Immunomodulatory effects of alcohol use involve regulation of innate immune cell function leading to liver disease. Alteration of inflammatory responses by alcohol is linked to dysregulated TNF-alpha production. Alcohol-induced oxidative stress also contributes to alterations in inflammatory cell activity. Heat shock proteins (hsps) and the heat shock transcription factor-1 (HSF-1) induced by oxidative stress regulate NF-kappaB activation and TNF-alpha gene expression in monocytes and macrophages. Here, we report that in vitro alcohol treatment induced and augmented LPS-induced HSF-1 nuclear translocation and DNA-binding activity in monocytes and macrophages. Supershift analysis revealed that alcohol regulated HSF-1- and not HSF-2-binding activity. Hsp70, a target gene induced by HSF-1, was transiently increased within 24 h by alcohol, but extended alcohol exposure decreased hsp70 in macrophages. The alcohol-induced alteration of hsp70 correlated with a concomitant change in hsp70 promoter activity. Hsp90, another HSF-1 target gene, was decreased during short-term alcohol but increased after prolonged alcohol exposure. Decreased hsp90-HSF-1 complexes after short-term alcohol indicated dissociation of HSF-1 from hsp90. On the other hand, hsp90 interacted with client protein IkappaB kinase beta, a signaling intermediate of the LPS pathway, followed by IkappaBalpha degradation and increased NF-kappaB activity after chronic alcohol exposure, indicating that hsp90 plays an important role in supporting inflammatory cytokine production. Inhibition of hsp90 using geldanamycin prevented prolonged alcohol-induced elevation in LPS-induced NF-kappaB and TNF-alpha production. These results suggest that alcohol exposure differentially regulates hsp70 and hsp90 via HSF-1 activation. Further, hsp90 regulates TNF-alpha production in macrophages contributing to alcohol-induced inflammation.

Acute alcohol use is associated with impaired immune responses and decreased proinflammatory cytokine production. Our earlier studies have shown that acute alcohol intake inhibits NF-κB DNA binding in an IκBα-independent manner. We report using human peripheral blood monocytes and Chinese hamster ovary cells transfected with CD14 cells that acute alcohol treatment in vitro exerts NF-κB inhibition by disrupting phosphorylation of p65. Immunoprecipitation of p65 and IκBα revealed that acute alcohol exposure for 1 h decreased NF-κB-IκBα complexes in the cytoplasm. Phosphorylation of p65 at Ser536 is mediated by IκB kinase (IKK)β and is required for NF-κB-dependent cellular responses. We show that acute alcohol treatment decreased LPS-induced IKKα and IKKβ activity resulting in decreased phosphorylation of p65 at Ser536. Furthermore, nuclear expression of IKKα increased after alcohol treatment, which may contribute to inhibition of NF-κB. Decreased phosphorylation of nuclear p65 at Ser276 was likely not due to alcohol-induced inhibition of protein kinase A and mitogen- and stress-activated protein kinase-1 activity. Although decreased IκBα phosphorylation after acute alcohol treatment was attributable to reduced IKKβ activity, degradation of IκBα during alcohol exposure was IKKβ-independent. Alcohol-induced degradation of IκBα in the presence of a 26S proteasome inhibitor suggested proteasome-independent IκBα degradation. Collectively, our studies suggest that acute alcohol exposure modulates IκBα-independent NF-κB activity primarily by affecting phosphorylation of p65. These findings further implicate an important role for IKKβ in the acute effects of alcohol in immune cells.

New therapies for leukemia are urgently needed due to adverse side effects, tumor resistance and lack of selectivity of many chemotherapeutic agents in clinical use. Ageratum conyzoides has been used in folklore medicine for managing leukemia and other cancers. Thus, this study aimed to investigate the effects of fractions, sub-fractions and purified compounds from the ethanol leaf extracts of A. conyzoides against Jurkat cells-model for acute T cell leukemia. A two-dimensional purification process using normal phase flash, followed by reverse phase purification was necessary to isolate pure methoxy-flavones, which were further characterized by Nuclear Magnetic Resonance (NMR) and MS-MS. The effect of fractions or pure compounds on cell viability was determined using either the MTT reagent or CellTiter-Blue® assay, while the caspase-3 and -7 activation was tested with Caspase-Glo® 3/7 assay. Prediction of compounds' drug disposition profiles in vivo was measured with biomimetic affinity chromatography methodologies.

A soluble impure acetylcholinesterase (AChE), (EC 3.1.1. 7) was isolated from rat brain. The enzyme showed tendency to form aggregates in a medium of low ionic strength that leaded to a decrease of the acetylcholinesterase activity. After an increase of the protein concentration of the obtained soluble enzyme, was observed a decrease or absolute loss of the activity which regenerated after diluting with water. Aforeover detachment of the enzyme from the membrane (.<;o/ubilization) leaded up to raising of the total AChE activity as well as the activity of the residual membrane bound AChE. Possible reason might be the transition of the enzyme to more active form with lower degree of aggregation. The soluble AChE was further purified by ion exchange chromatography (FPLC), but the obtained enzyme was not stable enough and rapidly lost its activity in a high ionic strength environment.

No abstract