CD19+CD5+ B-cell expansion and risk of pre-eclampsia

Rawhia H Eledel; Maha A Bassuoni; Wafaa M Radwan; Alaa Masoud; Sara M Eldeeb

doi:10.4103/1110-2098.192433

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ORIGINAL ARTICLE

Year : 2016 | Volume : 29 | Issue : 2 | Page : 319-323

CD19⁺CD5⁺ B-cell expansion and risk of pre-eclampsia

Rawhia H Eledel¹, Maha A Bassuoni¹, Wafaa M Radwan¹, Alaa Masoud², Sara M Eldeeb¹
¹ Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Menoufia, Egypt
² Department of Obstetrics and Gynecology, Faculty of Medicine, Menoufia University, Menoufia, Egypt

Date of Submission	20-Oct-2014
Date of Acceptance	18-Dec-2014
Date of Web Publication	18-Oct-2016

Correspondence Address:
Sara M Eldeeb
Department of Clinical Pathology, Faculty of Medicine, Menoufia University, 83 Saad Zaghlol St, El Bar El Sharky, Shebin El Kom, Menofia, 32511
Egypt

Source of Support: None, Conflict of Interest: None

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DOI: 10.4103/1110-2098.192433

Abstract

Background:
Pre-eclampsia (PE) is a devastating pregnancy-associated disorder affecting 5–8% of pregnant women worldwide. It emerges as an autoimmune-driven disease. The autoantibodies against angiotensin type 1 receptor II have been proposed to account for PE symptoms. CD19^(+)CD5(+) B-1a B cells constitute the main source of natural and polyreactive antibodies, which can be directed against own structures.
Objectives:
The aim of this study was to identify the B-cell subpopulation responsible for autoantibody production during PE and its correlation with b subunit of hCG levels.
CD19⁺CD5⁺ B-cell percentages were measured using flow cytometry in 36 PE women in their third trimester (20 severe and 16 mild) and 15 age-matched normotensive pregnant women with normally developing pregnancies in their third trimester.
Patients and methods:
We observed significantly higher levels of frequency of CD19⁺/CD5⁺ B lymphocytes in the mild and severe PE groups compared with the levels observed in normotensive women with normally developing pregnancies in the third trimester (mild PE, 28.7 ± 14.3%, n = 16; severe PE, 29.9 ± 9.8%, n = 20; control, 11.3 ± 6.4%, n = 15; P < 0.01 and P < 0.001, respectively). However, comparison of the mild PE group with the severe PE group showed comparable levels of CD19⁺/CD5⁺ B cells (P > 0.05).
Results:
Correlation between the percentages of CD19⁺/CD5⁺ B lymphocytes and b hCG levels was not significant. Moreover, correlations between the percentages of CD19⁺/CD5⁺ B lymphocytes and other laboratory parameters were not significant.
Conclusion:
CD19⁺/CD5⁺ cells emerge as a novel PE marker. Their frequency does not seem to be regulated by the increased serum hCG levels. The detection and quantification of CD19⁺/CD5⁺ cells in maternal blood may open vast new therapeutic opportunities.

Keywords: AT1-AAs, autoantibodies, CD19⁺CD5⁺ cells, pre-eclampsia, β hCG

How to cite this article:
Eledel RH, Bassuoni MA, Radwan WM, Masoud A, Eldeeb SM. CD19⁺CD5⁺ B-cell expansion and risk of pre-eclampsia. Menoufia Med J 2016;29:319-23

How to cite this URL:
Eledel RH, Bassuoni MA, Radwan WM, Masoud A, Eldeeb SM. CD19⁺CD5⁺ B-cell expansion and risk of pre-eclampsia. Menoufia Med J [serial online] 2016 [cited 2023 Mar 21];29:319-23. Available from: http://www.mmj.eg.net/text.asp?2016/29/2/319/192433

Introduction

Pre-eclampsia (PE) is a devastating pregnancy-associated disorder in humans, characterized by the onset of hypertension, proteinuria, and edema after the 20th week of pregnancy. It represents one of the most frequent medical complications of pregnancy, affecting about 5–8% of all pregnant women ^[1]. Limited progress has been achieved in identifying the causes, with the induction of delivery being the only available treatment for the clinical symptoms ^[2]. In recent times, PE has been suggested to represent an autoimmune disease ^[3]. This is based on the fact that many autoantibodies were identified in PE patients and suspected to account for the symptoms of the disease ^[4],[5],[6]. Much attention was focused on autoantibodies against angiotensin type 1 receptor II (AT1-AAs) because they have been detected in the serum of PE patients ^[7]. These autoantibodies are able to bind to and activate the AT1 angiotensin receptor, leading to the release of antiangiogenic factors from the placenta, which are reportedly involved in the origin of the disease ^[8],[9],[10].

Despite the efforts taken on identifying autoantibodies related to PE, no attention has been so far focused on the cell population responsible for their production. This is a very important piece of information because it may provide a novel target of actions in preventing the onset of the disease in patients at risk. B-1a B cells double positive for CD19 and CD5 are a distinctive population of B cells originated during the embryonic life and represent≈20% of the circulating B cells during adulthood ^[1]. CD19⁺/CD5⁺ cells are responsible for the production of so-called natural antibodies, which are characterized by their low-affinity, polyreactivity, and self-reactivity.

B-1a B cells possess characteristics that reflect their strong link to autoimmunity ^[2]. In fact, B-1a B levels have been shown to be elevated in several human autoimmune diseases such as Sjogren syndrome ^[3], rheumatoid arthritis ^[4], lupus erythematosus ^{[15],[16],[17]}, and type 1 diabetes mellitus ^[8].

The aim of this work was to analyze the dynamics of B-1a B cells in pregnant women with PE versus normal pregnancies and its correlation with b subunit of hCG levels.

Participants and Methods

Study participants

This study was carried out on 36 PE women in their third trimester, with an age range of 20–41 years old, attending the obstetric and gynecology department, Menoufia University hospitals. Of the 36 PE women, 20 were diagnosed with severe PE and the remaining 16 were diagnosed with mild PE. A total of 15 age-matched and sex-matched normotensive pregnant women in their third trimester were included as controls. The characteristics of the recruited participants are summarized in [Table 1]. This study was approved by the ethical committee of the faculty of medicine, Menoufia University. All patients provided signed informed consent to provide a blood sample and to review the medical record for research purposes (according to Helsinki declaration).

Table 1: Patients' characteristics

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Immunophenotyping of peripheral blood B lymphocytes

Peripheral blood cells were stained with FITC-conjugated antihuman CD5 and PE-conjugated antihuman CD19 (BioLegend, San Diego, California, USA) for 30 min at 4°C, followed by whole-blood lysis. The data were acquired and analyzed with a BD FACSCalibur flow cytometry (BD Biosciences, Qume Drive, San Jose, CA, USA). The percentage of CD19⁺/CD5⁺ B lymphocytes was defined using gating strategy on CD19⁺ cells. The samples of controls and patients were examined using the same settings and conditions for comparison. Samples were analyzed within 24 h of collection.

Measuring β hCG levels

Serum b hCG concentrations were measured by VIDAS hCG (bioMérieux Inc., Chemin de l'Orme - 69 280 Marcy l'Etoile - France) according to the manufacturer's instructions.

Results

The frequency of CD19⁺/CD5⁺ B lymphocytes

We observed that peripheral blood samples from all PE patients (both mild and severe groups) in their third trimester presented significantly higher levels of frequency of CD19⁺/CD5⁺ B lymphocytes as compared with the levels observed in samples of normotensive women with normally developing pregnancies in the third trimester (percentage of CD19⁺/CD5⁺ B lymphocytes: all PE, 29.4 ± 11.8%, n = 36; control, 11.3 ± 6.4%, n = 15; P<0.001).

When we compared the percentages of CD19⁺/CD5⁺ B lymphocytes in the mild PE group with the control group (mild PE, 28.7 ± 14.3%, n = 16; control, 11.3 ± 6.4%, n = 15; P<0.01), or the severe PE group with the control group (severe PE, 29.9 ± 9.8%, n = 20; control,11.3 ± 6.4%, n = 15; P<0.001), the levels of these cells were significantly more higher ([Figure 1] and [Figure 2]). However, comparison of the mild PE group with the severe PE group showed similar levels of CD19⁺/CD5⁺ B cells (percentage of CD19⁺/CD5⁺ B lymphocytes: mild PE, 28.7 ± 14.3%, n = 16; severe PE 29.9 ± 9.8%, n = 20; P > 0.05; [Figure 1] and [Figure 2]).

Figure 1: Higher frequency of CD19⁺/CD5⁺ B lymphocytes in pre-eclampsia (PE) women (both mild and severe) compared with the levels observed in normotensive women in their third trimester. Representative dot plot of flow cytometric analysis of CD19⁺/CD5⁺ B lymphocytes is shown for each group (the top panel represents controls, whereas the lower panel represents the mild and severe PE groups)

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Figure 2: CD19⁺/CD5⁺ B lymphocyte percentages in the pre-eclampsia (PE) and the control groups. Significantly higher levels of CD19⁺/CD5⁺ B cells were found in the mild (n = 16) and severe PE (n = 20) groups compared with normotensive controls (n = 15), whereas the levels of CD19⁺/CD5⁺ B cells were comparable between the mild and severe PE groups

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The changes observed in the levels of CD19⁺/CD5⁺ B cells were not a consequence of variations in the levels of total peripheral B cells, because there was no significant difference in the levels of CD19⁺ B-cell in normotensive pregnancies and PE patients (total peripheral B cells: all PE, 6.61 ± 2.97%, n = 36; control, 7.86 ± 2.72%, n = 15; P > 0.05; [Figure 3]).

Figure 3: Levels of total B cells were comparable between pre-eclampsia patients (n = 36) and normotensive controls (n = 15)

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β hCG levels

Comparison between the mild PE, the severe PE, and the control group as regards b hCG levels revealed that significantly higher values were present in the mild PE and the severe PE group compared with the control group (mild PE, 16383.7 ± 11071.0 mIU/ml, n = 16; severe PE, 24131.9 ± 20088.2 mIU/ml, n = 20; control, 7396 ± 1601 mIU/ml, n = 15; P <0.05), whereas no significant difference was found on comparing the mild PE group with the severe PE group (P > 0.05) [Figure 4].

Figure 4: Comparison between the mild pre-eclampsia (PE) (n = 16), the severe PE (n = 20), and the control group (n = 15) as regards β hCG levels revealed that significantly higher values were present in the mild PE and the severe PE group compared with the control group

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Correlation between the percentages of CD19⁺/CD5⁺ B lymphocytes and b hCG levels was not significant. Moreover, correlations between the percentages of CD19⁺/CD5⁺ B lymphocytes and other laboratory parameters were not significant [Table 2].

Table 2: Correlation between CD19⁺/CD5⁺% in pre-eclampsia and other laboratory parameters

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Discussion

In the last years, the concept of PE as an autoimmune disease has been introduced ^[3]. This assertion is based on the fact that several autoantibodies were described to be present in PE patients. Among others, the autoantibodies against the AT1-AA have gained attention. Since Wallukat et al. ^[7] described the presence of AT1-AA in PE patients, many features of the disease has been explained by the ability of these autoantibodies to bind to and to activate the angiotensin receptor on a variety of cells ^{[19],[20],[21],[22],[23]}. However, despite much information concerning the presence of autoantibodies in PE patients, no attention has so far been focused on the cells producing them. Autoimmune diseases are mainly triggered by the production of autoantibodies by B cells. There are two different lineages of B cells: B1 and B2. B2 cells originate continuously from precursors present in the bone marrow and then migrate to the spleen, where they complete their maturation ^[4]. Under nonpathological situations, those B cells that recognize autoantigens are likely to be eliminated. We focused our attention on the B-1a B cells. B1, unlike B2 cells, originate from a progenitor in the fetal liver and are able to self-replenish during adulthood ^[2]. B-1a B cells are the primary source of natural antibodies ^[5]. Natural antibodies are produced in the complete absence of antigenic stimulation and are polyreactive and have been reported to cross-react with multiple antigens including autoantigens ^[6]. In fact, B-1a B cells and their produced polyreactive antibodies have been related to autoimmune diseases in humans ^{[13],[14],[15],[16]}; however, their ability to cause autoimmunity has been demonstrated in mouse models of disease ^[7]. B-1a B cells were classically defined as CD19⁺/CD5⁺ cells ^[2]. It has been shown that human CD19⁺/CD5⁺ B-1a B cells may be activated and produce autoantibodies ^[7].

In this study, we observed that the levels of CD19⁺/CD5⁺ cells in the mild and severe PE groups were significantly higher compared with normotensive women having normally progressing pregnancies in their third trimester. This was not because of an increase in the levels of CD19⁺ B cells, because they were comparable between the two groups. These results are in agreement with those reported by Jensen et al. ^[8]. However, the levels of CD19⁺/CD5⁺ cells were not related to disease severity, because they were similar in both the mild and severe PE groups.

In several well-characterized autoimmune diseases, a similar augmentation in the levels of these cells compared with the healthy controls could be observed ^{[13],[14],[15],[16]}. This interesting dynamic of CD19⁺/CD5⁺ cells in pregnancy raises the question concerning the physiological relevance of this phenomenon.

In-vitro studies by Jensen et al. ^[8] showed that the variations in CD19⁺/CD5⁺ cells strongly related to the high serum levels of hCG present in PE patients and that CD19⁺/CD5⁺ cells highly express the hCG receptor and expand in vitro on hCG influence. They also showed that placenta explants from PE patients released higher amounts of hCG compared with placenta explants from normotensive pregnant women. Furthermore, they demonstrated that isolated CD19⁺/CD5⁺ but not CD19⁺/CD5⁻ cells produce AT1-AA when cultured with serum from PE patients and that PE patients have very high levels of AT1-AA, whereas the levels of these antibodies are very low or absent in normotensive pregnant women in their third trimester.

In our study, we found significantly higher levels of serum hCG in PE patients than in normotensive pregnant women, but its levels were not correlated with the increased percentages of CD19⁺/CD5⁺ cells.

In PE patients, not only the production of AT1-AA by CD19⁺/CD5₊ cells was reported but also CD19⁺/CD5⁺ cells were detected in placental tissue ^[8]. In line with this, it has been shown that B-1a B cells in autoimmune diseases not only contribute to the onset of the pathology by the antibody production but also migrate to the sites of inflammation and are able to produce and secrete inflammatory and hazardous molecules — for example, cytokines, which will exacerbate the symptoms.

Conclusion

CD19⁺/CD5⁺ cells emerge as a novel PE marker and their levels correlate with the disease. Their frequency does not seem to be regulated by the increased serum hCG levels. The detection and quantification of CD19⁺/CD5⁺ cells in maternal blood may serve as a noninvasive diagnostic tool, which opens vast new therapeutic opportunities.

Conflicts of interest

There are no conflicts of interest.[28]

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Figures

[Figure 1], [Figure 2], [Figure 3], [Figure 4]

Tables

[Table 1], [Table 2]

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