ObjectiveCirculating lipids are linked with insulin resistance and increased cardiovascular disease risk. We previously reported that dihydroceramides, a specific type of sphingolipid, are elevated in insulin-resistant individuals; however, little is known regarding whether insulin-sensitizing lifestyle interventions can improve profiles of sphingolipids and other lipid species.MethodsA total of 21 individuals with obesity participated in a 3-month lifestyle intervention of combined weight loss and exercise training. Insulin sensitivity was assessed by hyperinsulinemic-euglycemic clamps, and serum lipidomics was conducted.ResultsFollowing the intervention, BMI was significantly reduced by 10%; VO2peak and insulin sensitivity increased by 12% and 57%, respectively; and total serum triacylglycerol (TAG), diacylglycerol, dihydroceramides, sphingosine-1-phosphate, and sphinganine-1-phosphate were significantly reduced, as were specific species of dihydroceramides (C18:0 and C24:1). Individuals with higher preintervention TAG concentrations had significant decreases in serum lipids, which were not significantly changed in individuals with lower preintervention TAG.ConclusionsThese data show that serum sphingolipid species previously linked to insulin resistance in humans can be reduced with insulin-sensitizing lifestyle interventions. Furthermore, individuals with elevated serum TAG may significantly benefit from lifestyle interventions that increase insulin sensitivity due to a greater decrease in serum lipids related to insulin resistance.

Background and aims: To prospectively investigate associations of plasma sphingolipids with insulin sensitivity, 8 -cell function, and incident diabetes in the Japanese American Community Diabetes Study. Methods and results: Baseline plasma samples from adults without diabetes (n = 349; mean age 56.7 years, 51 % men) were assayed for circulating ceramide and sphingomyelin species. Adjusted regression models examined cross-sectional and longitudinal associations with insulin sensitivity (HOMA2-%S), 8 -cell function (oral disposition index: DIo) and with incident diabetes over 5 years follow-up. Concentrations of four species (Ceramide C16:0, C18:0, C20:0, and C22:0) were inversely associated with HOMA2-%S at baseline (all P values < 0.05, Q values < 0.05) and change in HOMA2-%S over 5 years (all P values < 0.05, Q values < 0.05). No sphingolipids were associated with baseline or change in DIo. Of the four species associated with HOMA2-%S, only Ceramide C18:0 was significantly and positively associated with incident diabetes (RR/1SD 1.44, 95 % CI 1.10-1.80, P = 0.006, Q = 0.024). The association of plasma Ceramide C18:0 with the risk of diabetes was partially mediated by change in HOMA2-%S between baseline and 5 years (mediation proportion: 61.5 %, 95 % CI 21.1%-212.5 %). Conclusion: Plasma Ceramide C18:0 was associated with higher risk of incident diabetes which was partially mediated through a decrease in insulin sensitivity between baseline and five years. Circulating Ceramide C18:0 could be a potential biomarker for identifying those at risk of developing diabetes. (c) 2023 The Italian Diabetes Society, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

Understanding microglial states in the aging brain has become crucial, especially with the discovery of numerous Alzheimer's disease (AD) risk and protective variants in genes such as INPP5D and TREM2, which are essential to microglia function in AD. Here we present a thorough examination of microglia-like cells and primary mouse microglia at the proteome and transcriptome levels to illuminate the roles these genes and the proteins they encode play in various cell states. First, we compared the proteome profiles of wildtype and INPP5D (SHIP1) knockout primary microglia. Our findings revealed significant proteome alterations only in the homozygous SHIP1 knockout, revealing its impact on the microglial proteome. Additionally, we compared the proteome and transcriptome profiles of commonly used in vitro microglia BV2 and HMC3 cells with primary mouse microglia. Our results demonstrated a substantial similarity between the proteome of BV2 and mouse primary cells, while notable differences were observed between BV2 and human HMC3. Lastly, we conducted targeted lipidomic analysis to quantify different phosphatidylinositols (PIs) species, which are direct SHIP1 targets, in the HMC3 and BV2 cells. This in-depth omics analysis of both mouse and human microglia enhances our systematic understanding of these microglia models.SignificanceGiven the growing urgency of comprehending microglial function in the context of neurodegenerative diseases and the substantial therapeutic implications associated with SHIP1 modulation, we firmly believe that our study, through a rigorous and comprehensive proteomics, transcriptomics and targeted lipidomic analysis of microglia, contributes to the systematic understanding of microglial function in the context of neurodegenerative diseases.

Endoplasmic reticulum calcium (ER Ca2+) levels are primarily maintained through the sarco-ER Ca2+ ATPase (SERCA) pump. We have demonstrated reduced SERCA2 expression in islets from human cadaveric donors with Type 2 Diabetes (T2D) compared to normoglycemic controls and linked β-cell specific deletion of SERCA2 with impaired proinsulin processing. To test mechanisms of how reduced ER Ca2+ and SERCA2 deficiency regulate proinsulin processing, a SNAP-tagged human proinsulin adenovirus (SNAP-proinsulin) was overexpressed in rat insulinoma β-cells where SERCA2 had been deleted (S2KO INS-1) using CRISPR/Cas9 and in islets from C57BL/6 mice with SERCA2 haploinsufficiency fed a high fat diet or treated ex vivo with glucolipotoxicity (GLT). Immunoblotting SNAP-tagged cleavage products showed reduced proinsulin processing in SERCA2 deficient models compared to wildtype (WT), these defects were exacerbated by GLT treatment. To determine whether reduced processing could be related to impaired trafficking of proinsulin within the β-cell secretory pathway, SNAP-proinsulin was fluorescently labelled in C57BL/6 mouse islets treated with GLT. The colocalization of SNAP-proinsulin with Golgi immuno-marker, Giantin, suggested an accumulation in this compartment. The ER is the main site of lipid synthesis and lipid remodeling is known to regulate β-cell function. Preliminary mass spectrometry data revealed that loss of SERCA2 increases ER levels of phosphatidylcholine and decreases ceramide and sphingomyelin in S2KO INS-1 cells compared to WT cells. Taken together, these results demonstrate that ER Ca2+ deficiency changes ER lipid composition, which may affect proinsulin processing and trafficking. Characterizing this novel mechanism could provide valuable insight into β-cell dysfunction in T2D. Disclosure M.Hartley: Employee; Eli Lilly and Company. M.J.Pearson: None. F.Syed: None. H.Bui: None. T.Kono: None. K.D.Roth: Employee; Eli Lilly and Company. C.Evans-molina: Advisory Panel; Provention Bio, Inc., DiogenX, Avotres Inc., Neurodon, MaiCell Therapeutics, Other Relationship; Isla Technology, Bristol-Myers Squibb Company, Nimbus Therapeutics, Research Support; Lilly, Astellas Pharma Inc. Funding National Institute of Diabetes and Digestive and Kidney Diseases (R01DK093954, R01DK127308, R01DK127236)

Heterozygous variants in GBA1, encoding glucocerebrosidase (GCase), are the most common genetic risk factor for Parkinson's disease (PD). Moreover, sporadic PD patients also have a substantial reduction of GCase activity. Genetic variants of SMPD1 are also overrepresented in PD cohorts, whereas a reduction of its encoded enzyme (acid sphingomyelinase or ASM) activity is linked to an earlier age of PD onset. Despite both converging on the ceramide pathway, how the combined deficiencies of both enzymes might interact to modulate PD has yet to be explored. Therefore, we created a doubleknockout (DKO) zebrafish line for both gba1 (or gba) and smpd1 to test for an interaction in vivo, hypothesising an exacerbation of phenotypes in the DKO line compared to those for single mutants. Unexpectedly, DKO zebrafish maintained conventional swimming behaviour and had normalised neuronal gene expression signatures compared to those of single mutants. We further identified rescue of mitochondrial Complexes I and IV in DKO zebrafish. Despite having an unexpected rescue effect, our results confirm ASM as a modifier of GBA1 deficiency in vivo. Our study highlights the need for validating how genetic variants and enzymatic deficiencies may interact in vivo.

Serum ceramides, especially C16:0 and C18:0 species, are linked to CVD risk and insulin resistance, but details of this association are not well understood. We performed this study to quantify a broad range of serum sphingolipids in individuals spanning the physiologic range of insulin sensitivity and to determine if dihydroceramides cause insulin resistance in vitro. As expected, we found that serum triglycerides were significantly greater in individuals with obesity and T2D compared with athletes and lean individuals. Serum ceramides were not signifi-cantly different within groups but, using all ceramide data relative to insulin sensitivity as a continuous variable, we observed significant inverse relation-ships between C18:0, C20:0, and C22:0 species and in-sulin sensitivity. Interestingly, we found that total serum dihydroceramides and individual species were significantly greater in individuals with obesity and T2D compared with athletes and lean individuals, with C18:0 species showing the strongest inverse relationship to insulin sensitivity. Finally, we administered a physiological mix of dihydrocer-amides to primary myotubes and found decreased insulin sensitivity in vitro without changing the overall intracellular sphingolipid content, suggesting a direct effect on insulin resistance. These data extend what is known regarding serum sphingolipids and insulin resistance and show the importance of serum dihydroceramides to predict and promote in-sulin resistance in humans.

BACKGROUND Lipid alterations contribute to Alzheimer's disease (AD) pathogenesis. Lipidomics studies could help systematically characterize such alterations and identify potential biomarkers. OBJECTIVE To identify lipids associated with mild cognitive impairment and amyloid-β deposition, and to examine lipid correlation patterns within phenotype groupsMethods:Eighty plasma lipids were measured using mass spectrometry for 1,255 non-demented participants enrolled in the Mayo Clinic Study of Aging. Individual lipids associated with mild cognitive impairment (MCI) were first identified. Correlation network analysis was then performed to identify lipid species with stable correlations across conditions. Finally, differential correlation network analysis was used to determine lipids with altered correlations between phenotype groups, specifically cognitively unimpaired versus MCI, and with elevated brain amyloid versus without. RESULTS Seven lipids were associated with MCI after adjustment for age, sex, and APOE4. Lipid correlation network analysis revealed that lipids from a few species correlated well with each other, demonstrated by subnetworks of these lipids. 177 lipid pairs differently correlated between cognitively unimpaired and MCI patients, whereas 337 pairs of lipids exhibited altered correlation between patients with and without elevated brain amyloid. In particular, 51 lipid pairs showed correlation alterations by both cognitive status and brain amyloid. Interestingly, the lipids central to the network of these 51 lipid pairs were not significantly associated with either MCI or amyloid, suggesting network-based approaches could provide biological insights complementary to traditional association analyses. CONCLUSION Our attempt to characterize the alterations of lipids at network-level provides additional insights beyond individual lipids, as shown by differential correlations in our study.

Sex differences in insulin sensitivity are present throughout the lifespan, though mechanisms that account for these differences are unclear. Accumulation of diacylglycerol (DAG) and sphingolipids in skeletal muscle are thought to promote insulin resistance, and localization plays an important role. Other lipids and lipid intermediates found in muscle and plasma such as acylcarnitines also relate to insulin sensitivity. However, it is not known whether there are sex differences in accumulation of these lipids that may affect insulin sensitivity. We evaluated insulin sensitivity and sub-cellular localization of skeletal muscle DAG and sphingolipids, as well as muscle acylcarnitines and serum lipidomics, in 25 obese, nondiabetic men and women (13F). Insulin sensitivity was assessed with a hyperinsulinemic-euglycemic clamp. Muscle biopsies were taken during basal and insulin stimulated conditions and fractionated into sub-cellular compartments. Lipids were measured using LC/MS/MS. Insulin sensitivity was significantly lower in men (p=0.05), however we found no sex-differences in localization of DAG or sphingolipids in skeletal muscle. Men had higher total acylcarnitines in muscle tissue (p<0.05) and higher serum lysophosphatidylcholines (LPCs; p<0.01), and both were correlated with insulin sensitivity (r=-0.42 and -0.43, respectively; p<0.05). After correcting for multiple comparisons, the linoleoylcarnitine species (18:2AC) was significantly elevated in men during insulin stimulation (p=0.001), and was negatively associated with insulin sensitivity (r=-0.43; p<0.05). Combined, these data suggest differences in content of acylcarnitines between sexes consistent with differences in fatty acid processing and oxidation that may impact insulin sensitivity. Further, greater serum LPC species may play an important role in the decreased insulin sensitivity associated with male sex. Disclosure J.L. Broussard: None. L. Perreault: Advisory Panel; Self; Novo Nordisk A/S. Speaker's Bureau; Self; Novo Nordisk A/S. Advisory Panel; Self; Merck & Co., Inc.. Speaker's Bureau; Self; Merck & Co., Inc., AstraZeneca, Janssen Pharmaceuticals, Inc., Boehringer Ingelheim Pharmaceuticals, Inc.. Consultant; Self; Boehringer Ingelheim Pharmaceuticals, Inc.. Advisory Panel; Self; Sanofi. Speaker's Bureau; Self; Sanofi. S.A. Newsom: None. D.E. Kahn: None. A. Kerege: None. K.A. Harrison: None. B. Bergman: Research Support; Self; Eli Lilly and Company. Advisory Panel; Spouse/Partner; Novo Nordisk Inc., Merck & Co., Inc., AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Eli Lilly and Company.

AIMS:The role of ceramides in the pathogenesis of type 2 diabetes mellitus (T2DM) is incompletely characterized. Given that ceramides represent therapeutic targets to disrupt the euglycemia-T2DM transition, we aimed to characterize their association with prevalent and incident T2DM in a novel cohort. METHODS:We examined the cross-sectional and longitudinal association of baseline ceramides with prevalent and incident T2DM among 1423 adults (47% women; median (range) baseline age 72 (51-95) years) in the Mayo Clinic Study of Aging cohort. We examined the associations of ceramides with prevalent T2DM (adjusted odds ratio [95% confidence interval]) at baseline and incident T2DM (adjusted hazard ratio [95% confidence interval]) during median follow-up of 6.2 years, after adjusting for demographic and metabolic factors. RESULTS:Among 1423 adults, there were 222 prevalent and 37 incident cases of T2DM. In cross-sectional analyses, higher levels of ceramide C16:0 were associated with lower odds of prevalent T2DM (aOR 0.84 [0.71-0.99];P = 0.03) whereas C18:0 (aOR 1.27 [1.06-1.42];P = 0.01), C18:0/16:0 (aOR 1.41 [1.22-1.62]; P < 0.001) and C18:0/24:0 (aOR 1.22 [1.05-1.41]; P = 0.01) were associated with higher odds. In Cox hazard regression models, C18:0/16:0 (aHR 1.63 [1.26-2.10];P < 0.001) and C18:0 (aHR 1.53 [1.12-2.08];P = 0.01) were associated with increased risk of incident T2DM. CONCLUSIONS:In this prospective population-based cohort, ceramides were associated with prevalent T2DM (C16:0,C18:0, C18:0/C16:0 ratio, C18:0/C24:0 ratio) and incident T2DM (C18:0, C18:0/C16:0 ratio) and could suggest targets for the primary and secondary prevention of T2DM.

Background The additive mechanistic effect of genetic risk variants for Parkinson’s disease (PD) is a plausible but largely unproven hypothesis. We investigated the mechanistic interaction between the two lysosomal PD risk genes glucocerebrosidase 1 ( GBA1 ) and sphingomyelinase 1 ( SMPD1 ) in complementing model systems. Methods Using CRISPR/Cas gene editing, smpd1 mutant ( smpd1 -/- ) zebrafish were generated and crossed to our previously characterised gba1 -/- zebrafish line, generating double knockouts ( gba1 -/- ; smpd1 -/- ). Spontaneous motor behaviour and survival were assessed in WT, single mutants and double mutants. HPLC-based sphingolipid quantification was combined with RNAseq based pathway analysis, assessment of the mitochondrial respiratory chain and quantification of lipid membrane oxidation for in-depth assessment of cellular health across all four genotypes. We also determined the effect of combined glucocerebrosidase (GCase) and acid sphingomyelinase (ASM) inactivation on autophagy and alpha-synuclein homeostasis in the human neuronal cell line SH-SY5Y. Results Unexpectedly, ASM deficiency rescued the marked behavioural phenotype and prolonged survival in gba1 -/- ; smpd1 -/- double-mutant zebrafish compared to gba1 -/- . RNAseq-based pathway analysis confirmed a profound rescue of neuronal function and intracellular homeostasis. We identified complete reciprocal rescue of mitochondrial respiratory chain function and abolished lipid membrane oxidation in gba1 -/- ; smpd1 -/- compared to gba1 -/- or smpd1 -/- as the underlying rescue mechanism. The complementing in vitro experiments demonstrated an unexpected reduction of α-synuclein levels in human cell lines with combined GCase and ASM deficiency. Conclusions Our study highlights the importance of functional validation for any putative mechanistic interactions between genetic risk factors and their overall effect on disease-relevant mechanisms rather than readily assuming an additive effect.

Obesity and elevation of circulating free fatty acids are associated with an accumulation and proinflammatory polarization of macrophages within metabolically active tissues, such as adipose tissue, muscle, liver, and pancreas. Beyond macrophages, neutrophils also accumulate in adipose and muscle tissues during high-fat diets and contribute to a state of local inflammation and insulin resistance. However, the mechanisms by which neutrophils are recruited to these tissues are largely unknown. Here we used a cell culture system as proof of concept to show that, upon exposure to a saturated fatty acid, palmitate, macrophages release nucleotides that attract neutrophils. Moreover, we found that palmitate up-regulates pannexin-1 channels in macrophages that mediate the attraction of neutrophils, shown previously to allow transfer of nucleotides across membranes. These findings suggest that proinflammatory macrophages release nucleotides through pannexin-1, a process that may facilitate neutrophil recruitment into metabolic tissues during obesity.

The additive effect of genetic risk variants on overall disease risk is a plausible but frequently unproven hypothesis. Variants in the two lysosomal disease genes () and () are genetic risk factors for sporadic Parkinson’s disease (PD). We investigated the biological effect of combined glucocerebrosidase (GCase) and acid sphingomyelinase (ASM) deficiency in zebrafish (), an ideally suited vertebrate model system to study gene-gene interaction. Combined genetic inactivation of GCase and ASM resulted in marked synergistic increases of key sphingolipids in the ; double-mutant compared to or single mutant zebrafish. However, the ; double-mutant zebrafish unexpectedly displayed a complete reciprocal rescue of mitochondrial respiratory chain function compared to the or single mutant zebrafish. The mitochondrial rescue effect resulted in abolished lipid membrane peroxidation, normalisation of motor behaviour and prolonged survival in the ; double-mutant zebrafish compared to the single mutant zebrafish strain. RNAseq-based transcriptomic pathway analysis confirmed a profound rescue of neuronal function and intracellular homeostasis. Complementing experiments in a previously established model of GCase deficiency in dopaminergic SH-SY5Y cells demonstrated that the observed rescue effect is autophagy-independent. Our study highlights the importance of a functional validation for any putative interactions between PD risk genes and their overall effect on disease-relevant mechanisms rather than readily assuming an additive effect.

Objective: High plasma ceramide levels and ratios are associated with poor outcomes in individuals with cardiovascular disease; less is known about their relation to cerebral small vessel disease. We examined whether high plasma ceramide levels or ratios were associated with cerebral microbleeds (CMBs) and lacunes and whether associations differ by sex. Approach and Results: We included 548 participants enrolled in the MCSA (Mayo Clinic Study of Aging) with concurrent plasma ceramide assays and magnetic resonance imaging. CMBs were quantified on T2* magnetic resonance imaging and lacunes on T2 fluid-attenuated inversion recovery magnetic resonance imaging. Fasting plasma ceramides were assayed using liquid chromatography-electrospray ionization tandem mass spectrometry. We used logistic regression models adjusting for age, sex, hypertension, and diabetes mellitus to examine the relationship between ceramides and presence of a lacune; hurdle models were used for presence and number of CMBs. Each SD increase in the log ceramide C16:0/24:0 ratio was associated with greater odds of a CMB (odds ratio, 1.28 [95% CI, 1.01-1.64]). There was an interaction between sex and the ceramide C16:0/24:0 ratio (P=0.049). The association between this ratio and presence of a CMB was stronger for women (odds ratio, 1.87 [95% CI, 1.20-3.00]) than men (odds ratio, 1.09 [95% CI, 0.80-1.46]). Several ceramides and all ceramide ratios were associated with number of CMBs. We did not find associations between plasma ceramides and lacunes. Conclusions: In a population-based sample, the plasma ceramide C16:0/24:0 ratio was associated with CMBs and was stronger for women. Plasma ceramides are differentially associated with cerebral small vessel pathologies. Graphic Abstract: A is available for this article.

It is unclear whether cerebrospinal fluid (CSF) phosphatidylcholines (PCs) are associated with neuroimaging measures of amyloid deposition and neurodegeneration (glucose metabolism, cortical thickness, and hippocampal volume), cognitive decline, or risk of mild cognitive impairment (MCI) among cognitively unimpaired older adults. This study investigated the associations of 19 individual CSF PC concentrations and their total sum with cross-sectional and longitudinal measures of amyloid deposition and neurodegeneration, global and domain-specific cognitive z-scores, and risk of MCI among 655 cognitively unimpaired participants, mean age of 71 years, enrolled in the Mayo Clinic Study of Aging. Neither the CSF total PC concentration nor individual CSF PCs were cross-sectionally or longitudinally associated with neuroimaging measures, cognition, or risk of MCI.

Serum lipids and sphingolipids circulate in plasma and are linked with insulin resistance and increased cardiovascular disease risk. We previously reported that dihydroceramides (DHCer) are elevated in people with insulin resistance compared to individuals with obesity and trained athletes. Despite associations with insulin resistance, little is known whether insulin sensitizing lifestyle interventions can improve sphingolipid profiles. Twenty-one individuals with obesity and prediabetes (18F; 46± 2y; 35.0± 0.9 kg/m2; fasting glucose: 113.8± 9.4 mg/dl, 2h OGTT: 143.0± 19.4 mg/dl) participated in a 3-month lifestyle intervention of combined weight loss and exercise. Insulin sensitivity (SI) was assessed by hyperinsulinemic-euglycemic clamps and both targeted and shotgun lipidomics were conducted before and after intervention. BMI and body fat percentage were reduced by 11+1% (mean± SEM) and 9± 1% after intervention (p<0.0001 for both). VO2 peak and SI increased by 14± 3% and 66± 12% following the intervention (p<0.0001 for both). Total serum triacylglyceride, diacylglyceride, DHCer and glucosylceramide were significantly reduced following intervention (p<0.05 for all). Reductions in specific species of ceramide (C18:0), lactosylceramide (C24:0), DHCer (C18:0, C24:1, C24:0) and glucosylceramide (C16:0, C22:0, C23:0, C24:1, C24:0) were also apparent. Of these lipids, glucosylceramide C20:0 and C22:0 were negatively associated with SI (r=-0.491, p<0.05 and r=-0.580, p<0.01, respectively). Furthermore, sphingomyelin C14:0, C16:1, C20:0, C21:0, C22:0, C23:0, C24:3 and C24:4 were negatively associated with SI. These data show that serum sphingolipids species previously linked to insulin resistance in humans can be improved with insulin sensitizing lifestyle interventions. Decreases in serum sphingolipids may explain increased SI and decreased cardiovascular disease risk associated with weight loss and exercise training. Disclosure J.L. Broussard: Advisory Panel; Self; National Institutes of Health. Research Support; Self; National Institute of Diabetes and Digestive and Kidney Diseases, Sleep Research Society, Society in Science-The Branco Weiss Fellowship. D.E. Kahn: None. S.A. Newsom: None. J.T. Brozinick: Employee; Self; Eli Lilly and Company. H. Bui: Employee; Self; Eli Lilly and Company. K.D. Roth: Employee; Self; Eli Lilly and Company. L. Perreault: Advisory Panel; Self; Novo Nordisk A/S, Sanofi. Speaker's Bureau; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Janssen Pharmaceuticals, Inc., Merck & Co., Inc., Novo Nordisk A/S. B.C. Bergman: Advisory Panel; Spouse/Partner; AstraZeneca, Merck & Co., Inc., Novo Nordisk Inc. Funding American Diabetes Association (1-14-CE-05 to B.B.); National Institutes of Health (RR-00036, K01DK110138); Colorado Nutrition Obesity Research Center (P30DK048520)

The additive effect of genetic risk variants on overall disease risk is a plausible but frequently unproven hypothesis. To test this hypothesis, we assessed the biological effect of combined glucocerebrosidase (GCase) and acid sphingomyelinase (ASM) deficiency. Variants in both glucocerebrosidase1 ( GBA1 ) and sphingomyelinase ( SMPD1 ) are genetic risk factors for Parkinson’s disease. Unexpectedly, ASM deficiency resulted in normalized behaviour and prolonged survival in gba1 −/−; smpd1 −/− double-mutant zebrafish compared to gba1 −/−. RNAseq-based pathway analysis confirmed a profound rescue of neuronal function and intracellular homeostasis. We identified complete reciprocal rescue of mitochondrial respiratory chain function and abolished lipid membrane oxidation in gba1 −/−; smpd1 −/− compared to gba1 −/− or smpd1 −/− as the underlying rescue mechanism. Complementing in vitro experiments demonstrated an unexpected reduction of α-synuclein levels in human cell lines with combined GCase and ASM deficiency. Our study highlights the importance of functional validation for any putative interactions between genetic risk factors and their overall effect on disease-relevant mechanisms rather than readily assuming an additive effect.Summary The additive effect of genetic risk variants on disease risk is a popular but typically unproven hypothesis. We investigated this hypothesis mechanistically for Parkinson’s disease risk factors and provide evidence of an unexpected rescue effect on neuronal function and survival.### Competing Interest StatementThe authors have declared no competing interest.

The de novo ceramide synthesis pathway is essential to human biology and health, but genetic influences remain unexplored. The core function of this pathway is the generation of biologically active ceramide from its precursor, dihydroceramide. Dihydroceramides have diverse, often protective, biological roles; conversely, increased ceramide levels are biomarkers of complex disease. To explore the genetics of the ceramide synthesis pathway, we searched for deleterious nonsynonymous variants in the genomes of 1,020 Mexican Americans from extended pedigrees. We identified a Hispanic ancestry-specific rare functional variant, L175Q, in delta 4-desaturase, sphingolipid 1 (DEGS1), a key enzyme in the pathway that converts dihydroceramide to ceramide. This amino acid change was significantly associated with large increases in plasma dihydroceramides. Indexes of DEGS1 enzymatic activity were dramatically reduced in heterozygotes. CRISPR/Cas9 genome editing of HepG2 cells confirmed that the L175Q variant results in a partial loss of function for the DEGS1 enzyme. Understanding the biological role of DEGS1 variants, such as L175Q, in ceramide synthesis may improve the understanding of metabolic-related disorders and spur ongoing research of drug targets along this pathway.

Plasma phosphatidylcholines (PCs) have been examined in the context of Alzheimer's disease dementia. However, their association with longitudinal changes in amyloid deposition remains unknown. This study investigated the associations of 8 plasma PC levels (PC aa [14:0_14:0], PC aa [16:0_16:0], PC aa [16:0_18:2], PC aa [16:0_22:6], PC aa [18:0_18:0], PC aa [18:0_18:1], PC aa [18:0_20:4], PC aa [18:1_18:1]) with cross-sectional and longitudinal measures of amyloid deposition, Alzheimer's disease-associated neurodegeneration (glucose metabolism and cortical thickness), and cognition (global- and domain-specific) of 1440 cognitively unimpaired participants (47% female, aged 50.7-95.3 years) in the Mayo Clinic Study of Aging. Longitudinally, higher baseline levels of PC aa [16:0_18:2], PC aa [18:0_18:1], and PC aa [18:1_18:1] were associated with slower decline in performance on tests of global cognition and specific cognitive domains. Furthermore, higher baseline levels of plasma PC aa (14:0_14:0) were associated with slower amyloid deposition and cortical thinning after multiple covariable adjustment (age, sex, education, medical comorbidity, dyslipidemia, statin use, and APOE4 allele presence). Our study findings support an independent association between plasma PC aa (14:0_14:0) with slower amyloid deposition and cortical thinning among cognitively unimpaired older adults. (C) 2019 Elsevier Inc. All rights reserved.