IL-6 Cytokines Research Articles

Phosphatidylserine Topically Attenuates Imiquimod-induced Psoriasis Through Inflammation Inhibition in Mice.

Psoriasis is a chronic skin condition that is associated with persistent inflammation and skin lesions. Topical therapy has been a promising approach to the alleviation of psoriasis through the application of anti-inflammatory agents. Phosphatidylserine (PS) administration has shown anti-inflammatory effects in the trials. Consequently, the objective of this study was to evaluate the effects of topical PS on the potential improvement of an imiquimod (IMQ)-induced psoriasis model. Additionally, cyclosporine A was utilized as a comparative anti-psoriatic agent in our study. The psoriasis model was established by topically applying IMQ to the dorsal skin of mice once daily for five consecutive days. The efficacy of topical PS was assessed using the Psoriasis Area and Severity Index (PASI) score to evaluate skin lesions. Subsequently, the skin samples were analyzed using Baker's scoring system, Masson's trichrome staining, immunohistochemistry, and real-time PCR analysis. IMQ-induced plaque-type psoriasis resulted in a significant increase (P<0.05) in dermal thickness, hyperkeratosis, PASI score, and inflammatory cytokines at the lesion site. The topical PS and cyclosporine A significantly (P<0.05) reduced PASI score and dermal thickness, while also alleviating erythema and scaling when compared to untreated mice. Furthermore, biomolecular assessments revealed that PS significantly (P<0.05) inhibited the gene expression of IL-17, IL-23, and TNF-α cytokines in the IMQ-induced lesions. Topical PS may pointedly alleviate psoriasis through the inhibition of inflammation. The beneficial effects of the PS recommend further investigation in both experimental and clinical studies in the control of skin psoriasis.

Efficient multiplex non-viral engineering and expansion of polyclonal γδ CAR-T cells for immunotherapy.

Gamma delta (γδ) T cells are defined by their unique ability to recognize a limited repertoire of non-peptide, non-MHC-associated antigens on transformed and pathogen-infected cells. In addition to their lack of alloreactivity, γδ T cells exhibit properties distinct from other lymphocyte subsets, prompting significant interest in their development as an off-the-shelf cellular immunotherapeutic. However, their low abundance in circulation, heterogeneity, limited methods for ex vivo expansion, and under-developed methodologies for genetic modification have hindered basic study and clinical application of γδ T cells. Here, we implement a feeder-free, scalable approach for ex vivo manufacture of polyclonal, non-virally modified, gene edited chimeric antigen receptor (CAR)-γδ T cells in support of therapeutic application. Engineered CAR-γδ T cells demonstrate high function in vitro and and in vivo. Longitudinal in vivo pharmacokinetic profiling of adoptively transferred polyclonal CAR-γδ T cells uncover subset-specific responses to IL-15 cytokine armoring and multiplex base editing. Our results present a robust platform for genetic modification of polyclonal CAR-γδ T cells and present unique opportunities to further define synergy and the contribution of discrete, engineered CAR-γδ T cell subsets to therapeutic efficacy in vivo .

Capsaicin attenuates the effect of inflammatory cytokines in a HaCaT cell model for basal keratinocytes.

The resolution of the skin's inflammatory response is only possible if its barrier function is restored. TRPV1 channel activation plays an important role during inflammation but the effect of this activation on the skin barrier under inflammatory conditions has not been clarified. We hypothesize that it could potentially aid the keratinocyte barrier by reducing inflammatory cytokine release and promoting tight junction development. To explore the role of TRPV1 activation in inflammation, we designed and optimized an in vitro model of keratinocytes with basal epidermal layer characteristics using HaCaT cells and TNFα to induce inflammation. TNFα increased the gene expression of tight junction protein claudin 1 (CLDN1) by at least 2.60 ± 0.16-fold, in a concentration-dependent manner, over a 48h period. The administration of a capsaicin pre-treatment reduced the CLDN1 expression to 1.51 ± 0.16-fold during the first 6h after TNFα induction, whereas IL-8 cytokine release was reduced 0.64 ± 0.17-fold. After 48h, CLDN1 protein levels increased by a factor of 6.57 ± 1.39 compared to cells only treated with TNFα. These results suggest that activation of TRPV1 by capsaicin can potentiate the increase in CLDN1 expression and CLDN1 protein synthesis induced by TNFα in cultured keratinocytes, while reducing the release of IL-8.

The Relationship Between Serum IgE Level and IL-4 and IL-13 Cytokines in Colorectal Cancer Patients

ABSTRACT Background Colorectal cancer (CRC) is the most common malignancy of the digestive system in the world. Immune cells and molecules in tumor microenvironment are crucial.Identifying immune system components in cancer aids in biomarker discovery. This study investigated the serum IgE levels and expression of IL-4 and IL-13 in the tissue and serum of CRC patients and explored their possible association with pathological and clinical factors. Materials and methods Thirty-six patients with CRC and 36 healthy individuals were involved in the study. Tissues and blood samples were collected. Serum levels of IgE and IL-4 and IL-13 were analyzed using the ELISA method. The quantitative Real-Time PCR (qRT-PCR) technique was used to assess the expression levels of the cytokines in CRC tissue samples in comparison with the adjacent control tissue. Results Our results revealed that the serum level of IL-4 and IL-13 and also their gene expression levels were significantly decreased in CRC patients compared to the controls. The results of this study revealed that there is no significant difference in the serum levels of IgE between CRC patients and the control group. Conclusion All in all, the results of the current research suggest that the expression levels of IL-13, IL-4, and IgE vary between CRC tissue.

Th2 and IL-17 responses inChlamydia pneumoniae-stimulated PBMC in asthmatic and non-asthmatic subjects

Abstract Background Chlamydia pneumoniae (C. pneumoniae) is a gram-negative intracellular bacterium that causes respiratory infections in humans, including asthmatic and non-asthmatic subjects. C. pneumoniae activates cells in vitro and produces cytokines that may contribute to the inflammatory responses observed in asthma. Different asthma endotypes are described, including Th2-high and Th2-low. Th2-low endotypes are characterized by IL-17 production and neutrophilic inflammation. The aim of this study was to investigate the role of C. pneumoniae in regulating Th2 versus Th17 responses in peripheral blood mononuclear (PBMC) from subjects with or without asthma. Methods PBM) (1×106/mL) from asthmatic (N=6) and non-asthmatic subjects (N=14) were infected +/- C. pneumoniae TW-183 at a multiplicity of infection (MOI) = 0.1, using dose responses (1:10, 1:100), and cultured 48 hrs. Cytokine responses (Interferon (IFN)-gamma, Interleukin (IL)-2, IL-4, IL-17 A/F) were measured in supernatants (ELISA). Results Comparison of cytokine responses (the mean differences in non-asthmatic versus asthmatic subjects) were significant for IFN gamma (unstimulated; P&amp;lt;0.0001), IL-2 (unstimulated and 1:100; P&amp;lt;0.0001, P=0.0002, respectively), IL-4 (unstimulated, 1:10, 1:100; P=0.0001, P&amp;lt;0.0001, P&amp;lt;0.0001, respectively) and IL-17 A/F (unstimulated, P&amp;lt;0.0001) (Wilcoxon signed-rank test). Cytokine levels were higher in asthmatic subjects for IFN-gamma (unstimulated,1:10, 1:100), IL-2 (unstimulated), and IL-17 A/F (unstimulated) compared with non-asthmatic subjects. However, IL-4 levels were higher in non-asthmatics (unstimulated, 1:10, 1:100). Conclusion Differences in Th2 and IL-17 cytokines responses in PBMC from subjects with and without asthma may indicate the involvement of cell-mediated immunity, while Th17 responses, which were increased in subjects with asthma at baseline, did not increase further in response to C. pneumoniae stimulation.

Protective Role of Eicosapentaenoic and Docosahexaenoic and Their N-Ethanolamide Derivatives in Olfactory Glial Cells Affected by Lipopolysaccharide-Induced Neuroinflammation

Neuroinflammation is a symptom of different neurodegenerative diseases, and growing interest is directed towards active drug development for the reduction of its negative effects. The anti-inflammatory activity of polyunsaturated fatty acids, eicosapentaenoic (EPA), docosahexaenoic (DHA), and their amide derivatives was largely investigated on some neural cells. Herein, we aimed to elucidate the protective role of both EPA and DHA and the corresponding N-ethanolamides EPA-EA and DHA-EA on neonatal mouse Olfactory Ensheathing Cells (OECs) after exposition to lipopolysaccharide (LPS)-induced neuroinflammation. To verify their anti-inflammatory effect and cell morphological features on OECs, the expression of IL-10 cytokine, and cytoskeletal proteins (vimentin and GFAP) was evaluated by immunocytochemical procedures. In addition, MTT assays, TUNEL, and mitochondrial health tests were carried out to assess their protective effects on OEC viability. Our results highlight a reduction in GFAP and vimentin expression in OECs exposed to LPS and treated with EPA or DHA or EPA-EA or DHA-EA in comparison with OECs exposed to LPS alone. We observed a protective role of EPA and DHA on cell morphology, while the amides EPA-EA and DHA-EA mainly exerted a superior anti-inflammatory effect compared to free acids.

Screening immunomodulatory Q-markers in Astragali Radix based on UHPLC-QTOF-MS analysis and spectrum-effect relationship.

Astragali Radix (AR) is one of the famous traditional Chinese medicines (TCMs) for boosting immunity, whereas the quality markers (Q-markers) of AR have not been clearly researched. The immunomodulatory activities of the bioactive extractions and components were evaluated by NO inhibition rate; phagocytic index; IL-10, TNF-α, IL-1β, and IL-6 cytokines in RAW264.7 cells; and the relative proliferation rate of spleen cells. The total saponins (TS) and the grade 2 (Xiaoxuan, XX) of AR showed the strongest immunomodulatory activities. At the concentration of 40 μg/mL, the TS increased spleen cells proliferation by 48.0% and upregulated the level of IL-1β and IL-6. Cytokines in the XX-treated group were at least 1.6 times higher than the control group. A total of 190 common peaks were detected in AR by ultrahigh-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UHPLC-QTOF-MS). The multivariate statistical analyses revealed that 41 compounds were positively correlated with immune responses, and bioactive compounds were verified by using RAW264.7 cell assay. Subsequently, the contents of six compounds in different commercial grades were determined, and the results showed the same trend in contents and activities. Finally, calycosin-7-O-β-D-glucoside, astragaloside IV, astragaloside II, astragaloside I, isomucronulatol-7-O-glucoside, and 9,10-dimethoxypterocarpan-3-O-glucoside were screened out as immunomodulatory Q-markers of AR.

Effects of Functionalized Carbon Nanotube on Cellular Homeostasis of Murine Hepatocytes

Nanotechnology in medicine represents a revolution offering significant advancements in the diagnosis, treatment, and prevention of diseases. This technology enables the development of devices and materials at the nanometer scale, allowing precise interactions with biological systems at the molecular and cellular levels. Among various nanomaterials, carbon nanotubes (CNTs) stand out due to their unique physical, chemical and mechanical properties. Due to their high strength, electrical and thermal conductivity and large surface area, CNTs have a wide range of applications. In healthcare they are explored for drug delivery and imaging diagnostics. CNTs can be functionalized to target drugs directly to diseased cells, minimizing side effects and enhancing treatment efficacy, additionally can be used in biosensors for early disease detection and in tissue engineering to cellular regeneration. The present study evaluated the effects of OCNT-TEPA, a multi-walled carbon nanotube functionalized, in murine hepatocytes AML-12. Cells were exposed to different concentrations of the sample for 12, 24, 48, and 72 hours. Cellular metabolism tests, cellular morphology by optical microscopy, synthesis of reactive oxygen species, changes in membrane potential and IL-6 cytokine secretion were performed. The results show that OCNT-TEPA altered the homeostasis of hepatocytes, as cellular metabolism decreased, cellular morphology was altered, the membrane experienced changes in its electrical potential, oxidative stress increased and inflammatory signaling molecules were synthesized. This alteration is dose-dependent, which may be harmful to hepatocytes at high concentrations but could be applied to the body at lower concentrations without causing harm.

The Role of Interleukin-24 and Downstream Pathways in Inflammatory and Autoimmune Diseases.

Inflammatory and autoimmune diseases are pathological immune disorders and pose significant public health challenges due to their impact on individuals and society. Cytokine dysregulation plays a critical role in the development of these disorders. Interleukin (IL)-24, a member of the IL-10 cytokine family, can be secreted by various cell types, including immune and non-immune cells. The downstream effects of IL-24 upon binding to its receptors can occur in dependence on, or independently of, the Janus kinase (JAK)/signal transducer and the activator of transcription (STAT) signaling pathway. IL-24 and its downstream pathways influence crucial processes such as cell differentiation, proliferation, apoptosis, and inflammation, with its role varying across different diseases. On the one hand, IL-24 can inhibit the activation of pathogenic cells and autoimmune responses in autoimmune ocular diseases; on the other hand, IL-24 has been also implicated in promoting tissue damage by fostering immune cell activation and infiltration in psoriasis and allergic diseases. It suggests that IL-24, as a multifunctional cytokine, has complex regulatory functions in immune cells and related diseases. In this paper, we summarize the current knowledge on IL-24's immunomodulatory actions and its involvement in inflammatory and autoimmune disorders. Such insights may pave the way for novel therapeutic strategies for these diseases.

Immuno-therapeutic and prophylactic potential of Trichinella spiralis antigens for inflammatory bowel diseases

Ulcerative colitis (UC), a severe chronic inflammatory disorder of the colon, is one of the inflammatory bowel diseases (IBD) that affects humans and several domestic animal species, including cats and dogs. Helminth infections and autoimmune diseases are inversely correlated, as explained by the hygiene hypothesis, which suggests that IBD is infrequent in countries where helminth infections are common but more prevalent in developed nations. This study investigated the therapeutic and prophylactic potential of Trichinella spiralis (T. spiralis) antigens in an experimental colitis model for IBD. Mice were divided into eight groups: normal model, colitis model, larval antigen prophylaxis, adult antigen prophylaxis, larval antigen therapeutic, adult antigen therapeutic, larval antigen prophylaxis and therapeutic, and adult antigen prophylaxis and therapeutic. Colitis was induced intrarectally by administering a single dose of 0.2 ml of acetic acid, except in the healthy group, which received PBS (0.2 ml). The mice were euthanized 12 days after colitis induction. The therapeutic and prophylactic potential of T. spiralis antigens were assessed through colitis severity and histopathological, immunological, and immunohistochemical examinations. The results showed a significant reduction in Disease Activity Index (DAI), an increase in goblet cells' acidic mucin levels, reduced iNOS and TNF-α expression, and decreased serum levels of IFN-γ and IL-10 cytokines in Groups IV-VIII compared to the colitis model, particularly in the group that received adult worm antigen both prophylactically and therapeutically. This study demonstrated that T. spiralis antigens, especially from adult worms, had protective and therapeutic effects on experimental colitis, with a superior effect when administered both before and after colitis induction by reducing inflammation and modulating the immune response. Thus, T. spiralis antigens may improve disease outcomes and provide a novel treatment approach for ulcerative colitis.

Serotonin Availability Shapes the Effects of Phenelzine on Inflammatory Response and Gene Expression in Macrophages.

Background Serotonin (5-HT) is a neurotransmitter with extensive physiological influence in the central nervous system (CNS) and various behavioral and biological functions, including immune regulation through 5-HT receptors (5-HTRs) expressed by immune cells. A variety of serotonin-modulating drugs have been developed to treat neurological disorders. Phenelzine, a drug indicated for the management of treatment-resistant depression, is a potent, non-selectiveinhibitor of monoamine oxidase (MAO), the enzyme that metabolizes serotonin to 5-hydroxyindole acetic acid (5-HIAA). Given the emerging evidence of a bidirectional link between depression and inflammation, as well as the potential therapeutic applications of serotonin-modulating drugs in autoimmune diseases and cancer, our study investigated the pro-inflammatory and anti-inflammatory factors influenced by serotonin and phenelzine. Methodology We conducted experiments on RAW264.7 macrophages, exposing them to various combinations and concentrations of serotonin, 5-HIAA, and phenelzine. We assessed the relative gene expression of monoamine oxidase-A (MAO-A), CYP1B1, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) using real-time PCR and measured the production of IL-6, TNF-α, and IL-10 cytokines using enzyme-linked immunosorbent assays (ELISA). Results Our findings revealed that phenelzine can downregulate genes associated with the production of reactive oxygen and nitrogen species, reduce aryl hydrocarbon receptor (AHR)-related gene expression induced by serotonin, and enhance the production of inflammatory cytokines. These effects were significantly influenced by the concentration of available serotonin. Conclusions Our study demonstrates that various mechanisms, including AHR activation, modulation of reactive oxygen and nitrogen speciesproduction, and others, in addition to the increased availability of serotonin due to phenelzine treatment, can significantly influence the inflammatory state.

Combined developmental exposure to estrogenic endocrine disruptor and nutritional imbalance induces long term adult prostate inflammation through inflammasome activation

Increasing number of studies suggested that environmental deleterious impacts (such as estrogen-like endocrine disruptors, EDCs, unhealthy diet) during early human development affect the risk of developing non-communicable diseases including prostate cancer (PCa) later in life. To test if the combination of EDCs and unhealthy induces adult prostate lesions, we developed an experimental model of adult male Sprague Dawley rats exposed during gestation (from day 7) to weaning to high fat diet (HFD 60 % fat), or to a xenoestrogen (estradiol benzoate, EB, 2.5 µg/d) from post-natal days 1–5, or to a combination of both. EB and EB+HFD exposures induced decreased prostate weight in adult rats along with inflammatory status. A white blood cell infiltrate was observed after EB exposure and more dramatic lesions were observed with the combined exposure, along with a gland destruction. The lesions, following EB or EB+HFD exposure, are associated with elevated mRNA levels for TNFa, IL6 and CCL2/MCP1 pro-inflammatory cytokines while the levels of the anti-inflammatory IL10 cytokine remained unchanged. This activation of NLRP3 and elevated levels of CASP1 were observed following EB or EB+HFD exposures associated with elevated mRNA levels for IL1b, substrates for the NLRP3 complex. HFD exposure alone has mild if not pro-inflammatory effects in adult prostate. In conclusion, we showed that developmental combined exposure to EB and HFD programmed prostate inflammatory lesions in adult prostate. Since proliferative inflammatory atrophy and chronic inflammation of prostate may drive cell to become cancer cells, our model might be useful for study onset of PCa.

Protective effect of recombinant interleukin-10 on newborn rat lungs exposed to short-term sublethal hyperoxia.

Lung injury imposed by hyperoxia is the main cause of bronchopulmonary dysplasia in newborns. These injuries are generated from the early stage of hyperoxia through the main biologic effects of cell death and inflammatory response. Interleukin (IL)-10 is a potent anti-inflammatory cytokine that may have the inhibitory effects on these biologic actions induced by hyperoxia. Based on our former in vitro studies investigating the effect of recombinant IL-10 (rIL-10) on protecting cultured alveolar type II cells exposed to short-term hyperoxia, we performed the in vivo study to investigate the effect of rIL-10 in newborn rats aged P4 exposed to hyperoxia. Rats were classified into 3 groups; the control group exposed to normoxia for 24 hours; the hyperoxia group exposed to 65% hyperoxia for 24 hours; and the IL10 group treated with intratracheal instillation of rIL-10 prior to exposure to 65% hyperoxia for 24 hours. Following each treatment, the rats were euthanized. Individual lobes of the right lung were prepared for hematoxyling and eosin (H&E) staining and immunohistochemical staining for thyroid transcription factor-1 (TTF1). Bronchoalveolar lavage (BAL) was performed in the left lung to analyze cell counts and cytokines. The IL10 group showed preserved air spaces similar to the control group, with decreased cellularity compared to the hyperoxia group, whereas the hyperoxia group showed markedly reduced air spaces with increased cellularity compared to the IL10 group. And, the IL10 group showed more TTF1-positive cells, which represented alveolar type II cells, compared to the hyperoxia group. Inflammatory cells, such as neutrophils and lymphocytes and proinflammatory cytokines of tumor necrosis factor-α, IL-1α, IL-8, and macrophage inflammatory protein-1α were significantly lower in BAL fluid of the IL10 group compared to the hyperoxia group. These results indicate that rIL-10 may be a promising pharmaceutical measure for protecting newborn lungs from injury induced at the early stage of hyper oxia.

Tumor Antigen-Tethered Spiked Virus-Like- Poly(Lactic-Co-Glycolic Acid)-Nanoparticle Vaccine Enhances Antitumor Ability Through Th9 Promotion in Mice.

Immunotherapy emerges as a promising frontier in cancer therapy and prevention. This study investigates the capacity of tumor-antigenic nanoparticles, specifically ovalbumin-tethered spiked virus-like poly(lactic-co-glycolic acid) nanoparticles (OVA-sVLNP), to effectively elicit humoral and cellular immune responses against tumors. OVA-sVLNP were synthesized through thiol-maleimide crosslinking using a single emulsion method. Comprehensive characterization was performed through Nuclear Magnetic Resonance (NMR), dynamic light scattering, Cryo-electron microscopy (Cryo-EM), confocal microscopy, and flow cytometry. Immunogenicity was evaluated using an enzyme-linked immunosorbent assay (ELISA) for quantifying immunoglobulin levels (IgG, IgG1, IgG2a) and cytokines in mouse sera. Flow cytometry profiled cellular immune responses in mouse spleens, and organ biosafety was assessed using immunohistochemistry and hematoxylin and eosin (H&E) staining. OVA-sVLNP had a mean particle size of 193.8 ± 11.9 nm, polydispersity index of 0.307 ± 0.04, and zeta potential of -39.6 ± 10.16 mV, remaining stable for one month at 4°C. In vitro studies revealed significant upregulation of CD80/CD86 in dendritic cells, indicating robust activation. In vivo, the optimal concentration (V25) induced potent IgG, IgG1, and IgG2a antibodies, significant populations of CD3+CD4+, CD3+CD8+, and a rare subset of CD3+CD4+CD8+ memory T cells. Notably, Th9 induction resulted in the secretion of IL-9, IL-10, and other cytokines, which are crucial for orchestrating cytotoxic T cell activity and antitumor effects. Overall, higher doses did not improve outcomes, highlighting the significance of optimal dosing. This study demonstrated potent immunogenicity of OVA-sVLNP, characterized by the induction of specific IgG antibodies and the stimulation of cellular immune responses, particularly tumor-killing Th9 cells. The simplicity and cost-effectiveness of the manufacturing process augment the potential of OVA-sVLNP as a viable candidate for antitumor vaccines, opening new avenues for cancer prevention and cell-based therapeutic strategies.

Oxidative stress in gut TH17 cells makes mice susceptible to bacterial infection.

A recent paper published in Cell Metabolism in August 2024 by Dirk Brenner's laboratory highlights the importance of effectively managing reactive oxygen species (ROS) in gut TH17 T cells for minimizing the damage caused by intestinal bacterial infection. This commentary will discuss the control of cellular ROS by glutathione and the emerging understanding that neutralizing ROS in immune cells is essential for the individualized functions of different immune subsets. In the case of this study, managing ROS within TH17 cells in the gut was shown to be essential to sustain the production of IL22 cytokine to maintain gut homeostasis in response to bacterial infection.

Targeting Kidney Inflammation After Brain Death and Cold Storage: Investigating the Potential of an NLRP3 Inflammasome Inhibitor (MCC950) for Preconditioning Donor Kidneys.

Brain death (BD) and cold storage (CS) are critical factors that induce inflammation in donor kidneys, compromising organ quality. We investigated whether treating kidneys from BD rats with an inflammasome Nod-like receptor family pyrin domain containing 3 (NLRP3) inhibitor (MCC950) followed by CS could reduce kidney inflammation. BD rats were assigned to MCC950-treated or nontreated (NT) groups. Kidneys were evaluated immediately before CS (T0) and after 12 h (T12) and 24 h (T24) of CS. Mean arterial pressure, serum creatinine, gene/protein expression, and histology were evaluated. At T0, MCC950 treatment did not affect mean arterial pressure but tended to reduce serum creatinine and ameliorated the histological score of acute tubular necrosis. However, MCC950 reduced NLRP3, caspase-1, interleukin (IL)-1β, IL-6, Kim-1, nuclear factor kappa B, tumor necrosis factor alpha, and caspase-3 gene expression while increasing IL-10 cytokine gene expression. After 12 h of CS, only the expression of the NLRP3 and caspase-1 genes decreased, and after 24 h of CS, no further changes in the gene expression profile were observed. The levels of the inflammasome proteins NLRP3, caspase-1, and IL-1β consistently decreased across all time points (T0, T12, and T24). These findings suggest that MCC950 treatment holds promise for mitigating the proinflammatory state observed in kidneys after BD and CS.

A multi-epitope protein vaccine encapsulated in alginate nanoparticles as a candidate vaccine against Shigella sonnei

Shigellosis, caused by the Gram-negative bacterium Shigella, is a major global health challenge. Despite extensive research over the past two decades, no commercial vaccine is available to prevent Shigella infection. Developing multi-epitope vaccines offers a promising and innovative approach to tackling infectious diseases. In this study, we produced a multi-epitope vaccine candidate using E. coli BL21 (DE3) plysS bacteria and purified the vaccine protein with Ni-NTA affinity chromatography. We then prepared alginate nanoparticles containing the vaccine protein, with a particle size of 122 ± 6 nm, PDI 0.17, SPAN 0.83, and zeta potential of -27 ± 2 mV. Successful protein loading was confirmed through nanodrop and ATR-FTIR analyses. To evaluate the immunogenicity of the encapsulated vaccine, mice were orally vaccinated, and their serum was analyzed for IgG, IL-4, and IFN-γ levels cytokines. The results showed a significant increase in IgG level in the vaccinated group compared to controls. Additionally, the vaccinated group exhibited a notable increase in IL-4 and IFN-γ cytokines, indicating a robust Th-cell-mediated immune response essential for combating Shigella. Our nano-vaccine demonstrated high efficacy in activating both humoral and cellular immunity, effectively protecting against the bacteria. The alginate-based oral vaccine candidate thus emerges as a promising strategy for developing a multi-epitope vaccine candidate against Shigella.

Cell-based in vitro hemoassay for evaluation of NLRP3-inflammasome activity

Currently, gouty arthritis is considered as a polygenic multifactorial autoinflammatory disease caused by activation of the NOD (nucleotide-binding domain) -like protein receptor 3 inflammasome NLRP3. The two cytokines IL-1β and IL-18 are considered important biomarkers of NLRP3 inflammasome activation. However, usually the concentration of IL-1β in donor sera is extremely low and found to be at the limit of detection level (1-3 pg/ml), while the concentration of circulating cytokine IL-18 in the sera of individual donors varies greatly. This results in difficulty using these biomarkers in the diagnosis of autoinflammatory diseases. We hypothesized that the patient’s blood cells which were sensitized in vivo to the presence of specific factors characteristic of autoinflammatory diseases, in particular, gouty arthritis, would produce increased amounts of the inflammasome-regulated cytokines compared to blood cells obtained from healthy donors. A comparison of the IL-18 cytokine in healthy donors and patients with gouty arthritis was carried out using 2 methods: a) by traditional analysis of the level of serum circulated IL-18 and b) by using a cell-based Hemoassay in vitro developed at the research institute “Biotech” SamGMU. The comparative analysis demonstrated the advantages of using an in vitro cell-based Hemoassay to assess the IL-18 cytokine status of patients. Serum IL-18 values varied widely and showed no significant difference between donors and patients with gouty arthritis. Using the developed cell-based Hemoassay in vitro, significant quantitative differences in the production of the inflammatory cytokine IL-18 produced by blood cells of potentially healthy donors and patients with gouty arthritis were detected. Blood cells of individual patients, sensitized in vivo with specific factors characteristic of gouty arthritis, produce increased concentrations of IL-18 in the cell growth medium in vitro compared to cells from healthy donors. Thus, the in vitro cell-based Hemoassay can be used for a more accurate assessment of the cytokine status of patients.

The role of epigenetic factors in childhood atopic dermatitis

Atopic dermatitis (AD) is a genetically determined chronic inflammatory skin disease characterized by itching, relapsing course, age-related features of the lesion morphology and localization. The etiological factors contributing to the development of AD are: allergic reactions, inflammatory reactions in the skin, leading to a barrier disfunction, the affect and interaction of genetic and environmental factors. Epigenetic mechanisms also play a key role in immune regulation. The aim of this work is to study the genome-wide methylation profile in the skin of children with AD in the acute stage, as well as to determine changes in the expression level of immune response genes associated with common signaling pathways during therapy. Whole-genome sequencing was conducted to examine methylation in the skin samples from AD patients and controls. The stages of bioinformatics data processing were carried out. Changes in the expression levels of the TLR2, TLR9, IL4, IL13, CAMP, and DEFB1 genes before and after treatment were carried out using RT-PCR-RT on samples of mononuclear blood cells. When comparing samples from patients with AD and healthy children, changes were shown in the methylation of 2364 regions of genomic DNA with their corresponding specific genes. Among the identified genes, those related to processes associated with the pathogenesis of AD were selected. At the next stage, changes in the expression of the described immunity factors (TLR2, TLR9, IL-4, IL-13, DEFB1, CAMP) were assessed over time according to the type of treatment. For this purpose, patients were divided into 2 groups depending on the prescription of systemic therapy. Three months after treatment, the level of expression for all studied immune factors decreased. However, a significant decrease in expression for almost all parameters (except for the DEFB1 gene) was recorded in the group in which systemic therapy (dupilumab) was used in addition to topical treatment. At the same time, the SCORAD index after treatment improved by an average of 63% and amounted to 17±6.0 points. Thus, systemic therapy aimed at suppressing signaling of IL-4 and IL-13 cytokines indirectly leads to normalization of the expression levels of innate immune factors. Studying the regulation of molecular genetic mechanisms of immunity involved in the processes of inflammation in AD helps to clarify the immunopathogenesis of this disease, determine diagnostic markers and targets for drug therapy.

Cyclic adenosine monophosphate critically modulates cardiac GLP-1 receptor's anti-inflammatory effects.

Glucagon-like peptide (GLP)-1 receptor (GLP1R) agonists exert a multitude of beneficial cardiovascular effects beyond control of blood glucose levels and obesity reduction. They also have anti-inflammatory actions through both central and peripheral mechanisms. GLP1R is a G protein-coupled receptor (GPCR), coupling to adenylyl cyclase (AC)-stimulatory Gs proteins to raise cyclic 3`-5`-adenosine monophosphate (cAMP) levels in cells. cAMP exerts various anti-apoptotic and anti-inflammatory effects via its effectors protein kinase A (PKA) and Exchange protein directly activated by cAMP (Epac). However, the precise role and importance of cAMP in mediating GLP1R`s anti-inflammatory actions, at least in the heart, remains to be determined. To this end, we tested the effects of the GLP1R agonist liraglutide on lipopolysaccharide (LPS)-induced acute inflammatory injury in H9c2 cardiac cells, either in the absence of cAMP production (AC inhibition) or upon enhancement of cAMP levels via phosphodiesterase (PDE)-4 inhibition with roflumilast. Liraglutide dose-dependently inhibited LPS-induced apoptosis and increased cAMP levels in H9c2 cells, with roflumilast but also PDE8 inhibition further enhancing cAMP production by liraglutide. GLP1R-stimulated cAMP markedly suppressed the LPS-dependent induction of pro-inflammatory tumor necrosis factor (TNF)-a, interleukin (IL)-1b, and IL-6 cytokine expression, of inducible nitric oxide synthase (iNOS) expression and nuclear factor (NF)-kB activity, of matrix metalloproteinases (MMP)-2 and MMP-9 levels and activities, and of myocardial injury markers in H9c2 cardiac cells. The effects of liraglutide were mediated by the GLP1R since they were abolished by the GLP1R antagonist exendin(9-39). Importantly, AC inhibition completely abrogated liraglutide`s suppression of LPS-dependent inflammatory injury, whereas roflumilast significantly enhanced the protective effects of liraglutide against LPS-induced inflammation. Finally, PKA inhibition or Epac1/2 inhibition alone only partially blocked liraglutide`s suppression of LPS-induced inflammation in H9c2 cardiac cells, but, together, PKA and Epac1/2 inhibition fully prevented liraglutide from reducing LPS-dependent inflammation. cAMP, via activation of both PKA and Epac, is essential for GLP1R`s anti-inflammatory signaling in cardiac cells and that cAMP levels crucially regulate the anti-inflammatory efficacy of GLP1R agonists in the heart. Strategies that elevate cardiac cAMP levels, such as PDE4 inhibition, may potentiate the cardiovascular, including anti-inflammatory, benefits of GLP1R agonist drugs.