New Mouse Lines for the Analysis of Neuronal Morphology Using CreER(T)/loxP-Directed Sparse Labeling

BACKGROUND: Pharmacologic control of Cre-mediated recombination using tamoxifen-dependent activation of a Cre-estrogen receptor ligand binding domain fusion protein [CreER(T)] is widely used to modify and/or visualize cells in the mouse. METHODS AND FINDINGS: We describe here two new mouse lines, constructed by gene targeting to the Rosa26 locus to facilitate Cre-mediated cell modification. These lines should prove particularly useful in the context of sparse labeling experiments. The R26rtTACreER line provides ubiquitous expression of CreER under transcriptional control by the tetracycline reverse transactivator (rtTA); dual control by doxycycline and tamoxifen provides an extended dynamic range of Cre-mediated recombination activity. The R26IAP line provides high efficiency Cre-mediated activation of human placental alkaline phosphatase (hPLAP), complementing the widely used, but low efficiency, Z/AP line. By crossing with mouse lines that direct cell-type specific CreER expression, the R26IAP line has been used to produce atlases of labeled cholinergic and catecholaminergic neurons in the mouse brain. The R26IAP line has also been used to visualize the full morphologies of retinal dopaminergic amacrine cells, among the largest neurons in the mammalian retina. CONCLUSIONS: The two new mouse lines described here expand the repertoire of genetically engineered mice available for controlled in vivo recombination and cell labeling using the Cre-lox system

Pets as Sentinels of Human Exposure to Neurotoxic Metals

The idea that animals may be used as sentinels of environmental hazards pending over humans and the associated public health implications is not a new one. Nowadays pets are being used as bioindicators for the effects of environmental contaminants in human populations. This is of paramount importance due to the large increase in the worldwide distribution of synthetic chemicals, particularly in the built environment. Companion animals share the habitat with humans being simultaneously exposed to and suffering the same disease spectrum as their masters. Moreover, their shorter latency periods (due to briefer lifespans) enable them to act as early warning systems, allowing timely public health interventions. The rise on ethical constraints on the use of animals and, consequently, on the sampling they can be subjected to has led to the preferential use of noninvasive matrices, and in this case we are looking into hair. This chapter focuses in three non-essential metals: mercury, lead, and cadmium, due to their ubiquitous presence in the built environment and their ability of affecting the mammal nervous system. There is a fairly short amount of studies reporting the concentrations of these metals in pets’ hair, particularly for cats. These studies are characterized, and the metal concentrations corresponding to different parameters (e.g., age, sex, diet, rearing) are described in order to provide the reader with a general vision on the use of this noninvasive matrix on the studies conducted since the last two decades of the twentieth century.publishe

The Type III Effectors NleE and NleB from Enteropathogenic E. coli and OspZ from Shigella Block Nuclear Translocation of NF-κB p65

Many bacterial pathogens utilize a type III secretion system to deliver multiple effector proteins into host cells. Here we found that the type III effectors, NleE from enteropathogenic E. coli (EPEC) and OspZ from Shigella, blocked translocation of the p65 subunit of the transcription factor, NF-κB, to the host cell nucleus. NF-κB inhibition by NleE was associated with decreased IL-8 expression in EPEC-infected intestinal epithelial cells. Ectopically expressed NleE also blocked nuclear translocation of p65 and c-Rel, but not p50 or STAT1/2. NleE homologues from other attaching and effacing pathogens as well OspZ from Shigella flexneri 6 and Shigella boydii, also inhibited NF-κB activation and p65 nuclear import; however, a truncated form of OspZ from S. flexneri 2a that carries a 36 amino acid deletion at the C-terminus had no inhibitory activity. We determined that the C-termini of NleE and full length OspZ were functionally interchangeable and identified a six amino acid motif, IDSY(M/I)K, that was important for both NleE- and OspZ-mediated inhibition of NF-κB activity. We also established that NleB, encoded directly upstream from NleE, suppressed NF-κB activation. Whereas NleE inhibited both TNFα and IL-1β stimulated p65 nuclear translocation and IκB degradation, NleB inhibited the TNFα pathway only. Neither NleE nor NleB inhibited AP-1 activation, suggesting that the modulatory activity of the effectors was specific for NF-κB signaling. Overall our data show that EPEC and Shigella have evolved similar T3SS-dependent means to manipulate host inflammatory pathways by interfering with the activation of selected host transcriptional regulators

Primary Postnatal Dorsal Root Ganglion Culture from Conventionally Slaughtered Calves

Neurological disorders in ruminants have an important impact on veterinary health, but very few host-specific in vitro models have been established to study diseases affecting the nervous system. Here we describe a primary neuronal dorsal root ganglia (DRG) culture derived from calves after being conventionally slaughtered for food consumption. The study focuses on the in vitro characterization of bovine DRG cell populations by immunofluorescence analysis. The effects of various growth factors on neuron viability, neurite outgrowth and arborisation were evaluated by morphological analysis. Bovine DRG neurons are able to survive for more than 4 weeks in culture. GF supplementation is not required for neuronal survival and neurite outgrowth. However, exogenously added growth factors promote neurite outgrowth. DRG cultures from regularly slaughtered calves represent a promising and sustainable host specific model for the investigation of pain and neurological diseases in bovines

Transcriptomic and metabolomic shifts in rice roots in response to Cr (VI) stress

<p>Abstract</p> <p>Background</p> <p>Widespread use of chromium (Cr) contaminated fields due to careless and inappropriate management practices of effluent discharge, mostly from industries related to metallurgy, electroplating, production of paints and pigments, tanning, and wood preservation elevates its concentration in surface soil and eventually into rice plants and grains. In spite of many previous studies having been conducted on the effects of chromium stress, the precise molecular mechanisms related to both the effects of chromium phytotoxicity, the defense reactions of plants against chromium exposure as well as translocation and accumulation in rice remain poorly understood.</p> <p>Results</p> <p>Detailed analysis of genome-wide transcriptome profiling in rice root is reported here, following Cr-plant interaction. Such studies are important for the identification of genes responsible for tolerance, accumulation and defense response in plants with respect to Cr stress. Rice root metabolome analysis was also carried out to relate differential transcriptome data to biological processes affected by Cr (VI) stress in rice. To check whether the Cr-specific motifs were indeed significantly over represented in the promoter regions of Cr-responsive genes, occurrence of these motifs in whole genome sequence was carried out. In the background of whole genome, the lift value for these 14 and 13 motifs was significantly high in the test dataset. Though no functional role has been assigned to any of the motifs, but all of these are present as promoter motifs in the Database of orthologus promoters.</p> <p>Conclusion</p> <p>These findings clearly suggest that a complex network of regulatory pathways modulates Cr-response of rice. The integrated matrix of both transcriptome and metabolome data after suitable normalization and initial calculations provided us a visual picture of the correlations between components. Predominance of different motifs in the subsets of genes suggests the involvement of motif-specific transcription modulating proteins in Cr stress response of rice.</p

Differential Gene Expression and Adherence of Escherichia coli O157:H7 In Vitro and in Ligated Pig Intestines

BACKGROUND: Escherichia coli O157:H7 strain 86-24 grown in MacConkey broth (MB) shows almost no adherence to cultured epithelial cells but adheres well in pig ligated intestines. This study investigated the mechanisms associated with the difference between in-vitro and in-vivo adherence of the MB culture. METHODOLOGY/PRINCIPAL FINDINGS: It was found that decreased adherence in vitro by bacteria grown in MB was mainly due to lactose, possibly implicating the involvement of carbon catabolite repression (CCR). Expression of selected virulence-related genes associated with adherence and CCR was then examined by quantitative PCR. When bacteria were grown in MB and Brain Heart Infusion with NaHCO(3) (BHIN) plus lactose, pH was reduced to 5.5-5.9 and there was a significant decrease in expression of the locus of enterocyte effacement (LEE) genes eae, tir, espD, grlA/R and ler, and an increase in cya (cAMP), and two negative regulators of the LEE, gadE and hfq. Putative virulence genes stcE, hlyA, ent and nleA were also decreased in vitro. Reversal of these changes was noted for bacteria recovered from the intestine, where transcripts for qseF and fis and putative virulence factors AidA(15), TerC and Ent/EspL2 were significantly increased, and transcripts for AIDA(48), Iha, UreC, Efa1A, Efa1B, ToxB, EhxA, StcE, NleA and NleB were expressed at high levels. CONCLUSIONS/SIGNIFICANCE: Presence of lactose resulted in decreased expression of LEE genes and the failure of EHEC O157:H7 to adhere to epithelial cells in vitro but this repression was overcome in vivo. CCR and/or acidic pH may have played a role in repression of the LEE genes. Bacterial pathogens need to integrate their nutritional metabolism with expression of virulence genes but little is known of how this is done in E. coli O157:H7. This study indicates one aspect of the subject that should be investigated further

Defective germline reprogramming rewires the spermatogonial transcriptome.

Defective germline reprogramming in Piwil4 (Miwi2)- and Dnmt3l-deficient mice results in the failure to reestablish transposon silencing, meiotic arrest and progressive loss of spermatogonia. Here we sought to understand the molecular basis for this spermatogonial dysfunction. Through a combination of imaging, conditional genetics and transcriptome analysis, we demonstrate that germ cell elimination in the respective mutants arises as a result of defective de novo genome methylation during reprogramming rather than because of a function for the respective factors within spermatogonia. In both Miwi2-/- and Dnmt3l-/- spermatogonia, the intracisternal-A particle (IAP) family of endogenous retroviruses is derepressed, but, in contrast to meiotic cells, DNA damage is not observed. Instead, we find that unmethylated IAP promoters rewire the spermatogonial transcriptome by driving expression of neighboring genes. Finally, spermatogonial numbers, proliferation and differentiation are altered in Miwi2-/- and Dnmt3l-/- mice. In summary, defective reprogramming deregulates the spermatogonial transcriptome and may underlie spermatogonial dysfunction

Combination of searches for heavy spin-1 resonances using 139 fb−1 of proton-proton collision data at s = 13 TeV with the ATLAS detector

A combination of searches for new heavy spin-1 resonances decaying into different pairings of W, Z, or Higgs bosons, as well as directly into leptons or quarks, is presented. The data sample used corresponds to 139 fb−1 of proton-proton collisions at = 13 TeV collected during 2015–2018 with the ATLAS detector at the CERN Large Hadron Collider. Analyses selecting quark pairs (qq, bb, , and tb) or third-generation leptons (τν and ττ) are included in this kind of combination for the first time. A simplified model predicting a spin-1 heavy vector-boson triplet is used. Cross-section limits are set at the 95% confidence level and are compared with predictions for the benchmark model. These limits are also expressed in terms of constraints on couplings of the heavy vector-boson triplet to quarks, leptons, and the Higgs boson. The complementarity of the various analyses increases the sensitivity to new physics, and the resulting constraints are stronger than those from any individual analysis considered. The data exclude a heavy vector-boson triplet with mass below 5.8 TeV in a weakly coupled scenario, below 4.4 TeV in a strongly coupled scenario, and up to 1.5 TeV in the case of production via vector-boson fusion

Measurement of the energy asymmetry in t(t)over-barj production at 13 TeV with the ATLAS experiment and interpretation in the SMEFT framework

A measurement of the energy asymmetry in jet-associated top-quark pair production is presented using $$139\,{\mathrm {fb}}^{-1}$$ 139 fb - 1 of data collected by the ATLAS detector at the Large Hadron Collider during pp collisions at $$\sqrt{s}=13\,\text {TeV}$$ s = 13 TeV . The observable measures the different probability of top and antitop quarks to have the higher energy as a function of the jet scattering angle with respect to the beam axis. The energy asymmetry is measured in the semileptonic $$t{\bar{t}}$$ t t ¯ decay channel, and the hadronically decaying top quark must have transverse momentum above $$350\,\text {GeV}$$ 350 GeV . The results are corrected for detector effects to particle level in three bins of the scattering angle of the associated jet. The measurement agrees with the SM prediction at next-to-leading-order accuracy in quantum chromodynamics in all three bins. In the bin with the largest expected asymmetry, where the jet is emitted perpendicular to the beam, the energy asymmetry is measured to be $$-0.043\pm 0.020$$ - 0.043 ± 0.020 , in agreement with the SM prediction of $$-0.037\pm 0.003$$ - 0.037 ± 0.003 . Interpreting this result in the framework of the Standard Model effective field theory (SMEFT), it is shown that the energy asymmetry is sensitive to the top-quark chirality in four-quark operators and is therefore a valuable new observable in global SMEFT fits