Magdalena Dudkowska

Agmatine modulates the in vivo biosynthesis and interconversion of polyamines and cell proliferation.

Agmatine has recently gained wide interest as a bioactive arginine metabolite with a multitude of physiological functions. This study evaluates the in vivo role of agmatine in the modulation of metabolism and intracellular level of polyamines. Here, we report that agmatine, administered to mice, differentially affects the renal and liver activity of the two key enzymes regulating polyamine biosynthesis and interconversion/degradation. Thus, agmatine exerts a negative regulation of ODC activity and protein content, and positive regulation of SSAT activity, having no effect on ODC and SSAT transcript level. Agmatine modulation of ODC and SSAT activities is noticeably augmented by the inhibitor of its catabolism, aminoguanidine. Antizyme and eIF4E protein content appears to be affected by agmatine only insignificantly and apparently do not contribute to agmatine-induced down-regulation of ODC content. The homeostasis of spermidine and spermine is preserved after agmatine injection, while the putrescine level decreases. Furthermore, when tested in a mouse kidney injury model, agmatine, partially but significantly, reduces [3H] thymidine incorporation into DNA. This is consistent with suppressed renal tubule epithelial cell proliferation. The findings provide in vivo evidence of a substantial role of agmatine as a modulator of polyamine biosynthesis and degradation and suggest its suppressive effect on cell proliferation.

Cross-talk between steroid-receptor-mediated and cell-membrane-receptor-mediated signalling pathways results in the in vivo modulation of c-Met and ornithine decarboxylase gene expression in mouse kidney

The cross-talk in vivo between two signalling pathways activated by testosterone via intracellular androgen receptor, and induced by damage to renal tubules evoked by anti-folate [N(10)-propargyl-5,8-dideazafolic acid (CB 3717)] or folate is reported. We show that CB 3717/folate induces the expression of the hepatocyte growth factor (HGF)/c-Met signalling system in injured kidneys in which a significant, but transient, elevation of the HGF mRNA level occurs. It is followed by a severalfold increase in the c-Met transmembrane receptor message that persists for up to 24 h. The c-Met expression is also positively controlled by testosterone, which induces a significant increase in its mRNA level that is abolished by an anti-androgen, casodex. However, when testosterone and anti-folate/folate are administered sequentially, a substantial (3.5-4.0-fold) decrease in the increase of c-Met expression caused by CB 3717/folate alone occurs. Similarly, testosterone-induced ornithine decarboxylase (ODC) mRNA level and activity are decreased 2.8-7.7-fold when the androgen is applied together with CB 3717. Antagonism between these pathways is also visible under physiological conditions in the kidneys of male mice in which, owing to elevated endogenous testosterone levels, neither the ODC activity nor the mRNA level is induced by anti-folate/folate, whereas the c-Met message response to these drugs is significantly decreased. Our results document a substantial negative regulation of c-Met and ODC gene expression as a result of the cross-talk between testosterone-activated and HGF-activated pathways and suggest a sex-differentiated response to injury of mouse kidneys.

Paradoxical action of growth factors: antiproliferative and proapoptotic signaling by HGF/c-MET

Hepatocyte growth factor (HGF)/mesenchymal–epithelial transition factor (c-MET) signaling is usually associated with the promotion of cellular growth and often with progression of tumors. Nevertheless, under certain conditions HGF can also act as an antiproliferative and proapoptotic factor and can sensitize various cancer cells, treated with anticancer drugs, to apoptosis. Not only HGF but also its various truncated forms as well as intracellular fragments of its membrane receptor, c-MET, may act as antiproliferative and proapoptotic factors toward various cells. This review focuses on different mechanisms responsible for such paradoxical action of the known typical growth factor. It also points toward the possibilities of usage of this information in anticancer therapy.

Catecholamines participate in the induction of ornithine decarboxylase gene expression in normal and hyperplastic mouse kidney

In the quinazoline antifolate (CB 3717)-induced hyperplastic kidney model, a remarkable increase of ornithine decarboxylase (ODC) activity was paralleled by a smaller, but highly significant augmentation of the ODC transcript level. Catecholamine depletion, evoked by reserpine, strongly impaired antifolate-induced ODC expression; the enzyme activity was almost completely abolished while the mRNA level decreased by 60%. Moreover, under conditions of a depleted catecholamine pool, kidney enlargement was significantly reduced confirming our earlier reports on the indispensability of ODC induction for renal hyperplasia (M. Manteuffel-Cymborowska et al. , Biochim. Biophys. Acta, 1182 (1993) 133-141[1]). In normal mouse kidney catecholamines appeared to be inducers of ODC expression. Use of selective agonists of catecholamine receptors demonstrated the importance of dopamine D2 receptors, and to a lower extent beta adrenoreceptors, in the catecholamine mediation of induction of ODC activity and of ODC mRNA levels. These increases were not abolished by an antiandrogen, casodex, suggesting that catecholamine control of ODC expression is an androgen receptor-independent process. The results obtained point to the critical role of renal catecholamines; these biogenic amines are not only involved in the regulation of ODC expression in normal kidney but are also required for the induction of ODC in hyperplastic kidney evoked by antifolate and, as shown recently (M. Manteuffel-Cymborowska et al., Biochim. Biophys. Acta, 1356 (1997) 292-298[2]), in testosterone-induced hypertrophic kidney.

Antifolate/folate-activated HGF/c-Met signalling pathways in mouse kidneys-the putative role of their downstream effectors in cross-talk with androgen receptor.

This in vivo study of mouse kidneys was focused on the identification of protein mediators involved in the cross-talk between two signalling pathways. One pathway was triggered by testosterone via an androgen receptor, AR, and the other induced by CB 3717/folate via HGF, and its membrane receptor c-Met. Sequential activation of these pathways leads to a drastic decrease of testosterone-induced ornithine decarboxylase, ODC, expression. We proved that CB 3717/folate-induced ODC expression is Akt-dependent. CB 3717/folate activates Akt and ERK1/2 kinases, PTEN phosphatase and also up-regulates cyclin D2 and PCNA, but decreases GSK3beta and cyclin D1 protein levels. Testosterone activation of AR induces GSK3beta and PTEN. Results of the sequential activation of the studied signalling pathways suggest that Akt, GSK3beta and possibly ERK1/2 kinases may participate in the negative cross-talk and attenuation of AR transactivity, while the involvement of PTEN and cyclin D1 seems to be doubtful.

Nuclear and membrane receptor-mediated signalling pathways modulate polyamine biosynthesis and interconversion

Polyamines play an important role in cell growth and differentiation, while their overproduction has potentially oncogenic consequences. Polyamine homoeostasis, a critical determinant of cell fate, is precisely tuned at the level of biosynthesis, degradation and transport. The enzymes ODC (ornithine decarboxylase), AdoMetDC (S-adenosylmethionine decarboxylase) and SSAT (spermidine/spermine N(1)-acetyltransferase) are critical for polyamine pool maintenance. Our experiments were designed to examine the expression of these enzymes in testosterone-induced hypertrophic and antifolate-induced hyperplastic mouse kidney, characterized by activation of AR (androgen receptor) and HGF (hepatocyte growth factor) membrane receptor c-Met respectively. The expression of these key enzymes was up-regulated by antifolate CB 3717 injury-evoked activation of HGF/c-Met signalling. In contrast, activation of the testosterone/AR pathway remarkably induced a selective increase in ODC expression without affecting other enzymes. Studies in catecholamine-depleted kidneys point to a synergistic interaction between the signalling pathways activated via cell membrane catecholamine receptors and AR, as well as c-Met. We found that this cross-talk modulated the expression of ODC and AdoMetDC, enzymes limiting polyamine biosynthesis, but not SSAT. This is in contrast with the antagonistic cross-talk between AR- and c-Met-mediated signalling which negatively regulated the expression of ODC, but affected neither AdoMetDC nor SSAT.

Simultaneous induction and blockade of autophagy by a single agent

Besides cell death, autophagy and cell senescence are the main outcomes of anticancer treatment. We demonstrate that tacrine-melatonin heterodimer C10, a potent anti-Alzheimer’s disease drug, has an antiproliferative effect on MCF-7 breast cancer cells. The main cell response to a 24 h-treatment with C10 was autophagy enhancement accompanied by inhibition of mTOR and AKT pathways. Significantly increased autophagy markers, such as LC3B- and ATG16L-positive vesicles, confirmed autophagy induction by C10. However, analysis of autophagic flux using mCherry-GFP-LC3B construct revealed inhibition of autophagy by C10 at the late-stage. Moreover, electron microscopy and analysis of colocalization of LC3B and LAMP-1 proteins provided evidence of autophagosome-lysosome fusion with concomitant inhibition of autolysosomal degradation function. After transient treatment with IC50 dose of C10 followed by cell culture without the drug, 20% of MCF-7 cells displayed markers of senescence. On the other hand, permanent cell treatment with C10 resulted in massive cell death on the 5th or 6th day. Recently, an approach whereby autophagy is induced by one compound and simultaneously blocked by the use of another one has been proposed as a novel anticancer strategy. We demonstrate that the same effect may be achieved using a single agent, C10. Our findings offer a new, promising strategy for anticancer treatment.

Erratum to: The role of gender and labour status in immunosenescence of 65+ Polish population

In the original publication, the affiliations of fourth and fifth authors have been incorrectly published. The correct affiliations are given below: K. Broczek Department of Geriatrics, Medical University of Warsaw, Oczki Str. 4, 02-007 Warsaw, Poland e-mail: kbroczek@gmail.com M. Dabrowski Laboratory of Bioinformatics, Nencki Institute of Experimental Biology of Polish Academy of Sciences, Pasteur Str. 3, 02-093 Warsaw, Poland e-mail: m.dabrowski@nencki.gov.pl

Ornithine and histidine decarboxylase: activities in hypertrophic and hyperplastic mouse kidney.

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