mTORC1 regulates high levels of protein synthesis in retinal ganglion cells of adult mice.

Mechanistic target of rapamycin (mTOR) and mTOR complex 1 (mTORC1), linchpins of the nutrient sensing and protein synthesis pathways, are present at relatively high levels in the ganglion cell layer (GCL) and retinal ganglion cells (RGCs) of rodent and human retinas. However, the role of mTORCs in the control of protein synthesis in RGC is unknown. Here, we applied the SUrface SEnsing of Translation (SUnSET) method of nascent protein labeling to localize and quantify protein synthesis in the retinas of adult mice. We also used intravitreal injection of an adeno-associated virus 2 vector encoding Cre recombinase in the eyes of mtor- or rptor-floxed mice to conditionally knockout either both mTORCs or only mTORC1, respectively, in cells within the GCL. A novel vector encoding an inactive Cre mutant (CreΔC) served as control. We found that retinal protein synthesis was highest in the GCL, particularly in RGC. Negation of both complexes or only mTORC1 significantly reduced protein synthesis in RGC. In addition, loss of mTORC1 function caused a significant reduction in the pan-RGC marker, RNA-binding protein with multiple splicing, with little decrease of the total number of cells in the RGC layer, even at 25 weeks after adeno-associated virus-Cre injection. These findings reveal that mTORC1 signaling is necessary for maintaining the high rate of protein synthesis in RGCs of adult rodents, but it may not be essential to maintain RGC viability. These findings may also be relevant to understanding the pathophysiology of RGC disorders, including glaucoma, diabetic retinopathy, and optic neuropathies.

Invariant natural killer T cells recognize lipid self antigen induced by microbial danger signals.

Invariant natural killer T cells (iNKT cells) have a prominent role during infection and other inflammatory processes, and these cells can be activated through their T cell antigen receptors by microbial lipid antigens. However, increasing evidence shows that they are also activated in situations in which foreign lipid antigens would not be present, which suggests a role for lipid self antigen. We found that an abundant endogenous lipid, ß-D-glucopyranosylceramide (ß-GlcCer), was a potent iNKT cell self antigen in mouse and human and that its activity depended on the composition of the N-acyl chain. Furthermore, ß-GlcCer accumulated during infection and in response to Toll-like receptor agonists, contributing to iNKT cell activation. Thus, we propose that recognition of ß-GlcCer by the invariant T cell antigen receptor translates innate danger signals into iNKT cell activation.

Metastable 1T'-phase group VIB transition metal dichalcogenide crystals.

Metastable 1T'-phase transition metal dichalcogenides (1T'-TMDs) with semi-metallic natures have attracted increasing interest owing to their uniquely distorted structures and fascinating phase-dependent physicochemical properties. However, the synthesis of high-quality metastable 1T'-TMD crystals, especially for the group VIB TMDs, remains a challenge. Here, we report a general synthetic method for the large-scale preparation of metastable 1T'-phase group VIB TMDs, including WS2, WSe2, MoS2, MoSe2, WS2xSe2(1-x) and MoS2xSe2(1-x). We solve the crystal structures of 1T'-WS2, -WSe2, -MoS2 and -MoSe2 with single-crystal X-ray diffraction. The as-prepared 1T'-WS2 exhibits thickness-dependent intrinsic superconductivity, showing critical transition temperatures of 8.6 K for the thickness of 90.1 nm and 5.7 K for the single layer, which we attribute to the high intrinsic carrier concentration and the semi-metallic nature of 1T'-WS2. This synthesis method will allow a more systematic investigation of the intrinsic properties of metastable TMDs.

Optimal policies for mitigating pandemic costs: a tutorial model.

There have been a number of pharmaceutical and non-pharmaceutical interventions associated with COVID-19 over the past two years. Various non-pharmaceutical interventions were proposed and implemented to control the spread of the COVID-19 pandemic. Most common of these were partial and complete lockdowns that were used in an attempt to minimize the costs associated with mortality, economic losses and social factors, while being subject to constraints such as finite hospital capacity. Here, we use a minimal model posed in terms of optimal control theory to understand the costs and benefits of such strategies. This allows us to determine top-down policies for how to restrict social contact rates given an age-structured model for the dynamics of the disease. Depending on the relative weights allocated to mortality and socioeconomic losses, we see that the optimal strategies range from long-term social-distancing only for the most vulnerable, partial lockdown to ensure not over-running hospitals, and alternating-shifts, all of which lead to significant reduction in mortality and/or socioeconomic losses. Crucially, commonly used strategies that involve long periods of broad lockdown are almost never optimal, as they are highly unstable to reopening and entail high socioeconomic costs. Using parameter estimates from data available for Germany and the USA early in the pandemic, we quantify these policies and use sensitivity analysis in the relevant model parameters and initial conditions to determine the range of robustness of our policies. Finally we also discuss how bottom-up behavioral changes affect the dynamics of the pandemic and show how they can work in tandem with top-down control policies to mitigate pandemic costs even more effectively.

Signaling roles of phosphoinositides in the retina.

The field of phosphoinositide signaling has expanded significantly in recent years. Phosphoinositides (also known as phosphatidylinositol phosphates or PIPs) are universal signaling molecules that directly interact with membrane proteins or with cytosolic proteins containing domains that directly bind phosphoinositides and are recruited to cell membranes. Through the activities of phosphoinositide kinases and phosphoinositide phosphatases, seven distinct phosphoinositide lipid molecules are formed from the parent molecule, phosphatidylinositol. PIP signals regulate a wide range of cellular functions, including cytoskeletal assembly, membrane budding and fusion, ciliogenesis, vesicular transport, and signal transduction. Given the many excellent reviews on phosphoinositide kinases, phosphoinositide phosphatases, and PIPs in general, in this review, we discuss recent studies and advances in PIP lipid signaling in the retina. We specifically focus on PIP lipids from vertebrate (e.g., bovine, rat, mouse, toad, and zebrafish) and invertebrate (e.g., Drosophila, horseshoe crab, and squid) retinas. We also discuss the importance of PIPs revealed from animal models and human diseases, and methods to study PIP levels both in vitro and in vivo. We propose that future studies should investigate the function and mechanism of activation of PIP-modifying enzymes/phosphatases and further unravel PIP regulation and function in the different cell types of the retina.

Premature neonatal gut microbial community patterns supporting an epithelial TLR-mediated pathway for necrotizing enterocolitis.

BACKGROUND: Necrotising enterocolitis (NEC) is a devastating bowel disease, primarily affecting premature infants, with a poorly understood aetiology. Prior studies have found associations in different cases with an overabundance of particular elements of the faecal microbiota (in particular Enterobacteriaceae or Clostridium perfringens), but there has been no explanation for the different results found in different cohorts. Immunological studies have indicated that stimulation of the TLR4 receptor is involved in development of NEC, with TLR4 signalling being antagonised by the activated TLR9 receptor. We speculated that differential stimulation of these two components of the signalling pathway by different microbiota might explain the dichotomous findings of microbiota-centered NEC studies. Here we used shotgun metagenomic sequencing and qPCR to characterise the faecal microbiota community of infants prior to NEC onset and in a set of matched controls. Bayesian regression was used to segregate cases from control samples using both microbial and clinical data. RESULTS: We found that the infants suffering from NEC fell into two groups based on their microbiota; one with low levels of CpG DNA in bacterial genomes and the other with high abundances of organisms expressing LPS. The identification of these characteristic communities was reproduced using an external metagenomic validation dataset. We propose that these two patterns represent the stimulation of a common pathway at extremes; the LPS-enriched microbiome suggesting overstimulation of TLR4, whilst a microbial community with low levels of CpG DNA suggests reduction of the counterbalance to TLR4 overstimulation. CONCLUSIONS: The identified microbial community patterns support the concept of NEC resulting from TLR-mediated pathways. Identification of these signals suggests characteristics of the gastrointestinal microbial community to be avoided to prevent NEC. Potential pre- or pro-biotic treatments may be designed to optimise TLR signalling.

mTORC1 and mTORC2 expression in inner retinal neurons and glial cells.

The retina is one of the most metabolically active tissues, yet the processes that control retinal metabolism remains poorly understood. The mTOR complex (mTORC) that drives protein and lipid biogenesis and autophagy has been studied extensively in regards to retinal development and responses to optic nerve injury but the processes that regulate homeostasis in the adult retina have not been determined. We previously demonstrated that normal adult retina has high rates of protein synthesis compared to skeletal muscle, associated with high levels of mechanistic target of rapamycin (mTOR), a kinase that forms multi-subunit complexes that sense and integrate diverse environmental cues to control cell and tissue physiology. This study was undertaken to: 1) quantify expression of mTOR complex 1 (mTORC1)- and mTORC2-specific partner proteins in normal adult rat retina, brain and liver; and 2) to localize these components in normal human, rat, and mouse retinas. Immunoblotting and immunoprecipitation studies revealed greater expression of raptor (exclusive to mTORC1) and rictor (exclusive for mTORC2) in normal rat retina relative to liver or brain, as well as the activating mTORC components, pSIN1 and pPRAS40. By contrast, liver exhibits greater amounts of the mTORC inhibitor, DEPTOR. Immunolocalization studies for all three species showed that mTOR, raptor, and rictor, as well as most other known components of mTORC1 and mTORC2, were primarily localized in the inner retina with mTORC1 primarily in retinal ganglion cells (RGCs) and mTORC2 primarily in glial cells. In addition, phosphorylated ribosomal protein S6, a direct target of the mTORC1 substrate ribosomal protein S6 kinase beta-1 (S6K1), was readily detectable in RGCs, indicating active mTORC1 signaling, and was preserved in human donor eyes. Collectively, this study demonstrates that the inner retina expresses high levels of mTORC1 and mTORC2 and possesses active mTORC1 signaling that may provide cell- and tissue-specific regulation of homeostatic activity. These findings help to define the physiology of the inner retina, which is key for understanding the pathophysiology of optic neuropathies, glaucoma and diabetic retinopathy.

A non-canonical rhodopsin-mediated insulin receptor signaling pathway in retinal photoreceptor neurons.

We previously reported a ligand-independent and rhodopsin-dependent insulin receptor (IR) neuroprotective signaling pathway in both rod and cone photoreceptor cells, which is activated through protein-protein interaction. Our previous studies were performed with either retina or isolated rod or cone outer segment preparations and the expression of IR signaling proteins were examined. The isolation of outer segments with large portions of the attached inner segments is a technical challenge. Optiprep™ density gradient medium has been used to isolate the cells and subcellular organelles, Optiprep™ is a non-ionic iodixanol-based medium with a density of 1.320 g/mL. We employed this method to examine the expression of IR and its signaling proteins, and activation of one of the downstream effectors of the IR in isolated photoreceptor cells. Identification of the signaling complexes will be helpful for therapeutic targeting in disease conditions.

Defense involvement of piscidin from striped murrel Channa striatus and its peptides CsRG12 and CsLC11 involvement in an antimicrobial and antibiofilm activity.

In this study, we have evaluated bioinformatics characterization and antimicrobial role of two piscidin (Pi) peptide identified from the established transcriptome of striped murrel Channa striatus (Cs). The identified CsPi cDNA contains 256 nucleotides encode a protein with 70 amino acids in length which has two antimicrobial peptides and named CsRG12 and CsLC11. The gene expression analysis with various immune stimulants indicated an induced expression pattern of CsPi. Antibiogram showed that CsRG12 and CsLC11 was active against Staphylococcus aureus ATCC 33592, a major multi-drug resistant (MDR) bacterial pathogen and Bacillus cereus ATCC 2106. The minimum inhibitory concentration (MIC) and antibiofilm assays were conducted to observe the activity of pathogenic bacteria with these derived antimicrobial peptides. Flow cytometry analysis noticed that the CsRG12 and CsLC11 disrupt the membrane formation of S. aureus and B. cereus, which was further assured by scanning electron microscopic (SEM) images that bleb formation leads to disruption around the bacterial membrane. Overall, it is reported that CsPi is involved in innate immunity as the gene expression plays a remarkable role in up and down regulation during infection. In addition, the involvement of peptides in antibiofilm formation and bacterial membrane disruption support its immune character. This study leads to a possibility for the development of therapeutics in aquaculture biotechnology.

Antioxidant molecular mechanism of adenosyl homocysteinase from cyanobacteria and its wound healing process in fibroblast cells.

An antioxidant molecule namely, adenosyl homocysteinase (AHc) was identified from the earlier constructed transcriptome database of Spirulina, where it was cultured in a sulphur deprived condition. From the AHc protein, a small peptide NL13 was identified using bioinformatics tools and was predicted to have antioxidant property. Further, the peptide was synthesised and its antioxidant mechanism was addressed at molecular level. NL13 was subjected to various antioxidant assays including DPPH assay, HARS assay, SARS Assay, NO assay and ABTS assay, where NL13 exhibited significant (P < 0.05) potential antioxidant activity compared to its antioxidant control, Trolox. Cytotoxicity was performed on Human whole blood and the cell viability was performed on VERO fibroblast cells. In both assays, it was found that NL13 did not exhibit any cytotoxic effect towards the cells. Further, the intracellular ROS was performed on Multimode reader followed by imaging on fluorescence microscope which showed scavenging activity even at lower concentration of NL13 (31.2 µM). An effective wound healing property of NL13 on VERO cells was confirmed by analysing the cell migration rate at two different time intervals (24 and 48 h). Overall, the study shows that NL13 peptide scavenges the intracellular oxidative stress.

Palmitic Acid-Rich High-Fat Diet Exacerbates Experimental Pulmonary Fibrosis by Modulating Endoplasmic Reticulum Stress.

The impact of lipotoxicity on the development of lung fibrosis is unclear. Saturated fatty acids, such as palmitic acid (PA), activate endoplasmic reticulum (ER) stress, a cellular stress response associated with the development of idiopathic pulmonary fibrosis (IPF). We tested the hypothesis that PA increases susceptibility to lung epithelial cell death and experimental fibrosis by modulating ER stress. Total liquid chromatography and mass spectrometry were used to measure fatty acid content in IPF lungs. Wild-type mice were fed a high-fat diet (HFD) rich in PA or a standard diet and subjected to bleomycin-induced lung injury. Lung fibrosis was determined by hydroxyproline content. Mouse lung epithelial cells were treated with PA. ER stress and cell death were assessed by Western blotting, TUNEL staining, and cell viability assays. IPF lungs had a higher level of PA compared with controls. Bleomycin-exposed mice fed an HFD had significantly increased pulmonary fibrosis associated with increased cell death and ER stress compared with those fed a standard diet. PA increased apoptosis and activation of the unfolded protein response in lung epithelial cells. This was attenuated by genetic deletion and chemical inhibition of CD36, a fatty acid transporter. In conclusion, consumption of an HFD rich in saturated fat increases susceptibility to lung fibrosis and ER stress, and PA mediates lung epithelial cell death and ER stress via CD36. These findings demonstrate that lipotoxicity may have a significant impact on the development of lung injury and fibrosis by enhancing pro-death ER stress pathways.

Human autoreactive T cells recognize CD1b and phospholipids.

In contrast with the common detection of T cells that recognize MHC, CD1a, CD1c, or CD1d proteins, CD1b autoreactive T cells have been difficult to isolate in humans. Here we report the development of polyvalent complexes of CD1b proteins and carbohydrate backbones (dextramers) and their use in identifying CD1b autoreactive T cells from human donors. Activation is mediated by αß T-cell receptors (TCRs) binding to CD1b-phospholipid complexes, which is sufficient to activate autoreactive responses to CD1b-expressing cells. Using mass spectrometry and T-cell responses to scan through the major classes of phospholipids, we identified phosphatidylglycerol (PG) as the immunodominant lipid antigen. T cells did not discriminate the chemical differences that distinguish mammalian PG from bacterial PG. Whereas most models of T-cell recognition emphasize TCR discrimination of differing self and foreign structures, CD1b autoreactive T cells recognize lipids with dual self and foreign origin. PG is rare in the cellular membranes that carry CD1b proteins. However, bacteria and mitochondria are rich in PG, so these data point to a more general mechanism of immune detection of infection- or stress-associated lipids.

Loss of human disease protein retinitis pigmentosa GTPase regulator (RPGR) differentially affects rod or cone-enriched retina.

It is unclear how genes, such as RPGR (retinitis pigmentosa guanine triphosphatase regulator) that are expressed in both rods and cones, cause variable disease pathogenesis. Using transcriptomic analysis, we show that loss of RPGR in a rod-dominant mouse retina (Rpgr(ko)) results in predominant alterations in genes involved in actin cytoskeletal dynamics, prior to onset of degeneration. We validated these findings and found an increase in activated RhoA-GTP levels and polymerized F-actin in the Rpgr(ko) mouse retina. To assess the effect of the loss of RPGR in the all-cone region of the human retina, we used Nrl(-/-) (neural retina leucine zipper) mice, to generate Rpgr(ko)::Nrl(-/-) double-knock-out (Rpgr-DKO) mice. These mice exhibited supranormal cone response to light and substantially retained retinal architecture. Transcriptomic analysis revealed predominant up-regulation of retinal pigmented epithelium (RPE)-specific genes associated with visual cycle, whereas fatty acid analysis showed mild decrease in docosahexaenoic acid in the retina of the Rpgr-DKO mice when compared with the Nrl(-/-) mice. Our data reveal new insights into distinct intracellular pathways that are involved in RPGR-associated rod and cone dysfunction and provide a platform to design new treatment modalities.

Mechanochemically Processed Nd-Fe-Co-Cr-B Nanoparticles with High Coercivity and Reduced Spin Reorientation Transition Temperature.

Nd-Fe-B magnets, possessing the highest energy product, are extensively used in cutting-edge applications, including electrical machines and electrical vehicles. An environmentally benign and cost effective synthesis method of Cr alloyed Nd2 (Fe,Co)14 B magnetic nanoparticles using a dry mechanochemical process is reported. The method is solvent free, facile, energy efficient and scalable. The reduction of mixed oxides of Nd, Fe, Co, B and Cr is performed by using Ca. The coercivity (HC ) of the nanoparticles is found to depend on the dispersant content, with the highest value obtained for Nd2 (Fe11.25 Co2 Cr0.75 )B with 40 % CaO dispersant. The HC of isolated Nd2 (Fe11.25 Co2 Cr0.75 )B nanoparticles and nanoparticles embedded in a CaO matrix is found to be 11.5 kOe and 14.4 kOe, respectively, largest values for heavy rare earth free Nd-Fe-B nanoparticles with reasonable saturation and remanent magnetization, regardless of synthesis route. Considering the density of Nd2 Fe14 B, an energy product of 14.2 MGOe is obtained for the nanoparticles. The thermal coefficient of remanence and thermal coefficient of coercivity for aligned samples are -0.06 % and -0.29 %, respectively, in the temperature range between 100 K and 400 K. The spin reorientation temperature is found to be ∼30 K less than that of bulk Nd2 Fe14 B magnets.

Redundant and Nonredundant Functions of Akt Isoforms in the Retina.

Serine/threonine kinase Akt is a downstream effector of the phosphoinositide 3-kinase pathway that is involved in many processes, including providing neuroprotection to stressed photoreceptor cells. Akt exists in three isoforms designated as Akt1, Akt2, and Akt3. All of these isoforms are expressed in the retina. We previously reported that Akt2 knockout mice were susceptible to light stress-induced photoreceptor degeneration, whereas Akt1 deletion had no effect on the retina. We hypothesized that the phenotype of Akt2 knockout mice may be due to the inactivation of specific substrate(s) in the retina. Yeast two-hybrid screening of a bovine retinal cDNA library with Akt2 identified a multidomain protein, POSH (plenty of SH3s), that acts as a scaffold for the JNK pathway of neuronal death. Our results suggest a stable interaction between Akt2 and POSH. Previous studies show that overexpression of POSH leads to cell death. The cell death that we observed in Akt2 knockout mice could be due to the absence of inactivation of POSH-mediated JNK signaling in the retina.

Nanoparticles as Delivery Vehicles for the Treatment of Retinal Degenerative Diseases.

Over the last few years, huge progress has been made in the understanding of molecular mechanisms underlying the pathogenesis of retinal degenerative diseases. Such knowledge has led to the development of gene therapy approaches to treat these devastating disorders. Non-viral gene delivery has been recognized as a prospective treatment for retinal degenerative diseases. In this review, we will summarize the constituent characteristics and recent applications of three representative nanoparticles (NPs) in ocular therapy.

Constitutive Activation Mutant mTOR Promote Cone Survival in Retinitis Pigmentosa Mice.

Studies form our laboratory and others show that the oncogenic tyrosine kinase and serine threonine kinase signaling pathways are essential for cone photoreceptor survival. These pathways are downregulated in mouse models of retinal degenerative diseases. In the present study, we found that activation mutants of mTOR delayed the death of cones in a mouse model of retinal degeneration. These studies suggest that oncogenic protein kinases may be useful as therapeutic agents to treat retinal degenerations that affect cones.

Information technology in morbidity management of human lymphatic filariasis-A promising tool in global programme for elimination of lymphatic filariasis.

BACKGROUND & OBJECTIVES: India is a signatory to the 1997 World Health Assembly resolutions on lymphatic fila- riasis, and other neglected tropical diseases, and supports global elimination of lymphatic filariasis by 2020. The global programme to eliminate lymphatic filariasis (GPELF) has two main components, viz. interrupting transmission of LF through mass drug administration; and managing morbidity and preventing disability. Consorted efforts by the Public Health Department in Tamil Nadu state (India) for elimination of LF was launched in the year 1997 concentrating on both the components of the programme. The data on the prevalence of filarial morbidity and its entire management at present is based on manual reports and registers maintained by the field functionaries. To overcome the constraints in the manual reporting, an attempt was made to develop a web-based reporting system with different modules and user-friendly interface. METHODS: The Vellore and Thiruvannamalai districts were selected as a study area. The study was conducted between 2011 and 2014, which revealed that the entire morbidity management was based on the manual formats. Constraints in the present manual reporting were analysed. PHP and MySQL tools were used to generate user friendly modules. Feedback was collected from field functionaries at different health centre levels, on the adequacy in the modules and effectiveness of the web-based reporting system. RESULTS: The online reporting modules facilitated data entry at the health subcentre level itself. Analysis and retrieval of data was facilitated at all other levels in the public health system. The modules also covered details of surgical interventions, ex-gratia payments and other benefits extended to the lymphoedema patients by the Government. INTERPRETATION & CONCLUSION: The usage and feedback of the web-based reporting system appeared very encouraging and reliable, indicating that it can be implemented in health programmes for disease management. This web-based user-friendly online reporting system can contribute largely to achieve the goals of the GPELF; specially after MDA is withdrawn.

Identification of a Potent Microbial Lipid Antigen for Diverse NKT Cells.

Semi-invariant/type I NKT cells are a well-characterized CD1d-restricted T cell subset. The availability of potent Ags and tetramers for semi-invariant/type I NKT cells allowed this population to be extensively studied and revealed their central roles in infection, autoimmunity, and tumor immunity. In contrast, diverse/type II NKT (dNKT) cells are poorly understood because the lipid Ags that they recognize are largely unknown. We sought to identify dNKT cell lipid Ag(s) by interrogating a panel of dNKT mouse cell hybridomas with lipid extracts from the pathogen Listeria monocytogenes. We identified Listeria phosphatidylglycerol as a microbial Ag that was significantly more potent than a previously characterized dNKT cell Ag, mammalian phosphatidylglycerol. Further, although mammalian phosphatidylglycerol-loaded CD1d tetramers did not stain dNKT cells, the Listeria-derived phosphatidylglycerol-loaded tetramers did. The structure of Listeria phosphatidylglycerol was distinct from mammalian phosphatidylglycerol because it contained shorter, fully-saturated anteiso fatty acid lipid tails. CD1d-binding lipid-displacement studies revealed that the microbial phosphatidylglycerol Ag binds significantly better to CD1d than do counterparts with the same headgroup. These data reveal a highly potent microbial lipid Ag for a subset of dNKT cells and provide an explanation for its increased Ag potency compared with the mammalian counterpart.

Activation of iNKT cells by a distinct constituent of the endogenous glucosylceramide fraction.

Invariant natural killer T (iNKT) cells are a specialized T-cell subset that recognizes lipids as antigens, contributing to immune responses in diverse disease processes. Experimental data suggests that iNKT cells can recognize both microbial and endogenous lipid antigens. Several candidate endogenous lipid antigens have been proposed, although the contextual role of specific antigens during immune responses remains largely unknown. We have previously reported that mammalian glucosylceramides (GlcCers) activate iNKT cells. GlcCers are found in most mammalian tissues, and exist in variable molecular forms that differ mainly in N-acyl fatty acid chain use. In this report, we purified, characterized, and tested the GlcCer fractions from multiple animal species. Although activity was broadly identified in these GlcCer fractions from mammalian sources, we also found activity properties that could not be reconciled by differences in fatty acid chain use. Enzymatic digestion of ß-GlcCer and a chromatographic separation method demonstrated that the activity in the GlcCer fraction was limited to a rare component of this fraction, and was not contained within the bulk of ß-GlcCer molecular species. Our data suggest that a minor lipid species that copurifies with ß-GlcCer in mammals functions as a lipid self antigen for iNKT cells.