Effect of mechanical cycling on the stress-strain response of a martensitic Nitinol shape memory alloy

An experimental investigation into the ambient temperature, load-controlled tension�tension fatigue behavior of a martensitic Nitinol shape memory alloy (SMA) was conducted. Fatigue life for several stress levels spanning the critical stress for detwinning was determined and compared with that obtained on an alloy similar in composition but in the austenitic state at room temperature. Results show that the fatigue life of the pseudo-plastic alloy is superior to superelastic shape memory alloy. The stress�strain hysteretic response, monitored throughout the fatigue loading, reveals progressive strain accumulation with the cyclic loading. In addition, the area of hysteresis and recoverable and frictional energies were found to decrease with increasing number of fatigue cycles. Post-mortem characterization of the fatigued specimens through calorimetry and fractography was conducted in order to get further insight into the fatigue micromechanisms. These results are discussed in terms of reversible and irreversible microstructural changes that take place during cyclic loading. Aspects associated with self-heating of martensitic alloy undergoing high frequency stress cycling are discussed.

Arabidopsis thaliana J-class heat shock proteins: cellular stress sensors

Plants are sessile organisms that have evolved a variety of mechanisms to maintain their cellular homeostasis under stressful environmental conditions. Survival of plants under abiotic stress conditions requires specialized group of heat shock protein machinery, belonging to Hsp70:J-protein family. These heat shock proteins are most ubiquitous types of chaperone machineries involved in diverse cellular processes including protein folding, translocation across cell membranes, and protein degradation. They play a crucial role in maintaining the protein homeostasis by reestablishing functional native conformations under environmental stress conditions, thus providing protection to the cell. J-proteins are co-chaperones of Hsp70 machine, which play a critical role by stimulating Hsp70s ATPase activity, thereby stabilizing its interaction with client proteins. Using genome-wide analysis of Arabidopsis thaliana, here we have outlined identification and systematic classification of J-protein co-chaperones which are key regulators of Hsp70s function. In comparison with Saccharomyces cerevisiae model system, a comprehensive domain structural organization, cellular localization, and functional diversity of A. thaliana J-proteins have also been summarized. Electronic supplementary material The online version of this article (doi:10.1007/s10142-009-0132-0) contains supplementary material, which is available to authorized users.

Effect of metal coating and residual stress on the resonant frequency of MEMS resonators

MEMS resonators are designed for a fixed resonant frequency. Therefore, any shift in the resonant frequency of the final fabricated structure can be a denting factor for its suitability towards a desired application. There are numerous factors which alter the designed resonant frequency of the fabricated resonator such as the metal layer deposited on top of the beam and the residual stresses present in the fabricated structure. While the metal coating, which acts as electrode, increases the stiffness and the effective mass of the composite structure, the residual stress increases or decreases the net stiffness if it is a tensile or compressive type respectively. In this paper, we investigate both these cases by taking two different structures, namely, the micro cantilever beam with gold layer deposited on its top surface and the MEMS gyroscope with residual stresses. First, we carry out experiments to characterize both these structures to find their resonant frequencies. Later, we analytically model those effects and compare them with the experimentally obtained values. Finally, it is found that the analytical models give an error of less than 10% with respect to the experimental results in both the cases.

Disease-associated polymorphisms in ERAP1 do not alter endoplasmic reticulum stress in patients with ankylosing spondylitis

The mechanism by which human leukocyte antigen B27 (HLA-B27) contributes to ankylosing spondylitis (AS) remains unclear. Genetic studies demonstrate that association with and interaction between polymorphisms of endoplasmic reticulum aminopeptidase 1 (ERAP1) and HLA-B27 influence the risk of AS. It has been hypothesised that ERAP1-mediated HLA-B27 misfolding increases endoplasmic reticulum (ER) stress, driving an interleukin (IL) 23-dependent, pro-inflammatory immune response. We tested the hypothesis that AS-risk ERAP1 variants increase ER-stress and concomitant pro-inflammatory cytokine production in HLA-B27 + but not HLA-B27-AS patients or controls. Forty-nine AS cases and 22 healthy controls were grouped according to HLA-B27 status and AS-associated ERAP1 rs30187 genotypes: HLA-B27 + ERAP1 risk, HLA-B27 + ERAP1 protective, HLA-B27-ERAP1 risk and HLA-B27-ERAP1 protective. Expression levels of ER-stress markers GRP78 (8 kDa glucose-regulated protein), CHOP (C/EBP-homologous protein) and inflammatory cytokines were determined in peripheral blood mononuclear cell and ileal biopsies. We found no differences in ER-stress gene expression between HLA-B27 + and HLA-B27-cases or healthy controls, or between cases or controls stratified by carriage of ERAP1 risk or protective alleles in the presence or absence of HLA-B27. No differences were observed between expression of IL17A or TNF (tumour necrosis factor) in HLA-B27 + ERAP1 risk, HLA-B27 + ERAP1 protective and HLA-B27-ERAP1 protective cases. These data demonstrate that aberrant ERAP1 activity and HLA-B27 carriage does not alter ER-stress levels in AS, suggesting that ERAP1 and HLA-B27 may influence disease susceptibility through other mechanisms. © 2015 Macmillan Publishers Limited.

ERAP2 functional knockout in humans does not alter surface heavy chains or HLA-B27, inflammatory cytokines or endoplasmic reticulum stress markers

Introduction Single nucleotide polymorphisms in ERAP2 are strongly associated with ankylosing spondylitis (AS). One AS-associated single nucleotide polymorphism, rs2248374, causes a truncated ERAP2 protein that is degraded by nonsense-mediated decay. Approximately 25% of the populations of European ancestry are therefore natural ERAP2 knockouts. We investigated the effect of this associated variant on HLA class I allele presentation, surface heavy chains, endoplasmic reticulum (ER) stress markers and cytokine gene transcription in AS. Methods Patients with AS and healthy controls with either AA or GG homozygous status for rs2248374 were studied. Antibodies to CD14, CD19-ECD, HLA-A-B-C, Valpha7.2, CD161, anti-HC10 and anti-HLA-B27 were used to analyse peripheral blood mononuclear cells. Expression levels of ER stress markers (GRP78 and CHOP) and proinflammatory genes (tumour necrosis factor (TNF), IL6, IL17 and IL22) were assessed by qPCR. Results There was no significant difference in HLAclass I allele presentation or major histocompatibility class I heavy chains or ER stress markers GRP78 and CHOP or proinflammatory gene expression between genotypes for rs2248374 either between cases, between cases and controls, and between controls. Discussion Large differences were not seen in HLAB27 expression or cytokine levels between subjects with and without ERAP2 in AS cases and controls. This suggests that ERAP2 is more likely to influence AS risk through other mechanisms.

Ozone-induced signaling in Arabidopsis thaliana

Tropospheric ozone (O3) is one of the most common air pollutants in industrialized countries, and an increasing problem in rapidly industrialising and developing countries in Asia, Africa and South America. Elevated concentrations of tropospheric O3 can lead to decrease in photosynthesis rate and therefore affect the normal metabolism, growth and seed production. Acute and high O3 episodes can lead to extensive damage leading to dead tissue in plants. Thus, O3 derived growth defects can lead to reduction in crop yield thereby leading to economical losses. Despite the extensive research on this area, many questions remain open on how these processes are controlled. In this study, the stress-induced signaling routes and the components involved were elucidated in more detail starting from visual damage to changes in gene expression, signaling routes and plant hormone interactions that are involved in O3-induced cell death. In order to elucidate O3-induced responses in Arabidopsis, mitogen-activated protein kinase (MAPK) signaling was studied using different hormonal signaling mutants. MAPKs were activated at the beginning of the O3 exposure. The activity of MAPKs, which were identified as AtMPK3 and AtMPK6, reached the maximum at 1 and 2 hours after the start of the exposure, respectively. The activity decreased back to clean air levels at 8 hours after the start of the exposure. Both AtMPK3 and AtMPK6 were translocated to nucleus at the beginning of the O3 exposure where they most likely affect gene expression. Differences were seen between different hormonal signaling mutants. Functional SA signaling was shown to be needed for the full protein levels and activation of AtMPK3. In addition, AtMPK3 and AtMPK6 activation was not dependent on ethylene signaling. Finally, jasmonic acid was also shown to have an impact on AtMPK3 protein levels and AtMPK3 activity. To further study O3-induced cell death, an earlier isolated O3 sensitive Arabidopsis mutant rcd1 was mapped, cloned and further characterized. RCD1 was shown to encode a gene with WWE and ADP-ribosylation domains known to be involved in protein-protein interactions and cell signaling. rcd1 was shown to be involved in many processes including hormonal signaling and regulation of stress-responsive genes. rcd1 is sensitive against O3 and apoplastic superoxide, but tolerant against paraquat that produces superoxide in chloroplast. rcd1 is also partially insensitive to glucose and has alterations in hormone responses. These alterations are seen as ABA insensitivity, reduced jasmonic acid sensitivity and reduced ethylene sensitivity. All these features suggest that RCD1 acts as an integrative node in hormonal signaling and it is involved in the hormonal regulation of several specific stress-responsive genes. Further studies with the rcd1 mutant showed that it exhibits the classical features of programmed cell death, PCD, in response to O3. These include nuclear shrinkage, chromatin condensation, nuclear DNA degradation, cytosol vesiculation and accumulation of phenolic compounds and eventually patches of HR-like lesions. rcd1 was found to produce extensive amount of salicylic acid and jasmonic acid in response to O3. Double mutant studies showed that SA independent and dependent processes were involved in the O3-induced PCD in rcd1 and that increased sensitivity against JA led to increased sensitivity against O3. Furthermore, rcd1 had alterations in MAPK signature that resembled changes that were previously seen in mutants defective in SA and JA signaling. Nitric oxide accumulation and its impact on O3-induced cell death were also studied. Transient accumulation of NO was seen at the beginning of the O3 exposure, and during late time points, NO accumulation coincided with the HR-like lesions. NO was shown to modify defense gene expression, such as, SA and ethylene biosynthetic genes. Furthermore, rcd1 was shown to produce more NO in control conditions. In conclusion, NO was shown to be involved in O3-induced signaling leading to attenuation of SA biosynthesis and other defense related genes.

Stress and developmental responses in Arabidopsis thaliana: regulation through the transcription factor-interacting protein RCD1

Plants constantly face adverse environmental conditions, such as drought or extreme temperatures that threaten their survival. They demonstrate astonishing metabolic flexibility in overcoming these challenges and one of the key responses to stresses is changes in gene expression leading to alterations in cellular functions. This is brought about by an intricate network of transcription factors and associated regulatory proteins. Protein-protein interactions and post-translational modifications are important steps in this control system along with carefully regulated degradation of signaling proteins. This work concentrates on the RADICAL-INDUCED CELL DEATH1 (RCD1) protein which is an important regulator of abiotic stress-related and developmental responses in Arabidopsis thaliana. Plants lacking this protein function display pleiotropic phenotypes including sensitivity to apoplastic reactive oxygen species (ROS) and salt, ultraviolet B (UV-B) and paraquat tolerance, early flowering and senescence. Additionally, the mutant plants overproduce nitric oxide, have alterations in their responses to several plant hormones and perturbations in gene expression profiles. The RCD1 gene is transcriptionally unresponsive to environmental signals and the regulation of the protein function is likely to happen post-translationally. RCD1 belongs to a small protein family and, together with its closest homolog SRO1, contains three distinguishable domains: In the N-terminus, there is a WWE domain followed by a poly(ADP-ribose) polymerase-like domain which, despite sequence conservation, does not seem to be functional. The C-terminus of RCD1 contains a novel domain called RST. It is present in RCD1-like proteins throughout the plant kingdom and is able to mediate physical interactions with multiple transcription factors. In conclusion, RCD1 is a key point of signal integration that links ROS-mediated cues to transcriptional regulation by yet unidentified means, which are likely to include post-translational mechanisms. The identification of RCD1-interacting transcription factors, most of whose functions are still unknown, opens new avenues for studies on plant stress as well as developmental responses.

Elevated circulating sclerostin concentrations in individuals with high bone mass, with and without LRP5 mutations

CONTEXT: The role and importance of circulating sclerostin is poorly understood. High bone mass (HBM) caused by activating LRP5 mutations has been reported to be associated with increased plasma sclerostin concentrations; whether the same applies to HBM due to other causes is unknown. OBJECTIVE: Our objective was to determine circulating sclerostin concentrations in HBM. DESIGN AND PARTICIPANTS: In this case-control study, 406 HBM index cases were identified by screening dual-energy x-ray absorptiometry (DXA) databases from 4 United Kingdom centers (n = 219 088), excluding significant osteoarthritis/artifact. Controls comprised unaffected relatives and spouses. MAIN MEASURES: Plasma sclerostin; lumbar spine L1, total hip, and total body DXA; and radial and tibial peripheral quantitative computed tomography (subgroup only) were evaluated. RESULTS: Sclerostin concentrations were significantly higher in both LRP5 HBM and non-LRP5 HBM cases compared with controls: mean (SD) 130.1 (61.7) and 88.0 (39.3) vs 66.4 (32.3) pmol/L (both P < .001, which persisted after adjustment for a priori confounders). In combined adjusted analyses of cases and controls, sclerostin concentrations were positively related to all bone parameters found to be increased in HBM cases (ie, L1, total hip, and total body DXA bone mineral density and radial/tibial cortical area, cortical bone mineral density, and trabecular density). Although these relationships were broadly equivalent in HBM cases and controls, there was some evidence that associations between sclerostin and trabecular phenotypes were stronger in HBM cases, particularly for radial trabecular density (interaction P < .01). CONCLUSIONS: Circulating plasma sclerostin concentrations are increased in both LRP5 and non-LRP5 HBM compared with controls. In addition to the general positive relationship between sclerostin and DXA/peripheral quantitative computed tomography parameters, genetic factors predisposing to HBM may contribute to increased sclerostin levels.

Direct physical evidence for the back-transformation of stress-induced martensite in the vicinity of cracks in pseudoelastic NiTi shape memory alloys

Crack loading and crack extension in pseudoelastic binary NiTi shape memory alloy (SMA) miniature compact tension (CT) specimens with 50.7 at.% Ni (austenitic, pseudoelastic) was investigated using infrared (IR) thermography during in situ loading and unloading. IR thermographic measurements allow for the observation of heat effects associated with the stress-induced transformation of martensite from B2 to BIT during loading and the reverse transformation during unloading. The results are compared with optical images and discussed in terms of the crack growth mechanisms in pseudoelastic NiTi SMAs. Direct experimental evidence is presented which shows that crack growth occurs into a stress-induced martensitic microstructure, which immediately retransforms to austenite in the wake of the crack.

Allelopathic effects of filamentous cyanobacteria on phytoplankton in the Baltic Sea

Eutrophication and enhanced internal nutrient loading of the Baltic Sea are most clearly reflected by increased late-summer cyanobacterial blooms, which often are toxic. In addition to their toxicity to animals, phytoplankton species can be allelopathic, which means that they produce chemicals that inhibit competing phytoplankton species. Such interspecific chemical warfare may lead to the formation of harmful phytoplankton blooms and the spread of exotic species into new habitats. This is the first report on allelopathic effects in brackish-water cyanobacteria. The experimental studies presented in this thesis showed that the filamentous cyanobacteria Anabaena sp., Aphanizomenon flos-aquae and Nodularia spumigena are capable of decreasing the growth of other phytoplankton species, especially cryptophytes, but also diatoms. The detected allelopathic effects are rather transitory, and some co-occurring species show tolerance to them. The allelochemicals are excreted during active growth and they decrease cell numbers, chlorophyll a content and carbon uptake of the target species. Although the more specific modes of action or chemical structures of the allelochemicals remain to be studied, the results clearly indicate that the allelopathic effects are not caused by the hepatotoxin, nodularin. On the other hand, cyanobacteria stimulated the growth of bacteria, other cyanobacteria, chlorophytes and flagellates in a natural phytoplankton community. In a long-term data analysis of phytoplankton abundances and hydrography of the northern Baltic Sea, a clear change was observed in phytoplankton community structure, together with a transition in environmental factors, between the late 1970s and early 2000s. Surface water salinity decreased, whereas water temperature and the concentration of dissolved inorganic nitrogen increased. In the phytoplankton community, the biomass of cyanobacteria, chrysophytes and chlorophytes significantly increased, and the late-summer phytoplankton community became increasingly cyanobacteria-dominated. In contrast, the biomass of cryptophytes decreased. The increased temperature and nutrient concentrations probably explain most of the changes in phytoplankton, but my results suggest that the possible effect of chemically mediated biological interactions should also be considered. Cyanobacterial allelochemicals can cause additional stress to other phytoplankton in the nutrient-depleted late-summer environment and thus contribute to the formation and persistence of long-lasting cyanobacterial mass occurrences. On the other hand, cyanobacterial blooms may either directly or indirectly promote the growth of some phytoplankton species. Therefore, a further increase in cyanobacteria will probably shape the late-summer pelagic phytoplankton community by stimulating some species, but inhibiting others.

A novel fully validated LC–MS/MS method for quantification of pyridoxal-5′-phosphate concentrations in samples of human whole blood

Quantification of pyridoxal-5´-phosphate (PLP) in biological samples is challenging due to the presence of endogenous PLP in matrices used for preparation of calibrators and quality control samples (QCs). Hence, we have developed an LC-MS/MS method for accurate and precise measurement of the concentrations of PLP in samples (20 µL) of human whole blood that addresses this issue by using a surrogate matrix and minimizing the matrix effect. We used a surrogate matrix comprising 2% bovine serum albumin (BSA) in phosphate buffer saline (PBS) for making calibrators, QCs and the concentrations were adjusted to include the endogenous PLP concentrations in the surrogate matrix according to the method of standard addition. PLP was separated from the other components of the sample matrix using protein precipitation with trichloroacetic acid 10% w/v. After centrifugation, supernatant were injected directly into the LC-MS/MS system. Calibration curves were linear and recovery was > 92%. QCs were accurate, precise, stable for four freeze-thaw cycles, and following storage at room temperature for 17h or at -80 °C for 3 months. There was no significant matrix effect using 9 different individual human blood samples. Our novel LC-MS/MS method has satisfied all of the criteria specified in the 2012 EMEA guideline on bioanalytical method validation.