REGISTRO DOI: 10.5281/zenodo.8361711
Lucas Araujo Souza1
Anna Paula Duque2
Myttermayer B. Santiago3
ABSTRACT
INTRODUCTION– Tuberculosis is an infectious disease of complex interaction with the host, the understanding of this process can take major steps in knowledge towards diagnosis and treatment even more efficiently. The presence of antiphospholipid antibodies (aPL) has been shown on articles in various infections, as well as in tuberculosis and autoimmune disease, but the repercussions on patients is unclear. OBJECTIVE– This work studied the prevalence of three antibodies (antiphosphatidylserine, antiprothrombin and antiannexin V) in serum of fifty patients with lung tuberculosis in supervision at the service of infectology inSanta Izabel Hospital. METHODS– This is a descriptive study of cases series report. A sample of blood was collected from patients in supervision at the ambulatory service of the hospital from September 2009 to December 2009. The serum levels of the studied antibodies were determined and analyzed using descriptive statistics at the Excel 2016 software, determining its prevalence. RESULTS- Considering all fifty patients studied, thirteen (26%) had at least one antibody positive. The most prevalent antibody was antiprothrombin with 14%. Antiphosphatidylserine and antiannexin V respectively showed 6% and 8% of prevalence. Considering the sex, 20% of female patients had positive tests for antiprothrombin and 12% for antiphosphatidylserine, it wasn’t found positive tests to antiannexin V in the female population. In male sex the antibodies prevalence of antiprothrombin, antiphosphatidylserine and antiannexine was, respectively, 8%, 4% and 12%. LIMITATIONS OF THE STUDY AND CONCLUSION- The study presented limitations in order to investigate the clinical outcomes of the patients studied and it was not possible to access the epidemiological data of the patients. The objective to demonstrate the prevalence of antibodies was reached, but other studies are needed in light of the theme since that it was only possible to compare data of prevalence of only one antibody due to the lack of studies about the subject.
Keywords: 1. Tuberculosis; 2. Antiphospholipid antibody; 3. Antiphosphatidylserine; 4. Antiprothrombin 5. Antiannexin V.
1. INTRODUCTION
It is understood by phospholipids as the main molecule forming the cell membrane, it is an amphipathic compound derived from phosphatidic acid. This is formed, generically, by glycerol, fatty acids, a phosphate group and a polar molecule attached to it. There are several types of membrane phospholipids such as cardiolipin and phosphatidylserine (1). It is also known that on the surface of the cell membrane, forming the phospholipid complexes, there are coagulation factors linked to them, which are proteins responsible for the balance of thrombogenic phenomena (2).
Antiphospholipid antibodies (aPLs) are immunoglobulins that bind to membrane phospholipids and studies of these antibodies have made an important contribution to understanding the clinical conditions linked to this autoimmunity process (3). The first step in the discovery of such antibodies took place with serological tests (Bordet–Wasserman test) in the diagnosis of syphilis in 1952, which then gave rise to the VDRL test, still used in medical routine today. It was from this discovery that the anticardiolipin antibody (aCL) emerged and a dimension of the importance of studies on aPLs antibodies was given (4).
Still on the subject, thanks to false positive values in the VDRL test in patients with systemic lupus erythematosus (SLE) in the tegumentary form, the presence of a new antibody in the serum of these patients was discovered. Soon, a statistical association was made between the antibody and characteristic clinical findings, which instigated the discovery of a syndrome marked by a state of hypercoagulability in 1986 by Hughes (5). However, it was only in 1987 that antiphospholipid antibody syndrome (APS) was so named. Because the findings were originally described in patients with lupus, the antibody was then baptized as a lupus anticoagulant (6).
It is known that antibodies that bind to proteins of phospholipid complexes (antiprothrombin and anti-annexin) are related to a procoagulation state and to obstetric outcomes marked by recurrent miscarriage. Likewise, such antibodies titrated in patients with APS and SLE have already been found in the literature (7) (8) (9). One hypothesis is that these antibodies would cause an activation of the endothelial cells, culminating in the expression of adhesion molecules, secretion of cytokines and interference in the metabolism of prostaglandins and, consequently, of thromboxane (10). A second theory focuses on the injury of the vascular endothelium by an oxidative process associated with low molecular weight lipid (LDL) that would trigger thrombogenic effects (11). Mechanisms by which aPLs antibodies interfere with the regulatory function of prothrombin, annexin V,
It is possible to verify titers of these antibodies in patients using pharmacological therapies (15) and in some infections such as HIV (16), hepatitis C (17) and tuberculosis (18), although most of the studied patients did not present relevant clinical evolutions. The presence of antibodies in infectious diseases is not commonly associated with complications, when compared to SLE (19), however, thrombosis and APS have already been reported in patients with tuberculosis, chronic viral infections, skin infection, pneumonia and urinary tract infection, with high titles of aPLs (20) (21) (22) (23) (24) (25).
Regarding tuberculosis, it is seen that most studies do not demonstrate modified clinical outcomes in the presence of anticardiolipin antibodies and lupus anticoagulants, for example. It is commonly noted only a predilection of anticardiolipin in the serum of patients with the infection, but there is also no evidence that these bring prognostic value to the patient (18) (25). Therefore, it is necessary that the case reports of thromboembolic phenomena seen in the literature in patients with tuberculosis (1.5-3.4%), and even episodes of Catastrophic APS in patients with pulmonary TB, be better investigated in the in order to understand its etiopathogenesis (21) (26).
2. OBJECTIVES
To analyze the prevalence of antibodies: antiphosphatidylserine, antiannexin and antiprothrombin in the serum of patients with pulmonary tuberculosis being followed up at an Infectology service in Bahia.
3. METHODOLOGY
3.1 Study design
Descriptive study of the case series type, whose population is composed of patients with pulmonary TB (diagnosis based on clinical and radiological criteria and AFB positivity in sputum) before or up to one month before the start of treatment.
3.2 Place of study
The study was carried out with patients undergoing outpatient treatment at Hospital Santa Izabel in the State of Bahia located in the city of Salvador. The hospital was founded in 1549, as Charity hospital, and is currently a huge hospital and a reference in several areas.
3.3 Study population and study period
The population consisted of patients with pulmonary tuberculosis being followed up at the Hospital Santa Izabel service who had diagnosis based on clinical, radiological, BAAR (alcohol-acid resistant bacillus) positivity of sputum. Any rheumatological condition was ruled out. Patients under 18 years of age, patients who refused to participate in the research or to sign the informed consent form, patients who withdrew from participating during the development of the research were excluded from the sample.
Data collection was carried out from September 2009 to December 2009.
3.4 Data collection methods
It was made blood collection for the investigation of antiphosphatidylserine, antiannexin and antiprothrombin antibodies by the ELISA method according to the manufacturer’s instructions (INOVA Diagnostics Inc.®). The tests were carried out at the LBPC laboratories of the Gonçalo Muniz Foundation (Fiocruz) and at the Faculty of Pharmacy at the Federal University of Bahia.
3.5 Analysis plan
The collected data were tabulated and analyzed using descriptive statistics of the variables using Excel 2016 spreadsheets.
3.6 Sample size
It is a convenience sample composed of 50 patients, 25 male and 25 female.
3.7 Ethical considerations
The project was submitted to the Research Ethics Committee (CEP) of Hospital Santa Izabel, Salvador-BA, and data collection was only initiated upon approval, under protocol number: 39/2009.All patients were properly instructed to read and sign the Informed Consent Form before answering the questionnaire.
The research complied with the norms of resolution 466/12 of the National Health Council (CNS).
4. RESULTS
Of the 50 (fifty) patients studied, 13 (thirteen) (26%) had a positive reaction to at least one of the studied antibodies. The aPT IgG antibody had the highest absolute value, with 7 patients presenting serum values above the reference cutoff (10 u). The aPS IgG/M had a total number of patients above the reference cutoff value (also 10 u) equal to 4. Antiannexin V reached a serum value above the cutoff value (8 u) in only 3 of the patients. (FIGURE 1)
FIGURE 1. Absolute number of patients who presented a positive reaction for each test studied; reference cutoff value for antinexin V – 8u; reference cutoff value for prothrombin and phosphatidylserine – 10u; aPT IgG – Antiprothrombin Immunoglobulin G; aPS IgG/M – Antiphosphatidylserine Immunoglobulin G or M.
Evaluating the prevalence of each antibody, none reached a prevalence greater than 20%. With regard to sex, it was observed that antiannexin V IgG in the studied sample has a higher prevalence in men (12%) than in women (0%) with an overall prevalence of 6%. The antiphosphatidylserine IgG/M had a positive reaction in 3 women, which represents 12% of the number of women in the sample, whereas men only 1 (4%) had a serum antibody value above the reference cutoff value. The antiprothrombin antibody was the one that stood out the most in the sample, with a total prevalence of 14%, of which 4% were men and 10% women. Females represent 14% of the total number of positive tests in the 3 three tests, while males have a prevalence of 12%. (FIGURE 2)
FIGURE 2. Prevalence values of antibodies in the general study population and by gender of patients; AntiPS IgG/M: antiphosphatidylserine immunoglobulin G or M; AntiPT IgG: antiprothrombin immunoglobulin G.
Considering the serum values in units, it was noticed that most patients had low serum antibody values, with aPT being the most prominent in the group, followed by aPS IgG. Only 1 patient had a positive reaction to more than one antibody. (FIGURE 3)
FIGURE 3. Serum values of antiprothrombin, antiphosphatidylserine and antiannexin antibodies in patients with pulmonary tuberculosis. Chart reveals low serum values of most of the studied patients. A patient with elevated aPT stands out, as well as another with aPS IgG showing present IgM fraction. The thickest line corresponds to the cutoff point.
5. DISCUSSION
The association of infectious processes with thrombogenic phenomena is already well documented in the literature. There are studies that indicate tuberculosis as a predisposing factor to deep venous thrombosis, as well as reports of other infections that also presented thrombosis associated with the presence of antiphospholipid antibodies (25) (26) (27).
A publication by the European League of Rheumatology (EULLAR) showed the growing importance of membrane cofactors (prothrombin and annexin V) and the phospholipid phosphatidylserine in the etiopathogenesis of the thrombotic phenomena of APS (28). The findings of the present study show an expressive prevalence of these mentioned antibodies in patients with tuberculosis (total 26%), with aPT, aPS and anti-annexin presenting separately 14%, 8% and 6% of prevalence, respectively.
Considering that the prevalence values of aPL antibodies in the general population, they vary between 1% and 5%, the finding of an increased value in this study group (26%) brings an alert to the association of cause and effect between tuberculosis and antibodies with possible thrombotic outcomes(29). This is not the first study evaluating aPL in tuberculosis, Santiago et al. despite having found a high prevalence of aPL antibodies (53% for lupus anticoagulant and 47% for aCL), it did not find an association with thrombosis in its studied population, however it does not invalidate the possibility either (20).
Santiago et al. still claims that there is a difference in the biological role of aPL in infections. Possibly, the difference in the biological role and the low titers of aPL may contribute to the explanation of a lower clinical association with thrombosis in patients with infection. In addition, the author also demonstrates that aPL antibody titers change over time, which may interfere with clinical outcomes (20). The findings of this study corroborate the author’s findings, in which low titers were observed in the vast majority of the evaluated antibodies.
In fact, the literature has suggested that the presence of aPL antibodies in infections has no direct prognostic value for thrombosis, but that infections behave as risk factors for thrombogenic phenomena in patients who already have antibodies previously in your serum. Likewise, other conditions such as serum values above 40u of APLs, smoking, oral contraceptives and pregnancy also appear as risk factors (20) (29).
Considering the aPT antibody, corroborating what the literature has suggested, it is clear that most studies do not correlate it with modified clinical outcomes, contrary to what is seen with the antibody to the complex formed by the phosphatidylserine molecule directly linked to prothrombin (aPS/PT). For example, Bermudez-Bermejo et al. found a prevalence of 62% of these antibodies in patients with primary APS and an association of thrombosis with IgM antibodies in this group equal to 87.7% (p= 0.003) (30).
Atsumi et al. also brought correlation of the same complex with clinical manifestations of APS [OR 4.39, 95% CI] and demonstrated that the antibody can be as specific for APS as the aCL-dependent Beta-2-glycoprotein I (93% for both). In this same study, aPT prevalence equal to 15% is demonstrated in patients with primary APS, which demonstrates agreement with the value of the present study (14%). However, the author does not find statistical significance of the antibody as a predictor of thrombosis in his patients (31).
Now in patients with SLE, Nojima et al. showed a prevalence of the aPT antibody equal to 54% and there was also no association with thrombosis, unlike aPS/PT which, with a prevalence of 43%, showed thrombotic outcomes(9). In obstetric conditions, most studies also failed to demonstrate an association between aPT antibody and pregnancy morbidities. However, the clinical significance of the antibody in patients with adverse pregnancy outcomes has already been confirmed, and therefore attention should be paid to its presence in other conditions, such as tuberculosis seen in this study (32) (33) (34) (35 ).
It is noteworthy that there is already a real association of aPS/PT antibodies with thrombosis and even as a diagnostic test for APS, as discussed earlier. Some researchers already suggest the incorporation of such antibodies to the APS criteria, but there is still resistance due to the lack of reliable evidence of applicability for diagnostic use, which may change with the advances of new research and bring discoveries about the antibodies aPS, aPT and antiannexin in infectious conditions(36).
With regard to anti-annexin V, Nojima et al., in another study, tested the antibody as a risk factor for vein and/or artery thrombosis and for recurrent miscarriage in 168 patients (165 women and 3 men) with SLE. Only in recurrent abortions, anti-annexin V obtained statistical significance [OR 5.9, 95% CI] p= 0.0119 with a significant prevalence of 64% (37). This denotes a greater association of antiannexin V with obstetric morbidities. Sorice et al. still demonstrated for the first time a prevalence of 26% in mononucleosis infection, however the repercussions of the findings of these antibodies in infectious conditions are not known, which still needs to be clarified (38).
Likewise, Sater et al., in a retrospective study, found expressive results in 309 patients with a history of abortion for the aPS antibody [OR 18.56 (6.63 – 51.91), 95% CI] p= 0.001, suggesting it as a pathogenic agent in cases of abortion, with a prevalence of 19% in the study population (33).
There are several findings of thrombotic phenomena in tuberculosis, such as: cerebral vein thrombosis (39); deep vein thrombosis in endometrial tuberculosis (40); portal vein thrombosis in hepatic tuberculosis (41); subclavian thrombosis in pulmonary tuberculosis (42) and pulmonary thromboembolism in an HIV-positive patient with pulmonary tuberculosis (43). Publications in general do not clarify the etiology of the process, corroborating the idea that research needs to be done in light of the theme.
6. CONCLUSION AND LIMITATIONS OF THE STUDY
Understanding the thrombotic phenomena in tuberculosis and their peculiarities requires greater efforts. This study denotes that, considering the prevalence of aPL antibodies in the general population (1 to 5%), the finding of 26% in the group of patients with tuberculosis draws attention to the risk of developing thrombosis, considering the thrombogenic activity of these antibodies already well known. documented in the literature.
It was not possible to test the association of tuberculosis and thrombotic phenomena with antibodies to verify their relevance in the etiopathogenesis of thromboses, but the prevalence found in the studied group, mainly of aPT, does not rule out the possibility of a cause and effect relationship, which invites the scientific community to carry out research on the subject. Another limitation of the study was that it was not possible to reassess the serum values of antibodies to assess possible fluctuations in values and behavior after tuberculosis treatment.
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