A unique serpin P1′ glutamate and a conserved β-sheet C arginine are key residues for activity, protease recognition and stability of serpinA12 (vaspin)

SerpinA12 (vaspin) is thought to be mainly expressed in adipose tissue and has multiple beneficial effects on metabolic, inflammatory and atherogenic processes related to obesity. KLK7 (kallikrein 7) is the only known protease target of vaspin to date and is inhibited with a moderate inhibition rate. In the crystal structure, the cleavage site (P1-P1′) of the vaspin reactive centre loop is fairly rigid compared with the flexible residues before P2, possibly supported by an ionic interaction of P1′ glutamate (Glu379) with an arginine residue (Arg302) of the β-sheet C. A P1′ glutamate seems highly unusual and unfavourable for the protease KLK7. We characterized vaspin mutants to investigate the roles of these two residues in protease inhibition and recognition by vaspin. Reactive centre loop mutations changing the P1′ residue or altering the reactive centre loop conformation significantly increased inhibition parameters, whereas removal of the positive charge within β-sheet C impeded the serpin–protease interaction. Arg302 is a crucial contact to enable vaspin recognition by KLK7 and it supports moderate inhibition of the serpin despite the presence of the detrimental P1′ Glu379, which clearly represents a major limiting factor for vaspin-inhibitory activity. We also show that the vaspin-inhibition rate for KLK7 can be modestly increased by heparin and demonstrate that vaspin is a heparin-binding serpin. Noteworthily, we observed vaspin as a remarkably thermostable serpin and found that Glu379 and Arg302 influence heat-induced polymerization. These structural and functional results reveal the mechanistic basis of how reactive centre loop sequence and exosite interaction in vaspin enable KLK7 recognition and regulate protease inhibition as well as stability of this adipose tissue-derived serpin

Experimental realization of a thermal squeezed state of levitated optomechanics

We experimentally squeeze the thermal motional state of an optically levitated nanosphere by fast switching between two trapping frequencies. The measured phase-space distribution of the center of mass of our particle shows the typical shape of a squeezed thermal state, from which we infer up to 2.7 dB of squeezing along one motional direction. In these experiments the average thermal occupancy is high and, even after squeezing, the motional state remains in the remit of classical statistical mechanics. Nevertheless, we argue that the manipulation scheme described here could be used to achieve squeezing in the quantum regime if preceded by cooling of the levitated mechanical oscillator. Additionally, a higher degree of squeezing could, in principle, be achieved by repeating the frequency-switching protocol multiple times

Religious practices and long-term survival after hospital discharge for an acute coronary syndrome

BACKGROUND: Prior studies of healthy populations have found religious practices to be associated with survival. However, no contemporary studies have examined whether religiosity influences survival among patients discharged from the hospital after an acute coronary syndrome (ACS). The present study examined the relationship between religious practices and 2-year all-cause mortality among hospital survivors of an ACS. METHODS: Patients hospitalized for an ACS were recruited from 6 medical centers in Massachusetts and Georgia between 2011 and 2013. Study participants self-reported three items assessing religiosity: strength/comfort from religion, petition prayers for health, and awareness of intercessory prayers by others. All cause-mortality within 2-years of hospital discharge was ascertained by review of medical records at participating study hospitals and from death certificates. Cox proportional hazards models were used to estimate the multivariable adjusted risk of 2-year all-cause mortality. RESULTS: Participants (n = 2,068) were on average 61 years old, 34% were women, and 81% were non-Hispanic White. Approximately 85% derived strength/comfort from religion, 61% prayed for their health, and 89% were aware of intercessions. Overall, 6% died within 2 years post-discharge. After adjusting for sociodemographic variables (age, sex, and race/ethnicity), petition prayers were associated with an increased risk of 2-year all-cause mortality (HR: 1.64; 95% CI: 1.01-2.66). With further adjustment for several clinical and psychosocial measures, this association was no longer statistically significant. Strength and comfort from religion and intercessory prayers were not significantly associated with mortality. CONCLUSIONS: Most ACS survivors acknowledge deriving strength and comfort from religion, praying for their health, and intercessions made by others for their health. Although the reported religious practices were not associated with post-discharge survival after multivariable adjustment, acknowledging that patients utilize their religious beliefs and practices as strategies to improve their health would ensure a more holistic approach to patient management and promote cultural competence in healthcare

Application to the Analysis of Germinal Center Reactions In Vivo

Simultaneous detection of multiple cellular and molecular players in their native environment, one of the keys to a full understanding of immune processes, remains challenging for in vivo microscopy. Here, we present a synergistic strategy for spectrally multiplexed in vivo imaging composed of (i) triple two-photon excitation using spatiotemporal synchronization of two femtosecond lasers, (ii) a broad set of fluorophores with emission ranging from blue to near infrared, (iii) an effective spectral unmixing algorithm. Using our approach, we simultaneously excite and detect seven fluorophores expressed in distinct cellular and tissue compartments, plus second harmonics generation from collagen fibers in lymph nodes. This enables us to visualize the dynamic interplay of all the central cellular players during germinal center reactions. While current in vivo imaging typically enables recording the dynamics of 4 tissue components at a time, our strategy allows a more comprehensive analysis of cellular dynamics involving 8 single-labeled compartments. It enables to investigate the orchestration of multiple cellular subsets determining tissue function, thus, opening the way for a mechanistic understanding of complex pathophysiologic processes in vivo. In the future, the design of transgenic mice combining a larger spectrum of fluorescent proteins will reveal the full potential of our method

Crystal structure of cleaved vaspin (serpinA12)

The adipokine vaspin (serpinA12) is mainly expressed in white adipose tissue and exhibits various beneficial effects on obesity-related processes. Kallikrein 7 is the only known target protease of vaspin and is inhibited by the classical serpin inhibitory mechanism involving a cleavage of the reactive center loop between P1 (M378) and P1′ (E379). Here, we present the X-ray structure of vaspin, cleaved between M378 and E379. We provide a comprehensive analysis of differences between the uncleaved and cleaved forms in the shutter, breach, and hinge regions with relation to common molecular features underlying the serpin inhibitory mode. Furthermore, we point out differences towards other serpins and provide novel data underlining the remarkable stability of vaspin. We speculate that the previously reported FKGx1Wx2x3 motif in the breach region may play a decisive role in determining the reactive center loop configuration in the native vaspin state and might contribute to the high thermostability of vaspin. Thus, this structure may provide a basis for future mutational studies

Search for Charginos with a Small Mass Difference with the Lightest Supersymmetric Particle at \sqrt{s} = 189 GeV

A search for charginos nearly mass-degenerate with the lightest supersymmetric particle is performed using the 176 pb^-1 of data collected at 189 GeV in 1998 with the L3 detector. Mass differences between the chargino and the lightest supersymmetric particle below 4 GeV are considered. The presence of a high transverse momentum photon is required to single out the signal from the photon-photon interaction background. No evidence for charginos is found and upper limits on the cross section for chargino pair production are set. For the first time, in the case of heavy scalar leptons, chargino mass limits are obtained for any \tilde{\chi}^{+-}_1 - \tilde{\chi}^0_1 mass difference

Search for Low Scale Gravity Effects in e+e- Collisions at LEP

Recent theories propose that quantum gravity effects may be observable at LEP energies via gravitons that couple to Standard Model particles and propagate into extra spatial dimensions. The associated production of a graviton and a photon is searched for as well as the effects of virtual graviton exchange in the processes: e+e- -> gamma gamma, ZZ, WW, mu mu, tau tau, qq and ee No evidence for this new interaction is found in the data sample collected by the L3 detector at LEP at centre-of-mass energies up to 183 GeV. Limits close to 1 TeV on the scale of this new scenario of quantum gravity are set

Imidazol-1-ylethylindazole Voltage-Gated Sodium Channel Ligands Are Neuroprotective during Optic Neuritis in a Mouse Model of Multiple Sclerosis

[Image: see text] A series of imidazol-1-ylethylindazole sodium channel ligands were developed and optimized for sodium channel inhibition and in vitro neuroprotective activity. The molecules exhibited displacement of a radiolabeled sodium channel ligand and selectivity for blockade of the inactivated state of cloned neuronal Na(v) channels. Metabolically stable analogue 6 was able to protect retinal ganglion cells during optic neuritis in a mouse model of multiple sclerosis