CARDIOVASCULAR JOURNAL OF AFRICA • Vol 24, No 5, June 2013
AFRICA
175
were centrifuged within two hours at 3 500 rpm for 10 min
at 4°C. Serum aliquots were then carefully transferred into
new tubes and stored at –70°C until analysis. Enzyme-linked
immunoassays for PlGF (1:2), TGF-
β
1
(1:40), sFlt1 (1:5)
and sEng (1:5) were performed in triplicate according to the
manufacturer’s instructions (R&D Systems, Minneapolis, MN).
Fifty-two of the blood samples were obtained from
pre-eclamptics and 58 from normotensive pregnant women.
These groups were further subdivided into HIV-negative and
HIV-positive sub-groups.
Inclusion criteria for pre-eclampsia were persistent systolic
blood pressure 140 mmHg and diastolic blood pressure 90
mmHg, taken at least six hours apart, after 20 weeks’ gestation
in a previously normotensive patient. Proteinuria was defined
as urine protein concentration of
≥
300 mg/dl or 1
+
on a urine
dipstick in at least two random specimens collected at least four
hours apart.
Exclusion criteria for all groups was chorio-amnionitis,
chronic hypertension, eclampsia and abruptio placentae;
intrauterine death, pre-gestational diabetes, gestational diabetes
and chronic renal disease; systemic lupus erythematous, sickle
cell disease and anti-phospholipid antibody syndrome; thyroid
disease, cardiac disease and active asthma requiring medication
during pregnancy and pre-existing seizure disorders.
Statistical analysis
Inter-group analysis was conducted using the non-parametric
Kruskal-Wallis test. Descriptive statistics were utilised and
outcome variables are presented as median (interquartile range).
Where differences were found in the Kruskal-Wallis test, Dunn’s
post hoc
test was used for multiple comparisons. A probability
level of
p
<
0.05 was considered statistically significant. All
statistical analyses were conducted using GraphPad Prism
®
version 5.01.
Results
Clinical characteristics for the pre-eclamptic and normotensive
participants (
n
=
110) were divided into HIV-positive (
n
=
56)
and HIV-negative groups (
n
=
54), respectively, namely, (1)
HIV-negative normotensive (N–): BP
≤
120/80 mmHg (
n
=
27);
(2) HIV-positive normotensive (N
+
): BP
≤
120/80 mmHg; CD
4
<
200 cells/
µ
l (
n
=
31); (3) HIV-negative pre-eclamptic (P–): BP
140/90 mmHg (
n
=
27) and (4) HIV-positive pre-eclamptic (P
+
):
BP 140/90 mmHg; CD
4
<
200 cells/
µ
l (
n
=
25) (Table 1).
A significant difference was detected for maternal and
gestational age, parity, maternal and placental weight, and
systolic and diastolic blood pressure (
p
<
0.05) between the four
groups (Kruskal-Wallis test, Table 1). Mean maternal age ranged
between 23 and 30 years while the mean gestational age ranged
between 37 and 39 weeks (Table 1).
For maternal weight, the Kruskal-Wallis test showed an overall
significance (
p
<
0.05). The Dunn’s multiple comparison tests
identified a significant difference between only the HIV-positive
pre-eclamptic and the HIV-negative normotensive pregnant
women (
p
=
0.0321;Table 1). However, for placental weight (Table
1), a significant difference was evident between the HIV-positive
pre-eclamptic and HIV-negative normotensive pregnant women
(
p
<
0.0001), the HIV-negative pre-eclamptic and HIV-negative
normotensive pregnant women (
p
<
0.0001) and the HIV-positive
normotensive and HIV-negative normotensive pregnant women
(
p
<
0.0001; Table 1).
For systolic blood pressure (Table 1), a significant difference
was evident between the HIV-positive pre-eclamptic and
HIV-negative normotensive pregnant women (
p
<
0.0001), the
HIV-positive pre-eclamptic and the HIV-positive normotensive
pregnant women (
p
<
0.0001), the HIV-negative pre-eclamptic
and the HIV-negative normotensive pregnant women (
p
<
0.0001) and the HIV-negative pre-eclamptic and HIV-positive
normotensive pregnant women (
p
<
0.0001). A similar pattern
was observed for diastolic blood pressure, as indicated in Table 1.
Pro-angiogenic and anti-angiogenic factors
Serum concentrations for all evaluated pro-angiogenic (PlGF
and TGF-
β
1
) and anti-angiogenic (sFlt1 and sEng) factors
varied (Table 2, Figs 1a–d and 2a–c). A significant difference
was observed for sFlt1, sEng and PlGF (
p
<
0.05) between the
groups (Figs 1a–d). For sFlt1, the Kruskal-Wallis test showed an
overall significance (
p
<
0.05). The Dunn’s multiple comparison
test revealed a significant difference between HIV-negative
pre-eclamptic and HIV-negative normotensive pregnant women
(
p
=
0.0061), and HIV-negative pre-eclamptic and HIV-positive
normotensive pregnant women (
p
=
0.0061).
A significant difference for sEng (Table 2, Fig. 1) was
evident between HIV-positive pre-eclamptic and HIV-positive
normotensive pregnant women (
p
=
0.0017), and HIV-negative
pre-eclamptic and HIV-positive normotensive pregnant women
(
p
=
0.0017). Likewise for PlGF, a significant difference was
found between the HIV-negative pre-eclamptic and HIV-negative
normotensive pregnant women (
p
=
0.021), and the HIV-negative
TABLE 1. DEMOGRAPHICAND CLINICAL PROFILE OF PATIENTS RECRUITED FOR IMMUNOASSAYS
Normotensive
pregnant women (N–)
HIV normotensive
pregnant women (N+)
Pre-eclamptic
pregnant women (P–)
HIV pre-eclamptic
pregnant women (P+)
p
-value
Number
27
31
27
25
Age (years)
24 (21–26)
27 (24–30)
25 (20–32)
32 (25.5–34)
0.0009*
Gestational age (weeks)
38 (38–39)
39 (38–40)
38 (37–40)
38 (36–38)
0.0026*
Parity
0 (0–1)*
1 (1–2)
1 (0–1)
1 (0.5–3)
0.0174*
Maternal weight (kg)
66 (59–74)
74 (65–82)
75 (65–96)
82 (64–106)
0.0321*
Birth weight (kg)
3.2 (3–3.4)
3.4 (3–3.7)
3.2 (2.6–3.8)
2.9 (2.7–3.4)
ns
Placental weight (g)
360 (300–400)
470 (380–500)
480 (430–510)
480 (380–515)
< 0.0001*
Systolic BP (mmHg)
110 (108–115)
112 (107–120)
154 (150–162)
150 (145.5–159)
< 0.0001*
Diastolic BP (mmHg)
70 (67–73)
70 (67–74)
94 (90–104)
9 6 (87–99.5)
< 0.0001*
Medians (range) are presented; Kruskal-Wallis test and the
post hoc
Dunns multiple comparison test was used for statistical analysis,
n
=
110. *
p
<
0.05
