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1.
Saturated fatty acids, such as palmitate, promote accumulation of ceramide, which impairs activation and signalling of PKB (protein kinase B; also known as Akt) to important end points such as glucose transport. SPT (serine palmitoyl transferase) is a key enzyme regulating ceramide synthesis from palmitate and represents a potential molecular target in curbing lipid-induced insulin resistance. In the present study we explore the effects of palmitate upon insulin action in L6 muscle cells in which SPT expression/activity has been decreased by shRNA (small-hairpin RNA) or sustained incubation with myriocin, an SPT inhibitor. Incubation of L6 myotubes with palmitate (for 16 h) increases intramyocellular ceramide and reduces insulin-stimulated PKB activation and glucose uptake. PKB inhibition was not associated with impaired IRS (insulin receptor substrate) signalling and was ameliorated by short-term treatment with myriocin. Silencing SPT expression (approximately 90%) by shRNA or chronic cell incubation with myriocin (for 7 days) markedly suppressed SPT activity and palmitate-driven ceramide synthesis; however, challenging these muscle cells with palmitate still inhibited the hormonal activation of PKB. This inhibition was associated with reduced IRS1/p85-PI3K (phosphoinositide 3-kinase) coupling that arises from diverting palmitate towards greater DAG (diacylglycerol) synthesis, which elevates IRS1 serine phosphorylation via activation of DAG-sensitive PKCs (protein kinase Cs). Treatment of SPT-shRNA cells or those treated chronically with myriocin with PKC inhibitors antagonized palmitate-induced loss in insulin signalling. The findings of the present study indicate that SPT plays a crucial role in desensitizing muscle cells to insulin in response to incubation with palmitate. While short-term inhibition of SPT ameliorates palmitate/ceramide-induced insulin resistance, sustained loss/reduction in SPT expression/activity promotes greater partitioning of palmitate towards DAG synthesis, which impacts negatively upon IRS1-directed insulin signalling. 相似文献
2.
Cantrell Stanford J Morris AJ Sunkara M Popa GJ Larson KL Özcan S 《The Journal of biological chemistry》2012,287(16):13457-13464
Recent studies suggest that sphingolipid metabolism is altered during type 2 diabetes. Increased levels of the sphingolipid ceramide are associated with insulin resistance. However, a role for sphingolipids in pancreatic beta cell function, or insulin production, and release remains to be established. Our studies in MIN6 cells and mouse pancreatic islets demonstrate that glucose stimulates an intracellular rise in the sphingolipid, sphingosine 1-phosphate (S1P), whereas the levels of ceramide and sphingomyelin remain unchanged. The increase in S1P levels by glucose is due to activation of sphingosine kinase 2 (SphK2). Interestingly, rises in S1P correlate with increased glucose-stimulated insulin secretion (GSIS). Decreasing S1P levels by treatment of MIN6 cells or primary islets with the sphingosine kinase inhibitor reduces GSIS. Moreover, knockdown of SphK2 alone results in decreased GSIS, whereas knockdown of the S1P phosphatase, Sgpp1, leads to a rise in GSIS. Treatment of mice with the sphingosine kinase inhibitor impairs glucose disposal due to decreased plasma insulin levels. Altogether, our data suggest that glucose activates SphK2 in pancreatic beta cells leading to a rise in S1P levels, which is important for GSIS. 相似文献
3.
Background
Sustained exposure of pancreatic β cells to an increase in saturated fatty acids induces pleiotropic effects on β-cell function, including a reduction in stimulus-induced insulin secretion. The objective of this study was to investigate the effects of chronic over supply of palmitate upon glucose- and amino acid-stimulated insulin secretion (GSIS and AASIS, respectively) and autocrine-dependent insulin signalling with particular focus on the importance of ceramide, ERK and CaMKII signalling.Principal Findings
GSIS and AASIS were both stimulated by >7-fold resulting in autocrine-dependent activation of protein kinase B (PKB, also known as Akt). Insulin release was dependent upon nutrient-induced activation of calcium/calmodulin-dependent protein kinase II (CaMKII) and extracellular signal-regulated kinase (ERK) as their pharmacological inhibition suppressed GSIS/AASIS significantly. Chronic (48 h, 0.4 mM) palmitate treatment blunted glucose/AA-induced activation of CaMKII and ERK and caused a concomitant reduction (∼75%) in GSIS/AASIS and autocrine-dependent activation of PKB. This inhibition could not be attributed to enhanced mitochondrial fatty acid uptake/oxidation or ceramide synthesis, which were unaffected by palmitate. In contrast, diacylglycerol synthesis was elevated suggesting increased palmitate esterification rather than oxidation may contribute to impaired stimulus-secretion coupling. Consistent with this, 2-bromopalmitate, a non-oxidisable palmitate analogue, inhibited GSIS as effectively as palmitate.Conclusions
Our results exclude changes in ceramide content or mitochondrial fatty acid handling as factors initiating palmitate-induced defects in insulin release from MIN6 β cells, but suggest that reduced CaMKII and ERK activation associated with palmitate overload may contribute to impaired stimulus-induced insulin secretion. 相似文献4.
SphK1 and SphK2, sphingosine kinase isoenzymes with opposing functions in sphingolipid metabolism 总被引:9,自引:0,他引:9
Maceyka M Sankala H Hait NC Le Stunff H Liu H Toman R Collier C Zhang M Satin LS Merrill AH Milstien S Spiegel S 《The Journal of biological chemistry》2005,280(44):37118-37129
The potent sphingolipid metabolite sphingosine 1-phosphate is produced by phosphorylation of sphingosine catalyzed by sphingosine kinase (SphK) types 1 and 2. In contrast to pro-survival SphK1, the putative BH3-only protein SphK2 inhibits cell growth and enhances apoptosis. Here we show that SphK2 catalytic activity also contributes to its ability to induce apoptosis. Overexpressed SphK2 also increased cytosolic free calcium induced by serum starvation. Transfer of calcium to mitochondria was required for SphK2-induced apoptosis, as cell death and cytochrome c release was abrogated by inhibition of the mitochondrial Ca(2+) transporter. Serum starvation increased the proportion of SphK2 in the endoplasmic reticulum and targeting SphK1 to the endoplasmic reticulum converted it from anti-apoptotic to pro-apoptotic. Overexpression of SphK2 increased incorporation of [(3)H]palmitate, a substrate for both serine palmitoyltransferase and ceramide synthase, into C16-ceramide, whereas SphK1 decreased it. Electrospray ionizationmass spectrometry/mass spectrometry also revealed an opposite effect on ceramide mass levels. Importantly, specific down-regulation of SphK2 reduced conversion of sphingosine to ceramide in the recycling pathway and conversely, down-regulation of SphK1 increased it. Our results demonstrate that SphK1 and SphK2 have opposing roles in the regulation of ceramide biosynthesis and suggest that the location of sphingosine 1-phosphate production dictates its functions. 相似文献
5.
Acid ceramidase overexpression prevents the inhibitory effects of saturated fatty acids on insulin signaling 总被引:3,自引:0,他引:3
Chavez JA Holland WL Bär J Sandhoff K Summers SA 《The Journal of biological chemistry》2005,280(20):20148-20153
Recent studies indicate that insulin resistance and type 2 diabetes result from the accumulation of lipids in tissues not suited for fat storage, such as skeletal muscle and the liver. To elucidate the mechanisms linking exogenous fats to the inhibition of insulin action, we evaluated the effects of free fatty acids (FFAs) on insulin signal transduction in cultured C2C12 myotubes. As we described previously (Chavez, J. A., and Summers, S. A. (2003) Arch. Biochem. Biophys. 419, 101-109), long-chain saturated FFAs inhibited insulin stimulation of Akt/protein kinase B, a central regulator of glucose uptake and anabolic metabolism. Moreover, these FFAs stimulated the de novo synthesis of ceramide and sphingosine, two sphingolipids shown previously to inhibit insulin action. To determine the contribution of either sphingolipid in FFA-dependent inhibition of insulin action, we generated C2C12 myotubes that constitutively overexpress acid ceramidase (AC), an enzyme that catalyzes the lysosomal conversion of ceramide to sphingosine. AC overexpression negated the inhibitory effects of saturated FFAs on insulin signaling while blocking their stimulation of ceramide accumulation. By contrast, AC overexpression stimulated the accrual of sphingosine. These results support a role for aberrant accumulation of ceramide, but not sphingosine, in the inhibition of muscle insulin sensitivity by exogenous FFAs. 相似文献
6.
Karolina Konstantynowicz-Nowicka Ewa Harasim Marcin Baranowski Adrian Chabowski 《PloS one》2015,10(1)
Aim
There are few and contradictory data on the role of excessive accumulation of intracellular sphingolipids, particularly ceramides, in the development of hepatic insulin resistance. In our study we assessed accumulated sphingolipid fractions and clarify the mechanisms of hepatic insulin resistance development as well as involvement of fatty acid and ceramide transporters in this process.Methods
In culture of primary rat hepatocytes, exposed to high concentration of palmitic acid (0.75mM) during short and prolonged incubation, high performance liquid chromatography was used to assess intra- and extracellular sphingolipid fractions content. Degree of palmitate-induced insulin resistance was estimated by measuring changes in phosphorylation of insulin pathway proteins by western blotting as well as changes in expression of different type of transporters.Results
In our study short and prolonged exposure of primary hepatocytes to palmitic acid resulted in increased intracellular accumulation of ceramide which inhibited insulin signaling pathway. We observed a significant increase in the expression of fatty-acid transport protein (FATP2) and ceramide transfer protein (CERT) what is consistent with enhanced intracellular ceramide content. The content of extracellular ceramide was increased nearly threefold after short and twofold after long incubation period. Expression of microsomal triglyceride transfer protein (MTP) and ATP-binding cassette transporter (ABCA1) was increased significantly mainly after short palmitate incubation.Conclusion
Our data showed that increase in intarcellular ceramide content contributes to the development of hepatic insulin resistance. We suggest pivotal role of transporters in facilitating fatty acid influx (FATP2), accumulation of ceramides (CERT) and export to the media (MTP and ABCA1). 相似文献7.
Huan Yu Xiangxin Che Xiaoyuan Xu Meirong Zheng Yong Zhao Wei He Jingmou Yu Jianjun Xiong Weidong Li 《PloS one》2013,8(12)
Objective
Experimental and clinical studies have shown that administration of insulin during reperfusion is cardioprotective, but the mechanisms underlying this effect are still unknown. In this study, the ability of insulin to protect apoptotic cardiomyocytes from hypoxia/reoxygenation injury using the sphingosine kinase/sphingosine 1-phosphate axis was investigated.Methods and Results
Rat cardiomyocytes were isolated and subjected to hypoxia and reoxygenation. [γ-32P] ATP was used to assess sphingosine kinase activity. Insulin was found to increase sphingosine kinase activity. Immunocytochemistry and Western blot analysis showed changes in the subcellular location of sphingosine kinase 1 from cytosol to the membrane in cardiomyocytes. Insulin caused cardiomyocytes to accumulate of S1P in a dose-dependent manner. FRET efficiency showed that insulin also transactivates the S1P1 receptor. TUNEL staining showed that administration of insulin during reoxygenation could to reduce the rate of reoxygenation-induced apoptosis, which is a requirement for SphK 1 activity. It also reduced the rate of activation of the S1P receptor and inhibited hypoxia/reoxygenation-induced cell death in cardiomyocytes.Conclusion
The sphingosine kinase 1/sphingosine 1-phosphate/S1P receptor axis is one pathway through which insulin protects rat cardiomyocytes from apoptosis induced by hypoxia/reoxygenation injury. 相似文献8.
Suman Bhattacharya Rajeshwary Ghosh Smarajit Maiti Gausal Azam Khan Asru K. Sinha 《PloS one》2013,8(12)
Introduction
Glucose has been reported to have an essential role in the synthesis and secretion of insulin in hepatocytes. As the efflux of glucose is facilitated from the liver cells into the circulation, the mechanism of transportation of glucose into the hepatocytes for the synthesis of insulin was investigated.Methods
Grated liver suspension (GLS) was prepared by grating intact liver from adult mice by using a grater. Nitric oxide (NO) was measured by methemoglobin method. Glucose transporter-4 (Glut-4) was measured by immunoblot technique using Glut-4 antibody.Results
Incubation of GLS with different amounts of glucose resulted in the uptake of glucose by the suspension with increased NO synthesis due to the stimulation of a glucose activated nitric oxide synthase that was present in the liver membrane. The inhibition of glucose induced NO synthesis resulted in the inhibition of glucose uptake. Glucose at 0.02M that maximally increased NO synthesis in the hepatocytes led to the translocation and increased synthesis of Glut-4 by 3.3 fold over the control that was inhibited by the inhibition of NO synthesis. The glucose induced NO synthesis was also found to result in the synthesis of insulin, in the presence of glucose due to the expression of both proinsulin genes I and II in the liver cells.Conclusion
It was concluded that glucose itself facilitated its own transportation in the liver cells both via Glut-4 and by the synthesis of NO which had an essential role for insulin synthesis in the presence of glucose in these cells. 相似文献9.
Sattar Gorgani-Firuzjaee Saeideh Ahmadi Reza Meshkani 《Biochemical and biophysical research communications》2014
Aims
Elevated plasma free fatty acids impair the insulin signaling by induction of the expression of protein phosphatases. However, the effect of palmitate on SH2-containing inositol 5′-phosphatase 2 (SHIP2) expression has not been investigated. Here we investigated the effects of palmitate on SHIP2 expression and elucidated the underlying mechanisms in skeletal muscle cells.Main methods
SHIP2 mRNA and protein levels were measured in C2C12 myotubes exposed to palmitate. Specific inhibitors were used to identify the signaling pathways involved in SHIP2 expression.Key findings
The results showed that 0.5 mM palmitate significantly upregulates the mRNA and protein levels of SHIP2 in C2C12 cells. To address the role of palmitate intracellular metabolites in SHIP2 expression, the myotubes were treated with palmitate in the presence of ceramide and diacylglycerol synthesis inhibitors. The results demonstrated that only ceramide synthesis inhibition could prevent palmitate-induced SHIP2 expression in these cells. In addition, the incubation of muscle cells with different concentrations of C2-ceramide dose-dependently enhanced SHIP2 expression. Furthermore, the inhibition of both JNK and NF-κB pathways could prevent ceramide-induced SHIP2 expression in myotubes.Significance
These findings suggest that palmitate contributes to SHIP2 overexpression in skeletal muscle via the mechanisms involving the activation of ceramide-JNK and NF-κB pathways. 相似文献10.
Turpin SM Lancaster GI Darby I Febbraio MA Watt MJ 《American journal of physiology. Endocrinology and metabolism》2006,291(6):E1341-E1350
Fatty acid-induced apoptosis occurs in pancreatic beta-cells and contributes to the metabolic syndrome. Skeletal muscle insulin resistance is mediated by fatty acid oversupply, which also contributes to the metabolic syndrome. Therefore, we examined whether fatty acids induce apoptosis in skeletal muscle myotubes, the proapoptotic signaling involved, and the effects on insulin sensitivity. Exposure of L6 myotubes to palmitate induced apoptosis, as demonstrated by increased caspase-3 activation, phosphatidylserine exposure on the plasma membrane, and terminal deoxynucleotide transferase dUTP nick end labeling and DNA laddering, both markers of DNA fragmentation. Ceramide content was concomitantly increased, indicating a potential role for ceramides in palmitate-induced apoptosis. Supporting this notion, reducing stearoyl-CoA desaturase-1 (SCD-1) protein content with short interfering RNA resulted in ceramide accumulation and was associated with increased apoptosis in the absence of palmitate. Furthermore, the membrane-permeable C(2)-ceramide enhanced apoptosis in myotubes, whereas the ceramide synthase inhibitor, fumonisin B(1), abrogated the proapoptotic effects of palmitate. Insulin-stimulated glucose uptake was inhibited by palmitate treatment, whereas the addition of effector caspase inhibitors [Ac-DEVD-aldehyde (DEVD-CHO), Z-DQMD-FMK] independently restored >80% of the insulin-stimulated glucose uptake. These effects were observed independently from changes in the protein content of insulin signaling proteins, suggesting that proteosomal degradation is not involved in this process. We conclude that lipoapoptosis occurs in skeletal muscle myotubes, at least partially via de novo ceramide accumulation, and that inhibiting downstream apoptotic signaling improves glucose uptake in vitro. 相似文献
11.
Evgeny V. Berdyshev Irina Gorshkova Anastasia Skobeleva Robert Bittman Xuequan Lu Steven M. Dudek Tamara Mirzapoiazova Joe G. N. Garcia Viswanathan Natarajan 《The Journal of biological chemistry》2009,284(9):5467-5477
Novel immunomodulatory molecule FTY720 is a synthetic analog of myriocin,
but unlike myriocin FTY720 does not inhibit serine palmitoyltransferase.
Although many of the effects of FTY720 are ascribed to its phosphorylation and
subsequent sphingosine 1-phosphate (S1P)-like action through
S1P1,3–5 receptors, studies on modulation of intracellular
balance of signaling sphingolipids by FTY720 are limited. In this study, we
used stable isotope pulse labeling of human pulmonary artery endothelial cells
with l-[U-13C, 15N]serine as well as in
vitro enzymatic assays and liquid chromatography-tandem mass spectrometry
methodology to characterize FTY720 interference with sphingolipid de
novo biosynthesis. In human pulmonary artery endothelial cells, FTY720
inhibited ceramide synthases, resulting in decreased cellular levels of
dihydroceramides, ceramides, sphingosine, and S1P but increased levels of
dihydrosphingosine and dihydrosphingosine 1-phosphate (DHS1P). The
FTY720-induced modulation of sphingolipid de novo biosynthesis was
similar to that of fumonisin B1, a classical inhibitor of ceramide synthases,
but differed in the efficiency to inhibit biosynthesis of short-chain
versus long-chain ceramides. In vitro kinetic studies
revealed that FTY720 is a competitive inhibitor of ceramide synthase 2 toward
dihydrosphingosine with an apparent Ki of 2.15
μm. FTY720-induced up-regulation of DHS1P level was mediated by
sphingosine kinase (SphK) 1, but not SphK2, as confirmed by experiments using
SphK1/2 silencing with small interfering RNA. Our data demonstrate for the
first time the ability of FTY720 to inhibit ceramide synthases and modulate
the intracellular balance of signaling sphingolipids. These findings open a
novel direction for therapeutic applications of FTY720 that focuses on
inhibition of ceramide biosynthesis, ceramide-dependent signaling, and the
up-regulation of DHS1P generation in cells.FTY7202 is a
synthetic analog of sphingosine and is currently being studied as a potent
immunosuppressive and immunomodulatory agent
(1–3).
FTY720-induced immunosuppression is ascribed, in part, to its protective
effect on endothelial cell barrier function that results in inhibition of
lymphocyte egress from lymph nodes and down-regulation of innate and adaptive
immune responses (4). As
endothelial cells predominantly express the sphingosine 1-phosphate 1
(S1P1) receptor and its activation initiates signaling that results
in the assembly of VE-cadherin-based adherens junctions
(5), it is thought that the
phosphorylation of FTY720 and the binding of FTY720-P to the S1P1
receptor determine its effect on vasculature
(1). Recently it became evident
that the action of FTY720 is more complex as several other direct protein
targets were identified. Thus, FTY720 was found to bind to and inhibit the
cannabinoid CB1 receptor (6),
to inhibit cytosolic phospholipase A2 (cPLA2), and to
counteract ceramide 1-phosphate-induced cPLA2 activation
(7). Additionally FTY720 but
not FTY720-P was shown to inhibit S1P lyase
(8), which degrades S1P to
ethanolamine phosphate and (E)-2-hexadecenal and regulates the
removal of sphingoid bases from the cumulative pool of sphingolipids. These
findings characterize FTY720 as a molecule with a multitargeted mode of action
whose cellular effects are complicated by its metabolic transformation to
FTY720-P, a structural and functional analog of S1P.Phosphorylation of FTY720 to FTY720-P by sphingosine kinases (SphKs) is the
only reported metabolic transformation of FTY720 and has been actively
explored because of its link to S1P-mediated signaling
(1,
2,
9,
10). Recent studies suggest
that the endogenous balance between S1P and ceramide molecules regulates
prosurvival and proapoptotic signaling cascades, which determine the outcome
of cellular response to different stress conditions
(11,
12) or the efficiency of
anticancer therapy
(12–14).
However, despite the fact that FTY720 resembles sphingosine (Sph) and is a
substrate of SphK2
(15–17),
there are no reported studies on the effect of FTY720 on the intrinsic balance
of signaling sphingolipids. Metabolic interconnections between proapoptotic
(ceramides) and prosurvival (dihydrosphingosine 1-phosphate (DHS1P)) molecules
are expected because it is known that fumonisin B1 (FB1), an inhibitor of
(dihydro)ceramide synthases, not only blocks the formation of ceramides and
up-regulates the intracellular content of dihydrosphingosine (DHSph) but also
increases the cellular level of DHS1P
(19,
20).In view of these considerations, it is important to know how compounds with
a potential ability to interfere with the sphingolipidome turnover affect the
DHS1P-S1P/ceramide balance in cells. To address this question we have
investigated the effect of FTY720 on metabolic pathways leading to ceramide
and sphingoid base 1-phosphate generation in human pulmonary artery
endothelial cells (HPAECs) by using a stable isotope pulse labeling approach
and quantitative liquid chromatography-tandem mass spectrometry of signaling
sphingolipids. We demonstrate that treatment of HPAECs with FTY720 results in
the inhibition of de novo ceramide formation with a concomitant
increase in DHSph and DHS1P content in cells. Moreover FTY720 showed a direct
inhibition of ceramide synthases in an in vitro assay, albeit it was
less efficient compared with the classical inhibitor of ceramide synthases,
FB1. Our present findings have identified ceramide synthase isozymes as a
novel molecular target for FTY720 action, opening a new direction for its
potential therapeutic application through the inhibition of ceramide
biosynthesis, ceramide-dependent signaling, and the up-regulation of DHS1P
generation in cells. 相似文献
12.
Cornelia Then Simone Wahl Anna Kirchhofer Harald Grallert Susanne Krug Gabi Kastenmüller Werner R?misch-Margl Melina Claussnitzer Thomas Illig Margit Heier Christa Meisinger Jerzy Adamski Barbara Thorand Cornelia Huth Annette Peters Cornelia Prehn Ina Heukamp Helmut Laumen Andreas Lechner Hans Hauner Jochen Seissler 《PloS one》2013,8(10)
13.
《The Journal of nutritional biochemistry》2014,25(12):1309-1316
Interaction between adipocytes and macrophages has been suggested to play a central role in the pathogenesis of obesity. Ceramide, a sphingolipid de novo synthesized from palmitate, is known to stimulate pro-inflammatory cytokine secretion from multiple types of cells. To clarify whether de novo synthesized ceramide contributes to cytokine dysregulation in adipocytes and macrophages, we observed cytokine secretion in mature 3T3-L1 adipocytes (L1) and RAW264.7 macrophages (RAW) cultured alone or co-cultured under the suppression of de novo ceramide synthesis.Palmitate enhanced ceramide accumulation and stimulated the expression and secretion of interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1) in L1. The suppression of serine-palmitoyl transferase, a rate-limiting enzyme of de novo ceramide synthesis, by myriocin or siRNA attenuated those palmitate-induced alterations, and a ceramide synthase inhibitor fumonisin B1 showed similar results. In contrast, the inhibitor of sphingosine kinase or a membrane-permeable ceramide analogue augmented the cytokine secretion. Myriocin effects on the palmitate-induced changes were not abrogated by toll-like receptor-4 blockade. Although palmitate stimulated RAW to secrete tumor necrosis factor-α (TNF-α), it did not significantly increase ceramide content, and neither myriocin nor fumonisin B1 attenuated the TNF-α hypersecretion. The co-culture of L1 with RAW markedly augmented IL-6 and MCP-1 levels in media. Myriocin or fumonisin B1 significantly lowered these cytokine levels and suppressed the gene expression of TNF-α and MCP-1 in RAW and of IL-6 and MCP-1 in L1.In conclusion, de novo synthesized ceramide partially mediates the palmitate effects on pro-inflammatory adipokines and is possibly involved in the interaction with macrophages. 相似文献
14.
Irina Gorshkova Tong Zhou Biji Mathew Jeffrey R. Jacobson Daisuke Takekoshi Palash Bhattacharya Brett Smith Bulent Aydogan Ralph R. Weichselbaum Viswanathan Natarajan Joe G. N. Garcia Evgeny V. Berdyshev 《Journal of lipid research》2012,53(8):1553-1568
The enforcement of sphingosine-1-phosphate (S1P) signaling network protects from radiation-induced pneumonitis. We now demonstrate that, in contrast to early postirradiation period, late postirradiation sphingosine kinase-1 (SphK1) and sphingoid base-1-phosphates are associated with radiation-induced pulmonary fibrosis (RIF). Using the mouse model, we demonstrate that RIF is characterized by a marked upregulation of S1P and dihydrosphingosine-1-phosphate (DHS1P) levels in the lung tissue and in circulation accompanied by increased lung SphK1 expression and activity. Inhibition of sphingolipid de novo biosynthesis by targeting serine palmitoyltransferase (SPT) with myriocin reduced radiation-induced pulmonary inflammation and delayed the onset of RIF as evidenced by increased animal lifespan and decreased expression of markers of fibrogenesis, such as collagen and α-smooth muscle actin (α-SMA), in the lung. Long-term inhibition of SPT also decreased radiation-induced SphK activity in the lung and the levels of S1P-DHS1P in the lung tissue and in circulation. In vitro, inhibition or silencing of serine palmitoyltransferase attenuated transforming growth factor-β1 (TGF-β)-induced upregulation of α-SMA through the negative regulation of SphK1 expression in normal human lung fibroblasts. These data demonstrate a novel role for SPT in regulating TGF-β signaling and fibrogenesis that is linked to the regulation of SphK1 expression and S1P-DHS1P formation. 相似文献
15.
Lawrence G. MillerJr. Jennifer A. Young Swapan K. Ray Guanghu Wang Sharad Purohit Naren L. Banik Somsankar Dasgupta 《Neurochemical research》2017,42(10):2755-2768
Multiple sclerosis (MS) is a demyelinating disorder characterized by massive neurodegeneration and profound axonal loss. Since myelin is enriched with sphingolipids and some of them display toxicity, biological function of sphingolipids in demyelination has been investigated in MS brain tissues. An elevation of sphingosine with a decrease in monoglycosylceramide and psychosine (myelin markers) was observed in MS white matter and plaque compared to normal brain tissue. This indicated that sphingosine toxicity might mediate oligodendrocyte degeneration. To explain the source of sphingosine accumulation, total sphingolipid profile was investigated in Lewis rats after inducing experimental autoimmune encephalomyelitis (EAE) and also in human oligodendrocytes in culture. An intermittent increase in ceramide followed by sphingosine accumulation in EAE spinal cord along with a stimulation of serine-palmitoyltransferase (SPT) activity was observed. Apoptosis was identified in the lumbar spinal cord, the most prominent demyelinating area, in the EAE rats. TNFα and IFNγ stimulation of oligodendrocytes in culture also led to an accumulation of ceramide with an elevation of sphingosine. Ceramide elevation was drastically blocked by myriocin, an inhibitor of SPT, and also by FTY720. Myriocin treatment also protected oligodendrocytes from cytokine mediated apoptosis or programmed cell death. Hence, we propose that sphingosine toxicity may contribute to demyelination in both EAE and MS, and the intermittent ceramide accumulation in EAE may, at least partly, be mediated via SPT activation, which is a novel observation that has not been previously reported. 相似文献
16.
Background
Reactive astrocytes are implicated in the development and maintenance of neuroinflammation in the demyelinating disease multiple sclerosis (MS). The sphingosine kinase 1 (SphK1)/sphingosine1-phosphate (S1P) receptor signaling pathway is involved in modulation of the inflammatory response in many cell types, but the role of S1P receptor subtype 3 (S1P3) signaling and SphK1 in activated rat astrocytes has not been defined.Methodology/Principal Findings
Using immunohistochemistry we observed the upregulation of S1P3 and SphK1 expression on reactive astrocytes and SphK1 on macrophages in MS lesions. Increased mRNA and protein expression of S1P3 and SphK1, as measured by qPCR and Western blotting respectively, was observed after treatment of rat primary astrocyte cultures with the pro-inflammatory stimulus lipopolysaccharide (LPS). Activation of SphK by LPS stimulation was confirmed by SphK activity assay and was blocked by the use of the SphK inhibitor SKI (2-(p-hydroxyanilino)-4-(p-chlorphenyl) thiazole. Treatment of astrocytes with a selective S1P3 agonist led to increased phosphorylation of extracellular signal-regulated kinase (ERK)-1/2), which was further elevated with a LPS pre-challenge, suggesting that S1P3 upregulation can lead to increased functionality. Moreover, astrocyte migration in a scratch assay was induced by S1P and LPS and this LPS-induced migration was sensitive to inhibition of SphK1, and independent of cell proliferation. In addition, S1P induced secretion of the potentially neuroprotective chemokine CXCL1, which was increased when astrocytes were pre-challenged with LPS. A more prominent role of S1P3 signaling compared to S1P1 signaling was demonstrated by the use of selective S1P3 or S1P1 agonists.Conclusion/Significance
In summary, our data demonstrate that the SphK1/S1P3 signaling axis is upregulated when astrocytes are activated by LPS. This signaling pathway appears to play a role in the establishment and maintenance of astrocyte activation. Upregulation of the pathway in MS may be detrimental, e.g. through enhancing astrogliosis, or beneficial through increased remyelination via CXCL1. 相似文献17.
Sebastián D Herrero L Serra D Asins G Hegardt FG 《American journal of physiology. Endocrinology and metabolism》2007,292(3):E677-E686
Oversupply of lipids to skeletal muscle causes insulin resistance by promoting the accumulation of lipid-derived metabolites that inhibit insulin signaling. In this study, we tested the hypothesis that overexpression of carnitine palmitoyltransferase I (CPT I) could protect myotubes from fatty acid-induced insulin resistance by reducing lipid accumulation in the muscle cell. Incubation of L6E9 myotubes with palmitate caused accumulation of triglycerides, diacylgycerol, and ceramide, produced an activation of PKCtheta and PKCzeta, and blocked insulin-stimulated glucose metabolism, reducing insulin-stimulated PKB activity by 60%. Transduction of L6E9 myotubes with adenoviruses encoding for liver CPT I (LCPT I) wild-type (WT), or a mutant form of LCPT I (LCPT I M593S), which is insensitive to malonyl-CoA, produced a twofold increase in palmitate oxidation when LCPT I activity was increased threefold. LCPT I WT and LCPT I M593S-overexpressing L6E9 myotubes showed normal insulin-stimulated glucose metabolism and an improvement in PKB activity when pretreated with palmitate. Moreover, LCPT I WT- and LCPT I M593S-transduced L6E9 myotubes were protected against the palmitate-induced accumulation of diacylglycerol and ceramide and PKCtheta and -zeta activation. These results suggest that LCPT I overexpression protects L6E9 myotubes from fatty acid-induced insulin resistance by inhibiting both the accumulation of lipid metabolites and the activation of PKCtheta and PKCzeta. 相似文献
18.
Emilie Pepin Arisa Higa Carole Schuster-Klein Catherine Bernard Thierry Sulpice Beatrice Guardiola Eric Chevet Thierry Alquier 《PloS one》2014,9(11)
Background
Type 2 diabetes is characterized by pancreatic beta-cell dysfunction and is associated with low-grade inflammation. Recent observations suggest that apoptosis signal-regulating kinase 1 (ASK1) is involved in beta-cell death in response to different stressors. In this study, we tested whether ASK1 deficiency protects beta-cells from glucolipotoxic conditions and cytokines treatment or from glucose homeostasis alteration induced by endotoxemia.Methodology/Principal Findings
Insulin secretion was neither affected upon shRNA-mediated downregulation of ASK1 in MIN6 cells nor in islets from ASK1-deficient mice. ASK1 silencing in MIN6 cells and deletion in islets did not prevent the deleterious effect of glucolipotoxic conditions or cytokines on insulin secretion. However, it protected MIN6 cells from death induced by ER stress or palmitate and islets from short term caspase activation in response to cytokines. Moreover, endotoxemia induced by LPS infusion increased insulin secretion during hyperglycemic clamps but the response was similar in wild-type and ASK1-deficient mice. Finally, insulin sensitivity in the presence of LPS was not affected by ASK1-deficiency.Conclusions/Significance
Our study demonstrates that ASK1 is not involved in beta-cell function and dysfunction but controls stress-induced beta-cell death. 相似文献19.
Christine Bernsmeier Anne C. Meyer-Gerspach Lea S. Blaser Lia Jeker Robert E. Steinert Markus H. Heim Christoph Beglinger 《PloS one》2014,9(1)
Background & Aims
The incretins glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are gastrointestinal peptide hormones regulating postprandial insulin release from pancreatic β-cells. GLP-1 agonism is a treatment strategy in Type 2 diabetes and is evaluated in Non-alcoholic fatty liver disease (NAFLD). However, the role of incretins in its pathophysiology is insufficiently understood. Studies in mice suggest improvement of hepatic steatosis by GLP-1 agonism. We determined the secretion of incretins after oral glucose administration in non-diabetic NAFLD patients.Methods
N = 52 patients (n = 16 NAFLD and n = 36 Non-alcoholic steatohepatitis (NASH) patients) and n = 50 matched healthy controls were included. Standardized oral glucose tolerance test was performed. Glucose, insulin, glucagon, GLP-1 and GIP plasma levels were measured sequentially for 120 minutes after glucose administration.Results
Glucose induced GLP-1 secretion was significantly decreased in patients compared to controls (p<0.001). In contrast, GIP secretion was unchanged. There was no difference in GLP-1 and GIP secretion between NAFLD and NASH subgroups. All patients were insulin resistant, however HOMA2-IR was highest in the NASH subgroup. Fasting and glucose-induced insulin secretion was higher in NAFLD and NASH compared to controls, while the glucose lowering effect was diminished. Concomitantly, fasting glucagon secretion was significantly elevated in NAFLD and NASH.Conclusions
Glucose-induced GLP-1 secretion is deficient in patients with NAFLD and NASH. GIP secretion is contrarily preserved. Insulin resistance, with hyperinsulinemia and hyperglucagonemia, is present in all patients, and is more severe in NASH compared to NAFLD. These pathophysiologic findings endorse the current evaluation of GLP-1 agonism for the treatment of NAFLD. 相似文献20.
Berdyshev EV Gorshkova I Usatyuk P Kalari S Zhao Y Pyne NJ Pyne S Sabbadini RA Garcia JG Natarajan V 《PloS one》2011,6(1):e16571