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Using fluorescence methods for visualization of human tumor cell apoptosis and necrosis in vitro, the antitumor effects of flavonoid-containing Gratiola officinalis L. and green tea extracts at ultralow concentrations on A498 human kidney carcinoma cells after 24 and 48 h were compared. Fluorescent dyes (propidium iodide and acridine orange) were used in the "living and dead" test, which made it possible to identify the total number of cells that died by necrosis and apoptosis, as well as cells in which apoptosis has started and apoptotic bodies or pycnosis of the nucleus were detected. Gratiola officinalis L. extract had the most pronounced cytostatic activity 48 h after exposure; the number of cells per field of view decreased by a factor of 1.5 compared with the control. Green tea extract at a concentration of 0.0288 mg/mL had the most pronounced cytotoxic activity; its manifestation was growing 48 h after exposure. Gratiola officinalis L. extract exhibited cytotoxic activity at lower concentrations (0.0036 mg/mL). [ABSTRACT FROM AUTHOR]

The purpose of the work is to study antitumor activity of flavonoid-containing extracts of anthocyan maize and gratiola in experiments in vivo and to estimate their effects on the internal organs, tumors and blood of rats with transplanted liver cancer PC-1. Materials and methods: Effect of extracts of anthocyan maize and gratiola has been studied in experiments in vivo in 30 laboratory rats with transplanted liver cancer PC-1 using morphological and biochemical methods. Results: It has been found that extract of gratiola received by a certain way has selective cytotoxic effect on tumor cells. Anthocyan maize extract reduces endogenous intoxication. Conclusion: These results open the prospect of further study of the extract of gratiola in various experimental models of cancer and anthocyan maize extract use may be developed as a means of endogenous intoxication reducing for application in the treatment of cancer. Investigation of anthocyan maize extract may assist in the development of drugs that reduce the endogenous intoxication at the treatment of cancer.

Introduction: Gratiola officinalis L. (hedge hyssop), a medicinal plant of the Scrophulariaceae family, has diuretic, purgative, and vermifuge properties. It is used as a herbal tea to treat chronic gastroenteritis, renal colic, jaundice, and intestinal worms. Previously, we have found that an extract from G. officinalis is nontoxic and has antitumor, antioxidant, antimicrobial, antiinflammatory, anticachexic, and other properties. Our aims in this study were to separate the G. officinalis extract into individual fractions, to identify the most biologically active fractions, and to examine the chemical composition of these fractions and their biological activity toward A498 renal carcinoma cells., Methods: The G. officinalis extract was fractionated by reversed-phase high-performance liquid chromatography, and each fraction was tested for antitumor activity. The active fractions were characterized by UV-visible electron spectral analysis, circular dichroism analysis, Fourier transform infrared spectroscopy, high-performance liquid chromatography, electrospray ionization tandem mass spectrometry, and nuclear magnetic resonance spectroscopy., Results: Two antitumor-active fractions of a flavonoid nature were isolated and chromatographically purified. On the basis of the nuclear magnetic resonance data, the aglycone fragment of the main component of one fraction was found to be structured as 2-(3,4-dimethoxyphenyl)-7-hydroxychroman-4-one, or 3',4'-dimethoxy-7- hydroxyflavanone., Conclusion: The antitumor effect of the most active fraction containing 7-O-glucoside of apigenin, glycoside 7,3'-di-O-luteolin and trace amounts of eupatilin against renal carcinoma A498 cells was manifested in its cytotoxic, cytostatic, apoptotic and autophagosomal activities. In addition, we found 3-(1-2)-glucoside of soyaspogenol B, which is a pentacyclic triterpenoid in the structure., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Flavonoid-containing Gratiola officinalis extract has been studied in relation to breast carcinoma and human cervical cancer cells in encapsulated and native form. Encapsulation was realized in polymer shells, which were formed by the layer-by-layer method using sequential adsorption of poly(allylamine hydrochloride) and poly(sodium 4-styrenesulfonate) on the destructible cores. The extract was prepared by the author's method and characterized using high performance liquid chromatography. By means of optical and fluorescent microscopy, cell changes under the action of pure and encapsulated extracts were comprehensively studied, and statistical analysis was carried out. Cells were stained with propidium iodide, acridine orange, and Hoechst 33258. A fluorescence microscope with a digital video camera were used for cell imaging. The encapsulated extract caused 100% death of breast cancer SKBR-3 cells and 34% death of cervical cancer HeLa cells and prevented the formation of autophagosomes in both cultures. Analysis of the viability and morphological features of tumor cells under the action of microencapsulated extract allows us to consider microencapsulation as an effective strategy for delivering Gratiola officinalis extract to tumor cells and a promising way to overcome the protective autophagy.

Urolithiasis is one of the most urgent problems of clinical urology. Currently, there is no consensus on the causes of stone formation, as well as the role of various factors in the development of urolithiasis, however, increasingly, according to various studies, the leading role is given to genetic causes. The article presents a modern review of data on genetic polymorphisms associated with ICD: rs1801197 and rs6776158 of the CASR gene; TaqI of the VDR gene; rs1801197 of the CALCR gene, rs3752472, rs650439, rs2853744 of the Klotho gene.

The karyotype of Camptochironomus pallidivittatus Edwards, 1929 (Diptera, Chironomidae) from five populations of the Lower Volga region and Central Caucasus (the northern macroslope) has been studied. In populations of S. pallidivittatus from the Central Caucasus, 11 banding sequences (BS) were found; one sequence, pal B10, was new to the species. In the Saratov population, 11 BS were also found, three of which were new for the species-pal A3, pal B11, and pal B12. The banding sequences detected for the first time have not yet been found in other parts of the habitat of this species and may be endemic to these regions. In the studied populations ofS. pallidivittatus, banding sequences were found that were nonstandard but fixed in the karyotype. This is indicative of some degree of chromosomal divergence. These banding sequences include pal A2.2 in arm A and pal B10.10 in arm B in the Central Caucasus region, as well as pal B2.2 and pal G2.2 in the Lower Volga region. Arms A, B, D, and G in the Central Caucasian populations and A, B, and D in the Saratov oblast were polymorphic. The composition of heterozygous sequences between populations from different regions coincided only in arm D (pal D 1.2). In arms A and B, the set of heterozygous BS was different: pal A1.2 and pal B1.10 sequences were found in the Central Caucasian populations, and pal A1.3 and B11.12 were found in Saratov oblast. The number of genotypic combinations of S. pallidivittatus was higher in the Central Caucasus region, whereas the number of zygotic combinations was higher in the Saratov population. The percentage of heterozygous larvae in the Central Caucasian populations varied from 20 to 80, whereas all individuals in the Saratov population had heterozygous inversions. Zygotic combinations of larvae in all the studied populations were different.

The extract of Gratiola officinalis L. has been obtained by an original method ensuring the maximum yield of flavonoids. The extract simultaneously exhibits high anti-inflammatory activity, selective antimicrobial properties (with respect to Staphylococcus aureus and Pseudomonas aeruginosa, but not to conditionally pathogenic E. coli) and antipyretic effect (observed for the first time in Gratiola officinalis L. preparations). Advantages of the proposed preparation are low toxicity, availability of the raw material, and broad spectrum of therapeutic effects.

Bacteriostatic and bactericidal activity of aqueous solution (50 mg/mL) of alcoholic extract of Helichrýsum arenárium (L.) dried flowers, prepared by a special technique so as to increase the yield of flavonoids, was studied in vitro with respect to Mycobacterium tuberculosis (MBT) strains possessing varying degrees of drug resistance, as characterized by replacements Ser R Leu (modification of b-subunit RNA-polymerase of MBT) and Ser R Thr (inactivation of MBT catalase-peroxidase enzyme). The mechanism of this drug action is clearly distinguished from that of the first-line drugs, since strains resistant to these reference drugs have proved susceptible to extract H. arenárium extract. This extract can be recommended for preclinical and clinical studies in the search for new antituberculous drugs and for studying new mechanisms of drug action on MBT. It may also be an effective drug for the treatment of multidrug-resistant MBT strains.

The karyotype of Camptochironomus tentans (Fabricius), 1805 (Diptera, Chironomidae) from the populations of the Lower Volga region and Central Caucasus (the northern macroslope) has been studied. In the karyotype of C. tentans from Saratov oblast, 14 banding sequences (BS), 13 genotypic combinations, and 14 zygotic combinations were identified. Four new BSs were described: ten B12, ten F4, ten F5, and ten G4, which are distinguished by simple paracentric inversions from those already known for the species. The chromosomal polymorphism belongs to the inversion type; its level in the Saratov population is generally the same as in populations of other parts of the habitat. The number of heterozygous inversions per specimen was 1.3; the number of heterozygous inversions per arm was 0.86; the percentage of heterozygous larvae was 84.5. The polymorphic arms in C. tentans were A, B, C, and F. Nine previously known BS were revealed in the karyotype of C. tentans from Central Caucasus, and no new sequences were found. Arm B in C. tentans was polymorphic. The zygotic combinations in all the studied populations were distinct.

Karyotypes of the Chironomus nuditarsis Str. (Chironomidae, Diptera) (Keyl, 1962) chironomids from central Caucasian populations located at different altitudes above the sea level (a.s.l.) were examined. Specific features of the inversion polymorphism of the high-mountain (more than 1000 m a.s.l.), foothill (500-1000 m a.s.l.), and plain (up to 500 m a.s.l.) populations were identified. A new chromosome banding sequence, ndt F2 (lg-e 7a-8c), which was endemic to Caucasian populations, was described. With the increase in the altitude of the reservoir a.s.l., the decrease in the frequency ofndt G1.2 and ndt B2.2 inversions and the number of inversions per individual and per arm was observed. In the high-mountain population, only the ndt G2.2 homozygotes were detected. The plain population is an interstitial population, while foothill and high-mountain populations are terminal. Cytogenetic distances between the high-mountain and other samples range from 0.174 to 0.223, which is higher than the mean interpopulation value for this species (0.138). The allochrony of the life cycle in aborigines from the foothill population and aborigines from the high-mountain population can be caused by factors such as low high-mountain reservoir temperature. This factor increases the duration of the stages of larvae developmental, which leads to a reduced number of generations and results in a shift in the imago flight and mating timing.

Contemporary systematics of insects is based mainly on morphological traits. However, their usage is limited both by high variability and complications in comparisons of remote taxa due to low number of common traits. In whole, this leads to a somewhat subjective view when elaborating the system. Unlike morphological ones, molecular traits of taxa, revealed by use of marker genes such as gene cytochrome-c-oxidase I (COI), are less variable and more uniform, which allows them to be used as a criterion of genus, tribe, and subfamily for a wide range of organisms. Application of molecular criteria appears to be all the more important when constructing the system for groups of organisms with high morphological and specific diversity, such as midges (Chironomidae, Diptera). Last years, the DNA-sequence of gene COI is becoming widely used for species identification as a bar-coding one. Its use as a criterion for taxa of super-species level is hampered by its high nucleotide variability. We established the bounds of COI nucleotide and aminoacid divergence between midge species of Chironominae subfamily belonging to the same genus, same tribe, different tribes, as well as between species of Chironominae and Orthocladiinae subfamilies. It is shown that the level of aminoacid divergence reflects molecular boundaries of genus and tribe better than nucleotide one. It can be stated that if the level of aminoacid divergence falls within the limits from 0 to 1.7% then a pair of species compared belongs to the same genus; if it falls within the limits from 1.7 to 4.0% then they belong to the same tribe; within the limits from 4.6 to 6.3%--to different tribes; if it exceeds 7.9%--to different subfamilies. The accuracy of identification when using these ranges turns out to be not less than 75%. In this regard, bounds of COI sequence aminoacid divergence may be used as taxonomic criteria for midge genus, tribe or subfamily.

This is the first study to infer the phylogenetic structure of minges of the subfamily Chironominae from the amino acid sequence of cytochrome oxidase I (COI). The subdivision of Chironominae into two tribes, Chironomini and Tanytarsini, has been confirmed. The segregation of the genera Pseudochironomus and Riethia into a separate tribe has not been confirmed. Stenochironomus gibbus forms a branch considerably deviating from the subfamily Chironominae. The genus Micropsectra is formed by a large polyphyletic cluster that also includes the genera Virgotanytharsus, Reotanytharsus, Kenopsectra, and Parapsectra. Tanytarsus is the basal genus of the tribe Tanytarsini. The times of divergence of the main taxa of Chironominae have been estimated. The calculated time of divergence of the genus Chironomus disproves the assumption that it is phylogenetically old.

The sequence of a 595-bp fragment of the mitochondrial COI gene was determined for the species Chironomus usenicus and Chironomus curabilis of the genus Chironomus. Phylogenetic reconstructions based on the analysis of the COI gene sequence coincide on the whole with cytogenetic data, permitting Ch. usenicus and Ch. curabilis to be regarded as members of the group plumosus. Chironomus usenicus and Ch. plumosus have identical COI gene sequences. Two hypotheses explaining this identity are considered: inheritance of mtDNA from one of the parental species during hybridogenesis and horizontal transfer of mitochondrial genes.

Morphometric analysis of changes in nucleolar organizer (NO), Balbiani rings (BR)--BR(B), BR(1G), BR(2G) and chromosome I arm B puff activities, and in chromosome compactness of Chironomus plumosus (Diptera) polytene chromosomes was carried out in acute period under separate and combined influence of atropine and pilocarpine. Supression effect of cholinotropic preparations mixture was revealed. Suppression of NO activity with atropine concentration increase in the mixture served as criterion of toxicity.

Karyotype and morphology of Chironomus sp. larvae from the Caspian Sea have been described. 2n = 8. Chromosomal arm combination is AB, CD, EF, G (cytocomplex thummi). All chromosomes display conjugation of homologues. Centromere areas are of s-type. The nucleolar organizer and two Balbiani rings are disposed in arm C, and another Balbiani ring is in arm B. Half larvae have heterozygotic paracentric inversions in arm D. The larva belongs to salinarius form. One scleritis is colorless, an occipital scleritis is light and irregularly colored. Premandible has 4-5 cogs. Epipharingeal ridge has 18-23 dens. This new species is close to salinarius group, especially to Ch. albidus Konst., but differs from it at the larva stage in the structure of non-massive basal antenna segment, in the presence of large quantity of premandible denses, in light color of occipital scleritis, and in a series of morphometric rates. A similarity berween Ch. paraalbidus and Ch. albidus, on the one hand, and Boeotendipes, on the other one, has been found out.

Hybridogenesis as a possible way of speciation in Chironomidae was considered with special reference to the species Chironomus usenicus resulting from hybridization between C. plumosus and C. behningi. The three species had 2n = 8 and belonged to the thummi cytocomplex with chromosome arm combinations AB, CD, EF, and G. Arm G had a marker chromosome disk sequence (CDS) and was used to demonstrate the hybrid origin of C. usenicus. Most C. usenicus larvae were heterozygous in CDS of arm G. CDS use G2 proved to be identical to CDS beh G1 of C. behningi and CDS use G1, to CDS plu G1 of C. plumosus. It was assumed that C. usenicus results from hybridization between eurybiont C. plumosus and stenobiont C. behnigni at the boundary of their species areas, in freshwater or brackish water bodies of the southern Saratov oblast and northern Kazakhstan. Morphologically and karyotypically, the hybrid was probably similar to C. plumosus. Crosses with C. plumosus eliminated virtually all C. behningi chromosome sequences from the karyotype of the hybrid. Further chromosome divergence resulting in C. usenicus involved a number of chromosome rearrangements, including duplication of pericentric heterochromatin and other chromosome regions; inversion, which occurred in arm F (regions 13-16) and was fixed in the karyotype; and other paracentric inversions and deletions accumulated in heterozygote in the karyotype pool of the species. Since C. behningi was eliminated from the introgression zone and its species are reduced, the assimilation character was assumed for introgressive hybridization of C. behningi and C. plumosus.

The karyotype of Chironomus heterodentatus, belonging to the obtusidens-group described by Konstantinov from the Volga in 1956, was studied in detail. Combinations of chromosomal arms are AB, CD, EF and G (cytocomplex thummi). The Ch. heterodentatus karyotype cleary differs from those of other members of the obtusidens-group with a species-specific banding pattern in arms A, B, C and D. Chromosomal polymorphism on homo- and heterozygous inversions was found in arms, A, B, D, E and G. 19 inversion banding sequences and their 27 genotypic combinations have been recorded. The shortest arm G is highly polymorphic. Heterozygotes on the Balbiani ring activity were found in arm G along with homo- and heterozygotes on inversions. B-chromosomes with a frequency equal to 2.7-25.0% were recorded in some Volga populations.