Glycosphingolipid expression in spontaneously aborted fetuses and placenta from blood group p women. Evidence for placenta being the primary target for anti-Tja-antibodies

A 12-week-old fetus and one 17-week-old fetus + placenta were obtained after spontaneous abortions from two women of blood group p. The 17-week-old fetus was dissected into intestine, liver, brain and residual tissue. Nonacid glycosphingolipid fractions were prepared from the tissues. Glycolipid characterization was carried out using thin layer chromatography immunostained with monoclonal antibodies and bacteria and by¹H NMR spectroscopy and mass spectrometry. In the placental fraction substantial amounts of globotetraosylceramide (P-antigen) and globotriaosylceramide (P^k-antigen) were identified. In contrast, the fetuses contained only trace amounts of these structures, as revealed by immunostaining. These results indicate that the primary target for the antibodies of the anti-Tj^a serum is the placenta tissue, resulting in termination of the pregnancy.     

Soil acidification as caused by the nitrogen uptake pattern of Scots pine (Pinus sylvestris)

Three-year-old Scots pine (Pinus sylvestris) trees were grown on a sandy forest soil in pots, with the objective to determine their NH₄/NO₃ uptake ratio and proton efflux. N was supplied in three NH₄-N/NO₃-N ratios, 3:1, 1:1 and 1:3, either as ¹⁵NH₄+¹⁴NO₃ or as ¹⁴NH₄+¹⁵NO₃. Total N and ¹⁵N acquisition of different plant parts were measured. Averaged over the whole tree, the NH₄/NO₃ uptake ratios throughout the growing season were found to be 4.2, 2.5, and 1.5 for the three application ratios, respectively. The excess cation-over-anion uptake value (C_a-A_a) appeared to be linearly related to the natural logarithm of the NH₄/NO₃ uptake ratio. Further, this uptake ratio was related to the NH₄/NO₃ ratio of the soil solution. From these relationship it was estimated that Scots pine exhibits an acidifying uptake pattern as long as the contribution of nitrate to the N nutrition is lower than 70%. Under field circumstances root uptake may cause soil acidification in the topsoil, containing the largest part of the root system, and soil alkalization in deeper soil layers.     

Applications of mass spectrometry in metabolomic studies of animal model and invertebrate systems

Metabolomics provides rich datasets for systems biology. Massspectrometric (MS) techniques are rapidly gaining in importancefor untargeted metabolic profiling. In this review, we surveythe various techniques for sample preparation and analysis relatingto the various MS techniques and illustrate the potential ofthese techniques for both observing complete metabolomes anddetecting changes in the metabolism resulting from genetic mutationof other perturbations. The use of some of these techniquesin the study of model organisms including rodent and variousinvertebrate models is described. The invertebrate systems areof particular interest since such organisms have valuable mutantresources, such as RNAi panels directed against nearly all thegenes in the genome. The demonstration that they are readilycompatible with metabolomic approaches is particularly importantfor systems approaches to metabolic pathways.      

Femur/stature ratio and estimates of stature in children.

The present study examines the relationship between femur length and stature in children between the ages of 8 and 18 years. In previous investigations, my colleagues and I reported the surprising finding that femur length bears a nearly constant relationship to stature in adult humans regardless of ethnicity or gender. This earlier study revealed that the femur/stature ratio averages 26.74% in adult humans, and that using the ratio to predict stature from femur length yields remarkably accurate estimates. The current study shows that femur/stature ratios of children between the ages of 8 and 11 differ significantly from their older counterparts. Between the ages of 12 and 18, there are no significant differences due to age in the femur/stature ratio; however, there are significant differences in this age group attributable to gender. This study also shows that the worldwide average adult femur/stature ratio does not adequately describe children in this age range. This study strongly documents the adolescent growth spurt in the femur/stature ratios of both males and females at the precise time one would expect to see the spurt occur (10-12 in females; 12-14 in males). This growth follows a nearly identical trajectory in both genders, with relative femur growth dominating before the peak years of the growth spurt, and relative stature growth dominating afterward. This accounts for the ratio's rise to maximum values just before peak growth, and its decline toward the adult ratio thereafter. These findings require us to use separate adolescent femur/stature ratios of 27.16 (females) and 27.44 (males) to estimate the stature of children between the ages of 12 and 18. Preliminary testing shows these ratios to be more accurate in estimating stature than the properly selected Trotter and Gleser adult regression equation. Use of the adolescent male ratio with the Homo erectus juvenile WT 15000 results in a lower stature estimate (157.4 cm) than previously reported. It is suggested that continued testing of the ratio occur, but that the values herein derived may be useful in routine forensic cases involving children in this age range, and with subadult paleontological specimens.     

Deep sequencing reveals a novel closterovirus associated with wild rose leaf rosette disease

A bizarre virus‐like symptom of a leaf rosette formed by dense small leaves on branches of wild roses (Rosa multiflora Thunb.), designated as ‘wild rose leaf rosette disease’ (WRLRD), was observed in China. To investigate the presumed causal virus, a wild rose sample affected by WRLRD was subjected to deep sequencing of small interfering RNAs (siRNAs) for a complete survey of the infecting viruses and viroids. The assembly of siRNAs led to the reconstruction of the complete genomes of three known viruses, namely Apple stem grooving virus (ASGV), Blackberry chlorotic ringspot virus (BCRV) and Prunus necrotic ringspot virus (PNRSV), and of a novel virus provisionally named ‘rose leaf rosette‐associated virus’ (RLRaV). Phylogenetic analysis clearly placed RLRaV alongside members of the genus Closterovirus, family Closteroviridae. Genome organization of RLRaV RNA (17 653 nucleotides) showed 13 open reading frames (ORFs), except ORF1 and the quintuple gene block, most of which showed no significant similarities with known viral proteins, but, instead, had detectable identities to fungal or bacterial proteins. Additional novel molecular features indicated that RLRaV seems to be the most complex virus among the known genus members. To our knowledge, this is the first report of WRLRD and its associated closterovirus, as well as two ilarviruses and one capilovirus, infecting wild roses. Our findings present novel information about the closterovirus and the aetiology of this rose disease which should facilitate its control. More importantly, the novel features of RLRaV help to clarify the molecular and evolutionary features of the closterovirus.     

Comparative transcriptome analysis of developmental stages of the Limonium bicolor leaf generates insights into salt gland differentiation

With the expansion of saline land worldwide, it is essential to establish a model halophyte to study the salt‐tolerance mechanism. The salt glands in the epidermis of Limonium bicolor (a recretohalophyte) play a pivotal role in salt tolerance by secreting excess salts from tissues. Despite the importance of salt secretion, nothing is known about the molecular mechanisms of salt gland development. In this study, we applied RNA sequencing to profile early leaf development using five distinct developmental stages, which were quantified by successive collections of the first true leaves of L. bicolor with precise spatial and temporal resolution. Specific gene expression patterns were identified for each developmental stage. In particular, we found that genes controlling salt gland differentiation in L. bicolor may evolve in a trichome formation, which was also confirmed by mutants with increased salt gland densities. Genes involved in the special ultrastructure of salt glands were also elucidated. Twenty‐six genes were proposed to participate in salt gland differentiation. Our dataset sheds light on the molecular processes underpinning salt gland development and thus represents a first step towards the bioengineering of active salt‐secretion capacity in crops.     

Exploration of peptides bound to MHC class I molecules in melanoma

Advancements in high‐resolution HPLC and mass spectrometry have reinvigorated the application of this technology to identify peptides eluted from immunopurified MHC class I molecules. Three melanoma cell lines were assessed using w6/32 isolation, peptide elution and HPLC purification; peptides were identified by mass spectrometry. A total of 13 829 peptides were identified; 83–87% of these were 8–11 mers. Only approximately 15% have been described before. Subcellular locations of the source proteins showed even sampling; mRNA expression and total protein length were predictive of the number of peptides detected from a single protein. HLA‐type binding prediction for 10 078 9/10 mer peptides assigned 88–95% to a patient‐specific HLA subtype, revealing a disparity in strength of predicted binding. HLA‐B*27‐specific isolation successfully identified some peptides not found using w6/32. Sixty peptides were selected for immune screening, based on source protein and predicted HLA binding; no new peptides recognized by antimelanoma T cells were discovered. Additionally, mass spectrometry was unable to identify several epitopes targeted ex vivo by one patient's T cells.     

A proteomic profile of synoviocyte lesions microdissected from formalin-fixed paraffin-embedded synovial tissues of rheumatoid arthritis

Background

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by chronic inflammation of the synovial joints. Early intervention followed by early diagnosis can result in disease remission; however, both early stage diagnosis and provision of effective treatment have been impeded by the heterogeneity of RA, which details of pathological mechanism are unclear. Regardless of numerous investigations of RA by means of genomic and proteomic approaches, proteins interplaying in RA synovial tissues that contain various types of synoviocytes, are not yet sufficiently understood. Hence we have conducted an HPLC/mass spectrometry-based exploratory proteomic analysis focusing on synoviocyte lesions laser-microdissected (LMD) from formalin-fixed paraffin-embedded (FFPE) synovial tissues (RA, n = 15; OA, n = 5), where those of Osteoarthritis (OA) were used as the control.

Results

A total of 508 proteins were identified from the RA and OA groups. With the semi-quantitative comparisons, the spectral index (SpI), log2 protein ratio (R_SC) based on spectral counting, and statistical G-test, 98 proteins were found to be significant (pair-wise p < 0.05) to the RA synovial tissues. These include stromelysin-1 (MMP3), proteins S100-A8 and S100-A9, plastin-2, galectin-3, calreticulin, cathepsin Z, HLA-A, HLA-DRB1, ferritin, neutrophil defensin 1, CD14, MMP9 etc.

Conclusions

Our results confirmed the involvement of known RA biomarkers such as stromelysin-1 (MMP3) and proteins S100-A8 and S100-A9, and also that of leukocyte antigens such as HLA-DRB1. Network analyses of protein–protein interaction for those proteins significant to RA revealed a dominant participation of ribosome pathway (p = 5.91 × 10⁻⁴⁵), and, interestingly, the associations of the p53 signaling (p = 2.34 × 10⁻⁵). An involvement of proteins including CD14, S100-A8/S100-A9 seems to suggest an activation of the NF-kB/MAPK signaling pathway. Our strategy of laser-microdissected FFPE-tissue proteomic analysis in Rheumatoid Arthritis thus demonstrated its technical feasibility in profiling proteins expressed in synovial tissues, which may play important roles in the RA pathogenesis.

Electronic supplementary material

The online version of this article (doi:10.1186/s12014-015-9091-8) contains supplementary material, which is available to authorized users.     

Application of inductively coupled plasma sector field mass spectrometry for elemental analysis of urine

An analytical method using double focusing sector field inductively — coupled plasma mass spectrometry (sector field ICPMS) for rapid simultaneous determination of 42 elements in urine is described. Sample preparation consisted of 20-fold dilution with 0.14 mol/l nitric acid in ultrapure water. The importance of controlling possible contamination sources at different sample preparation and analysis stages in order to achieve adequate method detection limits (DL) is emphasized. Correction for matrix effects was made using indium and lutetium as internal standards. Different approaches for accuracy assessment in urine analysis are evaluated. Additional information on trace element concentrations in a urine reference material is given. Between-batch precision was assessed from the analysis of separately prepared aliquots of the reference material and was better than 10% RSD for 32 of the elements. The robustness of the procedure was tested by analysis of about 250 samples in one analytical run lasting more than 50 hours. A statistical summary of results for 19 urine samples from non-exposed subjects is presented. For a majority of the elements tested concentrations were higher than the detection limit of the method.