The Stability of Lipid Rafts-Like Micro-Domains Is Dependent on the Available Amount of Cholesterol

Lipid rafts are sterol and sphingolipid rich membrane domains that possibly may play roles in multiple cellular processes. These domains are still the matter of debate and it is still unknown by which mechanism if any and organisms promote their formation. This study centers on the ease of in vitro formation of lipid rafts-like structures as it relates to the relative availability of sphingolipids, phospholipids, cholesterol, and membrane proteins. Following a 12 h incubation period, isolation and extraction of the lipid rafts-like assemblies, the composition of the structures was evaluated using HPLC. Cholesterol and sphingomyelin were detected at 206 nm and phosphatidylcholine was detected at 254 nm. Identification of lactose permease, a typical membrane protein, was done using FTIR. The thermal stability of the produced structures was also determined. Results show that the addition of cholesterol significantly increased both the amount of insoluble lipid rafts-like structures and their stability, and that the availability of a minimum amount of sphingolipid was necessary to produce larger amounts of more stable structures. However, the addition of phospholipids hindered the formation of lipid rafts-like assemblies and those formed were generally less stable.

Emodin suppresses LPS-induced proinflammatory responses and nuclear factor-B activation by disruption of lipid rafts and TLR-4 recruitment in endothelial cells

Emodin [1,3,8-Trihydroxy-6-methylanthraquinone] has been reported to exhibit vascular anti-inflammatory properties.However,the relevant anti-inflammatory mechanisms are not well understood.The present study was design to explore the molecular target(s) of emodin

Membrane lipid raft organization during cotton fiber development

Background:Cotton fiber is a single-celled seed trichome that originates from the ovule epidermis.It is an excellent model for studying cell elongation.Along with the elongation of cotton fiber cell,the plasma membrane is also extremely expanded.Despite progress in understanding cotton fiber cell elongation,knowledge regarding the relationship of plasma membrane in cotton fiber cell development remains elusive.Methods:The plasma membrane of cotton fiber cells was marked with a low toxic fluorescent dye,di-4-ANEPPDHQ,at different stages of development.Fluorescence images were obtained using a confocal laser scanning microscopy.Subsequently,we investigated the relationship between lipid raft activity and cotton fiber development by calculating generalized polarization(GP values)and dual-channel ratio imaging.Results:We demonstrated that the optimum dyeing conditions were treatment with 3μmol·L-1 di-4-ANEPPDHQ for 5 min at room temperature,and the optimal fluorescence images were obtained with 488 nm excitation and500–580 nm and 620–720 nm dual channel emission.First,we examined lipid raft organization in the course of fiber development.The GP values were high in the fiber elongation stage(5–10 DPA,days past anthesis)and relatively low in the initial(0 DPA),secondary cell wall synthesis(20 DPA),and stable synthesis(30 DPA)stages.The GP value peaked in the 10 DPA fiber,and the value in 30 DPA fiber was the lowest.Furthermore,we examined the differences in lipid raft activity in fiber cells between the short fiber cotton mutant,Li-1,and its wild-type.The GP values of the Li-1 mutant fiber were lower than those of the wild type fiber at the elongation stage,and the GP values of 10 DPA fibers were lower than those of 5 DPA fibers in the Li-1 mutant.Conclusions:We established a system for examining membrane lipid raft activity in cotton fiber cells.We verified that lipid raft activity exhibited a low-high-low change regularity during the development of cotton fiber cell,and the pattern was disrupted in the short lint fiber Li-1 mutant,suggesting that membrane lipid order and lipid raft activity are closely linked to fiber cell development.

Glut-4 is translocated to both caveolae and non-caveolar lipid rafts, but is partially internalized through caveolae in insulin-stimulated adipocytes

Caveolae 和 non-caveolar 类脂化合物椽子是膜 lipidmicrodomains 的二种类型在 adipocytes 在刺激胰岛素的葡萄糖举起起重要作用。为了在这查明他们的特定的功能，处理， caveolae 被 caveolin-1 RNAinterference 切除。在 Cav-1 RNAi adipocytes，也不刺激胰岛素的葡萄糖举起也不 Glut-4 (葡萄糖 transporter 4 ) translocation 到膜类脂化合物微领域被脱离 ofcaveolae 影响。与一个修改蔗糖密度坡度， caveolae 和 non-caveolar 类脂化合物椽子能被分开。在野类型的 3T3-L1 adipocytes， Glut-4 被发现是进 bothcaveolae 和 non-caveolar 类脂化合物椽子的 translocated。在 Cav-1 RNAi adipocytes，然而， Glut-4 在 non-caveolar 类脂化合物椽子主要是局部性的。在胰岛素的移动以后， caveolae 局部性的 Glut-4was 比 non-caveolar 类脂化合物快使内在化联系木排的 Glut-4。从质膜的成为主观 ofGlut-4 显著地在 Cav-1 RNAi adipocytes 被减少。这些结果建议那刺激胰岛素的 Glut-4 translocation 和葡萄糖举起是 caveolae-independentevents。Caveolae 玩在从在胰岛素的移动以后的质膜的 Glut-4 的成为主观的一个角色。

Lipid rafts participate in aberrant degradative autophagic-lysosomal pathway of amyloid-beta peptide in Alzheimer's disease

Amyloid-beta peptide is the main component of amyloid plaques,which are found in Alzheimer’s disease.The generation and deposition of amyloid-beta is one of the crucial factors for the onset and progression of Alzheimer’s disease.Lipid rafts are glycolipid-rich liquid domains of the plasma membrane,where certain types of protein tend to aggregate and intercalate.Lipid rafts are involved in the generation of amyloid-beta oligomers and the formation of amyloid-beta peptides.In this paper,we review the mechanism by which lipid rafts disturb the aberrant degradative autophagic-lysosomal pathway of amyloid-beta,which plays an important role in the pathological process of Alzheimer’s disease.Moreover,we describe this mechanism from the view of the Two-system Theory of fasciology and thus,suggest that lipid rafts may be a new target of Alzheimer’s disease treatment.

Involvement of Lipid Rafts and Cellular Actin in AcMNPV GP64 Distribution and Virus Budding

GP64 is the major envelope glycoprotein associated with the budded virus (BV) of Autographa californica nucleopolyhedrovirus (AcMNPV) and is essential for attachment and budding of BV particles. Confocal microscopy and flotation assays established the presence of lipid raft domains within the plasma membranes of AcMNPV-infected Sf9 cells and suggested the association of GP64 with lipid rafts during infection. GP64 and filamentous actin (F-actin) were found to co-localise at the cell cortex at 24 and 48 hpi and an additional restructuring of F-actin during infection was visualised, resulting in a strongly polarised distribution of both F-actin and GP64 at the cell cortex. Depletion of F-actin, achieved by treatment of Sf9 cells with latrunculin B (LB), resulted in the redistribution of GP64 with significant cytoplasmic aggregation and reduced presence at the plasma membrane. Treatment with LB also resulted in reduced production of BV in Sf9 cells. Analysis of virus gene transcription confirmed this reduction was not due to decreased trafficking of nucleocapsids to the nucleus or to decreased production of infectious progeny nucleocapsids. Reduced BV production due to a lack of GP64 at the plasma membrane of AcMNPV-infected Sf9 cells treated with LB, suggests a key role for F-actin in the egress of BV.

Designing the lipid raft marker protein for synaptic vesicles

Lipid rafts are cholesterol-enriched microdomains and implicated in many essential physiological ac-tivities such as the neurotransmitter release.Many studies have been carried out on the function of rafts inthe plasma membranes,whereas little is known about the information of such microdomains in subcellularcompartments especially synaptic vesicles(SVs).In the well-studied plasma membranes,several proteinshave been recognized as raft markers,which are used to label or trace rafts.But the raft marker proteinon SVs has not been identified yet.Although some SV proteins,including VAMP and CPE,have beenfound in raft fractions,they cannot be used as markers due to their low abundance in rafts.In this work,we designed several chimera proteins and tested their characteristics for using as SV raft makers.First,we detected whether they located in SVs,and then the chimeras exhibiting the better localization in SVswere further examined for their enrichment in raft using detergent treatment and gradient density floatationanalysis.Our results indicate that one of the chimeric proteins is primarily located in SVs and distributedin raft microdomains,which strongly suggests that it could be served as a raft marker for SVs.

Toy model that explains the regulation of cholesterol on lipid rafts

Cholesterol,as a common lipid on mammalian cell membranes,plays an important role in the formation of lipid rafts.Recent experiments suggest that the strength of cholesterol's regulation on lipid rafts can be affected by the length of the unsaturated phospholipid acyl chain on the membrane.In order to understand this observation,a simplified toy model containing three different molecules is proposed in this paper,where the tail length of phospholipids is considered.This model shows the regulation of membrane cholesterol on the phase separation of the lipid mixture and the formation of nano-domains,and also suggests that the configuration entropy of phospholipid tails is an essential factor.

Single virus tracking of Ebola virus entry through lipid rafts in living host cells

Ebola virus(EBOV)is one of the most pathogenic viruses in humans which can cause a lethal hemorrhagic fever.Understanding the cellular entry mechanisms of EBOV can promote the development of new therapeutic strategies to control virus replication and spread.It has been known that EBOV virions bind to factors expressed at the host cell surface.Subsequently,the virions are internalized by a macropinocytosis-like process,followed by being trafficked through early and late endosomes.Recent researches indicate that the entry of EBOV into cells requires integrated and functional lipid rafts.Whilst lipid rafts have been hypothesized to play a role in virus entry,there is a current lack of supporting data.One major technical hurdle is the lack of effective approaches for observing viral entry.To provide evidence on the involvement of lipid rafts in the entry process of EBOV,we generated the fluorescently labeled Ebola virus like particles(VLPs),and utilized single-particle tracking(SPT)to visualize the entry of fluorescent Ebola VLPs in live cells and the interaction of Ebola VLPs with lipid rafts.In this study,we demonstrate the compartmentalization of Ebola VLPs in lipid rafts during entry process,and inform the essential function of lipid rafts for the entry of Ebola virus.As such,our study provides evidence to show that the raft integrity is critical for Ebola virus pathogenesis and that lipid rafts can serve as potential targets for the development of novel therapeutic strategies.

The Functional Roles of Lipid Rafts in T Cell Activation,Immune Diseases and HIV Infection and Prevention

The first appearance of lipid rafts,or lipid rafts-like structure,was occasionally observed by cryo-electronic microscopy in 1980s as cavity,such as caveolae. However,the fully understanding of lipid raft was attributed by the studies of T cell activation,virus entry/budding,and other membrane events. During the interaction of T cell and antigen presenting cell,a highly organized structure is formed at the interface of the two cells,where cholesterol and sphingolipids are enriched,and form a liquid ordered phase that facilitates the signaling proteins on and off. Lipid rafts are also involved in virus entry and assembly. In this review,we will discuss cholesterol-sphingolipid floating microdomain,the lipid raft as a unique compartment of the plasma membrane,with biological functions that ensure correct intracellular traffic of proteins and lipids,such as protein-protein interactions by concentrating certain proteins in these microdomains,while excluding others. We also discuss the disruption of lipid rafts is related to different diseases and aging,and we especially exploit the lipid rafts as pharmaceutical targets for anti-virus and anti-inflammation,particularly a new approach to control HIV infection for AIDS prevention and protection by inhibition or disruption of lipid rafts.

Effect of docosahexaenoic acid on interleukin-2 receptor signaling pathway in lipid rafts

Recent studies have shown that polyunsaturated fatty acids (PUFA) regulated the func-tions of membrane receptors in T cells and suppressed T cell -mediated immune responses. But the molecular mechanisms of immune regulation are not yet elucidated. Lipid rafts are plasma membrane microdomains, in which many receptors localized. The purpose of this study was to investigate the effect of DHA on IL-2R signaling pathway in lipid rafts. We isolated lipid rafts by discontinuous sucrose density gradient ultracentrifugation, and found that DHA could change the composition of lipid rafts and alter the distribution of key molecules of IL-2R signaling pathway, which transferred from lipid rafts to detergent-soluble membrane fractions. These results revealed that DHA treatment increased the proportion of polyunsaturated fatty acids especially n?3 polyunsaturated fatty acids in lipid rafts and changed the lipid environment of membrane microdomains in T cells. Compared with controls, DHA changed the localization of IL-2R, STAT5a and STAT5b in lipid rafts and suppressed the expression of JAK1, JAK3 and tyrosine phosphotyrosine in soluble membrane fractions. Summarily, this study con-cluded the effects of DHA on IL-2R signaling pathway in lipid rafts and explained the regulation of PUFAs in T cell-mediated immune responses.

C2 domain of synaptotagmin I associates with lipid rafts of plasma membrane

In this paper we report that the C2 domain of synaptotagmin I (syt I) could associate with lipid rafts of plasma membrane. We demonstrate that phosphatidylinositol 4,5-bisphosphate (PIP2) in the target membrane and Ca2+ are the key factors to enhance the raft association of the C2 domain. We also found that the raft association of the C2 domain could be fulfilled by either C2A or C2B alone, suggesting that their raft association might be complementary. Finally, we indicate that destroying lipid rafts or blocking syt I-raft association could significantly reduce the Ca2+-driven release of glutamates. Our data indicate that the raft association of the C2 domain might play an important role in the regulated exocytosis.

Lipid Rafts Identified on Synaptic Vesicles from Rat Brain

For a long time, lipid rafts have been thought to participate in regulating neurotransmitter release. However,the existence of lipid rafts on synaptic vesicles (SVs) and the mechanism by which exocytosis-relative proteins distribute on this structure have not been fully investigated. There is also much controver-sial data concerning rafts on SVs and synaptic vesicle proteins which makes the results difficult to interpret. This study systematically analyzed the existence and properties of lipid rafts on purified SVs by sucrose density gradient centrifugation, cholesterol depletion, and temperature variation. The data reveals that typi-cal lipid rafts on SVs are both cholesterol dependent and temperature sensitive. Previous confusing results may have been caused by improper treatment or side effects of particular reagent. We also screened the lateral distribution of major exocytosis-related SV proteins and found that only the synaptobrevin (syb) and synaptotagmin (syt) produce detectable association with lipid rafts in 1% Triton X-100.

Interaction of hyperlipidemia and reactive oxygen species:Insights from the lipid-raft platform

Reactive oxygen species(ROS) and oxidative stress are closely associated with the development of atherosclerosis,and the most important regulator of ROS production in endothelial cells is NADPH oxidase.Activation of NADPH oxidase requires the assembly of multiple subunits into lipid rafts,which include specific lipid components,including free cholesterol and specific proteins.Disorders of lipid metabolism such as hyperlipidemia affect the cellular lipid components included in rafts,resulting in modification of cellular reactions that produce ROS.In the similar manner,several pathways associating ROS production are affected by the presence of lipid disorder through raft compartments.In this manuscript,we review the pathophysiological implications of hyperlipidemia and lipid rafts in the production of ROS.

外周血液中脂筏标记蛋白表达水平与重性抑郁障碍全病程相关性的研究

目的探讨外周血液中脂筏标记蛋白(FLOT)1表达水平与重性抑郁障碍(MDD)全病程的相关性,以及是否能起到预测MDD复发与难治性MDD的作用。方法选取福建省福州神经精神病防治院2022年9月至2023年8月收治的100例MDD患者为试验组,以同期50名健康体检者为对照组。分别采取不同时期(急性期、巩固期、稳定期)MDD患者及对照组的外周血样本,采用实时荧光定量PCR(qRT-PCR)法测定外周血单核细胞FLOT1表达水平,分析FLOT1表达水平变化与生活事件(主要为负性事件)的相关性。结果试验组FLOT1 mRNA相对表达量高于对照组,差异有统计学意义(P<0.05);与治疗前比较,试验组治疗后的FLOT1 mRNA相对表达量明显下降,差异有统计学意义(P<0.05);试验组急性期的FLOT1 mRNA相对表达量明显高于巩固期与稳定期,差异有统计学意义(P<0.05)。试验组急性期汉密顿抑郁量表-17(HAMD-17)评分均高于巩固期、稳定期,差异有统计学意义(P<0.05);稳定期与巩固期比较,差异无统计学意义(P>0.05)。FLOT1 mRNA相对表达量与MDD患者不同时期的HAMD-17评分呈正相关(P<0.05),FLOT1 mRNA相对表达量与负性事件刺激量、生活事件应激总分呈正相关(P<0.05)。结论外周血液中FLOT1表达水平与MDD全病程有相关性,可能是MDD发病的一种生物学标志物。

Linking Lipid Metabolism with Cell Transformation and Tumor Progression

Carbohydrates,lipids,and proteins are the three major nutrients required by the human body.The lipids,comprising triglycerides,phospholipids,and sterols,provide energy and essential fatty acids for the body,and are required for the growth and maintenance of human cells and tissues.A variety of lipid molecules and their intermediates are involved in cell signaling and inflammation,and have been reported to promote tumor transformation and progression.Fatty acid biosynthetic enzymes are also involved in the lipid metabolism of tumors.Dyslipidemia is closely related to many solid tumors,and may both play a role in both tumorigenesis and be a consequence of tumor development.Therefore,abnormal lipid metabolism is strongly associated with tumor transformation and progression.This review discusses the signaling pathways,related genes,enzymes,and inflammatory cell factors involved in tumor lipid metabolism,as well as the roles of dyslipidemia in tumor transformation and progression.We believe the information provided will serve as valuable reference highlighting molecules that can be targeted to improve the treatment of tumors.

The differential protein and lipid compositions of noncaveolar lipid microdomains and caveolae

词法上， caveolae 和类脂化合物椽子是二不同的膜结构。他们经常被报导分享类似的类脂化合物和蛋白质作文，并且被认为是膜类脂化合物 microdomains 的二种子类型。由修改蔗糖密度坡度筹款 centrifugation，它被用来孤立类脂化合物 microdomains，我们能把 caveolae 和 noncaveolar 类脂化合物 microdomains 分开成二不同部分。caveolar 膜是 100-nm 直径的膜泡，与 caveolin-1 和 flotillin-1 充实。noncaveolar 类脂化合物 microdomains 是非结晶的膜并且很可能异构的类脂化合物椽子的结合。他们 caveolin-1 被弄空，更多比 caveolae 与胆固醇和 sphingolipids 被充实。许多膜蛋白质例如像胰岛素的生长 factor-1 受体(膜受体) ， aquaporin-1 (膜 transporter )(抛锚 glycosylphosphatidylinositol 的膜蛋白质和膜 glycoprotein ) ， Thy-1 和 N-cadherin，明确地与 noncaveolar 类脂化合物 microdomains，然而并非与 caveolae 被联系。这些结果显示 caveolae 的类脂化合物和蛋白质作文 noncaveolar 类脂化合物 microdomains 不同于那些。在他们的蛋白质作文的差别暗示这二膜 microdomains 可以有不同细胞的功能。

微RNA-152靶向调控内质网脂质Raft关联蛋白1对甲状腺乳头状癌细胞增殖和迁移的影响

目的探讨微RNA-152(miR-152)对甲状腺乳头状癌(PTC)细胞增殖、侵袭和迁移的影响及其作用机制。方法将对数生长期PTC TPC-1细胞随机分为空白对照组、miR-con组(转染miRNA-mimics阴性对照)和miR-152组(转染miR-152-mimics),另取对数生长期正常甲状腺细胞株Nthy-ori 3-1作为Nthy-ori 3-1组。采用实时荧光定量聚合酶链反应法检测各组细胞中miR-152、内质网脂质Raft关联蛋白1(ERLIN1)mRNA表达,四唑盐比色法及平板克隆形成实验检测各组TPC-1细胞增殖能力,流式细胞仪检测各组TPC-1细胞凋亡情况,Transwell实验检测各组TPC-1细胞迁移、侵袭能力,Western blot法检测各组TPC-1细胞中ERLIN1蛋白表达。结果空白对照组与miR-con组TPC-1细胞中miR-152、ERLIN1 mRNA和蛋白相对表达量、TPC-1细胞增殖抑制率、细胞克隆数、细胞凋亡率、细胞迁移数及侵袭数比较差异均无统计学意义(P>0.05)。空白对照组、miR-con组、miR-152组TPC-1细胞中miR-152相对表达量显著低于Nthy-ori 3-1组(P<0.05);miR-152组TPC-1细胞中miR-152相对表达量显著高于空白对照组和miR-con组(P<0.05)。与空白对照组、miR-con组比较,miR-152组TPC-1细胞增殖抑制率、细胞凋亡率显著升高,细胞克隆数、细胞迁移数及侵袭数显著降低(P<0.05)。miR-152组TPC-1细胞中ERLIN1 mRNA及蛋白相对表达量均显著低于空白对照组和miR-con组(P<0.05)。结论过表达miR-152可下调TPC-1细胞中ERLIN1的表达,抑制TPC-1细胞增殖、侵袭和迁移,miR-152可能通过靶向调控ERLIN1参与PTC进展和转移。

Curvature-induced lipid segregation

We investigate how an externally imposed curvature influences lipid segregation on two-phase-coexistent membranes.We show that the bending-modulus contrast of the two phases and the curvature act together to yield a reduced effective line tension.On largely curved membranes,a state of multiple domains(or rafts) forms due to a mechanism analogous to that causing magnetic-vortex formation in type-II superconductors.We determine the criterion for such a multi-domain state to occur;we then calculate respectively the size of the domains formed on cylindrically and spherically curved membranes.