[Morpho-densitometric researches of cell nuclei at atypical hyperplasia and initial breast cancer].

There were performed researches on morphometric (the nuclear area, perimeter, the maximum and minimum diameters) and densitometric (optical density, integrated optical density) parameters of nuclei of cells on the cytologic smears received from 28 patients with atypical hyperplasia and an initial breast cancer. Significant reduction of measured nuclei parameters was noted among: moderately - and high-grade ductal carcinoma in situ, atypical ductal hyperplasia, lobular carcinoma in situ, an invasive papillary cancer, intraductal papilloma. The lowest value of the measured parameters was noted at love-grade ductal carcinoma in situ and intraductal papillary cancer. It was established that morphometric and densitometric indicators at atypical hyperplasia of an epithelium and an initial breast cancer in some cases overlap each other that is at the bottom of difficulties of differential cytomorphological diagnostics and quite often forces to formulate the cytologic conclusion in the presumable form.

Effectiveness of cytological diagnosis of breast cancer metastases in regional lymph nodes.

Results are presented of preoperative and intraoperative cytological diagnosis of breast cancer metastases in regional lymph nodes in 84 patients. Of them in 42 there was studied material of fine-needle aspiration biopsy, in 42 - smears of sentinel lymph nodes. In all cases cytohistological comparisons were carried out. The effectiveness of cytological examination of material of fine-needle aspiration biopsy was 90.5%. Urgent cytology of smears of sentinel lymph nodes allowed revealing metastases in 5 of 10 patients. The use of immunohistochemical examination increases the likelihood of detection of micrometastases and isolated tumor cell clusters.

[An evaluation of effectiveness of cytological method in diagnostics of breast tumors].

This paper presents an analysis of the results of cytological diagnostics of breast tumors in 3415 patients who underwent tests in the Laboratory of Cytology for 2009-2013. In 1434 patients there were performed cytohistological correlations. Sensitivity of cytology was 99.1%, specificity--93.5%, predictive value of the positive test--98.5%, predictive value of the negative test--95.9%, accuracy--98%. Non-informative material was obtained in 13.5% of cases. The main factors that reduced effectiveness of cytological method were non-informative and little informative material and also difficulties in conducting of differential cytomorphological diagnostics.

Platinum coat color in red fox (Vulpes vulpes) is caused by a mutation in an autosomal copy of KIT.

The red fox (Vulpes vulpes) demonstrates a variety of coat colors including platinum, a common phenotype maintained in farm-bred fox populations. Foxes heterozygous for the platinum allele have a light silver coat and extensive white spotting, whereas homozygosity is embryonic lethal. Two KIT transcripts were identified in skin cDNA from platinum foxes. The long transcript was identical to the KIT transcript of silver foxes, whereas the short transcript, which lacks exon 17, was specific to platinum. The KIT gene has several copies in the fox genome: an autosomal copy on chromosome 2 and additional copies on the B chromosomes. To identify the platinum-specific KIT sequence, the genomes of one platinum and one silver fox were sequenced. A single nucleotide polymorphism (SNP) was identified at the first nucleotide of KIT intron 17 in the platinum fox. In platinum foxes, the A allele of the SNP disrupts the donor splice site and causes exon 17, which is part of a segment that encodes a conserved tyrosine kinase domain, to be skipped. Complete cosegregation of the A allele with the platinum phenotype was confirmed by linkage mapping (LOD 25.59). All genotyped farm-bred platinum foxes from Russia and the US were heterozygous for the SNP (A/G), whereas foxes with different coat colors were homozygous for the G allele. Identification of the platinum mutation suggests that other fox white-spotting phenotypes, which are allelic to platinum, would also be caused by mutations in the KIT gene.

Measurement of segregating behaviors in experimental silver fox pedigrees.

Strains of silver foxes, selectively bred at the Institute of Cytology and Genetics of the Russian Academy of Sciences, are a well established, novel model for studying the genetic basis of behavior, and the processes involved in canine domestication. Here we describe a method to measure fox behavior as quantitative phenotypes which distinguish populations and resegregate in experimental pedigrees. We defined 50 binary observations that nonredundantly and accurately distinguished behaviors in reference populations and cross-bred pedigrees. Principal-component analysis dissected out the independent elements underlying these behaviors. PC1 accounted for >44% of the total variance in measured traits. This system clearly distinguished tame foxes from aggressive and wildtype foxes. F1 foxes yield intermediate values that extend into the ranges of both the tame and aggressive foxes, while the scores of the backcross generation resegregate. These measures can thus be used for QTL mapping to explore the genetic basis of tame and aggressive behavior in foxes, which should provide new insights into the mechanisms of mammalian behavior and canine domestication.

[Silencing of c-myc gene expression by enzymatically and chemically synthesized siRNAs].

Small interfering RNAs (siRNA) provide a powerful approach for sequence-specific silencing of gene expression. In the present study we investigated inhibition of c-myc gene expression by siRNAs targeted to the sequence 1452-1470 b. in third exon of c-myc mRNA and to homologous regions in second exons of c-myc (697-715 b.) and N-myc (302-320 b.) mRNAs. siRNAs were prepared enzymatically according to the scheme, including dsDNA-templates preparation using Klenow fragment, separate in vitro transcription of each RNA strand with subsequent hybridization and removal of leader sequences by T1 RNase. Investigation of c-myc gene silencing by siRNAs revealed that enzymatically prepared siRNAs induce stronger inhibition of c-myc expression, than siRNA with the same sequence prepared by chemical synthesis. It was found that down-regulation of c-myc gene expression by investigated siRNAs results in efficient inhibition and even complete arrest of carcinoma cell proliferation, moreover, the extend of growth inhibition correlates with the level of siRNA-mediated reduction of c-myc mRNA.

Silencing of MDR 1 gene in cancer cells by siRNA.

Inhibition of p-glycoprotein (PGP) expression and reverse of multidrug resistance (MDR) phenotype in KB-8-5 cells by synthetic 21-bp double-stranded oligoribonucleotides were investigated. siRNA constructs for the efficient down regulation of MDR1 that are active in nanomolar concentrations and cause reversal of MDR phenotype in cells were developed.

Silencing of c-myc expression in tumor cells by siRNA.

Suppression of c-myc protooncogene expression in KB-3-1 cells by siRNA was investigated. The siRNA duplex targeted to the exon 3 of c-myc mRNA was prepared by in vitro transcription with T7 RNA polymerase on short dsDNA-templates. It was found that incubation of KB-3-1 cells in the presence of 75 nM siRNA results in decrease of the c-myc mRNA level down to 5% of the level in the control cells and significant decline of KB-3-1 cell proliferation rate. Using 200 nM siRNA four-fold decrease of KB-3-1 cells proliferation rate was observed and this effect was stable at least 96 h after transfection.

[Binding of transcription factors with functionally significant regions of MDR1 gene promoter during its activation]. / Izuchenie sviazyvaniia transkriptsionnykh faktorov s funktsional'no znachimymi raonami promotora gena MDR1 pri ego aktivatsii.

Interaction of transcription factors with functional regions of gene mdr1 promoter has been studied. Binding of transcription factors to double-stranded (ds) oligonucleotides, mimicking four important regulatory regions before and after treatment of KB and K-562 cell lines with doxorubicin, cytarabine, and vinblastine was analysed by gel-shift assay. Gene induction resulted in enhancement of the main complex A(A1) proteins formation on the all four regions in both the cell lines. Inhibition of each complex formation by an excess of non-radiolabelled ds oligonucleotides suggests the possibility of regions interaction during promoter activation. Another indication of interaction of regulatory regions during activation is overlapping minor complexes sets. The results suggest that the four studied regulatory regions are important for mdr1 gene activation. Corresponding oligonucleotides mimicking these regions can be employed as inhibitors of gene transcription that bind specific transcriptional factors.