A moderate inflammation caused by the deterioration of housing conditions modifies Trp metabolism but not Trp requirement for growth of post-weaned piglets

Deterioration of the environment in which piglets are housed after weaning induces a moderate inflammatory response and modifies tryptophan (Trp) metabolism that can, in turn, decrease Trp availability for growth. We hypothesised that a Trp supply above the current recommendations may be required to preserve Trp availability and to maximise the growth of pigs suffering from moderate inflammation. The aim of this experiment was to compare growth performance and plasma concentrations of Trp and some of its metabolites in piglets, suffering or not from moderate inflammation, when they were fed diets containing graded levels of standardised ileal digestible (SID) Trp, obtained with the addition of crystalline l-Trp to the same basal diet (15%, 18%, 21% or 24%, relative to SID lysine). Differences in inflammatory status were obtained by housing the pigs under different sanitary conditions. Forty blocks of four littermate piglets each were selected and weaned at 4 weeks of age. The experimental design consisted of a split plot where the housing conditions (moderate inflammation v. control) were used as the main plot and dietary Trp content as the subplot. Body weight gain and feed intake were recorded 3, 5 and 7 weeks after weaning. Blood was sampled 13, 36 and 43 days after weaning to measure plasma concentrations of Trp, kynurenine and nicotinamide (i.e. two metabolites of Trp catabolism) and haptoglobin, a major acute phase protein in pigs. There was no interaction between dietary Trp and inflammatory status, irrespective of the response criterion. Compared with control pigs, pigs housed in poor housing conditions consumed less feed (P < 0.0001), had a lower growth rate (P < 0.001), higher plasma concentrations of haptoglobin (P < 0.05) and lower concentrations of plasma Trp irrespective of the Trp content in the diet. Increasing the Trp content in the diet improved feed intake (P < 0.05), growth rate and feed/gain (P < 0.05), but did not prevent the deterioration of performance induced by moderate inflammation because of poor housing conditions. The results of this study suggest that an inflammatory response caused by poor housing sanitary conditions altered Trp metabolism and growth performance, but this was not prevented by additional dietary crystalline l-Trp.     

mtDNA from hair and nail clarifies the genetic relationship of the 15th century Qilakitsoq Inuit mummies

The 15th century Inuit mummies excavated at Qilakitsoq in Greenland in 1978 were exceptionally well preserved and represent the largest find of naturally mummified specimens from the Arctic. The estimated ages of the individuals, their distribution between two adjacent graves, the results of tissue typing, and incomplete STR results led researchers to conclude that the eight mummies formed two distinct family groups: A grandmother (I/5), two daughters (I/3, I/4), and their two children (I/1, I/2) in one grave, and two sisters (II/6, II/8) and a daughter (II/7) of one of them in the other. Using mtDNA from hair and nail, we have reanalyzed the mummies. The results allowed the unambiguous assignment of each of the mummies to one of three mtDNA haplogroups: A2b (I/5); A2a (I/2, I/3, II/6, II/8); A2a-311 (I/1, I/4, II/7), excluded some of the previous relations, and pointed to new ones. I/5 is not the grandmother/mother of the individuals in Grave I, and she is not maternally related to any of the seven other mummies; I/3 and I/4 are not sisters and II/7 is neither the daughter of II/6 nor of II/8. However, I/1 may be the child of either I/4 or II/7 and these two may be sisters. I/2 may be the son of I/3, who may be the daughter of either II/6 or II/8, and these two may be sisters. The observation of haplogroups A2a and A2b amongst the 550-year-old Inuit puts a lower limit on the age of the two lineages in Greenland.     

Protocol of a prospective study on the diagnostic value of transcranial duplex scanning of the substantia nigra in patients with parkinsonian symptoms

Background  

Parkinson's disease (PD) is the second most common neurodegenerative disorder. As there is no definitive diagnostic test, its diagnosis is based on clinical criteria. Recently transcranial duplex scanning (TCD) of the substantia nigra in the brainstem has been proposed as an instrument to diagnose PD. We and others have found that TCD scanning of substantia nigra duplex is a relatively accurate diagnostic instrument in patients with parkinsonian symptoms. However, all studies on TCD so far have involved well-defined, later-stage PD patients, which will obviously lead to an overestimate of the diagnostic accuracy of TCD.     

Quantitative SUMO-1 modification of a vaccinia virus protein is required for its specific localization and prevents its self-association

Vaccinia virus (VV), the prototype member of the Poxviridae, a family of large DNA viruses, carries out DNA replication in specialized cytoplasmic sites that are enclosed by the rough endoplasmic reticulum (ER). We show that the VV gene product of A40R is quantitatively modified by SUMO-1, which is required for its localization to the ER-enclosed replication sites. Expression of A40R lacking SUMO-1 induced the formation of rod-shaped cytoplasmic aggregates. The latter likely consisted of polymers of nonsumoylated protein, because unmodified A40R interacted with itself, but not with the SUMO-1-conjugated protein. Using a bacterial sumoylation system, we furthermore show that unmodified A40R is mostly insoluble, whereas the modified form is completely soluble. By electron microscopy, the A40R rods seen in cells were associated with the cytosolic side of the ER and induced the apposition of several ER cisternae. A40R is the first example of a poxvirus protein to acquire SUMO-1. Its quantitative SUMO-1 modification is required for its proper localization to the viral "mini-nuclei" and prevents its self-association. The ability of the nonsumoylated A40R to bring ER membranes close together could suggest a role in the fusion of ER cisternae when these coalesce to enclose the VV replication sites.     

PcMtr, an aromatic and neutral aliphatic amino acid permease of Penicillium chrysogenum

The gene encoding an aromatic and neutral aliphatic amino acid permease of Penicillium chrysogenum was cloned, functionally expressed and characterized in Saccharomyces cerevisiae M4276. The permease, designated PcMtr, is structurally and functionally homologous to Mtr of Neurospora crassa, and unrelated to the Amino Acid Permease (AAP) family which includes most amino acid permeases in fungi. Database searches of completed fungal genome sequences reveal that Mtr type permeases are not widely distributed among fungi, suggesting a specialized function.     

A fluorescence cell biology approach to map the second integrin-binding site of talin to a 130-amino acid sequence within the rod domain

The cytoskeletal protein talin, which provides a direct link between integrins and actin filaments, has been shown to contain two distinct binding sites for integrin beta subunits. Here, we report the precise delimitation and a first functional analysis of the talin rod domain integrin-binding site. Partially overlapping cDNAs covering the entire human talin gene were transiently expressed as DsRed fusion proteins in Chinese hamster ovary cells expressing alpha(IIb)beta(3), linked to green fluorescent protein (GFP). Two-color fluorescence analysis of the transfected cells, spread on fibrinogen, revealed distinct subcellular staining patterns including focal adhesion, actin filament, and granular labeling for different talin fragments. The rod domain fragment G (residues 1984-2344), devoid of any known actin- or vinculin-binding sites, colocalized with beta(3)-GFP in focal adhesions. Direct in vitro interaction of fragment G with native platelet integrin alpha(IIb)beta(3) or with the recombinant wild type, but not the Y747A mutant beta(3) cytoplasmic tail, linked to glutathione S-transferase, was demonstrated by surface plasmon resonance analysis and pull-down assays, respectively. Here, we demonstrate for the first time the in vivo relevance of this interaction by fluorescence resonance energy transfer between beta(3)-GFP and DsRed-talin fragment G. Further in vitro pull-down studies allowed us to map out the integrin-binding site within fragment G to a stretch of 130 residues (fragment J, residues 1984-2113) that also localized to focal adhesions. Finally, we show by a cell biology approach that this integrin-binding site within the talin rod domain is important for beta(3)-cytoskeletal interactions but does not participate in alpha(IIb)beta(3) activation.     

Non-cell autonomous influence of the astrocyte system xc? on hypoglycaemic neuronal cell death

Despite longstanding evidence that hypoglycaemic neuronal injury is mediated by glutamate excitotoxicity, the cellular and molecular mechanisms involved remain incompletely defined. Here, we demonstrate that the excitotoxic neuronal death that follows GD (glucose deprivation) is initiated by glutamate extruded from astrocytes via system x_c⁻ – an amino acid transporter that imports l-cystine and exports l-glutamate. Specifically, we find that depriving mixed cortical cell cultures of glucose for up to 8 h injures neurons, but not astrocytes. Neuronal death is prevented by ionotropic glutamate receptor antagonism and is partially sensitive to tetanus toxin. Removal of amino acids during the deprivation period prevents – whereas addition of l-cystine restores – GD-induced neuronal death, implicating the cystine/glutamate antiporter, system x_c⁻. Indeed, drugs known to inhibit system x_c⁻ ameliorate GD-induced neuronal death. Further, a dramatic reduction in neuronal death is observed in chimaeric cultures consisting of neurons derived from WT (wild-type) mice plated on top of astrocytes derived from sut mice, which harbour a naturally occurring null mutation in the gene (Slc7a11) that encodes the substrate-specific light chain of system x_c⁻ (xCT). Finally, enhancement of astrocytic system x_c⁻ expression and function via IL-1β (interleukin-1β) exposure potentiates hypoglycaemic neuronal death, the process of which is prevented by removal of l-cystine and/or addition of system x_c⁻ inhibitors. Thus, under the conditions of GD, our studies demonstrate that astrocytes, via system x_c⁻, have a direct, non-cell autonomous effect on cortical neuron survival.