Investigating the mechanisms of the heat shock response impairment in Huntington’s disease mouse models

Huntington’s disease (HD) is a neurodegenerative disease caused by the expansion of a CAG repeat within exon 1 of the huntingtin (HTT) gene. This produces a mutant HTT protein with an abnormally long polyglutamine stretch, which is prone to misfold and form aggregates, as detected in HD patients and mouse models, and which have aberrant interactions with members of key cellular pathways that become disrupted. The heat shock response is a cytoprotective mechanism that aims to prevent and reduce aggregation and maintain proteome integrity. It is regulated by heat shock factor 1 (HSF1) which, under stress conditions, activates the transcription of the heat shock genes. In this thesis, a comprehensive analysis of the heat shock response has been described for HD mouse models R6/2 and zQ175, after in vivo pharmacological induction with the compound NVP-HSP990. The expression of Hsf1 and nine heat shock genes was measured from mouse tissue lysates by a QuantiGene multiplex assay, developed and validated for this purpose. Evidence of an impairment of the heat shock response was found in brain hemispheres, tibialis anterior and striatum of both models, with less consistent evidence in the cortex. As a potential cause of this dysregulation, HSF1 levels were analysed in brain and muscle, however, a decrease in HSF1 was not detected in either tissue. SIRT1 can deacetylate HSF1 and maintain it in a competent state, bound to heat shock gene promoters. SIRT1 levels decline with disease progression in HD mouse models, therefore, whether a decreased SIRT1 activity could be responsible for the impaired heat shock response was investigated. The analysis of the heat shock gene expression with QuantiGene revealed that Sirt1 overexpression on R6/2 mice did not significantly improve the heat shock response impairment in brain or tibialis anterior

Statistics of Core Lifetimes in Numerical Simulations of Turbulent, Magnetically Supercritical Molecular Clouds

We present measurements of the mean dense core lifetimes in numerical simulations of magnetically supercritical, turbulent, isothermal molecular clouds, in order to compare with observational determinations. "Prestellar" lifetimes (given as a function of the mean density within the cores, which in turn is determined by the density threshold n_thr used to define them) are consistent with observationally reported values, ranging from a few to several free-fall times. We also present estimates of the fraction of cores in the "prestellar", "stellar'', and "failed" (those cores that redisperse back into the environment) stages as a function of n_thr. The number ratios are measured indirectly in the simulations due to their resolution limitations. Our approach contains one free parameter, the lifetime of a protostellar object t_yso (Class 0 + Class I stages), which is outside the realm of the simulations. Assuming a value t_yso = 0.46 Myr, we obtain number ratios of starless to stellar cores ranging from 4-5 at n_thr = 1.5 x 10^4 cm^-3 to 1 at n_thr = 1.2 x 10^5 cm^-3, again in good agreement with observational determinations. We also find that the mass in the failed cores is comparable to that in stellar cores at n_thr = 1.5 x 10^4 cm^-3, but becomes negligible at n_thr = 1.2 x 10^5 cm^-3, in agreement with recent observational suggestions that at the latter densities the cores are in general gravitationally dominated. We conclude by noting that the timescale for core contraction and collapse is virtually the same in the subcritical, ambipolar diffusion-mediated model of star formation, in the model of star formation in turbulent supercritical clouds, and in a model intermediate between the previous two, for currently accepted values of the clouds' magnetic criticality.Comment: 25 pages, 8 figures, ApJ accepted. Fig.1 animation is at http://www.astrosmo.unam.mx/~e.vazquez/turbulence/movies/Galvan_etal07/Galvan_etal07.htm

Formation and Collapse of Quiescent Cloud Cores Induced by Dynamic Compressions

(Abridged) We present numerical hydrodynamical simulations of the formation, evolution and gravitational collapse of isothermal molecular cloud cores. A compressive wave is set up in a constant sub-Jeans density distribution of radius r = 1 pc. As the wave travels through the simulation grid, a shock-bounded spherical shell is formed. The inner shock of this shell reaches and bounces off the center, leaving behind a central core with an initially almost uniform density distribution, surrounded by an envelope consisting of the material in the shock-bounded shell, with a power-law density profile that at late times approaches a logarithmic slope of -2 even in non-collapsing cases. The resulting density structure resembles a quiescent core of radius < 0.1 pc, with a Bonnor-Ebert-like (BE-like) profile, although it has significant dynamical differences: it is initially non-self-gravitating and confined by the ram pressure of the infalling material, and consequently, growing continuously in mass and size. With the appropriate parameters, the core mass eventually reaches an effective Jeans mass, at which time the core begins to collapse. Thus, there is necessarily a time delay between the appearance of the core and the onset of its collapse, but this is not due to the dissipation of its internal turbulence as it is often believed. These results suggest that pre-stellar cores may approximate Bonnor-Ebert structures which are however of variable mass and may or may not experience gravitational collapse, in qualitative agreement with the large observed frequency of cores with BE-like profiles.Comment: Accepted for publication in ApJ. Associated mpeg files can be found in http://www.astrosmo.unam.mx/~g.gomez/publica.htm

Extensive Expression Analysis of Htt Transcripts in Brain Regions from the zQ175 HD Mouse Model Using a QuantiGene Multiplex Assay

Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by a CAG repeat expansion within exon 1 of the huntingtin (HTT) gene. HTT mRNA contains 67 exons and does not always splice between exon 1 and exon 2 leading to the production of a small polyadenylated HTTexon1 transcript, and the full-length HTT mRNA has three 3′UTR isoforms. We have developed a QuantiGene multiplex panel for the simultaneous detection of all of these mouse Htt transcripts directly from tissue lysates and demonstrate that this can replace the more work-intensive Taqman qPCR assays. We have applied this to the analysis of brain regions from the zQ175 HD mouse model and wild type littermates at two months of age. We show that the incomplete splicing of Htt occurs throughout the brain and confirm that this originates from the mutant and not endogenous Htt allele. Given that HTTexon1 encodes the highly pathogenic exon 1 HTT protein, it is essential that the levels of all Htt transcripts can be monitored when evaluating HTT lowering approaches. Our QuantiGene panel will allow the rapid comparative assessment of all Htt transcripts in cell lysates and mouse tissues without the need to first extract RNA

The heat shock response, determined by QuantiGene multiplex, is impaired in HD mouse models and not caused by HSF1 reduction.

Huntington's disease (HD) is a devastating neurodegenerative disorder, caused by a CAG/polyglutamine repeat expansion, that results in the aggregation of the huntingtin protein, culminating in the deposition of inclusion bodies in HD patient brains. We have previously shown that the heat shock response becomes impaired with disease progression in mouse models of HD. The disruption of this inducible arm of the proteostasis network is likely to exacerbate the pathogenesis of this protein-folding disease. To allow a rapid and more comprehensive analysis of the heat shock response, we have developed, and validated, a 16-plex QuantiGene assay that allows the expression of Hsf1 and nine heat shock genes, to be measured directly, and simultaneously, from mouse tissue. We used this QuantiGene assay to show that, following pharmacological activation in vivo, the heat shock response impairment in tibialis anterior, brain hemispheres and striatum was comparable between zQ175 and R6/2 mice. In contrast, although a heat shock impairment could be detected in R6/2 cortex, this was not apparent in the cortex from zQ175 mice. Whilst the mechanism underlying this impairment remains unknown, our data indicated that it is not caused by a reduction in HSF1 levels, as had been reported

Aedes albopictus diversity and relationships in south-western Europe and Brazil by rDNA/mtDNA and phenotypic analyses: ITS-2, a useful marker for spread studies

Background: Aedes albopictus is a very invasive mosquito, which has recently colonized tropical and temperate regions worldwide. Of concern is its role in the spread of emerging or re-emerging mosquito-borne diseases. Ae. albopictus from south-western Europe and Brazil were studied to infer genetic and phenetic diversity at intra-individual, intra-population and inter-population levels, and to analyse its spread. Methods: Genotyping was made by rDNA 5.8S-ITS-2 and mtDNA cox1 sequencing to assess haplotype and nucleotide diversity, genetic distances and phylogenetic networks. Male and female phenotyping included combined landmark-and outlined-based geometric morphometrics of wing size and shape. Results: Specimens from seven populations from Spain, France and Brazil provided 12 cox1 and 162 5.8S-ITS-2 haplotypes, with great genetic variability difference between both markers (0.9% vs 31.2%). Five cox1 haplotypes were shared with other countries, mainly Italy, USA and China, but none was shared between Europe and Brazil. The 5.8S-ITS-2 showed 2–7 intra-individual (mean 4.7) and 16–34 intra-/inter-population haplotypes (24.7), including haplotypes shared between Spain, France and Brazil. A 4.3% of ITS-2 haplotypes were shared, mainly with Italy, USA and Thailand, evidencing worldwide spread and introductions from areas where recent outbreaks of Ae. albopictus-transmitted pathogens occurred. Wing size showed sex differences. Wing shape distinguished between Brazilian and European specimens. Both genetic and morphometric markers showed differences between insular Spain and continental Spain, France and Brazil. Conclusions: ITS-2 proves to be a useful marker to assess Ae. albopictus spread, providing pronouncedly more information than cox1, including intra-individual, intra-population and inter-population levels, furnishing a complete overview of the evolutionary exchanges followed by this mosquito. Wing morphometry proves to be a useful phenotyping marker, allowing to distinguish different populations at the level of both male and female specimens. Results indicate the need for periodic surveillance monitorings to verify that no Ae. albopictus with high virus transmission capacity is introduced into Europe. Graphic Abstract: [Figure not available: see fulltext.] © 2021, The Author(s)

Dynamic star formation in the massive DR21 filament

The formation of massive stars is a highly complex process in which it is not clear whether the star-forming gas is in global gravitational collapse or in an equilibrium state, supported by turbulence. By studying one of the most massive and dense star-forming regions in the Galaxy at a distance of less than 3 kpc, the filament containing the well-known sources DR21 and DR21(OH), we expect to find observational signatures that allow to discriminate between the two views. We use molecular line data from our 13CO 1-0, CS 2-1, and N2H+ 1-0 survey of the Cygnus X region obtained with the FCRAO and high-angular resolution observations of CO, CS, HCO+, N2H+, and H2CO, obtained with the IRAM 30m telescope. We observe a complex velocity field and velocity dispersion in the DR21 filament in which regions of highest column-density, i.e. dense cores, have a lower velocity dispersion than the surrounding gas and velocity gradients that are not (only) due to rotation. Infall signatures in optically thick line profiles of HCO+ and 12CO are observed along and across the whole DR21 filament. From modelling the observed spectra, we obtain a typical infall speed of 0.6 km/s and mass accretion rates of the order of a few 10^-3 Msun/yr for the two main clumps constituting the filament. These massive (4900 and 3300 Msun) clumps are both gravitationally contracting. All observed kinematic features in the DR21 filament can be explained if it is formed by the convergence of flows at large scales and is now in a state of global gravitational collapse. Whether this convergence of flows originated from self-gravity at larger scales or from other processes can not be settled with the present study. The observed velocity field and velocity dispersion are consistent with results from (magneto)-hydrodynamic simulations where the cores lie at the stagnation points of convergent turbulent flows.Comment: Astronomy and Astrophysics, in pres

Diffusion in supersonic, turbulent, compressible flows

We investigate diffusion in supersonic, turbulent, compressible flows. Supersonic turbulence can be characterized as network of interacting shocks. We consider flows with different rms Mach numbers and where energy necessary to maintain dynamical equilibrium is inserted at different spatial scales. We find that turbulent transport exhibits super-diffusive behavior due to induced bulk motions. In a comoving reference frame, however, diffusion behaves normal and can be described by mixing length theory extended into the supersonic regime.Comment: 11 pages, incl. 5 figures, accepted for publication in Physical Review E (a high-resolution version is available at http://www.aip.de./~ralf/Publications/p21.abstract.html

Holographic flavor on the Higgs branch

In this paper we study the holographic dual, in several spacetime dimensions, of the Higgs branch of gauge theories with fundamental matter. These theories contain defects of various codimensionalities, where the matter fields are located. In the holographic description the matter is added by considering flavor brane probes in the supergravity backgrounds generated by color branes, while the Higgs branch is obtained when the color and flavor branes recombine with each other. We show that, generically, the holographic dual of the Higgs phase is realized by means of the addition of extra flux on the flavor branes and by choosing their appropriate embedding in the background geometry. This suggests a dielectric interpretation in terms of the color branes, whose vacuum solutions precisely match the F- and D-flatness conditions obtained on the field theory side. We further compute the meson mass spectra in several cases and show that when the defect added has codimension greater than zero it becomes continuous and gapless.Comment: 59 pages, 1 figure;v2: references adde

Activity of the DNA minor groove cross-linking agent SG2000 (SJG-136) against canine tumours

BACKGROUND: Cancer is the leading cause of death in older dogs and its prevalence is increasing. There is clearly a need to develop more effective anti-cancer drugs in dogs. SG2000 (SJG-136) is a sequence selective DNA minor groove cross-linking agent. Based on its in vitro potency, the spectrum of in vivo and clinical activity against human tumours, and its tolerability in human patients, SG2000 has potential as a novel therapeutic against spontaneously occurring canine malignancies. RESULTS: In vitro cytotoxicity was assessed using SRB and MTT assays, and in vivo activity was assessed using canine tumour xenografts. DNA interstrand cross-linking (ICL) was determined using a modification of the single cell gel electrophoresis (comet) assay. Effects on cell cycle distribution were assessed by flow cytometry and measurement of γ-H2AX by immunofluorescence and immunohistochemistry. SG2000 had a multi-log differential cytotoxic profile against a panel of 12 canine tumour cell lines representing a range of common tumour types in dogs. In the CMeC-1 melanoma cell line, DNA ICLs increased linearly with dose following a 1 h treatment. Peak ICL was achieved within 1 h and no removal was observed over 48 h. A relationship between DNA ICL formation and cytotoxicity was observed across cell lines. The formation of γ-H2AX foci was slow, becoming evident after 4 h and reaching a peak at 24 h. SG2000 exhibited significant anti-tumour activity against two canine melanoma tumour models in vivo. Anti-tumour activity was observed at 0.15 and 0.3 mg/kg given i.v. either once, or weekly x 3. Dose-dependent DNA ICL was observed in tumours (and to a lower level in peripheral blood mononuclear cells) at 2 h and persisted at 24 h. ICL increased following the second and third doses in a repeated dose schedule. At 24 h, dose dependent γ-H2AX foci were more numerous than at 2 h, and greater in tumours than in peripheral blood mononuclear cells. SG2000-induced H2AX phosphorylation measured by immunohistochemistry showed good correspondence, but less sensitivity, than measurement of foci. CONCLUSIONS: SG2000 displayed potent activity in vitro against canine cancer cell lines as a result of the formation and persistence of DNA ICLs. SG2000 also had significant in vivo antitumour activity against canine melanoma xenografts, and the comet and γ-H2AX foci methods were relevant pharmacodynamic assays. The clinical testing of SG2000 against spontaneous canine cancer is warranted. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12917-015-0534-2) contains supplementary material, which is available to authorized users