[Cytokines level in patients with drug-induced jaw necrosis].

The study included 15 patients with purulent inflammatory diseases of maxillofacial area and 25 patients with facial bone necrosis induced by synthetic drugs. Pro- and anti-inflammatory cytokines levels in saliva and wound fluid were analyzed in two groups. The results proved cytokines to play important role in jaw necrosis induced by drugs containing red phosphorus.

[Defensins level in patients with inflammatory diseases of maxillofacial area].

Molecular study of congenital immune factors was conducted in 45 patients with inflammatory diseases of maxillofacial area. The study focused on expression of cationic low-molecular peptides α- and Β-defensines in oral mucosa. These peptides are involved in antibacterial activity and regulation of immune reactions. The results showed 20-fold increase in defensines expression in oral mucosa of patients with inflammatory diseases of maxillofacial area when compared to control group (12 healthy individuals) thus proving an important role they play in the development of the disease. Therapy with cytokines may be indicated in these cases.

[Evaluation of expression of innate immunity component genes in mice leukocytes under the effect of synthetic ligands in vivo].

AIM: Study the effect of synthetic ligands on the dynamics of TLR9 and BD-2 gene expression in vivo. MATERIALS AND METHODS: Synthetic ligands were used whose sequence was homologous to DNA and RNA human viruses. At days 1, 7 and 28 blood sampling from BALB/c line mice was carried out. Determination of level of TLR9 and BD-2 gene expression was evaluated by using RT-PCR in real-time mode. RESULTS: The following ligands had the maximum effectiveness: tgg-ccc-ccc-ttg-tgg-acc-gg, ccg-gtc-cac-aag-ggg-ggc-ca and tcg-tcg-ttt-tgt-cgt-tgt-cg. At day 1 after the administration of ligands a significant increase of TLR9 gene expression was observed (62, 18 and 13 times, respectively), but at the same time defensin gene expression changed insignificantly. CONCLUSION: The approach developed may be used to evaluate the effect of these preparations on the system of innate immunity, namely TLR activation.

[Study of expression of cytokine genes in the process of cultivation of healthy donor leukocytes].

AIM: Study of expression of cytokine genes in the process of cultivation of healthy donor leukocytes. MATERIALS AND METHODS: RNA isolated before, after 3 and 24 hours of cultivation of leukocytes of 15 healthy donors aged 19 - 32 years was the object of the study. Study of the expression of genes of 8 cytokines by the level of their mRNA was performed by using polymerase chain reactionwith reverse transcription. RESULTS: In healthy donors among with individual differences, general regularities were traced: higher levels of IFNalpha and anti-inflammatory cytokine gene expression, as well as predominantly low expression of pro-inflammatory cytokines. CONCLUSION: In most of the cases in the process of cultivation of leukocytes isolated from blood donors the highest level of gene expression of pro-inflammatory cytokines is noted after 3 hours of cultivation, and expression of genes of anti-inflammatory cytokines remains constant for 24 hours of cultivation.

[Changes in expression level of TLR9 and BD-2 genes in cornea of mice with viral keratitis].

AIM: Study of innate immunity genes expression (TLR9 and BD-2) in epithelial cells of mice cornea on the model of viral keratitis during the application of immunotherapy. MATERIALS AND METHODS: RNA isolation from cornea cells was performed by using RNeasy Mini Kit (Qiagen) and "RIBO-sorb" (ILS, Russia) reagent kits. "Kit for performing PCR-RT in the presence of intercalating dye SYBR Green I" (Syntol, Russia) was used for performing PCR. Data on gene expression is presented in decimal logarithms (relatively to 1 million of beta-actin gene copies). RESULTS: In C57B1/6 mice line at day 1 post infection the virus was detected in 33% of animals, at 3 days - in 90%, at 7 days the number of infected mice started to decline. In cornea of the mice infected with HSV-1 a significant decrease of TLR9 gene expression was observed. Administration of a complex of natural cytokines and antimicrobial peptides (Superlimf preparation) caused statistically significant increase of expression levels of the studied TLR9 and BD-2 genes. CONCLUSION: The model of viral keratitis and expression analysis of recognizing receptor TLR9 and defensin (BD-2) genes can be used to study mechanism of action of various preparations during infection in eye tissues and evaluation of possibility of their use in complex therapy of ophthalmic herpes.

[Effect of combined therapy using complex of natural cytokines and antimicrobial peptides in urogenital infections caused by Chlamydia and Mycoplasma].

AIM: To study efficacy of complex therapy of urogenital infections caused by Chlamydia and Mycoplasma using immunomodulator Superlimph. MATERIALS AND METHODS: Fifty males and thirty six females ages 22 - 47 years old with chronic urogenital infections--cervicitis and vulvovaginitis (females), prostatitis (males)--were studied. PCR and bacteriologic methods were used for diagnostics of mixed infection and microbiota efficiency. Patients were divided on 3 groups according to treatment protocol. Twenty patients (group 1)--standard therapy (josamycin), 48 patients received immunomodulator (suppositorium) before treatment with josamycin (group 2), 18 patients were simultaneously treated with josamycin and immunomodulator (group 3). RESULTS: Combinations of Chlamydia trachomatis, Ureaplasma urealyticum, Gardnerella vaginalis and Mycoplasma genitalium were identified in 30 - 35% of cases before treatment. After treatment with josamycin (group 1) or simultaneous therapy with josamycin and immunomodulator (group 3) considerable suppression of growth and elimination of both main pathogens and members of microbiota. Use of immunomodulator (group 2) in some cases resulted elimination of main pathogens and associated opportunistic microflora. CONCLUSION: Microbiological monitoring in 97% cases demonstrated therapeutic effect of immunomodulator Superlymph on urogenital infections associated with Chlamydia and Mycoplasma and disbiotic microbiota of urogenital tract.

[Expression of the antimicrobial peptide beta-defensin-2 in the conjunctival epithelial cells in primary open-angle glaucoma and over time in the postoperative period].

The time course of changes in the postoperative expression of the antimicrobial peptide beta-defensin-2 (HBD-2) in the conjunctival epithelial cells was studied in 13 patients (13 eyes) with primary open-angle glaucoma (POAG) and 9 individuals (9 eyes) without ophthalmopathology. Scrapes from the eyelid conjunctiva and transient folds were used as a material for an immunological study. In the study group, the scrapes were taken a few hours before and at 7, 30, and 90 days after surgery. The expression of the HBD-2 gene in the study material was determined by real-time polymerase chain reaction. In the subjects without pathology of the organ of vision, the conjunctival epithelial cells exhibited HBD-2; in the patients with POAG, the baseline HBD-2 gene expression was significantly higher. The baseline HBD-2 expression did not depend on gender, the duration of local antihypertensive therapy, the stage of disease stage, and the level of ophthalmotonus. HBD-2 gene expression was by 5-10 times higher than that preoperatively in all the patients with POAG on postoperative day 7. One month after surgery, the HBD-2 gene expression considerably rose in 31% of the examined patients while it became normal in 38% of the patients.

[Regulatory effect of complex of natural cytokines and antimicrobial peptides on murine macrophages infected with multidrug-resistant Mycobacterium tuberculosis].

AIM: To study the effect of complex of natural cytokines and antimicrobial peptides (CNCAP) included in preparation Superlymph on growth of multidrug-resistant Mycobacterium tuberculosis CN-37 on the modem of murine peritoneal macrophages (MPh) cultivated in vitro. MATERIALS AND METHODS: Effect of CNCAP on peritoneal MPh of tuberculosis-susceptible mice C57BL/6 infected by M. tuberculosis CN-37 was studied using ex vivo model. Macrophages were preliminary incubated with CNCAP during one day. M. tuberculosis growth was assessed on 7th day by PCR. RESULTS: Preliminary incubation of infected MPh with CNCAP resulted in inhibition of M. tuberculosis CN-37 growth. CONCLUSION: Superlymph activates macrophages which lead to enhanced bactericidal action of MPh on M. tuberculosis CN-37.

[Changes in various immunological parameters of lacrimal fluid in excessive scarring after antiglaucoma surgery in patients with primary open-angle glaucoma].

Twenty-two patients in whom the expression of the human beta-defensin-2 (HBD-2) gene and the concentrations of human neutrophil peptides (HNP 1-3) and interleukin-17 (IL-17) were determined in the conjunctiva and lacrimal fluid were followed. In patients with primary open-angle glaucoma, the baseline concentrations of HNP 1-3 and IL-17 in lacrimal fluid and the expression of the HBD-2 gene in the conjunctival epithelial cells were higher than those in healthy individuals. The appearance of signs of pathological scarring was observed when HBD-2 expression in the conjunctival epithelial cells was lower and the rate of changes in the lacrimal fluid concentrations of HNP 1-3 and IL-17 was stable.

[TLR9 and HBD-2 gene expression by conjunctival and corneal cells in children with dendriform herpetic keratitis].

The current views of the role of innate immunity in protecting the organ of vision from viral infection offer scope for a search for new approaches to diagnosing and treating viral keratitis. TLR9 and HBD-2 gene expression in the conjunctiva and cornea was studied in children with dendriform herpetic keratitis. The latter showed a significant (more than 20-fold) increase in TLR9 gene expression as compared in healthy children. Conventional antiviral therapy for herpetic keratitis resulted in a significant decrease in the level of TLR9 gene expression (by more than two orders of magnitude). The use of the immunomodulator Superlymph in complex treatment normalized the level of TLR9 gene expression in the conjunctiva and cornea; moreover, there was a rapid relief of inflammation signs, an increase in visual acuity, and a reduction in the duration of treatment in patients with herpetic keratitis.

[Changes of expression levels of innate immunity signaling receptors during Candida albicans infections in vitro and in vivo].

AIM: To study levels of expression of Toll-like receptors genes in response to Candida albicans antigens in vitro (using Vero cell line as well as mononuclear cells) and in vivo (using cervical canal cells of pregnant women). MATERIALS AND METHODS: Test-systems for measurement of expression levels of such genes as TLR 1, TLR2, TLR6 as well as system for quantitative measurement of tumor necrosis factor a level, all of which were developed earlier, were used. RESULTS: It was shown that antigens of C. albicans stimulated early increase of expression of innate immunity genes both in studied in vitro models and in cells of cervical canal of pregnant women with candidosis. CONCLUSION: Results of performed study allow to suggest that activation of innate immunity factors resulted from Candida albicans infection.

[Role of microRNA in regulation of innate immunity mechanisms].

New class of regulatory molecules--microRNA--is of special interest now. Large amount of data about function of these molecules in immune response regulation is accumulated. Functions of several microRNA molecules (miRNA-146, miRNA-155, miRNA-223) contributing to such mechanisms of innate immunity regulation as transmission of signals from receptors, maturation, differentiation and proliferation of innate immunity cells are described in this review. It is necessary to underline the perspective of these molecules for development of drugs for correction of innate immunity abnormalities.

[Study of expression of TLR9, NF-kappaB, TNFalpha genes in cells of cervical canal mucosa in pregnant women with herpesvirus infection].

AIM: To study changes in expression of TLR9, NF-kappaB, TNFalpha genes in cervical canal mucosa in pregnant women with herpesvirus infection. MATERIALS AND METHODS: Using previously developed real-time polymerase chain reaction assays, levels of expression of genes, which corresponding proteins play important role in TLR9-dependent signaling pathway, was quantified. RESULTS: Two-fold, 4.4-fold, and 3-fold rise in expression of TLR9, NF-kappaB, and TNFalpha genes respectively was revealed in pregnant women with herpesvirus infection compared to women with physiologically occurring pregnancy. CONCLUSION: Obtained results demonstrated the activation of TLR-mediated mechanisms of innate immunity on the level of cervical canal mucosa during pregnancy complicated by viral infection.

[Toll-like receptors recognizing ligands of herpesvirus].

Innate immunity is the first line of host defense against infection including those caused by herpesviruses. Toll-like receptors (TLRs) are the main pathogen-recognizing receptors. TLR2 binds to glycoproteins of herpesvirus virion; intracellular TLR9 recognizes nonmethylated CpG repeats of herpesvirus' DNA, which lead to activation of receptor signaling pathways and synthesis of proinflammatory cytokines, chemokines, antimicrobial peptides and other molecules, which participate in induction of innate immunity mechanisms. Many viruses (measles virus, cytomegalovirus etc.) activate expression of TLRs for more effective replication of the virus in cells.

[Cytokine therapy in the regulation of corneal reparative processes: an experimental and clinical study].

Cytokine monotherapy with Superlymph used in combination with the keratoprotector Applicoll in the treatment of superficial corneal layer defect resulted in rapidly alleviated inflammatory reactions, accelerated reparative processes, more qualitative epithelial defect recovery, by forming a finer scar, without opacity whereas the traditional treatment led to long-term healing and keratoleukoma in an experiment. The use of cytokine therapy with Superlymph in the complex treatment of corneal injuries and inflammation in children promoted a prompt and stable arrest of a local inflammatory process, accelerated epithelization, reduced opacity intensity, higher visual acuity, and shorter treatment.

[Role of Toll-like receptors and defensins in antimicrobial protection of urogenital tract in females].

Levels of expression of hBD-1 gene (beta-defensin 1) and Toll-like receptors (TLR1, TLR2, TLR6) in cells of cervical mucosa in healthy nonpregnant and healthy pregnant women as well as in pregnant women with urogenital infection was measured by developed RT-PCR systems. During normal pregnancy compared with nonpregnant women, increase of TLRs genes expression which was correlated with increase of hBD-1 gene expression was observed. During urogenital infection in pregnant women compared with healthy pregnant, 10- fold and 50-fold increase of TLR1 and TLR2 genes expression respectively was associated with 2.5-fold decrease of hBD-1 gene expression in cervical mucosa. In group of women with untrauterine infection more marked increase of TLRs genes expression was observed. Thus significant changes (TLRs, antimicrobial peptides, cytokines etc.) in cells of cervical mucosa can be used as prognostic criteria for development of intrauterine infection.

[Inhibiting effect of complex of natural cytokines and cationic antimicrobial peptides on the growth of Mycobacterium tuberculosis H37Rv in vitro].

AIM: To assess direct antimicrobial effect of complex of natural cytokines (CNC) and antimicrobial peptides (Syperlymph preparation; CNC) on Mycobacterium tuberculosis H37Rv (H37Rv) and effect mediated by macrophages (MP) treated with the preparation. MATERIALS AND METHODS: Direct effect of CNC was studied during cultivation of H37Rv in the presence of preparation, whereas indirect effects--during simultaneous cultivation of H37Rv and mice peritoneal MP C57B1/6. Assessment of growth was performed on the 7th day using PCR. RESULTS: It was shown that CNC directly inhibits growth of H37Rv in vitro. Cultivation of H37Rv in culture of MP resulted in inhibition of M. tuberculosis. The most evident inhibition was noted after extension of time of preliminary treatment of MP with Syperlymph and simultaneous increase of its concentration. CONCLUSION: Antimycobacterial effect of Syperlymph preparation related to complex effect of cytokines and antimicrobial peptides directed to M. tuberculosis and macrophages, which forms the conditions for killing of mycobacteria.

[Analysis of cytokines in seminal plasma and blood sera of patients with chronic prostatitis during immunotherapy with natural complex of cytokines and antimicrobial peptides].

Levels of pro- and anti-inflammatory cytokines (TNFalpha, IL-6, IL-8, IL-10) were studied in blood sera and seminal plasma of healthy volunteers and patients with chronic bacterial prostatitis (CBP) or chronic abacterial prostatitis (CABP). Assessment of effect of immunotropic drug Superlymph with direct antimicrobial action for treatment of mentioned groups of patients was performed. It was shown that seminal plasma of patients with CBP and CABP contained higher levels of IL-6 compared with healthy subjects. IL-8 level was increased in small part of patients with CBP and CABP. Changes in cytokine status of patients with CBP and CABP that occurred during treatment and manifested in decrease of proinflammatory cytokines levels (IL-6 and IL-8) and increase of antiinflammatory cytokine level (IL-10) point to reduction of inflammatory process in prostate. Clinical effect of complex treatment in patients with CBP considering eradication of pathogen and shortening of duration of antibacterial treatment amounted 95%. Monotherapy of patients with CABP with Superlymph was effective in 72%.

[Natural immunity and herpes infection].

The review details natural immunological factors that are the first-line protection from herpes simplex virus. These reactions may be conventionally divided into 3 active phases. In Phase 1, the virus is attacked by proteins, such as complement of precursors of natural class M (IgM) antibodies and antimicrobial peptides. In Phase 2, the interferons produced by infected epithelial and resident dendritic cells go into action. And, finally, in Phase 3, effector cells, such as neutrophils, macrophages, and natural killer cells, show antiviral activity.