Apolipoprotein E alleles in women with pre-eclampsia.

AIMS: To investigate the frequency of three apolipoprotein E (apoE) alleles among women with pre-eclampsia. METHODS: The presence of the three most common apoE alleles (epsilon 2, epsilon 3, epsilon 4) was determined by polymerase chain reaction-restriction fragment length polymorphism in two groups of women: healthy pregnant women (n = 91) and pregnant women with a diagnosis of pre-eclampsia (n = 133). In addition, the frequencies of the alleles in the general population in this area are presented for comparison. RESULTS: The frequency of the apo epsilon 4 allele was 18.4% among women with pre-eclampsia and 18.7% among healthy pregnant women (Fisher's exact test; p = 0.941), which is close to the rate in the general population in this area (19%). None of the apolipoprotein E genotypes was significantly over-represented, and homozygous genotype epsilon 4 was not associated with more severe clinical disease than were the other genotypes. CONCLUSION: The observed profiles of allele and genotype frequencies confirm an equilibrium state between apoE polymorphism and pre-eclampsia and suggest that apoE does not play a major role in the development of pre-eclampsia.

Elevated maternal serum hCG in the second trimester increases prematurity rate and need for neonatal intensive care in primiparous preeclamptic pregnancies.

OBJECTIVE: This study was designed to investigate the association between the serum concentrations of maternal second trimester human chorionic gonadotropin (hCG) and the severity of preeclampsia. METHODS: At Kuopio University Hospital, a total of 487 preeclamptic primiparas had undergone maternal serum screening for Down's syndrome between January 1993 and December 1998. Of these, 37 women had unexplained elevated serum hCG concentrations [> 2.5 multiples of the median (MoM)], whereas the remaining 450 preeclamptic women had normal hCG results. Pregnancy characteristics and outcome measures in these groups were evaluated using logistic regression. RESULTS: Elevated midtrimester hCG concentrations were associated with higher rates of low-birth-weight infants, preterm delivery, and need for neonatal intensive care. The adjusted odds ratios was 2.11 [95% confidence interval (CI): 1.03-4.32], 2.08 (95% CI: 1.10-4.30), and 2.27 (95% CI: 1.14-4.51), respectively. CONCLUSIONS: In primiparous preeclamptic pregnancies, an elevated maternal serum hCG concentration is a marker of early-onset and severe disease with significant maternal and perinatal morbidity. This finding, in turn, reinforces the association between elevated hCG concentrations and placental damage in early pregnancy. Elevated maternal serum hCG levels identify a subgroup of preeclamptic patients who deserve more intensive observation.

The cellular uptake and metabolism of clodronate in RAW 264 macrophages.

PURPOSE: Non-nitrogen-containing bisphosphonates, such as clodronate (dichloromethylene bisphosphonate), appear to act as prodrugs, their active form being the AppCp-type analogues of ATP. To further elucidate this, we examined the cellular uptake of clodronate and intracellular accumulation of the metabolite of clodronate (AppCCl2p) in RAW 264 macrophages, the influence of clodronate metabolism on the intracellular ATP concentration, and the time course of clodronate metabolism and the effects of clodronate on cytokine secretion from macrophages. METHODS: The cellular uptake of clodronate was measured using 14C-labeled clodronate. AppCCl2p was determined in cell extracts by using an ion-pairing HPLC-ESI-MS. The cytokine concentrations in the culture supernatants were measured with time-resolved fluoroimmunoassay. Intracellular ATP concentration was measured with a luminometer using a luciferin-luciferase assay. RESULTS: Of the clodronate internalized by macrophages in vitro, 30-55% is metabolized to AppCCl2p, which accumulates to high intracellular concentrations during the first 12 h of exposure. This accumulation does not affect the ATP levels in the cells. The time course of metabolite appearance in the cells and the inhibition of cytokine secretion were very similar. CONCLUSIONS: These results strongly support the idea that clodronate acts as a prodrug, the active form being its intracellular AppCCl2p metabolite.

Obstetric prognosis in second pregnancy after preeclampsia in first pregnancy.

OBJECTIVE: To assess obstetric outcomes in women in their second pregnancy after preeclampsia in the first pregnancy. METHODS: We utilized population-based birth registry data of Kuopio University Hospital to investigate pregnancy outcome measures in 123 nonpreeclamptic parous women with prior preeclampsia and 21 women with repeat preeclampsia in their second pregnancy. The general obstetric population was used as a reference group in logistic regression. RESULTS: The development of recurrent preeclampsia in 15% of women is associated with adverse neonatal outcomes. A first preeclamptic pregnancy may offer protection against disease recurrence and a history of preeclampsia has no significant effects on birth weight, fetal distress, or prematurity rate. However, they have a higher rate of pregnancy-induced hypertension and abdominal deliveries, and, therefore, a greater proportion of newborns are referred to neonatal units for observation. CONCLUSIONS: Women in whom preeclampsia does not recur have good obstetric outcomes in their second delivery, almost comparable to that in the general obstetric population. A genetic susceptibility to preeclampsia alone has minor effects on pregnancy outcome in a second pregnancy if the disease does not recur.

Contrasting effects of alendronate and clodronate on RAW 264 macrophages: the role of a bisphosphonate metabolite.

Clodronate (dichloromethylidene-bisphosphonate), a halogen-containing bisphosphonate, can inhibit the release of cytokines from RAW 264 macrophages and has anti-inflammatory properties in rheumatoid arthritis, whilst amino-containing bisphosphonates such as alendronate (4-amino-1-hydroxybutylidene-bisphosphonate), have pro-inflammatory properties and can cause an acute phase response. The basis for these pharmacological properties is unclear. Recently, it was demonstrated that clodronate is metabolised by certain cell lines in vitro to an analogue of ATP, whereas amino-bisphosphonates are not. We therefore investigated whether clodronate can also be metabolised by RAW 264 macrophages and whether intracellular accumulation of the metabolite (AppCCl2p) could account for the anti-inflammatory properties of clodronate. The effect of alendronate and AppCCl2p on the release of cytokines (IL-1beta, IL-6, and TNFalpha) from RAW 264 cells was compared, and the effect of the bisphosphonates and AppCCl2p on the DNA binding activities of transcription factors, NF-kappaB and AP-1, was investigated. Pretreatment of RAW 264 macrophages with alendronate augmented the LPS-stimulated release of IL-1beta and increased the binding of NF-kappaB to DNA in an electrophoretic mobility shift assay. Without LPS-induction, alendronate did not affect cytokine release or NF-kappaB binding. Clodronate was metabolised by RAW 264 cells to AppCCl2p. Like clodronate, AppCCl2p inhibited the LPS-induced release of cytokines and NO from RAW 264 macrophages. Both clodronate and its metabolite also inhibited the LPS-stimulated binding of NF-kappaB to DNA. In conclusion, these results suggest that the metabolite of clodronate may be responsible for the anti-inflammatory properties of clodronate, and that the contrasting effects of different bisphosphonates on the release of cytokines could be mediated partly through changes in the DNA binding activity of NF-kappaB.

Streptomyces spores from mouldy houses induce nitric oxide, TNFα and IL-6 secretion from RAW264.7 macrophage cell line without causing subsequent cell death.

The current view is that only bacterial lipopolysaccharide (LPS) and gamma interferon (IFNγ) are able to alone activate macrophages to secrete nitric oxide (NO), probably a causative agent of cell death. Moreover, some cytokines and gram positive pathogens together with IFNγ induce NO-production. Surprisingly, spores of Streptomyces sp., which are mesophilic gram-positive bacteria found in mouldy houses, stimulated RAW264.7 macrophages to produce pro-inflammatory cytokines, tumor necrosis factor alpha (TNFα) and interleukin-6 (IL-6), and induced the expression of inducible NO-synthase (iNOS) with a subsequent NO-production. However, the Streptomyces spores did not kill NO-producing macrophages, as did both LPS and gram negative bacteria Pseudomonas fluorescens, strong inducers of cytokine- and NO-production. These results imply that Streptomyces sp., induced cytokine and NO-secretion, may play a role in the responses evoked by exposure to these microbes. Moreover, factors other than, or in addition to NO, are necessary for cytotoxicity in murine macrophages.

Induced production of nitric oxide, tumor necrosis factor, and interleukin-6 in RAW 264.7 macrophages by streptomycetes from indoor air of moldy houses.

Dampness and mold growth in buildings cause spore generation into indoor air, which is associated with respiratory tract disorders. Specific agents or cellular mechanisms of diseases have not yet been identified. In this study, airborne spores of Streptomyces sp., isolated from moldy houses, stimulated RAW264.7 macrophages, which produced tumor necrosis factor alpha and interleukin-6 and induced the expression of inducible nitric oxide synthase, with subsequent nitric oxide production. Spores of other microorganisms typically found in moldy houses did not markedly increase the production of these inflammatory mediators. The data implied a mechanism by which Streptomyces sp. may lead to respiratory tract disorders in individuals who live in moldy houses.

Different effects of three bisphosphonates on nitric oxide production by RAW 264 macrophage-like cells in vitro.

The macrophage-suppressive properties of three bisphosphonates were evaluated by studying their effect on nitric oxide (NO) production by activated RAW 264 macrophage-like cells. The cells were activated with 10 micrograms/ml of lipopolysaccharide, and NO was determined as nitrite in the cell culture supernatant. The effect of the drugs on inducible NO synthase was determined by Western blot analysis. As free drugs, clodronate and pamidronate inhibited NO secretion in a dose-dependent manner, whereas alendronate had no effect. Liposome encapsulation enhanced the effect of clodronate by a factor of 7, but the potency of pamidronate weakened slightly when encapsulated in liposomes. The inducible NO synthase expression inside the cells was also decreased by liposomal clodronate. In contrast to pamidronate, clodronate could affect the NO secretion when given to the cells simultaneously with lipopolysaccharide, and the inhibitory action was still seen when the drug was added 2 h after lipopolysaccharide induction. The viability of the cells was not affected by free or liposomal clodronate, whereas pamidronate showed considerable cytotoxicity. This study shows the different actions of these three bisphosphonates on NO production by macrophages and suggests that liposomal clodronate is the most promising bisphosphonate as an anti-inflammatory agent, whereas aminobisphosphonates do not possess anti-inflammatory properties.

Postpartum recovery after severe pre-eclampsia and HELLP-syndrome.

Postpartum recovery was examined in 100 pregnancies complicated by severe pre-eclampsia, and in 15 pregnancies in which HELLP-syndrome as present. Albuminuria disappeared and diastolic blood pressure returned to normal (< 90 mmHg) in half of the cases within one week postpartum. Postpartum recovery in the cases with HELLP-syndrome did not differ from that seen in the pre-eclamptic patients. Thrombocytopenia showed spontaneous resolution within three days after delivery. In stepwise discriminant analysis, the incidence of IUGR predicted a slow postpartum recovery, but this did not apply to other factors. Such as diastolic blood pressure before delivery, duration of subjective symptoms, duration of pregnancy or severity of albuminuria. Elevated diastolic pressure and/or albuminuria were diagnosed two months postpartum in one third of the patients. If the development of eclampsia is prevented by correctly timing the induction of delivery, relatively good short-term postpartum recovery is probable after severe pre-eclampsia and HELLP-syndrome.

The effect of free gallium and gallium in liposomes on cytokine and nitric oxide secretion from macrophage-like cells in vitro.

The aim of this study was to evaluate the effect of gallium nitrate, gallium-nitrilotriacetate (NTA) complex, and liposomal gallium-NTA on IL-6, TNF alpha, and nitric oxide (NO) release from activated macrophages. In addition, the expression of the inducible nitric oxide synthase (iNOS) was determined. Gallium inhibited dose-dependently the secretion of IL-6, TNF alpha, and NO from the LPS-induced macrophage-like RAW 264 cells. Encapsulation of gallium in negatively charged DSPG-liposomes increased its potency 10-50 times and 7-11 times compared to free gallium nitrate and gallium-NTA, respectively. Neither non-loaded liposomes nor NTA alone inhibited cytokine or NO secretion, demonstrating that the observed effects originated from gallium. Liposomal gallium-NTA inhibited the expression of iNOS by the macrophages, while other formulations of gallium had no effect. Thus, gallium, when delivered properly, suppresses macrophage functions by inhibiting the release of inflammatory mediators from the cells.