Characterization and cytotoxic effect of aqua-(2,2′,2′′-nitrilotriacetato)-oxo-vanadium salts on human osteosarcoma cells

The use of protonated N-heterocyclic compound, i.e. 2,2′-bipyridinium cation, [bpyH⁺], enabled to obtain the new nitrilotriacetate oxidovanadium(IV) salt of the stoichiometry [bpyH][VO(nta)(H₂O)]H₂O. The X-ray measurements have revealed that the compound comprises the discrete mononuclear [VO(nta)(H₂O)]^? coordination ion that can be rarely found among other known compounds containing nitrilotriacetate oxidovanadium(IV) moieties. The antitumor activity of [bpyH][VO(nta)(H₂O)]H₂O and its phenanthroline analogue, [phenH][VO(nta)(H₂O)](H₂O)_0.5, towards human osteosarcoma cell lines (MG-63 and HOS) has been assessed (the LDH and BrdU tests) and referred to cis-Pt(NH₃)₂Cl₂ (used as a positive control). The compounds exert a stronger cytotoxic effect on MG-63 and HOS cells than in untransformed human osteoblast cell line. Thus, the [VO(nta)(H₂O)]^? containing coordination compounds can be considered as possible antitumor agents in the osteosarcoma model of bone-related cells in culture.     

The tumorigenic potential of human CX-1 colon adenocarcinoma cells depends on carcinoembryonic antigen (CEACAM5) expression

It was shown that CEACAM5 can mediate cell-cell adhesion through homotypic and heterotypic interactions; however, its role in the expression of the malignant phenotype remains obscure. To study whether the formation of both primary tumors and metastases is directly related to the presence or absence of CEACAM5, we applied the antisense RNA strategy. By transfecting human CX-1.1 colon carcinoma cells with CEACAM5 antisense-expressing vector or with the vector itself, cell variants with a highly decreased expression of CEACAM5 were obtained. Profound differences in proliferative abilities among parental and obtained subclones of CX-1.1 cells were revealed when cells were implanted subcutaneously into nude mice. In contrast to their highly tumorigenic parental CX-1.1 cells (with high expression of membrane-bound and secreted CEACAM5), two subclones (3E and AS6Q) with substantially decreased expression of membrane-bound and secreted CEA showed a considerably diminished growth rate. Even more striking results were obtained with AS8Q cells, producing a residual amount of this glycoprotein. However, 3B cells (producing a large amount of secreted CEACAM5) did not differ significantly in their tumorigenic properties from CX-1.1 cells. Our experiments performed in nu/nu mice suggest that CEACAM5 supports the growth of primary tumors, but is not involved in the formation of metastases by colon cancer cells.     

Optimisation of AP-PCR fingerprinting discriminatory power for clinical isolates of Pseudomonas aeruginosa

Recently methods based on analysis of arbitrarily amplified target sites of microorganism genomes have been extensively applied in microbiological studies. The range of their applications is limited by problems with discrimination and reproducibility resulting from lack of standardised and reliable methods of optimisation. By orthogonal-array optimisation most advantageous and optimal parameters for highly discriminatory primers (CagA2+CMVin2) were selected and efficient AP-PCR (arbitrarily primed-polymerase chain reaction) fingerprinting conditions for Pseudomonas aeruginosa isolates were set up. Stable and multiplex amplicon profiles obtained in this study revealed high level of intraspecies DNA polymorphism among 20 analysed clinical strains of P. aeruginosa proving optimised AP-PCR fingerprinting to be useful in epidemiological typing of the species.     

Pivotal participation of nitrogen dioxide in L-arginine induced acute necrotizing pancreatitis: protective role of superoxide scavenger 4-OH-TEMPO

For the first time, a direct sensitive method of *NO(2) detection and measurement in biological material has been established. It is based on the interaction of this radical with the coordination compound of Cr(III) with aminodeoxysugar as biosensor. Our new method makes it possible to precisely assess *NO(2) level in experimental acute necrotizing pancreatitis induced by L-arginine, where oxidative and nitrosative stresses are supposed to play a key role in the pathomechanism of the disease. As much as 20 nmol of *NO(2)/mg protein was detected which correlated with severe deterioration of pancreatic acinar cell ultrastructure. Protective effect of superoxide radical scavenger 4-OH-TEMPO expressed as *NO(2) level decrease confirmed by preserved acinar cell ultrastructure and decreased pancreatic amylase release to blood serum is demonstrated. This study reveals a possible pathomechanism of L-arginine induced acute pancreatitis.     

Overexpression of lumican affects the migration of human colon cancer cells through up-regulation of gelsolin and filamentous actin reorganization

Cell migration is a multistep process initiated by extracellular matrix components that leads to cytoskeletal changes and formation of different protrusive structures at the cell periphery. Lumican, a small extracellular matrix leucine-rich proteoglycan, has been shown to inhibit human melanoma cell migration by binding to α2β1 integrin and affecting actin cytoskeleton organization. The aim of this study was to determine the effect of lumican overexpression on the migration ability of human colon adenocarcinoma LS180 cells. The cells stably transfected with plasmid containing lumican cDNA were characterized by the increased chemotactic migration measured on Transwell filters. Lumican-overexpressing cells presented the elevated filamentous to monomeric actin ratio and gelsolin up-regulation. This was accompanied by a distinct cytoskeletal actin rearrangement and gelsolin subcellular relocation, as observed under laser scaning confocal microscope. Moreover, LS180 cells overexpressing lumican tend to form podosome-like structures as indicated by vinculin redistribution and its colocalization with gelsolin and actin at the submembrane region of the cells. In conclusion, the elevated level of lumican secretion to extracellular space leads to actin cytoskeletal remodeling followed by an increase in migration capacity of human colon LS180 cells. These data suggest that lumican expression and its presence in ECM has an impact on colon cancer cells motility and may modulate invasiveness of colon cancer.     

Cell division and DNA segregation in Streptomyces: how to build a septum in the middle of nowhere?

Streptomycetes are antibiotic-producing filamentous microorganisms that have a mycelial life style. In many ways streptomycetes are the odd ones out in terms of cell division. While the basic components of the cell division machinery are similar to those found in rod-shaped bacteria such as Escherichia coli and Bacillus subtilis, many aspects of the control of cell division and its co-ordination with chromosome segregation are remarkably different. The rather astonishing fact that cell division is not essential for growth makes these bacteria unique. The fundamental difference between the cross-walls produced during normal growth and sporulation septa formed in aerial hyphae, and the role of the divisome in their formation are discussed. We then take a closer look at the way septum site localization is regulated in the long and multinucleoid Streptomyces hyphae, with particular focus on actinomycete-specific proteins and the role of nucleoid segregation and condensation.     

Stability of Tandemly Repetitive Subelement PCR Patterns in Trichophyton rubrum over Serial Passaging and with Respect to Drug Pressure

Trichophyton rubrum is the most significant agent of dermatomycoses worldwide, primarily causing tinea pedis and tinea unguium. PCR analysis of tandemly repetitive subelements (TRS) within the rDNA nontranscribed spacer region is a major tool for molecular typing of T. rubrum. The aim of this study was to investigate the stability of TRS PCR patterns by analyzing isogenic strains of T. rubrum. Twenty-seven groups of isogenic T. rubrum strains were examined, each composed of an original clinical isolate and its 3 subcultures, maintained on a drug-free medium, a medium containing fluconazole and itraconazole. TRS typing was performed for the original strains and their subcultures grown after 12 passages, at 4-week intervals, on respective media. To add more objectivity to the results, TRS typing for each of the isogenic strain was performed three times, using DNA isolated from three different colonies. Among 27 groups of isogenic strains, all but one were exclusively composed of strains with identical TRS-1 and TRS-2 PCR patterns. In one group, 3 isolates from the last, twelfth passage had identical TRS-1 PCR profiles (type 1), yet different TRS-2 PCR profiles, as compared with the original strain (type I vs. type II). The mechanism underlying the genotype switch was a deletion of a single repeat unit in the TRS-2 locus, as evidenced by sequence analysis. In the interpretation of TRS typing results, microevolutionary events need to be taken into account, urging drawing epidemiological conclusions with caution and in conjunction with other genotyping data and traditional contact tracing information.