The CD1 family is a large cluster of non-polymorphic, major histocompatibility complex (MHC) class-I-like molecules that bind distinct lipid-based antigens that are recognized by T cells. The most studied group of T cells that interact with lipid antigens are natural killer T (NKT) cells, which characteristically express a semi-invariant T-cell receptor (NKT TCR) that specifically recognizes the CD1 family member, CD1d. NKT-cell-mediated recognition of the CD1d-antigen complex has been implicated in microbial immunity, tumour immunity, autoimmunity and allergy. Here we describe the structure of a human NKT TCR in complex with CD1d bound to the potent NKT-cell agonist a-galactosylceramide, the archetypal CD1d-restricted glycolipid. In contrast to T-cell receptor-peptide-antigen-MHC complexes, the NKT TCR docked parallel to, and at the extreme end of the CD1d-binding cleft, which enables a lock-and-key type interaction with the lipid antigen. The structure provides a basis for the interaction between the highly conserved NKT TCR a-chain and the CD1d-antigen complex that is typified in innate immunity, and also indicates how variability of the NKT TCR b-chain can impact on recognition of other CD1d-antigen complexes. These findings provide direct insight into how a T-cell receptor recognizes a lipid-antigen-presenting molecule of the immune system.

RATIONALE: Pressure ulcers are a major economic and clinical problem in hospitals and intensive care units. Current methods to detect and grade pressure ulcers use subjective measures, and, presently, there are few ways to detect developing pressure ulcers which are not clinically evident. Here we aimed to employ a model to identify if volatile organic compounds (VOCs) released from skin could Leporidae be used to biochemically separate intact skin from skin with experimentally-induced ulcers. METHODS: Four New Zealand female white rabbits underwent experimental ulcer formation on their right ear while their left ear was used as a control. Ulcers were created using 7/16 x 1/16-inch neodymium magnets (KJ Magnetic Inc, Pipersvile, PA) for 60 minutes. The goal was to achieve an erythematous ring without causing skin breakage or necrosis (i.e., an early-stage ulcer). Following ulcer formation, 5-mm diameter circular polydimethylsiloxane sorbent patches (PDMS; 0.6-mm thick, Goodfellow, Coraopolis, PA) were placed in close proximity to the ulcer on the experimental ears and in a similar location on the control ears. These were covered with polytetrafluoroethylene patches and affixed with Tegaderm® dressings. Skin VOCs were sampled with the PDMS for 30 minutes, and patches were removed, placed into sterile, nitrogen-purged borosolicate vials, capped, and routed for gas chromatography-mass spectrometry (GC/MS) analysis. For analysis, the sampled chemicals were desorbed off the patch at 200C, and 500 µL of headspace was aspirated from the vials and injected into a GC/MS instrument. Two rabbits underwent necropsy for ear histology. Data from each group was compared using Student's test for each VOC. t RESULTS: A significant number of emitted VOCs that were clearly different for the background were detected for each category of samples (ulcer vs non-ulcer). We identified two compounds which were unique to the ulcer group, one of which we tentatively identified as benzyl alcohol. A number of compounds were found to differ in abundance between ulcer and non-ulcer samples. Histologic analysis of two rabbits showed 1-1.5 cm erythematous lesions with acute heterophilic dermatitis, edema, and micro-hemorrhage on the ulcerated ears with normal findings on the control ears. CONCLUSIONS: In this small animal model study, we showed that VOCs are released from rabbit skin, VOCs change as a result of induced experimental ulcers, and VOCs likely correlate with histologic evidence of early-stage ulcers. VOCs may be useful for future, non-invasive monitoring of pressure ulcers in humans.

Dupuytren's disease (DD) is an ill-defined fibroproliferative disorder affecting the palm of the hand, resulting in progressive and irreversible digital contracture. In view of the abnormal gene dysregulation found in DD, and its potential effect on metabolites at a functional level, we chose to examine the metabolic profile involved in DD. Using Fourier transform infrared (FT-IR) spectroscopy to generate metabolic fingerprints of cultured cells, we compared the profiles of DD cords and nodules (1) against the unaffected transverse palmar fascia (internal control), (2) against carpal ligamentous fascia (external control), and (3) against fibroblasts from fat surrounding the nodule and skin overlying the nodule (environmental control). We also determined the effects of serial passaging of the cells on DD fingerprints. Subsequently, gas chromatographymass spectrometry (GC-MS) was employed for metabolic profiling in order to identify metabolites characteristic of the DD tissue phenotypes. We developed a robust metabolomic analysis procedure of DD using cultured fibroblasts derived from DD tissues. Our carefully controlled culture conditions, combined with assessment of metabolic phenotypes by FT-IR and GC-MS, enabled us to demonstrate metabolic differences between DD and unaffected transverse palmar fascia and between DD and healthy control tissue. In early passage (0-3) the metabolic differences were clear, but cells from subsequent passages (4-6) started to lose this distinction between diseased and non-diseased origen. The dysregulated metabolites we identified were leucine, phenylalanine, lysine, cysteine, aspartic acid, glycerol-3-phosphate and the vitamin precursor to coenzyme A. Early passage DD cells exhibit a clear metabolic profile, in which central metabolic pathways appear to be involved. Experimental conditions have been identified in which these DD data are reproducible. The experimental reproducibility will be useful in DD diagnostics and for DD systems biology.