Stable isotope dilution assay for prostaglandin E metabolite: 24-hour urinary output in healthy male subjects

A stable isotope dilution assay for the major urinary metabolite of prostaglandins E1 and E2 (PGE-M) was developed. After extraction and purification, the compound was converted to the dimethyl ester bis(O-methyloxime)-trimethylsilyl ether derivative and quantified with the 13C-labeled dimethyl ester analog as internal standard. The interassay coefficient of variation was 0.8%, and the intraassay coefficient of variation ranged from 0.9 to 1.7%. The recovery of authentic, underivatized PGE-M added to urine was 99.5 +/- 3.2% (mean +/- SE, N = 9). The equation of the regression curve correlating the amounts added and recovered was Y = 1.05X - 5.84, with R = 0.998. The level of excreted metabolite in healthy male subjects was 13.86 +/- 1.46 micrograms/24 h (N = 24), in good agreement with data from other sources.     

Recombination by resolvase is inhibited by lac repressor simultaneously binding operators between res sites

The Tn3 resolvase requires that the two recombination (res) sites be aligned as direct repeats on the same molecule for efficient recombination to occur. To test whether resolvase must contact the DNA between res sites as predicted by tracking models, we have determined the sensitivity of recombination to protein diffusion blockades. Recombination between two res sites is unaffected either by lac repressor or bacteriophage T7 RNA polymerase being bound between them. Yet recombination is inhibited by lac repressor if the res site is bounded by a lac operator on both sides. We demonstrate that lac repressor will bind to more than one DNA site under the conditions used to assay recombination. This result suggests that lac repressor can inhibit resolvase by forming a DNA loop that isolates a res site topologically. These results do not support a tracking model for resolvase but suggest that the structure and topology of the DNA substrate is important in the formation of a synapse between res sites.     

Association of restriction fragment length polymorphisms of swine leucocyte antigen class I genes with production traits of Duroc and Hampshire boars

Restriction fragment length polymorphism analyses of swine leucocyte antigen (SLA) class I genes were performed on 70 Duroc and 38 Hampshire boars from the 1986-87 national performance tests of each breed in the USA. Few boars were inbred. Southern blotting and hybridization procedures were performed on genomic DNA, isolated from white blood cells, using PvuII endonuclease and a swine major histocompatibility complex (MHC) class I probe. Durocs had an average of 11 restriction fragments, with the most common being in 63% of the boars and the least common appearing in only one boar. Hampshire boars had an average of 12 restriction fragments, with the most common appearing in 73% of the boars and the least common appearing in only one boar. Least squares procedures and stepwise regression methods were used to examine the association between DNA restriction fragments and the selection index (INDEX), average daily gain (ADG), average backfat thickness (BF), loin muscle area (LEA), and age at 104 kg (DAY104). In the Duroc breed one DNA restriction fragment was associated with decreased INDEX (P less than 0.05) and decreased ADG (P less than 0.05) whereas two other fragments were associated with increased BF (P less than 0.05). In the Hampshire breed two restriction fragments were associated with an increase in INDEX (P less than 0.05). Cluster analyses were used to group pigs of each breed on the basis of similar RFLP patterns. One cluster group in the Duroc breed was associated with lower average INDEX values (P less than 0.05), greater average DAY104 (P less than 0.05), and a larger mean LEA (P less than 0.05). In the Hampshire breed one cluster group was associated with lower INDEX (P less than 0.05). These results suggest there may be an association between swine MHC class I genes and performance traits in swine. The use of SLA class I restriction fragments, as genetic markers, may have potential in the future for improving pig performance.     

Modification of prostaglandin metabolism in vivo by long-chain omega-3 polyunsaturates

In a pilot study conducted with a healthy male volunteer we determined that short-term dietary supplementation with fish oil markedly suppresses the systemic production of prostaglandin E (P = 0.04). This biochemical effect is observable after an incubation period of several days. The potential consequences of a reduced PGE synthetic rate on renal function, immune system and vascular dynamics, must be considered in the overall evaluation of the safety of fish oil supplements.     

Species distribution models for Peruvian plantcutter improve with consideration of biotic interactions

Biotic interactions have been controversial in distributional ecology, mainly in regards to whether they have effects over broad extents, with the negative view known as the Eltonian noise hypothesis (ENH). In this study, we evaluated the ENH for Phytotoma raimondii, a restricted‐range Peruvian endemic bird species: we developed models based on 1) only abiotic conditions, 2) only host plant distributions, and 3) both abiotic conditions and host plant distributions; models were evaluated with partial receiver operating characteristic test and Akaike information criteria metrics. We rejected the ENH for this case: biotic interactions improved the model. The frequency with which exceptions to the ENH are detected has important implications for distributional ecology and methods for estimating distributions of species.     

Male reproductive health statement (XIIIth international symposium on Spermatology,may 9th–12th 2018, Stockholm,Sweden

On the occasion of the XIIIth International Symposium on Spermatology held from 9 to 13 May 2018 in Stockholm (Sweden), participants (guest speakers and audience) collectively felt the need to make a public statement on the general issue of male reproductive health. Our intention is to raise awareness of what we believe is a neglected area of research despite alarming situations around the world. The disclosure strategy desired by the co-authors is to bring it to the attention of the greatest number partly by considering co-publication in the various periodicals dealing with Reproductive Biology and Andrology. BaCA’s editorial office accepted this mission and found it natural that our periodical, the official journal of the French Andrology Society (SALF), should carry this message.     

Substantial differences in bias between single‐digest and double‐digest RAD‐seq libraries: A case study

The trade‐offs of using single‐digest vs. double‐digest restriction site‐associated DNA sequencing (RAD‐seq) protocols have been widely discussed. However, no direct empirical comparisons of the two methods have been conducted. Here, we sampled a single population of Gulf pipefish (Syngnathus scovelli) and genotyped 444 individuals using RAD‐seq. Sixty individuals were subjected to single‐digest RAD‐seq (sdRAD‐seq), and the remaining 384 individuals were genotyped using a double‐digest RAD‐seq (ddRAD‐seq) protocol. We analysed the resulting Illumina sequencing data and compared the two genotyping methods when reads were analysed either together or separately. Coverage statistics, observed heterozygosity, and allele frequencies differed significantly between the two protocols, as did the results of selection components analysis. We also performed an in silico digestion of the Gulf pipefish genome and modelled five major sources of bias: PCR duplicates, polymorphic restriction sites, shearing bias, asymmetric sampling (i.e., genotyping fewer individuals with sdRAD‐seq than with ddRAD‐seq) and higher major allele frequencies. This combination of approaches allowed us to determine that polymorphic restriction sites, an asymmetric sampling scheme, mean allele frequencies and to some extent PCR duplicates all contribute to different estimates of allele frequencies between samples genotyped using sdRAD‐seq versus ddRAD‐seq. Our finding that sdRAD‐seq and ddRAD‐seq can result in different allele frequencies has implications for comparisons across studies and techniques that endeavour to identify genomewide signatures of evolutionary processes in natural populations.     

The structure of divalent cation-induced aggregates of PIP2 and their alteration by gelsolin and tau.

Phosphatidylinositol bisphosphate (PIP2) serves as a precursor for diacylglycerol and inositol trisphosphate in signal transduction cascades and regulates the activities of several actin binding proteins that influence the organization of the actin cytoskeleton. Molecules of PIP2 form 6-nm diameter micelles in water, but aggregate into larger, multilamellar structures in physiological concentrations of divalent cations. Electron microscopic analysis of these aggregates reveals that they are clusters of striated filaments, suggesting that PIP2 aggregates form stacks of discoid micelles rather than multilamellar vesicles or inverted hexagonal arrays as previously inferred from indirect observations. The distance between striations within the filaments varies from 4.2 to 5.4 nm and the diameter of the filaments depends on the dehydrated ionic radius of the divalent cation, with average diameters of 19, 12, and 10 nm for filaments formed by Mg2+, Ca2+, and Ba2+, respectively. The structure of the divalent cation-induced aggregates can be altered by PIP2 binding proteins. Gelsolin and the microtubule associated protein tau both affect the formation of aggregates, indicating that tau acts as a PIP2 binding protein in a manner similar to gelsolin. In contrast, another PIP2 binding protein, profilin, does not modify the aggregates.     

Determination of 5-methyltetrahydrofolic acid in human serum by stable-isotope dilution high-performance liquid chromatography-mass spectrometry.

The need for specific and sensitive methods for the determination of distinct serum folates is of high priority in clinical research settings. A stable-isotope liquid chromatography-mass spectrometry (LC/ESI-MS) assay was developed for the quantitative determination of the monoglutamyl form of 5-methyltetrahydrofolic acid (5-MTHFA) in human serum. Serum samples (0.5 ml) were amended with the internal standard, [5-13C5]MTHFA that had been labeled on the glutamic acid portion of the molecule and allowed to equilibrate. The analyte was trapped onto a solid-phase cartridge and then eluted with the HPLC mobile phase. Forty microliters was taken for LC/ESI-MS analysis using electrospray ionization operated in the positive ion mode. Using the standard method of addition of 5-MTHFA to serum, a linear dilution curve (y = 12.777x - 1.404; range 0.94-97 ng x ml(-1)) was constructed. The precision of the method was 5.3% (CV) based on the analysis of four sample replicates. The mass spectrum produced upon collision induced dissociation of the analyte in serum was used to confirm the identity of the 5-MTHFA. The method was applied to the analysis of a set of serum samples that contained standardized concentrations of 5-MTHFA. The determinations of 5-MTHFA in these samples using the LC/ESI-MS procedure were found to be in good agreement with other folate methods. A highly accurate and specific method for the analysis of 5-MTHFA in serum has been developed utilizing stable isotope dilution mass spectrometry.