Growth and differentiation signals regulated by the M-CSF receptor

The normal proto-oncogene c-fms encodes the macrophage growth factor (M-CSF) receptor involved in growth, survival, and differentiation along the monocyte-macrophage lineage of hematopoietic cell development. A major portion of our research concerns unraveling the temporal, molecular, and structural features that determine and regulate these events. Previous results indicated that c-fms can transmit a growth signal as well as a signal for differentiation in the appropriate cells. To investigate the role of the Fms tyrosine autophosphorylation sites in proliferation vs. differentiation signaling, four of these sites were disrupted and the mutant receptors expressed in a clone derived from the myeloid FDC-P1 cell line. These analyses revealed that: (1) none of the four autophosphorylation sites studied (Y697, Y706, Y721, and Y807) are essential for M-CSF-dependent proliferation of the FDC-P1 clone; (2) Y697, Y706, and Y721 sites, located in the kinase insert region of Fms, are not necessary for differentiation but their presence augments this process; and (3) the Y807 site is essential for the Fms differentiation signal: its mutation totally abrogates the differentiation of the FDC-P1 clone and conversely increases the rate of M-CSF-dependent proliferation. This suggests that the Y807 site may control a switch between growth and differentiation. The assignment of Y807 as a critical site for the reciprocal regulation of growth and differentiation may provide a paradigm for Fms involvement in leukemogenesis, and we are currently investigating the downstream signals transmitted by the tyrosine-phosphorylated 807 site. In Fms-expressing FDC-P1 cells, M-CSF stimulation results in the rapid (30 sec) tyrosine phosphorylation of Fms on the five cytoplasmic tyrosine autophosphorylation sites, and subsequent tyrosine phosphorylation of several host cell proteins occurs within 1–2 min. Complexes are formed between Fms and other signal transduction proteins such as Grb2, Shc, Sos1, and p85. In addition, a new signal transduction protein of 150 kDa is detectable in the FDC-P1 cells. The p150 is phosphorylated on tyrosine, and forms a complex with Shc and Grb2. The interaction with Shc occurs via a protein tyrosine binding (PTB) domain at the N-terminus of Shc. The p150 is not detectable in Fms signaling within fibroblasts, yet the PDGF receptor induces the tyrosine phosphorylation of a similarly sized protein. In hematopoietic cells, this protein is involved in signaling by receptors for GM-CSF, IL-3, KL, MPO, and EPO. We have now cloned a cDNA for this protein and found at least one related family member. The related family member is a Fanconia Anemia gene product, and this suggests potential ways the p150 protein may function in Fms signaling. Mol Reprod Dev 46:96–103, 1997. © 1997 Wiley-Liss, Inc.     

Mitigation of reactive metabolite formation for a series of 3-amino-2-pyridone inhibitors of Bruton’s tyrosine kinase (BTK)

Reactive metabolites have been putatively linked to many adverse drug reactions including idiosyncratic toxicities for a number of drugs with black box warnings or withdrawn from the market. Therefore, it is desirable to minimize the risk of reactive metabolite formation for lead molecules in optimization, in particular for non-life threatening chronic disease, to maximize benefit to risk ratio. This article describes our effort in addressing reactive metabolite issues for a series of 3-amino-2-pyridone inhibitors of BTK, e.g. compound 1 has a value of 459 pmol/mg protein in the microsomal covalent binding assay. Parallel approaches were taken to successfully resolve the issues: establishment of a predictive screening assay with correlation association of covalent binding assay, identification of the origin of reactive metabolite formation using MS/MS analysis of HLM as well as isolation and characterization of GSH adducts. This ultimately led to the discovery of compound 7 (RN941) with significantly reduced covalent binding of 26 pmol/mg protein.     

Discovery of the type VII ESX‐1 secretion needle?

Mycobacterium tuberculosis, the etiological agent of human tuberculosis, harbours five ESAT‐6/type VII secretion (ESX/T7S) systems. The first esx gene clusters were identified during the genome‐sequencing project of M. tuberculosis H37Rv. Follow‐up studies revealed additional genes playing important roles in ESX/T7S systems. Among the latter genes, one can find those that encode Pro‐Glu (PE) and Pro‐Pro‐Glu (PPE) proteins as well as a gene cluster that is encoded >260 kb upstream of the esx‐1 locus and encodes ESX‐1 secretion‐associated proteins EspA (Rv3616c), EspC (Rv3615c) and EspD (Rv3614c). The espACD cluster has been suggested to have an important function in ESX‐1 secretion since EspA‐EspC and EsxA–EsxB are mutually co‐dependent on each other for secretion. However, the molecular mechanism of this co‐dependence and interaction between the substrates remained unknown. In this issue of Molecular Microbiology, Lou and colleagues show that EspC forms high‐molecular weight polymerization complexes that resemble selected components of type II, III and/or IV secretion systems of Gram‐negative bacteria. Indeed, EspC‐multimeric complexes form filamentous structures that could well represent a secretion needle of ESX‐1 type VII secretion systems. This exciting observation opens new avenues for research to discover and characterize ESX/T7S components and elucidates the co‐dependence of EsxA/B secretion with EspA/C.     

Usefulness of 1,3 Beta-<Emphasis Type="SmallCaps">d</Emphasis>-Glucan Detection in non-HIV Immunocompromised Mechanical Ventilated Critically Ill Patients with ARDS and Suspected <Emphasis Type="Italic">Pneumocystis jirovecii</Emphasis> Pneumonia

Introduction

Pneumocystis jirovecii pneumonia (PCP) is a major cause of disease in immunocompromised individuals. Diagnosis is typically obtained by microscopy and/or PCR. For ambiguous PCR results, we evaluated the new biomarker 1,3-Beta-d-Glucan (BDG).

Methods

BDG serum levels were assessed and correlated to PCR results in immunosuppressed patients with ARDS.

Results

11 (22%) out of 50 patients had suspected PCP. APACHE II (26 vs. 24; p < 0.002), SOFA score (16 vs. 14; p < 0.010) and mortality rate (34 vs. 69% p < 0.004; 34 vs. 80% p < 0.003) were significantly altered in patients with positive (pPCR) and slightly positive (spPCR) PCJ PCR as compared to patients with no-PCP (nPCP). BDG levels were significantly lower in patients with nPCP (86; 30–315 pg/ml) than in patients with pPCR (589; 356–1000 pg/ml; p < 0.001) and spPCP (398; 297–516 pg/ml; p < 0.004) referring to the cutoff in this study for PCP of 275 pg/ml. An overall sensitivity (S) of 92% (95% CI 86–96%) and specificity (SP) of 84% (95% CI 79–85%) for PCP were found for the BDG Fungitell assay. In detail, S of 98% (95% CI 94–100%) and SP of 86% (95% CI 82–92%) for pPCP and S of 98% (95% CI 96–100%) and SP of 88% (95% CI 86–96%) for spPCO were found.

Conclusion

Serum BDG levels were strongly elevated in PCP, and the negative predictive value is high. BDG could be used as a preliminary test for patients with suspected PCP, especially in patients with slightly positive PCR results.

     

Phylogenetic signals in thermal traits remain stronger in the tropics if we can believe published physiological data. A reply to McKechnie et al., “Data quality problems undermine analyses of endotherm upper critical temperatures”

Due to concerns about data quality, McKechnie, Coe, Gerson, and Wolf ( 2016 ) questioned the conclusions of our study (Khaliq et al., 2015 ) published in this journal. Here, we argue that most of the questioned data points are in fact useful for macrophysiological analyses, mostly because the vast majority of data are explicitly reported in the peer‐reviewed physiological literature. Furthermore, we show that our conclusions remain largely robust irrespective of the data inclusion criterion. While we think that constructive debates about the adequate use of primary data in meta‐studies as well as more transparency in data inclusion criteria are indeed useful, we also emphasize that data suitability should be evaluated in the light of the scope and scale of the study in which they are used. We hope that this discussion will not discourage the exchange between disciplines such as biogeography and physiology, as this integration is needed to address some of the most urgent scientific challenges.     

Cerium Oxide Decorated Polymer Nanofibers as Effective Membrane Reinforcement for Durable,High‐Performance Fuel Cells

High‐power, durable composite fuel cell membranes are fabricated here by direct membrane deposition (DMD). Poly(vinylidene fluoride‐co ‐hexafluoropropylene) (PVDF‐HFP) nanofibers, decorated with CeO₂ nanoparticles are directly electrospun onto gas diffusion electrodes. The nanofiber mesh is impregnated by inkjet‐printed Nafion ionomer dispersion. This results in 12 µm thin multicomponent composite membranes. The nanofibers provide membrane reinforcement, whereas the attached CeO₂ nanoparticles promote improved chemical membrane durability due to their radical scavenging properties. In a 100 h accelerated stress test under hot and dry conditions, the reinforced DMD fuel cell shows a more than three times lower voltage decay rate (0.39 mV h^?1) compared to a comparably thin Gore membrane (1.36 mV h^?1). The maximum power density of the DMD fuel cell drops by 9%, compared to 54% measured for the reference. Impedance spectroscopy reveals that ionic and mass transport resistance of the DMD fuel cell are unaffected by the accelerated stress test. This is in contrast to the reference, where a 90% increase of the mass transport resistance is measured. Energy dispersive X‐ray spectroscopy reveals that no significant migration of cerium into the catalyst layers occurs during degradation. This proves that the PVDF‐HFP backbone provides strong anchoring of CeO₂ in the membrane.     

Real‐time pH monitoring of industrially relevant enzymatic reactions in a microfluidic side‐entry reactor (μSER) shows potential for pH control

Monitoring and control of pH is essential for the control of reaction conditions and reaction progress for any biocatalytic or biotechnological process. Microfluidic enzymatic reactors are increasingly proposed for process development, however typically lack instrumentation, such as pH monitoring. We present a microfluidic side‐entry reactor (μSER) and demonstrate for the first time real‐time pH monitoring of the progression of an enzymatic reaction in a microfluidic reactor as a first step towards achieving pH control. Two different types of optical pH sensors were integrated at several positions in the reactor channel which enabled pH monitoring between pH 3.5 and pH 8.5, thus a broader range than typically reported. The sensors withstood the thermal bonding temperatures typical of microfluidic device fabrication. Additionally, fluidic inputs along the reaction channel were implemented to adjust the pH of the reaction. Time‐course profiles of pH were recorded for a transketolase and a penicillin G acylase catalyzed reaction. Without pH adjustment, the former showed a pH increase of 1 pH unit and the latter a pH decrease of about 2.5 pH units. With pH adjustment, the pH drop of the penicillin G acylase catalyzed reaction was significantly attenuated, the reaction condition kept at a pH suitable for the operation of the enzyme, and the product yield increased. This contribution represents a further step towards fully instrumented and controlled microfluidic reactors for biocatalytic process development.     

Impacts of natural factors and farming practices on greenhouse gas emissions in the North China Plain: A meta‐analysis

Requirements for mitigation of the continued increase in greenhouse gas (GHG ) emissions are much needed for the North China Plain (NCP ). We conducted a meta‐analysis of 76 published studies of 24 sites in the NCP to examine the effects of natural conditions and farming practices on GHG emissions in that region. We found that N₂O was the main component of the area‐scaled total GHG balance, and the CH ₄ contribution was <5%. Precipitation, temperature, soil pH , and texture had no significant impacts on annual GHG emissions, because of limited variation of these factors in the NCP . The N₂O emissions increased exponentially with mineral fertilizer N application rate, with y  =  0.2389e^0.0058x for wheat season and y  =  0.365e^0.0071x for maize season. Emission factors were estimated at 0.37% for wheat and 0.90% for maize at conventional fertilizer N application rates. The agronomic optimal N rates (241 and 185 kg N ha^?1 for wheat and maize, respectively) exhibited great potential for reducing N₂O emissions, by 0.39 (29%) and 1.71 (56%) kg N₂O‐N ha^?1 season^?1 for the wheat and maize seasons, respectively. Mixed application of organic manure with reduced mineral fertilizer N could reduce annual N₂O emissions by 16% relative to mineral N application alone while maintaining a high crop yield. Compared with conventional tillage, no‐tillage significantly reduced N₂O emissions by ~30% in the wheat season, whereas it increased those emissions by ~10% in the maize season. This may have resulted from the lower soil temperature in winter and increased soil moisture in summer under no‐tillage practice. Straw incorporation significantly increased annual N₂O emissions, by 26% relative to straw removal. Our analysis indicates that these farming practices could be further tested to mitigate GHG emission and maintain high crop yields in the NCP .     

Establishing Life Cycle Inventories of Chemicals Based on Differing Data Availability (9 pp)

Goal, Scope and Background In contrast to inventory data of energy and transport processes, public inventory data of chemicals are rather scarce. Chemicals are important to consider in LCA, because they are used in the production of many, if not all, products. Moreover, they may cause considerable environmental impacts. For these reasons, it was one goal of the new ecoinvent database to provide LCI data on chemicals. In this paper, the methods and procedures used for establishing LCIs of chemicals in ecoinvent are presented.Methods Three different approaches are suggested for situations of differing data availability. First, in the case of good data availability, the general quality guidelines of ecoinvent can be followed. Second, a procedure is proposed for the translation of aggregated inventory data (cumulative LCI results) from industry into the ecoinvent format. This approach was used, if adequate unit process data was not available. Third, a procedure is put forward for estimating inventory data using stoichiometric equations from technical literature as a main information source. This latter method was used if no other information was available. The application of each of the three procedures is illustrated with the help of a case study.Results and Conclusion When sufficient information is available to follow the general guidelines of ecoinvent, the resulting dataset is characterized by a high degree of detail, and it is thus of high quality. For chemicals, however, the application of the standard procedure is possible in only a few cases. When using industrial data, the main drawback is the fact that those data are often available only as aggregated data, thus being out of tune with the quality guidelines of ecoinvent and its main aim, the harmonization of LCI data. As a third approach, the use of the stoichiometric reaction equation is used for the compilation of LCI datasets of chemicals. This approach represents an alternative to neglecting chemicals completely, but it contains a high risk to not consider important aspects of the life cycle of the respective substance.Outlook Further work in the area of chemicals should focus on an improvement of datasets, so far established by either of the two estimation procedures (APME method; estimation based on technical literature) described. Besides the improvement of already established inventories, the compilation of further harmonized inventories of specific types of chemicals (e.g. solvents) or of chemicals for new industrial sectors (e.g. electronics industry) are in discussion.