Biochemistry (Moscow) (v.81, #11)

Molecular and cellular mechanisms of inflammation by D. V. Kuprash; S. A. Nedospasov (1237-1239).
Inflammation is one of the most fundamental and pronounced protective reactions of the organism. From ancient times to the present day, complex and diverse patterns of inflammation development and their role in various diseases have attracted attention of investigators. This issue of Biokhimiya/Biochemistry (Moscow) includes experimental studies and reviews dedicated to various aspects of this important and interesting problem.
Keywords: signs of inflammation; infection; tissue damage; phagocytosis; innate immunity; inflammatory mediators; polymorphism

Mechanisms of inflammatory injury of renal tubular cells in a cellular model of pyelonephritis by M. A. Morosanova; E. Y. Plotnikov; L. D. Zorova; I. B. Pevzner; V. A. Popkov; D. N. Silachev; S. S. Jankauskas; V. A. Babenko; D. B. Zorov (1240-1250).
Previously, we have assembled a cellular model of pyelonephritis which contains a primary culture of renal tubular epithelial cells, mononuclear leukocytes, and bacterial lysate or lipopolysaccharide. After cocultivation of renal cells with leukocytes and bacterial lysate, proinflammatory changes were observed in the renal cells, followed by nitrosative and oxidative stress and cell death. The interaction of bacterial antigens not only with leukocytes, but also with epithelial cells of the renal tubules, was partially mediated by signaling pathways involving Toll-like receptors (TLR2 and TLR4). Activation of these receptors led to increased levels of oxidative stress and synthesis of proinflammatory cytokines (TNF, IL-6, IL-1α) in the renal epithelium, while TLR4 blockade decreased the severity of these processes. Apart from the fact that activation of inflammatory signaling in response to bacterial antigens is observed directly in the renal cells, the presence of leukocytes significantly amplifies the inflammatory response as measured by the level of cytokines generated in the ensemble. In the presence of activated leukocytes, higher expression of TLR2 on the surface of renal cells was observed in response to exposure to bacterial components, which might explain the increased inflammatory response in the presence of leukocytes. The synthesis of IL-1α in the epithelial cells of the renal tubules in this inflammatory model leads to its accumulation in the nuclei, which has been reduced by the TLR4 antagonist polymyxin. TLR2 agonists also led to increased levels of IL-1α. The elevation in the content of IL-1α in nuclei was accompanied by increased acetylation of nuclear proteins, which has been reduced to control values after exposure to protective agents (Trolox, mitochondria-targeted antioxidant SkQR1 or LiCl). The high level of acetylation of histones is probably regulated by proinflammatory cytokines, and to some extent it is a marker of inflammation, which can indirectly be reduced by protective agents.
Keywords: cytokines; acetylation; inflammation; pyelonephritis

Effects of fibroin microcarriers on inflammation and regeneration of deep skin wounds in mice by A. Y. Arkhipova; M. A. Nosenko; N. V. Malyuchenko; R. V. Zvartsev; A. M. Moisenovich; A. S. Zhdanova; T. V. Vasil’eva; E. A. Gorshkova; I. I. Agapov; M. S. Drutskaya; S. A. Nedospasov; M. M. Moisenovich (1251-1260).
The process of tissue regeneration following damage takes place with direct participation of the immune system. The use of biomaterials as scaffolds to facilitate healing of skin wounds is a new and interesting area of regenerative medicine and biomedical research. In many ways, the regenerative potential of biological material is related to its ability to modulate the inflammatory response. At the same time, all foreign materials, once implanted into a living tissue, to varying degree cause an immune reaction. The modern approach to the development of bioengineered structures for applications in regenerative medicine should be directed toward using the properties of the inflammatory response that improve healing, but do not lead to negative chronic manifestations. In this work, we studied the effect of microcarriers comprised of either fibroin or fibroin supplemented with gelatin on the dynamics of the healing, as well as inflammation, during regeneration of deep skin wounds in mice. We found that subcutaneous administration of microcarriers to the wound area resulted in uniform contraction of the wounds in mice in our experimental model, and microcarrier particles induced the infiltration of immune cells. This was associated with increased expression of proinflammatory cytokines TNF, IL-6, IL-1β, and chemokines CXCL1 and CXCL2, which contributed to full functional recovery of the injured area and the absence of fibrosis as compared to the control group.
Keywords: proinflammatory cytokines; microparticles; fibroin; IL-6; TNF

The cellular microenvironment directly and indirectly influences tumor development and possesses prognostic and in some cases diagnostic value. Over the years, understanding of structural organization of the immune/inflammatory moiety of neoplasms as well as in-depth phenotypic and transcriptomic profiling of its cellular components together provide more and more insights in both basic and translational medical science. In this review, we will discuss the specific roles of various stromal cells and their impact on neoplastic progression as well as address the use of quantitative and phenotypic analysis of immune/inflammatory infiltrate for diagnostics and predicting the clinical course of human malignancies.
Keywords: cellular microenvironment of tumors; immune infiltration; melanoma; breast cancer; thyroid cancer; colorectal cancer

Myeloid-derived suppressor cells represent a heterogeneous population of immature myeloid cells. Under normal conditions, these cells differentiate into macrophages, dendritic cells, and granulocytes. However, in pathological states such as inflammation, infection, or tumor growth, there is an arrest of their differentiation that results in the accumulation of immature myeloid cells in the organism. In addition, these cells acquire a suppressor phenotype, expressing anti-inflammatory cytokines and reactive oxygen and nitrogen species, and suppress T-cell immune response. Myeloid-derived suppressor cells (MDSC) contribute to cancerogenesis by forming a favorable microenvironment for tumor growth. Proinflammatory cytokines, secreted by tumor cells and the tumor microenvironment, induce angiogenesis and metastasis and promote tumor growth. They also provide signals necessary for survival, accumulation, and function of MDSC. Understanding the mechanisms of myeloid suppressor cell development and the use of proinflammatory cytokine inhibitors may prove beneficial for tumor therapy.
Keywords: MDSC; TNF; IL-6; IL-1; tumor microenvironment; tumor-associated inflammation

“Suppressor factor” of neutrophils: A short story of a long-term misconception by I. A. Linge; E. V. Kondratieva; T. K. Kondratieva; V. A. Makarov; V. I. Polshakov; O. Yu. Savelyev; A. S. Apt (1284-1292).
A large body of evidence obtained during the last decade has demonstrated that neutrophils suppress T cell proliferation in different models of inflammation and cell interaction. The commonly used method for assessing cell proliferation and proliferation inhibition is measuring [3H]thymidine incorporation into cells. Earlier, we observed inhibition of [3H]thymidine uptake in experiments on neutrophil-mediated regulation of T cell response in tuberculosis immunity. Here, we used different types of proliferating cells to analyze the nature of the soluble “neutrophil factor” by a variety of methods (dialysis, HPLC, mass spectrometry, and NMR) and unambiguously demonstrated that neutrophils do not synthesize a specific factor inhibiting cell proliferation, but secrete high concentrations of extracellular thymidine that competitively inhibit [3H]thymidine incorporation. Although the physiological significance of thymidine secretion by neutrophils remains unknown, this phenomenon should be carefully considered when designing test systems for studying cell–cell interactions.
Keywords: neutrophils; thymidine; proliferation inhibition

Mediators and biomarkers of inflammation in meningitis: Cytokine and peptidome profiling of cerebrospinal fluid by A. A. Belogurov Jr.; O. M. Ivanova; Y. A. Lomakin; R. H. Ziganshin; M. I. Vaskina; V. D. Knorre; E. A. Klimova; A. G. Gabibov; V. T. Ivanov; V. M. Govorun (1293-1302).
Differential diagnosis of bacterial and viral meningitis is an urgent problem of the modern clinical medicine. Early and accurate detection of meningitis etiology largely determines the strategy of its treatment and significantly increases the likelihood of a favorable outcome for the patient. In the present work, we analyzed the peptidome and cytokine profiles of cerebrospinal fluid (CSF) of 17 patients with meningitis of bacterial and viral etiology and of 20 neurologically healthy controls. In addition to the identified peptides (potential biomarkers), we found significant differences in the cytokine status of the CSF of the patients. We found that cut-off of 100 pg/ml of IL-1β, TNF, and GM-CSF levels discriminates bacterial and viral meningitis with 100% specificity and selectivity. We demonstrated for the first time the reduction in the level of two cytokines, IL-13 and GM-CSF, in the CSF of patients with viral meningitis in comparison with the controls. The decrease in GM-CSF level in the CSF of patients with viral meningitis can be explained by a disproportionate increase in the levels of cytokines IL-10, IFN-γ, and IL-4, which inhibit the GM-CSF expression, whereas IL-1, IL-6, and TNF activate it. These observations suggest an additional approach for differential diagnosis of bacterial and viral meningitis based on the normalized ratio IL-10/IL-1β and IL-10/TNF > 1, as well as on the ratio IFN-γ/IL-1β and IFN-γ/ TNF < 0.1. Our findings extend the panel of promising clinical and diagnostic biomarkers of viral and bacterial meningitis and reveal opposite changes in the cytokine expression in meningitis due to compensatory action of proand antiinflammatory factors.
Keywords: inflammation; meningitis; cytokines; biomarkers; peptidome; cerebrospinal fluid

Microbiota induces expression of tumor necrosis factor in postnatal mouse skin by D. V. Yuzhakova; M. V. Shirmanova; A. A. Bocharov; I. V. Astrakhantseva; E. A. Vasilenko; E. N. Gorshkova; M. S. Drutskaya; E. V. Zagaynova; S. A. Nedospasov; A. A. Kruglov (1303-1308).
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 naive 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 naive 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.
Keywords: tumor necrosis factor (TNF); TNF-2A-Kat reporter mice; in vivo ; fluorescent imaging

Tumor necrosis factor and lymphotoxin in regulation of intestinal inflammation by E. O. Gubernatorova; A. V. Tumanov (1309-1325).
Ulcerative colitis and Crohn’s disease are the major forms of inflammatory bowel disease. Cytokines of the tumor necrosis factor (TNF) family play an important role in the regulation of intestinal inflammation. In this review, we discuss the function of key cytokines of this family–TNF and lymphotoxin (LT)–in mucosal healing, IgA production, and in control of innate lymphoid cells (ILCs), novel regulators of mucosal homeostasis in the gut. TNF plays a central role in the pathogenesis of inflammatory bowel diseases (IBD). LT regulates group 3 of ILCs and IL-22 production and protects the epithelium against damage by chemicals and mucosal bacterial pathogens. In addition, we discuss major mouse models employed to study the mechanism of intestinal inflammation, their advantages and limitations, as well as application of TNF blockers in the therapy for IBD.
Keywords: tumor necrosis factor; lymphotoxin; intestinal inflammation; inflammatory bowel disease; mouse models

Inactivation of inflammasomes by pathogens regulates inflammation by F. Yu. Garib; A. P. Rizopulu; A. A. Kuchmiy; V. F. Garib (1326-1339).
Inflammatory response is initiated and sustained by the action of quintessential pro-inflammatory cytokines of immune system namely IL-1β and IL-18. The maturation process of those cytokines is ensured by caspase-1 enzymatic activity, that is in turn is tightly controlled by multiprotein complexes called inflammasomes. Inflammasomes are activated in cells of innate immune system in response to recognition of conservative parts of microbes (pathogen-associated molecular patterns) or by sensing molecular signs of tissue damage (damage-associated molecular patterns). Inflammasome activation apart of cytokines secretion leads to pro-inflammatory cell death, so-called pyroptosis. That culminates in release of cytoplasmatic content of cells including cytokines and alarmins that boost immune response against pathogens, as well as pyroptosis destroys replicative niches of intracellular pathogens. During co-evolution with the host, bacterial and viral pathogens developed a range of molecular inhibitors targeting each step of inflammasome activation. In current review, we will discuss the latest knowledge of inflammasomes’ signaling pathways and tricks that pathogens use to avoid immune recognition and clearance. Our better understanding of inflammasome inhibition by pathogens can lead to better therapeutic approaches for the treatment of infectious diseases.
Keywords: inflammasome; IL-1β; IL-18; evasion; bacterial and viral pathogens; pyroptosis

Mechanisms of changes in immune response during bacterial coinfections of the respiratory tract by E. N. Sviriaeva; K. V. Korneev; M. S. Drutskaya; D. V. Kuprash (1340-1349).
Acute diseases of the respiratory tract are often caused by viral pathogens and accompanying secondary bacterial infections. It is known that the development of such bacterial complications is caused mainly by a decreased infiltration with immune system cells and by suppressed inflammation in the lungs. There are significant advances in understanding the mechanisms of secondary infections, although many details remain unclear. This review summarizes current knowledge of the molecular and cellular changes in the host organism that can influence the course of bacterial coinfections in the respiratory tract.
Keywords: secondary bacterial coinfection; influenza; bacteria; pneumonia; macrophages; Toll-like receptor

Possible mechanisms of acquisition of herpesvirus virokines by E. A. Gorshkova; E. S. Shilov (1350-1357).
The genomes of certain types of human and primate herpesviruses contain functional homologs of important host cytokines (IL-6, IL-17, and IL-10), or so-called virokines. Virokines can interact with immune cell receptors, transmit a signal to them, and thus switch the type of immune response that facilitates viral infection development. In this work, we have summarized possible ways of virokine origin and proposed an evolutionary scenario of virokine acquisition with involvement of retroviral coinfection of the host. This scenario is probably valid for vIL-6 of HHV-8 and MRV-5 viruses, vIL-17 of HVS virus, and vIL-10 of HHV-4, Bonobo-HV, RhLCV, and BaLCV viruses. The ability to acquire cytokine genes allows herpesviruses to implement unique strategies of avoiding the immune response and provides them an evolutionary advantage: more than 90% of the host population can be chronically infected with different herpesviruses. It is possible that the biological success of herpesviruses can be partially due to their cooperation with another group of viruses. This hypothesis emphasizes the importance of studies on the reciprocal influence of pathogens on their coinfection, as well as their impact on the host organism.
Keywords: virokines; IL-6; IL-10; IL-17; herpesviruses; retroviruses; horizontal gene transfer

The role of cytokines in the development of atherosclerosis by A. R. Fatkhullina; I. O. Peshkova; E. K. Koltsova (1358-1370).
Atherosclerosis contributes to the development of many cardiovascular diseases, which remain the leading cause of death in developed countries. Atherosclerosis is a chronic inflammatory disease of large and medium-sized arteries. It is caused by dyslipidemia and mediated by both innate and adaptive immune responses. Inflammation is a key factor at all stages of atherosclerosis progression. Cells involved in pathogenesis of atherosclerosis were shown to be activated by soluble factors, cytokines, that strongly influence the disease development. Pro-inflammatory cytokines accelerate atherosclerosis progression, while anti-inflammatory cytokines ameliorate the disease. In this review, we discuss the latest findings on the role of cytokines in the development and progression of atherosclerosis.
Keywords: cardiovascular diseases; atherosclerosis; inflammation; immune cells; adhesion molecules; cytokines

Recombinant MHC tetramers for isolation of virus-specific CD8+ cells from healthy donors: Potential approach for cell therapy of posttransplant cytomegalovirus infection by A. S. Vdovin; S. Y. Filkin; P. R. Yefimova; S. A. Sheetikov; N. M. Kapranov; Y. O. Davydova; E. S. Egorov; E. G. Khamaganova; M. Y. Drokov; L. A. Kuzmina; E. N. Parovichnikova; G. A. Efimov; V. G. Savchenko (1371-1383).
Patients undergoing allogeneic hematopoietic stem cell transplantation have a high risk of cytomegalovirus reactivation, which in the absence of T-cell immunity can result in the development of an acute inflammatory reaction and damage of internal organs. Transfusion of the virus-specific donor T-lymphocytes represents an alternative to a highly toxic and often ineffective antiviral therapy. Potentially promising cell therapy approach comprises transfusion of cytotoxic T-lymphocytes, specific to the viral antigens, immediately after their isolation from the donor’s blood circulation without any in vitro expansion. Specific T-cells could be separated from potentially alloreactive lymphocytes using recombinant major histocompatibility complex (MHC) multimers, carrying synthetic viral peptides. Rapid transfusion of virus-specific T-cells to patients has several crucial advantages in comparison with methods based on the in vitro expansion of the cells. About 30% of hematopoietic stem cell donors and 46% of transplant recipients at the National Research Center for Hematology were carriers of the HLA-A*02 allele. Moreover, 94% of Russian donors have an immune response against the cytomegalovirus (CMV). Using recombinant HLA-A*02 multimers carrying an immunodominant cytomegalovirus peptide (NLV), we have shown that the majority of healthy donors have pronounced T-cell immunity against this antigen, whereas shortly after the transplantation the patients do not have specific T-lymphocytes. The donor cells have the immune phenotype of memory cells and can be activated and proliferate after stimulation with the specific antigen. Donor lymphocytes can be substantially enriched to significant purity by magnetic separation with recombinant MHC multimers and are not activated upon cocultivation with the antigen-presenting cells from HLA-incompatible donors without addition of the specific antigen. This study demonstrated that strong immune response to CMV of healthy donors and prevalence of HLA-A*02 allele in the Russian population make it possible to isolate a significant number of virus-specific cells using HLA-A*02–NLV multimers. After the transfusion, these cells should protect patients from CMV without development of allogeneic immune response.
Keywords: cytomegalovirus; cell therapy; adoptive transfer; allogeneic hematopoietic stem cell transplantation; MHC multimers

Study of immunomodulatory effects of extracellular HSP70 in a mouse model of allergic airway inflammation by M. A. Shevchenko; N. I. Troyanova; E. A. Servuli; E. L. Bolkhovitina; A. S. Fedorina; A. M. Sapozhnikov (1384-1395).
Immunostimulatory properties of extracellular heat shock proteins 70 kDa (HSP70) became interesting for investigators a long time ago. However, in recent years a series of works showing a significant relation of the immunostimulating effects of recombinant HSP70 to contamination of the protein samples with bacterial endotoxins (lipopolysaccharide, LPS) has been published. The authors showed that intensive elimination of LPS from the protein samples resulted in inversion of immunostimulating effects of HSP70 to immunosuppressive activity of the protein. Nevertheless, at present the conception of immunostimulating, proinflammatory action of extracellular HSP70 is the most common. In this work, we studied immunomodulatory effects of exogenous HSP70 in a mouse model of allergic inflammation of airways. We also analyzed the dynamics of the level of the extracellular pool of HSP70 in the site of inflammation. The results demonstrated a considerable content of extracellular HSP70 in bronchoalveolar lavages with dynamics reflecting the stages of development of the induced inflammation. Oropharyngeal injection of exogenous HSP70 in the acute phase of allergic inflammation of airways resulted in significant suppression of the inflammatory process, which conforms to published data demonstrating an immunosuppressive activity of the extracellular pool of HSP70.
Keywords: heat shock protein 70 kDa; HSP70; airway allergic inflammation; immunomodulatory effects

Immunogenicity of human interferon-beta-containing pharmaceuticals by V. D. Nazarov; S. V. Lapin; A. V. Mazing; E. P. Evdoshenko; A. A. Totolian (1396-1400).
Multiple sclerosis is a severe autoimmune disease with inflammatory component that continues to be resistant to treatment. One of the approaches retarding its progression is based on using nonspecific therapy with human interferon-beta (IFN-β)-containing pharmaceuticals. Neutralizing antibodies (NAbs) against genetically engineered pharmaceuticals developed by the patient’s immune system, which reduce their therapeutic and biological activity, pose a serious problem. Cell lines sensitive to IFN-β activity also quantifying NAb level are applied because direct measurement of IFN-β antiviral activity is complicated. This study was aimed at standardization and validation of a reporter cell system for measuring antihuman IFN-β NAb titers, and evaluation data were obtained with samples from 33 patients with multiple sclerosis.
Keywords: multiple sclerosis; immunogenicity; interferon-beta; neutralizing antibodies; titer