|Year : 2021 | Volume
| Issue : 2 | Page : 122-128
The Impact of IgG administration on the cellular immunity status in the patients with multidrug-resistant tuberculosis/HIV with CD4 + lymphocyte cells below 50 cells/ μl<
Nina Matsegora1, Antonina Kaprosh1, Petro Antonenko2
1 Department of Phthisiopulmonology, Odesa National Medical University, Odessa, Ukraine
2 Department of Pharmacology and Pharmacognosy, Odesa National Medical University, Odessa, Ukraine
|Date of Submission||07-Feb-2021|
|Date of Acceptance||03-Mar-2021|
|Date of Web Publication||14-Jun-2021|
Department of Pharmacology and Pharmacognosy, Odesa National Medical University, Valihovsky Lane, 2, Odesa 65082
Source of Support: None, Conflict of Interest: None
Background: Treatment of the patients with multidrug-resistant tuberculosis (MDR-TB)/HIV coinfection in a state of severely suppressed immune system remains unsatisfactory. Methods: The study involved 52 HIV-positive patients with MDR-TB and CD4+ lymphocyte cells below 50 cells/μCL. Patients in control group (Group 1) and in basic group (Group 2) received standard treatment with second-line antituberculosis agents and antiretroviral agents. In addition, the patients in basic group were treated by immunoglobulin G (IgG) intravenously. Immunological diagnostics with the determination of the level of lymphocytes subgroups (CD3+, CD4+, CD8+, CD4+/CD8+) was carried out using an AQUIOS™ CL flow cytometry device at the beginning and after 3–20 months of treatment. Statistical analysis was performed using the Statistica 10.0 software (Stat. Soft Inc., USA). Results: In the patients of Group 2, the absolute number of CD3+ and CD4+ cells at the end of the 20th month of the treatment normalized in 26.9% (absolute amount) and 42.3% (relative amount) of subjects, while in Group 1, this indicator remained below the normal level (P < 0.05). The addition of IgG into standard therapy caused normalization of CD8+ count in 76.9% of patients, while in the control group-only in 46.2% of patients (P < 0.05). Conclusions: The administration of IgG in combination with standard anti-tuberculosis and antiretroviral therapy (ART) contributes to the normalization of the cellular immunity status in patients with MDR-TB/HIV coinfection and severe immunosuppression and allows you to start ART earlier than in patients with single standard therapy
Keywords: Drug resistance, HIV, immunodeficiency, immunoglobulin, tuberculosis
|How to cite this article:|
Matsegora N, Kaprosh A, Antonenko P. The Impact of IgG administration on the cellular immunity status in the patients with multidrug-resistant tuberculosis/HIV with CD4 + lymphocyte cells below 50 cells/ μl<. Int J Mycobacteriol 2021;10:122-8
|How to cite this URL:|
Matsegora N, Kaprosh A, Antonenko P. The Impact of IgG administration on the cellular immunity status in the patients with multidrug-resistant tuberculosis/HIV with CD4 + lymphocyte cells below 50 cells/ μl<. Int J Mycobacteriol [serial online] 2021 [cited 2021 Jul 25];10:122-8. Available from: https://www.ijmyco.org/text.asp?2021/10/2/122/318376
| Introduction|| |
The clinical course of tuberculosis in HIV-positive people with severe immunodeficiency often takes on an aggressive character with severe immunodeficiency often takes on an aggressive character, makes rapid progress, and is accompanied by a generalized infection, which often leads to fatal outcome., According to the WHO, the average success rate of such patients' treatment is only 55%. A lot of authors report that the localization of tuberculosis (pulmonary, extra-pulmonary) and the recovery rate depend on the degree of immune suppression.,,
In recent years, the positive results of immunoglobulin G (IgG) administration in the severe course of numerous infection diseases have been shown.,,, The authors point out that after the administration of IgG, ready-to-use antibodies enter the patients' circulated blood, which bind and remove antigens of a viral and bacterial origin, thereby reducing the infectious and toxic load on the organ systems of the patients.
Previous researches detected an increase in the clinical effectiveness of the second-line anti-tuberculosis agents and antiretroviral therapy (ART) in case of additional IgG administration in the patients with multidrug-resistant tuberculosis (MDR-TB)/HIV coinfection with CD4+ lymphocyte cells level below 50 cells/μL, which was manifested in an improvement in hematological and biochemical parameters.,, However, the effect of IgG on the indexes of the cellular immunity in the complex treatment of such patients (during 20 months) has not been studied before, which makes the chosen research aim as relevant and appropriate.
The aim of the research includes the study of the influence of IgG administration in combination with standard second-line anti-tuberculosis drugs and ART on the cellular immunity status in the patients with MDR-TB/HIV coinfection at the level of CD4+ lymphocytes below 50 cells/μCL.
Clinical rationale for the study
This research is aimed to justify the effectiveness of IgG administration in MDR-TB/HIV patients with severe immunodeficiency.
| Methods|| |
The study involved 52 MDR-TB/HIV patients both genders with CD4+ lymphocytes below 50 cells/μL; the average female age was 39.20 ± 3.17 years, male age –36.50 ± 2.84 years. All patients were HIV-positive with laboratory-confirmed (cultural method or GeneXpert method) MDR-TB with different types of M. tuberculosis resistance to isoniazid- and rifampicin-resistant obligatory. Patients with MDR-TB/HIV were divided as follows:
- Group 1 (control) – 26 patients, who received standard treatment with second-line antituberculosis agents and ARV
- Group 2 (basic) – 26 patients, who also received the standard treatment by antituberculosis second-line agents with ARV treatment, but with the addition of intravenous IgG. It was prescribed according to the following scheme: On the 1st day before the beginning of anti-tuberculosis treatment (ATT), IgG was administered at the rate of 4 ml/kg intravenously; on the 2nd day, the second-line antituberculosis agents were added, according to M. tuberculosis susceptibility; and finally, after 2 weeks, the ARV treatment was added. The following IgG injections were performed every 4 weeks for 3 months, followed by the 5th and 8th months of the intensive MDR-TB/HIV treatment phase.
Immunological diagnostics with the determination of the level of lymphocytes subgroups (CD3+, CD4+, CD8+, CD4+/CD8+) was carried out in the clinical laboratory of the Odessa Regional AIDS Center, using an AQUIOS™ CL flow cytometry device manufactured by Beckman Coulter (certified according to ISO 15189 standard) at the beginning and after 3–20 months of treatment. This is a direct volumetric method for a single platform. The sample was processed using two multitasking probes: One probe pricked the lid and prepared the sample into a 96-pits microplate, while the other aspirate prepared the sample for analysis. While the first sample was incubated, the system continued to prepare additional samples and add them to the queue. Whole blood (140 μL) was added to every pit; then, specific white blood cell staining was performed by incubating whole blood with a monoclonal antibody reagent. Then the red blood cells were removed by their lysis without washing, and the remaining leukocytes were analyzed by flow cytometry. A 488 nm solid-state diode laser was used to measure the light diffraction, fluorescence, and electron volume, which estimated the relative size of cells. It was used a ready-to-use antibodies mixture (AQUIOS Tetra-1 panel, BeckmanCoulter): CD45-isothiocyanate fluorescein (FITC) (clone B3,821F4A)/CD4-phycoerythrin (PE) (clone SFCI12T4D11)/CD8-phycoerythrin TexasRed®-X (ECD) (clone SFCI21Ch3-Cyrin3 5 (clone UCHT1) for staining of the cells. The strategy of gating includes side and forward scattering, SS/CD45 strobing and electric volume (EV) evaluation with 2 parameters EV/SS that promote purification and better restoration of total lymphocytes.
Statistical analysis was performed using the Statistica 10.0 software (Dell Software, Austin, TX, USA). It was determined whether there is a significant difference concerning frequency of studied criteria between the two studied groups. Quantitative indicators in the text and tables are presented in the form M ± m (M– arithmetic average and m– standard deviation); quality indicator a represented in the form Q ± mq (Q is the frequency of occurrence of the trait and mq is the standard deviation). Statistical significance was assumed at the P < 0.05. Kruskal–Wallis, ANOVA, and Chi-square tests were used in this study.
Ethical issued and date of approval for project
The project was approved by the Ethics Committee of Odessa National Medical University, Ukraine (protocol N84, June 16, 2015). It was conducted according to the Declaration of Helsinki standards. All of the patients gave written informed consent and explicitly provided permission for treatment and blood analyses, as well as for the collection of relevant clinical data.
| Results|| |
As a result of dynamic observation, it was noted that even after the first administration of IgG in the patients of basic group (Group 2), the positive clinical changes like a decreasing of intoxication occurred. For instance, totally 16 patients (61.5%) suffered from fever that was 1.3 times less comparatively to the initial level. On the contrary, in the control group (Group 1), the manifestation of intoxication increased; hyperthermia after the appointment of anti-tuberculosis agents was observed in all patients, which we assumed as a consequence of increased bacillemia.
The analysis of immunogram test parameters included determining the level of lymphocytes subsets with the help of monoclonal antibodies (CD markers) by absolute and percentage values, which reflected the state of cellular immunity. Studies of the immune state of patients with MDR-TB/HIV, in which the level of CD4+ lymphocytes below 50 cells/μL, were carried out in dynamics during 20 months of treatment [Table 1] and [Table 2].
|Table 1: Absolute level of T-lymphocytes subsets in the patients with multidrugresistant tuberculosis-tuberculosis/human immunodeficiency virus with СD4 + lymphocytes count below 50 per μ l and immunoglobulin G (0–20 months) (M±m)|
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|Table 2: Relative level of T-lymphocytes subsets in the patients with multidrugresistant tuberculosis-tuberculosis/human immunodeficiency virus with СD4 + lymphocytes count below 50 per μL and immunoglobulin G (0–20 months) (M±m)|
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Initially, the level of CD3+ lymphocytes in the blood of all patients in both the groups was 2 times lower than normal (normal value is 690–2540 cells/μL) and was 372 ± 68.1 cells/μL and 380 ± 71.2 cells/μL, respectively, in the Group 1 and Group 2 or 32.5 ± 5.1% and 32.0 ± 4.9% correspondently (normal value is 55%–84%) [Table 1] and [Table 2]. Even at the 3rd month of IgG administration, the normal level of CD3+ lymphocytes was observed in 30.8 ± 1.85% of the patients of the Group 2 versus 7.7 ± 1.07% of the patients of the Group 1. Hence, the number of the patients in Group 2 with normal CD3+ count was in 4 times more than in Group 1 (P < 0.01) [Figure 1]. At the 5th month of IgG additional administration, the amount of the patients with normal level of CD3+ lymphocytes occurred in 2.4 times more frequently than in control group (73.1 ± 1.77% vs. 30.8 ± 1.85%, P < 0.05). At the 8 h month of studies, the normal level of CD3+ lymphocytes appeared in 100% of the patients in Group 2, while in the control group, it was in 61.5 ± 1.95% or in 1.6 times less (P < 0.05). The obtained positive results of the treatment with IgG of the patients in Group 2 sustained at the 14th and 20th months of treatment, the number of patients with normal CD3+ count in the Group 2 was (P < 0.05). For example, in Group 1, the normal level of CD3+ lymphocytes was determined in 73.1 ± 1.77% of the individuals at the 14th month, and in 76.9 ± 1.69% of the patients at 20th months or in 1.4 times and in 1.1 times less correspondently comparatively to the Group 2 (P < 0.05) [Figure 1]. At the same time during 20 months of treatment, the average level of CD3+ lymphocytes (absolute and relative) increased in both groups, but especially in Group 2 [Table 1] and [Table 2]; the number of patients with decreased CD3+ count gradually decreased in both groups, mostly in Group 1 [Figure 2].
|Figure 1: Number of patients with normal CD3+ and CD4+ (absolute) count according to the treatment strategy. *: The difference is significant between Group 1 and Group 2 (P < 0.05), **: The difference is significant between Group 1 and Group 2 (P < 0.01), ***: The difference is significant between Group 1 and Group 2 (P < 0.001)|
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|Figure 2: Number of patients with decreased CD3+ and CD4+ (absolute) count according to the treatment strategy. *: The difference is significant between Group 1 and Group 2 (P < 0.05), **: The difference is significant between Group 1 and Group 2 (P < 0.01), ***: The difference is significant between Group 1 and Group 2 (P < 0.001)|
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During 8 months of treatment, the level of T-helpers (T-h or CD4+) remained below normal values in all patients of both groups [Figure 1] and [Figure 2], but at the end of the 8th month, there was a gradual increase of the average CD4+ level to 395.00 ± 8.41 cells/μL in Group 2 and only to 115.00 ± 11.2 cells/μL in Group 1, that was in 3.4 times more (P < 0.001) [Table 1]. In the end of 14th month of combined therapy, the normal level of CD4+ count was revealed in 19.20 ± 1.58% of the patients in Group 2 and in rest in 80.8 ± 1.58% of patients, it reached the subnormal values (403 ± 4.2 cells/μL). In patients of the control group, this indicator at the 14th month of the treatment reached a level of 135.00 ± 14.80 cells/μL, which was significantly lower from the normal value (410–1590 cells/μL) and in 3 times lower than in the Group 2 (P < 0.001) [Table 1]. At the end of the 20th month of treatment, around 26.9 ± 1.77% of people in Group 2 had normal level of CD4+ with average values of 522 ± 11.3 cells/μL and in rest– in 73.1 ± 1.77%, it increased to subnormal levels (407 ± 2.6 cells/μL) [Table 1] and [Figure 1]. However, this indicator remained reduced in the control group even at the 20th month of treatment and reached 254.00 ± 12.30 cells/μL, which is 2 times less than the normal in 1.6 times less than in Group 2 (P < 0.001).
In addition, at the 14th month of treatment, around 19.2% of the patients in Group 2 had normal level of CD4+ (the normal relative level for CD4+ is 31%–60%); at the 20th month of treatment– 42.3%. In the same time in Group 1, there were no cases of normal CD4+ level [Table 2].
The absolute level of T-suppressors (T-s or CD8+) before the beginning of treatment in 100% of patients in both groups exceeded the normal level [Table 1] and [Table 2] and [Figure 3]. At the 5th month of treatment in 4 patients (15.4 ± 1.44%) of the Group 2 the normal level of CD8 + was determined (372–974 cells/μL); however, in patients of the Group 1, the level of this indicator remained beyond the normal level (P < 0.001). At the 8th month of treatment, the normal level of CD8+ level has been determined in 34.6 ± 1.90% of the Group 2 and in 23.1 ± 1.69% of the Group 1 or in 1.5 less (P < 0.05). Further, at the 14th month, CD8+ count reached the normal level in 61.5 ± 1.95% in the Group 2 and in 34.6 ± 1.90% of patients in the Group 1 that was in 1.8 times less (P < 0.05). Finally, at the end of the 20th month, the majority of the Group 2 (76.9 ± 1.69%) and almost half of the Group 1 (46.2 ± 1.99%) showed normal CD8+ level or in 1.7 times less (P < 0.05) [Figure 3].
The relative (percentage) level of CD8+ at the 14th month of treatment reached the normal count (19%–35%) in 7.7 ± 1.07% of patients of Group 2 and at 20th month-in 26.9 ± 1.77% of the same group; however, in the Group 1 only at the 20th month, the normal level of T-s was observed in 7.7 ± 1.07% of the patients that was in 3.5 times less than in Group 1 (P < 0.01) [Figure 3].
The immune regulatory index (CD4+/CD8+) in all studied patients at the beginning of treatment was lower than normal-average 0.02 ± 0.01 while the normal range 1.2–2.1 [Table 1]. The low level of CD4+/CD8+ ratio remained almost without changes for 14 months of treatment in both studied groups. In the Group 2 at the 20th month of the treatment around 26.9 ± 1.77% individuals reached the normal level (1.23 ± 0.03), however, in the rest individuals in 73.1 ± 1.77% the immune regulatory index rose to 0.68 ± 0.11. In 100% of patients in Group 1, the CD4+/CD8+ ratio even at 20 month of treatment remained below 1, with a low mean value 0.25 ± 0.08 (Tab. 2).
In the conducted study in the patients with MDR-TB/HIV the immunograms results matched with the clinical course of the disease, medical history, and the results of hematological and biochemical studies.,, The resorption of pulmonary TB-lesions and closing of TB-cavities in both groups started since 4 months of the treatment, but in Group 2 these changes were more intensive than in Group 1. For example, at the 20th month of treatment, the resorption of TB-lesions in Group 2 developed in 96.2% individuals, while in Group 1 – in 80.8% (P < 0.05).
Due to recommended immune stimulation, the patients of the basic group (2) could start ART from the 2 week of ATT. In the same time, in Group 1 the ART started much later-after 1 month of ATT it has been started in 4 individuals out of 26; after 2 months – in 11 persons, after 3 and 4 months – in 4 and 7 individuals correspondently. It is extremely important because the early initiation of ART reduces all-cause mortality in TB/HIV positive patients.
| Conclusions|| |
The obtained data concerning immunological parameters based the following conclusions:
- In the patients with MDR-TB/HIV co-infection and CD4+ lymphocyte levels below 50 cells/μL and who received IgG in complex antiretroviral and anti-tuberculosis therapy, the absolute number of T-helper cells (CD3+ CD4+) in the peripheral blood at the end of the 20th month of the treatment normalized in 26.9% of subjects; the rest– 73.1% had a subnormal level. However, in the group with standard therapy, this indicator remained below the normal level even at the 20th month of the treatment and reached 254 cells/μL, which is almost 2 times lower than normal. In addition, the relative amount of the T-helpers at the 20th month of treatment turned to normal in 42.3% of patients with the addition of IgG, while in individuals with standard therapy, a normal level of T-helpers was not observed at the 20th month of standard treatment
- At the 20th month of treatment the addition of IgG to the standard antiretroviral and anti-tuberculosis therapies caused normalization of T-suppressors count in 76.9% of patients, while in the control group-only in 46.2% of patients. At the same time, the combination of standard antiretroviral and anti-tuberculosis therapy with IgG accompanied by normal relative T-suppressors level in 26.9%, however, in isolated standard therapy-only in 7.7%
- At the beginning of treatment, the immune regulatory index (CD4+/CD8+) was reduced in all patients with average meaning 0.02 that was 60 times more than normal. After 20 months of combined treatment of standard antiretroviral and anti-tuberculosis therapy with IgG, a normal level of immune regulatory index was observed in 26.9% of the patients, while in rest– in 73.1% of the patients, it increased to 0.68. Simultaneously, in 100% of patients with standard antiretroviral and anti-tuberculosis therapy, the CD4+/CD8 + ratio remained below normal, with an average level 0.25
- The intravenous administration of IgG in combination with standard anti-tuberculosis and ART allows you to start ART earlier than in patients with single standard therapy and also contributes to the normalization of the cellular immunity status in patients with MDR-TB/HIV co-infection and severe immunosuppression.
Clinical implications/future directions
The obtained results proved the effectiveness of intravenous administration of IgG in the complex of MDR-TB/HIV-positive patients with severe immunodeficiency. In future we are planning to study IgG impact in complex therapy on the effectiveness of ATT (closing of TB-cavity, conversion of smear-positive status into smear-negative).
We would like to acknowledge the support of the clinicians, nurses, and laboratory personnel who contributed their efforts and made this study possible.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2]