Influence of polyamines of bacterial origin on the production of key cytokines in the culture of human mononuclear leukocytes

To date, participation of biogenic polyamines has been studied in details, with respect to regulation of microbial gene expression, interrelations between bacteria, development of their persistence state. Opportunity of their use as markers of human pathological conditions is being actively evaluated. The aim of our study was to assess the effect of bacterial diamines, i.e., cadaverine and putrescine, upon production of key cytokines (IFNγ and IL-4) in the culture of human mononuclear leukocytes. We studied leukocytes of peripheral venous blood obtained from 18 healthy male volunteers (mean age 24.0±0.6 years). The leukocytes were isolated by means of gradient centrifugation using a Ficoll-Verografin mixture. For the cultivation of lymphocytes, a micro-method and plastic round-bottom 96-well plates were used. Concanavalin A at a concentration of 5 μg/ml was used as a T cell mitogen. Polyamines were used at final concentrations of 5, 25, 50, 75 and 100 μM/L. The cultivation was carried out in humidified atmosphere with 5% CO2, at 37 °C for 72 hours. At the end of incubation, the culture medium was collected and frozen for subsequent quantitative enzyme immunoassays of cytokine concentrations (IFNγ and IL-4) (Russia). Viable cells were counted using Goryaev chamber after staining with 0.1% trypan blue solution. Statistical analysis was performed using Student’s t test or Mann–Whitney test. Addition of cadaverine at all concentrations reduced IFNγ production in the culture of mitogen-activated cells. When culturing leukocytes supplied with putrescine (5 to 50 μM/L), a dose-dependent decrease of IFNγ was observed. Upon further increase of putrescine concentrations, the IFNγ production is restored to the values of the control samples. Direct toxic effect of polyamines upon the cells was revealed. Both the diamines, at the doses of 50, 75, and 100 μM/L, caused increase of IL-4 production by the mitogen-activated cells. Such changes can be associated both with direct cytotoxic effect of cadaverine and putrescine, being mediated by changes of some metabolic pathways. In addition, the effects of polyamines upon monocytes present in culture can include their anti-inflammatory state, e.c., an increased IL-4 production. In general, cadaverine and putrescine, produced by microorganisms of various taxonomic groups, regulate the effectiveness of compensatory-adaptive reactions that ensure adaptation of microbial populations to changing or unfavorable environmental conditions

TECHNICAL ASPECTS OF EVALUATING CYTOKINE PROFILE AND LEUKOCYTE REACTION IN BILE DUCT OBTURATION OF DIFFERENT ORIGIN

The study was carried out to assess the prospects for the combined use of leukocyte and cytokine indices in the bile duct obstruction of different genesis.We have performed a study of the patients with mechanical jaundice, either due to tumor (14 persons), or non-tumor genesis (16 persons). Concentrations of cytokines (IL-1, IL-4, IL-8, TNFα) were determined in blood serum before surgical intervention. The total activity of endogenous inflammatory mediators in the serum of patients was estimated as an index of inflammatory activity (IVA), which was calculated using the following formula: IVA = (IL-1 + TNFα)/IL-4. In addition, a number of leukocyte indices were calculated. The cellular component in the index estimation makes it possible to clarify the intensity of the inflammatory reaction, and to suggest the direction of the functional changes among the  immunocompetent cells.It was shown that IVA in presence of malignancy is significantly higher than in benign conditions. Overactivation of immune response in tumors may be caused by a significant predominance of pro-nflammatory cytokines. Moreover, in cases of malignant tumor growth, both with or without cholangitis, the index scores of humoral and cellular components let us suggest the some problems with regeneration, due to pro-inflammatory domination of immune response, which does not ensure activation of the corresponding reactions. It has been established that the leukocyte intoxication index (LII) is increased,in patients with a combination of malignant disorder and cholangitis, and the immunoreactivity index (MDI) is lowered. In absence of cholangitis, the opposite pattern is observed, when LII is slightly lowered and MDI is elevated. Correlation analysis for the patients with cholangitis revealed a high association of cytokine imbalance and endogenous intoxication. A complex of cytokine and leukocyte indices is promising, when each of them reflects a certain part of the process, and their combination gives an integral picture of the pathology and allows to predict development of the disease. Such an approach, seems to increase the diagnostic significance of these immunological indices, primarily, the cytokine production, and may be used when predicting currence of complications in the surgical treatment of the disease

Биогенные полиамины при генитальной гонококковой инфекции: факты и гипотезы

Genital gonorrhea is one of the most common sexually transmitted diseases with significant gender differences in its clinical course. Laboratory verification of the diagnosis is associated with great difficulties in the cultivation and identification of the pathogen. Moreover, the diagnosis of female gonorrhea is a serious problem due to mild symptoms of the disease. Currently, a promising trend in the diagnosis of inflammatory diseases of reproductive organs is biochemical analysis of vaginal and sperm fluids, which have a rich component composition. Biogenic polyamines can be synthesized by both pro- and eukaryotic cells. These polycations are present in semen and vaginal fluid and can have a significant effect on various cell structures and functions. In this regard, the qualitative and quantitative composition, the level and ratio of these components and their changes can have a diagnostic value for infections of the genital tract.The aim of the review was to analyze current information on the role of biogenic polyamines in the physiological and biochemical potential of Neisseria gonorrhoeae and their participation in the development of genital gonococcal infection, taking into account the influence of sexual differences and a number of related factors. Special attention was paid to the origin and possible functional role of polyamines in the genital tract of men and women. As a result, taking into account the spectrum, origin and ratio of polyamines in the corresponding fluids, we formulated a hypothesis: the manifestation of the process in case of infection in men is largely determined by the reactivity of eukaryotic cells, but not the metabolic activity of the microbiota of the reproductive tract. At the same time, the development of “female” gonorrhea is primarily determined by the state of the microbiocenosis of the cervical vaginal biotope.Генитальная гонорея является одним из наиболее распространенных венерических заболеваний и характеризуется существенными гендерными различиями в его клиническом течении. Лабораторное подтверждение диагноза сопряжено с большими сложностями культивирования и идентификации возбудителя, а диагностика «женской» гонореи представляет серьезную проблему еще и в связи со стертой симптоматикой инфекционного процесса. На современном этапе перспективным направлением для диагностики воспалительных заболеваний репродуктивных органов признается изучение биохимического состава влагалищной и спермальной жидкостей, имеющих богатейший компонентный состав. Биогенные полиамины, которые могут синтезироваться как про-, так и эукариотическими клетками и в значительных количествах обнаруживаться в этих секретах, являются низкомолекулярными соединениями, оказывающими разнообразные эффекты на жизненно важные структуры и функции клеток обоих типов. В этой связи качественный и количественный состав, уровень и соотношение этих компонентов в секретах, с учетом изменения соответствующих показателей в динамике, могут иметь диагностический смысл при инфекционной патологии генитального тракта. Целью обзора явилось рассмотрение накопленной к настоящему времени информации о возможной роли биогенных полиаминов в физиолого-биохимическом потенциале Neisseria gonorrhoeae и их участии в развитии генитальной гонококковой инфекции с учетом влияния половых различий и ряда сопутствующих факторов. Особое внимание уделено происхождению и возможной функциональной роли полиаминов в генитальном тракте мужчин и женщин. В результате, с учетом спектра, происхождения и соотношения полиаминов, доминирующих в составе соответствующих секретов, сформулирована гипотеза о том, что манифестация процесса в случае инфицирования мужчин в большей степени обусловлена реактивностью эукариотических клеток, но не метаболической активностью микробиоты их репродуктивного тракта. В то время как развитие «женской» гонореи в первую очередь определяет состояние микробиоценоза цервикально-вагинального биотопа

Estimation of social, psychosomatic and gynecologic status in women with non-specific vaginitis

There was carried out the estimation of social, gynecologic and psychosomatic status of 50 women with the non-specific vaginitis. The study of the clinical and social parameters was carried out under the specially developed form for evaluating the state of the somatic and reproductive health of women. The characteristics of psycho-emotional status were obtained with the help of the Spielberger’s questionnaire and method of the quantitative assessment of mental phenomena with the application of the scales of the subjective evaluations of functional state (test SAN). Women with non-specific vaginitis were characterized by emotional instability, high level of situational and personal anxiety, which spptomized about psychological and vegetatic disregulation. The revealed special features make it possible to develop the differentiated therapeutic and prophylactic measures for the patients with the non-specific vaginitis.Резюме: Проведена оценка социального, гинекологического и психосоматического статуса 50 женщин с неспецифическим вагинитом. Изучение медико-социальных параметров проведено по специально разработанной анкете для оценки состояния соматического и репродуктивного здоровья женщин. Характеристика психоэмоционального статуса была получена с помощью тестирования пациенток по опроснику Спилбергера - Ханина и метода количественной оценки психических явлений с применением шкал субъективных оценок функционального состояния (тест САН). Показано, что женщин с неспецифическим вагинитом характеризует эмоциональная неустойчивость, высокий уровень ситуативной и личностной тревожности, что может свидетельствовать о наличии психо-вегетативной дизрегуляции. Выявленные особенности позволяют разработать дифференцированные лечебно-профилактические мероприятия для таких пациенток

Влияние комплекса хитозан-ß-циклодекстрин с левофлоксацином на микрофлору ран и толстого кишечника осетра

The authors in the article presented the results of determining the effectiveness of using the chitosan-ß-cyclodextrin complex with levofloxacin in the healing of mechanical wounds of valuable commercial fish sturgeons and their feeding. The experiment was conducted based on the “Progressive biotechnologies in aquaculture” research laboratory of the Saratov State University of Genetics, Biotechnology and Engineering. N.I. Vavilov. The microflora of incised wounds and the large intestine of sturgeon fingerlings under the influence of fluoroquinolone, represented by levofloxacin based on cyclodextrin, included in the shell of high-molecular chitosan, was studied. The studied microbiological indicators were chosen to determine that changes in the total number of microorganisms show the nature of the course of the inflammatory/pathological process, which contributes to the development of microorganisms (including opportunistic pathogens) and lactic acid bacteria in the intestine. Therefore, they are essential physiological indicators of the formation of “ intestinal immunity. It was found that the use of cyclodextrin with levofloxacin in the treatment of incised wounds in sturgeons leads to a significant decrease in the total microbial number (TMC) on their surface (by 10 thousand times compared to the group without treatment). It has been shown that using cyclodextrin with levofloxacin in feeding sturgeons reduces the total microbial number in the large intestine. This complex is characterised by good solubility and bioavailability for fish. The future study results can be used in aquaculture to treat mechanical injuries received during transportation and sorting in the rearing process in fish.Представлены результаты определения эффективности применения комплекса хитозан-ß-циклодекстрин с левофлоксацином в заживлении механических ран ценных промысловых рыб – осетров и их кормлении. Эксперимент проводили на базе научно-исследовательской лаборатории «Прогрессивные биотехнологии в аквакультуре» Саратовского государственного университета генетики, биотехнологии и инженерии им. Н.И. Вавилова. Исследовалась микрофлора резаных ран и толстого кишечника сеголетков осетров под действием фторхинолона, представленного левофлоксацином на основе циклодекстрина, включенного в оболочку высокомолекулярного хитозана. Исследуемые микробиологические показатели были выбраны для определения на том основании, что изменения общего количества микроорганизмов показывают характер течения воспалительного/патологического процесса, способствующего развитию микроорганизмов (в т.ч. условно-патогенных) и молочнокислых бактерий в кишечнике, поэтому являются важными физиологическими показателями формирования «кишечного иммунитета». Было обнаружено, что применение при лечении резаных ран у осетров циклодекстрина с левофлоксацином приводит к значительному уменьшению общего микробного числа (ОМЧ) на их поверхности (в 10 тыс. раз по сравнению с группой без лечения). Показано, что использование в кормлении осетров циклодекстрина с левофлоксацином также снижает общее микробное число в толстом кишечнике. Данный комплекс характеризуется хорошей растворимостью и биодоступностью для рыб. Результаты исследования в перспективе могут найти применение в аквакультуре при лечении механических травм, полученных при перевозке и сортировке в процессе выращивания, у рыб

The Microbiota Continuum along the Reproductive Tract in Women with Infertility

Introduction. At present, the question of commensal, including opportunistic, microflora participation in  infertility development remains debatable. In a number of studies, the translocation of the vaginal microflora into the endometrial tissue is considered as a factor contributing to inflammation development. In addition, the connection of some reproductive losses with the persistence of certain conditionally pathogenic microorganisms is shown. Today, to solve this issue, molecular genetic research methods are being actively introduced that surpass the routine cultivation techniques in a number of positions.The aim of the study was to assess the taxonomic diversity of microorganisms in the vaginal biotope with infertility.Material and methods. For the study samples of vaginal contents obtained from the posterior vaginal fornix of 15 women, consisting of barren marriage, were used. A metagenomic study of 16S ribosomal RNA samples was carried out on the Illumina MiSeq platform, using the MiSeq Reagent Kits v3 kit (600-Cycle Kit), as recommended by the manufacturer. Libraries for sequencing plots of the V3-V4 gene of the 16S ribosomal RNA were prepared according to the 16S Metagenomic Sequencing Library Preparation Illumina. In bioinformatics assessment, Kraken Metagenomics version 2.0.0 software for metagenomic analysis (classifier of reads — short nucleotide sequences) was used using a standard database.Results. It was shown that the occurrence of representatives of the Lactobacillaceae family in the vaginal biotope varies from 12 to 84%. The genus Lactobacillus with the dominance of L. jensenii, L. delbrueckii and L. amylolyticus occupied the leading position among the members of the family. In all samples, Moraxella spp. was found in large numbers, with M. osloensis leading among the representatives of the entire community. In half of the cases, the joint presence of M. osloensis and G. vaginalis was revealed.Conclusion. It has been established that in infertility in the vaginal microbiota the number of representatives of the genus Lactobacillus is significantly reduced, and there is also a change in the leading species to L. jensenii, whose functional activity does not fully ensure the colonization resistance of the vaginal biotope, allowing for excessive reproduction of oppotrunistic microorganisms, in particular, M. osloensis

Prevalence and Possible Causes of Infertility in the Perm Region

The aim of investigation was to assess the characteristics of the epidemic process of the incidence and prevalence of infertility and gonococcal infection, taking into account the demographic situation in Perm region.Materials and methods. The official statistics of the Center for Hygiene and Epidemiology in the Perm Region for 2003–2017 were analyzed (form 2), the territorial body of the Federal State Statistics Service in the Perm Region. The long-term dynamics of general morbidity, distribution of the patients by age, sex, social status, morbidity in combination with HIV infection, the detection of gonococcal infection by various specialists and methods of laboratory diagnostics were studied using the method of retrospective epidemiological analysis. Statistical processing was carried out using the program Statistica 7.0. To determine the relationship between the individual parameters, a simple correlation analysis was used with the calculation of the conjugacy coefficient of the Pearson traits (r).Results. In the Perm region, despite the positive rate of natural population growth, the number of inhabitants in 2010-2016 decreased by 14.5 thousand people. Every year the number of women in adolescent and youthful age falls. It has been shown that the leading factor in the formation of infertility is inflammatory diseases of the reproductive organs and the presence of comorbid pathology. It was established leading value in the pathogenesis of infertility of gonococcal infection. The features of the epidemic process of gonorrhea in the Perm region was shown.Conclusion. In a large number of cases, the complication of the demographic situation in the province is associated with infection of the genital tract. It is necessary to use more widely the possibilities of microbiological diagnostics