The Adhesion of Human Dermal Fibroblasts on Anodized Nanotube-layered Titanium, Modified for Implantology Application.

Anodization of titanium implants is accompanied by the formation of titanium oxide nanotubes improving osseointegration. An excessive fibroblast adhesion on the surface might lead to the formation of fibrous capsule resulting in implant rejection. In our research, we demonstrated that the adhesion activity of human dermal fibroblasts on anodized surface was not elevated, which is promising for the use of titanium with nanotube-layered surface for implantology.

Isolation of primary osteoblast cell lines from adult rat and rat embryos and their use as models for in vitro biocompatibility tests of nanostructured titanium-based implants.

Methods for obtaining osteoblast cultures from the calvaria of adult Wistar rats and 12-day-old embryos of these rats have been adapted for studying the biocompatibility and ossointegration of titanium-based implants. The osteoblast morphology and their differentiation into osteocytes on a titanium matrix with specially treated surface have been studied. It has been confirmed that two cultures of diploid rat cells obtained in the study can serve as efficient models for preclinical in vitro testing of nanostructured titanium implants for biocompatibility and osseointegration.

Sequence-specific minor groove binding ligands as potential regulators of gene expression in Xenopus laevis oocytes.

The mouse mammary tumor virus (MMTV) promoter is induced by glucocorticoid hormone. A robust hormone- and receptor-dependent gene activation could be reproduced in Xenopus laevis oocytes. The homogeneous response in this system allowed a detailed analysis of the DNA-protein interactions following hormone activation. The strategy of artificial regulating of gene activity by sequence-specific minor groove binding ligands is very attractive. We have synthesized and studied the interaction with DNA of bis-linked netropsin derivatives in which two monomers are attached via short linkers in head-to-head and tail-to-tail manners. We have found that cis-diammine-platinum bridged bis-netropsin added to Xenopus oocytes media penetrates cellular and nuclear membrane and binds selectively to the MMTV promoter at the DNA segment that partly overlaps with the site recognized by glucocorticoid receptor. DNase I footprinting studies demonstrate that there are more stronger binding sites for cis-diammine-platinum bridged bis-netropsin on the naked MMTV DNA which are found to be inaccessible for its binding in oocytes.

[Ultramorphometric study of the plastids in apical tuber cells of original and transgenic potato plants treated with ambiol].

Ultramorphometric characteristics of plastids in cells of apical tuber meristems of original and defensin gene-transfected potato (Solanum tuberosum L.) plants, either maintained under normal conditions or subjected to treatment with the antioxidant ambiol, were compared. Under normal conditions, the tuber cells of the original and transgenic potato plants differed in neither the number nor size of the plastids. Only certain quantitative distinctions in the development of individual ultrastructural characteristics of plastids were detected. Treatment with ambiol enhanced the differentiation of the internal membrane system of plastids in the cells of original and transgenic plants, especially the tubular membrane systems. Certain differences in the responses to ambiol of cell plastids of original and transgenic plants were related to plastid sizes and development of individual intraplastid structures. The results comply with earlier data on varying responses of mitochondria of original and transgenic plants to ambiol treatment.

Mapping protein-DNA interactions with CIS-DDP: chromatin structure of promoter region of D. Melanogaster hsp 70 gene.

DNA-protein crosslinking by cis-dichlorodiammineplatinum (II) (cis-DDP) was applied to study chromatin structure in situ. Histone H1 (H5) is crosslinked to DNA in significant amounts whereas core histones remain practically unattached. "Protein image hybridization" experiments show that the 5'-region of the D metanogaster hsp 70 gene is free of histone H1 in both control nuclei and nuclei isolated from heat-shocked embryos.

Mapping of DNA-binding proteins along the yeast genome by UV-induced DNA-protein crosslinking.

UV-induced crosslinking of DNA-binding proteins to DNA in intact nuclei of Saccharomyces cerevisiae and subsequent 'protein image' hybridization were applied to map non-histone proteins along single-copy genes of yeast. We detected two polypeptides that most probably correspond to core subunits of yeast RNA-polymerase II in the coding region of transketolase gene (TKL2). Several non-histone proteins were also detected which bind to the upstream region of TKL2 gene, and to the intergenic spacer between calmodulin (CMD1) and beta-mannosyl transferase (ALG1) genes.

Mapping and positioning DNA-binding proteins along genomic DNA. Structure of D. melanogaster ribosomal 'Alu-repeats' and 1.688 satellite chromatin.

Chromatin structure of so-called 'Alu-repeat' in D. melanogaster ribosomal non-transcribed spacer that contains sequences homologous to the promoter of ribosomal genes has been studied. Using the 'protein image' hybridization assay based on UV-light-induced DNA-protein crosslinking and 2-D gel retardation electrophoresis, two proteins of the molecular mass of 50 kD (rABP50) and 70 kD (rABP70), associated with 'Alu-repeat' DNA have been found. Exo III mapping of crosslinking sites and DNase I footprinting have provided a detailed map of H1, rABP50 and rABP70 contacts within the 'Alu-repeat' and H1 and a non-histone protein contacts on satellite DNA. These data indicate precise positioning of non-histone proteins, histone H1 and nucleosomes within genomic regions studied and account for the presence of unusual 240 bp long nucleosomal particles in 'Alu-repeats'. The same approach can be adapted for successive mapping and positioning proteins on genomic DNA.

Two non-histone proteins are associated with the promoter region and histone HI with the transcribed region of active hsp-70 genes as revealed by UV-induced DNA-protein crosslinking in vivo.

We described here an approach for mapping proteins on any sequence of genomic DNA. UV-induced DNA-protein crosslinking within whole cells and the 'protein image' hybridization technique (1) were applied to test the proteins bound to different regions of the D. melanogaster hsp-70 gene. The histone H1-DNA association with the coding region is shown to be maintained, even during very intensive transcription, but is absent in the promoter. Two non-histone proteins with apparent molecular masses of 50 kD (p50) and 100 kD (p100) are crosslinked only to the active hsp-70 gene regulatory region and preferentially bind to its complementary and coding DNA strands, respectively.

Chromatin structure of Drosophila melanogaster ribosomal genes.

The chromatin structure of ribosomal genes of D. melanogaster has been studied by crosslinking proteins to DNA. We found that a number of histone contacts with DNA through histidine in the approximately 1 kb-long region surrounding the transcription initiation site, coding regions and the region of 240 bp-long repeats from the intergenic spacers (Alu-repeats) were weakened as compared to the inactive chromatin of the type II insertion. A protein with the molecular mass of 50 kDa (p50), associated with all DNA sequences analysed, has been discovered. Another protein with molecular mass of about 70 kDa (p70) has been found to be specific only for the Alu-repeats.