Inhibitory effects of the beta-adrenergic receptor agonist zilpaterol on the LPS-induced production of TNF-alpha in vitro and in vivo.

In this study the anti-inflammatory properties of zilpaterol, a beta2-adrenergic receptor (AR) agonist specifically developed as a growth promoter in cattle were investigated. Although zilpaterol has a different structure compared with the beta2-AR agonists known to date, it was noted that it was able to bind to both the beta2-AR (Ki = 1.1 x 10(-6)) and the beta1-AR (Ki = 1.0 x 10(-5)). Using lipopolysaccharide (LPS)-exposed U937 macrophages, the production of cyclic adenosine-3',5'-cyclic monophosphate (cAMP) and tumor necrosis factor alpha (TNF-alpha) were investigated. Zilpaterol inhibited TNF-alpha release and induced intracellular cAMP levels in a dose-dependent manner. The inhibition of TNF-alpha release and induction of cAMP production was mainly mediated via the beta2-AR, as indicated by addition of beta1- and beta2-specific antagonists. The effects of zilpaterol were investigated in LPS-treated male Wistar rats after pretreatment with zilpaterol. Zilpaterol dosed at 500 microg/kg body weight reduced the TNF-alpha plasma levels. In conclusion, zilpaterol is a beta2-adrenergic agonist and an inhibitor of TNF-alpha production induced by LPS both in vivo and in vitro.

Do aberrant crypt foci have predictive value for the occurrence of colorectal tumours? Potential of gene expression profiling in tumours.

The effects of different dietary compounds on the formation of aberrant crypt foci (ACF) and colorectal tumours and on the expression of a selection of genes were studied in rats. Azoxymethane-treated male F344 rats were fed either a control diet or a diet containing 10% wheat bran (WB), 0.2% curcumin (CUR), 4% rutin (RUT) or 0.04% benzyl isothiocyanate (BIT) for 8 months. ACF were counted after 7, 15 and 26 weeks. Tumours were scored after 26 weeks and 8 months. We found that the WB and CUR diets inhibited the development of colorectal tumours. In contrast, the RUT and BIT diets rather enhanced (although not statistically significantly) colorectal carcinogenesis. In addition, the various compounds caused different effects on the development of ACF. In most cases the number or size of ACF was not predictive for the ultimate tumour yield. The expression of some tumour-related genes was significantly different in tumours from the control group as compared to tumours from the treated groups. It was concluded that WB and CUR, as opposed to RUT and BIT, protects against colorectal cancer and that ACF are unsuitable as biomarker for colorectal cancer. Effects of the different dietary compounds on metalloproteinase 1 (TIMP-1) expression correlated well with the effects of the dietary compounds on the ultimate tumour yield.

Protein kinase Czeta is a negative regulator of protein kinase B activity.

Protein kinase B (PKB), also known as Akt or RAC-PK, is a serine/threonine kinase that can be activated by growth factors via phosphatidylinositol 3-kinase. In this article we show that PKCzeta but not PKCalpha and PKCdelta can co-immunoprecipitate PKB from CHO cell lysates. Association of PKB with PKCzeta was also found in COS-1 cells transiently expressing PKB and PKCzeta, and moreover we found that this association is mediated by the AH domain of PKB. Stimulation of COS-1 cells with platelet-derived growth factor (PDGF) resulted in a decrease in the PKB-PKCzeta interaction. The use of kinase-inactive mutants of both kinases revealed that dissociation of the complex depends upon PKB activity. Analysis of the activities of the interacting kinases showed that PDGF-induced activation of PKCzeta was not affected by co-expression of PKB. However, both PDGF- and p110-CAAX-induced activation of PKB were significantly abolished in cells co-expressing PKCzeta. In contrast, co-expression of a kinase-dead PKCzeta mutant showed an increased induction of PKB activity upon PDGF treatment. Downstream signaling of PKB, such as the inhibition of glycogen synthase kinase-3, was also reduced by co-expression of PKCzeta. A clear inhibitory effect of PKCzeta was found on the constitutively active double PKB mutant (T308D/S473D). In summary, our results demonstrate that PKB interacts with PKCzeta in vivo and that PKCzeta acts as a negative regulator of PKB.

Melanocortin receptors mediate alpha-MSH-induced stimulation of neurite outgrowth in neuro 2A cells.

Melanocortins (MC), neuropeptides derived from pro-opiomelanocortin, have been implicated in enhancing neurite outgrowth via an as yet unknown mechanism. Recently, five MC receptors have been identified, three of which, the MC3-R, the MC4-R and the MC5-R, are expressed in the nervous system. In this study, alpha-MSH and the melanocortin analog [D-Phe7]ACTH (4-10) were able to stimulate neurite outgrowth in the neuroblastoma cell line Neuro 2A. ACTH (4-10), gamma2-MSH and ORG2766 were inactive. In addition, the MC4-R antagonist [D-Arg8]ACTH (4-10), inhibited the alpha-MSH effect, indicating that the MC4-R mediated stimulation of neurite outgrowth by alpha-MSH. Indeed, the presence of MC4-R mRNA in Neuro 2A cells was demonstrated by a RNase protection assay. Heterologous expression of the MC5-R in Neuro 2A cells lead to the recruitment of a responsiveness to gamma2-MSH, but did not increase the effect of alpha-MSH on neurite outgrowth. This finding indicated that the function of MC4-R can also be exerted by another MC receptor, suggesting that the coupling to Gs, which they have in common, plays an essential role in the neurite outgrowth promoting effect. This was further substantiated by the fact that forskolin treatment per se induced neurite outgrowth in a similar fashion. These data imply that the neurotrophic properties of alpha-MSH are likely to result from Gs-coupled MC receptor activity in neuronal cells.