Neutrophil function in women with pre-eclampsia.

OBJECTIVE: To investigate the function of neutrophils in normal pregnancy and pre-eclampsia. DESIGN: Baseline levels and activated responses of peripheral blood neutrophils were measured in response to the physiological agonists, n-formyl-met-leu-phe (fMLP) and zymosan activated serum. SAMPLE: Neutrophils of 16 pre-eclamptic, 17 normal pregnant (third trimester) and 15 nonpregnant age-matched control women were calculated. SETTING: Antenatal Clinic, City Hospital, Nottingham. METHODS: Neutrophil superoxide anion production was determined by lucigenin-enhanced chemiluminescence; the release of secondary granule lactoferrin by ELISA; and the expression of cell surface adhesion molecules CD11b, CD18 and L-selectin (CD62L) by flow cytometric analysis. RESULTS: Superoxide anion generation was reduced in the pregnant group compared with nonpregnant controls [fMLP by 51% (P = 0.03) and zymosan activated serum by 56% (P = 0.01)] but pre-eclamptic measurements did not show a similar reduction. There were no differences between the three study groups in the plasma levels of lactoferrin, or in the stimulated expression and release of CD11b and CD18, or lactoferrin and L-selectin. The base-line levels for the production of superoxide anions; the expression of CD11b or CD18 or L-selectin; and the release of lactoferrin showed no significant differences. CONCLUSIONS: Circulating neutrophils in pregnancy and pre-eclampsia are neither activated nor primed in vivo, however the release of reactive oxygen species is diminished in normal pregnancy. In comparison, an elevation of reactive oxygen generation in pre-eclampsia may highlight a role for neutrophils in the oxidative stress and pathophysiology of this disease.

Endothelial monocyte-activating polypeptide-2 is increased in pregnancy but is not further increased in preeclampsia.

OBJECTIVES: Endothelial monocyte-activating polypeptide-2 (EMAP-2) is a novel protein that demonstrates potent proinflammatory activity in vivo and in vitro. The purpose of this study was to investigate the expression of EMAP-2 in normal pregnancy and in pregnancies complicated with preeclampsia and to test the hypothesis that EMAP-2 is a causative agent of the endothelial activation of preeclampsia. METHODS: Expression of EMAP-2 in the placenta was investigated using reverse transcriptase-polymerase chain reaction to detect mRNA, and immunohistochemistry with an EMAP-2-specific polyclonal antiserum was carried out to detect protein. Levels of circulating EMAP-2 protein were measured in the blood of nonpregnant, normal pregnant, and preeclamptic individuals using a specific enzyme-linked immunoabsorbent assay and the molecular forms present assessed by Western blotting. RESULTS: EMAP-2 is transcribed and translated by the placenta troughout pregnancy and in preeclampsia and can be detected in the plasma of nonpregnant and pregnant individuals. Levels of circulating EMAP-2 antigen are raised in pregnancy compared with nonpregnant controls; however, levels in patients with preeclampsia are identical to those in normal pregnant individuals. CONCLUSION: While circulating levels of EMAP-2 are increased in pregnancy, there is no evidence that EMAP-2 is involved directly in the pathogenesis of preeclampsia.

The role of the endogenous anti-inflammatory compound gravidin in pre-eclampsia.

OBJECTIVES: The study's aims were to investigate the levels of gravidin, an endogenous phospholipase A2 inhibitor, in pregnancy and pre-eclampsia and to establish its effects on neutrophil function. STUDY DESIGN: Serum samples were collected from 9 nonpregnant, 15 preeclamptic, and 10 healthy pregnant women and assayed for free gravidin by enzyme-linked immunosorbent assay. Neutrophil phospholipase A2 and respiratory burst activities were determined in the presence of isolated free gravidin by cellular arachidonic acid release and superoxide anion production. RESULTS: Levels of free gravidin were higher in the healthy pregnant (36.1 +/- 5.5 ng/mL) and preeclamptic (17.8 +/- 2.8 ng/mL) groups than in the nonpregnant control group (3.9 +/- 0.5 ng/mL) and were significantly different between pregnancy groups (P <.01, Mann-Whitney U test). Free gravidin caused a concentration dependent decrease in N-formyl-methionyl-leucyl-phenylalanine-stimulated neutrophil arachidonic acid release (inhibitory concentration of 50% 25 nmol/L) and superoxide anion generation (inhibitory concentration of 50% 32 nmol/L). CONCLUSIONS: Circulating levels of free gravidin are reduced in pre-eclampsia compared with normal pregnancy. This may encourage an increase in the respiratory burst of neutrophils in pre-eclampsia and could contribute to the oxidative stress and vascular damage that characterize this disease.

Activation of endothelial cells by plasma from women with preeclampsia: differential effects on four endothelial cell types.

OBJECTIVE: To compare the ability of plasma from a population of women with preeclampsia and a population of plasma from women with normal pregnancies to activate four different endothelial cell types. METHODS: The secretion of nitrite and 6-keto prostaglandin F1 alpha by four endothelial cell types (isolated from the microvasculature of human decidua and skin, as well as a human umbilical vein endothelial cell line and a bovine coronary microvascular cell line) was assessed following a 24-hour incubation with plasma samples from the two groups. RESULTS: Nitrite production (an indicator of nitric oxide release) was detectable in only the decidual endothelial cells and the bovine microvascular endothelial cells (B-88), whereas 6-keto prostaglandin F1 alpha (stable metabolite of prostacyclin) was detectable in all cells. Only in the B-88 cells was there a greater production of nitrite or 6-keto prostaglandin F1 alpha in response to incubation with plasma from the preeclamptic patients when compared to plasma from the normotensive controls. CONCLUSION: The different responses of various endothelial cell types to the activating effects of plasma from preeclamptic women indicate that another important caveat to be considered when bioassaying for the circulating factor(s) of preeclampsia is the choice of endothelial cell to be studied.

Lack of effect of ET antibody or SB 209670 on endotoxin-induced renal failure.

Elevated plasma levels of endothelin-1 (ET-1) in patients with septic shock have implicated ET-1 in the vascular hypoperfusion and organ dysfunction, including renal failure, that occur in this condition. Administration of endotoxin to rats induces renal dysfunction characterized by increased blood urea nitrogen (BUN) and plasma creatinine (Cr) levels. Here, we have found that neither SB 209670 (an ETA and ETB receptor antagonist) nor an affinity-purified ET antibody blocked the endotoxin-induced changes in renal function, suggesting that ET-1 is not involved in the pathophysiology of endotoxin-induced renal dysfunction.

Antagonism of renal and systemic responses to endothelin-1 infusion with PD 145065.

The antagonism by PD 145065 (Ac-D-Bhg-L-Leu-L-Asp-L-Ile-L-Ile-L-Trp; D-Bhg = 5H-dibenzyl[a,d]cycloheptene-10,11-dihydroglycine), a non-selective endothelin ETA and ETB receptor antagonist, of the pressor and renal haemodynamic responses to an infusion of endothelin-1 was investigated in the anaesthetised rat. Infusion of endothelin-1 at 2 or 4 pmol/min/rat for 2 h produced a significant increase in mean arterial blood pressure and renal vascular resistance. Glomerular filtration rate was reduced by both infusion rates of endothelin-1 (-34% for the lower rate and -72% for the higher rate), but only the high rate significantly reduced renal blood flow (-64%). These effects of endothelin-1 were completely blocked by infusion of PD 145065 (0.05 mg/min/rat), demonstrating the efficacy of this antagonist in preventing the actions of endothelin-1 in vivo. These results lend support to an involvement of endothelin ETB receptors in the renal effects of endothelin-1.

Evidence from receptor antagonists of an important role for ETB receptor-mediated vasoconstrictor effects of endothelin-1 in the rat kidney.

1. To characterize the receptor subtype(s) mediating the renal vasoconstrictor effects of the endothelin (ET) and sarafotoxin (SX) peptides in the isolated perfused kidney of the rat, we have examined the effects of endothelin-1 (ET-1), sarafotoxin 6b (SX6b) and sarafotoxin 6c (SX6c) as agonists, BQ-123 and FR 139317 as selective ETA receptor antagonists, and PD 145065 as a non-selective (ETA and ETB) receptor antagonist. We have also compared in the anaesthetized rat the systemic pressor and renal vasoconstrictor effects of ET-1 and SX6c alone or after pretreatment with PD 145065. 2. In the isolated perfused kidney, ET-1, SX6b and SX6c all gave similar concentration-dependent increases in perfusion pressure. The ETA receptor selective antagonists, BQ-123 and FR 139317, both partially blocked the increase in perfusion pressure induced by ET-1. In contrast, PD 145065 completely blocked the increase in perfusion pressure caused by ET-1. 3. Indomethacin (10 microM) had no effect on the ET-1-induced increases in perfusion pressure but significantly reduced the vasoconstriction induced by low concentrations of SX6c, without affecting responses to high concentrations. In the anaesthetized rat, indomethacin (5 mg kg-1) did not modify the systemic pressor or renal vasoconstrictor effects of ET-1 or SX6c. 4. In anaesthetized rats, bolus intravenous injections of ET-1 or SX6c (0.1, 0.25, 0.5 or 1.0 nmol kg-1) produced initial transient depressor responses followed by sustained and dose-dependent increases in mean arterial pressure (MAP). Both peptides caused an equipotent fall in renal blood flow (RBF).PD 145065 (5 mg kg-1) partially antagonized the systemic pressor effects of ET-1 and SX6c but completely blocked the fall in RBF and rise in renal vascular resistance (RVR) induced by ET-1 and SX6c. PD 145065 also antagonized the transient depressor effect following the bolus administration of either ET-1 or SX6c.5. These results indicate that ET/SX induced renal vasoconstriction is mediated via ETA and ETB-like receptors with ETB receptors having a predominant role in vivo. This may be of therapeutic relevance for an ETA receptor-selective antagonist may offer only limited protection against the deleterious renal effects of endogenous ETs.

Vasoconstriction in the rat kidney induced by endothelin-1 is blocked by PD 145065.

We have previously shown that the receptors mediating the renal and systemic vasoconstrictor effects of endothelin-1 (ET-1) are of two distinct endothelin receptor subtypes. Here, we evaluate the effect of PD 145065, a nonselective endothelin receptor antagonist, on the renal vasoconstrictor effects of ET-1 in the rat kidney. ET-1 induced concentration-dependent increases in perfusion pressure in the isolated perfused kidney of the rat. The ETA receptor-selective antagonists BQ-123 (10 microM) and FR 139317 (10 microM) lowered the ET-1-induced rise in perfusion pressure by 57% and 61%, respectively, at 3 x 10(-10) M ET-1. By comparison, the ET-1-induced vasoconstriction was fully antagonized (96% inhibition) by PD 145065 (10 microM). In the anesthetized rat, ET-1 produced a dose-dependent increase in mean arterial pressure, which was attenuated by PD 145065. The initial depressor response to ET-1 was completely blocked by PD 145065, as were the reduction in renal blood flow and increase in renal vascular resistance induced by ET-1. These results suggest that both the ETA and a non-ETA receptor subtype play an important role in mediating the vasoconstrictor effects of ET-1 in the rat kidney.