SIMPL is a tumor necrosis factor-specific regulator of nuclear factor-kappaB activity.

The IL-1 receptor-associated kinase (IRAK/mPLK) is linked to the regulation of nuclear factor-kappaB (NF-kappaB)-dependent gene expression. Here we describe a novel binding partner of IRAK/mPLK that we term SIMPL (signaling molecule that associates with the mouse pelle-like kinase). Overexpression of SIMPL leads to the activation of NF-kappaB-dependent promoters, and inactivation of SIMPL inhibits IRAK/mPLK as well as tumor necrosis factor receptor type I-induced NF-kappaB activity. Dominant inhibitory alleles of IkappaB kinase (IKKalpha or IKKbeta) block the activation of NF-kappaB by IRAK/mPLK and SIMPL. Furthermore, SIMPL binds IRAK/mPLK and the IKKs in vitro and in vivo. In the presence of antisense mRNA to SIMPL, the physical association between IRAK/mPLK and IKKbeta but not IRAK/mPLK and IKKalpha is greatly diminished. Moreover, dominant-negative SIMPL blocks IKKalpha- or IKKbeta-induced NF-kappaB activity. These results lead us to propose a model in which SIMPL functions to regulate NF-kappaB activity by linking IRAK/mPLK to IKKbeta/alpha-containing complexes.

Lead toxicity in older adults.

Recent studies have shown that lead, even at relatively low levels of exposure, has the potential to harm not only the young and the occupationally-exposed, but also older people. Because they have been alive for a longer period of time, older adults have had more potential exposures to lead. They may have been exposed to lead while working in unregulated occupations, or they may have encountered more lead in the environment on a daily basis. Several large epidemiological studies have found that older people have higher blood and bone lead levels than younger adults. Additionally, sporadic clusters of acute lead exposure among older adults as a result of activities such as ceramic glaze hobby work and consumption of moonshine whiskey continue to be reported. After lead enters the body, it circulates in the blood reaching the soft tissues and bone. Researchers have learned that lead can hibernate within bone for decades. Although lead within bone is of uncertain toxicity to bone tissue, conditions of bone resorption, such as osteoporosis, can cause bone lead to reenter the bloodstream where it can then re-expose the soft tissue, and, potentially, exert delayed deleterious effects. Evidence is emerging that blood and bone lead levels, reflecting relatively modest exposures, are associated with hypertension, renal insufficiency, and cognitive impairment. Medical treatments that now exist to slow the rate of bone resorption may maintain lead within bones. On-going studies evaluating the relationship between body lead stores and both cognitive and renal impairment, as well as the potential modifying effect of bone resorption, will help determine whether bone resorption should be retarded specifically to preserve organ function. Physicians should be aware of potential past and present lead exposures among their older patients. Ongoing lead exposure should be prevented. In the future, treatment of osteoporosis may be undertaken not only to improve bone health but also to prevent mobilization of bone lead stores and subsequent toxicity.

Repression of transforming-growth-factor-beta-mediated transcription by nuclear factor kappaB.

Activation of transforming growth factor-beta (TGF-beta) and activin receptors leads to phosphorylation of Sma- and Mad-related protein 2 (Smad2) and Smad3, which function as transcription factors to regulate gene expression. Smad7 is a regulatory protein which is able to inhibit TGF-beta and activin signalling in a negative-feedback loop, mediated by a direct regulation by Smad3 and Smad4 via a Smad-binding element (SBE) in the Smad7 promoter. Interestingly, we found that the Smad7 promoter was also regulated by nuclear factor kappaB (NF-kappaB), a transcription factor which plays an important role in inflammation and the immune response. Expression of NF-kappaB p65 subunit was able to inhibit the Smad7 promoter activity, and this inhibition could be reversed by co-expression of IkappaB, an inhibitor of NF-kappaB. In addition, the inhibitory activity of p65 was observed in a minimal promoter that contained only the Smad7 SBE and a TATA box, without any consensus NF-kappaB binding site. This inhibitory effect appeared to be common to other TGF-beta- and activin-responsive promoters, since p65 also inhibited the forkhead-activin-signal-transducer-2-mediated activation of a Xenopus Mix.2 promoter, as well as the Smad3-mediated activation of 3TP-lux which contains PMA-responsive elements and a plasminogen-activator-inhibitor-1 promoter. Activation of endogenous NF-kappaB by tumour necrosis factor-alpha (TNF-alpha) was also able to inhibit the Smad7 promoter in human embryonic kidney 293 cells. In human hepatoma HepG2 cells, TNF-alpha was able to inhibit TGF-beta- and activin-mediated transcriptional activation. Furthermore, overexpression of the transcription co-activator p300 could abrogate the inhibitory effect of NF-kappaB on the Smad7 promoter. Taken together, these data have indicated a novel mode of crosstalk between the Smad and the NF-kappaB signalling cascades at the transcriptional level by competing for a limiting pool of transcription co-activators.

[Rheographic monitoring of intraoperative hemodynamics]. / Reograficheskii monitoring intraoperatsionnoi gemodinamiki.

The authors summed up the experiences with using noninvasive monitoring of hemodynamics during 420 operations. The method was based on computed procession in the real time of the integral rheogram of the body. The advantages, shortcomings and technological details of the method were analyzed.

Modulation of tumor necrosis factor and interleukin-1-dependent NF-kappaB activity by mPLK/IRAK.

The innate immune response is an important defense against pathogenic agents. A component of this response is the NF-kappaB-dependent activation of genes encoding inflammatory cytokines such as interleukin-8 (IL-8) and cell adhesion molecules like E-selectin. Members of the serine/threonine innate immune kinase family of proteins have been proposed to mediate the innate immune response. One serine/threonine innate immune kinase family member, the mouse Pelle-like kinase/human interleukin-1 receptor-associated kinase (mPLK/IRAK), has been proposed to play an obligate role in promoting IL-1-mediated inflammation. However, it is currently unknown whether mPLK/IRAK catalytic activity is required for IL-1-dependent NF-kappaB activation. The present study demonstrates that mPLK/IRAK catalytic activity is not required for IL-1-mediated activation of an NF-kappaB-dependent signal. Intriguingly, catalytically inactive mPLK/IRAK inhibits type 1 tumor necrosis factor (TNF) receptor-dependent NF-kappaB activation. The pathway through which mPLK/IRAK mediates this TNF response is TRADD- and TRAF2-independent. Our data suggest that in addition to its role in IL-1 signaling, mPLK/IRAK is a component of a novel signal transduction pathway through which TNF R1 activates NF-kappaB-dependent gene expression.

Coordinate regulation of glucocorticoid receptor and c-jun gene expression is cell type-specific and exhibits differential hormonal sensitivity for down- and up-regulation.

We have previously proposed a novel mechanism for the coupled regulation of glucocorticoid receptor (GR) and c-jun transcription in triamcinolone acetonide (TA)-treated AtT-20 cells. This involved transcriptional interference of AP-1 (Fos/Jun)-driven gene transcription by the formation of inactive GR/Jun heterodimers. To further elucidate the molecular mechanism for GR autoregulation, the expression of GR and c-jun mRNA and protein levels were examined in both mouse L929 fibroblast cells and human CEM-C7 acute lymphoblastic leukemia cells. A rapid down-regulation of both GR and c-jun mRNA and protein levels occurs in TA-treated L929 cells. All-trans-retinoic acid (RA) treatment of Jun-deficient, mouse F9, teratocarcinoma cells causes the induction of c-jun expression. The increased expression of both c-jun mRNA and protein is accompanied by the induction of GR expression. These data further suggest that functional cJun is needed for the expression of the GR and c-jun genes in F9 cells. CEM-C7 cells undergo apoptosis after exposure to glucocorticoids. There is a parallel up-regulation of GR and c-jun mRNA levels in TA-treated CEM-C7 cells. This is accompanied by a concomitant increase in GR and cJun protein levels. Dose-response analyses reveal the expected coordinate regulation of both GR and c-jun mRNA and protein in L929 cells (decreasing) and in CEM-C7 cells (increasing). However, approximately 20-fold less TA is required for the inhibition of GR and c-jun expression as compared to that required for the stimulation of these two genes. These data demonstrate that the coordinate regulation of GR and c-jun gene expression is dose-dependent and cell type-specific. These results, along with previously reported data, suggest that GR complex formation with itself or with another transcription factor is important for the coordinate up- and down-regulation, respectively, of the GR and c-jun genes.

Coordinate regulation of glucocorticoid receptor and c-jun mRNA levels: evidence for cross-talk between two signaling pathways' at the transcriptional level.

Autologous regulation of steroid receptors by their cognate ligands has been demonstrated for a number of nuclear receptor family members. To determine the molecular mechanism for glucocorticoid receptor (GR) autoregulation, the expression of glucocorticoid receptor mRNA and protein levels were examined in the mouse AtT-20 pituitary tumor cell line. The expression of c-jun and c-fos mRNA and protein was also examined in the same cell extracts. A rapid down-regulation of the GR protein was observed after treatment with the glucocorticoid analog, triamcinolone acetonide (TA). An oscillatory, parallel regulation of both GR and c-jun mRNA levels occurred. In contrast, POMC mRNA levels remained at a stable, low level during chronic TA treatment. Dose-response analyses also revealed a coordinate down-regulation of GR and c-jun (but not POMC or c-fos) mRNA levels. FOS protein levels were unaffected by TA treatment. Surprisingly, JUN protein levels were increased by TA, even when the c-jun mRNA levels were decreasing. Perhaps a derepression of c-jun mRNA translation occurs after TA treatment, and this may be due to GR/JUN heteromer formation interfering with JUN repression of c-jun mRNA translation. The effect of TA on GR and c-jun gene expression was a primary effect, as it occurred rapidly and was not inhibited by cycloheximide (CHX). Nuclear run-on transcription assays revealed a rapid (15 min) down-regulation in both GR and c-jun gene transcription rates, while POMC gene transcription was unaffected at this early time. Treatment of AtT-20 cells with all-trans retinoic acid gave different kinetics for GR and c-jun mRNA regulation than obtained with TA; however, the GR and c-jun mRNA levels were still coordinately regulated after retinoic acid treatment. Based upon these data, the promoter structures of the GR and c-jun genes, and previously published results, a novel mechanism for the coupled regulation of GR and c-jun transcription, via a direct transcriptional interference with AP-1 (FOS/JUN) activity, is proposed.

Reduced chloramphenicol acetyltransferase activity observed with vectors containing an upstream SphI recognition sequence.

Chloramphenicol acetyltransferase (CAT) is the most commonly used reporter gene for studying the regulation of mammalian gene transcription. Some of the currently available CAT vectors contain the recognition sequence for the restriction endonuclease SphI within the multiple cloning site. This sequence introduces an ATG triplet that is out of frame with the initiation codon of the CAT gene. Transient expression of CAT fusion genes, constructed using three different cellular promoters, demonstrates that the presence of the upstream AUG triplet in the CAT transcript reduces CAT activity, presumably by interfering with the translation of the coding sequence. Deletion of the SphI site from each of the plasmids increased CAT activity between 4-fold and 5-fold. From these results, we conclude that upstream, out-of-frame ATG triplets must be avoided in order to achieve maximum expression of the reporter gene.

Purification and characterization of Cu,Zn-superoxide dismutase from common carp liver.

1. Common carp (Cyprinus carpio L.) liver Cu,Zn-superoxide dismutase (Cu,Zn-SOD) was purified and characterized. 2. Its molecular weight, isoelectric point, electrophoretic mobility, amino acid pattern and some other characteristics were determined.

The effect of an organophosphorus pesticide on the enzymes of carp (Cyprinus carpio L.).

1. The effect of 1.5, 2.0 and 5.0 ppm methidation was studied on the ASAT, ALAT, LDH and AChE enzymes, as well as the blood sugar and adrenaline levels in carp. 2. According to our studies methidation implies a potential hazard on the normal biochemical processes of fishes: causing tissue necrosis indicated by the increase ASAT, ALAT and LDH activities, inducing continual stress effect reflected by the increased blood sugar and adrenaline levels, inhibiting AChE activity in the various organs, the consequence under acute effects being fish kill.

Localization and quantitative distribution of biogenic monoamines in the intestinal tract of locust, snail and carp.

The localization and quantitative distribution of the biogenic monoamines of the intestinal tract has been studied in Locusta migratoria, Helix pomatia and Cyprinus carpio with morphological and biochemical methods. Electron microscopically dense-core vesicles of aminergic character were found in the varicose nerve fibres located in the intestinal muscles of all three animal species. Intensive green fluorescence characteristic of catecholamines was detectable in both the varicose nerve fibres and perikarya. Using a fluorimetric method the quantity of biogenic monoamines was measured in the gastrointestinal tract of all three species. The amount of adrenaline and noradrenaline was rather low: 0.03-0.29 ?g/g. The dopamine content reached the value of 6 ?g/g in the locust gut, and a significant amount of serotonin was present in the intestinal tract of all three species: 1.00-2.59 ?g/g. Compared to the adrenaline and noradrenaline serotonin proved to be higher by more than one order in each case. On the basis of morphological and biochemical results the authors suggest that biogenic monoamines are involved in the regulation of the gut muscle functioning both in the form of transmitters as well as neurohormones.