Pregnancy Hypertension: An International Journal of Women’s Cardiovascular Health 1 (2011) 129–131

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Pregnancy Hypertension: An InternationalJournal of Women’s Cardiovascular Health

journal homepage: www.elsevier .com/locate /preghy

Short Communication

sFlt-1 and IP-10 in women with early-onset preeclampsia

Leandro De Oliveira a,b,c,⇑, Niels Olsen S. Câmara b, José C. Peraçoli d, Maria T. Peraçoli d,Antonio F. Moron a, Nelson Sass a,c

a Obstetrics Department, Universidade Federal de São Paulo, Brazilb Laboratory of Immunobiology of Transplantation, Universidade de São Paulo, Brazilc Laboratory of Clinical and Experimental Investigation, Hospital e Maternidade Vila Nova Cachoeirinha, São Paulo, Brazild Obstetrics Department, Universidade Estadual de São Paulo, Botucatu, Brazil

a r t i c l e i n f o a b s t r a c t

Article history:Available online 23 February 2011

Keywords:Angiogenic factorssFlt-1IP-10PlGFPreeclampsiaInflammation

2210-7789/$ - see front matter � 2011 Internationadoi:10.1016/j.preghy.2011.02.002

⇑ Corresponding author at: Obstetrics DepartmeBarros, 875, Universidade Federal de São Paulo, VPaulo 04024-002, Brazil. Tel.: +55 11 9949 8110; fa

E-mail address: Leminas@ig.com.br (L. De Olivei

Intense inflammatory response and an anti-angiogenic state have been implicated in thepathogenesis of preeclampsia. Here, we investigated this hypothesis evaluating the serumconcentrations of CXCL10/IP-10, sFlt-1, and PlGF in women with early-onset preeclampsia.CXCL10/IP-10 was measured by Cytometric Bead Array. sFlt-1 and PlGF were measured byautomated electrochemiluminescence immunoassay. The median serum concentration ofCXCL10/IP-10 was 109.5 pg/mL in preeclamptic women, as compared with 52.28 pg/mLin the controls (P = 0.0028). The mean serum level of sFlt-1 was 13,636 pg/mL in pre-eclamptic women, as compared with 2020 pg/mL in the controls (P < 0.0001). PlGF levelswere significantly lower in women with preeclampsia.

� 2011 International Society for the Study of Hypertension in Pregnancy. Published byElsevier B.V. All rights reserved.

1. Introduction

Preeclampsia occurs in about 5–8% of all pregnanciesand it constitutes a major cause of maternal and fetal mor-bidities and mortality [1]. The pathogenesis of preeclamp-sia has its roots on deficient trophoblast invasion and onthe failure in spiral artery remodeling. This incompletetransformation of the spiral arteries leads to inadequateplacental perfusion and consequently to placental oxida-tive stress [2]. The altered placenta then releases greatamounts of microparticles, debris, and anti-angiogenic fac-tors into the maternal circulation [3,4]. All these factors aresupposed to act in synergy to initiate and to maintain anintense inflammatory response and an anti-angiogenicstate. These alterations are responsible for the widespectrum of clinical manifestations of preeclampsia.

l Society for the Study of Hy

nt, Rua Napoleão deila Clementino, São

x: +55 11 3815 6308.ra).

Recently, Borzychowski et al. demonstrated the sys-temic inflammation of preeclampsia and emphasized theparticipation of the innate immune system in this inflam-matory scenario [5]. Additionally, Germain et al. demon-strated that maternal leukocytes in co-culture withsyncytial microparticles secrete higher amount of inflam-matory cytokines [6].

The anti-angiogenic state of preeclampsia is character-ized by increased endothelial permeability, vascular throm-bosis, platelet activation, and glomerular endotheliosis(that is a typical glomerular alteration of preeclampsia)[7]. Soluble fms-like tyrosine kinase 1 (sFlt-1) is ananti-angiogenic factor released in higher amounts bypreeclamptic placentas and it has been implicated in theendothelial dysfunction observed in preeclampsia. sFlit-1acts as a soluble receptor for VEGF and PlGF, interceptingthese angiogenic proteins before their binding to their nor-mal receptor at the cell membranes [7].

IP-10 is a chemokine of the CXC family that is inducedin the presence of inflammatory cytokines, such as TNF-aand INF-c [8]. IP-10 has been correlated with Th1

pertension in Pregnancy. Published by Elsevier B.V. All rights reserved.

130 L. De Oliveira et al. / Pregnancy Hypertension: An International Journal of Women’s Cardiovascular Health 1 (2011) 129–131

responses and with increased risk of rejection after renaltransplantation [9]. In addition, IP-10 is a potent inhibitorof angiogenesis in vivo [10]. Recently, Gotsch et al. demon-strated that higher serum concentration of IP-10 is foundin women with preeclampsia [11].

Considering the combined effect of inflammatory andanti-angiogenic factors in the clinical manifestation of pre-eclampsia, we decided to investigate the serum concentra-tions of sFlt-1, PlGF, and IP-10 in women with early-onsetpreeclampsia.

2. Materials and methods

2.1. Study design and sample collection

This study included 10 pregnant women with early-on-set preeclampsia (before 34 weeks gestation) and 10 normalpregnant women. Preeclampsia was diagnosed in the pres-ence of new-onset hypertension (blood pressure P140 �90 mm Hg) and significant proteinuria (P300 mg/24 h ordipstick measurement P2+). Each patient in the preeclamp-sia group was matched to a patient in the control groupaccording to the gestational age at the time of sample collec-tion. All patients in the control group were followed untilterm and had normal babies. Maternal blood samples wereobtained by venipuncture and drained into serum collectiontubes. Blood samples were centrifuged immediately aftercollection and sera were stored at �80 �C until requiredfor analyses. Our work has been approved by the ethicscommittee of our institution.

Table 1Clinical characteristics and perinatal outcomes from women with preeclampsia a

Variable Normal pregnancy

Maternal age 26.7 (3.2)Race White: 7

Not white: 3BMI 25.2 (3.1)Parity Primiparas: 3

Multiparas: 7Gestational age at blood draw 28.5 (2.6)Gestational age at delivery 38.5 (1.0)Neonatal birthweight (g) 3346 (291.3)24-h urine protein (g) Not evaluated

Fig. 1. Serum concentration (pg/mL) of sFlt-1 and PlGF in normal pregnant wo

2.2. Immunoassays

Serum concentrations of CXCL10/IP-10 were measuredin duplicate samples by Cytometric Bead Array using akit for chemokines (BD Biosciences, CA). Data were ana-lyzed using BD FACSCanto II (BD Biosciences, CA) equippedwith BD FACSDiva V6.1.1 operating software and FACAPArray software V1.0.

Serum concentrations of sFlt-1 and PlGF were measuredby automated electrochemiluminescence immunoassay(Roche Diagnostics, IN). Data were analyzed using Elecsys2010/Hitachi.

2.3. Statistical analyses

The statistical analyses were performed using the Prismsoftware (version 4.02, GraphPad Software Inc., San Diego,CA). Differences between results of CXCL10/IP-10, sFlt-1and PlGF were evaluated using Mann–Whitney U-test. Stu-dent t-test and exact Fisher test were used in the analysesof maternal and neonatal characteristics. Differences wereconsidered statistically significant at P < 0.05.

3. Results

The clinical characteristics and neonatal outcomes fromthe 20 patients enrolled in this study are summarized inTable 1.

nd normal controls (n = 10 in each group).

Preeclampsia P value

25.8 (4.7) 0.182White: 5Not white: 526.2 (4.0) 0.407Primiparas: 6Multiparas: 429.8 (2.5) 0.14630.8 (2.3) <0.00011457 (475.1) <0.00013.26 (2.2)

men and women with early-onset preeclampsia (n = 10 in each group).

L. De Oliveira et al. / Pregnancy Hypertension: An International Journal of Women’s Cardiovascular Health 1 (2011) 129–131 131

3.1. CXCL10/IP-10

The median serum concentration of CXCL10/IP-10was 109.5 pg/mL (42.21–216.2 pg/mL) in women withpreeclampsia, as compared with 52.28 pg/mL (12.58–104.8 pg/mL) in the controls (P = 0.0028).

3.2. sFlt-1 and PlGF

The mean serum concentration of sFlt-1 was 13,636 pg/mL (2803–26,498 pg/mL) in women with preeclampsia, ascompared with 2020 pg/mL (951.8–4045 pg/mL) inthe controls (P < 0.0001). The mean serum concentrationof PlGF was 87.54 pg/mL (17.46–239.0 pg/mL) in thepreeclampsia group, as compared with 479.8 pg/mL(64.5–1987 pg/mL) in the control group (P = 0.0068).Fig. 1 shows the distribution of sFlt-1 and PlGF in boththe groups. Our data showed correlation between sFlt-1and IP-10 levels in both the groups evaluated.

4. Discussion

In this work, we investigated serum concentrations ofIP-10, sFlt-1, and PlGF during the clinical manifestation ofearly-onset preeclampsia. The concentrations of IP-10and sFlt-1 were higher in preeclamptic women than in nor-mal controls. Inversely, the levels of PlGF were lower in thepreeclampsia group.

The mechanisms used for IP-10 to promote inflamma-tory response include the stimulation of T cell and mono-cyte migration and the promotion of T cell adhesion tothe endothelium [12]. In correlation with this effect, a re-cent work demonstrated that women with preeclampsiahave higher cytotoxic T-cell response to paternal antigenswhen compared with pregnant controls [13]. The effectof IP-10 as an anti-angiogenic factor may be an additionalcontribution for its inflammatory properties. Angiolilloet al. [10] demonstrated that this chemokine is able to im-pair the tubule-like formation of the human umbilical veincells in ‘‘ in vitro’’ cultures. These characteristics suggestthat IP-10 may contribute to the physiopathology ofpreeclampsia.

The role of sFlt-1 as an anti-angiogenic factor in thepathogenesis of preeclampsia has been extensively de-scribed in the last decade [14,15]. Levine et al. demon-strated that changes in the serum concentration of sFlt-1and PlGF can occur 5 weeks before the clinical manifesta-tion of preeclampsia [16]. Therefore, these factors havebeen interpreted as important markers for the disease. Inour work, we evaluated the serum concentration of sFlt-1in women with early-onset preeclampsia, and we alsofound significant increase in its concentrations. An inverseresult was found in terms of PlGF levels, which were signif-icantly lower in the study group when compared to normalpatients. It is important to note that cases of early-onset

preeclampsia are in general marked by severe clinicalmanifestations of the disease. This condition may be corre-lated with the high concentrations of sFlt-1 found here.

In conclusion, our data corroborate with other authorsin the literature and support the evidence that preeclamp-sia is a combined result of inflammatory responses with ananti-angiogenic state.

References

[1] American College of Obstetricians and Gynecologists. Committee onObstetric Practice. Diagnosis and management of preeclampsia andeclampsia: ACOG practice bulletin, No. 33. Int J Gynaecol Obstet2002;77:67–75.

[2] Pijnenborg R, Dixon G, Robertson WB, Brosens I. Trophoblasticinvasion of human decidua from 8 to 18 weeks of pregnancy.Placenta 1980;1:3–19.

[3] Burton GJ, Charnock-Jones DS, Jauniaux E. Regulation of vasculargrowth and function in the human placenta. Reproduction2009;138(6):895–902.

[4] Redman CW, Sargent IL. Circulating microparticles in normalpregnancy and pre-eclampsia. Placenta 2008;29(Suppl. A):S73–7.

[5] Borzychowski AM, Sargent IL, Redman CW. Inflammation andpreeclampsia. Semin Fetal Neonatal Med 2006;11(5):309–16.

[6] Germain SJ, Sacks GP, Soorana SR, Sargent IL, Redman CW. Systemicinflammatory priming in normal pregnancy and preeclampsia: therole of circulating syncytiotrophoblast microparticles. J Immunol2007;178:5949–56.

[7] Wang A, Rana S, Karumanchi SA. The role of angiogenic factors in itspathogenesis. Physiology 2009;24:147–58.

[8] Hardaker EL, Bacon AM, Carlson K, Roshak AK, Foley JJ, Schmidt DB,et al. Regulation of TNF-alpha- and IFN-gamma-induced CXCL10expression: participation of the airway smooth muscle in thepulmonary inflammatory response in chronic obstructivepulmonary disease. FASEB J 2004;18:191–3.

[9] Rotondi M, Rosati A, Buonamano A, Lasagni L, Lazzeri E, Pradella F,et al. High pretransplant serum levels of CXCL10/IP-10 are related toincreased risk of renal allograft failure. Am J Transplant 2004;4:1466–74.

[10] Angiolillo AL, Sgadari C, Taub DD, Liao F, Farber JM, Maheshwari S,et al. Human interferon-inducible protein 10 is a potent inhibitor ofangiogenesis in vivo. J Exp Med 1995;182:155–62.

[11] Gotsch F, Romero R, Friel L, Kusanovic JP, Espinoza J, Erez O, et al.CXCL10/IP-10: a missing link between inflammation and anti-angiogenesis in preeclampsia? J Matern Fetal Neonatal Med2007;20:777–92.

[12] Taub DD, Lloyd AR, Conlon K, Wang JM, Ortaldo JR, Harada A, et al.Recombinant human interferon-inducible protein 10 is achemoattractant for human monocytes and T lymphocytes andpromotes T cell adhesion to endothelial cells. J Exp Med 1993;177:1809–14.

[13] de Groot CJM, van der Mast BJ, Visser W, De Kuiper B, Weimar W,Van Besouw NM. Preeclampsia is associated with increasedcytotoxic T-cell capacity to paternal antigens. Am J Obstet Gynecol2010;203. 496.e1–6.

[14] Maynard SE, Min JY, Merchan J, Lim KH, Li J, Mondal S, et al. Excessplacental soluble fms-like tyrosine kinase 1 (sFlt1) may contribute toendothelial dysfunction, hypertension, and proteinuria inpreeclampsia. J Clin Invest 2003;111:649–58.

[15] Romero R, Nien JK, Espinoza J, Todem D, Fu W, Chung H, et al. Alongitudinal study of angiogenic (placental growth factor) and anti-angiogenic (soluble endoglin and soluble VEGF receptor-1) factors innormal pregnancy and patients destined to develop preeclampsiaand deliver a small-for-gestational-age neonate. J Matern FetalNeonatal Med 2008;21(1):9–23.

[16] Levine RJ, Maynard SE, Qian C, Lim KH, England LJ, Yu KF, et al.Circulating angiogenic factors and the risk of preeclampsia. N Engl JMed 2004;350:672–83.