Hepatic phosphorylation status of serine/threonine kinase 1, mammalian target of rapamycin signaling proteins, and growth rate in Holstein heifer calves in response to maternal supply of methionine.

The study investigated whether methionine supply during late pregnancy is associated with liver mammalian target of rapamycin (MTOR) pathway phosphorylation, plasma biomarkers, and growth in heifer calves born to cows fed a control diet (CON) or the control diet plus ethylcellulose rumen-protected methionine (MET; 0.09% of dry matter intake) for the last 28 d prepartum. Calves were fed and managed similarly during the first 56 d of age. Plasma was harvested at birth and 2, 7, 21, 42, and 50 d of age and was used for biomarker profiling. Liver biopsies were harvested at 4, 14, 28, and 50 d of age and used for protein expression. Body weight, hip height, hip width, wither height, body length, rectal temperature, fecal score, and respiratory score were measured weekly. Starter intake was measured daily, and average daily gain was calculated during the first 8 wk of age. During the first 7 wk of age, compared with calves in the CON group, calves in the MET group had greater body weight, hip height, wither height, and average daily gain despite similar daily starter intake. Concentration of methionine in plasma was lower at birth but increased markedly at 2 and 7 d of age in MET calves. Plasma insulin, glucose, free fatty acids, and hydroxybutyrate did not differ. A greater ratio of phosphorylated α-serine/threonine kinase (AKT):total AKT protein expression was detected in MET calves, namely due to differences at 4 d of age. The phosphorylated MTOR:total MTOR ratio also was greater in MET calves due to differences at 28 and 50 d (8 d postweaning). The decrease in phosphorylated MTOR:total MTOR between 14 and 28 d in CON calves agreed with the increase in phosphorylated eukaryotic translation initiation factor 4E binding protein 1 (EIF4EBP1):total EIF4EBP1 ratio during the same time frame. The overall expression of phosphorylated ribosomal protein S6 kinase B1 (RPS6KB1):total RPS6KB1 and phosphorylated eukaryotic translation elongation factor 2 (EEF2):total EEF2 was lower in MET calves. Regardless of methionine supply prepartum, there was an 11-fold temporal decrease from 4 to 50 d in phosphorylated AKT:total AKT. Similarly, regardless of methionine supply, there were overall decreases in phosphorylation ratios of AKT, MTOR, RPS6KB1, and eukaryotic translation initiation factor 2A (EIF2A) over time. Data provide evidence of a positive effect of methionine supply during the last month of pregnancy on rates of growth during the first 7 wk of age. Phosphorylation status of some components of the MTOR pathway in neonatal calf liver also was associated with greater maternal supply of methionine. Thus, the data suggest that molecular mechanisms in the liver might be programmed by supply of methionine during late pregnancy. The exact mechanisms coordinating the observed responses remain to be determined

Molecular basis for passive immunotherapy of Alzheimer's disease

Amyloid aggregates of the amyloid-{beta} (A{beta}) peptide are implicated in the pathology of Alzheimer's disease. Anti-A{beta} monoclonal antibodies (mAbs) have been shown to reduce amyloid plaques in vitro and in animal studies. Consequently, passive immunization is being considered for treating Alzheimer's, and anti-A{beta} mAbs are now in phase II trials. We report the isolation of two mAbs (PFA1 and PFA2) that recognize A{beta} monomers, protofibrils, and fibrils and the structures of their antigen binding fragments (Fabs) in complex with the A{beta}(1–8) peptide DAEFRHDS. The immunodominant EFRHD sequence forms salt bridges, hydrogen bonds, and hydrophobic contacts, including interactions with a striking WWDDD motif of the antigen binding fragments. We also show that a similar sequence (AKFRHD) derived from the human protein GRIP1 is able to cross-react with both PFA1 and PFA2 and, when cocrystallized with PFA1, binds in an identical conformation to A{beta}(1–8). Because such cross-reactivity has implications for potential side effects of immunotherapy, our structures provide a template for designing derivative mAbs that target A{beta} with improved specificity and higher affinity

Loss of Npn1 from motor neurons causes postnatal deficits independent from Sema3A signaling

AbstractThe correct wiring of neuronal circuits is of crucial importance for the function of the vertebrate nervous system. Guidance cues like the neuropilin receptors (Npn) and their ligands, the semaphorins (Sema) provide a tight spatiotemporal control of sensory and motor axon growth and guidance. Among this family of guidance partners the Sema3A-Npn1 interaction has been shown to be of great importance, since defective signaling leads to wiring deficits and defasciculation. For the embryonic stage these defects have been well described, however, also after birth the organism can adapt to new challenges by compensational mechanisms. Therefore, we used the mouse lines Olig2-Cre;Npn1cond and Npn1Sema− to investigate how postnatal organisms cope with the loss of Npn1 selectively from motor neurons or a systemic dysfunctional Sema3A-Npn1 signaling in the entire organism, respectively. While in Olig2-Cre+;Npn1cond−/− mice clear anatomical deficits in paw posturing, bone structure, as well as muscle and nerve composition became evident, Npn1Sema− mutants appeared anatomically normal. Furthermore, Olig2-Cre+;Npn1cond mutants revealed a dysfunctional extensor muscle innervation after single-train stimulation of the N.radial. Interestingly, these mice did not show obvious deficits in voluntary locomotion, however, skilled motor function was affected. In contrast, Npn1Sema− mutants were less affected in all behavioral tests and able to improve their performance over time. Our data suggest that loss of Sema3A-Npn1 signaling is not the only cause for the observed deficits in Olig2-Cre+;Npn1cond−/− mice and that additional, yet unknown binding partners for Npn1 may be involved that allow Npn1Sema− mutants to compensate for their developmental deficits

Summary of the ISEV workshop on extracellular vesicles as disease biomarkers, held in Birmingham, UK, during December 2017

This report summarises the presentations and activities of the ISEV Workshop on extracellular vesicle biomarkers held in Birmingham, UK during December 2017. Among the key messages was broad agreement about the importance of biospecimen science. Much greater attention needs to be paid towards the provenance of collected samples. The workshop also highlighted clear gaps in our knowledge about pre-analytical factors that alter extracellular vesicles (EVs). The future utility of certified standards for credentialing of instruments and software, to analyse EV and for tracking the influence of isolation steps on the structure and content of EVs were also discussed. Several example studies were presented, demonstrating the potential utility for EVs in disease diagnosis, prognosis, longitudinal serial testing and stratification of patients. The conclusion of the workshop was that more effort focused on pre-analytical issues and benchmarking of isolation methods is needed to strengthen collaborations and advance more effective biomarkers