Reverse Micelles as a Tool to Probe the Synergy Between Confinement and Osmolytes with Respect to Protein Hydration Properties

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Polyethylene Glycol Camouflaged Earthworm Hemoglobin.

Nearly 21 million components of blood and whole blood and transfused annually in the United States, while on average only 13.6 million units of blood are donated. As the demand for Red Blood Cells (RBCs) continues to increase due to the aging population, this deficit will be more significant. Despite decades of research to develop hemoglobin (Hb) based oxygen (O2) carriers (HBOCs) as RBC substitutes, there are no products approved for clinical use. Lumbricus terrestris erythrocruorin (LtEc) is the large acellular O2 carrying protein complex found in the earthworm Lumbricus terrestris. LtEc is an extremely stable protein complex, resistant to autoxidation, and capable of transporting O2 to tissue when transfused into mammals. These characteristics render LtEc a promising candidate for the development of the next generation HBOCs. LtEc has a short half-life in circulation, limiting its application as a bridge over days, until blood became available. Conjugation with polyethylene glycol (PEG-LtEc) can extend LtEc circulation time. This study explores PEG-LtEc pharmacokinetics and pharmacodynamics. To study PEG-LtEc pharmacokinetics, hamsters instrumented with the dorsal window chamber were subjected to a 40% exchange transfusion with 10 g/dL PEG-LtEc or LtEc and followed for 48 hours. To study the vascular response of PEG-LtEc, hamsters instrumented with the dorsal window chamber received multiple infusions of 10 g/dL PEG-LtEc or LtEc solution to increase plasma LtEc concentration to 0.5, then 1.0, and 1.5 g/dL, while monitoring the animals' systemic and microcirculatory parameters. Results confirm that PEGylation of LtEc increases its circulation time, extending the half-life to 70 hours, 4 times longer than that of unPEGylated LtEc. However, PEGylation increased the rate of LtEc oxidation in vivo. Vascular analysis verified that PEG-LtEc showed the absence of microvascular vasoconstriction or systemic hypertension. The molecular size of PEG-LtEc did not change the colloid osmotic pressure or blood volume expansion capacity compared to LtEc, due to LtEc's already large molecular size. Taken together, these results further encourage the development of PEG-LtEc as an O2 carrying therapeutic

Nitric Oxide Releasing Nanoparticles Are Therapeutic for Staphylococcus aureus Abscesses in a Murine Model of Infection

Staphylococcus aureus (SA) is a leading cause of a diverse spectrum of bacterial diseases, including abscesses. Nitric oxide (NO) is a critical component of the natural host defense against pathogens such as SA, but its therapeutic applications have been limited by a lack of effective delivery options. We tested the efficacy of a NO-releasing nanoparticle system (NO-np) in methicillin-resistant SA (MRSA) abscesses in mice. The results show that the NO-np exert antimicrobial activity against MRSA in vitro and in abscesses. Topical or intradermal NO-np treatment of abscesses reduces the involved area and bacterial load while improving skin architecture. Notably, we evaluated pro- and anti-inflammatory cytokines that are involved in immunomodulation and wound healing, revealing that NO-np lead to a reduction in angiogenesis preventing bacterial dissemination from abscesses. These data suggest that NO-np may be useful therapeutics for microbial abscesses

Nitric Oxide-Releasing Nanoparticles Prevent Propionibacterium acnes-Induced Inflammation by Both Clearing the Organism and Inhibiting Microbial Stimulation of the Innate Immune Response.

Propionibacterium acnes induction of IL-1 cytokines through the NLRP3 (NLR, nucleotide oligomerization domain-like receptor) inflammasome was recently highlighted as a dominant etiological factor for acne vulgaris. Therefore, therapeutics targeting both the stimulus and the cascade would be ideal. Nitric oxide (NO), a potent biological messenger, has documented broad-spectrum antimicrobial and immunomodulatory properties. To harness these characteristics to target acne, we used an established nanotechnology capable of generating/releasing NO over time (NO-np). P. acnes was found to be highly sensitive to all concentrations of NO-np tested, although human keratinocyte, monocyte, and embryonic zebra fish assays revealed no cytotoxicity. NO-np significantly suppressed IL-1β, tumor necrosis factor-α (TNF-α), IL-8, and IL-6 from human monocytes, and IL-8 and IL-6 from human keratinocytes, respectively. Importantly, silencing of NLRP3 expression by small interfering RNA did not limit NO-np inhibition of IL-1 β secretion from monocytes, and neither TNF-α nor IL-6 secretion, nor inhibition by NO-np was found to be dependent on this pathway. The observed mechanism by which NO-np impacts IL-1β secretion was through inhibition of caspase-1 and IL-1β gene expression. Together, these data suggest that NO-np can effectively prevent P. acnes-induced inflammation by both clearing the organism and inhibiting microbial stimulation of the innate immune response

Robust convergence in pulse coupled oscillators with delays

We show that for pulse coupled oscillators a class of phase response curves with both excitation and inhibition exhibit robust convergence to synchrony on arbitrary aperiodic connected graphs with delays. We describe the basins of convergence and give explicit bounds on the convergence times. These results provide new and more robust methods for synchronization of sensor nets and also have biological implications.Comment: http://prl.aps.org/abstract/PRL/v106/i19/e19410

Semihemoglobins, High Oxygen Affinity Dimeric Forms of Human Hemoglobin Respond Efficiently to Allosteric Effectors without Forming Tetramers

Significant reduction in oxygen affinity resulting from interactions between heterotropic allosteric effectors and hemoglobin in not only the unligated derivative but also the fully ligated form has been reported (Tsuneshige, A., Park, S. I., and Yonetani, T. (2002) Biophys. Chem. 98, 49-63; Yonetani, T., Park, S. I., Tsuneshige, A., Imai, K., and Kanaori, K. (2002) J. Biol. Chem. 277, 34508-34520). To further investigate this effect in more detail, alpha- and beta-semihemoglobins, namely, alpha(heme)beta(apo) and alpha(apo)beta(heme), respectively, were prepared and characterized with respect to the impact of allosteric effectors on both conformation and ligand binding properties. Semihemoglobins are dimers characterized by a high affinity for oxygen and lack of cooperativity. We found that, compared with stripped conditions, semihemoglobins responded to effectors (inositol hexaphosphate and L35) by decreasing the affinity for oxygen by 60- and 130-fold for alpha- and beta-semihemoglobins, respectively. 1H NMR and sedimentation velocity experiments carried out with their ligated and unligated forms in the absence and presence of effectors revealed that semihemoglobins always remain as single-heme-carrying dimers. Recombination kinetics of their photolyzed CO derivatives showed that effectors did indeed interact with their ligated forms. Measurements of the Fe-His stretching mode show that the semihemoglobins undergo a large ligand binding-induced conformational shift and that both ligand-free and ligand derivatives respond to the presence of effectors. Contradictions to the Monod-Wyman-Changeaux/Perutz allosteric model arise since 1) the modulation of ligand affinity is not achieved in semihemoglobins by the formation of a low affinity T conformation (quaternary effect) but by direct interaction with effectors, 2) effectors do interact significantly with ligated forms of high affinity semihemoglobins, and 3) modulation of the ligand affinity and the cooperativity are not necessarily linked but instead can be separated into two distinct phenomena that can be isolated

Nonlinear Dynamical Stability of Newtonian Rotating White Dwarfs and Supermassive Stars

We prove general nonlinear stability and existence theorems for rotating star solutions which are axi-symmetric steady-state solutions of the compressible isentropic Euler-Poisson equations in 3 spatial dimensions. We apply our results to rotating and non-rotating white dwarf, and rotating high density supermassive (extreme relativistic) stars, stars which are in convective equilibrium and have uniform chemical composition. This paper is a continuation of our earlier work ([28])

Existence and Nonlinear Stability of Rotating Star Solutions of the Compressible Euler-Poisson Equations

We prove existence of rotating star solutions which are steady-state solutions of the compressible isentropic Euler-Poisson (EP) equations in 3 spatial dimensions, with prescribed angular momentum and total mass. This problem can be formulated as a variational problem of finding a minimizer of an energy functional in a broader class of functions having less symmetry than those functions considered in the classical Auchmuty-Beals paper. We prove the nonlinear dynamical stability of these solutions with perturbations having the same total mass and symmetry as the rotating star solution. We also prove local in time stability of W^{1, \infty}(\RR^3) solutions where the perturbations are entropy-weak solutions of the EP equations. Finally, we give a uniform (in time) a-priori estimate for entropy-weak solutions of the EP equations