Reduced fetal growth velocity precedes antepartum fetal death.

OBJECTIVES: To determine whether decreased fetal growth velocity precedes antepartum fetal death and to evaluate whether fetal growth velocity is a better predictor of antepartum fetal death compared to a single fetal biometric measurement at the last available ultrasound scan prior to diagnosis of demise. METHODS: This was a retrospective, longitudinal study of 4285 singleton pregnancies in African-American women who underwent at least two fetal ultrasound examinations between 14 and 32 weeks of gestation and delivered a liveborn neonate (controls; n = 4262) or experienced antepartum fetal death (cases; n = 23). Fetal death was defined as death diagnosed at ≥ 20 weeks of gestation and confirmed by ultrasound examination. Exclusion criteria included congenital anomaly, birth at < 20 weeks of gestation, multiple gestation and intrapartum fetal death. The ultrasound examination performed at the time of fetal demise was not included in the analysis. Percentiles for estimated fetal weight (EFW) and individual biometric parameters were determined according to the Hadlock and Perinatology Research Branch/Eunice Kennedy Shriver National Institute of Child Health and Human Development (PRB/NICHD) fetal growth standards. Fetal growth velocity was defined as the slope of the regression line of the measurement percentiles as a function of gestational age based on two or more measurements in each pregnancy. RESULTS: Cases had significantly lower growth velocities of EFW (P < 0.001) and of fetal head circumference, biparietal diameter, abdominal circumference and femur length (all P < 0.05) compared to controls, according to the PRB/NICHD and Hadlock growth standards. Fetuses with EFW growth velocity < 10th percentile of the controls had a 9.4-fold and an 11.2-fold increased risk of antepartum death, based on the Hadlock and customized PRB/NICHD standards, respectively. At a 10% false-positive rate, the sensitivity of EFW growth velocity for predicting antepartum fetal death was 56.5%, compared to 26.1% for a single EFW percentile evaluation at the last available ultrasound examination, according to the customized PRB/NICHD standard. CONCLUSIONS: Given that 74% of antepartum fetal death cases were not diagnosed as small-for-gestational age (EFW < 10th percentile) at the last ultrasound examination when the fetuses were alive, alternative approaches are needed to improve detection of fetuses at risk of fetal death. Longitudinal sonographic evaluation to determine growth velocity doubles the sensitivity for prediction of antepartum fetal death compared to a single EFW measurement at the last available ultrasound examination, yet the performance is still suboptimal. © 2020 International Society of Ultrasound in Obstetrics and Gynecology.

Prediction of adverse perinatal outcome by fetal biometry: comparison of customized and population-based standards.

OBJECTIVE: To compare the predictive performance of estimated fetal weight (EFW) percentiles, according to eight growth standards, to detect fetuses at risk for adverse perinatal outcome. METHODS: This was a retrospective cohort study of 3437 African-American women. Population-based (Hadlock, INTERGROWTH-21st , World Health Organization (WHO), Fetal Medicine Foundation (FMF)), ethnicity-specific (Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD)), customized (Gestation-Related Optimal Weight (GROW)) and African-American customized (Perinatology Research Branch (PRB)/NICHD) growth standards were used to calculate EFW percentiles from the last available scan prior to delivery. Prediction performance indices and relative risk (RR) were calculated for EFW < 10th and > 90th percentiles, according to each standard, for individual and composite adverse perinatal outcomes. Sensitivity at a fixed (10%) false-positive rate (FPR) and partial (FPR < 10%) and full areas under the receiver-operating-characteristics curves (AUC) were compared between the standards. RESULTS: Ten percent (341/3437) of neonates were classified as small-for-gestational age (SGA) at birth, and of these 16.4% (56/341) had at least one adverse perinatal outcome. SGA neonates had a 1.5-fold increased risk of any adverse perinatal outcome (P < 0.05). The screen-positive rate of EFW < 10th percentile varied from 6.8% (NICHD) to 24.4% (FMF). EFW < 10th percentile, according to all standards, was associated with an increased risk for each of the adverse perinatal outcomes considered (P < 0.05 for all). The highest RRs associated with EFW < 10th percentile for each adverse outcome were 5.1 (95% CI, 2.1-12.3) for perinatal mortality (WHO); 5.0 (95% CI, 3.2-7.8) for perinatal hypoglycemia (NICHD); 3.4 (95% CI, 2.4-4.7) for mechanical ventilation (NICHD); 2.9 (95% CI, 1.8-4.6) for 5-min Apgar score < 7 (GROW); 2.7 (95% CI, 2.0-3.6) for neonatal intensive care unit (NICU) admission (NICHD); and 2.5 (95% CI, 1.9-3.1) for composite adverse perinatal outcome (NICHD). Although the RR CIs overlapped among all standards for each individual outcome, the RR of composite adverse perinatal outcome in pregnancies with EFW < 10th percentile was higher according to the NICHD (2.46; 95% CI, 1.9-3.1) than the FMF (1.47; 95% CI, 1.2-1.8) standard. The sensitivity for composite adverse perinatal outcome varied substantially between standards, ranging from 15% for NICHD to 32% for FMF, due mostly to differences in FPR; this variation subsided when the FPR was set to the same value (10%). Analysis of AUC revealed significantly better performance for the prediction of perinatal mortality by the PRB/NICHD standard (AUC = 0.70) compared with the Hadlock (AUC = 0.66) and FMF (AUC = 0.64) standards. Evaluation of partial AUC (FPR < 10%) demonstrated that the INTERGROWTH-21st standard performed better than the Hadlock standard for the prediction of NICU admission and mechanical ventilation (P < 0.05 for both). Although fetuses with EFW > 90th percentile were also at risk for any adverse perinatal outcome according to the INTERGROWTH-21st (RR = 1.4; 95% CI, 1.0-1.9) and Hadlock (RR = 1.7; 95% CI, 1.1-2.6) standards, many times fewer cases (2-5-fold lower sensitivity) were detected by using EFW > 90th percentile, rather than EFW < 10th percentile, in screening by these standards. CONCLUSIONS: Fetuses with EFW < 10th percentile or EFW > 90th percentile were at increased risk of adverse perinatal outcomes according to all or some of the eight growth standards, respectively. The RR of a composite adverse perinatal outcome in pregnancies with EFW < 10th percentile was higher for the most-stringent (NICHD) compared with the least-stringent (FMF) standard. The results of the complementary analysis of AUC suggest slightly improved detection of adverse perinatal outcome by more recent population-based (INTERGROWTH-21st ) and customized (PRB/NICHD) standards compared with the Hadlock and FMF standards. Published 2019. This article is a U.S. Government work and is in the public domain in the USA.

Funisitis and chorionic vasculitis: the histological counterpart of the fetal inflammatory response syndrome.

OBJECTIVE: To determine whether there is a relationship between the presence of histological signs of inflammation in the extraplacental membranes and umbilical cord and the concentrations of fetal plasma interleukin-6 (IL-6). METHODS: The study examined a cohort of patients who were admitted with preterm labor or preterm premature rupture of the membranes (PROM) and who underwent cordocentesis. Inclusion criteria included fetal plasma available for IL-6 determination, histological examination of the umbilical cord and placenta, and delivery within 48 h of the procedure. This last criterion was used to preserve a meaningful temporal relationship between fetal plasma IL-6 and the results of histological examination of the placenta. Fetal plasma IL-6 was determined by a high sensitivity ELISA. Forty-five patients were available for study: 18 patients had preterm labor with intact membranes and 27 had preterm PROM. RESULTS: The incidence of funisitis was 44.4% (20/45): 27.8% (5/18) in patients with preterm labor and intact membranes and 55.6% (15/27) in patients with preterm PROM. The median values of fetal plasma IL-6 in patients with funisitis, chorioamnionitis without funisitis, and non-inflamed membranes were 51.4, 18.4 and 5.2 pg/ml, respectively. After log transformation of the fetal plasma IL-6 concentration, the means differed significantly from each other (ANOVA, p < 0.02). There was no difference in log fetal plasma IL-6 concentration between patients with funisitis and those with chorioamnionitis without funisitis. The difference in mean concentration of log fetal plasma IL-6 between patients with funisitis or chorionic vasculitis and those without inflammation was highly significant (post-hoc test, p = 0.01 and p < 0.01, respectively). Fetuses with fetal plasma IL-6 > 11 pg/ml had a significantly higher rate of histological signs of inflammation in the extra-placental membranes and umbilical cord than those with fetal plasma IL-6 < 11 pg/ml (funisitis: 55.6% (15/27) vs. 27.8% (5/18), p < 0.05; chorionic vasculitis: 55.6% (15/27) vs. 12.5% (2/16), p < 0.01; chorioamnionitis only: 25.9% (7/27) vs. 16.7% (3/18), p < 0.05; no inflammation: 18.5% (5/27) vs. 55.6% (10/18), p < 0.05, respectively). Fetuses with funisitis had significantly higher rates of clinical and histological chorioamnionitis, and neonatal infectious morbidity (proven + suspected sepsis) than fetuses without funisitis (40% (8/20) vs. 8% (2/25), 90% (18/20) vs. 36% (9/25), and 40% (8/20) vs. 4% (1/25), respectively; p < 0.01 for each). Fetuses with chorionic vasculitis had significantly higher rates of clinical and histological chorioamnionitis as well as neonatal infectious morbidity (proven + suspected sepsis) than fetuses without chorionic vasculitis (100% (17/17) vs. 42.3% (11/26), p < 0.01; 82.4% (14/17) vs. 50.0% (13/26), p = 0.05; and 41.2% (7/17) vs. 7.7% (2/26), p = 0.01). CONCLUSION: Fetal plasma IL-6 concentration is significantly associated with the presence of inflammatory lesions in the extraplacental membranes and umbilical cord. Fetuses with fetal plasma IL-6 > 11 pg/ml had a significantly higher rate of funisitis and/or chorionic vasculitis than fetuses with fetal plasma IL-6 < 11 pg/ml. These findings suggest that funisitis/chorionic vasculitis is the histological manifestation of the fetal inflammatory response syndrome.

Maternal intravascular inflammation in preterm premature rupture of membranes.

OBJECTIVE: Intrauterine inflammation has been implicated in the mechanisms responsible for preterm premature rupture of membranes (PROM). However, it is unclear whether this inflammatory process remains localized to the uterus, at the site of membrane rupture, or extends to the maternal compartment. Flow cytometric analysis is a sensitive method to assess the presence and magnitude of in vivo inflammation. This study was conducted to determine whether preterm PROM is associated with changes in the phenotypic and metabolic characteristics of maternal granulocytes and monocytes consistent with the presence of maternal intravascular inflammation. STUDY DESIGN: A prospective cross-sectional study was performed including patients with preterm PROM (n = 43) and normal pregnancy (n = 51). Maternal intravascular inflammation was studied using flow cytometry. Maternal blood was assayed to determine granulocyte and monocyte phenotype using monoclonal antibodies, which included cluster differentiation (CD) markers CD11b, CD14, CD15, CD16, CD18, CD49d, CD62L, CD64, CD66b and human leukocyte antigen (HLA)-DR. The quantities of basal intracellular reactive oxygen species (iROS) and oxidative burst was assessed. Statistical analysis was conducted with the use of non-parametric methods. A p value < 0.01 was considered significant. RESULTS: Preterm PROM was associated with a significant increase in the median mean channel brightness (MCB) of CD11b, CD14, CD64 and CD66b on granulocytes and median MCB of CD11b on monocytes. The oxidative burst and the stimulation index in both cell types were higher in preterm PROM than in normal pregnancy (p < 0.01). CONCLUSION: Preterm PROM is associated with phenotypic and metabolic changes in circulating granulocytes and monocytes.

Phenotypic and metabolic characteristics of monocytes and granulocytes in preeclampsia.

OBJECTIVE: The maternal syndrome of preeclampsia has recently been attributed to a systemic intravascular inflammatory response and endothelial cell activation and dysfunction. This novel hypothesis has considerable clinical and biological implications. This study was designed to determine whether women with preeclampsia have evidence of intravascular inflammation by examination of the phenotypic and metabolic activity of granulocytes and monocytes. STUDY DESIGN: A cross-sectional study was performed that included patients with preeclampsia (n = 31) and normal pregnancies (n = 58) matched for gestational age at blood draw. Intravascular inflammation was studied with use of flow cytometry. Peripheral venous blood was assayed to determine granulocyte and monocyte phenotype with the use of monoclonal antibodies for selective cluster differentiation (CD) antigens. The panel of antibodies included CD11b, CD14, CD16, CD18, CD49d, CD62L, CD64, CD66b, and HLA-DR. The quantity of basal intracellular reactive oxygen species and oxidative burst was assessed. Results were reported as mean channel brightness or intensity of detected fluorescence. Analysis was conducted with nonparametric statistics. A P value <.01 was considered to be significant. RESULTS: Preeclampsia was associated with a significant increase in mean channel brightness for CD11b on granulocytes and monocytes but lower mean channel brightness for CD62L on granulocytes than those from women with normal pregnancy (P <.01 for each). Basal intracellular reactive oxygen species were increased in monocytes but not in granulocytes. The oxidative burst was higher in both cell types. CONCLUSION: Preeclampsia is associated with phenotypic and metabolic changes in granulocytes and monocytes.

A role for the 72 kDa gelatinase (MMP-2) and its inhibitor (TIMP-2) in human parturition, premature rupture of membranes and intraamniotic infection.

OBJECTIVE: Degradation of the extracellular matrix in fetal membranes has been implicated in the process of parturition and rupture of membranes. Matrix metalloproteinases (MMPs) are enzymes capable of degrading extracellular matrix including collagen. Tissue inhibitors of matrix metalloproteinases (TIMPs) inhibit the activity of MMPs by covalently binding to the enzymes. MMP-2 degrades Type IV collagen and TIMP-2 is its specific inhibitor. The objective of this study was to determine if human parturition, rupture of membranes (term and preterm) and microbial invasion of the amniotic cavity (MIAC) are associated with changes in the concentrations of MMP-2 and TIMP-2 in amniotic fluid. STUDY DESIGN: A cross-sectional study was conducted with women in the following categories: 1) term with intact membranes, in labor and not in labor; 2) preterm labor and intact membranes who delivered at term, who delivered preterm and preterm labor with MIAC; 3) preterm premature rupture of membranes (PROM) with and without infection; 4) term and preterm PROM not in labor; and 5) midtrimester. MMP-2 and TIMP-2 concentrations in amniotic fluid were determined using sensitive and specific immunoassays. RESULTS: The concentration of TIMP-2 increased with advancing gestational age (r = 0.6, p < 0.001). No correlation was found between MMP-2 concentrations and gestational age. Human parturition and rupture of membranes (term and preterm) and in patients with intact membranes were not associated with changes in the amniotic fluid MMP-2 concentrations. In contrast, 1) patients with spontaneous labor (term and preterm) had significantly lower median concentrations of TIMP-2 compared to those not in labor (p < 0.05 for both); 2) MIAC in women with preterm labor and preterm PROM was associated with a significant decrease in amniotic fluid TIMP-2 concentrations (p < 0.04 for both comparisons); 3) Rupture of the membranes (term and preterm) was also associated with a significant decrease in the amniotic fluid TIMP-2 concentrations (p < 0.05 and p < 0.03, respectively). CONCLUSIONS: Human parturition (preterm and term), rupture of fetal membranes (term and preterm) and intraamniotic infection are associated with a significant decrease in amniotic fluid TIMP-2 concentrations.

Further observations on the fetal inflammatory response syndrome: a potential homeostatic role for the soluble receptors of tumor necrosis factor alpha.

OBJECTIVE: The fetal inflammatory response syndrome is a subclinical condition frequently present in preterm labor and preterm premature rupture of the membranes and is associated with increased perinatal morbidity and mortality. Tumor necrosis factor alpha is a mediator of septic shock and death, and it exerts its biologic effects by interacting with 2 receptors, TNF-R1 and TNF-R2. Soluble tumor necrosis factor receptors can buffer the biologic activity and protect against the deleterious effects of tumor necrosis factor alpha. The purpose of this study was to determine the behavior of soluble tumor necrosis factor receptors in fetuses with and without fetal inflammatory response syndrome. STUDY DESIGN: Fetal blood sampling was performed in patients with preterm labor (n = 95) and preterm premature rupture of the membranes (n = 39). Control samples were obtained from fetuses who were undergoing blood sampling for clinical indications and had normal outcomes (n = 21). Fetal inflammatory response syndrome was defined as a fetal plasma interleukin 6 concentration >11 pg/mL. Concentrations of interleukin 6 and TNF-R1 and TNF-R2 were determined by use of sensitive and specific immunoassays. Analysis of covariance was used for statistical analysis. RESULTS: (1) TNF-R1 and TNF-R2 were detectable in all samples, and their concentrations decreased with advancing gestational age (r = -0.8 and r = -0.7; P<.0001 and P<.001, respectively). (2) The mean fetal plasma concentrations of TNF-R1 and TNF-R2 were significantly higher in fetuses with fetal inflammatory response syndrome than in those without the syndrome after adjustment for gestational age and fetal membrane status (TNF-R1: no fetal inflammatory response syndrome, mean +/- SE, 3473.7+/-128.8 pg/mL; vs fetal inflammatory response syndrome, mean +/- SE, 4079.9+/-190.7 pg/mL; P<.005; TNF-R2: no fetal inflammatory response syndrome, mean +/- SE, 6033.2+/-235.4 pg/mL; vs. fetal inflammatory response syndrome, mean +/- SE, 7783.1+/-342.8 pg/mL; P<.0001). (3) Fetuses of patients who delivered within 72 hours of cordocentesis had significantly higher concentrations of TNF-R1 and TNF-R2 receptors than those with longer latency periods (P<.05 for each). CONCLUSION: The fetal inflammatory response syndrome is associated with increased availability of the soluble receptors of tumor necrosis factor alpha in fetal plasma. These factors may attenuate the deleterious effects of tumor necrosis factor alpha.

Participation of the novel cytokine interleukin 18 in the host response to intra-amniotic infection.

OBJECTIVE: Interleukin 18 is a proinflammatory pleiotropic cytokine that has been implicated in the host defense against infection. This study was undertaken to determine whether interleukin 18 concentrations change in the maternal, fetal, and amniotic fluid compartments with labor (term and preterm) and microbial invasion of the amniotic cavity. STUDY DESIGN: Amniotic fluid was assayed for interleukin 18 in samples obtained from 285 patients in the following groups: (1) term not in labor (n = 22), in labor (n = 19), and with microbial invasion of the amniotic cavity (n = 16); (2) preterm labor who delivered at term (n = 38), who delivered preterm but without microbial invasion of the amniotic cavity (n = 41), and preterm labor with microbial invasion of the amniotic cavity (n = 24); (3) preterm premature rupture of membranes without microbial invasion of the amniotic cavity (n = 30) and with microbial invasion of the amniotic cavity (n = 34); (4) term premature rupture of membranes not in labor (n = 20) and term premature rupture of membranes in labor (n = 19); and (5) midtrimester (n = 22). In addition, cord and maternal plasma samples from women at term not in labor (n = 20) and in labor (n = 20) were assayed for interleukin 18. RESULTS: (1) Interleukin 18 was detectable in all amniotic fluid samples and maternal and umbilical cord blood samples. (2) Interleukin 18 concentrations increased with advancing gestational age (r = 0.47; P <.0001). (3) Microbial invasion of the amniotic cavity in either preterm or term parturition was associated with a significant increase in the amniotic fluid concentration of interleukin 18 (preterm labor without microbial invasion of the amniotic cavity: median, 14.95 pg/mL; range, 3.9-277.0 pg/mL; vs preterm labor with microbial invasion of the amniotic cavity: median, 20.75 pg/mL; range, 5.53-160.21 pg/mL; P <.02; term labor without microbial invasion of the amniotic cavity: median, 18.73 pg/mL; range, 5.09-95.44 pg/mL; vs term labor with microbial invasion of the amniotic cavity: median, 24.35 pg/mL; range, 10.07-144.42 pg/mL; P<.004). (4) Both term and preterm parturition were associated with a modest increase in amniotic fluid interleukin 18 concentrations, although this trend did not reach statistical significance. (5) Rupture of membranes at term was associated with a significant decrease in amniotic fluid interleukin 18 concentrations (intact membranes: median, 14.96 pg/mL; range, <3.89-26.07 pg/mL; vs rupture of membranes: median, 10.1 pg/mL; range, 4.29-21.44 pg/mL; P <.001). CONCLUSION: (1) Interleukin 18 is increased in cases of microbial invasion of the amniotic cavity. (2) Interleukin 18 is detectable in the amniotic, maternal, and fetal compartments. (3) We propose that this novel cytokine plays a role in the host defense against infection.

Evidence of in vivo differential bioavailability of the active forms of matrix metalloproteinases 9 and 2 in parturition, spontaneous rupture of membranes, and intra-amniotic infection.

OBJECTIVE: Matrix metalloproteinases (MMP-9 and MMP-2) have been implicated in the digestion of fetal membranes. The purpose of this study was to determine the amniotic fluid concentrations of active forms of MMP-2 and MMP-9 and to explore the participation of these enzymes in labor (term and preterm), rupture of membranes (term and preterm), and microbial invasion of the amniotic cavity. STUDY DESIGN: A cross-sectional study was conducted with 291 women in the following categories: (1) term not in labor, (2) term in labor, (3) preterm labor and intact membranes who delivered at term, (4) preterm labor who delivered preterm, (5) preterm labor with microbial invasion of the amniotic cavity, (6) preterm premature rupture of membranes without microbial invasion of the amniotic cavity, (7) preterm premature rupture of membranes with microbial invasion of the amniotic cavity, (8) term premature rupture of membranes not in labor, and (9) mid trimester. Active forms of MMP-2 and MMP-9 were measured by a novel assay that uses a substrate developed by protein engineering. RESULTS: (1) MMP-2 and MMP-9 were detected in 88% and 96% of amniotic fluid samples, respectively (255/291 and 279/291). (2) The concentrations of active forms of MMP-2 and MMP-9 changed with advancing gestational age. (3) Spontaneous term parturition was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) and a significant decrease in the median concentration of the active forms of MMP-2 (P <.003). (4) Preterm labor with intact membranes leading to preterm delivery in the absence of infection was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.2). (5) Rupture of membranes (either term or preterm) was associated with a significant increase in the concentration of the active forms of MMP-9 and with a significant decrease in the concentration of the active forms of MMP-2 (P <.005 for term and P <.03 and P <.003 for preterm, respectively). (6) Microbial invasion of the amniotic cavity in women with preterm premature rupture of membranes was also associated with a significant increase in the concentration of the active forms of MMP-9 (P <.03) and a decrease in the concentration of the active forms of MMP-2 (P <.05). (7) Microbial invasion of the amniotic cavity in patients with preterm labor was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.6). CONCLUSION: Spontaneous rupture of membranes (either term or preterm), parturition (either term or preterm), and microbial invasion of the amniotic cavity were associated with significant increases in the amniotic fluid concentration of the active forms of MMP-9. In contrast, the concentration of the active forms of MMP-2 either decreased or remained the same in these conditions. Our observations provide evidence for a novel regulation of gelatinolytic activity in vivo.

Lactoferrin in intrauterine infection, human parturition, and rupture of fetal membranes.

OBJECTIVE: Lactoferrin is an iron-binding protein with antimicrobial properties. This study was undertaken to determine whether amniotic fluid concentrations of this protein change with gestational age, infection, labor, and rupture of membranes. STUDY DESIGN: This cross-sectional study included women who underwent transabdominal amniocentesis (n = 268) in the following groups: (1) mid trimester of pregnancy; (2) preterm labor who delivered at term, preterm labor who delivered preterm with intra-amniotic infection, and preterm labor who delivered preterm without intra-amniotic infection; (3) preterm premature rupture of membranes in the presence or absence of intra-amniotic infection; (4) term with intact membranes not in labor, in labor, and in labor with intra-amniotic infection; and (5) premature rupture of membranes at term not in labor. In addition, lactoferrin concentrations were determined in maternal plasma and cord blood of patients at term not in labor. Lactoferrin concentration was measured with an immunoassay. RESULTS: (1) Lactoferrin was detectable in 85.4% (229/268) of amniotic fluid samples, not detectable in all fluid obtained in the mid trimester, and detectable in all maternal and cord plasma samples. (2) The concentration of lactoferrin increased with advancing gestational age (r = 0.68; P <.0001). (3) Intra-amniotic infection was associated with significant increases in amniotic fluid lactoferrin concentrations in patients with preterm labor (no intra-amniotic infection median, 1641.2 ng/mL; range, <1.24-35,090.0 ng/mL; vs intra-amniotic infection median, 3833.6 ng/mL; range, 746.0-47,020.0 ng/mL; P <.001), term labor (no intra-amniotic infection median, 2085.8 ng/mL; range, 425.0-23,230.0 ng/mL; vs intra-amniotic infection median, 5627.0 ng/mL; range, <1.24-19,220.0 ng/mL; P <. 001), and preterm premature rupture of membranes (no intra-amniotic infection median, 2190 ng/mL; range, <1.24-7456.1 ng/mL; vs intra-amniotic infection median, 3449.3 ng/mL; range, <1.24-83,600. 0; P <.01). (4) Spontaneous labor at term but not preterm was associated with a significant decrease in amniotic fluid lactoferrin concentration (P <.05). (5) Spontaneous term parturition was associated with a significant increase in umbilical cord plasma lactoferrin concentration (P <.005). CONCLUSION: (1) Intra-amniotic infection was consistently associated with dramatically increased concentrations of lactoferrin in amniotic fluid. (2) Term parturition was associated with a significant increase in lactoferrin concentration in the fetal compartment (umbilical cord blood) and a decrease in the amniotic compartment. We propose that lactoferrin is part of the repertoire of host defense mechanisms against intra-amniotic infection.

Evidence for the participation of interstitial collagenase (matrix metalloproteinase 1) in preterm premature rupture of membranes.

OBJECTIVE: Rupture of membranes is thought to result from the effects of physical forces in localized areas of the membranes weakened by the degradation of structural collagens. Matrix metalloproteinases are enzymes that degrade extracellular matrix components and have been implicated in membrane rupture. The objective of this study was to determine whether spontaneous rupture of membranes is associated with a change in the amniotic fluid concentration of interstitial collagenase (matrix metalloproteinase 1 [MMP-1]), a major collagenase. STUDY DESIGN: A cross-sectional study was conducted to determine MMP-1 concentrations in amniotic fluid from 353 women in the following categories: (1) term with intact membranes not in labor and in labor, (2) preterm labor who delivered at term, (3) preterm labor who delivered preterm without microbial invasion of the amniotic cavity, (4) preterm labor who delivered preterm with microbial invasion of the amniotic cavity, (5) preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, (6) term premature rupture of membranes not in labor and in labor, and (7) mid trimester of pregnancy. Microbial invasion of the amniotic cavity was determined by an amniotic fluid culture positive for microorganisms. MMP-1 concentrations in amniotic fluid were determined by means of sensitive and specific immunoassays. RESULTS: (1) MMP-1 was detectable in 81.3% of amniotic fluid samples (287/353), and its concentrations increased with advancing gestational age (r = 0.4; P <.001). (2) Preterm premature rupture of membranes was associated with a significant increase in the median amniotic fluid concentration of MMP-1 (P =.02). (3) Women with term premature rupture of membranes had a significantly lower amniotic fluid MMP-1 concentration than those with intact membranes at term not in labor (P <.001). (4) Microbial invasion of the amniotic cavity in patients in preterm labor with intact membranes and in patients with preterm premature rupture of membranes was also associated with significant increases in the median amniotic fluid MMP-1 concentrations (P <.05 and P <.01, respectively). (5) Patients with preterm premature rupture of membranes and microbial invasion of the amniotic cavity had a significantly higher median amniotic fluid MMP-1 concentration than those with intact membranes and microbial invasion of the amniotic cavity (P =.01). (6) Neither term nor preterm parturition was associated with changes in amniotic fluid MMP-1 concentrations (P =.6 and P =.3, respectively). CONCLUSION: (1) Collagenase 1 (MMP-1) is a physiologic constituent of amniotic fluid. (2) Preterm premature rupture of membranes (in both the presence and absence of infection) was associated with an increase in the amniotic fluid MMP-1 concentrations. (3) Neither term nor preterm parturition was associated with a significant increase in the amniotic fluid concentration of MMP-1.

Human neutrophil collagenase (matrix metalloproteinase 8) in parturition, premature rupture of the membranes, and intrauterine infection.

OBJECTIVES: The mechanisms by which microbial invasion of the amniotic cavity leads to membrane weakening and rupture are poorly understood. Recently, endogenous host enzymes have been implicated in this process. Matrix metalloproteinases are a family of potent enzymes that degrade components of the extracellular matrix. Collagen type I provides the main tensile strength of the fetal membranes. Matrix metalloproteinase 8 (MMP-8), or neutrophil collagenase, degrades interstitial collagens, acting preferentially on collagen type I. This study was undertaken (1) to determine whether MMP-8 is present in amniotic fluid and whether its concentrations are changed in preterm and term labor and membrane rupture with and without intra-amniotic infection and (2) to determine whether the amniotic fluid concentrations of MMP-8 in labor at term are different in the lower and upper uterine compartments. STUDY DESIGN: A cross-sectional study was conducted and transabdominal amniocentesis was performed in women in the following categories: (1) midtrimester (n = 25), (2) preterm labor in the presence and absence of microbial invasion of the amniotic cavity (n = 86), (3) preterm premature rupture of the membranes in the presence and absence of microbial invasion of the amniotic cavity (n = 51), (4) term patients in labor and not in labor (n = 51), and (5) term premature rupture of membranes (n = 20). Additional paired samples of amniotic fluid were retrieved by transabdominal amniocentesis (upper compartment) and transvaginal amniocentesis (lower or forebag compartment) from 14 term patients (28 samples) in spontaneous labor with intact membranes. Amniotic fluid MMP-8 concentrations were determined with a sensitive and specific immunoassay. RESULTS: MMP-8 was detected in 95.4% (249/261) of all samples. (1) Spontaneous human parturition was associated with a significant increase in amniotic fluid concentrations of MMP-8 in both term and preterm gestation. Term (no labor median, 3.3 ng/mL; range, <0.06-38.6 ng/mL; vs labor median, 16.6 ng/mL; range, 0. 33-1650 ng/mL; P <.05). Patients with preterm labor who delivered preterm (in the absence of microbial invasion of the amniotic cavity) had a significantly higher median amniotic fluid MMP-8 concentration than those with preterm labor who delivered at term (preterm labor, term delivery median, 3.1 ng/mL; range, <0.06-415.1 ng/mL; vs preterm labor, preterm delivery median, 32.5 ng/mL; range, <0.06-6006.6 ng/mL;P <.003). (2) Spontaneous rupture of membranes in preterm gestation but not in term gestation was associated with elevated amniotic fluid concentrations of MMP-8. Preterm gestation (preterm labor, intact membranes median, 3.1 ng/mL; range, <0.06-415. 1 ng/mL; vs preterm premature rupture of membranes median, 35.1 ng/mL; range, 0.71-1184.1 ng/mL; P <.05). Term gestation (intact membranes median, 3.3 ng/mL; range, 0.24-38.6 ng/mL; vs rupture of membranes median, 5.6 ng/mL; range, 0.22-19.8 ng/mL; P =.9). (3) Microbial invasion of the amniotic cavity was associated with a significant increase in amniotic fluid MMP-8 concentration in patients with preterm labor and intact membranes, as well as in patients with preterm premature rupture of membranes. Preterm labor (no microbial invasion of the amniotic cavity, preterm delivery median, 32.5 ng/mL; range, <0.06-6006.6 ng/mL; vs microbial invasion of the amniotic cavity median, 208.1 ng/mL; range, 4.2-14,600 ng/mL; P <.001). Preterm premature rupture of membranes (no microbial invasion of the amniotic cavity median, 35.1 ng/mL; range, 0.71-1184. 1 ng/mL; vs microbial invasion of the amniotic cavity median, 317.9 ng/mL; range, 2.16-14,500 ng/mL; P <.01). (4) The median amniotic fluid MMP-8 concentrations were significantly higher in fluid obtained from the forebag compartment than in that obtained from the upper compartment (median, 66.2 ng/mL; range, 7.4-170 ng/mL; vs median, 13.3 ng/mL; range, 2-170 ng/mL; respectively; P <.01). (ABSTRACT TRUNCATED)

Matrilysin (matrix metalloproteinase 7) in parturition, premature rupture of membranes, and intrauterine infection.

OBJECTIVE: Matrix metalloproteinases are enzymes capable of degrading extracellular matrix components. Matrilysin (matrix metalloproteinase 7), a novel member of this family, degrades fibronectin and proteoglycans. The objective of this study was to determine whether parturition (either term or preterm), premature rupture of the membranes, and microbial invasion of the amniotic cavity are associated with changes in the amniotic fluid concentration of matrilysin. STUDY DESIGN: A cross-sectional study was conducted with 275 women in the following categories: (1) second trimester, (2) term not in labor, (3) term in labor, (4) term with microbial invasion of the amniotic cavity, (5) preterm labor with intact membranes without microbial invasion of the amniotic cavity who delivered at term, (6) preterm labor without microbial invasion of the amniotic cavity who delivered preterm, (7) preterm labor with microbial invasion of the amniotic cavity, (8) preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, and (9) term premature rupture of membranes not in labor and without microbial invasion of the amniotic cavity. Matrilysin concentrations were measured with a sensitive specific immunoassay that was validated for amniotic fluid. RESULTS: Matrilysin was detectable in 97.4% (268/275) of the samples. The concentration of matrilysin increased with advancing gestational age (r = 0.8; P <.001). Parturition at term was not associated with a significant increase in amniotic fluid concentration of matrilysin. Preterm parturition in the absence of microbial invasion of the amniotic cavity was associated with a significant increase in amniotic fluid concentration of matrilysin (preterm labor with preterm delivery: median, 1.7 ng/mL; range, 0.45-21.6 mg/mL; vs preterm labor with term delivery: median, 1.2 ng/mL; range, 0.17-42. 1 ng/mL; P <.05). Premature rupture of membranes without microbial invasion of the amniotic cavity (either term or preterm) was not associated with a significant change in the amniotic fluid matrilysin concentration. Intra-amniotic infection was associated with a significant increase in amniotic fluid matrilysin among both patients with preterm labor and patients with preterm premature rupture of membranes (preterm labor with microbial invasion of the amniotic cavity: median, 3.2 ng/mL; range, 0.16-21.9 ng/mL; vs preterm labor and delivery without microbial invasion of the amniotic cavity: median, 1.7 ng/mL; range, 0.45-21.6 ng/mL; vs preterm labor with term delivery: median, 1.2 ng/mL; range, 0.17-42. 1 ng/mL; P <.01 for each comparison; and preterm premature rupture of membranes without microbial invasion of the amniotic cavity: median, 1.7 ng/mL; range, 0.29-13.9 ng/mL; vs preterm premature rupture of membranes with microbial invasion of the amniotic cavity: median, 3.6 ng/mL; range, 0.59-20.3 ng/mL; P <.01). CONCLUSION: Matrilysin is a physiologic constituent of amniotic fluid, and its concentration increases with advancing gestational age. Microbial invasion of the amniotic cavity in preterm gestations was associated with a significant increase in amniotic fluid concentration of matrilysin. Matrilysin therefore may play a role in the host defense mechanism.

A study of the relationship between placenta growth factor and gestational age, parturition, rupture of membranes, and intrauterine infection.

OBJECTIVE: Placenta growth factor is a potent angiogenic factor produced by the human placenta that has been implicated in the pathogenesis of preeclampsia and intrauterine growth restriction. Placenta growth factor belongs to the vascular endothelial growth factor family and is capable of inducing proliferation, migration, and activation of endothelial cells. The objective of this study was to determine the relationship between amniotic fluid concentration of placenta growth factor and gestational age, parturition (term and preterm), spontaneous rupture of the membranes, and intra-amniotic infection. STUDY DESIGN: Amniotic fluid samples obtained from 273 pregnant patients were assayed in the following clinical groups: midtrimester pregnancy, preterm labor who delivered at term, preterm labor without microbial invasion of the amniotic cavity who delivered preterm, preterm labor with microbial invasion of the amniotic cavity, term not in labor, term in labor, term with microbial invasion of the amniotic cavity, preterm premature rupture of membranes with and without microbial invasion of the amniotic cavity, and term with premature rupture of membranes without microbial invasion of the amniotic cavity. The placenta growth factor concentrations were determined by an immunoassay that is both sensitive and specific. RESULTS: Placenta growth factor was detectable in 96.3% (263/273) of samples. Amniotic fluid placenta growth factor concentration decreased with advancing gestational age (r = -0.42; P <.001). Amniotic fluid placenta growth factor concentrations were significantly higher in women in midtrimester pregnancy than in those at term not in labor (midtrimester pregnancy: median, 43.1 pg/mL; range, 22.9-69.8 pg/mL; vs term not in labor: median, 28.7 pg/mL; range, 16.1-82.7 pg/mL; P <.01). Neither term nor preterm parturition was associated with a change in amniotic fluid placenta growth factor concentrations. Term premature rupture of membranes was associated with a significant decrease in amniotic fluid placenta growth factor concentration (term premature rupture of membranes: median, 16.5 pg/mL; range <5.2-195.1 pg/mL; vs term intact membranes: median, 28.7 pg/mL; range, 16.1-822.7 pg/mL; P <.005). Preterm premature rupture of membranes was not associated with changes in amniotic fluid placenta growth factor concentrations. Intra-amniotic infection in preterm labor, term labor with intact membranes, and preterm premature rupture of membranes were not associated with changes in amniotic fluid placenta growth factor concentrations. CONCLUSION: Placenta growth factor is a physiologic constituent of amniotic fluid. Amniotic fluid concentrations of placenta growth factor decrease with advancing gestational age. Neither parturition nor infection affects amniotic fluid placenta growth factor concentrations.

Interleukin 16 in pregnancy, parturition, rupture of fetal membranes, and microbial invasion of the amniotic cavity.

OBJECTIVE: Interleukin 16 is a proinflammatory cytokine that promotes the recruitment of nonclonotypic T cells and eosinophils to sites of inflammation and induces resistance to activation-induced apoptosis. This peptide has no homology with members of the chemokine family and is produced by epithelial cells. No information is available about the expression of this cytokine during human pregnancy. This study was conducted to determine whether interleukin 16 is present in amniotic fluid and to examine the effects of labor and intrauterine infection on the concentrations of this cytokine. STUDY DESIGN: A cross-sectional study was constructed with 230 women in the following groups: (1) mid- trimester (n = 25); (2) term not in labor (n = 25), term in labor (n = 25), and term premature rupture of membranes not in labor (n = 40); (3) preterm labor and intact membranes with (n = 21) and without (n = 42) intra-ammiotic infection; and (4) preterm premature rupture of membranes in the presence (n = 29) and absence (n = 23) of microbial invasion of the amniotic cavity. Interleukin 16 concentration was measured with sensitive and specific immunoassays validated for amniotic fluid. Data were analyzed with nonparametric statistics. RESULTS: (1) Interleukin 16 was detected in 87.8% (202/230) of amniotic fluid samples. (2) Amniotic fluid interleukin 16 concentrations were significantly higher in women in the midtrimester than in those at term not in labor (median, 321.5 pg/mL; range, 146.9-1185.8 pg/mL; vs median, 85.9 pg/mL; range, <25-409.8 pg/mL; P <.001). (3) Labor at term was not associated with a significant increase in the median amniotic fluid interleukin 16 concentration. (4) Patients with preterm labor who delivered preterm had a significantly higher median amniotic fluid interleukin 16 concentration than those with preterm labor who delivered at term (median, 328.1 pg/mL; range, 38. 9-4660 pg/mL; vs median, 119.8 pg/mL; range, <25-558.5 pg/mL;P <.05). (5) Microbial invasion of the amniotic cavity was associated with a significant increase in median amniotic fluid interleukin 16 concentration in patients with preterm labor and intact membranes (microbial invasion of the amniotic cavity: median, 839 pg/mL; range, <25-8620 pg/mL; vs no microbial invasion of the amniotic cavity: median, 119.8 pg/mL; range, <25-558.5 pg/mL; P <.001) and in patients with preterm premature rupture of the membranes (microbial invasion of the amniotic cavity: median, 1005.8 pg/mL; range, <25-4590 pg/mL; vs no microbial invasion of the amniotic cavity: median, 204.9 pg/mL; range, 42.6-2347 pg/mL; P <.05). (6) Spontaneous rupture of the membranes at term but not preterm was associated with a significant decrease in amniotic fluid concentrations of interleukin 16 (term premature rupture of the membranes: median, <25 pg/mL; range, <25-231.2 pg/mL; vs term intact membranes: median, 85.9 pg/mL; range, <25-409.8 pg/mL; P <.05). CONCLUSIONS: Amniotic fluid interleukin-16 concentrations decreased with advancing gestational age. Women with preterm labor that led to preterm delivery and women with microbial invasion of the amniotic cavity had higher interleukin 16 amniotic fluid concentrations than those with preterm labor who delivered at term or those with sterile amniotic fluid. Microbial invasion of the amniotic cavity but not spontaneous labor at term or rupture of membranes was associated with increased concentrations of interleukin 16 in amniotic fluid.

A role for the novel cytokine RANTES in pregnancy and parturition.

OBJECTIVE: RANTES (regulated on activation, normal T cell expressed and secreted), a potent and versatile chemokine, is capable of attracting monocytes, lymphocytes, basophils, and eosinophils. This cytokine has been implicated in the regulation of the inflammatory response and in the recruitment of macrophages to the implantation site in early pregnancy. RANTES messenger ribonucleic acid and protein have been detected in fetal tissue and first-trimester trophoblast in response to bacterial endotoxin. The purpose of this study was to determine whether intrauterine infection, parturition (preterm and term), and gestational age affect the amniotic fluid concentrations of RANTES in human pregnancy. STUDY DESIGN: A cross-sectional study was designed to examine the relationship between labor, microbial invasion of the amniotic cavity, gestational age, and RANTES expression in amniotic fluid. Amniotic fluid was obtained from 214 women in the following groups: (1) midtrimester (n = 22), (2) preterm labor with intact membranes in the presence (n = 20) or absence (n = 74) of microbial invasion of the amniotic cavity, (3) term, not in labor (n = 44) and term, in labor in the presence (n = 27) and absence (n = 27) of microbial invasion of the amniotic cavity. Microbial invasion of the amniotic cavity was defined as a positive amniotic fluid culture for microorganisms. RANTES concentrations were determined by use of a sensitive and specific immunoassay. RESULTS: (1) Amniotic fluid RANTES concentrations decrease with advancing gestational age (r = 0. 43; P <.01). (2) Labor at term was associated with an increase in median concentrations of RANTES (labor-median, 8.4 pg/mL; range, <1.3-94.4 vs no labor-median, <1.3 pg/mL; range, <1.3-230.3; P <.01). (3) Women with preterm labor who delivered preterm (no microbial invasion of the amniotic cavity) had a higher median concentration of amniotic fluid RANTES than those who delivered at term (median, 12.7 pg/mL; range, <1.3-928 vs median, <1.3 pg/mL; range, <1.3-127. 5; P <.001). (4) Microbial invasion of the amniotic cavity was associated with a significant increase in median amniotic fluid RANTES in both preterm and term labor (preterm labor with microbial invasion of the amniotic cavity-median, 51.6 pg/mL; range, <1.3-2290 vs preterm labor without microbial invasion of the amniotic cavity-median, 12.7 pg/mL; range, <1.3-928 and vs preterm labor with delivery at term-median, <1.3 pg/mL; range, <1.3-127.5; P <.001 for each; term labor with microbial invasion of the amniotic cavity-median, 16.8 pg/mL; range, <1.3-171.4 vs term labor without microbial invasion of the amniotic cavity-median, 8.4 pg/mL; range, <1.3-94.4; P <.05 and vs no labor and no microbial invasion of the amniotic cavity-median, 1.4 pg/mL; range, <1.3-230.3; P <.001 and P <.05, respectively). CONCLUSION: These results support a role for RANTES in the mechanisms of human parturition and in the regulation of the host response to intrauterine infection.

Matrix metalloproteinases-9 in preterm and term human parturition.

OBJECTIVE: Spontaneous rupture of the fetal membranes occurs after the commencement of labor in 90% of cases. Recent evidence indicates that the process of parturition requires not only an increase in myometrial contractility and cervical ripening, but also degradation of extracellular matrix in fetal membranes (i.e., leakage of fibronectin into cervico-vaginal secretions). This study was undertaken to determine if parturition is associated with in vivo evidence of increased bioavailability of matrix metalloproteinases-9 (MMP-9) and its inhibitor, tissue inhibitor of metalloproteinases-1(TIMP-1). METHODS: A cross-sectional study was conducted with women in the following categories: 1) midtrimester (n = 25); 2) preterm labor and intact membranes in the absence of intraamniotic infection (n = 78); 3) term not in labor (n = 25); and 4) term with intact membranes in labor (n = 25). MMP-9, and TIMP-1 were measured using sensitive and specific immunoassays. RESULTS: 1) Spontaneous labor at term was associated with a significant increase in MMP-9 but not in TIMP-1.2) Women with preterm labor who delivered prematurely had significantly higher concentrations of MMP-9 but not TIMP-1 in amniotic fluid than those with preterm labor who delivered at term. 3) The concentrations of TIMP-1 decreased with advancing gestational age. In contrast, MMP-9 concentrations did not change with advancing gestational age. CONCLUSIONS: Spontaneous human parturition is associated with specific changes in the enzymatic machinery responsible for extracellular matrix degradation.

A role for matrix metalloproteinase-9 in spontaneous rupture of the fetal membranes.

OBJECTIVES: Preterm premature rupture of fetal membranes is responsible for 30% to 40% of preterm deliveries. Fetal membranes are composed primarily of collagen. Matrix metalloproteinases are enzymes capable of degrading extracellular matrix macromolecules, including collagens. Expression of matrix metalloproteinase-9 (gelatinase B, 92 kd) and its tissue inhibitor (tissue inhibitor of metalloproteinase-1) has been localized in amnion and chorion. The objective of this study was to determine whether rupture of fetal membranes and intrauterine infection are associated with changes in the expression of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1. STUDY DESIGN: Two hundred one women in the following categories had amniotic fluid retrieved: (1) preterm labor and intact membranes in the presence (n = 42) or absence (n = 21) of microbial invasion of the amniotic cavity, (2) preterm premature rupture of the membranes with (n = 29) or without (n = 23) microbial invasion of the amniotic cavity, and (3) term gestation with intact membranes (n = 50) or with premature rupture of the membranes (n = 40). Women in groups 1 and 2 were matched for gestational age at amniocentesis. Microbial invasion of the amniotic cavity was defined by a positive amniotic fluid culture for micro-organisms. Matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 were measured with use of sensitive and specific immunoassays that were validated for amniotic fluid. RESULTS: Spontaneous rupture of membranes at term is associated with a significant increase in the amniotic fluid concentrations of matrix metalloproteinase-9 (premature rupture of membranes, no labor: median 3.9 ng/mL, range 2. 7 to 11.1 ng/mL vs no premature rupture of membranes, no labor: median <0.4 ng/mL, range <0.4 to 22.4 ng/mL; P <.001). Patients with preterm premature rupture of the membranes had higher median matrix metalloproteinase-9 concentrations than those with preterm labor and intact membranes who were delivered at term (7.6 ng/mL, range <0.4 to 230.81 ng/mL vs <0.4 ng/mL, range <0.4 to 1650 ng/mL; P =.06). Women with microbial invasion of the amniotic cavity had higher median matrix metalloproteinase-9 concentrations than did those without microbial invasion regardless of membrane status (preterm labor: 54.5 ng/mL, range <0.4 to 3910 ng/mL vs <0.4 ng/mL, range <0. 4 to 1650 ng/mL; P <.01; preterm premature rupture of membranes: 179. 8 ng/mL, range <0.4 to 611 ng/mL vs 7.6 ng/mL, range <0.4 to 230.81; P <.001). CONCLUSION: Our data support a role for matrix metalloproteinase-9 in the mechanisms responsible for membrane rupture in term and preterm gestations.

[Sensitivity of the parietal cell to the effect of pentagastrin in men living at high altitude]. / Sensibilidad de la célula parietal a la acción de la pentagastrina en el hombre de altura.

This study evaluates the response of the gastric mucosa to pentagastrin in a group (HAG) of 17 andean subjects compared to a similar group of 17 subjects at sea level (LAG). Both groups had normal fundic, body and antrum mucosa as demonstrated by endoscopic and histological means. Each group underwent two assays, the first one with a dose of 6 micrograms/kg and 72 hours later, the second administration with 3 micrograms/kg. In the HAG the acid response expressed as concentration and output was significantly greater with 6 micrograms/kg that with 3 micrograms/kg. In the LAG the difference was not statistically significant. The ANOVA analysis showed, for both determinations, free and total acidity, a different response pattern between the HAG and LAG for both doses. The response of both groups to the 3 micrograms/kg reveals a lesser sensitivity to pentagastrin of the parietal cell in HAG.