Enhanced or Reduced Fetal Growth Induced by Embryo Transfer into Smaller or Larger Breeds Alters Post-Natal Growth and Metabolism in Pre-Weaning Horses

In equids, placentation is diffuse and nutrient supply to the fetus is determined by uterine size. This correlates with maternal size and affects intra-uterine development and subsequent post-natal growth, as well as insulin sensitivity in the newborn. Long-term effects remain to be described. In this study, fetal growth was enhanced or restricted through ET using pony (P), saddlebred (S) and draft (D) horses. Control P-P (n = 21) and S-S (n = 28) pregnancies were obtained by AI. Enhanced and restricted pregnancies were obtained by transferring P or S embryos into D mares (P-D, n = 6 and S-D, n = 8) or S embryos into P mares (S-P, n = 6), respectively. Control and experimental foals were raised by their dams and recipient mothers, respectively. Weight gain, growth hormones and glucose homeostasis were investigated in the foals from birth to weaning. Fetal growth was enhanced in P-D and these foals remained consistently heavier, with reduced T₃ concentrations until weaning compared to P-P. P-D had lower fasting glucose from days 30 to 200 and higher insulin secretion than P-P after IVGTT on day 3. Euglycemic clamps in the immediate post-weaning period revealed no difference in insulin sensitivity between P-D and P-P. Fetal growth was restricted in S-P and these foals remained consistently lighter until weaning compared to S-D, with elevated T₃ concentrations in the newborn compared to S-S. S-P exhibited higher fasting glycemia than S-S and S-D from days 30 to 200. They had higher maximum increment in plasma glucose than S-D after IVGTT on day 3 and clamps on day 200 demonstrated higher insulin sensitivity compared to S-D. Neither the restricted nor the enhanced fetal environment affected IGF-1 concentrations. Thus, enhanced and restricted fetal and post-natal environments had combined effects that persisted until weaning. They induced different adaptive responses in post-natal glucose metabolism: an early insulin-resistance was induced in enhanced P-D, while S-P developed increased insulin sensitivity.     

Effects of Moderate Amounts of Barley in Late Pregnancy on Growth,Glucose Metabolism and Osteoarticular Status of Pre-Weaning Horses

In stud management, broodmares are commonly fed concentrates in late pregnancy. This practice, however, was shown to correlate with an increased incidence of osteochondrosis in foals, which may be related to insulin sensitivity. We hypothesized that supplementation of the mare with barley in the last trimester of pregnancy alters the pre-weaning foal growth, glucose metabolism and osteoarticular status. Here, pregnant multiparous saddlebred mares were fed forage only (group F, n=13) or both forage and cracked barley (group B, n=12) from the 7^th month of pregnancy until term, as calculated to cover nutritional needs of broodmares. Diets were given in two daily meals. All mares and foals returned to pasture after parturition. Post-natal growth, glucose metabolism and osteoarticular status were investigated in pre-weaning foals. B mares maintained an optimal body condition score (>3.5), whereas that of F mares decreased and remained low (<2.5) up to 3 months of lactation, with a significantly lower bodyweight (-7%) than B mares throughout the last 2 months of pregnancy. B mares had increased plasma glucose and insulin after the first meal and after the second meal to a lesser extent, which was not observed in F mares. B mares also had increased insulin secretion during an intravenous glucose tolerance test (IVGTT). Plasma NEFA and leptin were only temporarily affected by diet in mares during pregnancy or in early lactation. Neonatal B foals had increased serum osteocalcin and slightly increased glucose increments and clearance after glucose injection, but these effects had vanished at weaning. Body measurements, plasma IGF-1, T₄, T₃, NEFA and leptin concentrations, insulin secretion during IVGTT, as well as glucose metabolism rate during euglycemic hyperinsulinemic clamps after weaning, did not differ between groups. Radiographic examination of joints indicated increased osteochondrosis relative risk in B foals, but this was not significant. These data demonstrate that B or F maternal nutrition has very few effects on foal growth, endocrinology and glucose homeostasis until weaning, but may induce cartilage lesions.     

Key Role of the GITR/GITRLigand Pathway in the Development of Murine Autoimmune Diabetes: A Potential Therapeutic Target

Background

The cross-talk between pathogenic T lymphocytes and regulatory T cells (Tregs) plays a major role in the progression of autoimmune diseases. Our objective is to identify molecules and/or pathways involved in this interaction and representing potential targets for innovative therapies. Glucocorticoid-induced tumor necrosis factor receptor (GITR) and its ligand are key players in the T effector/Treg interaction. GITR is expressed at low levels on resting T cells and is significantly up-regulated upon activation. Constitutive high expression of GITR is detected only on Tregs. GITR interacts with its ligand mainly expressed on antigen presenting cells and endothelial cells. It has been suggested that GITR triggering activates effector T lymphocytes while inhibiting Tregs thus contributing to the amplification of immune responses. In this study, we examined the role of GITR/GITRLigand interaction in the progression of autoimmune diabetes.

Methods and Findings

Treatment of 10-day-old non-obese diabetic (NOD) mice, which spontaneously develop diabetes, with an agonistic GITR-specific antibody induced a significant acceleration of disease onset (80% at 12 weeks of age). This activity was not due to a decline in the numbers or functional capacity of CD4⁺CD25⁺Foxp3⁺ Tregs but rather to a major activation of ‘diabetogenic’ T cells. This conclusion was supported by results showing that anti-GITR antibody exacerbates diabetes also in CD28^−/− NOD mice, which lack Tregs. In addition, treatment of NOD mice, infused with the diabetogenic CD4⁺BDC2.5 T cell clone, with GITR-specific antibody substantially increased their migration, proliferation and activation within the pancreatic islets and draining lymph nodes. As a mirror image, blockade of the GITR/GITRLigand pathway using a neutralizing GITRLigand-specific antibody significantly protected from diabetes even at late stages of disease progression. Experiments using the BDC2.5 T cell transfer model suggested that the GITRLigand antibody acted by limiting the homing and proliferation of pathogenic T cells in pancreatic lymph nodes.

Conclusion

GITR triggering plays an important costimulatory role on diabetogenic T cells contributing to the development of autoimmune responses. Therefore, blockade of the GITR/GITRLigand pathway appears as a novel promising clinically oriented strategy as GITRLigand-specific antibody applied at an advanced stage of disease progression can prevent overt diabetes.     

A configuration space of homologous proteins conserving mutual information and allowing a phylogeny inference based on pair-wise Z-score probabilities

Background  

Popular methods to reconstruct molecular phylogenies are based on multiple sequence alignments, in which addition or removal of data may change the resulting tree topology. We have sought a representation of homologous proteins that would conserve the information of pair-wise sequence alignments, respect probabilistic properties of Z-scores (Monte Carlo methods applied to pair-wise comparisons) and be the basis for a novel method of consistent and stable phylogenetic reconstruction.     

VE-statin/egfl7 regulates vascular elastogenesis by interacting with lysyl oxidases

We previously characterized VE-statin/egfl7, a protein that is exclusively secreted by endothelial cells and modulates smooth muscle cell migration. Here, we show that VE-statin/egfl7 is the first known natural negative regulator of vascular elastogenesis. Transgenic mice, expressing VE-statin/egfl7 under the control of keratin-14 promoter, showed an accumulation of VE-statin/egfl7 in arterial walls where its presence correlated with an impaired organization of elastic fibres. In vitro, fibroblasts cultured in the presence of VE-statin/egfl7 were unable to deposit elastic fibres due to a deficient conversion of soluble tropoelastin into insoluble mature elastin. VE-statin/egfl7 interacts with the catalytic domain of lysyl oxidase (LOX) enzymes and, in endothelial cells, endogenous VE-statin/egfl7 colocalizes with LoxL2 and inhibits elastic fibre deposition. In contrast, mature elastic fibres are abundantly deposited by endothelial cells that are prevented from producing endogenous VE-statin/egfl7. We propose a model where VE-statin/egfl7 produced by endothelial cells binds to the catalytic domains of enzymes of the LOX family in the vascular wall, thereby preventing the crosslink of tropoelastin molecules into mature elastin polymers and regulating vascular elastogenesis.     

Analysis of the compositional biases in Plasmodium falciparum genome and proteome using Arabidopsis thaliana as a reference

Comparative genomic analysis of the malaria causative agent, Plasmodium falciparum, with other eukaryotes for which the complete genome is available, revealed that the genome from P. falciparum was more similar to the genome of a plant, Arabidopsis thaliana, than to other non-apicomplexan taxa. Plant-like sequences are thought to result from horizontal gene transfers after a secondary endosymbiosis involving an algal ancestor. The use of the A. thaliana genome and proteome as a reference gives an opportunity to refine our understanding of the extreme compositional bias in the P. falciparum genome that leads to a proteome-wide amino acid bias. A set of pairs of non-redundant protein homologues was selected owing to rigorous genome-wide sequence comparison methods. The introduction of A. thaliana as a reference was a mean to weight the magnitude of the protein evolutionary divergence in P. falciparum. The correlation of the amino acid proportions with evolutionary time supports the hypothesis that amino acids encoded by GC-rich codons are directionally substituted into amino acids encoded by AT-rich codons in the P. falciparum proteome. The long-term deviation of codons in malarial sequences appears as a possible consequence of a genome-wide tri-nucleotidic signature imprinting. Additionally, this study suggests possible working guidelines to improve the accuracy of P. falciparum sequence comparisons, for homology searches and phylogenetic studies.     

Selective and effective bactericidal activity of the cobalt (II) cation against Helicobacter pylori

BACKGROUND: Although the anti-Helicobacter pylori activity of bismuth is well established, the therapeutic potential of other metal ions against the organism is not known. MATERIALS AND METHODS: We measured the minimum inhibitory concentrations of a series of metal ions, including several cobalt (II) compounds against four type strains and seven clinical isolates of H. pylori using three standard broth culture media and a defined medium. Other intestinal bacteria were also investigated for specificity of action. RESULTS: Cobalt chloride had marked activity against H. pylori (minimum inhibitory concentration range was 0.03-1.0 mg/l). The effect was specific because other transition metals had no effect and other intestinal bacteria were not affected by cobalt chloride. Activity was attributable to free cobalt ions as ligands inhibited activity in proportion to their affinity for the ions. Inhibition of cobalt activity was also observed in the presence of nickel, in a dose dependent fashion. However, cobalt activity was not directed towards the nickel-dependent urease enzyme because its effect was similar in wild-type and urease negative mutant strains of H. pylori. Finally, the viability of H. pylori was reduced at the same rate with 2 mg/l cobalt as with 1 mg/l amoxicillin. CONCLUSIONS: Cobalt competes for nickel in its acquisition by H. pylori, but mediates toxicity in a nonurease dependent fashion. As cobalt MIC is similar to some antibiotics and 10 to a hundred times lower than for bismuth, cobalt may represent an effective form of therapy for H. pylori infection.     

Construction of non-symmetric substitution matrices derived from proteomes with biased amino acid distributions

Automatic comparison of compositionally biased genomes, such as that of the malarial causative agent Plasmodium falciparum (82% adenosine + thymidine), with genomes of average composition, is currently limited. Indeed, popular tools such as BLAST require that amino acid distributions be similar in aligned sequences. However, the P. falciparum genome is so biased that six amino acids account for more than 50% of the protein composition. One reason for the comparison methods failure lies in the compositional difference between the query and the subject proteomes, which is not taken into account in the amino acid substitution matrices. This paper introduces a method to derive substitution matrices, in particular BLOSUM 62, in the frame of the information theory. It allows the construction of non-symmetrical matrices, taking into account the non-symmetric amino acid distributions. The dirAtPf family of matrices allowing the comparison of P. falciparum and A. thaliana is given as an example. This paper further provides an analysis of the obtained matrices in the frame of the information theory, supporting the discrimination advantage they bring.     

Structure of the BgK-Kv1.1 complex based on distance restraints identified by double mutant cycles. Molecular basis for convergent evolution of Kv1 channel blockers

A structural model of BgK, a sea anemone toxin, complexed with the S5-S6 region of Kv1.1, a voltage-gated potassium channel, was determined by flexible docking under distance restraints identified by a double mutant cycles approach. This structure provides the molecular basis for identifying the major determinants of the BgK-Kv1.1 channel interactions involving the BgK dyad residues Lys(25) and Tyr(26). These interactions are (i) electrostatic interactions between the extremity of Lys(25) side chain and carbonyl oxygen atoms of residues from the channel selectivity filter that may be strengthened by solvent exclusion provided by (ii) hydrophobic interactions involving BgK residues Tyr(26) and Phe(6) and Kv1.1 residue Tyr(379) whose side chain protrudes in the channel vestibule. In other Kv1 channel-BgK complexes, these interactions are likely to be conserved, implicating both conserved and variable residues from the channels. The data suggest that the conservation in sea anemone and scorpion potassium channel blockers of a functional dyad composed of a lysine, and a hydrophobic residue reflects their use of convergent binding solutions based on a crucial interplay between these important conserved interactions.     

A variable residue in the pore of Kv1 channels is critical for the high affinity of blockers from sea anemones and scorpions

Animal toxins are associated with well defined selectivity profiles; however the molecular basis for this property is not understood. To address this issue we refined our previous three-dimensional models of the complex between the sea anemone toxin BgK and the S5-S6 region of Kv1.1 (Gilquin, B., Racape, J., Wrisch, A., Visan, V., Lecoq, A., Grissmer, S., Ménez, A., and Gasparini, S. (2002) J. Biol. Chem. 277, 37406-37413) using a docking procedure that scores and ranks the structures by comparing experimental and back-calculated values of coupling free energies DeltaDeltaGint obtained from double-mutant cycles. These models further highlight the interaction between residue 379 of Kv1.1 and the conserved dyad tyrosine residue of BgK. Because the nature of the residue at position 379 varies from one channel subtype to another, we explored how these natural mutations influence the sensitivity of Kv1 channel subtypes to BgK using binding and electrophysiology experiments. We demonstrated that mutations at this single position indeed suffice to abolish or enhance the sensitivity of Kv1 channels for BgK and other sea anemone and scorpion toxins. Altogether, our data suggest that the residue at position 379 of Kv1 channels controls the affinity of a number of blocking toxins.