Chronic granulomatous disease: a review of the infectious and inflammatory complications

Chronic Granulomatous Disease is the most commonly encountered immunodeficiency involving the phagocyte, and is characterized by repeated infections with bacterial and fungal pathogens, as well as the formation of granulomas in tissue. The disease is the result of a disorder of the NADPH oxidase system, culminating in an inability of the phagocyte to generate superoxide, leading to the defective killing of pathogenic organisms. This can lead to infections with Staphylococcus aureus, Psedomonas species, Nocardia species, and fungi (such as Aspergillus species and Candida albicans). Involvement of vital or large organs can contribute to morbidity and/or mortality in the affected patients. Major advances have occurred in the diagnosis and treatment of this disease, with the potential for gene therapy or stem cell transplantation looming on the horizon.     

Modulation of ROS/MAPK signaling pathways by okadaic acid leads to cell death via,mitochondrial mediated caspase-dependent mechanism

Okadaic acid (OA) is a specific and potent protein phosphatase inhibitor and tumor promoter. The present study establishes the role of reactive oxygen species (ROS) and mitogen activated protein kinases in cell death induced by okadaic acid. The study showed that okadaic acid is cytotoxic at 10 nM with an IC50 of 100 nM in U-937 cells. The CVDE assay and mitochondrial dehydrogenase assay showed a time dependent cytotoxicity. The phase contrast visualization of the OA treated cells showed the apoptotic morphology and was confirmed with esterase staining for plasma membrane integrity. OA activated caspases-7, 9 and 3, PARP cleavage and induced nuclear damage in a time and dose dependent manner. Compromised mitochondrial membrane potential, release of cytochrome-c and apoptosis inducing factor confirms the involvement of mitochondria. A time dependent decrease in glutathione levels and a dose dependent increase in ROS with maximum at 30 min were observed. ROS scavenger-N-acetyl cysteine, mitochondrial stabilizer-cyclosporin-A, and broad spectrum caspase inhibitor Z-VAD-FMK inhibited the OA induced caspase-3 activation, DNA damage and cell death but caspase-8 inhibitor had no effect. OA activated p38 MAPK and JNK in a time dependent manner, but not ERK½. MAP kinase inhibitors SB203580, SP600125 and PD98059 confirm the role of p38 MAPK and JNK in OA induced caspase-3 activation and cell death. Over all, our results indicate that OA induces cell death by generation of ROS, and activation of p38 MAPK and JNK, and executed through mitochondrial mediated caspase pathway.     

Interaction of glycated human serum albumin with endothelial cells in a hemodynamic environment: structural and functional correlates

Advanced glycation end products (AGEs) are known to be involved in the pathogenesis of several diseases, in particular diabetes, via signaling through their receptor. Numerous studies have been carried out on protein-sugar interactions at very high concentrations of the latter. The objective of this investigation was to determine the effects of nonenzymatic glycation induced by reducing sugars on the secondary structure of human serum albumin (HSA) under different physiological conditions and to correlate that with expression of RAGE (receptor for advanced glycation end products) on HUVECs (human umbilical vein endothelial cells) in a controlled hemodynamic environment. Our results indicate that RAGE expression is shear stress modulated and that glycated HSA enhances the expression further. The secondary structure of AGE-HSA derived from glucose at 20 mM contains higher α-helical content and elicits maximum expression of the receptor. The effect of shear stress at 10 dynes cm(-2) is independent of AGE-HSA.     

IL-1β promotes TGF-β1 and IL-2 dependent Foxp3 expression in regulatory T cells

Earlier, we have shown that GM-CSF-exposed CD8α- DCs that express low levels of pro-inflammatory cytokines IL-12 and IL-1β can induce Foxp3+ Tregs leading to suppression of autoimmunity. Here, we examined the differential effects of IL-12 and IL-1β on Foxp3 expression in T cells when activated in the presence and absence of DCs. Exogenous IL-12 abolished, but IL-1β enhanced, the ability of GM-CSF-exposed tolerogenic DCs to promote Foxp3 expression. Pre-exposure of DCs to IL-1β and IL-12 had only a modest effect on Foxp3- expressing T cells; however, T cells activated in the absence of DCs but in the presence of IL-1β or IL-12 showed highly significant increase and decrease in Foxp3+ T cell frequencies respectively suggesting direct effects of these cytokines on T cells and a role for IL-1β in promoting Foxp3 expression. Importantly, purified CD4+CD25+ cells showed a significantly higher ability to maintain Foxp3 expression when activated in the presence of IL-1β. Further analyses showed that the ability of IL-1β to maintain Foxp3 expression in CD25+ T cells was dependent on TGF-β1 and IL-2 expression in Foxp3+Tregs and CD25- effectors T cells respectively. Exposure of CD4+CD25+ T cells to IL-1β enhanced their ability to suppress effector T cell response in vitro and ongoing experimental autoimmune thyroidits in vivo. These results show that IL-1β can help enhance/maintain Tregs, which may play an important role in maintaining peripheral tolerance during inflammation to prevent and/or suppress autoimmunity.     

Exogenous supply of pantoyl lactone to excised leaves increases their pantothenate levels

BACKGROUND AND AIMS: All plants synthesize pantothenate but its synthesis and regulation are not well understood. The aim of this work is to study the effect of exogenous supply of precursor compounds on pantothenate levels in leaves. METHODS: Precursor compounds were supplied in solution to excised leaves and the pantothenate content was measured using a microbial method. KEY RESULTS: Pantothenate levels in excised leaves of Limonium latifolium, tomato (Lycopersicon esculentum), bean (Phaseolus vulgaris) and grapefruit (Citrus x paradisi) were examined following an exogenous supply of the precursor compounds pantoyl lactone or beta-alanine. Significantly higher levels of extractable pantothenate were found when pantoyl lactone was supplied, but not when beta-alanine was supplied despite a measurable uptake of beta-alanine into the leaf. CONCLUSIONS: The results suggested that the pantoate supply may be rate-limiting or regulating pantothenate synthesis in leaves.     

Evidence that protein kinase Calpha interacts with and regulates the glial glutamate transporter GLT-1

Many of the sodium‐dependent neurotransmitter transporters are rapidly (within minutes) regulated by protein kinase C (PKC), with changes in activity being correlated with changes in transporter trafficking to or from the plasma membrane. Our recent studies suggest that one of the classical subtypes of PKC, PKCα, may selectively mediate redistribution of the neuronal glutamate transporter, excitatory amino acid carrier (EAAC)1, and show that PKCα can be co‐immunoprecipitated with EAAC1. When the glial glutamate transporter GLT‐1a is transfected into C6 glioma cells, this transporter is internalized in response to activation of PKC, but the PKC subtype involved in this regulation is unknown. In the present study, expression of the phorbol ester‐activated subtypes of PKC was examined in C6 glioma transfected with GLT‐1. Of the classical subtypes, only PKCα was detected, and of the non‐classical subtypes, PKCδ and PKCε were detected. In this system, phorbol ester‐dependent internalization of GLT‐1 was blocked by a general inhibitor of PKCs (bisindolylmaleimide II) and by concentrations of Gö6976 that selectively block classical PKCs, but not by an inhibitor of PKCδ (rottlerin). PKCα immunoreactivity was found in GLT‐1 immunoprecipitates obtained from transfected C6 cells and from crude rat brain synaptosomes, a milieu that better mimics in vivo conditions. The amount of PKCα in both types of immunoprecipitate was modestly increased by phorbol ester, and this increase was blocked by a PKC antagonist. These studies suggest that PKCα may be required for the regulated redistribution of GLT‐1.     

2-(Aryl)-3-furan-2-ylmethyl-thiazolidin-4-ones as selective HIV-RT inhibitors

A series of 4-thiazolidinones were evaluated as selective inhibitors of the HIV-RT enzyme. Our attempt in correlating the derived physicochemical properties with the HIV-RT inhibitory activity resulted in some statistically significant QSAR models with good predictive ability. The QSAR studies indicated the role of lipophilicity, dipole moment and out-of-plane potential energy of the compounds in rationalizing the activity. One of the compounds, 1, inhibited the enzyme at 0.204 μM concentration with minimal toxicity to MT-4 cells.     

A directed screen for chlamydial proteins secreted by a type III mechanism identifies a translocated protein and numerous other new candidates

Chlamydiae are strict intracellular parasites that induce their internalization upon contact with the host cell and grow inside an intracellular compartment called an inclusion. They possess a type III secretion (TTS) apparatus, which allows for the translocation of specific proteins in the host cell cytosol. In particular, chlamydial proteins of the Inc family are secreted to the inclusion membrane by a TTS mechanism; other TTS substrates are mostly unknown. Using a secretion assay based on the recognition of TTS signals in Shigella flexneri, we searched for TTS signals in the proteins of unknown function, conserved between three different chlamydial species, Chlamydia pneumoniae, C. trachomatis and C. caviae. We identified 24 new candidate proteins which did not belong to the Inc family. Four of these proteins were also secreted as full-length proteins by a TTS mechanism in S. flexneri, indicating that their translocation does not require other chlamydial proteins. One of these proteins was detected in the cytosol of infected cells using specific antibodies, directly demonstrating that it is translocated in the host cell during bacterial proliferation. More generally, this work represents the first directed search for TTS effectors not based on genetic information or sequence similarity. It reveals the abundance of proteins secreted in the host cell by chlamydiae.     

5-Amidinobenzo[b]thiophenes as dual inhibitors of factors IXa and Xa

Syntheses and SAR studies of 5-amidinobenzo[b]thiophene analogs provided compounds with low submicromolar factor IXa activity and equal or slightly better selectivity relative to factor Xa.