Calpain inhibitor I reduces the activation of nuclear factor-kappaB and organ injury/dysfunction in hemorrhagic shock.

There is limited evidence that inhibition of the activity of the cytosolic cysteine protease calpain reduces ischemia/reperfusion injury. The multiple organ injury associated with hemorrhagic shock is due at least in part to ischemia (during hemorrhage) and reperfusion (during resuscitation) of target organs. Here we investigate the effects of calpain inhibitor I on the organ injury (kidney, liver, pancreas, lung, intestine) and dysfunction (kidney) associated with hemorrhagic shock in the anesthetized rat. Hemorrhage and resuscitation with shed blood resulted in an increase in calpain activity (heart), activation of NF-kappaB (kidney), expression of iNOS and COX-2 (kidney), and the development of multiple organ injury and dysfunction, all of which were attenuated by calpain inhibitor I (10 mg/kg i.p.), administered 30 min prior to hemorrhage. Chymostatin, a serine protease inhibitor that does not prevent the activation of NF-kappaB, had no effect on the organ injury/failure caused by hemorrhagic shock. Pretreatment (for 1 h) of murine macrophages or rat aortic smooth muscle cells (activated with endotoxin) with calpain inhibitor I attenuated the binding of activated NF-kappaB to DNA and the degradation of IkappaBalpha, IkappaBbeta, and IkappaBvarepsilon. Selective inhibition of iNOS activity with L-NIL reduced the circulatory failure and liver injury, while selective inhibition of COX-2 activity with SC58635 reduced the renal dysfunction and liver injury caused by hemorrhagic shock. Thus, we provide evidence that the mechanisms by which calpain inhibitor I reduces the circulatory failure as well as the organ injury and dysfunction in hemorrhagic shock include 1) inhibition of calpain activity, 2) inhibition of the activation of NF-kappaB and thus prevention of the expression of NFkappaB-dependent genes, 3) prevention of the expression of iNOS, and 4) prevention of the expression of COX-2. Inhibition of calpain activity may represent a novel therapeutic approach for the therapy of hemorrhagic shock.     

A human NK cell activation/inhibition threshold allows small changes in the target cell surface phenotype to dramatically alter susceptibility to NK cells

NK cell activation is negatively regulated by the expression of target cell MHC class I molecules. We show that this relationship is nonlinear due to an NK cell activation/inhibition threshold. Ewing's sarcoma family tumor cell monolayers, which were highly susceptible to NK cells in vitro, developed a highly resistant phenotype when cultured as three-dimensional multicellular tumor spheroid structures. This suggested that tumor architecture is likely to influence the susceptibility to NK cells in vivo. Resistance of the multicellular tumor spheroid was associated with the increased expression of MHC class I molecules and greatly reduced NK cell activation, implying that a threshold of NK cell activation/inhibition had been crossed. Reducing MHC class I expression on Ewing's sarcoma family tumor monolayers did not alter their susceptibility to NK cells, whereas increased expression of MHC class I rendered them resistant and allowed the threshold point to be identified. This threshold, as defined by MHC class I expression, was predictive of the number of NK-resistant target cells within a population. A threshold permits modest changes in the target cell surface phenotype to profoundly alter the susceptibility to NK cells. Whereas this allows for the efficient detection of target cells, it also provides a route for pathogens and tumors to evade NK cell attack.     

Modelling viral and CD4 cellular population dynamics in HIV: approaches to evaluate intervention strategies.

Computational models, such as in epidemiology, provide a powerful tool that can be used to systematically examine an array of dynamic interactions among populations as well as to evaluate altemate disease intervention strategies. The specific objectives in this study were to: a/ examine the interaction of cellular (CD4) and HIV population dynamics and evaluate the impact of the use of combination chemotherapies on viral and CD4 populations (Experiment #1), b/ demonstrate how modelling can be used to evaluate the impact of an intervention (condom use) on reducing the rate of HIV/AIDS (Experiment #2). In this study, we used state transition models and conducted simulation experiments to evaluate various alternatives for the control and/or prevention of HIV/AIDS. The result indicated that combination therapy (double or triple drug therapies) was very effective. The HIV viral population decreased rapidly and remained suppressed for years. On the other hand, the CD4 cell population increased above 400 cells per ml and was maintained above that level for many years. Mono-therapy was not as effective; although the viral load decreased rapidly, it increased to its original levels within a few months. Since condom use is one of the key interventions of HIV/AIDS, we evaluated its use in 25%, 50% and 75% of an adult, sexually active population. Increasing condom use by 50% and 75% above an estimated baseline of 25% reduced the incidence of AIDS by 53% in Blacks, 49% in Hispanics and 43% in Whites. The study shows how a cellular/molecular level model can be incorporated within a macro-epidemiologic systems dynamics model to evaluate a variety of scientific questions such as to see if cellular/molecular level interventions reduce morbidity and mortality rates in HIV.     

Synthesis and activity against HBV of novel Tetra-seconucleoside analogues of dyphlline having the acyclic chains of ACV and HBG

Selective alkylation of dyphylline (1) with (2-acetoxyethoxy)methyl bromide (2a) or 4-acetoxybutyl bromide (2b) afforded 3'-O-[(acetoxyethoxy)methyl]dyphylline (3a) and 3'-O-(4-acetoxybutyl)-dyphylline (3b), respectively. A trans esterification process rather than alkylation of the dihydroxy-propyl side chain in 1 had taken place during the reaction with 2-p-toluoyloxy)ethyl chloride (5) to afford the respective 3'-toluoyloxy derivative 7 and not the anticipated 3'-O-[(p-toluoyloxy)ethyl]-dyphylline (6). Deacylation of 3a,b and 7 afforded 4a,b and 1, respectively. Viral screening of selected compounds against HBV has been investigated.