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Rakhimov Abdulla Khaitovich, researcher of the Department of Immunoprophylaxis, Republican Specialized Research-Practical Medical Center of Pediatrics, Republic of Uzbekistan.
E-mail: rabdulla@mail.ru Khalilova Gulnara Mukhamedovna, Head of the Department of Immunoprophylaxis, Doctor of med. sciences, Republican Specialized Research-Practical Medical Center of Pediatrics, Republic of Uzbekistan
IMMUNOLOGICAL PARAMETERS IN CHILDREN WITH SECONDARY IMMUNODEFICIENCY
Abstract: This article deals with immunological investigations carried out in 70 children with clinical signs of secondary immunodeficiency. The parameters of cellular and humoral immunity in children with HIV are characterized by discoordinated disorders, which are related to immune deficiency state. The results obtained show necessity of the following study on serocontrol of the antibody titres and cells of immunologic memory for the further development of individual scheme of buster performance of vaccination for children with HIV.
Keywords: children, secondary immunodeficiency, immunity, vaccination, immunological parameters.
Immunodeficiency or syndrome of immunological insufficiency is a disorder of normal immune status, related to defect of one or several mechanisms of immune responses. It should be noted that secondary immunodeficiency is a pathological state developing on the background of normally functioning system with persistent marked lowering ofquantitative and functional characteristics, relating to various chains of immune system [2, 4].
Three forms of secondary immunodeficiency (SID) are classified as [5]:
- Induced SID-states, when the concrete reason is present, inducing their appearance (roentgen radiation, im-munodepressants, traumas, hormone therapy, tumors etc.)
- Acquired SID-HIV-infected (AIDS)
- Spontaneous SID-states, characterized by absence of clear reason, inducing disturbance of immunological reactivity.
- The secondary immunodeficiency states are characterized by reversible dysfunctions of immune system, changes of the processes of differentiation, proliferation and its cell adaptation, resulting to attenuation of immune response, expressing by impairment of humoral
and cellular immunity, synthesis of the complement components, absence or reducing of the activity of the cytotoxic lymphocytes and macrophages [1, 3, 6, 7].
Therefore the study of immunological parameters of immune system in children of this category appeared to be topical for determination of the category of deviations of immunologic parameters.
Materials and methods
The immunological investigations were performed in 70 children with clinical signs of secondary immunodeficiency. All children were of the age from 6 months to 5 years. Of them 34 (48.5%) children were with induced form of SID and 36 (51.5%) with spontaneous form of SID. Control group included 30 practically health children. In connection with that secondary immunodeficiencies are not independent diseases, and the whole complex of the features indicates about impairment of functioning of the immune system and diversity of the existing classifications of SID, we used screening clinical-anamnestic criteria for clinical diagnosis of immunodefi-cient diseases in the target selection of the patients (Ka-zmirchuk V. E., Drannic G. N., 2003, National Medical
University after O. O. Bogomolets, The Ukraine). These criteria were based on the association of various signs in 5 groups: 1. Clinical symptoms, including 32 signs. 2. Anamnesis of the life and disease, comprising of 19 signs. 3. Antenatal immunological anamnesis, having 12 signs. 4. Family anamnesis (congenital and hereditary diseases) including 12 signs. 5. Laboratory data of 9 findings. Totally 74 signs.
Diagnosis of SID was determined at presence more than 3 clinical criteria of group 1, absence of data of group 4 (family anamnesis(, presence of more than one sign of group 3 (antenatal anamnesis) and presence of 3 and more criteria of group 2 (anamnesis of disease).
In order to study immune status in children with SID the following immunological investigations were performed: Lymphocytes were isolated by the common method in the gradient ofdensity of ficoll-verografin. Phe-notyping of the lymphocytes was carried out with use of monoclonal antibodies of series LT ("Sorbent", Moscow, Russia) and there were revealed numbers of CD4+, CD8+, +CD16+. The contents of immunoglobulins was measured with method of radial immunodiffusion by Man-chini with use of monospecific serums against IgA, IgM, IgG of Russian production of Moscow Institute of Microbiology and Epidemiology after N. F. Gamaleya. Phagocytic activity of neutrophils (FAN) was determined in the test with latex (LLC"Biopreparations" MH RF). 2. Determination of the levels of interleukins: IL-1^, 1L-8, IL-4, IL-10 was performed in the blood serum with method of
immune enzymatic analysis-IFA with use of test-systems LLC"Cytokin" (Saint-Petersburg, Russia).
Analytical researches. The data of complex investigation of the patients were processed with use of appropriate computed program. The analysis of the obtained data was carried out with application of the modern statistic mathematic methods. There were used universal methods of statistical process of the data, evaluation of the statistic reliability with use of criterion Student t and statistic methods of comparison of non parametric data.
Results. In order to study character of immunological disorders in children with HIV infection there was carried out study of cellular, humoral parts and intrasystemic connections of immune system in 70% of children with seronegative and low titres of the seroconversion response. This was made because identification of the changes in these parameters reflects functional peculiarities of the immune system disturbances in children with HIV and in the further they will promote to give more precise definition of the pathogenic regularities of the immune system functioning. As it is known many secondary disorders of immune system are reversible (beside HIV-infection) therefore comparison of immunological disorders between these two forms of HIV will allow choice of the tactic of immunization of these children. This, in its turn, will help to form adequate immune response and reduction of the postvaccinal responses and complications.
Table 1.- Immunological parameters in children with spontaneous and induced forms of the secondary immunodeficiency
Parameters Groups of studied children P
Children with HIV spontaneous form (SP) (n = 36) Children with HIV induced form (IF) (n - 34) Control group (relatively healthy children) (n = 3o)
1 2 3 4 5
Leucocytes 4500±157 4400 ± 229 7925 ±148 > 0.05* < 0.001** < 0.001***
Lymphocytes.% 36 ± 0.7 33 ± 0.8 43 ± 0.7 < 0.01* < 0.001** < 0.001***
Lymphocytes. abs. 2700 ± 95 2500±106 3600 ±108 > 0.05* < 0.001** < 0.001***
1 2 3 4 5
T-lymphocytes.% 56 ± 0.5 55 ± 0.6 65 ± 0.4 > 0.05* < 0.001** < 0.001***
T-lymphocytes. abs. 1750 ± 49 1700±53 1950±65 > 0.05* < 0.05** < 0.05***
B-lymphocytes.% 28 ± 0.07 32 ± 0.16 24 ± 0.4 < 0.001* < 0.001** < 0.001***
B-lymphocytes. abs. 1200 ± 4.0 1250 ± 2.0 960 ± 12 < 0.001* < 0.001** < 0.001***
CD4.% 30 ± 0.8 29 ± 0.45 40 ± 0.08 > 0.05* < 0.001** < 0.001***
CD4. abs. 1200±24 1100±29 1450± 14 < 0.05* < 0.001** < 0.001***
CD8.% 28 ± 0.15 31 ± 0.04 25 ± 0.45 < 0.001* < 0.001** < 0.001***
CD8. abs. 1200±12 1250±10 890 ± 18 < 0.01* < 0.001** < 0.001***
CD4+/ CD8+ 1.07 ± 0.04 0.93 ± 0.05 1.60 ± 0.02 > 0.05* < 0.001** < 0.001***
CD16.% 19 ± 0.15 22 ± 0.30 13 ± 0.07 < 0.01* < 0.001** < 0.001***
CD16. abs. 600 ± 4.0 650 ± 2.0 410 ± 11 < 0.001* < 0.001** < 0.001***
PAN.% 58 ± 0.07 45 ± 0.6 57 ± 0.11 < 0.001* > 0.05** < 0.001***
IgA. mg% 150 ± 2.0 160 ± 1.6 116 ± 2.9 < 0.001* < 0.001** < 0.001***
IgG. mg% 1200 ± 8.0 1250 ± 6.0 1100 ± 11.0 > 0.05* < 0.001** < 0.001***
< 0.05*
IgM. mg% 150 ± 19.5 110 ± 4.0 115 ± 2.0 < 0.05**
> 0.05***
Note: * The value is reliable between data of spontaneous and induced form. ** - between spontaneous form and control
group, *** - between induced form and control group
Immunological investigations in children were performed after complete course of vaccination with use of The National Calendar of Vaccinations of the Republic of Uzbekistan. The investigations of the cellular immunity gave the results presented on the (table № 1). It was revealed that the total number of leucocytes had no reliable differences between groups with HIV, but in comparison with healthy children these parameters had reliable differences (P < 0.001). The number of lymphocytes expressed in percentage ratio had reliable differences between two groups with HIV (P < 0.01) and healthy children (P < 0.001).
The quantity of T-lymphocytes between groups with HIV had no differences (P<0,05), reliability was noted only in comparison with group of healthy children (P<0,001). The reliable increase both in percent and absolute quantity among B-lymphocytes (P<0,001) was noted in groups of children with IF in comparison with HIV SF and group of control, that shows activation of B-cellular part of immunity with low level of T-cellular immunity. This revealed more deep disorders in functioning of the immune system at the early stage of the body defense. It should be also noted that B-lymphocytes were reliably higher in group of SF in comparison with group of control (P < 0.001), however with preserving functioning phagocytic activity of neutrophiles (PAN). The ratio of CD4 cells between groups HIV were reliably increased (P < 0.05) in relation to absolute number in favour of group SF HIV, that is, immune reaction is characterized by helper type of immune disorders.
Analyzing the results obtained on study of the humoral immunity it was revealed that the content of IgA was reliably increased (P < 0.001) in the both groups with HIV in comparison with group of control (P < 0.001). There was also noted reliable rising (P < 0.001) of this immunoglobulin in group IF in comparison with group of SF. Similar reliable increase (P < 0.001) was noted during evaluation of the content of IgG in the both groups with HIV in comparison with group of control. This immunoglobulin had no reliable differences between
groups with HIV (P < 0.05). IgM was reliably increased only in group of SF in comparison with IF and group of control (P < 0.05). It should be noted that the content of this immunoglobulin in group with induced form was relatively reduced (P > 0.05) in comparison with group of control. but reliably decreased in relation to group of SF (P < 0.05). This fluctuation of immunoglobulin M. in our opinion. was connected with coincidence of the conversion of the II-critical period of the making of immune system into III critical period and development of the child with period of vaccination.
In the second period the IgM-mediated immune response develops to the majority of antigens without formation of immunological memory and the defects of immune system appeared in this period. In the third critical period of the children development the primary character of immune response has been still preserved to the majority of pathogens. However the ability appeared for transfer of the immunoglobulin synthesis to the class of IgG consequently reducing production of Ig M. The change of the suppressive type of immune response to helper and formation of adequate humoral immune response in norm has been made.
Thus, the analysis of the state of cellular and humoral immunity in children with HIV shows that in children with induced form of HIV-infection there are found deeper disorders both in the cellular and humoral part of immune system characterized by more severe disturbances in the functioning of immune system. We consider that these functional shifts are connected with etiopathogenic character of various forms of secondary immunodeficiency.
The following stage of researches included study of cytokine profile in 50 children with secondary immunodeficiency at the age from 6 months to 5 years (Table 2). There were studied parameters of the intra-systemic regulation of the immunity. There were studied proinflammatory cytokines IL1^, IL-8 and proinflammatory cytokines IL-4, IL-10. Among the children with spontaneous form HIV the proinflammatory cytokines
accounted for IL10 35.5 ± 2.2 pg/ml and IL-8 22.5 ± ± 2.0 pg/ml. and in children with induced form of HIV IL1p 36.5 ± 2.2 pg/ml and IL-8 23.0 ± 2.0 pg/ml. During evaluating proinflammatory cytokines in children
Table 2.- The contents of cytokines in the blood
with spontaneous HIV form IL-4 was 3.0 ± 0.5 pg/ml and IL-10 was 11.0 ± 1.5 pg/ml. and in children with induced form of HIV IL-4 was 2.8 0.5 pg/ml and IL-10 was 10.5 1.5 pg/ml.
serum of studied children with HIV (pg/ml), (M ± m)
Immunologic parameters Spontaneous form Induced form
IL-13 35. 5 ± 2. 2 36. 5 ± 2.2
IL-8 22. 5 ± 2. 0 23. 0 ± 2. 0
IL-4 3. 0 ± 0. 5 2. 8 ± 0. 5
IL-10 11. 0 ± 1. 5 10. 5 ± 1. 5
Discussion
Analyzing the results of cellular and humoral parts of immunity in children with HIV it may be concluded that in children with induced form of HIV there are found deeper disorders both in cellular and humoral parts of immune system, characterized by more serious disturbances of the immune system functioning. In children with induced form of HIV the levels ofproinflammatory cytokines are higher, than in spontaneous form, but the reliable differences were not found (P > 0.05). The values of the levels of proinflammatory cytokines had practically no differences between the both groups. The obtained data of the cytokine status correlate with duration of the inflammatory processes in the body of the children with HIV-infection that enhances picture of immune status. In our opinion, these functional changes are connected with etiopathogenic character of different forms of secondary immunodeficiency. Consequently, this situation dictates individual approach to immunocorrection, rehabilitation of immunodeficiency states in children for the following performance of effective vaccination and requires the further scientific researches.
Conclusions:
1. The parameters of cellular and humoral immunity in children with HIV characterized by discoordinated disorders which are characteristic for immunodeficiency state.
2. The ratio of CD4 cells between groups with HIV was reliably increased (P < 0.05) by absolute number in favor of group HIV SF, that is, immune reaction is characterized by helper type of immune disorders.
3. The quantity of CD8 cells as in percentage, so as in absolute ratio show reliable differences between groups HIV. This reliable rising of CD8 cells in induced form of HIV indicated about immune disorders by suppressor type.
4. In children with induced form of HIV the levels of proinflammatory cytokines was higher than in spontaneous form, however there were no reliable differences (P > 0.05). The values of the levels of proinflammatory cytokines between the both groups have almost similar parameters.
5. The results of immunological investigations dictate obligatory following study of serocontrol of the titres of antibodies and cells of immunological memory for the further development of the individual scheme ofbooster performance of vaccination in children with HIV.
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