Thajasvarie Naicker

Quantitative analysis of trophoblast invasion in preeclampsia

Background.  The process of physiological conversion of spiral arteries is dependent on the invasion of the interstitium and spiral arteries of the uterine wall by invasive extravillous trophoblast thereby creating a high flow–low resistance vessel. Quantitative data on restriction of trophoblast invasion and failure of spiral artery transformation are limited in preeclampsia.

Pilot study of comparative placental morphometry in pre-eclamptic and normotensive pregnancies suggests possible maladaptations of the fetal component of the placenta

OBJECTIVE Adequate maternal, intervillous and fetal blood flow are all necessary for fetal well-being. Compromise to any part of this exchange would be detrimental to pregnancy outcome. Pre-eclampsia is associated with reduced maternal spiral artery flow, resulting in reduced placental perfusion. This in turn creates an ischaemic environment, which may predispose to morphological changes in placental villi. This pilot study sought to assess whether there were morphological alterations in the fetal component of the placenta which could be detrimental to exchange and therefore pregnancy outcome. STUDY DESIGN This study utilized morphometric image analysis to examine some features of the fetal component of the placenta in normotensive (NT) and pre-eclamptic (PE) groups. The features examined included: density of placental villi (expressed as percentage of field area occupied by placental tissue); stem vessel carrying capacity (expressed as percentage of stem villus area occupied by vessel lumina); the thickness of the stem arterial walls relative to artery size (expressed as percentage of artery area occupied by arterial wall) and the extent of fibrosis associated with villi (expressed as percentage of field area occupied by fibrosis). RESULTS There were significant differences between NT and PE placentae in density of placental villus arrangement NT: 51.89 ± 6.19, PE: 64.78 ± 6.93 (P<0.001); carrying capacity of stem villi NT: 17.20 ± 11.78, PE: 8.67 ± 8.51 (P<0.001); relative thickness of stem villi arterial walls NT: 74.08 ± 12.92, PE: 86.85 ± 10.55 (P<0.001); and extent of fibrosis NT: 0.727 ± 0.310, PE: 1.582 ± 0.707 (P<0.001). CONCLUSION These significant differences between normotensive and pre-eclamptic placentae suggest possible fetal maladaptations in response to the intervillous ischaemia, compounding the existing maternal compromise to materno-fetal exchange. Further investigations would, however, be necessary in order to make more conclusive deductions.

Maternal imbalance between pro-angiogenic and anti-angiogenic factors in HIV-infected women with pre-eclampsia.

Abstract Angiogenic imbalance contributes to the development of preeclampsia. We evaluated the protein expression of the proangiogenic placental growth factor (PlGF) and transforming growth factor beta 1 (TGF-β1) compared with the anti-angiogenic soluble fms-like tyrosine kinase receptor (sFlt1) and soluble endoglin (sEng) in HIV-infected normotensive and pre-eclamptic pregnancies. Blood was obtained from 110 pregnant women, enrolled in four groups, namely, HIV-negative normotensives (27); HIV-positive normotensives (31); HIV-negative pre-eclamptics (27) and HIV-positive pre-eclamptics (25), and was used to measure PlGF, TGF-β1, sFlt1 and sEng levels. Increased sFlt1 and sEng levels were associated with the pre-eclamptics (HIV negative and positive) compared with their counterparts. Decreased PlGF levels were observed between the HIV-negative pre-eclamptics versus HIV-negative normotensives, but levels differed significantly (p = 0.02) among the normotensives (HIV negative and positive). TGF-β1 remained unchanged across all groups. Higher sEng/TGF-β1 ratios were associated with the pre-eclamptics (HIV negative and positive) compared with their counterparts. This study demonstrated increased sFlt1 and sEng levels in pre-eclamptic compared with normotensive pregnancies, irrespective of the HIV status.

The Role of Apoptosis on Trophoblast Cell Invasion in the Placental Bed of Normotensive and Preeclamptic Pregnancies

Background. Placental development depends on careful coordination of trophoblast proliferation and apoptosis; however, the synchrony of its effect on trophoblast invasion is unknown. Objective. To examine the relationship between trophoblast apoptosis and proliferation in placental bed tissue of preeclamptic and normotensive pregnancies. Methods. Serial sections from archived placental bed biopsies of 12 normotensive (group 1) and 12 preeclamptic (group 2) were immunolabeled with a rabbit anti-Ki67 antibody, a mouse anti-cytokeratin 18 and its neo-epitope, and a monoclonal cytodeath M30 antibody. Results. The immunoexpression of Ki67 for all trophoblast cell subpopulations within the myometrium was non-reactive in both study groups. Smooth muscle cells of the microvasculature reflected a moderate degree of proliferation in both groups. Morphometric image analysis of the wall of the spiral artery revealed a mean area of 31,1729 ± 51,180 µm2 compared to 35,795 ± 8045 µm2 in groups 1 and 2, respectively. An elevation of intramural trophoblast was evident within the spiral artery of group 1 (13%). Comparative analyses of M30 distribution on corresponding serial sections were 0.06% versus 0% in groups 1 and 2, respectively. The mean field area percentage of interstitial trophoblast invasion was 10.79% versus 2.87% with corresponding areas of apoptosis been 0.8 % versus 1.9 % in groups 1 and 2, respectively. Conclusions. This study demonstrates an increased trophoblast apoptosis in placental bed of preeclamptic compared to normotensive pregnancies with concurrent absence of proliferation at term.

Soluble fms-like tyrosine kinase-1 and soluble endoglin in HIV-associated preeclampsia

OBJECTIVE Preeclampsia is characterized by endothelial dysfunction combined with increased concentrations of sFlt1, which antagonizes the biological effects of VEGF and PlGF, and of sEng, which antagonizes TGFβ1. This angiogenic imbalance may have a role in its etiology. This study evaluated the expression of VEGF, PlGF, sFlt1 and sEng amongst third trimester pregnancies in women with HIV-associated pre-eclampsia. METHOD Serum and placental tissue were obtained from 76 pregnancies in women who were normotensive and HIV negative (N-) or positive (N+), and in women who were pre-eclamptic and HIV negative (P-) or positive (P+). The serum and placental samples were quantitatively evaluated using ELISAs and RT-PCR respectively. RESULTS Placental sFlt1 expression differed significantly between the N- and P- groups (p=0.001). Similarly, sEng expression differed between the N- and P- groups (p=0.001). No significant effect was shown between HIV status and pregnancy. Serum sFlt1 (p=0.02) and sEng (p=0.001) were up-regulated in the P- compared to the N- groups. Similarly, no significant effect was shown between HIV status and pregnancy. Both VEGF and PlGF did not differ significantly between groups. Notably, sEng expression was elevated in both placenta and serum, whilst placental sFlt1 differed from serum. A weak but significant correlation between serum and placental concentration for sFlt1, sEng and PlGF (r=0.26, p=0.031; r=0.42, p<0.001 and r=-0.3, p=0.014) was observed. CONCLUSIONS This novel study demonstrates an up-regulation of serum sFlt1 and sEng in preeclamptic compared to normotensive groups irrespective of the HIV status of the pregnancy. This implicates a contributory role of sFlt1 and sEng in preeclampsia development. The serum reduction of sFlt1 and sEng within the HIV positive compared to HIV negative cohorts may imply a neutralization of the immune hyperreactivity of preeclampsia.

Relationship between histopathological changes in post partum renal biopsies and renal function tests of African women with early onset pre-eclampsia

Background. To improve the diagnostic accuracy of concurrent renal disease in hypertension of pregnancy, biopsy evaluation is essential. In addition, establishing underlying renal disease is important for prognosis on future pregnancies. We therefore designed a study to determine the diagnostic yield of postpartum renal biopsy and the nature and frequency of complications associated with this procedure. Also, to determine relationships, if any, between renal function tests and ultrastructural and histopathological findings.

Soluble fms-like tyrosine kinase-1 in HIV infected pre-eclamptic South African Black women

INTRODUCTION Hypertensive disorders of pregnancy are the commonest direct cause of maternal deaths in South Africa, 83% being attributed to pre-eclampsia. Elevated placental sFlt-1 levels are linked with angiogenic disruption and subsequent pre-eclampsia development. The impact of HIV infection on pre-eclampsia is controversial. Its effect on angiogenic imbalance in both normotensive and pre-eclamptic pregnancies remains unknown. METHODS We examined the immunolocalisation of both membrane bound and soluble forms of Flt-1, within placentae of HIV negative and positive normotensive and pre-eclamptic pregnancies at term using immunohistochemistry and immuno-electron microscopy. RESULTS Strong Flt-1 and sFlt-1 immunoreactivity was observed within endothelial, syncytio and cytotrophoblast cells. Subcellularly, gold particles were localised predominantly within the endoplasmic reticulum and mitochondria and occurring free within the cytoplasm. There was no significant effect of HIV on Flt-1 and sFlt-1 immunoexpression in both exchange and stem villi. A significant effect of type of pregnancy (normotensive vs pre-eclamptic) on Flt-1 and sFlt-1 immunoexpression (p = 0.003) within exchange rather than stem villi, indicated that the pre-eclamptic had elevated Flt-1 and sFlt-1 expressions compared to the normotensive pregnant women. There was no interaction between HIV and pregnancy type (normotensive vs pre-eclampsia) for Flt-1 and sFlt-1 expressions in both exchange and stem villi. A weak correlation of Flt-1 and sFlt-1 intensity between the exchange and stem villi was noted. DISCUSSION Elevated immunoexpression of Flt-1 and sFlt-1 within trophoblasts suggests an autocrine mode of action on trophoblast invasion and differentiation thereby contributing to abnormal placentation with consequential endothelial dysfunction in pre-eclampsia. CONCLUSION Irrespective of the HIV status, placental Flt-1 and sFlt-1 expressions remain elevated in pre-eclampsia compared to normotensive pregnancies.

Placental leptin in HIV-associated preeclampsia

OBJECTIVE HIV-associated preeclampsia reflects a combination of opposing influences on the immune status. The adipocyte hormone leptin has been implicated in the pathophysiology of preeclampsia and in enhancing immunity. This study is the first, to our knowledge, to determine whether leptin levels in the placenta differ between HIV-associated normotensive and preeclamptic pregnancies. The study also compares leptin levels between the exchange and conducting areas of the placenta. STUDY DESIGN Pregnant women were recruited antenatally and grouped as follows: normotensive HIV uninfected (n=30), normotensive HIV infected (n=60), preeclamptic HIV uninfected (n=30) and preeclamptic HIV infected (n=60). Anthropometric data were collected and placental leptin was analysed by immunohistochemistry and ELISA. RESULTS Leptin levels were similar in the central and peripheral regions of the placenta. Leptin immunoreactivity was observed amongst the different trophoblast cell populations. Both ELISA and immunohistochemistry of the placental exchange villi indicated that leptin levels were higher in preeclampsia compared to normotensive pregnancies (p<0.001). HIV status had no effect on leptin levels but levels were higher in participants on highly active antiretroviral treatment (HAART) compared to those on prophylaxis for prevention of mother to child transmission (PMTCT) with normotensive (p=0.006) and preeclamptic (p=0.002) pregnancies. The area of immunostaining was greater in the exchange compared to the conducting villi in HIV infected and uninfected preeclampsia. CONCLUSIONS This novel study establishes an elevation of leptin in preeclamptic placentae, irrespective of HIV status. Leptin elevation was not focal in that it occurred in both central and peripheral regions of the preeclamptic placenta. This suggests a role of leptin in the pathophysiology of preeclampsia.

Elemental analysis of serum and hair from pre-eclamptic South African women

Pre-eclampsia is a hypertensive disorder that is associated with adverse maternal and perinatal outcomes. It has been proposed that specific trace and macro elements associated with antioxidant activities may also play a contributory role in aetiology of pre-eclampsia. The aim of this study was to measure the concentrations of thirteen different elements in hair and serum samples from women with a diagnosis of pre-eclampsia and compare them with normotensive controls. Venous blood and pubic hair samples were collected from forty-three pre-eclamptic and twenty-three normotensive pregnant women. In each sample, the concentration of arsenic (As); calcium (Ca); cadmium (Cd); chromium (Cr); cobalt (Co); magnesium (Mg); manganese (Mn); iron (Fe); copper (Cu); lead (Pb); selenium (Se); nickel (Ni); zinc (Zn) were measured using inductively coupled plasma-optical emission spectrometry. Cobalt concentration in hair was significantly lower in the pre-eclampsia group (1.56±0.74μg/g) compared to the normotensive group (2.89±4.99μg/g) (p=0.02). The concentrations of Zn and Cr were significantly higher in hair samples from the pre-eclamptic group, compared to the normotensive control group (Zn, 395.99±48.60 vs 330.88±29.70μg/g; Cr, 13.31±2.67 vs 11.05±7.62μg/g: p≤0.05). There were no significant differences in the hair levels of other elements between groups. Serum Zn was significantly higher in the pre-eclamptic group (0.16-253.4mg/L) compared to the normotensive group (0.2-48.4mg/L) (p=0.01). Serum Ca, Co, Cu, Mg, Mn and Se levels were found to be significantly lower in the pre-eclamptic group compared to the normotensive group (p<0.05). This study confirms the association between pre-eclampsia and maternal trace as well as macro element levels.

The role of podocytes in the early detection of pre-eclampsia

INTRODUCTION Pre-eclampsia is a significant cause of maternal and neonatal mortality and morbidity in resource constrained countries. Because the exact aetiology is unknown, treatment of preeclampsia is empiric. Therefore, researchers have been investigating biomarkers for early detection of the syndrome to take steps to prevent complications. The kidney is reported to be affected by the preeclamptic process before clinical signs appear. Podocytes have been suggested as possible markers for this syndrome. However there is debate as to which is the best way to measure the amount of podocyturia. OBJECTIVE To determine the best method to estimate podocyturia as a biomarker. METHODS Midstream urine specimens were collected from 18 normotensive healthy primigravidae at their first antenatal visit. Urinary podocyte immunolabelling was performed by two techniques viz., culture and cytospin on urine from normotensive and clinically healthy pregnant women. MAIN OUTCOME MEASURED Are the podocyte specific proteins, podocalyxin, podocin, nephrin and synaptopodin able to detect pre-eclampsia prior to the development of clinical signs as measured by two separate techniques. RESULTS The results suggest that the expression of podocyte specific proteins, podocalyxin, podocin, nephrin and synaptopodin, is identifiable and quantifiable from midstream urine in healthy normotensive pregnant women. Cytospin was more efficient in determining the podocyte specific protein expression levels and podocalyxin was the most sensitive marker, with a Kappa coefficient of 0.23. CONCLUSIONS These findings suggest that immuno-expression of podocyturia are best detected by the cytospin method.