DISSEMINATED PURULENT PERITONITIS OUTCOME AFFECTS NKT CELL PHENOTYPE

  • Authors: Savchenko A.A.1, Borisov A.G.2, Kudryavtsev I.V.3,4, Belenjuk V.D.2
  • Affiliations:
    1. Federal Research Center «Krasnoyarsk Science Center» of the Siberian Branch of the Russian Academy of Sciences, Scientific Research Institute of medical problems of the Northl Siberian Federal University
    2. Federal Research Center «Krasnoyarsk Science Center» of the Siberian Branch of the Russian Academy of Sciences, Scientific Research Institute of medical problems of the North
    3. Pavlov First St. Petersburg State Medical University, St.Petersburg, Russian Federation
    4. Federal State Budgetary Scientific Institution "Institute of Experimental Medicine" (FSBSI "IEM"), St.Petersburg, Russian Federation
  • Section: ORIGINAL ARTICLES
  • URL: https://iimmun.ru/iimm/article/view/2004
  • DOI: https://doi.org/10.15789/2220-7619-DPP-2004

Cite item

Abstract

The aim of our study was to investigate the main characteristics of peripheral blood NKT cell phenotype in patients with disseminated purulent peritonitis (DPP) in dynamics of postoperative period, depending on the disease outcome. Fifty-two patients with acute surgical diseases and injuries of the abdominal organs complicated by DPP, and 68 healthy individuals in control group, were examined. Blood sampling was performed before surgery (preoperative period), as well as on the day 7, 14 and 21 of postoperative period. All patients with DPP were divided into two groups depending on disease outcome in postoperative period: patients with favorable disease outcome (n = 34); and patients with unfavorable outcome (n = 18). Study of the phenotype of blood NKT lymphocytes was performed by flow cytometry using direct immunofluorescence of whole peripheral blood samples with monoclonal antibodies. The low relative and absolute level of NKT cells was observed in DPP patients regardless of outcome disease in preoperative period. At the same time, the absolute level of NKT cells returned to normal only in patients with favorable DPP outcome and only by day 21 after surgery. Patients with favorable DPP outcome by the end of examination period had normalized quantity of mature NKT-lymphocytes and significantly decreased level of cytotoxic cells which was apparently associated with migration of such cell subsets to site of inflammation. A reduced level of non-classical (expressing CD8 marker) mature and cytokine-producing NKT cells was detected only in patients with favorable DPP outcome in preoperative period which returned to normal by the end of postoperative period. At the same time, patients with unfavorable disease outcome had reduced quantity of NKT cells of these subsets by day 21 of postoperative treatment. Patients with favorable outcome had high level of mature and cytotoxic CD11b+ NKT cells already in the preoperative period, while patients with unfavorable DPP outcome had increased level of cytotoxic CD11b+ NKT cells only by day 21 after surgery. The proportion of NKT cells expressing activation markers (CD28 and CD57) was reduced in patients in preoperative period that returned to normal immediately after surgery with favorable outcome, while it recovered with unfavorable outcome closer to the end of postoperative examination. The defined features of NKT cell phenotype in patients with unfavorable DPP outcome characterize disturbances in subset ratio and mechanisms of functioning of this cell fraction. This determines a need to develop immunotherapeutic methods aimed at stimulating immunoregulatory activity of NKT cells.

Full Text

FEATURES OF THE NKT CELL PHENOTYPE IN DEPENDING ON THE WIDESPREAD PURULENT PERITONITIS OUTCOME

 

Introduction

Widespread purulent peritonitis (WPP) remains one of the unsolved problems of modern abdominal surgery due to high morbidity and mortality. The disease is a complication of a number of surgical diseases or abdominal injuries (acute appendicitis, perforated gastric and duodenal ulcer, acute gangrenous cholecystitis, pancreatic necrosis, perforation of hollow organs, their damage during trauma, etc.) in the vast majority of cases [22, 27, 28]. Mortality in WPP is about 20-30% reaching the highest figures in the development of multiple organ failure and septic shock, the prevention and relief of which are key in the treatment of peritonitis [33, 34].

It has been proven that the course of the infectious process in the abdominal cavity, the nature and characteristics of the development of purulent postoperative complications are determined not only by the severity of the underlying disease, the adequacy of the surgical intervention performed and the completeness of the intensive care but also by the functional state of the immune system [3, 19, 24]. Yang L. et al. (2022) reported that patients with spontaneous bacterial peritonitis had IL-13 overexpression in their ascites and a reduced functional activity of CD8+ T cells [41]. There is evidence that an increased level of macrophage mannose receptor (CD206) expression on peritoneal macrophages was associated with an increased risk of an adverse outcome of peritonitis [38]. Previously, we found that the number of ‘naïve’ B lymphocytes and B2 cells non-expressing and expressing CD23 in patients with an unfavorable outcome of WPP was higher than in patients with a favorable outcome of this disease [2]. Violations of the mechanisms of the respiratory burst of neutrophils as well as a decrease in their phagocytic activity, and the level of TNF-α synthesis in patients with WPP were previously identified and presented in a number of publications [5, 21].

Natural killer T cells (NKT) are defined as a heterogeneous subset of T lymphocytes with a CD3+CD16/56+ phenotype, i.e. they combine the phenotypic characteristics of T and NK cells [11, 40]. Accordingly, the functional activity of NKT cells is realized in various mechanisms of the immune response, realizing the relationship between natural resistance and adaptive immunity. This lymphocyte subset was involved in the mechanisms of antiviral and antiparasitic protection, and also secreted different immunoregulatory cytokines in the site of inflammation [1, 10, 12]. In addition, NKT cells were able to stimulate and inhibit antitumor immune responses [9, 39].

Semi-invariant αβ-TCR expressed by NKT cells could recognize α-glycuronylceramides (one of the main components of gram-negative bacteria cell wall) that led to the formation of a complex of antimicrobial functions including those mediated by the induction of CD40L and pronounced stimulation of Th1 and Th2 lymphocytes [10, 16]. Therefore, NKT cells are also involved in immune-inflammatory processes development. For instance, Nilsson J. et al. demonstrated that NKT cells cytokine profile switching regulated liver sterile inflammation [31]. It was shown that NKT cells made a significant contribution to mucosal immunity regulation by intestinal homeostasis controlling and participating in the development of inflammatory diseases of the abdominal cavity [11].

Thus, the aim of our investigation was to study the characteristics of NKT cell phenotype in patients with WPP in the dynamics of the postoperative period in depending on the outcome of the disease.

 

Materials and Methods

Study participants. 52 patients with acute surgical diseases and injuries of the abdominal organs complicated by WPP aged 25-65 years (the mean age of the patients was 49.6 years) who were treated at the Krasnoyarsk regional purulent-septic center at the Regional Clinical Hospital were examined. Exclusion criteria from the study were the presence of acute destructive pancreatitis (pancreatic necrosis), total mesenteric thrombosis, oncological diseases and tuberculosis. The volume of surgical intervention and the number of sanations were determined by the attending physician depending on the patient's condition. Blood sampling was performed before the surgery (preoperative period) as well as on the 7th, 14th and 21st days of the postoperative period. All patients were divided into two groups depending on the outcome of peritonitis in the postoperative period: group 1 - patients with a favorable outcome of the disease (n=34), group 2 - patients with an unfavorable outcome (n=18). 68 healthy people were examined as a control group.

All studies were performed with the informed consent of the patients and in accordance with the Helsinki Declaration of the World Association «Ethical principles of scientific medical research involving human» as amended in 2013 and «Rules of clinical practice in the Russian Federation» approved by the Order of the Ministry of Health of Russia of 19.06.2003 (No. 266).

Flow cytometry. The study of the phenotype of AA cells was performed by flow cytometry using direct immunofluorescence of whole peripheral blood with monoclonal antibodies (Beckman Coulter, USA). The preparation of blood samples and the adjustment of the flow cytometer were carried out in accordance with the recommendations presented in the article by Khaidukov S.V. et al. [7]. The distribution of antibodies along the fluorescence channels was carried out in accordance with the principles of panel formation for multicolor cytofluorometric studies [4]. Immunophenotyping of cells was performed by staining 200 µl of whole EDTA-stabilized blood with the following combination of fluorochrome-conjugated monoclonal antibodies: anti-CD3 Alexa Fluor 700 (clone UCHT1, isotype – Mouse IgG1), anti-CD8 Allophycocyanin (clone B9.11, isotype – Mouse IgG1), anti-CD11b Phycoerythrin-Texas Red-X (clone J33, isotype – Mouse IgG1), anti-CD16 Phycoerythrin-Cyanin 7 (clone 3G8, isotype – Mouse IgG1), anti-CD28 Phycoerythrin (clone CD28.2, isotype – Mouse IgG1), anti-CD45 Alexa Fluor 750 (clone J33, isotype – Mouse IgG1), anti-CD56 Phycoerythrin-Ccyanin 5.5 (clone N901, isotype – Mouse IgG1) and anti-CD57 Fluorescein Isothiocyanate (clone NC1, isotype – Mouse IgM). Incubation of blood samples with antibodies was carried out for 15 min at room temperature and in the dark. Lysis of erythrocytes was carried out for 15 min using 2 ml of VersaLyse Lysing Solution (Beckman Coulter, Inc., USA) with the addition of 50 µl of IOTest 3 Fixative Solution (Beckman Coulter, Inc., USA). Stained cells were analyzed on a Navios flow cytometer (Beckman Coulter, Inc., USA) of the Krasnoyarsk Regional Center of Research Equipment of Federal Research Center «Krasnoyarsk Science Center SB RAS». At least 50.000 lymphocytes were analyzed for each blood sample. The obtained data were analyzed using the Kaluza software package (Beckman Coulter, Inc., USA).

Statistical analysis. The results were presented using the median (Me) and interquartile range as 25th (Q1) and 75th (Q3) percentiles. The significance of differences between the indicators of independent samples was assessed using the nonparametric Mann-Whitney test (Mann-Whitney U test). The significance of differences in indicators in the dynamics of treatment was determined by the Wilcoxon test (Wilcoxon matched pairs test). Friedman's rank analysis of variance (Friedman ANOVA by Ranks) was also used to assess changes in the studied parameters in the dynamics of postoperative treatment. Statistical analysis was carried out using the Statistica 8.0 software package (StatSoft Inc., USA, 2007).

 

Results

We noticed that the absolute and relative numders of NKT cell was decreased in patients in the preoperative period with a favorable outcome of WPP if compared to healthy controls, and this was mainly due to cells with CD3+CD16+CD56+ and CD3+CD16-CD56+ phenotypes (table 1). The absolute number of NKT lymphocytes in patients with a favorable outcome of WPP remained reduced in compared to control values on the 7th day after surgery. An increase in CD3+CD16-CD56+ cells levels if compared with the initial time point was also found, and this content remained until the end of the observed period. Moreover, the consistent increase of CD3+CD16-CD56+ cells frequency was also confirmed by the results of Friedman ANOVA test: c2=13,08, р=0,004. The absolute number of NKT cells in patients with a favorable outcome of WPP remained on the 14th day of postoperative treatment, but with a decrease in the percentage of CD3+CD16+CD56+ cells vs. control group. Patients with a favorable outcome of peritonitis showed a normalization of the absolute number of circulating NKT cells and a reduced percentages of CD3+CD16+CD56- cells if compared to control values by the end of the observed period.

We found that the level of CD8-expressing NKT cells was altered in patients with a favorable outcome of WPP (table 2). Thus, the percentage of peripheral blood CD3+CD8+CD16+CD56+, CD3+CD8+CD16+CD56- and CD3+CD8+CD16-CD56+ cells was reduced in this group of patients even in the preoperative period if compared to healthy controls. The relative number of NKT cells with CD3+CD8+CD16+CD56+ phenotype in patients with a favorable outcome of the disease was increased vs. control ranges on the 7th day after surgery, and it reached its maximum by the end of the observed period. The percentage of CD3+CD8+CD16+CD56- cells in patients of this group increased if compared to control values on the 7th day after surgery, but it decreased again at 14th day point and remained at the initial level until the end of the observed period. The frequency of CD3+CD8+CD16-CD56+ NKT cells was reduced on the 7th day post surgery, their number was increased vs. control values on the 14th day of treatment, and reached its maximum by the end of the observed period. The sequence of changes in the content of NKT cells with this phenotype was also confirmed using the Friedman ANOVA test (c2=9,60, р=0,022).

Next, we investigated the expression of cell adhesion molecules and activation markers by peripheral blood NKT cells (table 3). We found increased levels of CD3+CD16+CD56+CD11b+ and CD3+CD16+CD56-CD11b+ NKT cells in blood samples from WPP patients with a favorable outcome in compared to controls, while the frequencies of CD28- and CD57 expressing NKT cells were decreased (table 3). Moreover, CD3+CD16+CD56+CD11b+ cells in patients of this group were decreased if compared to control values from the 14th day, while the number of CD3+CD16+CD56-CD11b+ cells remained elevated by end of our observed period. The number of CD3+CD16-CD56+CD11b+ NKT cells in the current patients group increased significantly vs. healthy controls only at the end of the observation period, that was also confirmed by the Friedman ANOVA test (c2=12,60, р=0,006). The percentage of NKT cells expressing CD28 and CD57 increased from the beginning of postoperative treatment and remained at the control levels until the end of the examination period.

The percentage of NKT cells in the blood of patients with WPP with an unfavorable outcome of the disease was reduced vs. control levels before the surgery, while their relative number restored after the surgery (table 4). However, the absolute number of NKT cells in patients of this group was reduced in preoperative and postoperative periods if compared to control group. The percentage of CD3+CD16+CD56+ NKT cells in patients with an unfavorable outcome of WPP corresponded to the control values in the preoperative period and within 14 days of subsequent treatment, but it decreased by the end of the observed period. The level of CD3+CD16+CD56- cells in patients of this group also corresponded to the control values in the preoperative period, while it significantly increased by the 7th and 14th days of observation. Moreover, it returned to initial ranges by the 21st day of postoperative treatment. At the same time, the percentage of CD3+CD16-CD56+ NKT cells in patients with an unfavorable outcome of WPP was reduced in the preoperative period, but it reached the control values on the 14th day of postoperative treatment, and then decreased by the end observed period if compared to healthy controls.

The relative numbers of peripheral blood CD3+CD8+CD16+CD56+ and CD3+CD8+CD16+CD56- cells in patients with an unfavorable outcome of WPP in the preoperative period corresponded to the control values (table 5). The frequencies of CD3+CD8+CD16+CD56+ cells were increased by the 7th day after the surgery, but it significantly decreased by the end of the observed period if compared to controls. The level of CD3+CD8+CD16+CD56- cells was increased if compared to control values on the 7th and 14th days of postoperative treatment, but their number decreased to the initial range by the end of the observed period. The percentage of CD3+CD8+CD16-CD56+ cells in patients with an unfavorable outcome of the disease was reduced in the preoperative period, it increased to control values on the 7th and 14th days of treatment, but, finally, it was decreased by the end of the observed period if compared to controls.

The percentage of circulating CD3+CD16+CD56+CD11b+ cells in patients with an unfavorable outcome of WPP in the pre- and postoperative period corresponded to the control range, while the level of CD3+CD16-CD56+CD11b+ cells during the entire examination period exceeded the control values (table 6). The levels of CD3+CD16+CD56-CD11b+ NKT cells in patients of this group in the preoperative period showed no differences with control values. However, the level of this cell subset decreased on the 7th day after the surgery, but then increased by the end of the observed period if compared to healthy controls and initial ranges. The relative numbers of CD28-expressing NKT cells in patients with an unfavorable outcome of the disease were reduced compared to control values in the preoperative period and on the 7th day of the postoperative period, but they reached the control values by the end of the observed period significantly exceeding the initial level. Similarly, CD57-positive NKT cells in this group of patients were reduced in the preoperative period and during the first 14 days of postoperative treatment, but they increased to control values by the end of the observed period.

Differences in NKT cells content were found between patients with favorable and unfavorable outcomes of WPP (tables 1 and 4). Thus, the percentages of CD3+CD16+CD56+ cells were increased in patients with an unfavorable outcome of the disease on the 7th day after surgery (р=0,045), the level of CD3+CD16+CD56- cells was increased on the 14th day (р=0,014) and the relative content of CD3+CD16-CD56+ cells were reduced on the 21st day after surgery (р<0,001) if compared to patients with a favorable outcome of WPP. Additionally, the frequencies of circulating CD3+CD8+CD16+CD56+ cells were increased in the case of an unfavorable outcome of WPP during the preoperative period (р=0,025), and the level of CD3+CD8+CD16-CD56+ NKT cells was reduced if compared (р=0,043) to patients with a favorable outcome of the disease (tables 2 and 5). At the same time, an increased level of CD3+CD8+CD16+CD56+ cells in patients with an unfavorable outcome persisted on the 7th day after surgery (p=0,048), while a decrease in this NKT cell subset (p=0,008) was observed at 21st day of postoperative treatment if compared to patients with a favorable outcome of WPP. The percentage of CD3+CD8+CD16-CD56+ cells in patients with an unfavorable outcome also remained lower at the end of the observed period vs. patients with a favorable outcome of WPP (p<0.001). The relative numbers of CD3+CD8+CD16+CD56- cells in the blood of patients with an unfavorable outcome of WPP on the 7th and 14th days after surgery were also increased (р=0,017 и р<0,001, respectively).

Finally, the percentages of CD3+CD16+CD56+CD11b+ cells in patients with an unfavorable outcome of WPP on the 7th and 21st days of postoperative treatment were reduced vs. patients with a favorable outcome (p=0,003 and p=0,044, respectively) (tables 3 and 6). Similarly, the levels of CD3+CD16+CD56-CD11b+ NKT cells were reduced on the 7th and 14th days of postoperative treatment vs. patients with an unfavorable outcome (p<0,001 and p=0,002, respectively). An increase in relative numbers of CD3+CD16-CD56+CD11b+ cells in patients with an unfavorable outcome vs. patients with a favorable outcome were observed only in the preoperative period (p=0,002). In addition, CD28+ NKT cells were reduced in patients with an unfavorable outcome of WPP in the preoperative period and on the 7th day after surgery (р=0,020 и р=0,028, respectively), and the numbers of CD57+ NKT cells were reduced during the entire observation period (p=0,043, p=0,007, p=0,046 and p=0,039, respectively) relative to the indicators found in patients with a favorable course of this infectious and inflammatory disease.

 

Discussion

The functional activity of NKT cells is realized through effector (perforin/granzyme and/or FasL-mediated) mechanisms and regulatory (cytokine production) providing the relationship between innate and adaptive immunity [6, 12, 40]. The subset composition of NKT cells was determined by the CD16 and CD56 receptors expression. The CD16 is a low affinity immunoglobulin G receptor (FcγRIII) that is non-covalently bound to the CD3ζ molecule on the NKT cell membrane [17, 18]. CD56 (NCAM, Leu-19, NKH-1) is an immunoglobulin superfamily adhesion molecule that takes part in intercellular interaction [8, 23]. Mature NKT cells express both markers. Cells that exhibit CD16+CD56- phenotype were defined as cytotoxic cells, while NKT cells with CD16-CD56+ phenotype were defined as cytokine-producing cells [15, 35].

In general, the relative and absolute numbers of NKT cells in the peripheral blood of patients with WPP were reduced in the preoperative period, regardless of the outcome of the disease. Moreover, if their percentage was restored already on the 7th day after the surgery then the reduced absolute level of this fraction of lymphocytes remained in the postoperative period. Only patients with a favorable outcome of the disease by the end of the observed period (on the 21st day after surgery) had increased percentages of NKT cells similar to control ranges. A feature of the dynamics of the number of NKT cells in the blood in patients with a favorable outcome of WPP was that the reduced level of cells was associated with a low content of mature and cytokine-producing NKT cells in the preoperative period while their number was restored to control values by the end of the observed period and the number of cytotoxic NKT cells decreased. Patients with an unfavorable outcome of WPP in the preoperative period had a low level of NKT cells which was determined by a reduced content of cytokine-producing cells. The low level of cytokine-producing and mature NKT cells was also observed in patients of this group on the 21st day of postoperative treatment.

NKT cells expressing the CD8 marker are part of type II NKT cells (non-classical, non-invariant) [1, 32]. This cell fraction recognizes a wider range of antigenic molecules (compared to type I NKT cells), synthesizes cytokines that induce differentiation of Th1- and Th2-lymphocytes but also implement immunosuppressive functions [1, 14, 36]. In particular, type II NKT cells can stimulate the functional activity of myeloid suppressor cells, able to kill antigen-prescribing dendritic cells and to inhibit the functional activity of cytotoxic CD8+ T cells through the induction of TGF-β expression [29, 30]. The content of mature, cytotoxic and cytokine-producing fractions of NKT cells with CD8 expression was reduced in the blood of patients with WPP with a favorable outcome of the disease in the preoperative period compared with control values, however, the level of mature and cytokine-producing CD8+NKT cells recovered to the control range by the end of the observed period (on the 21st day after the surgery). At the same time, a decrease in the content of only cytokine-producing CD8+ NKT cells was found in the examined patients with an unfavorable outcome of WPP in the preoperative period relative to the control range and values detected with a favorable outcome, the level of mature CD8+ NKT cells even exceeded that detected in case of a favorable outcome of the disease. However, the number of all studied fractions of CD8+NKT cells was significantly reduced compared to control values by the 21st day of postoperative treatment.

The CD11b receptor is a type I glycoprotein defined as a subunit of the αM integrin and forms the Mac-1 integrin in complex with the CD18 molecule (CD11b/CD18) [25, 26]. Expression of this marker on the membrane of NKT cells increases the level of effector and migratory activity. An increased numbers of CD11b-expressing mature NKT cells were found in patients with a favorable outcome of WPP in the preoperative period, their content returned to normal by day 21 while the content of CD11b+ cytotoxic NKT cells in the pre- and postoperative period remained elevated. In addition, the level of cytokine-producing CD11b+NKT cells in individuals of this group increased towards the end of the observed period. The patients with an unfavorable outcome of the disease had a lower content of mature CD11b+ NKT cells by the 21st day of the postoperative period compared with the control values and the level of these cells with a favorable outcome of peritonitis. In addition, the content of cytotoxic NKT cells significantly increased in patients with an unfavorable outcome only at the end of the observed period while the level of cytokine-producing CD11b+ NKT cells was increased throughout the entire period of the study.

Also, the content of NKT cells expressing activation markers CD28 and CD57 was studied in patients with WPP in depending on the outcome of the disease. The CD28 antigen (Tp44) belongs to the immunoglobulin superfamily and is involved in the enhancement of T-cell receptor signals, which determines its role in the regulation of adaptive immunity [13, 20]. Blockade of CD28 on the membrane of NKT cells completely suppressesed cytokines production [37]. CD57 receptor (Leu-7, HNK-1, NK-1) was defined as an oligosaccharide with sulfated glucuronic acid residues which is expressed on membrane proteins, lipids, and proteoglycans, and its expression level is associated with the accumulation of perforin and granzyme B in the cytolytic killer granules cells [14, 32]. The frequencies of CD28+ and CD57+ NKT cells in the preoperative period was reduced relative to the control values in patients with a favorable outcome of WPP in the preoperative period. However, their number returned to normal immediately after the surgery and remained at the level of the control range until the end of the observed period. At the same time, patients with an unfavorable outcome of peritonitis had low levels of CD28+ and CD57+ NKT cells in the preoperative period which increased to control values only by the end of the observed period.

In general, it can be concluded that the systemic inflammatory response in WPP patients with an unfavorable outcome of the disease in the postoperative period was characterized by a violation of the ratio of the subset composition of NKT cells with a low level of non-classical NKT cells by the end of the observed period (day 21 after surgery) and a pronounced change in the content cells expressing adhesion and activation markers.

 

Conclusion

Thus, significant differences in the phenotype of peripheral blood NKT cells were found between patients with different outcome of WPP. The low relative and absolute levels of NKT cells were observed all patients with WPP regardless of the disease outcome in the preoperative period. At the same time, the absolute level of NKT cells returned to normal values only in patients with a favorable outcome of WPP post 21 days after the surgery. The content of mature NKT lymphocytes was normalized in the peripheral blood of patients with a favorable outcome of WPP by the end of the examination period. The number of cytotoxic cells in the blood of these patients significantly decreased by the 21st day of the examination which is apparently determined by their migration to the area of inflammation. At the same time, patients of this group had the level of cytokine-producing cells at the level of the control range during the entire postoperative period. Conversely, the level of mature and cytokine-producing NKT cells was reduced in the blood of patients with an unfavorable outcome of WPP by the 21st day of the postoperative period. A reduced level of non-classical (expressing the CD8 marker) mature and cytokine-producing NKT cells was detected only in patients with a favorable outcome of WPP in the preoperative period which returned to normal by the end of the postoperative period. At the same time, patients with an unfavorable outcome of the disease had a reduced number of NKT cells of these subsets by the 21st day of postoperative treatment. It can be assumed that a high level of systemic inflammatory response in the postoperative period in patients of this category was associated with a lack of regulatory processes in the immune system including a low level of non-classical NKT cells. In addition, it was found that a high level of NKT cells (compared to control values) expressing the CD11b receptor was observed in patients with WPP during the entire period of the study. However, only patients with a favorable outcome of the disease had a high level of mature and cytotoxic CD11b+ NKT cells already in the preoperative period while an increased content of cytotoxic CD11b+ NKT cells was found in patients with an unfavorable outcome of peritonitis only by the 21st day after surgery. The content of NKT cells expressing activation markers (CD28 and CD57) was reduced in patients in the preoperative period, it returned to normal with a favorable outcome immediately after surgery while patients with an unfavorable outcome had a recovery of these cell fractions towards the end of the postoperative examination. The established features of the phenotype of NKT cells in patients with an unfavorable outcome of WPP characterize disturbances in the ratio of the subset composition and the mechanisms of functioning of this cell fraction which determines the need to develop immunotherapeutic methods aimed at stimulating the immunoregulatory activity of NKT cells.

×

About the authors

Andrei Anatyevich Savchenko

Federal Research Center «Krasnoyarsk Science Center» of the Siberian Branch of the Russian Academy of Sciences, Scientific Research Institute of medical problems of the Northl Siberian Federal University

Email: aasavchenko@yandex.ru

doctor of medicine, professor, head of the cellular-molecular physiology and pathology laboratory; professor of the medical biology department

Russian Federation, 660022, Russia, Krasnoyarsk region, Krasnoyarsk, Partizana Zheleznjaka str., 3G. Scientific Research Institute of medical problems of the North

Alexandr Gennadyevich Borisov

Federal Research Center «Krasnoyarsk Science Center» of the Siberian Branch of the Russian Academy of Sciences, Scientific Research Institute of medical problems of the North

Email: 2410454@mail.ru

leading researcher of the cellular-molecular physiology and pathology laboratory

Russian Federation, 660022, Russia, Krasnoyarsk region, Krasnoyarsk, Partizana Zheleznjaka str., 3G. Scientific Research Institute of medical problems of the North

Igor Vladimirovich Kudryavtsev

Pavlov First St. Petersburg State Medical University, St.Petersburg, Russian Federation; Federal State Budgetary Scientific Institution "Institute of Experimental Medicine" (FSBSI "IEM"), St.Petersburg, Russian Federation

Author for correspondence.
Email: igorek1981@yandex.ru

PhD (Biology), assistant professor; head of laboratory of cellular immunology

Russian Federation, 197376, St. Petersburg, acad. Pavlov str., 12, Scientific Research Institute of Experimental Medicine

Vasilij Dmitrievich Belenjuk

Federal Research Center «Krasnoyarsk Science Center» of the Siberian Branch of the Russian Academy of Sciences, Scientific Research Institute of medical problems of the North

Email: dyh.88@mail.ru

Jr. researcher of the cellular-molecular physiology and pathology laboratory

660022, Russia, Krasnoyarsk region, Krasnoyarsk, Partizana Zheleznjaka str., 3G. Scientific Research Institute of medical problems of the North

References

  1. Акинфиева О.В., Бубнова Л.Н., Бессмельцев С.С. NKT-клетки: характерные свойства и функциональная значимость для регуляции иммунного ответа // Онкогематология, 2010, № 4, С. 39-47. [Akinfieva O.V., Bubnova L.N., Bessmeltsev S.S. NKT cells: characteristic properties and functional significance for the regulation of the immune response. Oncohematology, 2010, no. 4, pp. 39-47. (In Russ.)]. https://cyberleninka.ru/ article/n/nkt-kletki-harakternye-svoystva-i-funktsionalnaya-znachimost-dlya-regulyatsii-immunnogo-otveta
  2. Беленюк В.Д., Савченко А.А., Борисов А.Г., Кудрявцев И.В. Особенности фенотипа В-лимфоцитов крови в зависимости от исхода распространенного гнойного перитонита // Инфекция и иммунитет, 2021, Т. 11, № 3, C. 454-462. [Belenjuk V.D., Savchenko A.A., Borisov A.G., Kudryavtsev I.V. Features of peripheral blood B-cell subset phenotype are associated with clinical outcome of widespread purulent peritonitis. Russian Journal of Infection and Immunity, 2021, Vol. 11, no. 3, pp. 454-462. (In Russ.)]. doi: 10.15789/2220-7619-CBC-1397
  3. Борисов Р.Н., Здзитовецкий Д.Э., Каспаров Э.В., Савченко А.А., Борисов С.А., Бердников Д.С., Говоруха Е.С., Болдырев П.Н. Типы реакции иммунной системы и их характеристика у больных распространенным гнойным перитонитом // Сибирское медицинское обозрение, 2019, № 5, С. 80-87. [Borisov R.N., Zdzitovetskii D.E., Kasparov E.V., Savchenko A.A., Borisov S.A., Berdnikov D.S., Govorukha E.S., Boldyrev P.N. Types of immune system reactions and their characteristic in patients with generalized purulent peritonitis. Siberian Medical Review, 2019, no. 5, pp. 80-87. (In Russ.)]. doi: 10.20333/2500136-2019-5-80-87
  4. Кудрявцев И.В., Субботовская А.И. Опыт измерения параметров иммунного статуса с использованием шестицветного цитофлуориметрического анализа // Медицинская иммунология, 2015, Т. 17, № 1, С. 19-26. [Kudryavtsev I.V., Subbotovskaya A.I. Experience in measuring the parameters of the immune status using six-color cytofluorimetric analysis. Meditsinskaya immunologiya = Medical Immunology (Russia), 2015, Vol. 17, no. 1, pp. 19-26. (In Russ.)]. doi: 10.15789/1563-0625-2015-1-19-26]
  5. Савченко А.А., Борисов А.Г., Черданцев Д.В., Первова О.В., Кудрявцев И.В., Гвоздев И.И., Мошев А.В. Особенности фенотипа и активности NAD(P)-зависимых дегидрогеназ нейтрофилов у больных распространенным гнойным перитонитом в прогнозе развития сепсиса // Инфекция и иммунитета, 2018, Т. 8, № 3, С. 369-376. [Savchenko А.А., Borisov A.G., Cherdancev D.V., Pervova O.V., Kudryavtsev I.V., Gvozdev I.I., Moshev A.V. Features of the phenotype and NAD(P)-dependent dehydrogenases activity in neutrophil by patients with widespread purulent peritonitis in prognosis for sepsis development. Russian Journal of Infection and Immunity, 2018, Vol. 8, no. 3, pp. 369-376. (In Russ.)]. doi: 10.15789/2220-7619-2018-3-369-376
  6. Табаков Д.В., Заботина Т.Н., Борунова А.А., Панчук И.О., Короткова О.В., Кадагидзе З.Г. Гетерогенность популяций NK и NKT-лимфоцитов у здоровых доноров // Медицинская иммунология, 2017, Т. 19б № 4, С. 401-408. [Tabakov D.V., Zabotina T.N., Borunova A.A., Panchuk I.O., Korotkova O.V., Kadagidze Z.G. Heterogeneity of NK and NKT lymphocyte populations in healthy donors. Meditsinskaya immunologiya = Medical Immunology (Russia), 2017, Vol. 19, no. 4, pp. 401-408. (In Russ.)]. https://doi.org/10. 15789/1563-0625-2017-4-401-408
  7. Хайдуков С.В., Байдун Л.А., Зурочка А.В., Тотолян Арег А. Стандартизованная технология «Исследование субпопуляционного состава лимфоцитов периферической крови с применением проточных цитофлюориметрованализаторов» (проект) // Медицинская иммунология, 2012, Т. 14, № 3, С. 255-268. [Khaydukov S.V., Baydun L.A., Zurochka A.V., Totolian Areg A. Standardized technology «Research of lymphocytes subpopulation composition in peripheral blood using flow cytometry analyzers» (Draft). Meditsinskaya immunologiya = Medical Immunology (Russia), 2012, Vol. 14, no. 3, pp. 255-268. (In Russ.)]. doi: 10.15789/1563-0625-2012-3-255-268
  8. Almeida J.S., Couceiro P., López-Sejas N., Alves V., Růžičková L., Tarazona R., Solana R., Freitas-Tavares P., Santos-Rosa M., Rodrigues-Santos P. NKT-Like (CD3+CD56+) Cells in Chronic Myeloid Leukemia Patients Treated With Tyrosine Kinase Inhibitors. Front. Immunol., 2019, Vol. 10, pp. 2493. doi: 10.3389/fimmu. 2019.02493
  9. Bae E.A., Seo H., Kim I.K., Jeon I., Kang C.Y. Roles of NKT cells in cancer immunotherapy. Arch. Pharm. Res., 2019, Vol. 42, no. 7, pp. 543-548. doi: 10.1007/s12272-019-01139-8
  10. Bendelac A., Savage P.B., Teyton L. The biology of NKT cells. Annu. Rev. Immunol., 2007, Vol. 25, pp. 297-336. doi: 10.1146/annurev. immunol.25.022106. 141711
  11. Brailey P.M., Lebrusant-Fernandez M., Barral P. NKT cells and the regulation of intestinal immunity: a two-way street. FEBS J., 2020, Vol. 287, no. 9, pp. 1686-1699. doi: 10.1111/febs.15238
  12. Cairo C., Webb T.J. Effective Barriers: The Role of NKT Cells and Innate Lymphoid Cells in the Gut. J. Immunol., 2022, Vol. 208, no. 2, pp. 235-246. doi: 10.4049/jimmunol. 2100799
  13. Ceeraz S., Thompson C.R., Beatson R., Choy E.H. Harnessing CD8+CD28- Regulatory T Cells as a Tool to Treat Autoimmune Disease. Cells, 2021, Vol. 10, no. 11, pp. 2973. doi: 10.3390/cells10112973
  14. Chen Y., Tian Z. Innate lymphocytes: pathogenesis and therapeutic targets of liver diseases and cancer. Cell. Mol. Immunol., 2021, Vol. 18, no. 1, pp. 57-72. doi: 10.1038/s41423-020-00561-z
  15. Cichoż-Lach H., Grywalska E., Michalak A., Kowalik A., Mielnik M., Roliński J. Deviations in Peripheral Blood Cell Populations are Associated with the Stage of Primary Biliary Cholangitis and Presence of Itching. Arch. Immunol. Ther. Exp. (Warsz), 2018, Vol. 66, no. 6, pp. 443-452. doi: 10.1007/s00005-018-0515-9
  16. de Andrés C., Fernández-Paredes L., Tejera-Alhambra M., Alonso B., Ramos-Medina R., Sánchez-Ramón S. Activation of Blood CD3+CD56+CD8+ T Cells during Pregnancy and Multiple Sclerosis. Front. Immunol., 2017, Vol. 8, pp. 196. doi: 10.3389/fimmu. 2017.00196
  17. Farrington L.A., Callaway P.C., Vance H.M., Baskevitch K., Lutz E., Warrier L., McIntyre T.I., Budker R., Jagannathan P., Nankya F., Musinguzi K., Nalubega M., Sikyomu E., Naluwu K., Arinaitwe E., Dorsey G., Kamya M.R., Feeney M.E. Opsonized antigen activates Vδ2+ T cells via CD16/FCγRIIIa in individuals with chronic malaria exposure. PLoS Pathog., 2020, Vol. 16, no. 10, e1008997. doi: 10.1371/journal.ppat. 1008997
  18. Ferrari L., Martelli P., Saleri R., De Angelis E., Ferrarini G., Cavalli V., Passeri B., Bazzoli G., Ogno G., Magliani W., Borghetti P. An engineered anti-idiotypic antibody-derived killer peptide (KP) early activates swine inflammatory monocytes, CD3+CD16+ natural killer T cells and CD4+CD8α+ double positive CD8β+ cytotoxic T lymphocytes associated with TNF-α and IFN-γ secretion. Comp. Immunol. Microbiol. Infect. Dis., 2020, Vol. 72, pp. 101523. doi: 10.1016/j.cimid. 2020.101523
  19. Gao Y.L., Yao Y., Zhang X., Chen F., Meng X.L., Chen X.S., Wang C.L., Liu Y.C., Tian X., Shou S.T., Chai Y.F. Regulatory T Cells: Angels or Demons in the Pathophysiology of Sepsis? Front. Immunol., 2022, Vol. 13, pp. 829210. doi: 10.3389/fimmu. 2022.829210
  20. González-Osuna L., Sierra-Cristancho A., Cafferata E.A., Melgar-Rodríguez S., Rojas C., Carvajal P., Cortez C., Vernal R. Senescent CD4+CD28- T Lymphocytes as a Potential Driver of Th17/Treg Imbalance and Alveolar Bone Resorption during Periodontitis. Int. J. Mol. Sci., 2022, Vol. 23, no. 5, pp. 2543. doi: 10.3390/ ijms23052543
  21. Goswami M., Sharma D., Khan N.M., Checker R., Sandur S.K., Jawali N. Antioxidant supplementation enhances bacterial peritonitis in mice by inhibiting phagocytosis. J. Med. Microbiol., 2014, Vol. 63, Pt. 3, pp. 355-366. doi: 10.1099/jmm.0.067173-0
  22. Hu J., Yi B., Zhang H. Influence of climatic factors on single-center peritoneal dialysis-associated peritonitis. Zhong Nan Da Xue Xue Bao Yi Xue Ban, 2022, Vol. 47, no. 5, pp. 639-649. doi: 10.11817/j.issn.1672-7347.2022.210506
  23. Huang H., Liu Y., Ouyang X., Wang H., Zhang Y. Identification of a peptide targeting CD56. Immunobiology, 2020, Vol. 225, no. 4, pp. 151982. doi: 10.1016/j.imbio. 2020.151982
  24. Ibidapo-Obe O., Stengel S., Köse-Vogel N., Quickert S., Reuken P.A., Busch M., Bauer M., Stallmach A., Bruns T. Mucosal-Associated Invariant T Cells Redistribute to the Peritoneal Cavity During Spontaneous Bacterial Peritonitis and Contribute to Peritoneal Inflammation. Cell. Mol. Gastroenterol. Hepatol., 2020, Vol. 9, no. 4, pp. 661-677. doi: 10.1016/j.jcmgh. 2020.01.003
  25. Kabanov D.S., Grachev S.V., Prokhorenko I.R. Monoclonal Antibody to CD14, TLR4, or CD11b: Impact of Epitope and Isotype Specificity on ROS Generation by Human Granulocytes and Monocytes. Oxid. Med. Cell. Longev., 2020, Vol. 2020, pp. 5708692. doi: 10.1155/ 2020/5708692
  26. Khan S.Q., Khan I., Gupta V. CD11b Activity Modulates Pathogenesis of Lupus Nephritis. Front. Med. (Lausanne), 2018, Vol. 5, pp. 52. doi: 10.3389/fmed. 2018.00052
  27. Kumar V.V., Verma A., Thakur D.S., Somashekar U., Kothari R., Sharma D. Prophylactic mesh placement in emergency midline laparotomy for intestinal perforation peritonitis: An appeal for caution. Trop. Doct., 2022, Vol. 23, e494755221110831. doi: 10.1177/ 00494755221110831
  28. Lotte R., Courdurié A., Gaudart A., Emery A., Chevalier A., Tran A., Payen M., Ruimy R. Spontaneous Bacterial Peritonitis: The Incremental Value of a Fast and Direct Bacterial Identification from Ascitic Fluids Inoculated in Blood Culture Bottles by MALDI-TOF MS for a Better Management of Patients. Microorganisms, 2022, Vol. 10, no. 6, pp. 1188. doi: 10.3390/ microorganisms 10061188
  29. Lu Y., Li Y., Zhou W., Ding B., Yu Q. Regulatory T cells regulate the distribution of natural killer T cells through CD39 signal transduction in asthma. Hum. Cell., 2019, Vol. 32, no. 2, pp. 141-149. doi: 10.1007/s13577-018-00226-0
  30. Ngiow S.F., Young A. Re-education of the Tumor Microenvironment With Targeted Therapies and Immunotherapies. Front. Immunol., 2020, Vol. 11, pp. 1633. doi: 10.3389/fimmu. 2020.01633
  31. Nilsson J., Hörnberg M., Schmidt-Christensen A., Linde K., Nilsson M., Carlus M., Erttmann S.F., Mayans S., Holmberg D. NKT cells promote both type 1 and type 2 inflammatory responses in a mouse model of liver fibrosis. Sci. Rep., 2020, Vol. 10, no. 1, pp. 21778. doi: 10.1038/s41598-020-78688-2
  32. Noma H., Eshima K., Satoh M., Iwabuchi K. Differential dependence on nuclear factor-κB-inducing kinase among natural killer T-cell subsets in their development. Immunology, 2015, Vol. 146, no. 1, pp. 89-99. doi: 10.1111/imm.12484
  33. Pinson J., Tuech J.J., Ouaissi M., Mathonnet M., Mauvais F., Houivet E., Lacroix E., Rondeaux J., Sabbagh C., Bridoux V. Role of protective stoma after primary anastomosis for generalized peritonitis due to perforated diverticulitis-DIVERTI 2 (a prospective multicenter randomized trial): rationale and design (nct04604730). BMC Surg., 2022, Vol. 22, no. 1, pp. 191. doi: 10.1186/s12893-022-01589-w
  34. Rajabaleyan P., Michelsen J., Tange Holst U., Möller S., Toft P., Luxhøi J., Buyukuslu M., Bohm A.M., Borly L., Sandblom G., Kobborg M., Aagaard Poulsen K., Schou Løve U., Ovesen S., Grant Sølling C., Mørch Søndergaard B., Lund Lomholt M., Ritz Møller D., Qvist N., Bremholm Ellebæk M.; VACOR study group. Vacuum-assisted closure versus on-demand relaparotomy in patients with secondary peritonitis-the VACOR trial: protocol for a randomised controlled trial. World J. Emerg. Surg., 2022, Vol. 17, no. 1, pp. 25. doi: 10.1186/s13017-022-00427-x
  35. Senpuku H., Miyazaki H., Yoshihara A., Yoneda S., Narisawa N., Kawarai T., Nakagawa N., Miyachi M., Tada A., Yoshida G., Shimada M., Ohashi M., Nishimuta M., Kimura Y., Yoshitake Y. CD56(dim)CD16(high) and CD56(bright)CD16(-) cell percentages associated with maximum knee extensor strength and incidence of death in elderly. Springerplus, 2016, Vol. 5, pp. 244. doi: 10.1186/s40064-016-1884-3
  36. Shen H., Gu C., Liang T., Liu H., Guo F., Liu X. Unveiling the heterogeneity of NKT cells in the liver through single cell RNA sequencing. Sci. Rep., 2020, Vol. 10, no. 1, pp. 19453. doi: 10.1038/s41598-020-76659-1
  37. Shissler S.C., Singh N.J., Webb T.J. Thymic resident NKT cell subsets show differential requirements for CD28 co-stimulation during antigenic activation. Sci. Rep., 2020, Vol. 10, no. 1, pp. 8218. doi: 10.1038/s41598-020-65129-3
  38. Stengel S., Quickert S., Lutz P., Ibidapo-Obe O., Steube A., Köse-Vogel N., Yarbakht M., Reuken P.A., Busch M., Brandt A., Bergheim I., Deshmukh S.D., Stallmach A., Bruns T. Peritoneal Level of CD206 Associates With Mortality and an Inflammatory Macrophage Phenotype in Patients With Decompensated Cirrhosis and Spontaneous Bacterial Peritonitis. Gastroenterology, 2020, Vol. 158, no. 6, pp. 1745-1761. doi: 10.1053/j.gastro. 2020.01.029
  39. Terabe M., Berzofsky J.A. Tissue-Specific Roles of NKT Cells in Tumor Immunity. Front. Immunol., 2018, Vol. 9, pp. 1838. doi: 10.3389/fimmu. 2018.01838
  40. Vogt S., Mattner J. NKT Cells Contribute to the Control of Microbial Infections. Front. Cell. Infect. Microbiol., 2021, Vol. 11, pp. 718350. doi: 10.3389/fcimb. 2021.718350
  41. Yang L., Liu S., Zhang Q., Jia S., Qiu C., Jin Z. Overexpression of ascitic interleukin-35 induces CD8+ T cell exhaustion in liver cirrhotic patients with spontaneous bacterial peritonitis. Int. Immunopharmacol., 2022, Vol. 108, pp. 108729. doi: 10.1016/j.intimp. 2022.108729

Supplementary files

There are no supplementary files to display.


Copyright (c) Savchenko A.A., Borisov A.G., Kudryavtsev I.V., Belenjuk V.D.

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.

СМИ зарегистрировано Федеральной службой по надзору в сфере связи, информационных технологий и массовых коммуникаций (Роскомнадзор).
Регистрационный номер и дата принятия решения о регистрации СМИ: серия ПИ № ФС 77 - 64788 от 02.02.2016.


This website uses cookies

You consent to our cookies if you continue to use our website.

About Cookies