Brucellosis is considered to be a zoonotic disease with a high disability rate and great harm, which seriously threatens public health.
Host protection against
In our previous studies [
The specific mechanism of the immune function after
T lymphocyte is composed of different cell subsets, in which the number and function of different cell subsets determine the immune status of the host. T lymphocyte subsets form a complex immune network in vivo by secreting a variety of cytokines. The normal immune function of the human body is maintained with a certain balance ratio between different cells. Once this balance is broken, it can lead to immune disorders in the body [
Immune damage plays an important role in the occurrence and development of the disease, especially in the chronic process. T lymphocyte plays a key role in the elimination of pathogens after
The current literatures on the research of the expression of Th1, Th2, Th17, and Treg-related T cell subsets and PD-1 in brucellosis patients at different stages (acute, chronic, and convalescent stages) are rare or unreported. In order to further clarify the role of immune function in the pathogenesis of brucellosis, we explored the characteristics of the immune response of T cell subsets and related cytokines and PD-1 in patients with brucellosis and provided theoretical basis for the further study of the immunological pathogenesis of
125 cases of brucellosis in inpatient and outpatient departments of the First Affiliated Hospital of Xinjiang Medical University from March 2017 to December 2017 were collected. There were 45 patients of Han nationality, 65 patients of Kazakh nationality, 5 patients of Hui nationality, 7 patients of Uygur nationality, and 3 patients of Mongolian nationality. Among them, 47 cases were in the acute stage (age: 18-70 years; average age:
The criteria for diagnosis and clinical staging of brucellosis refer to the guidelines issued by China in 2012 for the diagnosis and treatment of brucellosis: (1) acute phase—the duration of disease was less than 6 months; (2) chronic phase—the disease had not recovered for more than 6 months; and (3) convalescence phase—at present, there is no definition of convalescent period in China and WHO guidelines. In this study, patients who completed the regular course of treatment, whose clinical symptoms disappeared, and whose laboratory inflammation markers returned to normal were regarded as convalescent patients.
Patients who met the diagnostic criteria for brucellosis guidelines issued in 2012 were included in the study.
The exclusion criteria are as follows: (1) patients with a history of serious diseases or dysfunction of other systems (cardiovascular, nervous, respiratory, liver, kidney, etc.); (2) patients with a history of tumor or immune system disorders; (3) patients who used immunosuppressive drugs, corticosteroids, and immunomodulators for a long time or nearly three months; and (4) patients with an immunity disorder.
This study was approved by the Ethics Committee of the First Affiliated Hospital of Xinjiang Medical University. All patients and healthy controls had signed informed consent forms.
Human antibodies anti-human CD4, anti-human CD25, anti-human CD127 (IL-7R
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200
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The expression levels of IL-5, IL-13, IFN-gamma, IL-2, IL-17A, IL-17F, and IL-4 in the peripheral serum of patients with brucellosis and healthy controls were detected using the Human Th Cytokine Panel (13-plex) Kit, according to the manufacturers’ instructions.
R version 3.4.2 was used to analyze the data. If the data obeyed normal distribution and homogeneity test of variance, a
Of the 125 brucellosis patients, 110 (88%) had a clear contact history, including 15 patients (12%) who had a history of veterinary occupational exposure, 8 patients (6.4%) who were animal skin and meat processors, and 87 patients (70%) who were cattle and sheep breeders with or without contact or eating uncooked beef, mutton, and dairy products; only 15 patients (12%) did not have a contact history.
Among the 125 patients with brucellosis, 47 were in an acute stage (the duration ranged from 1 week to 6 months), 43 were in a chronic stage (the duration ranged from 7 months to 2 months), and 35 were in a convalescent stage (the duration of treatment ended from 2 months to 2 years). The clinical characteristics of patients at different clinical stages were analyzed. The results showed that there were some differences in clinical manifestations between acute and chronic patients. The main clinical manifestations of acute stage patients were fever, fatigue, muscle and joint pain, and hyperhidrosis. The main clinical manifestations of chronic stage patients were fatigue, fever, and joint pain. Few patients showed sequelae after clinical cure, such as fatigue syndrome and intermittent joint pain, and few patients had hyperhidrosis (Table
Clinical characteristics of acute, chronic, and convalescent patients.
Clinical manifestations | Acute stage ( |
Chronic stage ( |
Convalescent stage ( |
---|---|---|---|
Fever | 46 (97.9%) | 39 (90.7%) | 0 |
Fatigue | 40 (85.1%) | 41 (95.3%) | 3 (8.6%) |
Chills | 20 (42.6%) | 5 (11.6%) | 0 |
Sweats | 30 (63.8%) | 10 (23.3%) | 1 (2.9%) |
Joint pain | 39 (83%) | 35 (81.4%) | 2 (5.7%) |
Headache | 15 (31.9%) | 5 (11.6%) | 0 |
Muscle pain | 40 (85.1%) | 20 (46.5%) | 0 |
Weight loss | 9 (21.3%) | 2 (4.7%) | 0 |
Cough | 7 (14.9%) | 0 | 0 |
Orchitis/epididymitis | 5 (14.7%) | 2 (7.7%) | 0 |
The laboratory indicators of all patients were analyzed. All patients were positive for erythrocyte plate test and tube agglutination test. The titers of antibodies in acute patients were 1 : 100-1 : 800, in chronic patients 1 : 50-1 : 800, and in convalescent patients 1 : 50-1 : 200. In acute stage, the positive rate of blood culture was relatively low, and there were no positive results in blood culture in patients at chronic stage. The abnormal rate of erythrocyte sedimentation and the C reactive protein levels in the chronic phase were lower than those in the acute phase.
Laboratory characteristics of acute, chronic, and convalescent patients.
Laboratory examinations | Acute stage ( |
Chronic stage ( |
Convalescent stage ( |
---|---|---|---|
Positive erythrocyte plate test | 47 (100%) | 43 (100%) | 35 (100%) |
Positive blood culture | 7 (14.9%) | 0 | 0 |
Leucocytopenia | 2 (4.3%) | 3 (7%) | 0 |
Hyperleukocytosis | 4 (8.5%) | 0 | 0 |
Anemia | 6 (12.8%) | 8 (18.6%) | 0 |
Thrombocytopenia | 2 (4.3%) | 5 (11.6%) | 0 |
Increased erythrocyte sedimentation rate | 40 (85.1%) | 28 (65.1%) | 0 |
Increased C reactive protein | 34 (72.3%) | 26 (60.5%) | 0 |
Enlargement of liver, spleen, and (or) lymph nodes | 15 (31.9%) | 16 (37.2%) | 0 |
Abnormal liver function | 13 (27.7%) | 3 (7%) | 0 |
Abnormal urine routine | 17 (36.2%) | 11 (25.6%) | 0 |
Abnormal chest X-ray | 8 (17%) | 2 (4.7%) | 0 |
The expressions of PD-1 on CD4+ and CD8+ T cells in the acute brucellosis group were significantly higher than those in the chronic brucellosis group (
Expression ratio of PD-1 on the CD4+ T cell surface in the peripheral blood of different groups.
Expression ratio of PD-1 on the CD8+ T cell surface in the peripheral blood of different groups.
The expressions of T lymphocyte subsets in the peripheral blood of patients with acute, chronic, and convalescent brucellosis and healthy controls were detected by flow cytometry. The results showed that (1) CD4+ IFN-gamma (Th1) lymphocyte expressions in the peripheral blood of patients in the acute stage were significantly higher than those of patients in the chronic, convalescent, and healthy control groups (
Expression ratio of Th1 in the peripheral blood of different groups.
Expression ratio of Th2 in the peripheral blood of different groups.
Expression ratio of Th17 in the peripheral blood of different groups.
Expression ratio of Treg in the peripheral blood of different groups.
The expression ratios of T lymphocyte subsets in convalescent patients with drug withdrawal less than 12 months and in those with drug withdrawal more than 12 months were compared and analyzed. The results showed that the expression ratio of Th1 cells in peripheral blood was significantly higher in convalescent patients with drug withdrawal over 12 months (
Expression ratio of Th1 in convalescent patients at different stages of treatment.
In this study, 35 patients who finished standard antimicrobial therapy had no clinical symptoms and normal laboratory tests. The withdrawal time ranged from 2 months to 24 months. Among them, 28 patients (80%) finished treatment within 12 months, and 7 patients finished treatment over 12 months. The correlation between the percentage of T lymphocyte in peripheral blood and the time of drug withdrawal showed that the percentage of Th1 cell expression in the peripheral blood of convalescent patients was negatively correlated with the time of drug withdrawal (
Correlation between T lymphocyte subsets and withdrawal time in convalescent patients.
Withdrawal time | Th1 | Th2 | Th17 | Treg | |
---|---|---|---|---|---|
Withdrawal time | 1.000 | ||||
Th1 | -0.679 |
1.000 | |||
Th2 | 0.056 | -0.232 | 1.000 | ||
Th17 | -0.168 | 0.036 | 0.109 | 1.000 | |
Treg | -0.256 | 0.088 | 0.047 | -0.136 | 1.000 |
Note.
Cytokines related to different T lymphocyte subsets in the peripheral serum of patients and healthy controls were detected by cytometric bead analysis. The following results were shown: (1) The expressions of Th1 cell-related cytokine IL-2 in acute and convalescent patients were higher than that in healthy controls (
Expression of IL-2 in the peripheral blood of different groups.
Expression of IFN-
Expression of IL-4 in the peripheral blood of different groups.
Expression of IL-5 in the peripheral blood of different groups.
Expression of IL-13A in the peripheral blood of different groups.
Expression of IL-17A in the peripheral blood of different groups.
Expression of IL-17F in the peripheral blood of different groups.
At present, the pathogenesis of brucellosis has not been fully understood. And T cell immune response is involved in the whole course of
The results show that the expression of Th1 cells in the peripheral blood of patients with brucellosis is significantly higher than that of healthy controls, and the expression level in patients at the acute stage is higher than that in patients at the chronic stage. Previous reports on the expression of Th1 cells in the peripheral blood of patients were controversial [
Th1 cells mainly secrete interferon gamma and IL-2. The results show that the expression of cytokine IL-2 in the peripheral serum of patients at the acute stage is the highest, which is significantly different from that of healthy people. The expression of IL-2 in convalescent patients after standard anti-
The mechanism of IFN-gamma in brucellosis infection is not completely clear yet. According to the results of this study, the secretion of IFN-gamma is upregulated in patients with
The main cytokines secreted by Th2 cells are interleukin-4 (IL-4), interleukin-5 (IL-5), and interleukin-13 (IL-13) [
We find that the expression of IL-4 is the highest in chronic and convalescent patients, which is significantly higher than that in acute patients. The increased expression of IL-4 in chronic patients is consistent with the change of Th2, suggesting that IL-4 is involved in the chronicity of
Th17 cells are important immune cells associated with infection [
IL-17A and IL-17F are the main effector molecules of Th17 cells [
Studies on the pathogenesis of PD-1 in infections indicate that PD-1 plays a role in mediating the progress of infection into chronicity [
Previous reports showed that
Our study has this limitation: we did not observe the dynamic changes of these routine parameters and levels of T cell subsets and cytokine in the course of drug treatment.
In summary, Th1, Th17, and Treg cell immunity are predominant in the acute phase and Th2, Th17, and Treg cell immunity predominated in the chronic phase after
The data used and/or analyzed in the current study are available from the corresponding author on reasonable request.
The authors declare no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
This work was supported by the Key Research and Development Projects of the Xinjiang Uygur Autonomous Region (Grant no. 2016B03047-1) and the National Natural Science Foundation of China (Grant no. 11461073).