Induction of cross-reactive, mucosal anti-SARS-CoV-2 antibody responses in rheumatoid arthritis patients after 3rd dose of COVID-19 vaccination.

Systemic vaccination against SARS-CoV-2 elicited high titers of specific antibodies in the blood and in the oral cavity. Preexisting autoimmune diseases, such as rheumatoid arthritis, and biological treatments, like B cell depletion, are known to exhibit higher risk of severe COVID-19 manifestation and increased frequency of breakthrough infections after vaccination. We hypothesized that such increased risk is associated with an aberrant induction of secreted antibodies in the oral cavity. Here we evaluated the levels of secreted antibodies in the oral cavity against the SARS-CoV-2 Spike protein during the course of vaccination in RA patients with or without B cell depletion. We found that total salivary IgG levels were correlated with number of B cells in the blood. Anti-Spike IgG responses 7 days after second vaccination were induced in the oral cavity of all healthy individuals, while only 6 out 23 RA patients exhibited anti-Spike IgG in their saliva regardless of B cell depleting therapy. Importantly, both salivary and serologic anti-Spike IgG and IgA responses towards WT and omicron Spike variants were efficiently induced by third vaccination in RA patients with or without B cell depletion to the levels that were similar to healthy individuals. Altogether, these data advocate for the necessity of three dose vaccination for RA patients to mount anti-Spike antibody responses at the mucosal surfaces and annotate the reduction of secreted salivary IgG by B cell depletion.

Cytokines as Mediators of Neuroinflammation in Experimental Autoimmune Encephalomyelitis.

Cytokines play a pivotal role in maintaining homeostasis of the immune system and in regulation of the immune response. Cytokine dysregulation is often associated with development of various pathological conditions, including autoimmunity. Recent studies have provided insights into the cytokine signaling pathways that are involved not only in pathogenesis of autoimmune neuroinflammatory disorders, such as multiple sclerosis, but also in neurodegenerative states, for example, Alzheimer's disease. Understanding the exact molecular mechanisms of disease pathogenesis and evaluation of relevant experimental animal models are necessary for development of effective therapeutic approaches.

[Analysis of the Specificity of IgA Antibodies Produced in the Mouse Small Intestine].

Intestinal microbiota controls multiple aspects of body homeostasis. The microbiota composition changes easily in response to internal or external factors, which may result in dysbiosis and associated inflammatory reactions. Thus, maintaining the microbiota composition by the host immune system is crucial, and one of the main mechanisms for microbiota control is production of immunoglobulin A (IgA) at mucosal surfaces. The molecular mechanisms regulating the interactions between the immune system and microbiota remain obscure. A panel of hybridoma cell lines was constructed to produce monoclonal IgA antibodies specific to various commensal bacteria present in intestinal microbiota. The panel can be used to further understand the mechanisms whereby the adaptive immune system controls the microbiota composition.

[Cytokine neutralization at specific cellular source : A new therapeutic paradigm? German Version]. / Zytokinneutralisierung an spezifischen zellulären Quellen : Ein neues Therapiemodell?

BACKGROUND: Currently, treatment of autoimmune diseases is based on manipulation of general control mechanisms, including those mediated by immunoregulatory cytokines. This approach is non-curative and may cause unwanted side effects due to numerous beneficial and non-redundant functions of a particular cytokine. METHODS: Techniques of reverse genetics, such as conditional gene targeting, were employed to uncover the contributions of two proinflammatory and immunomodulatory cytokines, tumour necrosis factor (TNF) and interleukin 6 (IL-6), in various disease states. RESULTS: Several non-redundant functions of TNF from distinct cellular sources were identified. TNF from myeloid cells is pathogenic in several autoimmune diseases, whereas TNF produced by T cells showed non-redundant protective functions in experimental arthritis and in a Mycobacterium tuberculosis infection model. To test the idea of selective pharmacological inhibition of "bad" TNF produced by myeloid cells while sparing "good" TNF produced by T lymphocytes, a myeloid-specific TNF inhibitor (MYSTI) was designed - a recombinant mini-antibody with dual specificity that can bind to the surface molecule F4/80 on myeloid cells and to TNF. In vitro experiments confirmed retention of TNF on the surface of TNF-producing cells and in vivo experiments indicated that MYSTI can protect mice from lethal TNF-mediated hepatotoxicity. MYSTI is also effective in experimental arthritis. CONCLUSION: The proposed therapeutic strategy may be more effective than systemic anti-cytokine therapy in several human autoimmune diseases, as it would preserve the potentially beneficial effects of the same cytokine produced by other cell types. Such bispecific biological agents may become interesting tools for experimental studies and, eventually, drug development.

Cytokine neutralization at specific cellular source : A new therapeutic paradigm?

BACKGROUND: Currently, treatment of autoimmune diseases is based on manipulation of general control mechanisms, including those mediated by immunoregulatory cytokines. This approach is non-curative and may cause unwanted side effects due to numerous beneficial and non-redundant functions of a particular cytokine. METHODS: Techniques of reverse genetics, such as conditional gene targeting, were employed to uncover the contributions of two proinflammatory and immunomodulatory cytokines, tumour necrosis factor (TNF) and interleukin 6 (IL-6), in various disease states. RESULTS: Several non-redundant functions of TNF from distinct cellular sources were identified. TNF from myeloid cells is pathogenic in several autoimmune diseases, whereas TNF produced by T cells showed non-redundant protective functions in experimental arthritis and in a Mycobacterium tuberculosis infection model. To test the idea of selective pharmacological inhibition of "bad" TNF produced by myeloid cells while sparing "good" TNF produced by T lymphocytes, a myeloid-specific TNF inhibitor (MYSTI) was designed-a recombinant mini-antibody with dual specificity that can bind to the surface molecule F4/80 on myeloid cells and to TNF. In vitro experiments confirmed retention of TNF on the surface of TNF-producing cells and in vivo experiments indicated that MYSTI can protect mice from lethal TNF-mediated hepatotoxicity. MYSTI is also effective in experimental arthritis. CONCLUSION: The proposed therapeutic strategy may be more effective than systemic anti-cytokine therapy in several human autoimmune diseases, as it would preserve the potentially beneficial effects of the same cytokine produced by other cell types. Such bispecific biological agents may become interesting tools for experimental studies and, eventually, drug development.

Microbiota Induces Expression of Tumor Necrosis Factor in Postnatal Mouse Skin.

Tumor necrosis factor (TNF) is a pleiotropic cytokine that regulates many important processes in the body. TNF production in a physiological state supports the structure of lymphoid organs and determines the development of lymphoid cells in hematopoiesis. However, chronic TNF overexpression leads to the development of various autoimmune disorders. Sites of TNF production in the naïve state remain unclear due to the lack of in vivo models. In the present study, we used TNF-2A-Kat reporter mice to monitor the expression of TNF in different tissues. Comparative analysis of tissue fluorescence in TNF-2A-Kat reporter mice and wild type mice revealed constitutive expression of TNF in the skin of naïve adult mice. In the skin of TNF-2A-Kat reporter mouse embryos, no statistically significant differences in the expression of TNF compared to wild type animals were observed. Furthermore, we established that local depletion of microflora with topical antibiotics leads to a reduction in the fluorescence signal. Thus, we assume that the skin microflora is responsible for the expression of TNF in the skin of mice.

An activated unfolded protein response promotes retinal degeneration and triggers an inflammatory response in the mouse retina.

Recent studies on the endoplasmic reticulum stress have shown that the unfolded protein response (UPR) is involved in the pathogenesis of inherited retinal degeneration caused by mutant rhodopsin. However, the main question of whether UPR activation actually triggers retinal degeneration remains to be addressed. Thus, in this study, we created a mouse model for retinal degeneration caused by a persistently activated UPR to assess the physiological and morphological parameters associated with this disease state and to highlight a potential mechanism by which the UPR can promote retinal degeneration. We performed an intraocular injection in C57BL6 mice with a known unfolded protein response (UPR) inducer, tunicamycin (Tn) and examined animals by electroretinography (ERG), spectral domain optical coherence tomography (SD-OCT) and histological analyses. We detected a significant loss of photoreceptor function (over 60%) and retinal structure (35%) 30 days post treatment. Analysis of retinal protein extracts demonstrated a significant upregulation of inflammatory markers including interleukin-1ß (IL-1ß), IL-6, tumor necrosis factor-α (TNF-α), monocyte chemoattractant protein-1 (MCP-1) and IBA1. Similarly, we detected a strong inflammatory response in mice expressing either Ter349Glu or T17M rhodopsin (RHO). These mutant rhodopsin species induce severe retinal degeneration and T17M rhodopsin elicits UPR activation when expressed in mice. RNA and protein analysis revealed a significant upregulation of pro- and anti-inflammatory markers such as IL-1ß, IL-6, p65 nuclear factor kappa B (NF-kB) and MCP-1, as well as activation of F4/80 and IBA1 microglial markers in both the retinas expressing mutant rhodopsins. We then assessed if the Tn-induced inflammatory marker IL-1ß was capable of inducing retinal degeneration by injecting C57BL6 mice with a recombinant IL-1ß. We observed ~19% reduction in ERG a-wave amplitudes and a 29% loss of photoreceptor cells compared with control retinas, suggesting a potential link between pro-inflammatory cytokines and retinal pathophysiological effects. Our work demonstrates that in the context of an established animal model for ocular disease, the persistent activation of the UPR could be responsible for promoting retinal degeneration via the UPR-induced pro-inflammatory cytokine IL-1ß.

Microbiota, intestinal immunity, and mouse bustle.

The composition of the intestinal microbiota is regulated by the immune system. This paper discusses the role of cytokines and innate immunity lymphoid cells in the intestinal immune regulation by means of IgA.

Distinct biological activity of lipopolysaccharides with different lipid A acylation status from mutant strains of Yersinia pestis and some members of genus Psychrobacter.

Correlation between the chemical structure of lipid A from various Gram-negative bacteria and biological activity of their lipopolysaccharide (LPS) as an agonist of the innate immune receptor Toll-like receptor 4 was investigated. Purified LPS species were quantitatively evaluated by their ability to activate the production of tumor necrosis factor (TNF) by murine bone marrow-derived macrophages in vitro. Wild-type LPS from plague-causing bacteria Yersinia pestis was compared to LPS from mutant strains with defects in acyltransferase genes (lpxM, lpxP) responsible for the attachment of secondary fatty acid residues (12:0 and 16:1) to lipid A. Lipid A of Y. pestis double ΔlpxM/ΔlpxP mutant was found to have the chemical structure that was predicted based on the known functions of the respective acyltransferases. The structures of lipid A from two members of the ancient psychrotrophic bacteria of the genus Psychrobacter were established for the first time, and biological activity of LPS from these bacteria containing lipid A fatty acids with shorter acyl chains (C10-C12) than those in lipid A from LPS of Y. pestis or E. coli (C12-C16) was determined. The data revealed a correlation between the ability of LPS to activate TNF production by bone marrow-derived macrophages with the number and the length of acyl chains within lipid A.

Modern anti-cytokine therapy of autoimmune diseases.

The emergence of genetically engineered biological agents opened new prospects in the treatment of autoimmune and inflammatory diseases. Cytokines responsible for regulation of a wide range of processes during development of the normal immune response are among the most successful therapeutic targets. Studies carried out in recent decades and accompanied by rapid development of biotechnology have promoted establishing in detail the role and place of cytokines in autoimmune and inflammatory pathologies. Nevertheless, mechanisms that underlie anti-cytokine therapy are still not fully understood. This review examines the role of such cytokines as TNF, IL-1, and IL-6 in the development of inflammatory processes and the action mechanisms of their inhibitors.

Experimental models of arthritis in which pathogenesis is dependent on TNF expression.

Rheumatoid arthritis (RA) is an autoimmune inflammatory disease characterized by joint damage as well as systemic manifestations. The exact cause of RA is not known. Both genetic and environmental factors are believed to contribute to the development of this disease. Increased expression of tumor necrosis factor (TNF) has been implicated in the pathogenesis of RA. Currently, the use of anti-TNF drugs is one of the most effective strategies for the treatment of RA, although therapeutic response is not observed in all patients. Furthermore, due to non-redundant protective functions of TNF, systemic anti-TNF therapy is often associated with unwanted side effects such as increased frequency of infectious diseases. Development of experimental models of arthritis in mice is necessary for studies on the mechanisms of pathogenesis of this disease and can be useful for comparative evaluation of various anti-TNF drugs. Here we provide an overview of the field and present our own data with two experimental models of autoimmune arthritis - collagen-induced arthritis and antibody-induced arthritis in C57Bl/6 and BALB/c mice, as well as in tnf-humanized mice generated on C57Bl/6 background. We show that TNF-deficient mice are resistant to the development of collagen-induced arthritis, and the use of anti-TNF therapy significantly reduces the disease symptoms. We also generated and evaluated a fluorescent detector of TNF overexpression in vivo. Overall, we have developed an experimental platform for studying the mechanisms of action of existing and newly developed anti-TNF drugs for the treatment of rheumatoid arthritis.

[Preparation and characterization of mouse embryonic fibroblasts with K72W mutation in somatic cytochrome C gene].

Mouse embryonic fibroblasts (MEF) with point mutation in somatic cytochrome C gene were generated and characterized. It was shown that substitution of lysine for tryptophan in position 72 (K72W) decreased the proapoptotic functions of cytochrome C in response to staurosporin treatment without disrupting its respiratory functions. The presence of this mutation did not affect the pattern of cytochrome C gene expression or its localization inside the cell. These cell cultures therefore represent an interesting model for study apoptotic signaling and physiological functions of cytochrome C.

Tumor Necrosis Factor and the consequences of its ablation in vivo.

Although TNF has been discovered due to anti-tumor activity, its physiological functions are different. Current knowledge places TNF downstream of many receptors of innate immunity, implying its primary role in host defense and inflammation. When overproduced systemically or locally, TNF may exert deleterious effects on the organism. Anti-TNF therapy is highly efficient in several autoimmune and inflammatory diseases. However, due to TNF unique beneficial functions in immune system, such therapy cannot be entirely free of adverse effects. We review the current status of the field with the focus on drugs and strategies used for TNF ablation in vivo.

[Chromosomal localization and molecular organization of human genomic fragment containing TNF/LT locus in transgenic mice].

Molecular organization, copy number and chromosomal localization of human TNF/LT locus fragment were determined in genomes of two transgenic mouse lines. Genome of the first one contains two copies, organized in head-to-tail manner and determined on eighth chromosome by karyotyping; single transgene copy of the second line is observed on the fifth chromosome. These mice could serve as valuable model for studying both human tumor necrosis factor and lymphotoxin physiological functions.

[Pharmacokinetics of azlocillin in critical conditions: an individualized schedule of drug administration in relation to anatomo-physiologic and pathologic factors]. / Farmakokinetika azlotsillina pri kriticheskikh sostoianiiakh: individualizatsiia skhemy vvedeniia preparata s uchetom anatomo- fiziologicheskikh i patologicheskikh faktorov.

Azlocillin pharmacokinetics was studied after a single intravenous injection of the antibiotic in a dose of 4 g in 20 patients in critical state. To elucidate the causes of significant individual variability of the antibiotic pharmacokinetics observed in the patients, multiple correlation analysis of the main pharmacokinetic parameters i. e. the area under the concentration/time curve, total clearance, steady-state volume of distribution and mean residence time was performed in regard to the "patient factors" such as sex, age, the volumes of transfused liquid, blood, plasma and blood substitutes, hemoglobin levels, erythrocyte count and ESR. Adequate correspondence of the predicted by the "patient factor" values of the areas under the concentration/time curve and the total clearance to the actually determined values was observed. Correspondence of the predicted values to the steady-state volume of distribution and the mean residence time to the actually determined values was satisfactory. A procedure for design of azlocillin individual dosage regimens based on calculating individual clearance by the "patient factors" is described.