Analysis of the Basic Characteristics of Osteogenic and Chondrogenic Cell Lines Important for Tissue Engineering Implants.

We isolated and characterized cultures of bone and cartilage tissue cells of laboratory minipigs. The size and morphological features of adherent osteogenic and chondrogenic cells were specified. During long-term culturing under standard conditions, the studied cultures expressed specific markers that were detected by immunohistochemical staining: alkaline phosphatase and calcium deposits in osteoblasts and type II collagen and cartilage extracellular matrix in chondrogenic cells. Proliferative potential (mitotic index) of both cell types was 4.64% of the total cell number. Cell motility, i.e. the mean velocity of cell motion was 49 pixels/h for osteoblasts and 47 pixels/h for chondroblasts; the mean migration distance was 2045 and 2118 pixels for chondroblasts and osteoblasts, respectively. The obtained cell lines are now used as the control for evaluation of optimal biocompatibility of scaffold materials in various models. Characteristics of the motility of the bone and cartilage tissue cells can be used for modeling and estimation of the rate of cells population of 3D scaffolds made of synthetic and biological polymers with different internal structure and physicochemical properties during designing in vitro tissue implants.

Comparative mapping of cattle chromosome 19: cytogenetic localization of 19 BAC clones.

Here we present the results of fluorescent in situ hybridization (FISH) mapping of a set of cattle BAC clones preselected for assignment on cattle chromosome 19 (BTA19). The BAC clones were anchored to human chromosome 17 (HSA17) sequences by BLASTn similarity search of cattle BAC-ends against the human genome sequence (NCBI build 33). Five blocks of homologous synteny were defined in the comparative map of BTA19 and HSA17 built with FISH data and the human genome coordinates. The positions for four evolutionary breakpoints in the bovine and human chromosomes were identified. Comparison of the FISH comparative map with previously published comparative RH, physical, and cytogenetic maps of BTA19 did not reveal major conflicts and allowed for the extension of the boundaries of homology between BTA19 and HSA17. Comparative analysis of HSA17, BTA19, and mouse chromosome 11 (MMU11) demonstrates that most likely mice retain the ancestral organization of the synteny group, and both cattle and human chromosomes underwent several major internal rearrangements after the divergence of Primates, Rodentia, and Cetartiodactyla.

[Use of PCR markers for mapping swine chromosome 12]. / Ispol'zovanie PTsR-markerov dlia kartirovaniia khromosomy 12 svin'i.

Using PCR analysis of pig-mink and pig-Chinese hamster hybrid cell lines and heterologous and homologous primers of various types, chromosomal and subchromosomal mapping of genes TOP2A, THRA, BRCA1, GAS, HLR1, MYL4, LIS1, MCP1, ENO3, CRYB1, P4HB, STAT5B, and H3F3B to pig chromosome 12 was carried out. The efficiency of using different types of heterologous primers for pig chromosome mapping was compared.

Zoo-FISH with region-specific paints for mink chromosome 5q: delineation of inter- and intrachromosomal rearrangements in human, pig, and fox.

Comparison of evolutionarily conserved mammalian chromosomes homologous to human chromosome 17, performed with microdissected painting probes, revealed rearrangements inside these chromosomes in mink and pig and a disruption of this conserved region in the fox. Detection of a homologous region on an Iberian shrew chromosome showed the efficiency of microdissected painting probes for delineation of homologous chromosome regions in species belonging to orders that diverged at least 100 million years ago.

[A new approach to the analysis of complex chromosomal rearrangements in cell hybrids]. / Novye vozmozhnosti analiza slozhnykh khromosomnykh perestroek v kletochnykh gibridakh.

The chromosomal complements of somatic cell pig-mink hybrids was determined by a new approach. This approach includes microdissection of metaphase chromosomes, generation of chromosome and region-specific DNA libraries, and fluorescence in situ hybridization of these libraries with pig lymphocyte chromosomes. The studied hybrid cells were shown to contain two small acrocentric chromosomes and a microchromosome of porcine origin. Identification of these chromosomes by differential GTG-staining was impossible. Chromosome isolation by a micromanipulation technique followed by DNA amplification in TOPO-DOP polymerase chain reaction provided chromosome-specific DNA libraries of the rearranged chromosomes. Based on these libraries, the labeled DNA probes were prepared and hybridized to pig chromosomes. This allowed us to determine the origin of the material contributing to the hybrid cell chromosomes. One of these chromosomes contained five pig chromosomal regions: 15cen-q2; 6q21-q23; 13q21; 13q22; 7q25-qter, while the other contained the following pig chromosomal regions: 4p12-p13; 16q12-q14; 12pter-p15. The microchromosome contained the Xp11-Xq11 region. The minimal size of the revealed chromosomal regions was about 3 to 4 x 10(6) bp. Segregation analysis of the thymidine kinase gene 1 (TK1), which was earlier localized to the pig 12p region, and the hybrid cell pig chromosomes in the hybrid subclones suggested that TK1 gene can be assigned to 12p15-pter. The results obtained demonstrate the efficiency of the applied approach in its detailed and reliable description of complex chromosomal rearrangements in hybrid clones, when differential chromosome staining failed to identify these chromosomes.

[Chromosome localization and analysis of synteny analysis of some genes in swine, cattle, and sheep (Artiodactyla)]. / Khromosomnaia lokalizatsiia i analiz sintenii nekotorykh genov u svin'i, korovy i ovtsy (otriad Artiodactyla)]

Using the hybrid cell lines pig-American mink, cow-American mink, and sheep-American mink, the localization of some genes included in a large conservative block localized on human chromosome (chr) 17 was performed by means of electrophoresis of proteins and Southern blot hybridization. Genes NF1, RARA, PRKCA, and ERBB2 were assigned to chr 12 in swine; TK1 and UMPH2, to chr 19 in cattle; and TK1, UMPH2, and PEPA, to chr 11 in sheep. The conserved synteny of these genes in three representatives of the order Artiodactyla was shown.

Characterization of pig-mink cell hybrids: assignment of the TK1 and UMPH2 genes to pig chromosome 12.

By fusion of thymidine kinase-deficient mink cells with pig leukocytes, a new type of cell hybrid was produced. It was demonstrated that pig chromosomes segregate in pig-mink hybrids and that hybrid cells contain no cytologically visible rearrangements between the chromosomes of parental species, or chromosome fragmentation. With a set of subclones of two primary hybrid clones, the genes for thymidine kinase-1 (TK1) and uridine 5'-monophosphate hydrolase-2 (UMPH2) were assigned to pig Chromosome (Chr) 12. A cell line with a single pig Chr 8 on the background of mink chromosomes was established. This clone could serve as a source of DNA for building a chromosome-specific library of pig Chr 8. The data obtained suggest that pig-mink cell hybrids can be used for mapping of pig chromosomes.

[Preliminary localization of TK1 and UMPH2 genes on swine chromosome 12]. / Predvaritel'naia lokalizatsiia genov TK1 i UMPH2 na khromosomu 12 svin'i.

A new type of pig-mink cell hybrids was produced. It was demonstrated that segregation of pig chromosomes occurs in these hybrids and that no rearrangements between the chromosomes of different species occurs; fragmentation of pig chromosomes is also absent. Using these hybrids, genes TK1 and UMPH2 were preliminarily located on pig chromosome 12. The obtained data demonstrate that these hybrids can be used for chromosome mapping of pig genes.

[Detailed analysis of a new translocation in a domestic pig]. / Detal'nyi analiz novoi translokatsii u domashnei svin'i.

A sterile minisibs (mini Siberian swine) boar with normal sexual behaviour was studied by the routine, C- and high resolution G-banding chromosome techniques. A new reciprocal translocation was identified involving chromosomes 12 and 13. The numbers of diakinesis and sperms were very low. The synaptonemal complex was typical of reciprocal translocations.