Savolyuk, Gudz, and Zhmur: Changes of the Immune Status in Patients with Purulent Peritonitis



Problem statement and analysis of the recent research

The development of purulent peritonitis (PP) is characterized by three stages - reactive, toxic and terminal which have significant differences concerning the clinical picture and perioperative treatment strategy [1]. Cellular and humoral immunity in patients with purulent peritonitis is closely associated with pathogenetic mechanisms of its development. Underestimation of immune disorders in the preoperative stage and after the surgery leads to progression of peritonitis, development of septic complications, occurrence of abdominal sepsis [2, 3]. The difficulty in treating these terrible processes largely depends on the understanding and knowledge of processes in the patient’s immune system. However, immunity changes depending on the PP stages are understudied.

The objective of the research was to examine the changes of cellular and humoral immunity depending on the stage of purulent peritonitis.

Materials and methods

The work was based on study of 126 patients with acute surgical diseases complicated by the development of PP, who were hospitalized in the clinic of the Department of Surgery No 2 at National Pirogov Memorial Medical University, at the City Clinical Emergency Hospital, Vinnytsya during 2012 - 2016.

Most PP cases occurred as complications of acute destructive appendicitis in 41 patients (32.5%), perforated gastric ulcer and duodenal ulcer in 30 cases (23.8%), acute intestinal obstruction in 12 patients (9.5%), acute destructive cholecystitis in 10 cases (7.9%), acute infected necrotizing pancreatitis in 8 patients (6.4%), mesenteric ischemia in 8 cases (12.3%), perforation of the colon and small intestine in 4 patients (3.2%), gynecological diseases in 7 cases (5.6%), traumatic injuries of hollow organs in 6 patients (4.8%). Analysis of patients was conducted taking into account the severity of the disease according to the classification adopted in protocols to provide surgical care to patients with peritonitis (MOH No297 from 02.04.2010.). All patients were divided into the PP stages according to clinical signs, namely reactive (49 patients), toxic (41 patient) and terminal (36 patients). A group of 10 volunteer donors was created to determine the normal indicators.

Immunological reactivity was assessed according to the PP stage in terms of humoral and cellular immunity. Analysis of cellular component of immune system was based on the number of lymphocytes (32.8±0.8%), with their population and subpopulation identification of surface structures of the general population of T-lymphocytes (CD3) (57.3±1.4%) and B-lymphocytes (CD19) (20.6 ± 0.8%), subpopulation of T-cell level (T-helpers (CD4) (36.7±0.92%), T-suppressors (CD8) (22.6±0.74%) of the immunoregulatory index (IRI = CD4 / CD8) 1.6 ± 0.1), natural killer (NK-cells) (CD16) (15.2±0,6%), activated lymphocytes with receptor for IL-2 (CD25) (35.2±0.8%) and dynamics mediated apoptosis of T-lymphocytes (CD95) (7.1±0.03%) using method of direct immunofluorescence by monoclonal antibodies of “Ortho Diagnostic System” company [4]. Determination of humoral component of immune system was performed by quantitative determination of serum concentrations of immunoglobulin (Ig) class G, M and A by ELISA using a test system of “Vector – Best”. The concentration of immunoglobulin (Ig) class G (9.2±0.24 g/l), M (1.6±0.15 g/l) and A (2.4±0.14 g/l) was performed by radial immunodiffusion by Mancini G. (1965).

Statistical calculations were performed using the program “STATISTIKA 6.0”, which includes tools for their analysis. Having proved the normality of data distribution and equality of general variances in samples of control and main groups that were compared, the following methods of statistical analysis were used: Student t-test (two-sample t-test with the same variances), paired Student t-test (paired two-sample t-test for medium). The data format in the Table 1 was M±m, where M meant arithmetic mean, m meant standard error of the arithmetic mean.

Results and Discussion

The immune system state is an important criterion in determining the progression of inflammation in the abdomen. PP course to a large extent depends on an adequate immune response. Correct immune response and sufficient functional reserves contribute to inflammation compensation. Immune deficiency as distress syndrome causes an unfavorable course, the progression of secondary acquired immune deficiency leading to local purulent complications, sepsis.

Analyzing the dynamics of T lymphocyte in blood, a tendency to progressive decrease in CD3 in reference to donors data (57.3±1.4%) was detected, patients data with reactive stage constituted (49.6±0.7%, p<0.01) with further reduction of concentrations in patients with toxic (44.2±0.44%, p<0.01) and terminal (33.6±0.52%, p<0.01) PP stages.

Assessing the dynamics of CD8, their number in relation to donor (22.6±0.74%) and patients with reactive stage data was found not to vary (p>0.05) Only a tendency to increase in the comparative analysis of patients with toxic (23.5±0.25, p<0.01) and terminal stage PP (23.9±0.49, p<0.01) was noted. Moreover, significant index difference between toxic and terminal PP was not observed.

Table 1

Dynamics of the indicators of the NIHSS scale when treating patients with ischemic stroke, points (M±m)

Indicator Reactive (n=49) Toxic (n=41) Terminal (n=36) Donors (n=10)
CD3, % 49.6±0.7*** 44.2±0.44*** 33.6±0.52*** 57.3±1.4
CD4, % 29.3±0.62*** 26.7±0.4*** 23.5±0.48*** 36.7±0.92
CD8, % 22.6±0.52* 23.5±0.25* 23.9±0.41* 22.6±0.74
CD4/CD8 1.29±0.024*** 1.13±0.018** 0.98±0.018*** 1.51±0.08
СD19, % 22.6±0.52 27.7±0.3*** 25.2±0.25*** 20.6±0.8
CD16, % 20.1±0.4%*** 21.6±0.2** 13.9±0.45** 15.6±0.6
CD25, % 42.3±0.55**** 36.8±0.48** 30.8±0.42** 35.2±0.8
CD95, % 7.3±0.02*** 7.9±0.02*** 8.4±0.03*** 4.7±0.51
IgG, g/l 17.8±0.24**** 20.1±0.30*** 14.1±0.21**** 9.2±0.24
IgM, g/l 2.40±0.22*** 3.62±0.12**** 1.88±0.15* 1.6±0.15
IgA, g/l 1.88±0.10** 1.42±0.08*** 0.90±0.1**** 2.3±0.12

Note.

* - p>0.05

** - p<0.05

*** - p<0.01

**** - p<0.001

Thus, the variety of trends of indicators of CD4 and CD8 was observed. If the number of CD4 progressively decreased, the CD8 in relative values was stable with a tendency to increase in patients with toxic and terminal PP, as reflected in the assessment of the IRI. IRI differed in the patients with reactive stage from the group of donors (1.51±0.08) and constituted 1.29±0.024 (p<0.01), patients with toxic stage had a difference of values with reactive stage and reached a minimum in the terminal stage (0.98±0.018, p˂0.05). IRI significant difference was stated in all groups being significant for clinical characteristics of T-cell immunity suppression. The opposite dynamics of progressive increase was observed after the comparison of B-cells number (CD19) with the donors indeices (20.6±0.8%) and patients with reactive and toxic stage where the index increase to maximum of 27.7±0.3% (p<0.01) and a gradual decrease in the terminal stage of 25.2±0.25% (p<0.001) was observed, which could indicate depletion of humoral immunity on the background of inflammation progression.

Assessing changes in NK-cells (CD16) presented in groups, their increase during the reactive phase (20.1±0.4%) with indices difference in comparison with donors (15.6±0.6%, p<0.01) and indices of reactive and toxic stages was observed. Their sharp decrease (13.9±0.5%, p<0,05) at PP terminal stage was stated in comparison with donors and with those of patients with reactive and toxic PP stages that characterized the acute degree of inhibition of cellular immunity. Analysis of activated lymphocytes dynamics with receptor for interleukin-2 (CD25) detected that their number was 42.3±0.55% at the reactive stage which exceeded donors indices (p<0.001) and gradually decreased at the toxic stage (36.8±0.48%, p<0.01) reaching a minimum of 30.8±0.42% in the terminal PP compared with the donors index (p<0.05), and with the difference between each other (p<0.001) being different from the dynamics of the increase in IL-2 in PP toxic stage. However, beginning with the terminal stage, a similar decrease in their number was observed, and it coincided with the dynamics of IL-2 reduction in patients with end-stage peritonitis. Changes of the T-lymphocyte apoptosis receptor (CD95) detected that its value was increasing progressively beginning with the reactive PP stage (7.3±0.02) when the difference of values was defined compared with donors (4.7±0.51%, p<0.01), reaching its maximum at terminal PP (8.4±0.03%) compared with PP toxic stage (7.9±0.02%, p<0.01), which directly correlated with the severity of PP.

Evaluation of humoral immunity detected patterns of its changes in patients with PP. The concentration of IgA decreased (1.88±0.10 g/l, p<0.05) starting from reactive stage compared with donors (2.3±0.12) reaching a maximum decrease in patients with terminal stage (0.9±0.1 g/l, p<0.001) characterizing the extreme degree of suppression of humoral factors of regional defence. The level of IgM tended to increase gradually in patients at the reactive stage (2.4±0.22 g/l, p<0.01) and reached its maximum at the toxic stage (3.62±0.12 g/l) comparing with a group of donors (1.6±0.15 g/l, p<0.01). Decrease in the concentration of highmolecular IgM (1.88±0.15 g/l) was observed in patients with terminal PP comparing with the toxic stage reaching donors’ indices (1.7±0.12 g/l, p>0.05) and indicating the severe PP and functional failure of the body to the synthesis of macromolecular defense factors in terminal PP.

A similar dynamic was noted during IgG levels analysis. Progressive increase in IgG at the reactive stages (17.8±0.24 g/l, p<0.001 at donors) was observed during the observation. This trend remained in the toxic stage as well, when maximum concentration was noted (20.1±0.30 g/l, p<0.001). However, the rapid decrease in IgG (14.1±0.21 g/l, p<0.001) in case of the terminal PP was marked comparing with the previous indices of the group, although this value was more than in donors (9.2±0.24 g/l, p<0.001) confirming the severe inhibition of synthetic function, stating functional failure of humoral immunity.

Thus, laboratory monitoring of immunological reactivity in patients with PP stated a number of regular and conditioned changes in cellular and humoral immunity. The progressive growth of PP severity caused by lifetime not eliminated source of infection led to increase in white blood cell count, CD95 indices. This was caused by a dose-dependent influence of increasing concentrations of IL-6, B-lymphocytes (CD19), production of Ig class M and G and the number of NK-cells (CD16 and CD25), the dynamics of which was closely related and directly dependent on fluctuations of IL-2 content where sharp decrease in CD25, CD16 and CD19, dysfunction of acute inhibition of the synthesis of antibodies, namely IgM and IgG concentrations decrease, and in the case of macromolecular IgM to a level below the group of donors was observed. This was accompanied by a progressive lymphocytopenia, decrease in CD3, CD4, leading to decrease in IRI on the background of constant concentration of CD8.

Conclusions

  1. The development and course of purulent peritonitis is accompanied by changes in the reactivity of the immune system towards secondary immunodeficiency.

  2. The depletion of immune system reserves at toxic and terminal stages of purulent peritonitis requires search for adequate methods of correction of patient’s cellular and humoral immunity.

Prospects for further research

Prospects for further research involve research on local directional correction of the immune system reactivity under the conditions of surgical treatment of purulent peritonitis depending on the stage of the disease.

References

1 

VV Boyko, VK Logachov, ME Timchenko. The problem of prediction of complications in the treatment of widespread peritonitis. Suchasni medychni tekhnolohiyi. 2011;3-4:44-47.

2 

VV Bilookiy, YE Rogoviy. Indicators immunological blood tests, depending on the severity of bile peritonitis. Klinichna ta eksperymental'na patolohiya. 2011;1 (35):6-8.

3 

NG Hagiev. Imunne status cytokine profile and endogenic intoxication in patients with general peritonitis. Khirurhiya Ukrayiny. 2012;4:67-68.

4 

DC Nascimento, JC Alves-Filho, F Sonego, SY Fukada, MS Pereira, et al. Role of regulatory T-cells in long-term immune dysfunction associated with severe sepsis. Crit Care Med. 2011;42 (8):1718-1725. doi:10.1097/CCM.0b013e3181e78ad0.



Copyright (c) 2017 Serhiy I Savolyuk, Maksim A Gudz, Andriy A Zhmur

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


IFNMU Logo

Free counters!