The modulation of actin polymerization is a common theme among microbial pathogens. Even though microorganisms show a wide repertoire of strategies to subvert the activity of actin, most of them ...converge in the ones that activate nucleating factors, such as the Arp2/3 complex. Brucella spp. are intracellular pathogens capable of establishing chronic infections in their hosts. The ability to subvert the host cell response is dependent on the capacity of the bacterium to attach, invade, avoid degradation in the phagocytic compartment, replicate in an endoplasmic reticulum-derived compartment and egress. Even though a significant number of mechanisms deployed by Brucella in these different phases have been identified and characterized, none of them have been described to target actin as a cellular component. In this manuscript, we describe the identification of a novel virulence factor (NpeA) that promotes niche formation. NpeA harbors a short linear motif (SLiM) present within an amphipathic alpha helix that has been described to bind the GTPase-binding domain (GBD) of N-WASP and stabilizes the autoinhibited state. Our results show that NpeA is secreted in a Type IV secretion system-dependent manner and that deletion of the gene diminishes the intracellular replication capacity of the bacterium. In vitro and ex vivo experiments demonstrate that NpeA binds N-WASP and that the short linear motif is required for the biological activity of the protein.IMPORTANCEThe modulation of actin-binding effectors that regulate the activity of this fundamental cellular protein is a common theme among bacterial pathogens. The neural Wiskott–Aldrich syndrome protein (N-WASP) is a protein that several pathogens target to hijack actin dynamics. The highly adapted intracellular bacterium Brucella has evolved a wide repertoire of virulence factors that modulate many activities of the host cell to establish successful intracellular replication niches, but, to date, no effector proteins have been implicated in the modulation of actin dynamics. We present here the identification of a virulence factor that harbors a short linear motif (SLiM) present within an amphipathic alpha helix that has been described to bind the GTPase-binding domain (GBD) of N-WASP stabilizing its autoinhibited state. We demonstrate that this protein is a Type IV secretion effector that targets N-WASP-promoting intracellular survival and niche formation.
Adult mice lacking functional GABAB receptors (GABAB1KO) show altered Gnrh1 and Gad1 expressions in the preoptic area-anterior hypothalamus (POA-AH) and females display disruption of cyclicity and ...fertility. Here we addressed whether sexual differentiation of the brain and the proper wiring of the GnRH and kisspeptin systems were already disturbed in postnatal day 4 (PND4) GABAB1KO mice.
PND4 wild-type (WT) and GABAB1KO mice of both sexes were sacrificed; tissues were collected to determine mRNA expression (qPCR), amino acids (HPLC), and hormones (RIA and/or IHC).
GnRH neuron number (IHC) did not differ among groups in olfactory bulbs or OVLT-POA. Gnrh1 mRNA (qPCR) in POA-AH was similar among groups. Gnrh1 mRNA in medial basal hypothalamus (MBH) was similar in WTs but was increased in GABAB1KO females compared to GABAB1KO males. Hypothalamic GnRH (RIA) was sexually different in WTs (males > females), but this sex difference was lost in GABAB1KOs; the same pattern was observed when analyzing only the MBH, but not in the POA-AH. Arcuate nucleus Kiss1 mRNA (micropunch-qPCR) was higher in WT females than in WT males and GABAB1KO females. Gad1 mRNA in MBH was increased in GABAB1KO females compared to GABAB1KO males. Serum LH and gonadal estradiol content were also increased in GABAB1KOs.
We demonstrate that GABABRs participate in the sexual differentiation of the ARC/MBH, because sex differences in several reproductive genes, such as Gad1, Kiss1 and Gnrh1, are critically disturbed in GABAB1KO mice at PND4, probably altering the organization and development of neural circuits governing the reproductive axis.
This work studies how the crystallinity of a raw kaolinite clay affects the properties of mullite ceramics produced from it. To this purpose, two well and poorly crystallized kaolinite samples from ...the repository of the Clay Mineral Society were assessed. Also, other two samples were produced in which the growth of mullite acicular grains were promoted by addition of a MoO3 precursor. The role of the kaolinite crystallinity and the MoO3 precursor on the properties of the final ceramics was studied. For each sample, the microstructure and the textural, crystalline, and mechanical properties were analyzed.
The poorly crystallized kaolinite is more reactive than the well crystallized one and promotes better mullitization. In addition, when the MoO3 precursor is used, the formation of grains with acicular shape inhibits the ceramic shrinkage and promotes higher porosities. On the other hand, the flexural strength is slightly lower for the mullite produced from poorly crystallized kaolinite. Nevertheless, when this property is extrapolated to the case of zero porosity it is found that in such cases the ceramic solid fraction presents higher flexural strength, indicating that the porous mullite with the acicular grain morphology is a reinforced lightweight ceramic. The obtained results show that the crystallinity of the starting clay should be considered in the ceramic fabrication process due to its effects on the properties of the final product.
•Well and poorly crystallized kaolinite were used to produce porous mullite ceramics.•Molybdenum additive was also considered to promote formation of acicular grains.•The microstructural and mechanical properties were comprehensively examined.•Poorly crystallized kaolinite can produce a reinforced lightweight ceramic.•Starting clay crystallinity should be considered for ceramic fabrication.
Kisspeptin, encoded by Kiss1, stimulates reproduction and is synthesized in the hypothalamic anteroventral periventricular and arcuate nuclei. Kiss1 is also expressed at lower levels in the medial ...amygdala (MeA) and bed nucleus of the stria terminalis (BNST), but the regulation and function of Kiss1 there is poorly understood. γ-Aminobutyric acid (GABA) also regulates reproduction, and female GABAB1 receptor knockout (KO) mice have compromised fertility. However, the interaction between GABAB receptors and Kiss1 neurons is unknown. Here, using double-label in situ hybridization, we first demonstrated that a majority of hypothalamic Kiss1 neurons coexpress GABAB1 subunit, a finding also confirmed for most MeA Kiss1 neurons. Yet, despite known reproductive impairments in GABAB1KO mice, Kiss1 expression in the anteroventral periventricular and arcuate nuclei, assessed by both in situ hybridization and real-time PCR, was identical between adult wild-type and GABAB1KO mice. Surprisingly, however, Kiss1 levels in the BNST and MeA, as well as the lateral septum (a region normally lacking Kiss1 expression), were dramatically increased in both GABAB1KO males and females. The increased Kiss1 levels in extrahypothalamic regions were not caused by elevated sex steroids (which can increase Kiss1 expression), because circulating estradiol and testosterone were equivalent between genotypes. Interestingly, increased Kiss1 expression was not detected in the MeA or BNST in prepubertal KO mice of either sex, indicating that the enhancements in extrahypothalamic Kiss1 levels initiate during/after puberty. These findings suggest that GABAB signaling may normally directly or indirectly inhibit Kiss1 expression, particularly in the BNST and MeA, and highlight the importance of studying kisspeptin populations outside the hypothalamus.
Abstract Porous mullite ceramics have good properties for high‐temperature applications, but porosity gives place to ceramics with low mechanical strength, which restricts the service life in their ...potential applications. Therefore, performing modifications at the microscale to increase the mechanical strength has become a current challenge to expand its application fields. This work describes the properties of a porous mullite–zirconia composite produced by ceramic processing, using industrial kaolin and stabilized zirconia as raw materials. The growth of mullite needle‐like grains to reinforce the ceramic was promoted by the addition of a molybdenum oxide precursor. The effect of zirconia on the composite was analyzed through an experimental multi‐technique approach and considering a pure mullite sample, identically processed, as a reference. The novel composite has a porosity of about 50%, and presents a homogeneous microstructure, with interlocked mullite needle‐like grains and dispersed rounded zirconia grains. This morphology restricts the mullite tendency to shrink during sintering, giving the material a higher stiffness. In particular, the presence of zirconia in the composite improves both the flexural strength and the apparent Young modulus of the material (about 20% and up to 600%, respectively). These results encourage further investigations to establish this composite for different technological applications.
Female mice lacking GABAB receptors, GABAB1KO, show disrupted oestrous cycles, reduced pregnancies and increased hypothalamic Gnrh1 mRNA expression, whereas anteroventral ...periventricular/periventricular preoptic nucleus (AVPV/PeN) Kiss1 mRNA was not affected. In the present study, we characterise the important components of the gonadotrophic preovulatory surge, aiming to unravel the origin of this reproductive impairment. In GABAB1KO and wild‐type (WT) females, we determined: (i) hypothalamic oestrogen receptor (ER)α and β and aromatase mRNA and protein expression; (ii) ovulation index and oestrus serum follicle‐stimulating hormone (FSH) and pituitary Gnrh1r expression; (iii) in ovariectomised‐oestradiol valerate‐treated mice, we evaluated ex vivo hypothalamic gonadotrophin‐releasing hormone (GnRH) pulsatility in the presence/absence of kisspeptin (Kiss‐10, constant or pulsatile) and oestradiol (constant); and (iv) in ovariectomised‐oestradiol silastic capsule‐treated mice (proestrous‐like environment), we evaluated morning and evening kisspeptin neurone activation (c‐Fos+) and serum luteinising homrone (LH). In the medial basal hypothalamus of oestrus GABAB1KOs, aromatase and ERα mRNA and protein were increased, whereas ERβ was decreased. In GABAB1KOs, the ovulation index was decreased together with decreased first oestrus serum FSH and increased pituitary Gnrh1r mRNA. Under constant Kiss‐10 stimulation, hypothalamic GnRH pulse frequency did not vary, although GnRH mass/pulse was increased in GABAB1KOs. In WTs, pulsatile Kiss‐10 together with constant oestradiol significantly increased GnRH pulsatility, whereas, in GABAB1KOs, oestradiol alone increased GnRH pulsatility and this was reversed by pulsatile Kiss‐10 addition. In GABAB1KOs AVPV/PeN kisspeptin neurones were similarly activated (c‐Fos+) in the morning and evening, whereas WTs showed the expected, marked evening stimulation. LH correlated with activated kisspeptin cells in WT mice, whereas GABAB1KO mice showed high, similar LH levels both in the morning and evening. Taken together, all of these alterations point to impairment in the trigger of the preovulatory GnRH surge that entails the reproductive alterations described.
Invasive candidiasis is an important cause of morbidity and mortality, and its occurrence is increasing due to the growing complexity of patients. In particular,
exhibits several virulence factors ...that facilitate yeast colonization in humans. In this sense, the photodynamic inactivation of yeasts is a promising new alternative to eliminate fungal infections. Herein, the photodynamic activity sensitized by a free-base chlorin (TPCF
) and its complexes with Zn(II) (ZnTPCF
) and Pd(II) (PdTPCF
) was investigated in order to eliminate
under different forms of cell cultures. A decrease in cell survival of more than 5 log was found in planktonic cells incubated with 5 μM TPCF
or ZnTPCF
upon 15 min of white-light irradiation. The mechanism of action mainly involved a type II pathway in the inactivation of
cells. In addition, the photodynamic action induced by these chlorins was able to suppress the growth of
in a culture medium. These photosensitizers were also effective to photoinactivate
pseudohyphae suspended in PBS. Furthermore, the biofilms of
that incorporated the chlorins during the proliferation stage were completely eradicated using 5 μM TPCF
or ZnTPCF
after 60 min of light irradiation. The studies indicated that these chlorins are effective photosensitizing agents to eliminate
as planktonic cells, pseudohyphae, and biofilms.
Aging is characterized by deterioration of biomolecules and impaired stress responses that make the elderly especially vulnerable to environmental pollutants. The pesticide p,p'-DDE is a DDT ...derivative that generates great concern because of its wide distribution and its harmful effects on both human health and the environment. We analyzed here the biological responses elicited by p,p'-DDE exposure in the liver of aged Mus spretus mice. Data demonstrate that the elderly constitute a population especially sensitive to this noxious environmental pollutant. We also demonstrated here that the daily consumption of sun-dried Pedro Ximénez (PX) white-grape must (PXM) protects the liver of aged mice from both the age and the damages caused by p,p'-DDE exposure. The PXM activity was exerted through the restoration of the hepatic metabolisms of lipids and carbohydrates and, probably, is a consequence of the ability of this polyphenol-rich mixture to avoid oxidative stress. Nutritional interventions including PXM, which ameliorates the effects of unavoidable exposure to pesticides in our food, are helpful tools that can help elderly populations to enjoy a healthy and expanded lifetime.
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•p,p'-DDE altered the plasma levels of glucose, triglycerides, and urea and the MDA concentration in mice.•p,p’-DDE perturbs the hepatic redox status, glucose physiology, several central signaling pathways, and hormone metabolism.•The antioxidant properties of Pedro Ximenez must (PXM) polyphenols prevent many of the toxic effects of p,p'-DDE.•PXM dietary intervention will improve the overall health of the elderly and promote healthier aging.
Boron oxide sources as fluxing agents for clay-based materials have been a problem for the high solubility of these additives, however the dry milling-mixing-granulating route for clay based ceramics ...has been proposed and implemented with noticeable energetic advances.
In the present study, the role of different boron oxide sources in the thermal behavior and sinterability of ceramic materials based on kaolinite clay was evaluated.
Four boron sources were evaluated: Boric acid, borax, colemanite and ulexite. Four binary mixtures were made in a planetary mill to obtain uniform mixtures. For systematic comparison, the boron content added in all samples was 1 wt%. The thermal behavior evaluation of the mixtures was carried out by a multi-technique approach accompanied by textural, microstructural characterization complemented by a deep crystalline and non-crystalline phase evolution.
Ulexite and borax presented the highest fluxing effect and colemanite has an even more moderate effect than boric acid. The observed microstructure was similar to other clay-based ceramics, with quartz grains, cristobalite and mullite embedded in the silica-based vitreous phase.
The present study provides information for the additional design of clay-based materials with boron oxides as fluxing agents. The dry route alternative for using these agents was effective.
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