Antiinflammatory Activity of a Novel Folic Acid Targeted Conjugate of the mTOR Inhibitor Everolimus

Folate receptor (FR)-β has been identified as a promising target for antimacrophage and antiinflammatory therapies. In the present study, we investigated EC0565, a folic acid–derivative of everolimus, as a FR-specific inhibitor of the mammalian target of rapamycin (mTOR). Because of its amphiphilic nature, EC0565 was first evaluated for water solubility, critical micelle formation, stability in culture and FR-binding specificity. Using FR-expressing macrophages, the effect of EC0565 on mTOR signaling and cellular proliferation was studied. The pharmacokinetics, metabolism and bioavailability of EC0565 were studied in normal rats. The in vivo activity of EC0565 was assessed in rats with adjuvant arthritis, a “macrophage-rich” model with close resemblance to rheumatoid arthritis. EC0565 forms micellar aggregates in physiological buffers and demonstrates good water solubility as well as strong multivalent FR-binding capacity. EC0565 inhibited mTOR signaling in rat macrophages at nanomolar concentrations and induced G0/G1 cell cycle arrest in serum-starved RAW264.7 cells. Subcutaneously administered EC0565 in rats displayed good bioavailability and a relatively long half-life (~12 h). When given at 250 nmol/kg, EC0565 selectively inhibited proliferating cell nuclear antigen expression in thioglycollate-stimulated rat peritoneal cells. With limited dosing regimens, the antiarthritic activity of EC0565 was found superior to that of etanercept, everolimus and a nontargeted everolimus analog. The in vivo activity of EC0565 was also comparable to that of a folate-targeted aminopterin. Folate-targeted mTOR inhibition may be an effective way of suppressing activated macrophages in sites of inflammation, especially in nutrient-deprived conditions, such as in the arthritic joints. Further investigation and improvement upon the physical and biochemical properties of EC0565 are warranted.     

Dietary exposure to an environmental toxin triggers neurofibrillary tangles and amyloid deposits in the brain

Neurofibrillary tangles (NFT) and β-amyloid plaques are the neurological hallmarks of both Alzheimer''s disease and an unusual paralytic illness suffered by Chamorro villagers on the Pacific island of Guam. Many Chamorros with the disease suffer dementia, and in some villages one-quarter of the adults perished from the disease. Like Alzheimer''s, the causal factors of Guamanian amyotrophic lateral sclerosis/parkinsonism dementia complex (ALS/PDC) are poorly understood. In replicated experiments, we found that chronic dietary exposure to a cyanobacterial toxin present in the traditional Chamorro diet, β-N-methylamino-l-alanine (BMAA), triggers the formation of both NFT and β-amyloid deposits similar in structure and density to those found in brain tissues of Chamorros who died with ALS/PDC. Vervets (Chlorocebus sabaeus) fed for 140 days with BMAA-dosed fruit developed NFT and sparse β-amyloid deposits in the brain. Co-administration of the dietary amino acid l-serine with l-BMAA significantly reduced the density of NFT. These findings indicate that while chronic exposure to the environmental toxin BMAA can trigger neurodegeneration in vulnerable individuals, increasing the amount of l-serine in the diet can reduce the risk.     

“One Health” or Three? Publication Silos Among the One Health Disciplines

The One Health initiative is a global effort fostering interdisciplinary collaborations to address challenges in human, animal, and environmental health. While One Health has received considerable press, its benefits remain unclear because its effects have not been quantitatively described. We systematically surveyed the published literature and used social network analysis to measure interdisciplinarity in One Health studies constructing dynamic pathogen transmission models. The number of publications fulfilling our search criteria increased by 14.6% per year, which is faster than growth rates for life sciences as a whole and for most biology subdisciplines. Surveyed publications clustered into three communities: one used by ecologists, one used by veterinarians, and a third diverse-authorship community used by population biologists, mathematicians, epidemiologists, and experts in human health. Overlap between these communities increased through time in terms of author number, diversity of co-author affiliations, and diversity of citations. However, communities continue to differ in the systems studied, questions asked, and methods employed. While the infectious disease research community has made significant progress toward integrating its participating disciplines, some segregation—especially along the veterinary/ecological research interface—remains.     

The influence of locus number and information content on species delimitation: an empirical test case in an endangered Mexican salamander

Perhaps the most important recent advance in species delimitation has been the development of model‐based approaches to objectively diagnose species diversity from genetic data. Additionally, the growing accessibility of next‐generation sequence data sets provides powerful insights into genome‐wide patterns of divergence during speciation. However, applying complex models to large data sets is time‐consuming and computationally costly, requiring careful consideration of the influence of both individual and population sampling, as well as the number and informativeness of loci on species delimitation conclusions. Here, we investigated how locus number and information content affect species delimitation results for an endangered Mexican salamander species, Ambystoma ordinarium. We compared results for an eight‐locus, 137‐individual data set and an 89‐locus, seven‐individual data set. For both data sets, we used species discovery methods to define delimitation models and species validation methods to rigorously test these hypotheses. We also used integrated demographic model selection tools to choose among delimitation models, while accounting for gene flow. Our results indicate that while cryptic lineages may be delimited with relatively few loci, sampling larger numbers of loci may be required to ensure that enough informative loci are available to accurately identify and validate shallow‐scale divergences. These analyses highlight the importance of striking a balance between dense sampling of loci and individuals, particularly in shallowly diverged lineages. They also suggest the presence of a currently unrecognized, endangered species in the western part of A. ordinarium's range.     

High levels of cyclic‐di‐GMP in plant‐associated Pseudomonas correlate with evasion of plant immunity

The plant innate immune system employs plasma membrane‐localized receptors that specifically perceive pathogen/microbe‐associated molecular patterns (PAMPs/MAMPs). This induces a defence response called pattern‐triggered immunity (PTI) to fend off pathogen attack. Commensal bacteria are also exposed to potential immune recognition and must employ strategies to evade and/or suppress PTI to successfully colonize the plant. During plant infection, the flagellum has an ambiguous role, acting as both a virulence factor and also as a potent immunogen as a result of the recognition of its main building block, flagellin, by the plant pattern recognition receptors (PRRs), including FLAGELLIN SENSING2 (FLS2). Therefore, strict control of flagella synthesis is especially important for plant‐associated bacteria. Here, we show that cyclic‐di‐GMP [bis‐(3′‐5′)‐cyclic di‐guanosine monophosphate], a central regulator of bacterial lifestyle, is involved in the evasion of PTI. Elevated cyclic‐di‐GMP levels in the pathogen Pseudomonas syringae pv. tomato (Pto) DC3000, the opportunist P. aeruginosa PAO1 and the commensal P. protegens Pf‐5 inhibit flagellin synthesis and help the bacteria to evade FLS2‐mediated signalling in Nicotiana benthamiana and Arabidopsis thaliana. Despite this, high cellular cyclic‐di‐GMP concentrations were shown to drastically reduce the virulence of Pto DC3000 during plant infection. We propose that this is a result of reduced flagellar motility and/or additional pleiotropic effects of cyclic‐di‐GMP signalling on bacterial behaviour.     

Uncovering species boundaries in the Neotropical ant complex Ectatomma ruidum (Ectatomminae) under the presence of nuclear mitochondrial paralogues

Nuclear mitochondrial (mt) paralogues (numts) are non‐functional fragments of mtDNA integrated into the nuclear genome that can overestimate the number of species in analyses based on mtDNA sequences. As numts have relatively slow mutation rates, they can pass undetected by conventional procedures such as inspecting for internal stop codons, indels or apparent polymorphism in chromatograms. Species boundaries based on mtDNA markers therefore require a thorough assessment of numts, especially in insects, where this phenomenon appears to be relatively frequent. Ectatomma ruidum is a widely distributed Neotropical ant species that is distributed from northern Mexico to northern Brazil. Previous behavioural and molecular evidence suggests that this species actually represents a composite taxon. Here we assessed the species boundaries in E. ruidum based on two mt (COI, cyt b) and one nuclear (H3) marker, as well as on external morphology. Ancient and recent mt paralogues were detected in several specimens, although pre‐PCR dilution of DNA template helped to recover most of the mt orthologues. Based on the congruence found between our species delineation obtained from the mt genealogies and the discriminated morphospecies, we propose that E. ruidum is actually composed of at least three species. Two of these species have a wide geographical distribution in the Neotropics, whereas the remaining one was restricted to localities situated near the Pacific coast in south‐east Mexico. We also found extensive intra‐ and interspecific variation in the barcoding locus. Moreover, the nuclear evidence suggests the existence of hybrids between two of these species in Oaxaca, south‐east Mexico. This study agrees with previous studies of other closely related animal taxa, which have revealed a complex evolutionary history and overlooked species diversity in the latter region.     

Pharmacological removal of serum amyloid P component from intracerebral plaques and cerebrovascular Aβ amyloid deposits in vivo

Human amyloid deposits always contain the normal plasma protein serum amyloid P component (SAP), owing to its avid but reversible binding to all amyloid fibrils, including the amyloid β (Aβ) fibrils in the cerebral parenchyma plaques and cerebrovascular amyloid deposits of Alzheimer''s disease (AD) and cerebral amyloid angiopathy (CAA). SAP promotes amyloid fibril formation in vitro, contributes to persistence of amyloid in vivo and is also itself directly toxic to cerebral neurons. We therefore developed (R)-1-[6-[(R)-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl]pyrrolidine-2-carboxylic acid (CPHPC), a drug that removes SAP from the blood, and thereby also from the cerebrospinal fluid (CSF), in patients with AD. Here we report that, after introduction of transgenic human SAP expression in the TASTPM double transgenic mouse model of AD, all the amyloid deposits contained human SAP. Depletion of circulating human SAP by CPHPC administration in these mice removed all detectable human SAP from both the intracerebral and cerebrovascular amyloid. The demonstration that removal of SAP from the blood and CSF also removes it from these amyloid deposits crucially validates the strategy of the forthcoming ‘Depletion of serum amyloid P component in Alzheimer''s disease (DESPIAD)’ clinical trial of CPHPC. The results also strongly support clinical testing of CPHPC in patients with CAA.