linked to or associated with amyotrophic lateral sclerosis, parkinsonism, dementia, and related neurodegenerative syndromes were sequenced in Chamorro subjects living in the Mariana Islands. Homozygous PINK1 p.L347P, heterozygous DCTN1 p.T54I, FUS p.P431L, and HTT (42 CAG repeats) were identified as pathogenic mutations. INTERPRETATION: The findings explain the clinical, pathologic, and genetic heterogeneity observed in some multi-incident families and contribute to the excess incidence of neurodegeneration previously reported on Guam.

Genetics and genomics of Parkinson's disease.

Parkinson's disease (PD) is a progressively debilitating neurodegenerative syndrome. Although best described as a movement disorder, the condition has prominent autonomic, cognitive, psychiatric, sensory and sleep components. Striatal dopaminergic innervation and nigral neurons are progressively lost, with associated Lewy pathology readily apparent on autopsy. Nevertheless, knowledge of the molecular events leading to this pathophysiology is limited. Current therapies offer symptomatic benefit but they fail to slow progression and patients continue to deteriorate. Recent discoveries in sporadic, Mendelian and more complex forms of parkinsonism provide novel insight into disease etiology; 28 genes, including those encoding alpha-synuclein (SNCA), leucine-rich repeat kinase 2 (LRRK2) and microtubule-associated protein tau (MAPT), have been linked and/or associated with PD. A consensus regarding the affected biological pathways and molecular processes has also started to emerge. In early-onset and more a typical PD, deficits in mitophagy pathways and lysosomal function appear to be prominent. By contrast, in more typical late-onset PD, chronic, albeit subtle, dysfunction in synaptic transmission, early endosomal trafficking and receptor recycling, as well as chaperone-mediated autophagy, provide a unifying synthesis of the molecular pathways involved. Disease-modification (neuroprotection) is no longer such an elusive goal given the unparalleled opportunity for diagnosis, translational neuroscience and therapeutic development provided by genetic discovery.

DNAJC13 mutations in Parkinson disease.

A Saskatchewan multi-incident family was clinically characterized with Parkinson disease (PD) and Lewy body pathology. PD segregates as an autosomal-dominant trait, which could not be ascribed to any known mutation. DNA from three affected members was subjected to exome sequencing. Genome alignment, variant annotation and comparative analyses were used to identify shared coding mutations. Sanger sequencing was performed within the extended family and ethnically matched controls. Subsequent genotyping was performed in a multi-ethnic case-control series consisting of 2928 patients and 2676 control subjects from Canada, Norway, Taiwan, Tunisia, and the USA. A novel mutation in receptor-mediated endocytosis 8/RME-8 (DNAJC13 p.Asn855Ser) was found to segregate with disease. Screening of cases and controls identified four additional patients with the mutation, of which two had familial parkinsonism. All carriers shared an ancestral DNAJC13 p.Asn855Ser haplotype and claimed Dutch-German-Russian Mennonite heritage. DNAJC13 regulates the dynamics of clathrin coats on early endosomes. Cellular analysis shows that the mutation confers a toxic gain-of-function and impairs endosomal transport. DNAJC13 immunoreactivity was also noted within Lewy body inclusions. In late-onset disease which is most reminiscent of idiopathic PD subtle deficits in endosomal receptor-sorting/recycling are highlighted by the discovery of pathogenic mutations VPS35, LRRK2 and now DNAJC13. With this latest discovery, and from a neuronal perspective, a temporal and functional ecology is emerging that connects synaptic exo- and endocytosis, vesicular trafficking, endosomal recycling and the endo-lysosomal degradative pathway. Molecular deficits in these processes are genetically linked to the phenotypic spectrum of parkinsonism associated with Lewy body pathology.

Haplotype co-segregation with attention deficit-hyperactivity disorder in unrelated German multi-generation families.

Complex disorders have proved to be elusive in the search for underlying genetic causes. In the presence of large multi-generation pedigrees with multiple affected individuals, heritable familial forms of the disorders can be postulated. Observations of particular chromosomal haplotypes shared among all affected individuals within pedigrees may reveal chromosomal regions, in which the disease-related genes may be located. Hence, the biochemical pathways involved in pathogenesis can be exposed. We have recruited eight large Attention Deficit-Hyperactivity Disorder (ADHD, OMIM: #143465) families of German descent. Densely spaced informative microsatellite markers with high heterozygosity rates were used to fine-map and haplotype chromosomal regions of interest in these families. In three subsets and one full family of the eight ADHD families, haplotypes co-segregating with ADHD-affected individuals were identified at chromosomes 1q25, 5q11-5q13, 9q31-9q32, and 18q11-18q21. Positive LOD scores supported these co-segregations. The existence of haplotypes co-segregating among affected individuals in large ADHD pedigrees suggests the existence of Mendelian forms of the disorder and that ADHD-related genes are located within these haplotypes. In depth sequencing of these haplotype regions can identify causative genetic mechanisms and will allow further insights into the clinico-genetics of this complex disorder.

The chromosome 15q14 locus for bipolar disorder and schizophrenia: is C15orf53 a major candidate gene?

Bipolar disorder (BD) and schizophrenia are complexly inherited and highly heritable disorders with currently unknown etiologies. Recently, two independent genome-wide association studies for BD identified a small region on chromosome 15q14-15.1, pointing to a locus close to the gene C15orf53. Previously, this genomic region was also found to co-segregate with periodic catatonia (SCZD10, OMIM %605419), an unsystematic schizophrenia according to Leonhard's classification, in several multiplex families, thus pointing to overlapping etiologies of both conditions. A susceptibility locus on chromosome 15q14-15.1 was narrowed down to a 4.38 Mb region in these affected families followed by mutation and segregation analyses of C15orf53. Association analysis of individuals affected by BD and/or SCZD10 (n = 274) and controls (n = 230) and expression analyses in distinct post-mortem human limbic brain tissues were conducted. C15orf53 revealed no mutations in our SCZD10 family members, but segregation of two common haplotypes was found. No association of identified haplotypes was found in our case-control samples. Gene expression could be demonstrated for immune-system-derived cells but not for the post-mortem human limbic brain tissue. Our results indicate that C15orf53 is probably neither causative for the etiology of BD nor for SCZD10 in our samples.

Characterizing the neurite outgrowth inhibitory effect of Mani.

Mani (myelin-associated neurite-outgrowth inhibitor) protein is implicated in both axonal guidance and axonal regeneration after central nervous system (CNS) injury. Here, we applied a neurite outgrowth assay, coupled with a siRNA-driven investigation and immunocytochemistry, to unveil Mani's axonal outgrowth inhibitory effect in embryonic rat cortical primary neurons in vitro. We further demonstrate Mani's neuronal localization in comparison with a principal subunit, Cdc27, of the anaphase promoting complex (APC). Considering the protein structure of Mani obtained via a series of bio-computational studies, we propose a Cdc27-Mani-APC-related signalling pathway may be involved in CNS axon regeneration.

A non-human primate system for large-scale genetic studies of complex traits.

Non-human primates provide genetic model systems biologically intermediate between humans and other mammalian model organisms. Populations of Caribbean vervet monkeys (Chlorocebus aethiops sabaeus) are genetically homogeneous and large enough to permit well-powered genetic mapping studies of quantitative traits relevant to human health, including expression quantitative trait loci (eQTL). Previous transcriptome-wide investigation in an extended vervet pedigree identified 29 heritable transcripts for which levels of expression in peripheral blood correlate strongly with expression levels in the brain. Quantitative trait linkage analysis using 261 microsatellite markers identified significant (n = 8) and suggestive (n = 4) linkages for 12 of these transcripts, including both cis- and trans-eQTL. Seven transcripts, located on different chromosomes, showed maximum linkage to markers in a single region of vervet chromosome 9; this observation suggests the possibility of a master trans-regulator locus in this region. For one cis-eQTL (at B3GALTL, beta-1,3-glucosyltransferase), we conducted follow-up single nucleotide polymorphism genotyping and fine-scale association analysis in a sample of unrelated Caribbean vervets, localizing this eQTL to a region of <200 kb. These results suggest the value of pedigree and population samples of the Caribbean vervet for linkage and association mapping studies of quantitative traits. The imminent whole genome sequencing of many of these vervet samples will enhance the power of such investigations by providing a comprehensive catalog of genetic variation.