NLRP3 inflammasome is required in murine asthma in the absence of aluminum adjuvant.

Background: Inflammasome activation with the production of IL-1 beta received substantial attention recently in inflammatory diseases. However, the role of inflammasome in the pathogenesis of asthma is not clear. Using an adjuvant-free model of allergic lung inflammation induced by ovalbumin (OVA), we investigated the role of NLRP3 inflammasome and related it to IL-1R1 signaling pathway.Methods: Allergic lung inflammation induced by OVA was evaluated in vivo in mice deficient in NLRP3 inflammasome, IL-1R1, IL-1 beta or IL-1 alpha. Eosinophil recruitment, Th2 cytokine, and chemokine levels were determined in bronchoalveolar lavage fluid, lung homogenates, and mediastinal lymph node cells ex vivo.Results: Allergic airway inflammation depends on NLRP3 inflammasome activation. Dendritic cell recruitment into lymph nodes, Th2 lymphocyte activation in the lung and secretion of Th2 cytokines and chemokines are reduced in the absence of NLRP3. Absence of NLRP3 and IL-1 beta is associated with reduced expression of other proinflammatory cytokines such as IL-5, IL-13, IL-33, and thymic stromal lymphopoietin. Furthermore, the critical role of IL-1R1 signaling in allergic inflammation is confirmed in IL-1R1-, IL-1 beta-, and IL-1 alpha-deficient mice.Conclusion: NLRP3 inflammasome activation leading to IL-1 production is critical for the induction of a Th2 inflammatory allergic response.

Liquid line of descent of a basanitic liquid at 1.5 GPa: Constraints for metasomatic vein formation

The metasomatism observed in the oceanic and continental lithosphere is generally interpreted to represent a continuous differentiation process forming anhydrous and hydrous veins plus a cryptic enrichment in the surrounding peridotite. In order to constrain the mechanisms of vein formation and potentially clarify the nature and origin of the initial metasomatic agent, we performed a series of high-pressure experiments simulating the liquid line of descent of a basanitic magma differentiating within continental or mature oceanic lithosphere. This series of experiments has been conducted in an end-loaded piston cylinder apparatus starting from an initial hydrous ne-normative basanite at 1.5 GPa and temperature varying between 1,250 and 980°C. Near-pure fractional crystallization process was achieved in a stepwise manner in 30°C temperature steps and starting compositions corresponding to the liquid composition of the previous, higher-temperature glass composition. Liquids evolve progressively from basanite to peralkaline, aluminum-rich compositions without significant SiO2 variation. The resulting cumulates are characterized by an anhydrous clinopyroxene + olivine assemblage at high temperature (1,250-1,160°C), while at lower temperature (1,130-980°C), hydrous cumulates with dominantly amphibole + minor clinopyroxene, spinel, ilmenite, titanomagnetite and apatite (1,130-980°C) are formed. This new data set supports the interpretation that anhydrous and hydrous metasomatic veins could be produced during continuous differentiation processes of primary, hydrous alkaline magmas at high pressure. However, the comparison between the cumulates generated by the fractional crystallization from an initial ne-normative liquid or from hy-normative initial compositions (hawaiite or picrobasalt) indicates that for all hydrous liquids, the different phases formed upon differentiation are mostly similar even though the proportions of hydrous versus anhydrous minerals could vary significantly. This suggests that the formation of amphibole-bearing metasomatic veins observed in the lithospheric mantle could be linked to the differentiation of initial liquids ranging from ne-normative to hy-normative in composition. The present study does not resolve the question whether the metasomatism observed in lithospheric mantle is a precursor or a consequence of alkaline magmatism; however, it confirms that the percolation and differentiation of a liquid produced by a low degree of partial melting of a source similar or slightly more enriched than depleted MORB mantle could generate hydrous metasomatic veins interpreted as a potential source for alkaline magmatism by various authors.

Use of quantum dots in aqueous solution to detect blood fingermarks on non-porous surfaces.

A new and original reagent based on the use of highly fluorescent cadmium telluride (CdTe) quantum dots (QDs) in aqueous solution is proposed to detect weak fingermarks in blood on non-porous surfaces. To assess the efficiency of this approach, comparisons were performed with one of the most efficient blood reagents on non-porous surfaces, Acid Yellow 7 (AY7). To this end, four non-porous surfaces were studied, i.e. glass, transparent polypropylene, black polyethylene, and aluminium foil. To evaluate the sensitivity of both reagents, sets of depleted fingermarks were prepared, using the same finger, initially soaked with blood, which was then successively applied on the same surface without recharging it with blood or latent secretions. The successive marks were then cut in halves and the halves treated separately with each reagent. The results showed that QDs were equally efficient to AY7 on glass, polyethylene and polypropylene surfaces, and were superior to AY7 on aluminium. The use of QDs in new, sensitive and highly efficient latent and blood mark detection techniques appears highly promising. Health and safety issues related to the use of cadmium are also discussed. It is suggested that applying QDs in aqueous solution (and not as a dry dusting powder) considerably lowers the toxicity risks.

Large fluctuations in the disassembly rate of microtubules revealed by atomic force microscopy.

Atomic force microscopy (AFM) in situ has been used to observe the cold disassembly dynamics of microtubules at a previously unrealised spatial resolution. Microtubules either electrostatically or covalently bound to aminosilane surfaces disassembled at room temperature under buffer solutions with no free tubulin present. This process was followed by taking sequential tapping-mode AFM images and measuring the change in the microtubule end position as a function of time, with an spatial accuracy down to +/-20nm and a temporal accuracy of +/-1s. As well as giving average disassembly rates on the order of 1-10 tubulin monomers per second, large fluctuations in the disassembly rate were revealed, indicating that the process is far from smooth and linear under these experimental conditions. The surface bound rates measured here are comparable to the rates for GMPCPP-tubulin microtubules free in solution, suggesting that inhibition of tubulin curvature through steric hindrance controls the average, relatively low disassembly rate. The large fluctuations in this rate are thought to be due to multiple pathways in the kinetics of disassembly with differing rate constants and/or stalling due to defects in the microtubule lattice. Microtubules that were covalently bound to the surface left behind the protofilaments covalently cross-linked to the aminosilane via glutaraldehyde during the disassembly process. Further work is needed to quantitatively assess the effects of surface binding on protofibril disassembly rates, reveal any differences in disassembly rates between the plus and minus ends and to enable assembly as well as disassembly to be imaged in the microscope fluid cell in real-time.

Direct visualization of the trimeric structure of the ASIC1a channel, using AFM imaging.

There has been confusion about the subunit stoichiometry of the degenerin family of ion channels. Recently, a crystal structure of acid-sensing ion channel (ASIC) 1a revealed that it assembles as a trimer. Here, we used atomic force microscopy (AFM) to image unprocessed ASIC1a bound to mica. We detected a mixture of subunit monomers, dimers and trimers. In some cases, triple-subunit clusters were clearly visible, confirming the trimeric structure of the channel, and indicating that the trimer sometimes disaggregated after adhesion to the mica surface. This AFM-based technique will now enable us to determine the subunit arrangement within heteromeric ASICs.

The inflammasomes: guardians of the body.

The innate immune system relies on its capacity to rapidly detect invading pathogenic microbes as foreign and to eliminate them. The discovery of Toll-like receptors (TLRs) provided a class of membrane receptors that sense extracellular microbes and trigger antipathogen signaling cascades. More recently, intracellular microbial sensors have been identified, including NOD-like receptors (NLRs). Some of the NLRs also sense nonmicrobial danger signals and form large cytoplasmic complexes called inflammasomes that link the sensing of microbial products and metabolic stress to the proteolytic activation of the proinflammatory cytokines IL-1beta and IL-18. The NALP3 inflammasome has been associated with several autoinflammatory conditions including gout. Likewise, the NALP3 inflammasome is a crucial element in the adjuvant effect of aluminum and can direct a humoral adaptive immune response. In this review, we discuss the role of NLRs, and in particular the inflammasomes, in the recognition of microbial and danger components and the role they play in health and disease.

Toll-like receptor 3 is necessary for dsRNA adjuvant effects.

Toll-like receptors (TLR) recognize pathogen associated molecular patterns, and the binding of their specific ligands triggers a proinflammatory response that helps to fight invading microorganisms, and can be harnessed to increase vaccine efficiency. The present study demonstrates that double-stranded RNA is a promising vaccine adjuvant able to increase both proliferation and activation of antigen-specific CD8(+) T cells. Importantly, TLR3 is required for this adjuvant effect, as TLR3 deficient recipients failed to enhance proliferation of adoptively transferred TCR transgenic CD8(+) T cells in the presence of double-stranded RNA. Finally, this study also shows that, in contrast to previous reports in humans, TLR3 does not exert direct costimulatory activity on CD8(+) T cells in mice.

Different PTH response to oral peptone load and oral calcium load in patients with normocalcemic primary hyperparathyroidism, primary hyperparathyroidism, and healthy subjects.

BACKGROUND: Normocalcemic primary hyperparathyroidism (PHPT-N) is a condition that may have similar long-term implications to primary hyperparathyroidism (PHPT); however, differential diagnosis and treatment for parathyroid disorders are not clearly defined. We investigated the effect of an oral peptone and an oral calcium load on calcium-regulating hormones in PHPT-N compared with PHPT and healthy controls to provide a new potential diagnostic tool. DESIGN: Case-control study. METHODS: We evaluated serum gastrin, PTH, ionized calcium, and phosphate responses to oral calcium (1 g) and peptone (10 g) load in 22 PHPT and 20 PHPT-N patients matched for PTH serum values. Moreover, 30 healthy subjects were enrolled as controls. In 12 patients for each group, we also performed the oral peptone test adding aluminum hydroxide (AH) to suppress phosphate absorption. RESULTS: In PHPT patients, PTH increased significantly 30 min after the oral peptone load, while no significant increase was found in PHPT-N and controls. After oral calcium load, PTH remained stable in PHPT while it decreased dramatically in PHPT-N patients, and ionized calcium increased significantly in each of the three groups. Peptones plus AH induced a blunted PTH increase in the three groups. CONCLUSIONS: Considering the marked difference in PTH response elicited by peptones in PHPT compared with PHPT-N, we suggest that the oral peptone test could be added to the diagnostic evaluation of PHPT patients. In case of absent response to peptones, patients should have their serum calcium levels assessed twice a year in accordance with recent guidelines.

Acute hepatitis A in a young returning traveler from Kenya despite immunization before departure.

Aluminum-adsorbed hepatitis A vaccines are known to be highly efficient. We present here the case of a patient who was immunized against hepatitis A before leaving for Kenya and who contracted an acute symptomatic hepatitis A during travel.

High-throughput and sensitive quantitation of plasma catecholamines by ultraperformance liquid chromatography-tandem mass spectrometry using a solid phase microwell extraction plate.

Plasma catecholamines provide a reliable biomarker of sympathetic activity. The low circulating concentrations of catecholamines and analytical interferences require tedious sample preparation and long chromatographic runs to ensure their accurate quantification by HPLC with electrochemical detection. Published or commercially available methods relying on solid phase extraction technology lack sensitivity or require derivatization of catecholamine by hazardous reagents prior to tandem mass spectrometry (MS) analysis. Here, we manufactured a novel 96-well microplate device specifically designed to extract plasma catecholamines prior to their quantification by a new and highly sensitive ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method. Processing time, which included sample purification on activated aluminum oxide and elution, is less than 1 h per 96-well microplate. The UPLC-MS/MS analysis run time is 2.0 min per sample. This UPLC-MS/MS method does not require a derivatization step, reduces the turnaround time by 10-fold compared to conventional methods used for routine application, and allows catecholamine quantification in reduced plasma sample volumes (50-250 μL, e.g., from children and mice).

Mineral zoning and geochemistry of epithermal polymetallic Zn-Pb-Ag-Cu-Bi mineralization at Cerro de Pasco, Peru

The large Cerro de Pasco Cordilleran base metal deposit in central Peru is located on the eastern margin of a middle Miocene diatreme-dome complex and comprises two mineralization stages. The first stage consists of a large pyrite-quartz body replacing Lower Mesozoic Pucara carbonate rocks and, to a lesser extent, diatreme breccia. This body is composed of pyrite with pyrrhotite inclusions, quartz, and black and red chalcedony (containing hypogene hematite). At the contact with the pyrite-quartz body, the diatreme breccia is altered to pyrite-quartz-sericite-pyrite. This body was, in part, replaced by pipelike pyrrhotite bodies zoned outward to carbonate-replacement Zn-Pb ores hearing Fe-rich sphalerite (up to 24 mol % Fes). The second mineralization stage is partly superimposed on the first and consists of zoned east-west-trending Cu-Ag-(Au-Zn-Pb) enargite-pyrite veins hosted in the diatreme breccia in the western part of the deposit and well-zoned Zn-Pb-(Bi-Ag-Cu) carbonate-replacement orebodies; in both cases, sphalerite is Fe poor and the inner parts of the orebodies show typically advanced argillic alteration assemblages, including aluminum phosphate Sulfate (APS) minerals. The zoned enargite-pyrite veins display mineral zoning, from a core of enargite-pyrite +/- alunite with traces of Au, through an intermediate zone of tennantite, chalcopyrite, and Bi minerals to a poorly developed Outer zone hearing sphalerite-galena +/- kaolinite. The carbonate-hosted replacement ores are controlled along N 35 degrees E, N 90 degrees E, N 120 degrees E, and N 170 degrees E faults. They form well-zoned upward-flaring pipelike orebodies with a core of famatinite-pyrite and alunite, an intermediate zone with tetrahedrite-pyrite, chalcopyrite, matildite, cuprobismutite, emplectite, and other Bi minerals accompanied by APS minerals, kaolinite, and dickite, and an outer zone composed of Fe-poor sphalerite (in the range of 0.05-3.5 mol % Fes) and galena. The outermost zone consists of hematite, magnetite, and Fe-Mn-Zn-Ca-Mg carbonates. Most of the second-stage carbonate-replacement orebodies plunge between 25 degrees and 60 degrees to the west, suggesting that the hydrothermal fluids ascended from deeper levels and that no lateral feeding from the veins to the carbonate-replacement orebodies took place. In the Venencocha and Santa Rosa areas, located 2.5 km northwest of the Cerro de Pasco open pit and in the southern part of the deposit, respectively, advanced argillic altered dacitic domes and oxidized veins with advanced argillic alteration halos occur. The latter veins are possibly the oxidized equivalent of the second-stage enargite-pyrite veins located in the western part of the deposit. The alteration assemblage quartz-muscovite-pyrite associated with the pyrite-quartz body suggests that the first stage precipitated at slightly, acidic fin. The sulfide mineral assemblages define an evolutionary path close to the pyrite-pyrrhotite boundary and are characteristic of low-sulfidation states; they suggest that the oxidizing slightly acidic hydrothermal fluid was buffered by phyllite, shale, and carbonate host rock. However, the presence in the pyrite-quartz body of hematite within quartz suggests that, locally, the fluids were less buffered by the host rock. The mineral assemblages of the second mineralization stage are characteristic of high- to intermediate-sulfidation states. High-sulfidation states and oxidizing conditions were achieved and maintained in the cores of the second-stage orebodies, even in those replacing carbonate rocks. The observation that, in places, second-stage mineral assemblages are found in the inner and outer zones is explained in terms of the hydrothermal fluid advancing and waning. Microthermometric data from fluid inclusions in quartz indicate that the different ores of the first mineralization stage formed at similar temperatures and moderate salinities (200 degrees-275 degrees C and 0.2-6.8 wt % NaCl equiv in the pyrite-quartz body; 192 degrees-250 degrees C and 1.1-4.3 wt % NaCl equiv in the pyrrhotite bodies; and 183 degrees-212 degrees C and 3.2-4.0 wt % NaCl equiv in the Zn-Pb ores). These values are similar to those obtained for fluid inclusions in quartz and sphalerite from the second-stage ores (187 degrees-293 degrees C and 0.2-5.2 wt % NaCl equiv in the enargite-pyrite veins: 178 degrees-265 degrees C and 0.2-7.5 wt % NaCl equiv in quartz of carbonate-replacement orebodies; 168 degrees-999 degrees C and 3-11.8 wt % NaCl equiv in sphalerite of carbonate-replacement orebodies; and 245 degrees-261 degrees C and 3.2-7.7 wt % NaCl equiv in quartz from Venencocha). Oxygen and hydrogen isotope compositions oil kaolinite from carbonate-replacement orebodies (delta(18)O = 5.3-11.5%o, delta D = -82 to -114%o) and on alunite from the Venencocha and Santa Rosa areas (delta(18)O = 1.9-6.9%o, delta D = -56 to -73%o). Oxygen isotope compositions of quartz from the first and second stages have 6180 values from 9.1 to 1.7.8 per mil. Calculated fluids in equilibrium with kaolinite have delta(18)O values of 2.0 to 8.2 and delta D values of -69 to -97 per mil; values in equilibrium with alunite are -1.4 to -6.4 and -62 to -79 per mil. Sulfur isotope compositions of sulfides from both stages have a narrow range of delta(34)S values, between -3.7 and +4.2 per mil; values for sulfates from the second stage are between 4.2 and 31.2 per mil. These results define two mixing trends for the ore-forming fluids. The first trend reflects mixing between a moderately saline (similar to 10 wt % NaCl equiv) magmatic end member that had degassed (as indicated by the low delta D values) and meteoric water. The second mixing indicates condensation of magmatic vapor with HCl and SO(2) into meteoric water, which formed alunite. The hydrothermal system at Cerro de Pasco was emplaced at a shallow depth (similar to 500 m) in the epithermal and upper part of a porphyry environment. The similar temperatures and salinities obtained for the first stage and second stages, together with the stable isotope data, indicate that both stages are linked and represent successive stages of epithermal polymetallic mineralization in the upper part of a porphyry system.

Cutting edge: alum adjuvant stimulates inflammatory dendritic cells through activation of the NALP3 inflammasome.

Adjuvants are vaccine additives that stimulate the immune system without having any specific antigenic effect of itself. In this study we show that alum adjuvant induces the release of IL-1beta from macrophages and dendritic cells and that this is abrogated in cells lacking various NALP3 inflammasome components. The NALP3 inflammasome is also required in vivo for the innate immune response to OVA in alum. The early production of IL-1beta and the influx of inflammatory cells into the peritoneal cavity is strongly reduced in NALP3-deficient mice. The activation of adaptive cellular immunity to OVA-alum is initiated by monocytic dendritic cell precursors that induce the expansion of Ag-specific T cells in a NALP3-dependent way. We propose that, in addition to TLR stimulators, agonists of the NALP3 inflammasome should also be considered as vaccine adjuvants.