Labor and infection. II. Bacterial endotoxin in amniotic fluid and its relationship to the onset of preterm labor
ABSTRACT We have previously reported the detection of endotoxin in the amniotic fluid of patients with gram-negative intraamniotic infection. Endotoxin or lipopolysaccharide is a potent biologic product capable of inducing prostaglandin release from several cell types and, therefore, may be involved in the onset of human parturition in the presence of intraamniotic infection. This article describes a technique for the quantification of endotoxin in amniotic fluid. The method uses a computer-assisted quantification of the turbidimetric reaction between the Limulus amebocyte lysate and endotoxin. Serial dilutions of Escherichia coli endotoxin in culture-negative amniotic fluid were prepared, and the samples were run in the assay. Amniotic fluid was found to enhance the reaction, and a dilution of 1:20 was required for this biologic fluid to behave similarly to pyrogen-free water. The sensitivity of this kinetic turbidimetric technique in the detection of endotoxin in amniotic fluid was 40 pg/ml. This method was applied to the quantification of endotoxin concentration in amniotic fluid in 26 patients with intraamniotic infection and premature rupture of membranes. Patients in active labor had higher concentrations of endotoxin (median = 47,514 pg/ml) than nonlaboring patients (median = 635 pg/ml) (p less than 0.025). Therefore, women with preterm labor had a higher median concentration of endotoxin in amniotic fluid than patients who were not in labor.
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- "Since this translational research study could have clinical implications for new BPD therapy, microarray methodology provided a screening tool for gene expression that could be important for effective and safe therapy. Fortunately, there has been previous work measuring plasma DEX levels in preterm infants being treated for BPD   as well as measurements of endotoxin levels in amniotic fluid . Therefore the present microarray studies used a clinically relevant stimulus dose of LPS (10ng/ml) with and without DEX (10 "
ABSTRACT: Bronchopulmonary dysplasia (BPD) is one of the most common causes of mortality and morbidity in neonatal intensive care units. Persistent inflammation, with an abnormal influx of polymorphonuclear leukocytes (PMNs) followed by monocytes (MONOs), occurs early in the pathogenesis of BPD. Anti-inflammatory therapy with better efficacy and safety than dexamethasone (DEX) is needed. In the present study we determined cell-specific gene expression and cytokine release in response to glucocorticoids versus interleukin-10 (IL-10). Subsequently, we hypothesized that the insensitivity of MONOs to DEX was associated with a failure of the glucocorticoid receptor to translocate to the nucleus. PMNs and MONOs were isolated from umbilical cord blood at birth, and pretreated with PBS vehicle, IL-10 or glucocorticoids prior to endotoxin (LPS)-stimulation for 4 and 18h. Genome-wide gene expressions were determined by microarray and validated by RT-qPCR. Interleukin 8 release in cell culture supernatant was measured by ELISA. To examine the mechanism of monocyte insensitivity to glucocorticoids, nuclear translocation of the glucocorticoid receptor was determined by Western blots. MONOs had 6 times the number of genes changing expression with IL-10 compared to PMNs at 4h. DEX at the therapeutic level for neonates with BPD had no effect on gene expression in MONOs. The order of potency for inhibition of interleukin-8 release from MONOs was IL-10 >betamethasone >dexamethasone and hydrocortisone. Glucocorticoid potency in MONOs was directly related to glucocorticoid receptor translocation to nucleus. Gene expression profiling for IL-10 versus glucocorticoids indicates there may be major differences in therapeutic efficacy for BPD.American Journal of Translational Research 01/2013; 5(1):103-15. · 3.23 Impact Factor
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- "Micro-organisms can be cultured from 10–40% of AFs collected during PTL (Romero et al., 1988a, 1994; Gomez et al., 1995). In addition, LPS (Cox et al., 1988; Romero et al., 1988b) as well as an impressive array of mediators of the inflammatory response, including IL-1β (Hillier et al., 1993; Romero et al., 1992; Gomez et al., 1995), are found in a sizeable proportion of AFs that are collected during PTL. Other cytokines, e.g. "
ABSTRACT: From the finding of micro-organisms or inflammatory mediators, or both, in amniotic fluid (AF), it has been proposed that intrauterine infection is one cause of preterm labour (PTL, intact fetal membranes). This theory, however, remains unproved, i.e. the accumulation of micro-organisms and inflammatory mediators in AF after labour is in progress may be the consequence, not the cause, of labour both at term and preterm. This study was conducted to evaluate this possibility by a comparison of the concentrations of interleukin (IL)-1beta and IL-6 in AFs collected before and during PTL (<34 weeks gestation) with those in AFs collected at term (before labour and from the forebag and upper compartments of the amniotic sac during labour). The concentrations of IL-1beta and IL-6 in AF were also analysed as a function of the duration of labour (term or preterm) before fluid collection. In addition, studies were conducted to define the source of IL-1beta in AF. A total of 666 AFs were evaluated. IL-1beta was not detected (<50 pg/ml) in AFs collected before the onset of labour at any stage of gestation (n = 320), including 170 fluids obtained at term. During labour, IL-1beta was detected (>50 pg/ml) in 58 out of 106 (54.7%), 17 out of 64 (26.6%) and 60 out of 176 (34%) of AF samples obtained during PTL, term labour (upper compartment) and term labour (forebag) respectively. AF sampling, as well as labour and delivery, were completed in <18 h in all term pregnancies. However, labour (with cervical dilation) was in progress for >18 h before AF was collected in 39 out of 106 (37%) PTL pregnancies. The incidence of IL-1beta-positive samples among AFs collected before 18 h of PTL (23 out of 67; 34%) was indistinguishable from that in AFs collected during labour at term. However, in AFs collected after >18 h PTL, the incidence of IL-1beta-positive samples was 35 out of 39 (89.7%) The concentrations of IL-1beta (pg/ml; mean +/- SEM) in AFs collected during PTL (2680 +/- 730; n = 106) were greater than those in AFs collected from the upper compartment and forebag during term labour (436 +/- 244, n = 64; and 468 +/- 119, n = 176) respectively; this difference, however, was attributable to very high concentrations of IL-1beta in AFs in which PTL was in progress for >18 h before AF collection (6021 +/- 1832; n = 39). The concentrations of IL-6 in AF were correlated with those of IL-1beta (P < 0.001). We conclude that IL-1beta and IL-6 accumulate in AF in a similar proportion of pregnancies during the first 18 h of term and preterm labour. Therefore, the accumulation of these cytokines in AF cannot be taken as evidence for a role for infection in the pathogenesis of PTL.Human Reproduction Update 09/1997; 3(5):517-27. DOI:10.1093/humupd/3.5.517 · 8.66 Impact Factor
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ABSTRACT: The main aim of this study was to determine the socioeconomic, demographic and environmental factors which may be associated with the occurrence of pathological microflora of the lower genital tract in early pregnancy. A group of 96 pregnant women was selected at random from the patients of 10 district maternity units in the Lodz region of Poland. Only singleton pregnancies below 24 weeks were qualified for inclusion in the survey. A standard questionnaire covering medical, socio-economic, demographic, constitutional, and environmental items was administered to every subject and checked against medical records. Based on microbiological results, two groups of pregnant women were distinguished: Group I, with normal cervicovaginal flora, predominantly Lactobacillus spp. with coagulase-negative staphylococci and viridans streptococci, and Group II, with abnormal flora. The latter included two subgroups: IIA, intermediate microbial flora, dominated by M. hominis, U. urealyticum, G. vaginalis, gram-negative anaerobic rods, Ch. trachomatis, and few Lactobacillus spp, and IIB, highly abnormal flora, containing similar microbial components as in IIB but without Lactobacillus spp. Based on the results of microbiological culturing, 18 (18.7%) of the 96 women examined were classified to Group I, and 78 (81.2%) to Group II: 32 (33.3%) in group IIA and 46 (47.9%) in IIB. Groups IIA and IIB were combined for further analysis. An excessive risk of abnormal vaginal flora was observed in connection with such socio-economic factors as marital status, unemployment, and smoking, Moreover, the first pregnancy was also found to be a potential risk factor for this pathology. The risk of developing abnormal vaginal flora, although exceeding unity for each of these factors, was not considered statistically significant. Socio-economic and environmental factors may influence the course and outcome of pregnancy. Pregnant women who present with risk factors for abnormal cervicovaginal microflora should be included in comprehensive prenatal surveillance, which enables early detection and treatment of this pathology.Medical science monitor: international medical journal of experimental and clinical research 7(6):1250-5. · 1.22 Impact Factor