Tadeusz Wilczok

Butyrate-Induced Differentiation of Colon Cancer Cells Is PKC and JNK Dependent

Butyric acid, a short–chain fatty acid physiologically present in human large gut, is derived from bacterial fermentation of complex carbohydrates. It has been shown to reduce the growth and motility of colon cancer cell lines and to induce cell differentiation and apoptosis. Apoptosis is considered a result of normal colonocyte terminal differentiation in vivo. The aim of this study was to characterize the cellular mechanisms regulating differentiation of colon cancer cells stimulated with sodium butyrate (NaB). The two human colon cancer cell lines Caco-2 and HT-29 were treated with NaB at physiologically relevant concentrations. Alkaline phosphatase (ALP) activity, a marker of colonocyte differentiation, was increased 48 hr after treatment with 1 mM NaB. Higher doses of NaB (5 and 10 mM) induced apoptosis of the cells and failed to stimulate the colonocyte differentiation. Therefore, we assumed that butyrate augments cell differentiation and induces apoptosis, acting via various intracellular mechanisms, and butyrate-mediated programmed cell death cannot be considered a consequence of colonocyte terminal differentiation. The effect of NaB on ALP activity was significantly attenuated in the presence of inhibitors of protein kinase C and JNK. Inhibition of MEK–ERK signal transduction pathways augmented the impact of butyrate on colonocyte differentiation. These results suggest that butyrate could influence the colonocyte differentiation via modulation of the activity of cellular protein kinases and signal transduction.

Structure, properties and cytostatic activity of tributyltin aminoarylcarboxylates

Abstract Properties of butyltin complexes [Sn(C4H9-n)3{OOCC6H3(NH2)2-3,4}]n (1), [Sn(C4H9-n)3{OOCC6H3(NH2)2-3,5}] (2), [Sn(C4H9-n)3{OOCC6H4NNC6H4N(CH3)2-4}] (3) and [Sn(C6H5)3{OOCC6H3(NH2)2-3,5}]n (4) have been investigated. 1H, 13C and 119Sn NMR spectra indicate that the compounds in chloroform are distorted tetrahedral and in strongly coordinating solvents trigonal-bipyramidal complexes. Structure of complex 3 has been determined by X-ray crystallography. This compound adopts trigonal bipyramidal structure with bridging carboxylato ligand bound asymmetrically with tin atoms in axial positions. The complexes are effective cytostatic agents.

Antioxidative activity of synthetic melanins. Cardiolipin liposome model

The inhibiting effect of melanin synthesized from dihydroxyphenylalanine (DOPA), dopamine, adrenaline and adrenolutin on the ultraviolet- or the Fe(2+)-ascorbic acid-induced peroxidation of cardiolipin liposomes has been studied. All these melanins are able to inhibit both the ultraviolet- and the Fe(2+)-ascorbic acid-induced lipid peroxidation. Antioxidative activity of melanins enhances in the order: dopamine-melanin less than melanin synthesized from dopamine in the presence of Cu(2+) less than DOPA--melanin less than melanin synthesized from adrenaline in the presence of Cu(2+) approximately equal to adrenolutin-melanin, and correlates with their ability to scavenge superoxide anion radical. The optical screening effect of the investigated melanins in the inhibition of lipid peroxidation was not higher than 15% for the most active melanins.

Human cytomegalovirus in patients with systemic lupus erythematosus

The objective of this study was to determine the frequencies of human cytomegalovirus (HCMV) infection and HCMV genome copy number in blood of consecutive (treated from several months to several years) systemic lupus erythematosus (SLE) patients (22 women). The obtained results were compared to the healthy controls (15 women). All patients fulfilled at least four of the 1982 revised American rheumatism association (ARA) classification criteria for SLE. Our patients demonstrated three or four of the nine possible organ systems involved and most of them had mild SLE with SLE disease activity index (SLEDAI) score < 10 at time when blood samples were collected to detect HCMV. Quantitative analysis of HCMV genome was performed with aid of sequence analyzer ABI PRISM TM 7700 Perkin Elmer. Primers and probe were constructed on the basis of IE4 region of HCMV genome. The viral load was expressed as log10 of calculated HCMV genome copy number. Qualitative analysis revealed that 100% of our SLE patients were infected with HCMV, whereas in the control group only 73% of persons were HCMV positive. Statistically significant difference was demonstrated when the strength of the association between SLE or controls and infection of HCMV was calculated (estimated by Fishers exact test, P value = 0.02). Higher viral DNA copy number was observed in whole blood of SLE patients than in the control group (338.45 ± 221.76 and 229.00 ±405.61 copies/ml respectively) but did not reach statistical significance level (95% confidence interval from 170.41 to 249.32, P = 0.71). Furthermore percentage of patients with HCMV-DNA copy number >2.0 × 102 copies/ml was statistically significantly higher than this one in controls. The data show association between HCMV infection and SLE, which should be taken into account during the course of SLE.

Application of EPR spectroscopy to examination of gentamicin and kanamycin binding to DOPA-melanin

Electron paramagnetic resonance (EPR) spectroscopy was applied to measure the influence of two aminoglycoside antibiotics: gentamicin and kanamycin on free radical propertis of DOPA-melanin. DOPA-melanin was formed by oxidative polymerization of 3,4-dihydroxyphenylalanine. Different concentrations of gentamicin and kanamycin (from 1·10−4 to 1·10−2 M) were used. o-Semiquinone free radicals with ag factor of 2.0043 were found in all studied melanin samples. Their concentrations in the DOPA-melanin-drug complexes were higher than in DOPA-melanin, and increased with the increase of gentamicin and kanamycin concentration. A single EPR line of the analyzed samples (ΔBpp, 0.48-0.52 mT) indicates that aminoglycoside antibiotics do not create a new type of free radicals in DOPA-melanin. Microwave saturation behavior of the experimental lines indicates the homogeneous broadening of resonance absorption curves for DOPA-melanin and its complexes with aminoglycosides. The EPR lines saturate at low microwave powers. Slow spin-lattice relaxation processes were characteristic for all studied melanin samples.

Structure, Properties and Cytostatic Activity of Triorganotin (Aminoaryl)carboxylates

The properties of vinyltin and phenyltin complexes [Sn(CH=CH2)3{μ-OOCC6H3(NH2)2-3,4}]n (1), [Sn(C6H5)3{OOCC6H3(NH2)2-3,4}] (2), [Sn(C6H5)3{OOC-2-C6H4N=NC6H4N(CH3)2-4}] (3) and [Sn(CH=CH2)3{OOC-2-C6H4N=NC6H4N(CH3)2-4}] (4) have been investigated. The structures of complexes 1, 2, and 3, have been determined by X-ray crystallography. Compound 1 is a distorted trigonal-bipyramidal complex and compounds 2 and 3 adopt a distorted tetrahedral structure. Complex 1 is a single-strand polymer with a bridging 3,4-diaminobenzoato ligand coordinating via the O(1) atom of the carboxylato group and the nitrogen atom of the para-amino group. The oxygen and nitrogen atoms occupy the axial coordination sites. The Sn(1)−N(2A) bond is weak. In complexes 2 and 3 the carboxylato ligands are strongly coordinated to the central atom via one oxygen atom, and the Sn(1)−O(2) distances are considerably longer. Weak interactions of the central atom with the amino group in complex 1, and with the O(2) atoms in complexes 2 and 3, as well as the hydrogen bonds, stabilize the crystal structure. The complexes are effective cytostatic agents. (© Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002)

Catecholamine melanins structural changes induced by copper ions

Melanins synthesized from adrenaline and dopamine in the presence or absence of copper ions were characterized by pyrolysis-gas chromatography-mass spectrometry and by IR and ESR methods. It was shown that Cu2+ are able to induce changes in the melanin structure. Melanins obtained from adrenaline-Cu2+ and dopamine-Cu2+ complexes are composed mainly from monomeric units of the indole type. Melanins synthesized from these catecholamines without Cu2+ contain additionally large amounts of unindolized monomeric units. The structure differences in both types of melanins are reflected in their sorptive abilities and spectroscopic characteristics.

Pyrolytic GC-MS analysis of melanin from black, gray and yellow strains of Drosophila melanogaster

Abstract Properties of melanins depend strongly on their chemical constitution. Melanins from differently colored strains of Drosophila melanogaster demonstrate various proprieties, and biological activity in the presence of toxic ions or radiation. In the presented work, we analyzed chemical constitution of the melanins derived from black, gray and yellow D. melanogaster strains. Analysis of the products forming during thermal degradation of the biopolymers by pyrolitic gas chromatography followed by mass spectrometry allowed us to determine chemical composition of these biopolymers and classified melanins derived from black and gray strains as eumelanins and derived from yellow strains as pheomelanins. These findings allow us to determine chemical and biological properties of the melanins in vivo, and can explain susceptibility of differently colored strains of D. melanogaster to metal ions.

Spectroscopic studies of chemically modified synthetic melanins

The infrared and electron spin resonance spectra of synthetic 3,4-dihydroxyphenylalanine (DOPA) and tyrosine melanins and chemically modified melanin samples were determined, and it was shown that unmodified and reduced DOPA melanins exhibited similar ir spectra. Oxidized DOPA melanins showed a higher number of carboxy groups in the sample. A significant increase of free radical content in reduced DOPA melanin and a decrease of free radical content in oxidized DOPA melanin in comparison to unmodified samples were demonstrated by the use of ESR methodology. Methylation of tyrosine melanin with an excess of diazomethane gave very rich ir spectra as compared to melanins methylated with methanol saturated by gaseous HCl. In tyrosine melanin samples the esterification of carboxy groups with methanol caused a decrease in the free radical content. When diazomethane was used, the methylated melanin samples had free radical levels reduced to only about 4% of the total observed for unmodified tyrosine melanin.

Structure, properties and in vitro cytotoxic activity of hexakis(2-cyanoethyl)ditin(III).

The structure of the tin(III) complex [Sn(2)(CH(2)CH(2)CN)(6)] has been determined. There are two independent molecules in the crystal, both adopt distorted eclipsed conformation. The molecular and electronic structures of this compound have been studied both at the semiempirical level and with the use of non-empirical ab initio methods. The calculated Sn-Sn distances agree well with those found crystallographically. The results of calculations showed that the eclipsed conformation of complex is more stable as compared with staggered conformation. The compound show modest cytotoxic activity against A549 and HSMC cells.