Sphingolipid Signaling Research Articles

Identification of lipidomic profiles associated with drug-resistant prostate cancer cells

BackgroundThe association of circulating lipids with clinical outcomes of drug-resistant castration-resistant prostate cancer (DR-CRPC) is not fully understood. While it is known that increases in select lipids correlate to decreased survival, neither the mechanisms mediating these alterations nor the correlation of resistance to drug treatments is well characterized.MethodsThis gap-in-knowledge was addressed using in vitro models of non-cancerous, hormone-sensitive, CRPC and drug-resistant cell lines combined with quantitative LC-ESI-Orbitrap-MS (LC-ESI-MS/MS) lipidomic analysis and subsequent analysis such as Metaboanalyst and Lipid Pathway Enrichment Analysis (LIPEA).ResultsSeveral lipid regulatory pathways were identified that are associated with Docetaxel resistance in prostate cancer (PCa). These included those controlling glycerophospholipid metabolism, sphingolipid signaling and ferroptosis. In total, 7460 features were identified as being dysregulated between the cell lines studied, and 21 lipid species were significantly altered in drug-resistant cell lines as compared to nonresistant cell lines. Docetaxel resistance cells (PC3-Rx and DU145-DR) had higher levels of phosphatidylcholine (PC), oxidized lipid species, phosphatidylethanolamine (PE), and sphingomyelin (SM) as compared to parent control cells (PC-3 and DU-145). Alterations were also identified in the levels of phosphatidic acid (PA) and diacylglyceride (DAG), whose levels are regulated by Lipin (LPIN), a phosphatidic acid phosphatase that converts PA to DAG. Data derived from cBioPortal demonstrated a population of PCa patients expressing mutations aligning with amplification of LPIN1, LPIN2 and LPIN3 genes. Lipin amplification in these genes correlated to decreased survival in these patients. Lipin-1 mRNA expression also showed a similar trend in PCa patient data. Lipin-1, but not Lipin-2 or − 3, was detected in several prostate cancer cells, and was increased in 22RV1 and PC-3 cell lines. The increased expression of Lipin-1 in these cells correlated with the level of PA.ConclusionThese data identify lipids whose levels may correlate to Docetaxel sensitivity and progression of PCa. The data also suggest a correlation between the expression of Lipin-1 in cells and patients with regards to prostate cancer cell aggressiveness and patient survivability. Ultimately, these data may be useful for identifying markers of lethal and/or metastatic prostate cancer.

The Influence of Coumestrol on Sphingolipid Signaling Pathway and Insulin Resistance Development in Primary Rat Hepatocytes

Coumestrol is a phytoestrogen widely known for its anti-diabetic, anti-oxidant, and anti-inflammatory properties. Thus, it gets a lot of attention as a potential agent in the nutritional therapy of diseases such as obesity and type 2 diabetes. In our study, we evaluated whether coumestrol affects insulin resistance development via the sphingolipid signaling pathway in primary rat hepatocytes. The cells were isolated from the male Wistar rat’s liver with the use of collagenase perfusion. Next, we incubated the cells with the presence or absence of palmitic acid and/or coumestrol. Additionally, some groups were incubated with insulin. The sphingolipid concentrations were assessed by HPLC whereas the expression of all the proteins was evaluated by Western blot. Coumestrol markedly reduced the accumulation of sphingolipids, namely, ceramide and sphinganine through noticeable inhibition of the ceramide de novo synthesis pathway in insulin-resistant hepatocytes. Moreover, coumestrol augmented the expression of fatty acid transport proteins, especially FATP5 and FAT/CD36, which also were responsible for excessive sphingolipid accumulation. Furthermore, coumestrol altered the sphingolipid salvage pathway, which was observed as the excessive deposition of the sphingosine-1-phosphate and sphingosine. Our study clearly showed that coumestrol ameliorated hepatic insulin resistance in primary rat hepatocytes. Thus, we believe that our study may contribute to the discovery of novel preventive and therapeutic methods for metabolic disorders.

SiO2 stimulates macrophage stress to induce the transformation of lung fibroblasts into myofibroblasts and its relationship with the sphingomyelin metabolic pathway.

Silicosis is a serious occupational disease with the highest incidence in China. However, its pathogenesis has not been fully elucidated. Studies have shown that the sphingomyelin signaling pathway may play an important role in different fibrotic diseases but its role in silicosis-mediated fibrosis is still unclear. In this study, the supernatant of human peripheral blood mononuclear cell line (THP-1)-derived macrophages exposed to silica (SiO2 ) was used to stimulate the transformation of human embryonic lung fibroblast cell line (HFL-1) into myofibroblasts, and the intervention effect of recombinant human acid ceramidase (rAC) was observed. The results showed that SiO2 stimulated the production of reactive oxygen species and malondialdehyde in the supernatant of THP-1-derived macrophages and increased the secretion of TGF-β1, TNF-α, and IL-8. In addition, we found that the expression levels of α-SMA, FN, Col I, and Col III in HFL-1 cells increased. Meanwhile, the activities of ASMase and ACase and the expression levels of Cer, Sph, and S1P were increased. Intervention by rAC can suppress these changes to different degrees. In conclusion, the present study shows that SiO2 dust poisoning may stimulate HFL-1 cell differentiation into myofibroblasts by inducing oxidative stress in THP-1-derived macrophages, thereby promoting the secretion of a variety of inflammatory factors and activating the sphingolipid signaling pathway in HFL-1 cells. Exogenous rAC can effectively interfere with the stimulation of HFL-1 cells by silica in vitro.

A Network Pharmacology Approach to Estimate Potential Targets of the Active Ingredients of Epimedium for Alleviating Mild Cognitive Impairment and Treating Alzheimer's Disease.

Background The present study made use of a network pharmacological approach to evaluate the mechanisms and potential targets of the active ingredients of Epimedium for alleviating mild cognitive impairment (MCI) and treating Alzheimer's disease (AD). Methods The active ingredients of Epimedium were acquired from the Traditional Chinese Medicine System Pharmacology database, and potential targets were predicted using the TCMSP target module, SwissTargetPrediction, and PharmMapper database. Target proteins correlating with MCI and AD were downloaded from the GeneCards, DisGeNet, and OMIM databases. The common targets of Epimedium, MCI, and AD were identified using the Jvenn online tool, and a protein-protein interaction (PPI) network was constructed using the String database and Cytoscape. Finally, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of the common targets was performed using DAVID, and molecular docking between active ingredients and target genes was modeled using AutoDock Vina. Results A total of 20 active ingredients were analyzed, and 337 compound-related targets were identified for Epimedium. Out of 236 proteins associated with MCI and AD, 54 overlapped with the targets of Epimedium. The top 30 interacting proteins in this set were ranked by topological analysis. GO and KEGG enrichment analysis suggested that the common targets participated in diverse biological processes and pathways, including cell proliferation and apoptosis, inflammatory response, signal transduction, and protein phosphorylation through cancer pathway, MAPK signaling pathway, PI3K-Akt signaling pathway, HIF-1 signaling pathway, sphingolipid signaling pathway, FoxO signaling pathway, and TNF signaling pathway. Molecular docking analysis suggested that the 20 active ingredients could bind to the top 5 protein targets. Conclusions The present study provides theoretical evidence for in-depth analysis of the mechanisms and molecular targets by which Epimedium protects against MCI, AD, and other neurodegenerative diseases and lays the foundation for pragmatic clinical applications and potential new drug development.

Lactation Associated Genes Revealed in Holstein Dairy Cows by Weighted Gene Co-Expression Network Analysis (WGCNA).

Simple SummaryWeighted gene coexpression network analysis (WGCNA) is a novel approach that can quickly analyze the relationships between genes and traits. In the past few years, studies on the gene expression changes of dairy cow mammary glands were only based on transcriptome comparisons between two lactation stages. Few studies focused on the relationships between gene expression of the dairy mammary gland and lactation stage or milk composition in a lactation cycle. In this study, we detected milk yield and composition in a lactation cycle. For the first time, we constructed a gene coexpression network using WGCNA on the basis of 18 gene expression profiles during six stages of a lactation cycle by transcriptome sequencing, generating 10 specific modules. Genes in each module were performed with gene ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Module–trait relationship analysis showed a series of potential candidates related to milk yield and composition. The current study provides an important theoretical basis for the further molecular breeding of dairy cows. Weighted gene coexpression network analysis (WGCNA) is a novel approach that can quickly analyze the relationships between genes and traits. In this study, the milk yield, lactose, fat, and protein of Holstein dairy cows were detected in a lactation cycle. Meanwhile, a total of 18 gene expression profiles were detected using mammary glands from six lactation stages (day 7 to calving, −7 d; day 30 post-calving, 30 d; day 90 post-calving, 90 d; day 180 post-calving, 180 d; day 270 post-calving, 270 d; day 315 post-calving, 315 d). On the basis of the 18 profiles, WGCNA identified for the first time 10 significant modules that may be related to lactation stage, milk yield, and the main milk composition content. Genes in the 10 significant modules were examined with gene ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The results revealed that the galactose metabolism pathway was a potential candidate for milk yield and milk lactose synthesis. In −7 d, ion transportation was more frequent and cell proliferation related terms became active. In late lactation, the suppressor of cytokine signaling 3 (SOCS3) might play a role in apoptosis. The sphingolipid signaling pathway was a potential candidate for milk fat synthesis. Dairy cows at 315 d were in a period of cell proliferation. Another notable phenomenon was that nonlactating dairy cows had a more regular circadian rhythm after a cycle of lactation. The results provide an important theoretical basis for the further molecular breeding of dairy cows.

Genome-wide methylation analysis of pre-pregnancy women in hypothyroidism

Background Hypothyroidism is a systemic metabolic deficiency syndrome caused by a deficiency in thyroid hormone or a decrease in the action of thyroid hormones. It has a high incidence among women of child-bearing age, and pregnant women with hypothyroidism may have a higher risk of birth defects. Objective To explore the specific biological mechanism affecting the occurrence of hypothyroidism. Methods This study determined key molecules by comparing and analyzing the difference in methylation levels between pre-pregnancy women and normal controls using the Illumina Infinium MethylationEPIC BeadChip. Results 3493 Differential methylation positions (DMPs) related genes and 47 differentially methylated regions (DMRs) related genes were found between the Hypothyroidism group and the control group. Among them, miR-21 has been found to be closely related to thyroid hormone regulation. The results of enrichment analysis showed that the DMPs or DMRs-related genes are both significantly enriched in human T-cell leukemia virus 1 infection, osteoclast differentiation and sphingolipid signaling pathway, which were also closely related to the occurrence and development of hypothyroidism. In addition, the combined analysis of CNVs and DMRs showed that significant differences occurred near the regions of 16 M bp in chromosome 1 between the two groups, and the genes involved in these regions included NDUFS2, FCER1G and SHC1. Conclusion This work screened molecular markers and key signaling pathways that are involved in the development of hypothyroidism in pre-pregnancy women, which may provide new targets for the research and diagnosis of hypothyroidism in the future.

A Novel Function of Sphingolipid Signaling via S1PR3 in Hematopoietic and Leukemic Stem Cells.

In this issue of Blood Cancer Discovery, Xie and colleagues describe a novel function of sphingosine-1-phosphate receptor 3 (S1PR3) to regulate myeloid differentiation and activate inflammatory programs in both human hematopoietic stem cells and leukemic stem cells. They propose S1PR3 as a major downstream signaling pathway of a TNFα-NF-κB axis in this study and unlock potential therapeutic opportunities to improve outcomes of patients with acute myeloid leukemia by modulating sphingolipid signaling via S1PR3. See related article by Xie et al., p. 32.

Very long-chain acyl-CoA synthetase 3 mediates onco-sphingolipid metabolism in malignant glioma.

Gliomas are the largest category of primary malignant brain tumors in adults, and glioblastomas account for nearly half of malignant gliomas. Glioblastomas are notoriously aggressive and drug-resistant, with a very poor 5 year survival rate of about 5%. New approaches to treatment are thus urgently needed. We previously identified an enzyme of fatty acid metabolism, very long-chain acyl-CoA synthetase 3 (ACSVL3), as a potential therapeutic target in glioblastoma. Using the glioblastoma cell line U87MG, we created a cell line with genomic deletion of ACSVL3 (U87-KO) and investigated potential mechanisms to explain how this enzyme supports the malignant properties of glioblastoma cells. Compared to U87MG cells, U87-KO cells grew slower and assumed a more normal morphology. They produced fewer, and far smaller, subcutaneous xenografts in nude mice. Acyl-CoA synthetases, including ACSVL3, convert fatty acids to their acyl-CoA derivatives, allowing participation in diverse downstream lipid pathways. We examined the effect of ACSVL3 depletion on several such pathways. Fatty acid degradation for energy production was not affected in U87-KO cells. Fatty acid synthesis, and incorporation of de novo synthesized fatty acids into membrane phospholipids needed for rapid tumor cell growth, was not significantly affected by lack of ACSVL3. In contrast, U87-KO cells exhibited evidence of altered sphingolipid metabolism. Levels of ceramides containing 18-22 carbon fatty acids were significantly lower in U87-KO cells. This paralleled the fatty acid substrate specificity profile of ACSVL3. The rate of incorporation of stearate, an 18-carbon saturated fatty acid, into ceramides was reduced in U87-KO cells, and proteomics revealed lower abundance of ceramide synthesis pathway enzymes. Sphingolipids, including gangliosides, are functional constituents of lipid rafts, membrane microdomains thought to be organizing centers for receptor-mediated signaling. Both raft morphology and ganglioside composition were altered by deficiency of ACSVL3. Finally, levels of sphingosine-1-phosphate, a sphingolipid signaling molecule, were reduced in U87-KO cells. We conclude that ACSVL3 supports the malignant behavior of U87MG cells, at least in part, by altering cellular sphingolipid metabolism.

GC-TOF-MS-Based Metabolomics Analyses of Liver and Intestinal Contents in the Overfed vs. Normally-Fed Geese.

Simple SummaryNon-alcoholic fatty liver disease has been considered as one of the most important causes of liver disease, and it is a threat to human and animal health worldwide. Interestingly, goose fatty liver can reach 8–10 times the weight of normal liver with no overt pathological symptoms, suggesting that there are some protective mechanisms. Scientists have indicated that gut microbiota participate in the formation of non-alcoholic fatty liver disease in human and mammalian animals. However, it is unclear whether gut microbiota and their metabolites contribute to goose fatty liver. The aim of the present study was to investigate the metabolomic analyses of liver and intestinal contents in overfed vs. normally fed geese. The results showed that the formation of goose fatty liver is accompanied by obvious changes in the metabolic profiles of liver and intestinal contents. The intestinal metabolites can affect the formation of goose fatty liver by affecting the metabolisms of glucose and fatty acid, oxidative stress, and inflammatory reactions. These findings provide a basis for future work addressing the relationship between intestinal metabolites and the development of non-alcoholic fatty liver disease.No overt pathological symptoms are observed in the goose liver with severe steatosis, suggesting that geese may host unique protective mechanisms. Gas chromatography time-of-flight mass spectrometry-based metabolomics analyses of liver and intestinal contents in overfed vs. normally fed geese (26 geese in each treatment) were investigated. We found that overfeeding significantly changed the metabolic profiles of liver and intestinal contents. The differential metabolites mainly belong to fatty acids, amino acids, organic acids, and amines. The differential metabolites were involved in glycolysis/gluconeogenesis, glycerolipid metabolism, the pentose phosphate pathway, fatty acid degradation, the sphingolipid signaling pathway, and the biosynthesis of unsaturated fatty acids. Moreover, we determined the biological effects of arachidonic acid (ARA) and tetrahydrocorticosterone (TD) in goose primary hepatocytes and intestinal cells. Data showed that the mRNA expression of arachidonate 5-lipoxygenase (ALOX5) in goose primary intestinal cells was significantly induced by 0.50 mM ARA treatment. Cytochrome P-450 27A1 (CYP27A1) mRNA expression was significantly inhibited in goose primary hepatocytes by 1 µM TD treatment. In conclusion, the formation of goose fatty liver is accompanied by significant changes in the metabolic profiles of liver and intestinal contents, and the changes are closely related to the metabolisms of glucose and fatty acids, oxidative stress, and inflammatory reactions.

Ceramide Dynamics and Prognostic Value in Acute and Subacute Ischemic Stroke: Preliminary Findings in a Clinical Cohort

Background: Ceramides are implicated in sphingolipid signaling. Elevated ceramide levels have been associated with increased cardiovascular risk, but information on their role in acute ischemic stroke (AIS) is limited. The purpose of this study is to investigate the temporal dynamics of ceramide levels in AIS and assess their prognostic utility for long-term outcomes. Methods: This is a prospective pilot study of patients with AIS admitted to Mayo Clinic within 12 h of last known well (LKW). Ceramides were assessed by liquid chromatography mass spectrometry at two time points: T1 (within 12 h of LKW) and T2 (1 - 7 days from LKW). Wilcoxon signed rank test was used to compare paired ceramide levels and ratios. Ordinal logistic regression was used for assessment of associations with long-term outcomes. Results: Twenty-three patients met inclusion criteria (median (range)): age (76 years (45 - 95)); body mass index (25.6 (20.5 - 46.6)); National Institutes of Health Stroke Scale (NIHSS) score (5 (0 - 27)); infarct volume (1.4 cm 3 (0.0 - 36.5)). Long-chain ceramides increased between T1 and T2 whereas very-long chain ceramides decreased, P < 0.05. Upon stratification of patients by prior statin exposure, increase in long-chain ceramide level was present only in statin-naive patients. Greater neurological disability at follow-up was associated with higher ceramide score, C(18:0)/C(24:0) ratio and higher levels of glycated hemoglobin. Conclusions: Long-chain and very-long-chain ceramide are actively implicated in pathologic processes in acute and subacute phases of stroke, with their dynamics being inversely related and potentially modulated by statin therapy. Ceramide levels and ratios might be useful for prognosis of long-term neurological outcomes. J Neurol Res. 2020;10(6):209-219 doi: https://doi.org/10.14740/jnr633

Metabolomic Analysis of the Ameliorative Effect of Enhanced Proline Metabolism on Hypoxia-Induced Injury in Cardiomyocytes.

Background Coronary heart disease is currently the leading cause of death in humans. Its poor prognosis and high mortality are associated with myocardial ischemia, which leads to metabolic disorder-related cardiomyocyte apoptosis and reactive oxygen species (ROS) production. Previous cardiovascular metabolomics studies in humans and mice have shown that proline metabolism is severely altered after cardiomyocyte hypoxia. Proline dehydrogenase (PRODH) is located on the inner mitochondrial membrane and is an enzyme that catalyzes the first step of proline catabolism, which plays an important role in improving the cellular redox state. In vitro oxygen-glucose deprivation can mimic in vivo myocardial ischemic injury. This study is aimed at investigating whether enhancing proline metabolism by overexpressing PRODH can ameliorate hypoxia-induced injury in cardiomyocytes and to reveal the related altered metabolites and mechanistic pathway via untargeted metabolomics analysis. Methods and Results First, through public database analysis and RT-qPCR and western blot analyses in a cardiomyocyte hypoxia model, we found that the expression of the proline-degrading enzyme PRODH was downregulated after myocardial infarction and hypoxia exposure. Second, LDH assays, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), DHE staining, flow cytometric apoptosis analysis with DCFH and Annexin V-FITC/PI, and western blot analysis were used to assess the injury level in cardiomyocytes. Enhanced proline metabolism induced by PRODH overexpression reduced the levels of reactive oxidative stress and apoptosis, whereas PRODH knockdown had the opposite effects. Third, untargeted metabolomics analysis revealed that the protective effect was associated with significant changes in metabolism linked to sphingolipid signaling pathways, unsaturated fatty acid biosynthesis, phosphocreatine, glutathione disulfide, aminoacyl-tRNA biosynthesis, and ABC transporters. Conclusions Our study demonstrated a protective effect of enhanced proline metabolism in cardiomyocytes under hypoxia, providing a novel strategy for exploring new treatments for coronary heart disease.

Sphingolipids and Inositol Phosphates Regulate the Tau Protein Phosphorylation Status in Humanized Yeast.

Hyperphosphorylation of protein tau is a hallmark of Alzheimer’s disease (AD). Changes in energy and lipid metabolism have been correlated with the late onset of this neurological disorder. However, it is uncertain if metabolic dysregulation is a consequence of AD or one of the initiating factors of AD pathophysiology. Also, it is unclear whether variations in lipid metabolism regulate the phosphorylation state of tau. Here, we show that in humanized yeast, tau hyperphosphorylation is stimulated by glucose starvation in coincidence with the downregulation of Pho85, the yeast ortholog of CDK5. Changes in inositol phosphate (IP) signaling, which has a central role in energy metabolism, altered tau phosphorylation. Lack of inositol hexakisphosphate kinases Kcs1 and Vip1 (IP6 and IP7 kinases in mammals) increased tau hyperphosphorylation. Similar effects were found by mutation of IPK2 (inositol polyphosphate multikinase), or PLC1, the yeast phospholipase C gene. These effects may be explained by IP-mediated regulation of Pho85. Indeed, this appeared to be the case for plc1, ipk2, and kcs1. However, the effects of Vip1 on tau phosphorylation were independent of the presence of Pho85, suggesting additional mechanisms. Interestingly, kcs1 and vip1 strains, like pho85, displayed dysregulated sphingolipid (SL) metabolism. Moreover, genetic and pharmacological inhibition of SL biosynthesis stimulated the appearance of hyperphosphorylated forms of tau, while increased flux through the pathway reduced its abundance. Finally, we demonstrated that Sit4, the yeast ortholog of human PP2A protein phosphatase, is a downstream effector of SL signaling in mediating the tau phosphorylation state. Altogether, our results add new knowledge on the molecular effectors involved in tauopathies and identify new targets for pharmacological intervention.

Effects of Yuanhu- Zhitong tablets on alcohol-induced conditioned place preference in mice

ObjectivesThis study aimed at determining the synergistic effects of Yuanhu Zhitong tablets (YHZTP) on alcohol-induced conditioned place preference (CPP) in mice, in addition, the intervention mechanism was preliminarily explored based on traditional Chinese Medicine (TCM) network pharmacology on alcohol addiction. MethodsAlcohol-induced CPP mice were used to evaluate the effects of either YHZTP or levo-tetrahydropalmatine (l-THP) plus imperatorin (IMP) administration on animal behavior. The network pharmacological strategy was used to establish the “compound-target” and “disease-drug-target” network. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed on the shared targets between the compound and the disease. Twelve algorithms on CytoHubba were used to find the hub genes that were verified by qPCR. ResultsSystemic administration (2 g/kg, i.p.) of ethanol (EtOH) to mice was used to induce CPP. YHZTP On its own did not induce CPP or conditioned place aversion (CPA) at the doses of 0.3 g/kg or 0.6 g/kg (i.g.), but attenuated the acquisition and expression of EtOH-induce CPP in mice. In addition, YHZTP (0.3 or 0.6 g/kg) did not exhibit any effect on the motor activity of mice. Acquisition of alcohol-induced CPP was blocked by a combination of l-THP (5 mg/kg, i.g.) + IMP (2.5 mg/kg, i.g.) or l-THP (10 mg/kg, i.g.) + IMP (5 mg/kg, i.g.). However, the combination of l-THP (2.5 mg/kg, i.g.) + IMP (1.25 mg/kg, i.g.) or mono-administration of l-THP and IMP did not exhibit any effect on alcohol-induced CPP. YHZTP was also shown to reverse the up-regulation of Gabra1, Ptgs2, Mapk1, Mapk8, Mapk14, Nr3c, Prkca and Sirt1 genes and the down-regulation of Hhtr2a and Drd2 genes in the prefrontal cortex of EtOH induced CPP mice. These genes were associated with neuroactive ligand-receptor interactions, activation of the sphingolipid, calcium, cAMP, ErbB, NF-kappa B and MAPK signaling pathways. ConclusionYHZTP inhibits EtOH-induced CPP behavior in mice while a combination of l-THP and IMP exerts a synergistic effect on the reduction of EtOH-induced CPP. Possible pharmacological mechanisms include inhibition of the expression of inflammatory factors and regulation of neurotransmitter receptor levels. Therefore, YHZTP is a novel candidate for the treatment of alcohol addiction.

Exploring polar headgroup interactions between sphingomyelin and ceramide with infrared spectroscopy

Ceramide is a major actor in the sphingolipid signaling pathway elicited by various kinds of cell stress. Under those conditions ceramide (Cer) is produced in the plasma membrane as a product of sphingomyelin (SM) hydrolysis, and this may lead to apoptosis. Thus, SM and Cer coexist in the membrane for some time, and they are known to separate laterally from the (more abundant) glycerolipids, giving rise to highly rigid domains or platforms. The properties of these domains/platforms are rather well understood, but the underlying SM:Cer molecular interactions have not been explored in detail. Infrared (IR) spectroscopy is a powerful analytical technique that provides information on all the chemical groupings in a molecule, and that can be applied to membranes and lipid bilayers in aqueous media. IR spectra can be conveniently retrieved as a function of temperature, thus revealing the thermotropic transitions of SM and its mixtures with Cer. Four regions of the IR spectrum of these sphingolipids have been examined, two of them dominated by the hydrophobic regions in the molecules, namely the C–H stretching vibrations (2800–3000 cm−1), and the CH2 scissoring vibrations (1455–1485 cm−1), and two others arising from chemical groups at the lipid-water interface, the sphingolipid amide I band (1600–1680 cm−1), and the phosphate vibrations in the 1000–1110 cm−1 region. The latter two regions have been rarely studied in the past. The IR data from the hydrophobic components show a gel (or ripple)-fluid transition of SM at 40 °C, that is shifted up to about 70 °C when Cer is added to the bilayers, in agreement with previous studies using a variety of techniques. IR information concerning the polar parts is more interesting. The amide I (carbonyl) band of pure SM exhibits a maximum at 1638 cm−1 at room temperature, and its position is shifted by about 10 cm−1 in the presence of Cer. Cer causes also a change in the overall band shape, but no signs of band splitting are seen, suggesting that SM and Cer carbonyl groups are interacting tightly, presumably through H-bonds. The 1086 cm−1 band, corresponding to PO2− vibrations, appears more stable in SM than in DPPC, and it is further stabilized by Cer, again suggesting an important role of H-bonds in the formation of SM:Cer clusters. Thus, SM and Cer can interact through their polar headgroups, in a way that is not accessible to other lipid classes.

Spatio-temporal correlates of gene expression and cortical morphology across lifespan and aging

Evidence from neuroimaging and genetic studies supports the concept that brain aging mirrors development. However, it is unclear whether mechanisms linking brain development and aging provide new insights to delay aging and potentially reverse it. This study determined biological mechanisms and phenotypic traits underpinning brain alterations across the lifespan and in aging by examining spatio-temporal correlations between gene expression and cortical volumes using datasets d with the age range from 2 to 82 years. We revealed that a large proportion of genes whose expression was associated with cortical volumes across the lifespan were in astrocytes. These genes, which showed up-regulation during development and down-regulation during aging, contributed to fundamental homeostatic functions of astrocytes. Included among these genes were those encoding components of cAMP, Ras, and retrograde endocannabinoid signaling pathways. Genes associated with cortical volumes in the same data aged above 55 years were also enriched for the sphingolipid, renin-angiotensin system (RAS), proteasome, and TGF-β signaling pathway, which is linked to senescence-associated secretory phenotypes. Neuroticism, drinking, and smoking were the common phenotypic traits in the lifespan and aging, while memory was the unique phenotype associated with aging. These findings provide biological mechanisms mirroring development and aging as well as unique to aging.

A Network Pharmacological Approach to Investigate the Mechanism of Action of Active Ingredients of Epimedii Herba and Their Potential Targets in Treatment of Alzheimer's Disease.

BackgroundEpimedii Herba is a traditional Chinese herbal medicine used to treat central nervous system diseases such as Alzheimer’s disease in China. However, the pharmacological mechanism is unclear. To investigate the mechanisms of Epimedii Herba in the treatment of Alzheimer’s disease, we assessed effective compounds, corresponding targets, and related pathways of Epimedii Herba in the treatment of Alzheimer’s disease based on network pharmacology.Material/MethodsThe active components and targets of Epimedii Herba were obtained through the TCMSP database and the DrugBank database. The DisGeNET database and GeneCards database were used to search for Alzheimer’s disease targets. The common targets of components and disease were obtained by Wayne diagram. Gene ontology (GO) analysis and enrichment analysis of the Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed using the DAVID database. The component-target-pathway interaction network model was constructed using Cytoscape software. Auto Duck Vina software was used for molecular docking to analyze the affinity of the key ingredients and the main targets.ResultsWe screened 17 active ingredients and 27 key targets of Epimedii Herba in the treatment of Alzheimer’s disease, which were related to the HIF-1 signaling pathway, TNF signaling pathway, PI3K-Akt signaling pathway, NF-κB signaling pathway, VEGF signaling pathway, and sphingolipid signaling pathway.ConclusionsBased on network pharmacology, the multi-component, multi-target, and multi-pathway characteristics of Epimedii Herba in the treatment of Alzheimer’s disease were explored. Our results provide new ideas for future pharmacological and experimental research on Epimedii Herba in the treatment of Alzheimer’s disease.

Analyzing Gene Expression Profiles from Ataxia and Spasticity Phenotypes to Reveal Spastic Ataxia Related Pathways.

Spastic ataxia (SA) is a group of rare neurodegenerative diseases, characterized by mixed features of generalized ataxia and spasticity. The pathogenetic mechanisms that drive the development of the majority of these diseases remain unclear, although a number of studies have highlighted the involvement of mitochondrial and lipid metabolism, as well as calcium signaling. Our group has previously published the GBA2 c.1780G > C (p.Asp594His) missense variant as the cause of spastic ataxia in a Cypriot consanguineous family, and more recently the biochemical characterization of this variant in patients’ lymphoblastoid cell lines. GBA2 is a crucial enzyme of sphingolipid metabolism. However, it is unknown if GBA2 has additional functions and therefore additional pathways may be involved in the disease development. The current study introduces bioinformatics approaches to better understand the pathogenetic mechanisms of the disease. We analyzed publicly available human gene expression datasets of diseases presented with ‘ataxia’ or ‘spasticity’ in their clinical phenotype and we performed pathway analysis in order to: (a) search for candidate perturbed pathways of SA; and (b) evaluate the role of sphingolipid signaling pathway and sphingolipid metabolism in the disease development, through the identification of differentially expressed genes in patients compared to controls. Our results demonstrate consistent differential expression of genes that participate in the sphingolipid pathways and highlight alterations in the pathway level that might be associated with the disease phenotype. Through enrichment analysis, we discuss additional pathways that are connected to sphingolipid pathways, such as PI3K-Akt signaling, MAPK signaling, calcium signaling, and lipid and carbohydrate metabolism as the most enriched for ataxia and spasticity phenotypes.

Protective effect of picroside I against hepatic fibrosis in mice via sphingolipid metabolism, bile acid biosynthesis, and PPAR signaling pathway

Picroside I, a hepatoprotectant isolated from Picrorhiza kurroa Royle ex Benth and P. scrophulariiflora Pennell, can reduce liver injury in humans and animals. However, its anti-fibrosis effect remains elusive. This work aimed to explore the mechanism underlying the hepatoprotective effect of picroside I against hepatic fibrosis. Male mice (12 mice per group) were randomly divided into six groups: the control group; the model group, which received thioacetamide (TAA); the positive group, which received TAA + S-(5′-adenosyl)-l-methionine (SAMe, 10 mg/kg); the low-dose group, which received TAA + picroside I (25 mg/kg); the middle-dose group, which received TAA + picroside I (50 mg/kg); and the high-dose group, which received TAA + picroside I (75 mg/kg). Serum biochemical indicators were detected, and histological evaluation was performed. Metabolomics and proteomic analyses were conducted via liquid-chromatography coupled with tandem mass spectrometry (LC-MS/MS). Data showed that picroside I could decrease the serum levels of alanine transaminase (ALT), aspartate transaminase (AST), collagen type IV (CIV), N-terminal peptide of type III procollagen (PIIINP), laminin (LN), and hyaluronic acid (HA) and reduced fibrosis area. Picroside I altered metabolomic profiles, including energy, lipid, and glutathione (GSH) metabolism, in ice with fibrosis. Additionally, 25 differentially expressed proteins in the picroside I high-dose-treated group were reversed relative to in the model group. These proteins were involved in the sphingolipid signaling pathway, primary bile acid biosynthesis, and peroxisome proliferator-activated receptor (PPAR) signaling pathway. Moreover, this study revealed how picroside I could protect against TAA-induced liver fibrosis in mice. Results indicated that picroside I can serve as a candidate drug for hepatic fibrosis.

Sepsis and Cerebral Dysfunction: BBB Damage, Neuroinflammation, Oxidative Stress, Apoptosis and Autophagy as Key Mediators and the Potential Therapeutic Approaches.

Sepsis-associated cerebral dysfunction is complex pathophysiology, generated from primary infections that are developed elsewhere in the body. The neonates, elderly population and chronically ill and long-term hospitalized patients are predominantly vulnerable to sepsis and related cerebral damage. Generally, electrophysiological recordings, severity and sedation scales, computerized imaging and spectroscopy techniques are used for its detection and diagnosis. About the underlying mechanisms, enhanced blood-brain barrier permeability and metalloprotease activity, tight junction protein loss and endothelial cell degeneration promote the influx of inflammatory and toxic mediators into the brain, triggering cerebrovascular damage. An altered neutrophil count and phenotype further dysregulate the normal neuroimmune responses and neuroendocrine stability via modulated activation of protein kinase C-delta, nuclear factor kappa-B and sphingolipid signaling. Glial activation, together with pro-inflammatory cytokines and chemokines and the Toll-like receptor, destabilize the immune system. Moreover, superoxides and hydroperoxides generate oxidative stress and perturb mitochondrial dynamics and ATP synthesis, propagating neuronal injury cycle. Activated mitochondrial apoptotic pathway, characterized by increased caspase-3 and caspase-9 cleavage and Bax/Bcl2 ratio in the hippocampal and cortical neurons, stimulate neurocognitive impairments. Additionally, altered LC3-II/I and P62/SQSTM1, p-mTOR, p-AMPK1 and p-ULK1 levels and dysregulated autophagosome-lysosome fusion decrease neuronal and glial energy homeostasis. The therapies and procedures for attenuating sepsis-induced brain damage include early resuscitation, cerebral blood flow autoregulation, implantable electric vagus nerve stimulation, antioxidants, statins, glucocorticoids, neuroimmune axis modulators and PKCδ inhibitors. The current review enumerates the pathophysiology of sepsis-induced brain damage, its diagnosis, the role of critical inducers and mediators and, ultimately, therapeutic measures attenuating cerebrovascular degeneration.

Role of Sphingolipid Signaling in Glomerular Diseases: Focus on DKD and FSGS.

Sphingolipids are well-recognized as major players in the pathogenesis of many human diseases, including chronic kidney disease. The kidney is a very sensitive organ to alterations in sphingolipid metabolism. The critical issues to be addressed in this review relate to the role of sphingolipids and enzymes involved in sphingolipid metabolism in the pathogenesis of glomerular diseases with a special focus on podocytes, a key cellular component of the glomerular filtration barrier. Among several sphingolipids, we will highlight the role of ceramide, sphingosine, sphingosine-1-phosphate and ceramide-1-phosphate. Additionally, we will summarize the current knowledge with regard to the use of sphingolipids as therapeutic agents for the treatment of podocyte injury in kidney disease.