The effect of endometriosis, its stage and activity, and of autoantibodies on in vitro fertilization and embryo transfer success rates.

OBJECTIVES: To analyze IVF cycle parameters, including pregnancy rates (PR), in women with and without endometriosis and to evaluate the effect of the stage and activity of endometriosis and of autoantibodies. DESIGN: A retrospective analysis of 237 consecutive IVF cycles (193 patients), 119 in women with and 118 without endometriosis. The endometriosis group was further subdivided according to the stage and activity of the disease and autoantibody positivity. SETTING: Hospital-based and freestanding IVF programs with the same IVF team. PATIENTS: One hundred ninety-three women of reproductive age undergoing IVF; 84 had prior diagnosis of endometriosis, and 109 had other indications for IVF. Within the endometriosis group, 40 did and 44 did not have evidence of active disease. Autoantibodies were measured in 50 patients. INTERVENTIONS: The IVF protocol was standard with GnRH agonist administered from the midluteal phase of the preceding cycle. Variables included the method of ET and the use of corticosteroids. MAIN OUTCOME MEASURES: Number of follicles produced, number of eggs retrieved, fertilization rates, number of embryos transferred, and PR per transfer. RESULTS: There was no difference between groups in the response to stimulation, number of oocytes retrieved, number fertilized, and number cleaved. The overall PR was 27% per transfer; it was similar in women with and without endometriosis (29% and 25%, respectively). There was also no difference in PR according to the stage or activity of the disease. However, PR in autoantibody-positive and -negative patients were significantly different (22.9% and 45.7%, respectively). Among autoantibody-positive patients treated with corticosteroids, 8 of 10 conceived. CONCLUSIONS: This study confirms previous reports that IVF success rates are comparable in women with and without endometriosis regardless of the activity and stage of the disease. However, our study also indicates that autoantibodies may affect adversely implantation of embryos and that this effect can be overcome by administration of corticosteroids.

Activation of the alternative complement pathway by red blood cells from patients with sickle cell disease.

Increased activation of the alternative pathway of complement in serum from sickle cell disease (SCD) patients has been reported. We now show that this selective activation is not an artifact of clotting by measuring increased plasma concentrations of Bb in sickle cell patients compared to controls. Furthermore, we show that red blood cells (RBC) from SCD patients activate the alternative complement pathway more than control RBC in an in vitro system. RBC were incubated in 50% normal human serum chelated with 3.5 mM MgCl2 and 10 mM EGTA to block activation by the classical pathway, but allow alternative pathway activation. SCD RBC yielded significantly more activation, as measured by an EIA for C3, P complexes, than control RBC. Denser SCD RBC produced more activation than control RBC, unfractionated SCD RBC, and less dense SCD RBC. These findings are consistent with the hypothesis that dense irreversibly sickled cells, or membrane spicules or vesicles derived from them, may result in complement activation.

Human IgG antibody to group b Streptococcus type III: comparison of protective levels in a murine model with levels in infected human neonates.

We determined the serum concentration of human IgG antibody to the native capsular polysaccharide of group B Streptococcus (GBS) type III needed to passively protect mice against lethal homologous challenge. Antibody was measured by an ELISA, standardized by two methods, and corrected for nonprecipitating antibody. A concentration of 1.3 micrograms of IgG antibody to GBS type III/ml protected 126 (97%) of 130 mice from an 80%-96% lethal dose bacterial challenge. Concentrations of IgG antibody to GBS type III in sera from 42 infected infants were less than or equal to 0.3 micrograms/ml. Concentrations of antibody ranged from less than 0.02 to 21.7 micrograms/ml in sera from 102 unselected pregnant women (median, 0.05 microgram/ml); 13% had concentrations greater than or equal to 1.3 microgram/ml. Levels in 25 women colonized with GBS type III who gave birth to normal infants were significantly higher and ranged from 0.1 to 10.7 microgram/ml (median, 0.78 micrograms/ml). In a study of transplacental passage of antibody, protective levels were found in a number of infants with gestational ages between 28 and 36 weeks.

Protective levels of human immunoglobulin G antibody to group B streptococcus type Ib.

We studied the concentration of circulating human immunoglobulin G (IgG) antibody to the native capsular polysaccharide of group B streptococcus (GBS) type Ib necessary to protect mice against lethal challenge by laboratory and clinical GBS Ib strains. Antibody was measured by an enzyme-linked immunosorbent assay in which native polysaccharide antigen coupled to human serum albumin was used. The assay was standardized by a quantitative precipitation test, using native antigen and specific human IgG antibody purified by affinity chromatography. IgG anti-GBS Ib antibody level-protection curves for 90% lethal dose challenge of mice were sigmoidal. The curves of whole serum and affinity-chromatographed IgG anti-GBS Ib were superimposable. The serum concentrations of human antibody required for complete protection of mice varied with the infecting strain and ranged from 0.038 to 0.175 microgram/ml. Protective levels of human IgG anti-GBS Ib were lower than those we found previously for homologous protection against GBS Ia challenge (range, 0.25 to 1.0 microgram/ml).

Type-specific capsular antigen is associated with virulence in late-onset group B Streptococcal type III disease.

Strain differences have been postulated to explain the observation that group B Streptococcus type III (GBS III) late-onset disease occurs in only a fraction of colonized infants. To determine the distribution of type-specific polysaccharide antigen (Ag) in GBS III, Ag was measured by rocket immunoelectrophoresis in both supernatant fluids and EDTA extracts and by radial immunodiffusion in multiple HCl extracts of the pellet from cultures of 10 strains of GBS III. Capsular Ag was defined as the sum of Ag in EDTA extracts + Ag in multiple HCl extracts. Both Ag in EDTA extracts and Ag in supernatant fluids correlated with capsular Ag (r = 0.94). GBS III strains were obtained from the blood of 19 infants with late-onset sepsis, from the cerebrospinal fluid or blood of 22 infants with late-onset meningitis, and from mucosal surfaces of both 18 infants and 12 mothers of infants with low levels of type-specific antibody and asymptomatic colonization. Mean values of Ag in supernatant fluids in strains from infants with late-onset sepsis (1.50 +/- 0.08 micrograms/ml) and late-onset meningitis (1.67 +/- 0.09 micrograms/ml) were significantly greater than those in asymptomatic colonization strains (1.14 +/- 0.05 micrograms/ml; P less than 0.001). The number of organisms required for a 50% lethal dose in the chick embryo, determined in 29 strains, was inversely related to Ag in supernatant fluids (r = -0.60). The demonstration that the quantity of capsular Ag produced by GBS III strains is related to their virulence in chick embryos and to their invasiveness in susceptible infants supports the hypothesis that Ag is a virulence factor in humans.

Estimation of the protective level of human IgG antibody to the type-specific polysaccharide of group B Streptococcus type Ia.

Human IgG antibody to the type-specific polysaccharide antigen of group B Streptococcus type Ia in the sera of mice was measured by enzyme-linked immunosorbent assay, previously standardized by quantitative precipitation, 24 hr after passive immunization with human serum or affinity-chromatographed antibody. The concentrations of antibody needed to protect mice against 90% lethal dose challenge varied with the bacterial inoculum and ranged from 0.25 to 1 microgram/ml using five strains of group B Streptococcus type Ia. Affinity-chromatographed antibody gave results comparable to serum, indicating the specificity of the antibody and the absence of other humoral factors in protection with this serum. Sera from 11 infected infants and their mothers had concentrations of antibody of less than or equal to 0.17 micrograms/ml, below the protective level delineated in the experimental model. Twelve percent of 50 adult women and 36% of 25 women colonized with group B Streptococcus type Ia had antibody levels of greater than or equal to 1 microgram/ml.

An enzyme-linked immunosorbent assay (ELISA) for human IgG antibody to the type Ia polysaccharide of group B streptococcus.

An ELISA was developed to measure human IgG antibody to the native polysaccharide antigen of GBS serotype Ia. Because the polysaccharide binds poorly to polyvinyl chloride, its adherence was enhanced by activation with cyanogen bromide and coupling to HSA in a molar ratio of polysaccharide to albumin of 1:4.5. There was minimal loss of sialic acid during coupling, and the coupled antigen showed identity with uncoupled native antigen by Ouchterlony analysis. OD values obtained by ELISA showed a log-linear relation to concentration of specific antibody in whole and affinity-chromatographed human sera measured by quantitative precipitation over a range of 0.25 to 3.5 microgram/ml. In replicate ELISA experiments using serially diluted human serum, dilutions with antibody content as low as 0.016 microgram/ml could be reliably differentiated from PBS or agammaglobulinemic serum. The concentration of antibody in 98 selected human sera measured by ELISA correlated well (r = 0.89, p less than 0.001) with results obtained by indirect IF assay. This quantitative ELISA for GBS antibody is rapid, convenient, economical, and suitable for routine use.

Quantitative nitrobluetetrazolium reduction by normal newborn monocytes.

The phagocytosis-induced metabolic burst of human newborn monocytes, as evaluated by their capacity to reduce NBT, was comparable to that of adult monocytes. The NBT reduction assay constitutes a simple method of ascertaining the functional capacity of human monocytes.

Technetium-99m labeling of human monocytes for chemotactic studies.

Although the current use of 51Cr for the evaluation of monocyte chemotaxis has yielded encouraging results, certain properties of this radionuclide leave room for improvement. Technetium-99m was evaluated as an alternative label for this purpose. A cell loss of 50% was found, but the recovered cells showed excellent viability and function. Chemotaxis was measured using a modified Boyden's chamber and a lymphocyte-derived chemotactic factor. The chemotactic properties of 99mTc-labeled human monocytes were preserved, and an excellent correlation between radioactive measurement and microscopic counting of migrating cells was observed.

Chemotactic and bactericidal capacities of human newborn monocytes.

We evaluated the chemotactic and bactericidal capacities of human cord blood monocytes, and the ability of cord lymphocytes and sera to generate chemotactic (LDCF) and opsonic factors for monocytes. Our results suggest that the intrinsic locomotive capacity, and the receptor system for LDCF, are adequately developed in the newborn monocyte. Moreover, newborn lymphocytes appear to produce adequate amounts of LDCF, capable of attracting adult monocytes. Poor chemotaxis was observed only when cord monocytes were exposed to supernatants of cord lymphocytes, which suggests that both contribute to this abnormal response. An inhibitory factor for which only cord monosensitivity of monocytes and strength of chemotactic factor between adults and newborns would explain these results. Bactericidal capactiy of cord monocytes against Escherichia coli K-12 opsonized by either cord or adult serum was comparable to that of adult monocytes.