Bathing suit ichthyosis is caused by transglutaminase-1 deficiency: evidence for a temperaturesensitive phenotype. Hum Mol Genet 15: 3083-97 Persikov AV, Pillitteri RJ, Amin P et al. (2004) Stability related bias in residues replacing glycines within the collagen triple helix (Gly-Xaa-Yaa) in inherited connective tissue disorders. Hum Mutat 24:330-7 Xu K, Nowak I, Kirchner M et al. (2008) Recombinant collagen studies link the severe conformational changes induced by osteo-genesis imperfecta mutations to the disruption of a set of interchain salt bridges. J Biol Chem 283:34337-44 Woodley DT, Hou Y, Martin S et al. (2008) Characterization of molecular mechanisms underlying mutations in dystrophic epidermolysis bullosa using site-directed mutagenesis.

Ceramides (CERs) in human stratum corneum (SC) play physicochemical roles in determining barrier and waterholding functions of the skin, and specific species might be closely related to the regulation of keratinization, together with other CER-related lipids. Structures of those diverse CER species, however, have not been comprehensively revealed. The aim of this study was to characterize overall CER species in the SC. First, we constructed 3D multi-mass chromatograms of the overall CER species, based on normalphase liquid chromatography (NPLC) connected to electrospray ionization-mass spectrometry (ESI-MS) using a gradient elution system and a postcolumn addition of a volatile saltcontaining polar solvent. The CERs targeted from the 3D chromatograms were structurally analyzed using NPLC-ESItandem mass spectrometry (MS/MS), which resulted in the identification of 342 CER species in the inner forearm SC. This led to the discovery of a new CER class consisting of ahydroxy fatty acid and dihydrosphingosine moieties, in addition to the 10 classes generally known. The results also revealed that those CERs contain long-chain (more than C 18 )-containing sphingoids and a great number of isobaric species. These novel results will contribute not only to physiochemical research on CERs in the SC but also to lipidomics approaches to CERs in the skin.-Masukawa, Y

This article is available online at http://www.jlr.org novo pathways, such as alterations of some species of the targeted lipidomes into others. Similar to common quantitative analyses using direct MS or LC-MS that need authentic standards together with internal ones, quantitative lipidomics may also require a standard set of all lipid species targeted, because each lipid species has a different molar response in MS detection except for special cases. However, it is practically impossible to obtain all authentic species, including not only different even straight carbon chains but also odd and/or branched ones. To overcome such limitations, lipidomics researchers have contrived a novel procedure to quantify lipidomes comprehensively. Han and Gross ( 3 ) proposed a quantitative method for analyzing triglyceride molecular species using direct MS, which involves a procedure to calculate response factors for species that are not available as authentic materials, based on differences of the responses in the numbers of total carbons and double bonds among the species. For quantitative determination of phospholipid molecular species, Koivusalo et al. ( 4 ) utilized the following parameAbstract One of the key challenges in lipidomics is to quantify lipidomes of interest, as it is practically impossible to collect all authentic materials covering the targeted lipidomes. For diverse ceramides (CER) in human stratum corneum (SC) that play important physicochemical roles in the skin, we developed a novel method for quantifi cation of the overall CER species by improving our previously reported profi ling technique using normal-phase liquid chromatography-electrospray ionization-mass spectrometry (NPLC-ESI-MS). The use of simultaneous selected ion monitoring measurement of as many as 182 kinds of molecular-related ions enables the highly sensitive detection of the overall CER species, as they can be analyzed in only one SC-stripped tape as small as 5 mm × 10 mm. To comprehensively quantify CERs, including those not available as authentic species, we designed a procedure to estimate their levels using relative responses of representative authentic species covering the species targeted, considering the systematic error based on intra-/inter-day analyses. The CER levels obtained by this method were comparable to those determined by conventional thin-layer chromatography (TLC), which guarantees the validity of this method. , ceramide class consisting of non-hydroxy fatty acids and 4-sphingenines; ESI, electrospray ionization; LOD, limit of detection; LOQ, limit of quantifi cation; NPLC, normal-phase liquid chromatography; RPLC, reversed-phase liquid chromatography; RSD, relative standard deviation; SC, stratum corneum; SIM, selected ion monitoring; S/N, signal to noise; TLC, thinlayer chromatography

Spinal and bulbar muscular atrophy (SBMA) is an inherited neurodegenerative disorder caused by the expansion of the polyglutamine (polyQ) tract of the androgen receptor (AR-polyQ). Characteristics of SBMA include proximal muscular atrophy, weakness, contraction fasciculation and bulbar involvement. MicroRNAs (miRNAs) are a diverse class of highly conserved small RNA molecules that function as crucial regulators of gene expression in animals and plants. Recent functional studies have shown the potent activity of specific miRNAs as disease modifiers both in vitro and in vivo. Thus, potential therapeutic approaches that target the miRNA processing pathway have recently attracted attention. Here we describe a novel therapeutic approach using the adeno-associated virus (AAV) vector–mediated delivery of a specific miRNA for SBMA. We found that miR-196a enhanced the decay of the AR mRNA by silencing CUGBP, Elav-like family member 2 (CELF2). CELF2 directly acted on AR mRNA and enhanced the stability of AR mRNA. Furthermore, we found that the early intervention of miR-196a delivered by an AAV vector ameliorated the SBMA phenotypes in a mouse model. Our results establish the proof of principle that disease-specific miRNA delivery could be useful in neurodegenerative diseases.

A novel method to quantify glycidol fatty acid esters (GEs), supposed to present as food processing contaminants in edible oils, has been developed in combination with double solid-phase extractions (SPEs) and LC-MS measurements. The analytes were five species of synthetic GEs: glycidol palmitic, stearic, oleic, linoleic and linolenic acid esters. The use of selected ion monitoring in a positive ion mode of atmospheric chemical ionization-MS with a reversed-phase gradient LC provided a limit of quantification of 0.0045-0.012 mg/mL for the standard GEs, which enables the detection of GEs in mg ranges per gram of edible oil. Using the double SPE procedure first in reversed-phase and then in normalphase second, allowed large amounts of co-existing acylglycerols in the oils to be removed, which improved the robustness and stability of the method in sequential runs of LC-MS measurements. When the method was used to quantify GEs in three commercial sources of edible oils, the recovery% ranged from 71.3 to 94.6% (average 79.4%) with a relative standard deviation of 2.9-12.1% for the two oils containing triacylglycerols as major components, and ranged from 90.8 to 105.1% (average 97.2%) with a relative standard deviation of 2.1-12.0% for the other, diacylglycerol-rich oil. Although the accuracy and precision of the method may not be yet sufficient, it is useful for determining trace levels of GEs and will be helpful for the quality control of edible oils.