Production Of Immunoregulatory Cytokines Research Articles

IMMUNOMODULATORY ACTIVITY OF THE LACTOBACILLUS DELBRUECKII SUBSP. BULGARICUS LB 86 ENZYME LYSATE

The aim of the study was to immunomodulatory activity of the enzyme lysate of probiotic culture Lactobacillus delbrueckii subsp. bulgaricus LB86 for the development of a functionally active substance based on it with a deliberate action forapplication in cosmetology and medicine. In vitro and in vivo experiments, using an experimental model of intact BALB/c mice, investigated the effect of enzyme lysate on the functional activity of animal peritoneal exudate macrophages and the production of immunoregulatory cytokines, using conventional research methods. The enzyme lysate of L. delbrueckii subsp. bulgaricus LB86 showed immunomodulatory activity aimed at altering the activity of macrophages, as well as the production of pro- and anti-inflammatory cytokines, which are involved in balancing the immune response of the Th1- and Th2- type. Under the action of this enzyme lysate in vitro, the indicator of phagocytosis (PhI) of the peritoneal cavity macrophages of intact BALB/c mice was enhanced and after its intragastric administration to these mice, the increase of the parameters of spontaneous and stimulated NBT test of macrophages was observed and increase (on the 6th day after its introduction) in the levels of IL-12 and IFN-γ in the blood serum of mice treated with this enzyme lysate was noted against the background of an increase in the ratio of INF-γ/IL-4, which also indicated its potential ability to activate the cellular immune response. It has been established that the enzyme lysate of L. delbrueckii subsp. bulgaricus LB86 after intragastric administration to intact mice does not directly participate in modulation of intestinal microbiota and throughout the entire observation period in the intestinal contents the number of lacto- and bifidobacteria in the intestinal contents was maintained at the level of control indicators. The number of aerobic and optional anaerobic bacteria, including staphylococci, streptococci, colimorphic bacteria and microscopic fungi also did not change. Obtained data of immunomodulatory activity of the enzyme lysate of L. delbrueckii subsp. bulgaricus LB86 may indicate the potential prospects for its practical use in therapeutic and prophylactic drugs.

Impact of Carao (Cassia grandis) on Lactobacillus plantarum Immunomodulatory and Probiotic Capacity

Lactobacillus plantarum has beneficial effects on the reduction of symptoms of poor lactose digestion and hypercholesterolemia, removal of the duration and severity of diarrheal processes, improvement of the intestinal permeability barrier, prevention of some types of cancer by adsorption or inactivation of genotoxic agents, increased resistance to intestinal and extraintestinal infections, attenuation of inflammatory bowel disease, and prevention of allergies (especially food). On the other hand, carao (Cassia grandis) has shown remarkable nutritious content with influential dietary applications. As a result, this investigation aimed to explore the effect of Cassia grandis pulp on viability of Lactobacillus plantarum under gastrointestinal conditions, immunomodulatory capacity, and probiotic potential. Adding carao to the medium under different experimental conditions, including rich and minimal culture media and a gastrointestinal digestion process of skimmed milk, did not substantially affect Lactobacillus plantarum’s growth but prolonged its viability. The administration of Lactobacillus plantarum with carao in mice did not induce a proinflammatory response at a systemic level. Still, it did cause an increase in the production of immunoregulatory cytokines. Also, the viability of TSB broth was improved by adding carao. Carao improved the growth of acid tolerance, bile tolerance, growth in TSB broth, and NaCl resistance. According to the results, carao may enhance the characteristics of L. plantarum when enriching fermented dairy products.

Tissue factor: A link between metastatic colorectal cancer and thrombosis in patients in the CALGB (Alliance)/SWOG 80405 trial.

198 Background: Tissue factor (TF) is a transmembrane protein that plays a crucial role in thrombosis. TF expression is elevated in various types of cancer, including colorectal cancer (CRC) and is associated with more aggressive disease and poor prognosis. The aim of this study was to evaluate TF expression in patients (pts) with metastatic CRC (mCRC) enrolled in the CALGB (Alliance)/SWOG 80405 clinical trial. Understanding the relationship between TF expression and mCRC will provide insights into the role of coagulation in the progression and prognosis of this disease and may have therapeutic implications, particularly in the setting of recent approval of a TF antibody drug conjugate for metastatic cervical cancer and other agents in the pipeline. Methods: 433 pts with mCRC treated with bevacizumab (Bev, n = 226) or cetuximab (Cet, n =207) in combination with first-line chemotherapy in CALGB/SWOG 80405 were included in the analysis. TF RNA was isolated from FFPE tumor samples and sequenced on the HiSeq 2500 (Illumina). Overall survival (OS) and progression-free survival (PFS) were compared between groups of pts categorized by tertiles of TF expression: high (H), medium (M) and low (L). Sensitivity analyses were conducted after stratifying by side. Logrank P-values describe differences without adjustment for pt characteristics. Transcriptome-wide gene association analysis was performed using linear regression, adjusting for age, sex, ethnicity, ECOG PS, location, number of metastases, KRAS, MSI status, treatment with FOLFOX or FOLFIRI, and the first 3 principal components from the RNA-seq data. Gene Ontology enrichment analysis was conducted with the top 100 TF-associated genes. Results: TF expression is associated with genes related to maintenance of epithelium, cell adhesion, and migration, immunoregulatory cytokine production, and metabolic, HER2, and MAP/Kinase pathways. TF-L showed significantly longer median (m)PFS (13.4 vs 10.1 vs 9.2 months (mo), p=0.0022) and mOS (35.4 vs 30.9 vs 25.2, p=0.0051) in the entire cohort. In Cet treated pts, TF-L had significantly longer mOS (41.3 vs 29.9 vs 26.1 mo, p=0.0013). In pts treated with FOLFOX, TF-L had significantly longer PFS (13.2 vs 10.9 vs 8.2 mo, p=0.0014) and OS (33.4 vs 30.9 vs 22.4 mo, p=0.0044). Pts with liver metastases and TF-H had worse PFS (9.2 vs 9.9 vs 13.5 mo, p=0.006) and OS (23.6 vs 29.9 vs 35.1 mo, p=0.016). Conclusions: Our results suggest that pts with TF-L tumors with liver metastases have improved prognosis compared to pts with TF-H and may benefit from FOLFOX and Cet-based treatment. Findings provide rationale for further evaluation of TF as a predictive biomarker and potential therapeutic target in CRC. Trial Identifier NCT00265850. Support: P30CA014089, U10CA180821, U10CA180882, U10CA180820 and UG1CA233277; UG1CA180830, U10CA180888; Pfizer, Genentech. https://acknowledgments.alliancefound.org.

Mesenchymal Stem/Stromal Cells Induce Myeloid-Derived Suppressor Cells in the Bone Marrow via the Activation of the c-Jun N-Terminal Kinase Signaling Pathway.

Our previous study demonstrated that mesenchymal stem/stromal cells (MSCs) induce the differentiation of myeloid-derived suppressor cells (MDSCs) in the bone marrow (BM) under inflammatory conditions. In this study, we aimed to investigate the signaling pathway involved. RNA-seq revealed that the mitogen-activated protein kinase (MAPK) pathway exhibited the highest number of upregulated genes in MSC-induced MDSCs. Western blot analysis confirmed the strong phosphorylation of c-Jun N-terminal kinase (JNK) in BM cells cocultured with MSCs under granulocyte-macrophage colony-stimulating factor stimulation, whereas p38 kinase activation remained unchanged in MSC-cocultured BM cells. JNK inhibition by SP600125 abolished the expression of Arg1 and Nos2, hallmark genes of MDSCs, as well as Hif1a, a molecule mediating monocyte functional reprogramming toward a suppressive phenotype, in MSC-cocultured BM cells. JNK inhibition also abrogated the effects of MSCs on the production of TGF-β1, TGF-β2 and IL-10 in BM cells. Furthermore, JNK inhibition increased Tnfa expression, while suppressing IL-10 production, in MSC-cocultured BM cells in response to lipopolysaccharides. Collectively, our results suggest that MSCs induce MDSC differentiation and promote immunoregulatory cytokine production in BM cells during inflammation, at least in part, through the activation of the JNK-MAPK signaling pathway.

Optimization of Selectivity and Pharmacokinetic Properties of Salt-Inducible Kinase Inhibitors that Led to the Discovery of Pan-SIK Inhibitor GLPG3312.

Salt-inducible kinases (SIKs) SIK1, SIK2, and SIK3 are serine/threonine kinases and form a subfamily of the protein kinase AMP-activated protein kinase (AMPK) family. Inhibition of SIKs in stimulated innate immune cells and mouse models has been associated with a dual mechanism of action consisting of a reduction of pro-inflammatory cytokines and an increase of immunoregulatory cytokine production, suggesting a therapeutic potential for inflammatory diseases. Following a high-throughput screening campaign, subsequent hit to lead optimization through synthesis, structure-activity relationship, kinome selectivity, and pharmacokinetic investigations led to the discovery of clinical candidate GLPG3312 (compound 28), a potent and selective pan-SIK inhibitor (IC50: 2.0 nM for SIK1, 0.7 nM for SIK2, and 0.6 nM for SIK3). Characterization of the first human SIK3 crystal structure provided an understanding of the binding mode and kinome selectivity of the chemical series. GLPG3312 demonstrated both anti-inflammatory and immunoregulatory activities in vitro in human primary myeloid cells and in vivo in mouse models.

Sitagliptin Induces Tolerogenic Human Dendritic Cells

Sitagliptin, an anti-diabetic drug, is a dipeptidyl peptidase (DPP)-4/CD26 inhibitor with additional anti-inflammatory and immunomodulatory properties. In this study, we investigated for the first time the effect of sitagliptin on the differentiation and functions of human dendritic cells generated from monocytes (MoDCs) for 4 days using the standard GM-CSF/IL-4 procedure. LPS/IFN-γ treatment for an additional 24 h was used for maturation induction of MoDCs. Sitagliptin was added at the highest non-cytotoxic concentration (500 µg/mL) either at the beginning (sita 0d protocol) or after MoDC differentiation (sita 4d protocol). Sitagliptin impaired differentiation and maturation of MoDCs as judged with the lower expression of CD40, CD83, CD86, NLRP3, and HLA-DR, retention of CD14 expression, and inhibited production of IL-β, IL-12p70, IL-23, and IL-27. In contrast, the expression of CD26, tolerogenic DC markers (ILT4 and IDO1), and production of immunoregulatory cytokines (IL-10 and TGF-β) were increased. Generally, the sita 0d protocol was more efficient. Sitagliptin-treated MoDCs were poorer allostimulators of T-cells in MoDC/T-cell co-culture and inhibited Th1 and Th17 but augmented Th2 and Treg responses. Tolerogenic properties of sitagliptin-treated MoDCs were additionally confirmed by an increased frequency of CD4+CD25+CD127- FoxP3+ Tregs and Tr1 cells (CD4+IL-10+FoxP3-) in MoDC/T-cell co-culture. The differentiation of IL-10+ and TGF-β+ Tregs depended on the sitagliptin protocol used. A Western blot analysis showed that sitagliptin inhibited p65 expression of NF-kB and p38MAPK during the maturation of MoDCs. In conclusion, sitagliptin induces differentiation of tolerogenic DCs, and the effect is important when considering sitagliptin for treating autoimmune diseases and allotransplant rejection.

Toxocara canis extract fractions promote mainly the production of Th1 and regulatory cytokines by human leukocytes in vitro

Helminths possibly down-modulate immune responses to airborne allergens through the induction of a regulatory network. The identification of helminths bioactive molecules is highly desirable, given their immunomodulatory potential which could be used in immunotherapies for allergy and autoimmune diseases. To investigate the immunoregulatory potential of the adult Toxocara canis crude extract and ten protein fractions of its extract, human peripheral blood mononuclear cells (PBMC) from 10 allergic and 9 non-allergic individuals were cultivated, in vitro, in the presence or absence of these antigens, and their supernatants were evaluated for cytokine production (TGF-β, IL-10, IL-12, TNF-α, IL-6, IL-5, IL13, and IL-17). To determine the cell viability, the PBMC were cultivated for 24 h in the presence of the antigens and, following, they were subjected to a cytotoxicity assay. The viability of the PBMC was not affected by incubation with the T. canis antigens. As some fractions stimulated the production of immunoregulatory (TGF-β and/or IL-10), IL-12 and Th1 (TNF-α) cytokines, without stimulating Th2 cytokines (IL-5 and IL13) and IL-17, it was proposed that they would be potential candidates for further studies, especially involving the purification and characterization of specific proteins, which could be tested separately to evaluate their specific role as adjuvants in immunotherapy for inflammatory diseases.

Assessment of the Correlation Between Energy Expenditure and Muscular Mass with Production of Immunoregulatory Cytokines in Ice Hockey Players

Strenuous physical activity may impact on immune response. The goal of the investigation was to study the impact of an endurance sport on the production of immunoregulatory cytokines and the correlation between cytokine production and resting energy expenditure (REE) and parameters of body composition of ice hockey players.

Optimization of the dental implantation protocol in patients with partial secondary edentulousness and vitamin D deficiency

Relevance. The processes of osseointegration in the area of dental implant placement are accompanied by the development of reactions associ-ated with inflammation and activation of local immune processes in peri-implant tissues. Vitamin D preparations can have a significant local effect on the cells of the innate immune defense of the periodontal tissues.The aim of this work is to study the effectiveness of local application of vitamin D on the immune status of the peri-implant sulcus in rehabilitation of patients with partial secondary edentulous and chronic periodontitis by the method of dental implantation.Materials and methods.Determination of the content of biomarkers (IL-1RA, MCP-1, VEGF, TGF-β1, sTNFR) in the crevicular fluid before the installation of implants and the peri-implantation furrow fluid was carried out by the method of enzyme-linked immunosorbent assay using commercial kits «Vector Best» reagents after 3 and 6 months after surgery. The assessment of the state of the bone tissue was carried out using the Orthopantomograph tm OP300 apparatus (KAVO Dental, Germany). Vitamin D has been used topically at a therapeutic dosage.Results.Vitamin D, by reducing Th-1 and Th-17 cell proliferation and differentiation, blocks the activity of the immune-inflammatory process that occurs during implant placement.Conclusion. Application of colecalciferol in patients with moderate periodontitis leads to the activation of the production of immunoregulatory cyto-kines by the cellular structures of the peri-implantation furrow and can be used in combination with the traditional protocol in this category of patients.

Immunotropic effects of vitamin D3 in original rectal suppositories in experimental ulcerative colitis

Increased incidence of ulcerative colitis (UC) is a prerequisite for searching new therapeutic approaches, primarily with an opportunity of site-directed impact on the colon lesion. UC pathogenesis is associated with dysregulated immune response, and limited effectiveness of basic therapy for the disorder. Vitamin D3 exhibits antioxidant, anti-inflammatory, immunomodulatory and other properties, it has been shown to be effective in some autoimmune diseases, thus prompting us to study its effect on immune status in UC. We aimed for studying the effect of vitamin D3, as a component of original rectal suppositories, upon clinical course and indexes of immune status in experimental UC. UC in rats was modeled with 3% oxazolone solution. The vitamin D3-containing suppositories (1500 IU) weighing 300 mg were administered per rectum every 12 hours for 6 days. On days 2, 4 and 6 of UC, the clinical features were assessed as well as blood leukocyte counts, numbers of CD3+, CD45RA+; absorbing and NBT-reducing abilities of blood neutrophils were determined; IgM, IgG, IL-6 and IL-8 concentrations in serum were also studied.The DAI index increased in non-treated UC, along with raised neutrophil numbers in blood, their absorption and NBT-reducing activity was also increased, the total number of lymphocytes, including CD3+, CD45RA+became higher, serum concentrations of IgM, IgG, IL-6, IL-8 increased. Local use of vitamin D3 in UC reduces DAI parameters, causes decrease in blood neutrophil counts, reducing and partially restoring absorptive and NBT-reducing abilities of neutrophils, decline of total lymphocyte counts in blood, partially restoring the CD3+and CD45RA+ numbers, causing decline and partial restoration of serum IgM, IgG, IL-6, IL-8 concentrations. An association between clinical signs and indexes of immune status in UC was established under the conditions of vitamin D3 use. Conclusions: The protective effect of vitamin D3 in UC can be mediated by its antioxidant effect, changes in production of immunoregulatory cytokines, modulation of Th1-, Th2-, Th17-dependent reactions and Treg activity, being a pre-requisite for further studies to clarify the mechanism of vitamin D3 immunotropic action in UC,with an opportunity of using it in clinical practice.

Age-related aspects of cytokine profile of immune response in children with pertussis

The article presents a comparative analysis of the cytokine network in 161 patients with pertussis under the age of 1 year and in 180 patients over the age of 1 year. The studies revealed lower production of immunoregulatory cytokines (interferon-γ and interleukin-2) in patients under the age of 1 year at all stages of the disease and with all variants of pertussis (both mono- and mixed infections). The researchers revealed the relationship between the level of interferon-γ production and the severity of pertussis. They revealed age differences in the interferon-8 production in patients with mixed infection, which can determine clinical features and cause bronchopulmonary complications. The authors demonstrated the features of the dynamics of spontaneous and induced production of interleukin-6 and anti-inflammatory cytokines (interleukin-4, interleukin-10) in children under the age of 1 year; these features can be considered as immunological markers determining the imperfection of the humoral response in patients with pertussis in this age group.

The Animal Lectin Galectin-8 Promotes Cytokine Expression and Metastatic Tumor Growth in Mice

Secreted animal lectins of the galectin family are key players in cancer growth and metastasis. Here we show that galectin-8 (gal-8) induces the expression and secretion of cytokines and chemokines such as SDF-1 and MCP-1 in a number of cell types. This involves gal-8 binding to a uPAR/LRP1/integrin complex that activates JNK and the NFkB pathway. Cytokine and chemokine secretion, induced by gal-8, promotes migration of cancer cells toward cells treated with this lectin. Indeed, immune-competent gal-8 knockout (KO) mice express systemic lower levels of cytokines and chemokines while the opposite is true for gal-8 transgenic animals. Accordingly, gal-8 KO mice experience reduced tumor size and smaller and fewer metastatic lesions when injected with cancer cells. These results suggest the existence of a ‘vicious cycle’ whereby gal-8 secreted by the tumor microenvironment, promotes secretion of chemoattractants at the metastatic niche that promote further recruitment of tumor cells to that site. This study further implicate gal-8 in control of cancer progression and metastasis through its effects on the production of immunoregulatory cytokines.

Innate immune receptors in development of Mycobacterium tuberculosis infection

According to the World Health Organization, over 10 million new tuberculosis cases are reported annually worldwide. According to the 2017 Federal State Statistics Service Report, incidence rate for active TB infection in the Russian Federation was 109.8 cases per 100,000 population, of which 41.3% accounted for chronic disease form. Regardless of climatic conditions, high prevalence of TB infection, is not only due to high Mycobacterium tuberculosis viability, but also its ability for long persistence in human body and reactivation after an unlimited period of dormancy. The outcome of infection is largely determined by host immunoreactivity and its ability to develop protective immune response. In addition, status of immune system also underlies tuberculosis course after the onset: either as a localized form, or as a form with extensive damage to the lungs and even other organs observed in generalized infection. In recent decades, a great attention was paid to examining mechanisms of adaptive cell immunity played in pathogenesis of TB infection. No doubt, adaptive immunity is a powerful defense system providing a targeted specific immune response, but now it is becoming clear that it represents solely an effector arm of innate immunity. Innate immunity is a phylogenetically more ancient, inherited system largely aimed at ensuring rapid pathogen elimination and preventing development of infection at early stages when adaptive immunity ongoing antigen-specific maturation. Mechanisms of innate immunity mediated by cells, diverse receptors, molecules and their complexes, found on various cells. Activation of innate immunity begins with recognition of conserved molecular groups present in various pathogens called pathogen-associated molecular patterns (PAMPs), which are sensed by pathogen recognition receptors (PRRs). Here, we review current data on the role of innate receptors in recognizing M. tuberculosis-derived PAMPs, production of immunoregulatory cytokines and activation of signaling pathways playing a crucial role in the regulation of necroptosis, apoptosis and autophagy of infected macrophages. Significance of innate mucosal factors in implementing immune response to M. tuberculosis is discussed. In particular, Toll-like receptors, scavenger-receptors, mannose receptor, DC-SIGN etc. were described to participate in development of M. tuberculosis immunity. The data on single nucleotide polymorphic variants for innate genes are shown, which predispose to developing tuberculosis and affecting its course.

Low-Dose IL-2 Therapy in Transplantation, Autoimmunity, and Inflammatory Diseases.

Regulatory T cells (Tregs) play a central role in the induction and maintenance of immune homeostasis and self-tolerance. Tregs constantly express the high-affinity receptor to IL-2. IL-2 is a pleiotropic cytokine and a key survival factor for Tregs. It maintains Tregs' suppressive function by promoting Foxp3 expression and subsequent production of immunoregulatory cytokines. Administration of low-dose IL-2 is shown to be a promising approach to prevent allograft rejection and to treat autoimmune and inflammatory conditions in experimental models. The combination of IL-2 with its mAb (JES6-1) has also been shown to increase the t 1/2 of IL-2 and further enhance Treg frequencies and function. Low-dose IL-2 therapy has been used in several clinical trials to treat conditions such as hepatitis C vasculitis, graft-versus-host disease, type 1 diabetes, and systemic lupus erythematosus. In this paper, we summarize our findings on low-dose IL-2 treatment in corneal allografting and review recent studies focusing on the use of low-dose IL-2 in transplantation, autoimmunity, and other inflammatory conditions. We also discuss potential areas of further investigation with the aim to optimize current low-dose IL-2 regimens.

THE LEVELS OF IL-4 AND IL-8 IN SERUM OF PATIENTS WITH CHRONIC GASTRITIS AND ULCERATIVE DISEASE OF STOMACH AND DUODENUM

Local inflammation in H. pylori infection is characterized by infiltration of the gastric mucosa by neutrophils and specific lymphocytes, an increase in the production of immunoregulatory, pro- and anti-inflammatory cytokines. IL-4 is the most important regulatory cytokine that plays a role in a huge number of immune and non-immune functions. IL-8 is a potent angiogenic factor and chemoattractant, its presence in the circulation can have a systemic pro-inflammatory effect on the body. IL-4 and IL-8 in H. pylori infection has been studied insufficiently. The goal is to establish serum levels of IL-4 and IL-8 in patients with chronic gastritis and peptic ulcer and duodenal ulcer. Materials and methods. 95 patients of the gastroenterological department were examined, of them 37 men and 58 women (mean age 59 years): 53 patients diagnosed with chronic active gastritis (CAG, including erosive gastroduodenitis) and 42 patients diagnosed with peptic ulcer (PU). Patients were conditionally divided into 4 subgroups: PU (without confirming the presence in patients with Hp according to generally accepted methods), PU + Hp (patients with the presence of Hp), CAG and CAG + Hp. Results. An increase in serum IL-4 levels (above 10 pg / ml) was observed in only 50.7% of patients, with approximately the same frequency in patients with CAG and PU, and an increase in serum IL-8 levels (above 0.062 ng / ml) - in most patients (90%), and the highest levels of IL8 were detected in patients with PU + Hp. In the first analysis, 31% of patients had elevated levels of IL-8 in the absence of IL-4, while in the second analysis - already 62% of patients (p ≤ 0.01). This indicated a sharp twofold decrease in the production of IL-4 in just a few days between the two analyzes. Conclusion. Considering that an increase in serum levels of IL-8 is found in a larger number of patients than IL-4, which, moreover, ceases to be detected in serum, it can be assumed that the production and anti-inflammatory effect of IL-4 are insufficient in some patients with gastritis and peptic ulcer disease. Against this background, elevated levels of IL-8 can help maintain inflammation in a significant number of patients, as well as reduce the effectiveness of eradication therapy.

Implications of altered O-glycosylation in tumour immune evasion.

Aberrant glycosylation on tumour cells has been implicated in tumour immune modulation. A recent article published in The Journal of Biochemistry (Sutoh Yoneyama et al., A mechanism for evasion of CTL immunity by altered O-glycosylation of HLA class I, J. Biochem. 2017;161:479-492) showed that bladder cancer cells evaded cytotoxic T lymphocyte-mediated antitumour immunity by a novel mechanism involving the loss of Core 2 structures on human leukocyte antigen Class I O-glycans and subsequent impairment of galectin-glycan lattice formation. The immunosuppressive action of O-glycans on natural killer cell-mediated tumour immunity is also considered an immune evasion system. Furthermore, sialylated O-glycans have been proposed to play a central role in tumour immune escape by modulating the production of immunoregulatory cytokines and growth factors through interactions with sialic acid-binding immunoglobulin-like lectins. Therefore, a better understanding of how alterations in O-glycosylation influence tumour immune evasion will enable the development of novel and more effective therapeutic options for cancer treatment.

Production of immunoregulatory cytokines in clinically asymptomatic periapical lesions depends on the lesions size

Bacground/Aim. Development of periapical lesions (PLs) involves a complex cross-talk between pathogenic microorganisms from the root canal and host immune mechanisms. In these processes proinflammatory cytokines are involved in stimulation of inflammation and osteodestructive mechanisms, whereas anti-inflammatory cytokines, with the immunoregulatory functions, have the opposite effects. How this balance is controlled is still the subject of numerous studies. The aim of this study was to examine whether the local production of interleukin (IL)-10, IL-27 and transforming growth factor (TGF)-? in human asymptomatic PLs depends on the lesion size and how levels of these immunoregulatory cytokines correlate with the cellular composition of PLs. Methods. The study was conducted on 30 PLs which were collected at the Clinic for Stomatology of the Military Medical Academy, Belgrade, Serbia. The PLs were divided according to their size into small- and largesize lesions (n = 12 and n = 18, respectively). The inflammatory cells (PL-ICs) were isolated from PLs and cultivated for 24 hours in culture medium supplemented with phorbol myristate acetate and Ca2+ ionophore. Cytospins were processed for immunocytology by using monoclonal antibodies to cell subsets. The levels of cytokines in culture supernatants were determined by the ELISA method. Statistical analysis was performed using the Student t-test and the Spearman?s correlation test. The values of p < 0.05 were considered to be significant. Results. The levels of IL-10 and TGF-? were significantly higher in the PL-ICs cultures of large-size lesions than in small ones (p < 0.01 and p < 0.05, respectively). In contrast, the levels of IL-27 were higher in the cultures of small-size lesions than in small ones (p < 0.05). Although the total number of PL-ICs and the proportion of mononuclear phagocytes were higher in the large-size PLs (p < 0.01 and p < 0.05, respectively), their main composition was not significantly different between the groups. The proportions of B cells/plasma cells (CD19/CD38+ cells), CD8+ T cells and CD14+ cells were significantly higher in the large-size PLs (p < 0.005; p < 0.05; p < 0.05, respectively). In contrast, the proportion of total T cells (CD3+ cells) was higher in the small-size lesions (p < 0.05). No statistically significant correlation was found between the levels of these cytokines and the composition/ phenotype of PL-ICs. Conclusion. This study suggests that IL-10, IL-27 and TGF-? may play different roles in suppression of unwanted immune/inflammatory reactions in asymptomatic PLs, depending on the extension of pathological process as judged by the size of lesions.

Recruitment of Antigen Presenting Cells to Skin Draining Lymph Node From HPV16E7-Expressing Skin Requires E7-Rb Interaction.

“High-risk” human papillomaviruses (HPV) infect keratinocytes of squamous epithelia. The HPV16E7 protein induces epithelial hyperplasia by binding Rb family proteins and disrupting cell cycle termination. Murine skin expressing HPV16E7 as a transgene from a keratin 14 promoter (K14.E7) demonstrates epithelial hyperplasia, dysfunctional antigen presenting cells, ineffective antigen presentation by keratinocytes, and production of immunoregulatory cytokines. Furthermore, grafted K14.E7 skin is not rejected from immunocompetent non-transgenic recipient animals. To establish the contributions of E7, of E7-Rb interaction and of epithelial hyperplasia to altered local skin immunity, K14.E7 skin was compared with skin from K14.E7 mice heterozygous for a mutant Rb unable to bind E7 (K14.E7xRbΔL/ΔL mice), that have normoplastic epithelium. Previously, we demonstrated that E7-speicfic T cells do not accumulate in K14.E7xRbΔL/ΔL skin grafts. Here, we further show that K14.E7xRbΔL/ΔL skin, like K14.E7 skin, is not rejected by immunocompetent non-transgenic animals. There were fewer CD11b+ antigen presenting cells in skin draining lymph nodes from animals recipient of K14.E7xRbΔL/ΔL grafts, when compared with animals receiving K14.E7 grafts or K5mOVA grafts. Maturation of migratory DCs derived from K14.E7xRbΔL/ΔL grafts found in the draining lymph nodes is significantly lower than that of K14.E7 grafts. Surprisingly, K14.E7xRbΔL/ΔL keratinocytes, unlike K14.E7 keratinocytes, are susceptible to E7 directed CTL-mediated lysis in vitro. We conclude that E7-Rb interaction and its associated epithelial hyperplasia partially contribute to the suppressive local immune responses in area affected by HPV16E7 expression.

Interferon-β inhibits inflammatory responses mediators via suppression of iNOS signaling pathway in PBMCs from patients with primary Sjögren's syndrome.

Primary Sjögren's syndrome (pSS) represents a chronic, systemic autoimmune disorder, characterized by lymphocytic infiltration of exocrine glands, inducing compromised secretory function and tissue destruction. Increasing evidence had revealed that inflammatory mediators, such as nitric oxide (NO) and pro-inflammatory cytokines, are critical in the development and perpetuation of pSS systemic manifestations. In our current study, we aimed to investigate the ex vivo immunomodulatory effect of interferon (IFN)-β on iNOS expression, as well as on pro-inflammatory (tumor necrosis factor (TNF)-α, interleukin (IL)-6) and immunoregulatory (IL-10) cytokine production. Furthermore, we examined potential associations between the influence of IFN-β treatment on NO production, and pSS clinical and serological manifestations. In 41 pSS patients documented for their clinical and serological features, NO and cytokines levels were measured by the Griess method and enzyme-linked immunosorbent assay, respectively. Inducible nitric oxide synthase expression was analyzed by fluorescence immunostaining assay, using peripheral blood mononuclear cells (PBMCs) isolated from healthy controls and pSS patients. Our results revealed a strong down-modulating effect of IFN-β in the secretion of pro-inflammatory mediators including TNF-α, IL-6, and NO production. Interestingly, IFN-β exerts an increase in IL-10 levels. The most suppressive effect exerted by IFN-β on NO production was importantly reported for patients with neurological manifestation. This immunomodulatory effect of IFN-β on NO production is highly related to the decrease of inducible nitric oxide synthase (iNOS) expression. Our findings highlight a consistent ex vivo inhibitory effect of IFN-β on pro-inflammatory cytokine production and NO pathway in pSS patients. Our data suggest that IFN-β could represent a potential candidate for targeting inflammation during pSS.

Endocrine disrupting activities and immunomodulatory effects in lymphoblastoid cell lines of diclofenac, 4-hydroxydiclofenac and paracetamol

A critical literature review reveals that knowledge of side effects of pharmaceuticals diclofenac and paracetamol is extremely important because of their widespread use and occurrence in the environment. In order to delineate whether these compounds have endocrine activity and influence on the immune system, we assessed the potential endocrine disrupting and immunomodulatory activities of: diclofenac (DIC), its metabolite 4-hydroxydiclofenac (4-HD) and paracetamol (PAR). Herein, we report on their impact on estrogen receptor (ER), androgen receptor (AR), glucocorticoid receptor (GR) and thyroid hormone receptor (TR). The endocrine disrupting effects were assessed in vitro in MDA-kb2 and GH3.TRE-Luc cell lines and by the XenoScreen YES/YAS assay. Moreover, binding affinity to nuclear receptors (GR and AR) was also measured. Immunomodulatory properties of the compounds were evaluated in lymphoblastoid cell lines. All the tested compounds showed endocrine disrupting and immunomodulatory activities. The results revealed that both DIC and its metabolite 4-HD exhibited significant estrogenic, anti-androgenic (in YAS assay), (anti)-androgenic, (anti)-glucocorticoid and anti-thyroid hormonal activities (in luciferase reporter gene assays). DIC showed direct binding to the GR, while its metabolite 4-HD to the GR and AR. Only metabolite 4-HD showed estrogenic, androgenic (in YAS assay) and thyroid-hormonal activities. PAR had anti-androgenic activity and anti-thyroid hormonal activity. PAR displayed GR agonist activity with competition to its receptor and agonistic activity to AR. All of the compounds significantly modulated pro-inflammatory and immunoregulatory cytokine production in lymphoblastoid cell lines and were thus proven immunomodulatory. The study is useful in determining toxicological effects and contributes to the knowledge of possible side effects of diclofenac, its metabolite and paracetamol.