Lack of hypoxia-inducible factor-1 alpha impairs midbrain neural precursor cells involving vascular endothelial growth factor signaling

Oxygen tension is critical for proliferation of human and murine midbrain-derived neural precursor cells (mNPCs). Here, we conditionally inactivated the hypoxia-responsive transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha) in murine NPCs to determine its role in proliferation, survival, and dopaminergic differentiation in vitro as well as survival of murine dopaminergic neurons in vivo. HIF-1alpha conditional knock-out (HIF-1alpha CKO) mNPCs showed midbrain-specific impairment of survival and proliferation. Dopaminergic differentiation of HIF-1alpha CKO mNPCs in vitro was markedly reduced. Expression of vascular endothelial growth factor (VEGF) mRNA was reduced in HIF-1alpha CKO mNPCs, whereas erythropoietin signaling was not affected. Treatment of HIF-1alpha CKO mNPCs with 50 ng/ml VEGF partially recovered proliferation and dopaminergic differentiation in vitro. In substantia nigra (SN) of adult HIF-1alpha CKO mice, protein levels of dopaminergic marker molecules such as tyrosine hydroxylase (TH) and aldehyde dehydrogenase were reduced by 41 and 61%, respectively. The cell survival marker Bcl-2 was reduced by 58% while caspase-3 was activated. Nonbiased stereological cell counts of TH-positive neurons in SN of young adult HIF-1alpha CKO mice revealed a reduction of 31% compared with cre/wt mice (in which the wild-type Hif1a allele is expressed in parallel with the Cre recombinase allele). However, we found no impairment of striatal dopamine concentrations or locomotor behavior. In conclusion, HIF-1alpha seems to be a transcription factor relevant to the development and survival of substantia nigra dopaminergic neurons involving VEGF signaling

NEOTROPICAL CARNIVORES: a data set on carnivore distribution in the Neotropics

Incluye contenido parcial de los autoresAbstract.Mammalian carnivores are considered a key group in maintaining ecologicalhealth and can indicate potential ecological integrity in landscapes where they occur. Carni-vores also hold high conservation value and their habitat requirements can guide managementand conservation plans. The order Carnivora has 84 species from 8 families in the Neotropicalregion: Canidae; Felidae; Mephitidae; Mustelidae; Otariidae; Phocidae; Procyonidae; andUrsidae. Herein, we include published and unpublished data on native terrestrial Neotropicalcarnivores (Canidae; Felidae; Mephitidae; Mustelidae; Procyonidae; and Ursidae). NEOTRO-PICAL CARNIVORES is a publicly available data set that includes 99,605 data entries from35,511 unique georeferenced coordinates. Detection/non-detection and quantitative data wereobtained from 1818 to 2018 by researchers, governmental agencies, non-governmental organi-zations, and private consultants. Data were collected using several methods including cameratrapping, museum collections, roadkill, line transect, and opportunistic records. Literature(peer-reviewed and grey literature) from Portuguese, Spanish and English were incorporated inthis compilation. Most of the data set consists of detection data entries (n=79,343; 79.7%) butalso includes non-detection data (n=20,262; 20.3%). Of those, 43.3% also include count data(n=43,151). The information available in NEOTROPICAL CARNIVORES will contribute tomacroecological, ecological, and conservation questions in multiple spatio-temporal perspec-tives. As carnivores play key roles in trophic interactions, a better understanding of their distri-bution and habitat requirements are essential to establish conservation management plans andsafeguard the future ecological health of Neotropical ecosystems. Our data paper, combinedwith other large-scale data sets, has great potential to clarify species distribution and relatedecological processes within the Neotropics. There are no copyright restrictions and no restric-tion for using data from this data paper, as long as the data paper is cited as the source of theinformation used. We also request that users inform us of how they intend to use the data

A large scale hearing loss screen reveals an extensive unexplored genetic landscape for auditory dysfunction

The developmental and physiological complexity of the auditory system is likely reflected in the underlying set of genes involved in auditory function. In humans, over 150 non-syndromic loci have been identified, and there are more than 400 human genetic syndromes with a hearing loss component. Over 100 non-syndromic hearing loss genes have been identified in mouse and human, but we remain ignorant of the full extent of the genetic landscape involved in auditory dysfunction. As part of the International Mouse Phenotyping Consortium, we undertook a hearing loss screen in a cohort of 3006 mouse knockout strains. In total, we identify 67 candidate hearing loss genes. We detect known hearing loss genes, but the vast majority, 52, of the candidate genes were novel. Our analysis reveals a large and unexplored genetic landscape involved with auditory function

Engineering Corynebacterium glutamicum for isobutanol production

The production of isobutanol in microorganisms has recently been achieved by harnessing the highly active 2-keto acid pathways. Since these 2-keto acids are precursors of amino acids, we aimed to construct an isobutanol production platform in Corynebacterium glutamicum, a well-known amino-acid-producing microorganism. Analysis of this host’s sensitivity to isobutanol toxicity revealed that C. glutamicum shows an increased tolerance to isobutanol relative to Escherichia coli. Overexpression of alsS of Bacillus subtilis, ilvC and ilvD of C. glutamicum, kivd of Lactococcus lactis, and a native alcohol dehydrogenase, adhA, led to the production of 2.6 g/L isobutanol and 0.4 g/L 3-methyl-1-butanol in 48 h. In addition, other higher chain alcohols such as 1-propanol, 2-methyl-1-butanol, 1-butanol, and 2-phenylethanol were also detected as byproducts. Using longer-term batch cultures, isobutanol titers reached 4.0 g/L after 96 h with wild-type C. glutamicum as a host. Upon the inactivation of several genes to direct more carbon through the isobutanol pathway, we increased production by ∼25% to 4.9 g/L isobutanol in a ∆pyc∆ldh background. These results show promise in engineering C. glutamicum for higher chain alcohol production using the 2-keto acid pathways

Comparison of Pharmacological Modulation of APP Metabolism in Primary Chicken Telencephalic Neurons and in a Human Neuroglioma Cell Line

Sequential cleavage of amyloid precursor protein (APP) by β- and γ-secretases and the formation of Aβ peptides are pivotal for Alzheimer's disease. Therefore, a large number of drugs has been developed targeting APP metabolism. However, many pharmacological compounds have been identified in vitro in immortalized APP overexpressing cell lines rather than in primary neurons. Here, we compared the effect of already characterized secretase inhibitors and modulators on Aβ formation in primary chicken telencephalic neurons and in a human neuroglioma cell line (H4) ectopically expressing human APP with the Swedish double mutation. Primary chicken neurons replicated the effects of a β-secretase inhibitor (β-secretase inhibitor IV), two γ-secretase inhibitors (DAPM, DAPT), two non-steroidal-anti-inflammatory drugs (sulindac sulfide, CW), and of the calpain inhibitor calpeptin. With the exception of the two γ-secretase inhibitors, all tested compounds were more efficacious in primary chicken telencephalic neurons than in the immortalized H4 cell line. Moreover, H4 cells failed to reproduce the effect of calpeptin. Hence, primary chicken telencephalic neurons represent a suitable cell culture model for testing drugs interfering with APP processing and are overall more sensitive to pharmacological interference than immortalized H4 cells ectopically expressing mutant human APP

Barnase as a New Therapeutic Agent Triggering Apoptosis in Human Cancer Cells

RNases are currently studied as non-mutagenic alternatives to the harmful DNA-damaging anticancer drugs commonly used in clinical practice. Many mammalian RNases are not potent toxins due to the strong inhibition by ribonuclease inhibitor (RI) presented in the cytoplasm of mammalian cells.In search of new effective anticancer RNases we studied the effects of barnase, a ribonuclease from Bacillus amyloliquefaciens, on human cancer cells. We found that barnase is resistant to RI. In MTT cell viability assay, barnase was cytotoxic to human carcinoma cell lines with half-inhibitory concentrations (IC(50)) ranging from 0.2 to 13 microM and to leukemia cell lines with IC(50) values ranging from 2.4 to 82 microM. Also, we characterized the cytotoxic effects of barnase-based immunoRNase scFv 4D5-dibarnase, which consists of two barnase molecules serially fused to the single-chain variable fragment (scFv) of humanized antibody 4D5 that recognizes the extracellular domain of cancer marker HER2. The scFv 4D5-dibarnase specifically bound to HER2-positive cells and was internalized via receptor-mediated endocytosis. The intracellular localization of internalized scFv 4D5-dibarnase was determined by electronic microscopy. The cytotoxic effect of scFv 4D5-dibarnase on HER2-positive human ovarian carcinoma SKOV-3 cells (IC(50) = 1.8 nM) was three orders of magnitude greater than that of barnase alone. Both barnase and scFv 4D5-dibarnase induced apoptosis in SKOV-3 cells accompanied by internucleosomal chromatin fragmentation, membrane blebbing, the appearance of phosphatidylserine on the outer leaflet of the plasma membrane, and the activation of caspase-3.These results demonstrate that barnase is a potent toxic agent for targeting to cancer cells

Rho GTPases as therapeutic targets in Alzheimer’s disease

The progress we have made in understanding Alzheimer’s disease (AD) pathogenesis has led to the identification of several novel pathways and potential therapeutic targets. Rho GTPases have been implicated as critical components in AD pathogenesis, but their various functions and interactions make understanding their complex signaling challenging to study. Recent advancements in both the field of AD and Rho GTPase drug development provide novel tools for the elucidation of Rho GTPases as a viable target for AD. Herein, we summarize the fluctuating activity of Rho GTPases in various stages of AD pathogenesis and in several in vitro and in vivo AD models. We also review the current pharmacological tools such as NSAIDs, RhoA/ROCK, Rac1, and Cdc42 inhibitors used to target Rho GTPases and their use in AD-related studies. Finally, we summarize the behavioral modifications following Rho GTPase modulation in several AD mouse models. As key regulators of several AD-related signals, Rho GTPases have been studied as targets in AD. However, a consensus has yet to be reached regarding the stage at which targeting Rho GTPases would be the most beneficial. The studies discussed herein emphasize the critical role of Rho GTPases and the benefits of their modulation in AD