Endosomes and lysosomes play distinct roles in sulfatide-induced neuroblastoma apoptosis: potential mechanisms contributing to abnormal sulfatide metabolism in related neuronal diseases

Alterations in sulfatide metabolism, trafficking and homoeostasis are present at the earliest clinically recognizable stages of Alzheimer's disease and are associated with metachromatic leukodystrophy. However, the role of sulfatide in these disease states remains unknown. In the present study, we investigated the sequelae of NB (neuroblastoma) cells upon sulfatide supplementation and the biochemical mechanisms contributing to the sulfatide-induced changes. By using shotgun lipidomics, we showed dramatic accumulations of sulfatide, ceramide and sphingosine in NB cells in a time- and dose-dependent manner. Further studies utilizing subcellular fractionation and shotgun lipidomics analyses demonstrated that most of the increased ceramide content was generated in the endosomal compartment, whereas sulfatides predominantly accumulated in lysosomes. In addition, we determined that the sulfatide-mediated increase in endosomal ceramide content mainly resulted from beta-galactosidase activity, which directly hydrolyses sulfatide to ceramide without a prior desulfation step. Substantial cell apoptosis occurred in parallel with the accumulation of sulfatides and ceramides, as revealed by mitochondrial membrane depolarization, by phosphatidylserine translocation and by the TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling) assay. These findings were also demonstrated with primary neuron cultures. Collectively, our results demonstrate that abnormal sulfatide metabolism can induce cell apoptosis due to endosome-mediated ceramide generation and the accumulation of cytotoxic levels of sulfatides in lysosomes.     

Plasmalogen deficiency in early Alzheimer's disease subjects and in animal models: molecular characterization using electrospray ionization mass spectrometry

To explore the hypothesis that alterations in ethanolamine plasmalogen may be directly related to the severity of dementia in Alzheimer's disease (AD), we performed a systematic examination of plasmalogen content in cellular membranes of gray and white matter from different regions of human subjects with a spectrum of AD clinical dementia ratings (CDR) using electrospray ionization mass spectrometry (ESI/MS). The results demonstrate: (1) a dramatic decrease in plasmalogen content (up to 40 mol% of total plasmalogen) in white matter at a very early stage of AD (i.e. CDR 0.5); (2) a correlation of the deficiency in gray matter plasmalogen content with the AD CDR (i.e. approximately 10 mol% of deficiency at CDR 0.5 (very mild dementia) to approximately 30 mol% of deficiency at CDR 3 (severe dementia); (3) an absence of alterations of plasmalogen content and molecular species in cerebellar gray matter at any CDR despite dramatic alterations of plasmalogen content in cerebellar white matter. Alterations of ethanolamine plasmalogen content in two mouse models of AD, APP(V717F) and APPsw, were also examined by ESI/MS. A plasmalogen deficiency was present (up to 10 mol% of total plasmalogen at the age of 18 months) in cerebral cortices, but was absent in cerebella from both animal models. These results suggest plasmalogen deficiency may play an important role in the AD pathogenesis, particularly in the white matter, and suggest that altered plasmalogen content may contribute to neurodegeneration, synapse loss and synaptic dysfunction in AD.     

Neuropathological correlates of amyloid PET imaging in Down syndrome

Down syndrome (DS) results in an overproduction of amyloid‐β (Aβ) peptide associated with early onset of Alzheimer's disease (AD). DS cases have Aβ deposits detectable histologically as young as 12–30 years of age, primarily in the form of diffuse plaques, the type of early amyloid pathology also seen at pre‐clinical (i.e., pathological aging) and prodromal stages of sporadic late onset AD. In DS subjects aged >40 years, levels of cortical Aβ deposition are similar to those observed in late onset AD and in addition to diffuse plaques involve cored plaques associated with dystrophic neurites (neuritic plaques), which are of neuropathological diagnostic significance in AD. The purpose of this review is to summarize and discuss findings from amyloid PET imaging studies of DS in reference to postmortem amyloid‐based neuropathology. PET neuroimaging applied to subjects with DS has the potential to (a) track the natural progression of brain pathology, including the earliest stages of amyloid accumulation, and (b) determine whether amyloid PET biomarkers predict the onset of dementia. In addition, the question that is still incompletely understood and relevant to both applications is the ability of amyloid PET to detect Aβ deposits in their earliest form.     

Specificity and potential mechanism of sulfatide deficiency in Alzheimer's disease: an electrospray ionization mass spectrometric study.

Recently, we have demonstrated that sulfatide content was substantially depleted in post-mortem brain samples from subjects with very mild Alzheimer's disease (AD) relative to age-matched controls. However, it is unknown if the observed sulfatide deficiency is AD-specific and what mechanism(s) lead to this depletion. By exploiting the advantages of electrospray ionization mass spectrometry techniques, we examined the specificity and a potential mechanism of sulfatide deficiency in AD in the study. In contrast to the sulfatide depletion observed in AD, it was found that the sulfatide content in post-mortem brain samples from subjects with Parkinson's disease and dementia with Lewy bodies was either higher than or comparable to that observed from controls, respectively, suggesting that sulfatide deficiency is likely specific to AD. Examination of lipid alterations in cultured embryonic rat brain oligodendrocytes treated with amyloid-beta peptide demonstrated that there was no alteration in sulfatide content up to a 24-hr interval after amyloid-beta addition/treatment. However, there were significant decreases in plasmenylethanolamine and increases in sphingomyelin content in the same study. These findings suggest that sulfatide deficiency in AD is unlikely mediated directly by amyloid-beta peptide accumulation. Thus, these results illustrate the specificity of sulfatide deficiency in AD and exclude amyloid-beta accumulation as a factor directly contributing to sulfatide deficiency in AD.     

The Pathogenic Implication of Abnormal Interaction Between Apolipoprotein E Isoforms, Amyloid-beta Peptides, and Sulfatides in Alzheimer’s Disease

Alzheimer’s disease (AD) is the most common cause of dementia in the aging population. Prior work has shown that the ε4 allele of apolipoprotein E (apoE4) is a major risk factor for “sporadic” AD, which accounts for >99% of AD cases without a defined underlying mechanism. Recently, we have demonstrated that sulfatides are substantially and specifically depleted at the very early stage of AD. To identify the mechanism(s) of sulfatide loss concurrent with AD onset, we have found that: (1) sulfatides are specifically associated with apoE-associated particles in cerebrospinal fluid (CSF); (2) apoE modulates cellular sulfatide levels; and (3) the modulation of sulfatide content is apoE isoform dependent. These findings not only lead to identification of the potential mechanisms underlying sulfatide depletion at the earliest stages of AD but also serve as mechanistic links to explain the genetic association of apoE4 with AD. Moreover, our recent studies further demonstrated that (1) apoE mediates sulfatide depletion in amyloid-β precursor protein transgenic mice; (2) sulfatides enhance amyloid β (Aβ) peptides binding to apoE-associated particles; (3) Aβ42 content notably correlates with sulfatide content in CSF; (4) sulfatides markedly enhance the uptake of Aβ peptides; and (5) abnormal sulfatide-facilitated Aβ uptake results in the accumulation of Aβ in lysosomes. Collectively, our studies clearly provide a link between apoE, Aβ, and sulfatides in AD and establish a foundation for the development of effective therapeutic interventions for AD.     

Substantial sulfatide deficiency and ceramide elevation in very early Alzheimer's disease: potential role in disease pathogenesis

In addition to pathology in the gray matter, there are also abnormalities in the white matter in Alzheimer's disease (AD). Sulfatide species are a class of myelin-specific sphingolipids and are involved in certain diseases of the central nervous system. To assess whether sulfatide content in gray and white matter in human subjects is associated with both the presence of Alzheimer's disease (AD) pathology as well as the stage of dementia, we analyzed the sulfatide content of brain tissue lipid extracts by electrospray ionization mass spectrometry from 22 subjects whose cognitive status at time of death varied from no dementia to very severe dementia. All subjects with dementia had AD pathology. The results demonstrate that: (i) sulfatides were depleted up to 93% in gray matter and up to 58% in white matter from all examined brain regions from AD subjects with very mild dementia, whereas all other major classes of lipid (except plasmalogen) in these subjects were not altered in comparison to those from age-matched subjects with no dementia; (ii) there was no apparent deficiency in the biosynthesis of sulfatides in very mild AD subjects as characterized by the examination of galactocerebroside sulfotransferase activities in post-mortem brain tissues; (iii) the content of ceramides (a class of potential degradation products of sulfatides) was elevated more than three-fold in white matter and peaked at the stage of very mild dementia. The findings demonstrate that a marked decrease in sulfatides is associated with AD pathology even in subjects with very mild dementia and that these changes may be linked with early events in the pathological process of AD.     

Sulfatides facilitate apolipoprotein E-mediated amyloid-beta peptide clearance through an endocytotic pathway

Amyloid-β (Aβ) accumulation and fibril formation are key pathologic characteristics of Alzheimer's disease (AD). We have previously found that sulfatide depletion occurs at the earliest stages of AD. To further identify the role of sulfatides in the pathogenesis of AD as well as the interactions between apolipoprotein E (apoE), sulfatides, and Aβ peptides, we examined alterations in the clearance of apoE-mediated Aβ peptides after sulfatide supplementation to cell culture systems. We demonstrated that sulfatides markedly facilitate apoE-mediated clearance of Aβ peptides endogenously generated from H4-APPwt cells through an endocytotic pathway. Moreover, we found that the uptake of Aβ42 mediated by sulfatides was selective in comparison to that of Aβ40. We excluded the possibility that the supplementation of sulfatides and/or apoE altered the production of Aβ peptides from H4-APPwt cells through determination of the clearance of Aβ peptides from conditioned H4-APPwt cell media by neuroblastoma cells which do not appreciably generate Aβ peptides. Finally, we demonstrated that the sulfate galactose moiety of sulfatides is essential for the sulfatide-facilitated clearance of Aβ peptides. Collectively, the current study provides insight into a molecular mechanism leading to Aβ clearance/deposition, highlights the significance of sulfatide deficiency at the earliest clinically recognizable stage of AD, and identifies a potential new direction for therapeutic intervention for the disease.     

Heme oxygenase-1 posttranslational modifications in the brain of subjects with Alzheimer disease and mild cognitive impairment

Alzheimer disease (AD) is a neurodegenerative disorder characterized by progressive cognitive impairment and neuropathology. Oxidative and nitrosative stress plays a principal role in the pathogenesis of AD. The induction of the heme oxygenase-1/biliverdin reductase-A (HO-1/BVR-A) system in the brain represents one of the earliest mechanisms activated by cells to counteract the noxious effects of increased reactive oxygen species and reactive nitrogen species. Although initially proposed as a neuroprotective system in AD brain, the HO-1/BVR-A pathophysiological features are under debate. We previously reported alterations in BVR activity along with decreased phosphorylation and increased oxidative/nitrosative posttranslational modifications in the brain of subjects with AD and those with mild cognitive impairment (MCI). Furthermore, other groups proposed the observed increase in HO-1 in AD brain as a possible neurotoxic mechanism. Here we provide new insights about HO-1 in the brain of subjects with AD and MCI, the latter condition being the transitional phase between normal aging and early AD. HO-1 protein levels were significantly increased in the hippocampus of AD subjects, whereas HO-2 protein levels were significantly decreased in both AD and MCI hippocampi. In addition, significant increases in Ser-residue phosphorylation together with increased oxidative posttranslational modifications were found in the hippocampus of AD subjects. Interestingly, despite the lack of oxidative stress-induced AD neuropathology in cerebellum, HO-1 demonstrated increased Ser-residue phosphorylation and oxidative posttranslational modifications in this brain area, suggesting HO-1 as a target of oxidative damage even in the cerebellum. The significance of these findings is profound and opens new avenues into the comprehension of the role of HO-1 in the pathogenesis of AD.     

Soluble interleukin-1 receptor type II, IL-18 and caspase-1 in mild cognitive impairment and severe Alzheimer's disease

In the present study, we have determined levels of soluble interleukin-1 (IL-1) receptor type II (sIL-1RII), interleukin-18 (IL-18) and caspase-1 in cerebrospinal fluid and serum from mild cognitive impairment patients that later progressed to Alzheimer's disease (AD) and severe AD patients. Previous studies have shown that a chronic local inflammatory process is a part of AD neuropathology. In this process, activated microglial production of IL-1 seems to play an important role. In a previous study, we have shown increased levels of sIL-1RII in CSF from AD patients in a mild-moderate disease stage. In the present study, we found no significant differences in CSF or serum levels of sIL-1RII in either mild cognitive impairment or advanced AD patients as compared to control subjects. Likewise, there was no significant difference between mild cognitive impairment and severe AD patients. The same was true for caspase-1 and IL-18 serum levels, whereas CSF levels of caspase-1 and IL-18 were below detection limits. Our data indicate that the IL-1 system is relatively intact in the early and late stages of AD.     

Neural membrane phospholipids in alzheimer disease

Phospholipids form the backbone of neural membranes, providing fluidity and permeability. Two plasma membrane fractions, one from synaptosomes (SPM), the other glial and neuronal cell bodies (PM), were prepared from different regions of autopsied Alzheimer disease (AD) brains. Corresponding fractions were prepared from age-matched control brains. All fractions from AD brains showed significantly lower levels of ethanolamine glycerophospholipids and significantly higher levels of serine glycerophospholipids than the control brain. No differences were observed in phosphatidylcholine levels among these membranes. These results suggest that altered phospholipid composition of plasma membranes may be involved in the abnormal signal transduction and neurodegeneration in AD.Special issue dedicated to Dr. Leon S. Wolfe.     

Brain ceramide hexosides in Tay-Sachs disease and generalized gangliosidosis (GM1-gangliosidosis)

The carbohydrate composition was determined for ceramide hexosides isolated from brains of patients with Tay-Sachs disease and generalized gangliosidosis (hereby named GM1-gangliosidosis). Gray matter of patients with each disease showed a characteristic abnormal ceramide hexoside pattern. In Tay-Sachs gray matter, ceramide trihexoside is the major component, whereas ceramide tetrahexoside is barely detectable. In GM1-gangliosidosis, ceramide tetrahexoside is the major ceramide hexoside, while ceramide trihexoside is present only in small amount. These two major components have been characterized as the asialo derivatives of, respectively, the "Tay-Sachs ganglioside" (GM2-ganglioside) and the normal major monosialoganglioside (GM1-ganglioside). In both diseases, more than half the ceramide monohexoside of gray matter was glucocerebroside. Gray matter ceramide dihexoside, present in both diseases at higher than normal levels, was mostly ceramide lactoside, with possibly a small amount of ceramide digalactoside. Sulfatide contained only galactose. The abnormal ceramide hexoside pattern is limited to gray matter: white matter showed normal ceramide hexosides, i.e. a preponderance of monohexosides and sulfatide, with no detectable glucocerebroside.     

Pineal and cortical melatonin receptors MT1 and MT2 are decreased in Alzheimer's disease

The pineal hormone melatonin is involved in physiological transduction of temporal information from the light dark cycle to circadian and seasonal behavioural rhythms, as well as possessing neuroprotective properties. Melatonin and its receptors MT1 and MT2, which belong to the family of G protein-coupled receptors, are impaired in Alzheimer's disease (AD) with severe consequences to neuropathology and clinical symptoms. The present data provides the first immunohistochemical evidence for the cellular localization of the both melatonin receptors in the human pineal gland and occipital cortex, and demonstrates their alterations in AD. We localized MT1 and MT2 in the pineal gland and occipital cortex of 7 elderly controls and 11 AD patients using immunohistochemistry with peroxidase-staining. In the pineal gland both MT1 and MT2 were localized to pinealocytes, whereas in the cortex both receptors were expressed in some pyramidal and non-pyramidal cells. In patients with AD, parallel to degenerative tissue changes, there was an overall decrease in the intensity of receptors in both brain regions. In line with our previous findings, melatonin receptor expression in AD is impaired in two additional brain areas, and may contribute to disease pathology.     

An investigation of the molecular mechanisms engaged before and after the development of Alzheimer disease neuropathology in Down syndrome: a proteomics approach

Down syndrome (DS) is one of the most common causes of intellectual disability, owing to trisomy of all or part of chromosome 21. DS is also associated with the development of Alzheimer disease (AD) neuropathology after the age of 40 years. To better clarify the cellular and metabolic pathways that could contribute to the differences in DS brain, in particular those involved in the onset of neurodegeneration, we analyzed the frontal cortex of DS subjects with or without significant AD pathology in comparison with age-matched controls, using a proteomics approach. Proteomics represents an advantageous tool to investigate the molecular mechanisms underlying the disease. From these analyses, we investigated the effects that age, DS, and AD neuropathology could have on protein expression levels. Our results show overlapping and independent molecular pathways (including energy metabolism, oxidative damage, protein synthesis, and autophagy) contributing to DS, to aging, and to the presence of AD pathology in DS. Investigation of pathomechanisms involved in DS with AD may provide putative targets for therapeutic approaches to slow the development of AD.     

Multi-dimensional mass spectrometry-based shotgun lipidomics and the altered lipids at the mild cognitive impairment stage of Alzheimer's disease

Multi-dimensional mass spectrometry-based shotgun lipidomics (MDMS-SL) is a well-developed technology for global lipid analysis, which identifies and quantifies individual lipid molecular species directly from lipid extracts of biological samples. By using this technology, we have revealed three marked changes of lipids in brain samples of subjects with mild cognitive impairment of Alzheimer's disease including sulfatides, ceramides, and plasmalogens. Further studies using MDMS-SL lead us to the identification of the potential biochemical mechanisms responsible for the altered lipids at the disease state, which are thoroughly discussed in this minireview. Specifically, in studies to identify the causes responsible for sulfatide depletion at the mild cognitive impairment stage of Alzheimer's disease, we have found that apolipoprotein E is associated with sulfatide transport and mediates sulfatide homeostasis in the nervous system through lipoprotein metabolism pathways and that alterations in apolipoprotein E-mediated sulfatide trafficking can lead to sulfatide depletion in the brain. Collectively, the results obtained from lipidomic analyses of brain samples provide important insights into the biochemical mechanisms underlying the pathogenesis of Alzheimer's disease.     

Decreased RNA, and Increased RNA Oxidation, in Ribosomes from Early Alzheimer’s Disease

All cells rely on efficient protein synthesis in order to maintain cellular homeostasis. Recent studies from our laboratory indicate that declines in protein synthesis and ribosome function occur in the earliest stage of Alzheimer’s disease (AD). Additional studies indicate a potential role for ribosomal RNA oxidation as a potential mediator of decreased protein synthesis in AD. The ribosome is a complex of proteins and nucleic acids that mediates all protein synthesis. At present it is unclear if significant alterations in ribosomal RNA occurs within the ribosome complex during the progression of AD. In this study we examined the amount of ribosomal RNA in the different ribosomal fractions generated from control subjects, individuals with mild cognitive impairment (MCI), and individuals with AD. Studies were conducted in the inferior parietal lobule of each subject. Together, these data demonstrate that during the progression of AD there is a gross decline in the amount of ribosomal RNA within the ribosome complex. Additionally, these studies provide evidence for gross elevations in RNA oxidation within the ribosome complex of MCI and AD. Together, these data strongly suggest a role for RNA alterations within the ribosome as a mediator of decreased protein synthesis in both MCI and AD.     

Variable phenotype of Alzheimer's disease with spastic paraparesis

Pedigrees with familial Alzheimer's disease (AD) show considerable phenotypic variability. Spastic paraparesis (SP), or progressive spasticity of the lower limbs is frequently hereditary and exists either as uncomplicated (paraparesis alone) or complicated (paraparesis and other neurological features) disease subtypes. In some AD families, with presenilin-1 (PSEN1) mutations, affected individuals also have SP. These PSEN1 AD pedigrees frequently have a distinctive and variant neuropathology, namely large, non-cored plaques without neuritic dystrophy called cotton wool plaques (CWP). The PSEN1 AD mutations giving rise to CWP produce unusually high levels of the amyloid β peptide (Aβ) ending at position 42 or 43, and the main component of CWP is amino-terminally truncated forms of amyloid β peptide starting after the alternative β-secretase cleavage site at position 11. This suggests a molecular basis for the formation of CWP and an association with both SP and AD. The SP phenotype in some PSEN1 AD pedigrees also appears to be associated with a delayed onset of dementia compared with affected individuals who present with dementia only, suggesting the existence of a protective factor in some individuals with SP. Variations in neuropathology and neurological symptoms in PSEN1 AD raise the prospect that modifier genes may underlie this phenotypic heterogeneity.     

Inhibition of phosphatidylcholine and phosphatidylethanolamine biosynthesis in rat-2 fibroblasts by cell-permeable ceramides.

Phospholipids and sphingolipids are important precursors of lipid-derived second messengers such as diacylglycerol and ceramide, which participate in several signal transduction pathways and in that way mediate the effects of various agonists. The cross-talk between glycerophospholipid and sphingolipid metabolism was investigated by examining the effects of cell-permeable ceramides on phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdEtn) synthesis in Rat-2 fibroblasts. Addition of short-chain C6-ceramide to the cells resulted in a dose- and time-dependent inhibition of the CDP-pathways for PtdCho and PtdEtn synthesis. Treatment of cells for 4 h with 50 microM C6-ceramide caused an 83% and a 56% decrease in incorporation of radiolabelled choline and ethanolamine into PtdCho and PtdEtn, respectively. Exposure of the cells for longer time-periods (>/= 16 h) to 50 microM C6-ceramide resulted in apoptosis. The structural analogue dihydro-C6-ceramide did not affect PtdCho and PtdEtn synthesis. In pulse-chase experiments, radioactive choline and ethanolamine accumulated in CDP-choline and CDP-ethanolamine under the influence of C6-ceramide, suggesting that synthesis of both PtdCho and PtdEtn were inhibited at the final step in the CDP-pathways. Indeed, cholinephosphotransferase and ethanolaminephosphotransferase activities in membrane fractions from C6-ceramide-treated cells were reduced by 64% and 43%, respectively, when compared with control cells. No changes in diacylglycerol mass levels or synthesis of diacylglycerol from radiolabelled palmitate were observed. It was concluded that C6-ceramide affected glycerophospholipid synthesis predominantly by inhibition of the step in the CDP-pathways catalysed by cholinephosphotransferase and ethanolaminephosphotransferase.     

Cyclooxygenase‐1 inhibition reduces amyloid pathology and improves memory deficits in a mouse model of Alzheimer's disease

Several epidemiological and preclinical studies suggest that non‐steroidal anti‐inflammatory drugs (NSAIDs), which inhibit cyclooxygenase (COX), reduce the risk of Alzheimer's disease (AD) and can lower β‐amyloid (Aβ) production and inhibit neuroinflammation. However, follow‐up clinical trials, mostly using selective cyclooxygenase (COX)‐2 inhibitors, failed to show any beneficial effect in AD patients with mild to severe cognitive deficits. Recent data indicated that COX‐1, classically viewed as the homeostatic isoform, is localized in microglia and is actively involved in brain injury induced by pro‐inflammatory stimuli including Aβ, lipopolysaccharide, and interleukins. We hypothesized that neuroinflammation is critical for disease progression and selective COX‐1 inhibition, rather than COX‐2 inhibition, can reduce neuroinflammation and AD pathology. Here, we show that treatment of 20‐month‐old triple transgenic AD (3 × Tg‐AD) mice with the COX‐1 selective inhibitor SC‐560 improved spatial learning and memory, and reduced amyloid deposits and tau hyperphosphorylation. SC‐560 also reduced glial activation and brain expression of inflammatory markers in 3 × Tg‐AD mice, and switched the activated microglia phenotype promoting their phagocytic ability. The present findings are the first to demonstrate that selective COX‐1 inhibition reduces neuroinflammation, neuropathology, and improves cognitive function in 3 × Tg‐AD mice. Thus, selective COX‐1 inhibition should be further investigated as a potential therapeutic approach for AD.     

Acute kidney injury induced by protein-overload nephropathy down-regulates gene expression of hepatic cerebroside sulfotransferase in mice, resulting in reduction of liver and serum sulfatides

Sulfatides, possible antithrombotic factors belonging to sphingoglycolipids, are widely distributed in mammalian tissues and serum. We recently found that the level of serum sulfatides was significantly lower in hemodialysis patients than that in normal subjects, and that the serum level closely correlated to the incidence of cardiovascular disease. These findings suggest a relationship between the level of serum sulfatides and kidney function; however, the molecular mechanism underlying this relationship remains unclear. In the present study, the influence of kidney dysfunction on the metabolism of sulfatides was examined using an established murine model of acute kidney injury, protein-overload nephropathy in mice. Protein-overload treatment caused severe proximal tubular injuries within 4 days, and this treatment obviously decreased both serum and hepatic sulfatide levels. The sphingoid composition of serum sulfatides was very similar to that of hepatic ones at each time point, suggesting that the serum sulfatide level is dependent on the hepatic secretory ability of sulfatides. The treatment also decreased hepatic expression of cerebroside sulfotransferase (CST), a key enzyme in sulfatide metabolism, while it scarcely influenced the expression of the other sulfatide-metabolizing enzymes, including arylsulfatase A, ceramide galactosyltransferase, and galactosylceramidase. Pro-inflammatory responses were not detected in the liver of these mice; however, potential oxidative stress was increased. These results suggest that down-regulation of hepatic CST expression, probably affected by oxidative stress from kidney injury, causes reduction in liver and serum sulfatide levels. This novel mechanism, indicating the crosstalk between kidney injury and specific liver function, may prove useful for helping to understand the situation where human hemodialysis patients have low levels of serum sulfatides.     

Increases in ceramide levels in normal human mesangial cells subjected to different cellular stresses result from changes in distinct enzyme activities and can influence cellular responses to other stimuli

Sphingolipids, ceramide in particular, have come to be regarded as having roles in cellular signaling, most recently being associated with stress and the cellular responses to stress. In the present study we first examined the mechanisms involved in the changes in cellular ceramide levels in normal human mesangial cells (NHMC) in the growth, quiescent, and senescent phases as well as those resulting from environmental stimuli. We found that in NHMC total ceramide levels increase in response to cellular stresses as a result of a combination of enzyme activities. Furthermore, different stresses cause different alterations in various enzyme activities, with age and growth influencing acidic enzymes, but cell density affecting neutral, resulting in final ceramide level increases which most likely are associated with distinct pools of ceramide. Secondly, we examined the influence of changes in ceramide levels on apoptosis induced by sphingosine and its methylated derivative N, N-dimethylsphingosine. We found that increases in cellular ceramide levels prohibited the apoptosis and caused a quiescent state in the cells. The data presented here provide additional insight into the roles of ceramide and related enzymes in cellular responses to stress and suggest a possible relevance to in vivo disease states.