The phosphoinositide 3-kinases (PI3Ks) are a conserved family of signal transduction enzymes that are involved in regulating cellular activation, inflammatory responses, chemotaxis, and apoptosis. We have discovered that a carbohydrate ligand, glucan, will stimulate the endogenous PI3K/Akt signaling pathway. This article reviews the current data on the role of the PI3K/Akt signaling pathway as a negative feedback mechanism or compensatory regulator of septic and inflammatory responses. Of greater importance, the data reviewed in this article suggest that modulation of the PI3K/ Akt signaling pathway can reduce the morbidity and mortality associated with septic and I/R injury. Thus, manipulation of the endogenous PI3K/Akt signaling pathway may represent a new and novel therapeutic approach to management of important diseases.

These studies evaluated the effects treatment with glucan phosphate, a soluble polysaccharide immunomodulator, on the inflammatory response induced by burn injury and on resistance to Pseudomonas aeruginosa burn wound infection. Mice were exposed to 35% total body surface area burns and were resuscitated with lactated Ringer's (LR) solution alone or LR supplemented with glucan phosphate (40 mg/kg). Glucan phosphate treatment attenuated burn-induced expression of interleukin (IL)-1β, IL-6, and IL-10 mRNAs in spleen, lung, and heart. Plasma concentrations of IL-1β, IL-6, macrophage inflammatory protein (MIP)-2, and IL-10 were also decreased in burned mice treated with glucan phosphate compared with vehicle-treated controls. Early postburn mortality was not significantly different between control (20%) and glucan phosphate-treated (10%) mice, but there was a small improvement in acid-base balance in the glucan phosphate-treated group. Mice received a second injection of glucan phosphate or LR on day 4 postburn and were infected by topical application of P. aeruginosa to the burn wound on day 5. Glucan phosphate treatment significantly improved survival in mice exposed to P. aeruginosa burn wound infection. The improved survival correlated with lower bacterial burden in the burn wound, attenuated production of proinflammatory cytokines, and enhanced production of Th1 cytokines. These studies show that glucan phosphate treatment attenuates burn-induced inflammation and increases resistance to P. aeruginosa burn wound infection in an experimental model of burn injury.

Pyrotechnic shock events used during stage separation on rocket vehicles produce high amplitude short duration structural response that can lead to malfunction or degradation of electronic components, cracks and fractures in brittle materials, local plastic deformation, and can cause materials to experience accelerated fatigue life. These transient loads propagate as waves through the structural media losing energy as they travel outward from the source. This work assessed available test data in an effort to better understand attenuation characteristics associated with wave propagation and attempted to update a historical standard defined by the Martin Marietta Corporation in the late 1960's using out of date data acquisition systems. Two data sets were available for consideration. The first data set came from a test that used a flight like cylinder used in NASA's Ares I-X program, and the second from a test conducted with a at plate. Both data sets suggested that the historical standard was not a conservative estimate of shock attenuation with distance, however, the variation in the test data did not lend to recommending an update to the standard. Beyond considering attenuation with distance an effort was made to model the at plate configuration using finite element analysis. The available at plate data consisted of three groups of tests, each with a unique charge density linear shape charge (LSC) used to cut an aluminum plate. The model was tuned to a representative test using the lowest charge density LSC as input. The correlated model was then used to predict the other two cases by linearly scaling the input load based on the relative difference in charge density. The resulting model predictions were then compared with available empirical data. Aside from differences in amplitude due to nonlinearities associated with scaling the charge density of the LSC, the model predictions matched the available test data reasonably well. Finally, modeling best practices were recommended when using industry standard software to predict shock response on structures. As part of the best practices documented, a frequency dependent damping schedule that can be used in model development when no data is available is provided.

Autism is a behavioral syndrome characterized by a severe impairment of reciprocal social relations, and of verbal and nonverbal communications. Many different etiologic factors such as viral infection and genetic predisposition have been proposed to explain the development of this disorder. Immune abnormalities, such as a decreased lymphoblastic response to T-cell mitogen, defective antibody responses to rubella vaccine, and decreased numbers of T lymphocytes, also have been identified in a subpopulation of patients with autism, which implies that the development of autism in some cases may be due to autoimmune mechanisms. Recent evidence suggests that immune response to the heat-shock proteins 60 and 70 is associated with several autoimmune diseases, including juvenile arthritis, type 1 diabetes, and multiple sclerosis. Therefore, in this study, the plasmas of patients with autism were examined by enzyme linked immunosorbent assay (ELISA) for antibodies to the heat-shock proteins 60 and 70. The autistic subjects were found to have increased levels of antibodies against heat-shock protein 70 as compared to that of age-matched controls (p=O. 0148). However, levels of antibodies to heat shock protein 60 in the autistic subjects showed considerable individual variation and no significant difference was found. Abnormal immune reactions to myelin basic protein have also been found in autistic subjects. Since epitopes on myelin basic protein have been shown to crossreact with determinants on heat-shock protein 60, the similarity between anti-myelin basic protein monoclonal antibodies and antiheat- shock protein 60 antibodies in the autistic subjects was also studied. The results showed no crossreactivity between these two antibodies. In conclusion, the data from the study of antibodies against heat-shock protein 70 suggest an elevated immune response to heat-shock protein 70 in autistic subjects. This result implies that autism could be an autoimmune disease.

Indiana University-Purdue University Indianapolis (IUPUI) / The ubiquitin-proteasome system is responsible for the targeted degradation of proteins within the cell. The 26S proteasome, which is the protease of this system, is a high molecular weight complex consisting of 33 subunits that arrange to form two smaller complexes the 19S regulatory particle (RP) and the 20S core particle (CP). The 19S RP can bind one or both ends of the 20S CP and is responsible for recognizing the ubiquitinated substrates. After recognition, the 19S RP will subsequently deubiquitinate, unfold, and translocate the substrates into the proteolytic 20S CP. The 20S CP consists of seven unique alpha and seven unique beta subunits that arrange into four stacked rings, with two alpha rings capping two beta rings. Assembly of the alpha(1-7)beta(1-7)beta(1-7)alpha(1-7) structure begins with the formation of an alpha ring and proceeds through specific assembly intermediates. This process is assisted by assembly chaperone proteins that promote on pathway interactions to efficiently construct the 20S CP. In this dissertation, three new findings are described which further characterize the proteasome assembly pathway. First, novel non-canonical complexes comprised of proteasome subunit alpha4 were identified in vivo, revealing proteasome subunits can assemble into complexes outside of the proteasome. Second, Hsp70 proteins, Ssa1/2, were shown to assist in the assembly of 20S CPs, adding to the growing list of proteins guiding proteasome assembly. Third, a novel complex was identified which is believed to represent a new proteasome assembly intermediate.

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In this study the mouse tumor necrosis factor-alpha (TNF-α) was prepared by stimulating macrophage cell line RAW 264.7 with lipopoly-saccharide (LPS) obtained from Escheria coli strain 055:B5.

In the fall of 2001, letters laced with anthrax spores were delivered to various news organizations in New York and Florida, as well as to two Senators in Washington, D.C. Over 22 anthrax infections and five deaths resulted from exposure to these spores, and decontamination of the affected buildings was both time consuming and costly. Since these attacks, interest in sporicidal disinfectants has increased greatly. Many chemical sporicidal disinfectants are available commercially, but the exposure time required to sterilize can be relatively long. In addition, some spores are simply injured or inhibited by chemical disinfectants, but not necessarily killed. Studies have shown that heat shocking spores after exposure to some disinfectants can aid in the recovery of injured spores, but these studies have not evaluated this effect on spores exposed to peracetic acid-based disinfectants. Recently, our lab has evaluated two novel peracetic acid-based chemical disinfectants, PeraDox™ and PeraDox Ultra™ for their activity against a variety of bacterial agents. Results indicated that the PeraDox™ solutions had extremely rapid cidal activity on a wide variety of microorganisms, especially those with innate germicide resistance, such as bacterial endospores. However, possible recovery of these spores after heat shock was not evaluated. The purpose of this study was to compare the sporicidal activity of three disinfectants: CIDEX™, PeraDox™, and PeraDox Ultra™ on three species of spores (Bacillus subtilis, Bacillus anthracis, and Clostridium sporogenes) in suspension, with and without heat shocking. Spores in suspension were exposed to disinfectants for specified times and assayed for viable spores. These spore suspensions were then heat shocked (80 ºC for 20 min) and assayed again. After exposure to peracetic acid-based disinfectants and subsequent heat shock, some B. subtilis spores recovered, resulting in up to a one log difference in viable spores. Other species and disinfectants did not show this effect. In addition, the activity of these disinfectants on spores dried onto a surface was evaluated using the standard AOAC sporicidal test. The current AOAC test specifies heat shocking after three weeks of incubation. In this study, we evaluated the AOAC sporicidal test by heat shocking immediately following disinfection and after three weeks of incubation as prescribed. Carrier tests showed a greater number of positive B. subtilis carriers when heat shocked immediately following PeraDox™, and PeraDox Ultra™ treatment, than when carriers were heat shocked after three weeks. In summary, results showed that heat shocking increases resuscitation of spores treated with some disinfectants, but not others. Spores in suspension and those dried onto carriers responded similarly to heat shocking. Finally, PeraDox™ formulations had surprisingly rapid sporicidal kinetics.