Amniotic fluid cathepsin-G in pregnancies complicated by the preterm prelabor rupture of membranes

Objective: The aim of this study was to evaluate the amniotic fluid cathepsin-G concentrations in women with preterm prelabor rupture of membranes (PPROM) based on the presence of the microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI).

Methods: A total of 154 women with singleton pregnancies complicated by PPROM were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis. Amniotic fluid cathepsin-G concentrations were assessed by ELISA. MIAC was determined using a non-cultivation approach. IAI was defined as an amniotic fluid bedside interleukin-6 concentration?≥?745?pg/mL.

Results: Women with MIAC had higher amniotic fluid cathepsin-G concentrations than women without MIAC (with MIAC: median 82.7?ng/mL, versus without MIAC: median 64.7?ng/mL; p?=?0.0003). Women with IAI had higher amniotic fluid cathepsin-G concentrations than women without this complication (with IAI: median 103.0?ng/mL, versus without IAI: median 66.2?ng/mL; p?p?Conclusions: The presence of either microbial-associated or sterile IAI was associated with increased amniotic fluid cathepsin-G concentrations in pregnancies complicated by PPROM. Amniotic fluid cathepsin-G appears to be a potential marker of IAI.     

Amniotic fluid clusterin in pregnancies complicated by the preterm prelabor rupture of membranes

Objective: The aim of this study was to evaluate clusterin concentrations in amniotic fluid in pregnancies complicated by preterm prelabor rupture of membranes (PPROM) with respect to the presence of the microbial invasion of the amniotic cavity (MIAC), intra-amniotic inflammation (IAI) and microbial-associated IAI.

Methods: One hundred thirty-six women with singleton pregnancies complicated by PPROM were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis. Amniotic fluid clusterin concentrations were assessed by enzyme-linked immunosorbent assay. MIAC was determined by a non-cultivation approach. IAI was defined as an amniotic fluid bedside interleukin-6 concentration?≥745?pg/mL. Microbial-associated IAI was characterized as the presence of both MIAC and IAI.

Result: Women with MIAC, IAI and microbial-associated IAI had lower amniotic fluid clusterin concentrations than women without these complications (with MIAC: median 1314?ng/mL versus without MIAC: median 1633?ng/mL, p?=?0.003; with IAI: median 1281?ng/mL versus without IAI: median 1575?ng/mL, p?=?0.04; with microbial associated-IAI: median 1220?ng/mL versus without microbial-associated IAI: median 1575?pg/mL; p?=?0.008). A week negative correlation between amniotic fluid clusterin concentrations and gestational age at sampling was revealed (rho=??0.30; p?=?0.0005).

Conclusions: The presence of MIAC, IAI and microbial-associated IAI was characterized by lower amniotic fluid clusterin concentrations in pregnancies complicated by PPROM.     

Cervical fluid interleukin 6 and intra-amniotic complications of preterm prelabor rupture of membranes

Objective: To determine if cervical fluid interleukin (IL)-6 concentrations in women with preterm prelabor rupture of membranes (PPROM) allows identification of microbial invasion of the amniotic cavity (MIAC) and/or intra-amniotic inflammation (IAI).

Methods: One hundred forty-four women with singleton pregnancies complicated by PPROM were included in this prospective cohort study. Cervical and amniotic fluids were collected at the time of admission and concentrations of IL-6 were measured using an ELISA and point-of-care test, respectively. Cervical fluid was obtained using a Dacron polyester swab and amniotic fluid was obtained by transabdominal amniocentesis. MIAC was diagnosed based on a positive PCR result for Ureaplasma species, M. hominis, and/or C. trachomatis and/or by positivity for the 16?S rRNA gene. IAI was defined as amniotic fluid point-of-care IL-6 concentrations ≥745?pg/mL. The women were assigned to four subgroups based on the presence of MIAC and/or IAI: microbial-associated IAI (both MIAC and IAI), sterile IAI (IAI alone), MIAC alone, and without either MIAC or IAI.

Results: (1) Women with microbial-associated IAI had higher cervical fluid IL-6 concentrations (median 560?pg/mL) than did women with sterile IAI (median 303?pg/mL; p?=?.001), women with MIAC alone (median 135?pg/mL; p?=?.0004), and women without MIAC and IAI (median 180?pg/mL; p?=?.0001). (2) No differences were found in cervical fluid IL-6 concentrations among women with sterile IAI, with MIAC alone, and without MIAC and IAI. (3) A positive correlation was observed between cervical fluid IL-6 concentrations and the amount of Ureaplasma species in amniotic fluid (copies DNA/mL; rho?=?0.57, p?p?Conclusions: The presence of microbial-associated IAI is associated with the highest cervical fluid IL-6 concentrations. Cervical IL-6 can be helpful in the identification of microbial-associated IAI.     

Cervical fluid calreticulin and cathepsin-G in pregnancies complicated by preterm prelabor rupture of membranes

Objective: The study aimed to determine the cervical calreticulin and cathepsin-G concentrations in pregnancies complicated by preterm prelabor rupture of membranes (PPROM) with respect to the presence of microbial invasion of the amniotic cavity (MIAC) and intra-amniotic inflammation (IAI).

Methods: Eighty women with singleton pregnancies complicated by PPROM were included in this study. Cervical and amniotic fluids were obtained at the time of admission, and concentrations of calreticulin and cathepsin-G in cervical fluid were determined using ELISA. The MIAC was defined as a positive PCR analysis for Ureaplasma species, Mycoplasma hominis, and/or Chlamydia trachomatis and/or by positivity for the 16S rRNA gene. IAI was defined as amniotic fluid bedside IL-6 concentrations ≥745?pg/mL

Result: Neither women with MIAC nor with IAI had different cervical fluid concentrations of calreticulin (with MIAC: median 18.9?pg/mL vs. without MIAC: median 14.7?pg/mL, p?=?0.28; with IAI: median 14.3?pg/mL vs. without IAI: median 15.6?pg/mL, p?=?0.57;) or of cathepsin-G (with MIAC: median 30.7?pg/mL vs. without MIAC: median 24.7?pg/mL, p?=?0.28; with IAI: median 27.3?pg/mL vs. without IAI: median 25.1?pg/mL, p?=?0.80) than women without those complications. No associations between amniotic fluid IL-6 concentrations, gestational age at sampling, and cervical fluid calreticulin and cathepsin-G concentrations were found.

Conclusions: Cervical fluid calreticulin and cathepsin-G concentrations did not reflect the presence of MIAC or IAI in women with PPROM.     

Intraamniotic inflammation and umbilical cord blood interleukin-6 concentrations in pregnancies complicated by preterm prelabor rupture of membranes

Objective: To evaluate umbilical cord blood interleukin (IL)-6 concentrations and the occurrence of fetal inflammatory response syndrome (FIRS) with respect to microbial invasion of the amniotic cavity (MIAC) and/or intraamniotic inflammation (IAI) in pregnancies complicated by preterm prelabor rupture of membranes (PPROM).

Methods: One-hundred-eighty-eight women with singleton pregnancies complicated by PPROM between gestational ages of 24?+?0 and 36?+?6 weeks were included in the study. Blood samples were obtained by venipuncture from the umbilical cord after the delivery of the newborn. The umbilical cord blood IL-6 concentrations were evaluated using ELISA kits. FIRS was defined as umbilical cord blood IL-6?>?11?pg/mL.

Result: Women with MIAC and IAI had higher IL-6 concentrations than women without these complications (with MIAC: median 18.1?pg/mL versus without MIAC: median 5.8; p?<?0.0001; with IAI: median 32.9?pg/mL, versus without IAI: median 5.8; p?<?0.0001). Women with IAI with MIAC and women with IAI without MIAC had the highest umbilical cord blood IL-6 concentrations (medians: 32.6 and 39.4?pg/mL) and rates of FIRS (78% and 67%).

Conclusions: IAI was associated with the highest umbilical cord blood IL-6 concentrations and rate of FIRS independent of the presence or absence of MIAC.     

Amniotic fluid prostaglandin E2 in pregnancies complicated by preterm prelabor rupture of the membranes

Objective: To determine amniotic fluid prostaglandin E2 concentrations in women preterm prelabor rupture of the membranes (PPROM) with respect to microbial invasion of the amniotic cavity (MIAC), intraamniotic inflammation (IAI), microbial-associated IAI, histological chorioamnionitis, and short-term neonatal morbidity.

Methods: One hundred forty-five women with singleton pregnancies were included in this study. Amniotic fluid samples were obtained by transabdominal amniocentesis and were assayed for prostaglandin E2 concentrations by ELISA. IAI was defined as amniotic fluid interleukin-6 >745?pg/mL. Microbial-associated IAI was defined as the presence of both MIAC and IAI.

Result: No differences in prostaglandin E2 concentrations were found between women with and without MIAC (p?=?0.27). Women with IAI (p?=?0.0008) and microbial-associated IAI (p?=?0.01) had higher prostaglandin E2 concentrations than women without these complications. Women with histological chorioamnionitis had higher prostaglandin E2 concentrations only in crude analysis (p?=?0.02), but not after adjustment for gestational age at sampling (p?=?0.10). No associations between amniotic fluid prostaglandin E2 concentrations and the selected conditions of severe neonatal morbidity were found.

Conclusions: The intraamniotic inflammatory response either to infectious or to non-infectious stimulus, but not MIAC per se, seems to be a main factor associated with the elevation of the amniotic fluid PGE2 concentrations in women with PPROM.     

Amniotic fluid soluble Toll-like receptor 2 in pregnancies complicated by preterm prelabor rupture of membranes

Objective: To determine amniotic fluid soluble Toll-like receptor 2 (sTLR2) levels in PPROM according to the presence of microbial invasion of the amniotic cavity (MIAC), histological chorioamnionitis (HCA), and both these conditions. To test the cutoff level of 222.7?ng/mL, as proposed in our previous study, in order to distinguish women with both MIAC and HCA. Methods: 169 women with a gestational age between 24+0 and 36+6 weeks were included in a prospective cohort study. Amniocenteses were performed, and sTLR2 in the amniotic fluid were determined using ELISA. Results: Women with MIAC had higher sTLR2 levels (median 113.2?ng/mL) than those without MIAC (median 47.1 ng/mL; p < 0.0001). Women with HCA did not have a higher sTLR2 level (median 52.6?ng/mL) compared with women without HCA (median 47.1?ng/mL; p = 0.23). Women with both MIAC and HCA had higher sTLR2 levels (median: 311.3?ng/mL) than other women (17.5?ng/mL; p < 0.0001). The cutoff level 222.7?ng/mL had a sensitivity of 63%, a specificity of 98%, and a likelihood ratio of 40.3 for the prediction of both MIAC and HCA. Conclusions: Amniotic fluid sTLR2 is a promising predictor of both MIAC and HCA with high specificity in PPROM.     

Amniotic fluid concentrations of soluble scavenger receptor for hemoglobin (sCD163) in pregnancy complicated by preterm premature rupture of the membranes and histologic chorioamnionitis

Objective.?To determine changes in the amniotic fluid, soluble form of scavenger receptor for hemoglobin (sCD163) concentrations during advancing gestation, and in patients with preterm premature rupture of membranes (PPROM) complicated by histological chorioamnionitis were studied.

Methods.?One hundred and fifty-two women with singleton pregnancies were enrolled. The concentration of sCD163 in amniotic fluid was determined using sandwich enzyme immunoassay technique.

Results.?Women in the midtrimester had a significantly higher median amniotic fluid sCD163 concentration than those at term not in labor (308 ng/ml vs. 217 ng/ml; p?=?0.04). Patients with PPROM and histological chorioamnionitis had a higher median amniotic fluid sCD163 level than those with PPROM without histological chorioamnionitis (885 ng/ml vs. 288 ng/ml; p?<?0.0001).

Conclusions.?Amniotic fluid sCD163 concentrations decrease with advancing gestation. Amniotic fluid sCD163 concentrations are significantly higher in women with PPROM between 24 and 36 gestational weeks with histological chorioamnionitis than those without histological signs of inflammation.     

Periodontal disease and intra-amniotic complications in women with preterm prelabor rupture of membranes

Objective: Periodontal disease is frequently suggested as a possible causal factor for preterm delivery. The link between periodontal disease and preterm delivery is a possible translocation of periopathogenic bacteria to the placenta and amniotic fluid as well as a systemic response to this chronic inflammatory disease. However, there is a lack of information on whether there is an association between clinical periodontal status in women with preterm prelabor rupture of membranes (PPROM) and the presence of microbial invasion of the amniotic cavity (MIAC) and intra-amniotic inflammation (IAI). Therefore, the main aim of this study was to evaluate the incidence and severity of periodontal disease in women with PPROM. The secondary aim was to characterize an association between periodontal status and the presence of intra-amniotic PPROM complications (MIAC and/or IAI).

Materials and methods: Seventy-eight women with PPROM at gestational ages between 24?+?0 and 36?+?6 weeks were included in this study. The samples of amniotic fluid were obtained at admission via transabdominal amniocentesis, and amniotic fluid interleukin (IL)-6 concentrations were determined using a point-of-care test. All women had a full-mouth recording to determine the periodontal and oral hygiene status. Probing pocket depth and clinical attachment loss were measured at four sites on each fully erupted tooth.

Results: In total, 45% (35/78) of women with PPROM had periodontal disease. Mild, moderate, and severe periodontal disease was present in 19% (15/78), 19% (15/78), and 6% (5/78) of women, respectively. The presence of MIAC and IAI was found in 28% (22/78) and 26% (20/78) of women, respectively. Periopathogenic bacteria (2?×?Streptococcus intermedius and 1?×?Fusobacterium nucleatum) was found in the amniotic fluid of 4% (3/78) of women. There were no differences in periodontal status between women with MIAC and/or IAI and women without these intra-amniotic complications.

Conclusions: The presence of MIAC and IAI was not related to the periodontal status of women with PPROM.     

Cervical and vaginal fluid soluble Toll-like receptor 2 in pregnancies complicated by preterm prelabor rupture of membranes

Objective: To determine the cervical and vaginal fluid soluble Toll-like receptor-2 (sTLR2) levels in pregnancies complicated by preterm prelabor rupture of membranes (PPROM) and their correlation to microbial invasion of the amniotic cavity (MIAC) and/or histological chorioamnionitis (HCA).

Methods: Sixty-eight women with singleton pregnancies complicated by PPROM were included in this study. Cervical and vaginal fluid was collected at the time of admission, and levels of sTLR2 in the cervical and vaginal fluid were determined using enzyme-linked immunosorbent assay.

Result: Women with MIAC and both MIAC and HCA did not have different cervical and vaginal fluid sTLR2 levels compared to those without MIAC and without both MIAC and HCA. Women with HCA had higher cervical fluid sTLR2 levels in crude analysis (with HCA: median 11.6?pg/mL versus without HCA: median 5.5?pg/mL; p?=?0.04) but not after adjustment for gestational age at sampling (p?=?0.19). No difference in vaginal fluid sTLR2 levels between women with and without HCA was found. A positive correlation between cervical and vaginal fluid sTLR2 levels was identified (rho?=?0.54; p?Conclusions: Cervical and vaginal fluid sTLR2 levels did not reflect the presence of MIAC and/or HCA.     

Microbial invasion and histological chorioamnionitis upregulate neutrophil-gelatinase associated lipocalin in preterm prelabor rupture of membranes

Our recent exploratory proteomic study suggested increased levels of neutrophil-gelatinase associated lipocalin (P80188, NGAL_HUMAN) due to microbial invasion of the amniotic cavity (MIAC) and histological chorioamnionitis (HCA) in women with preterm prelabor rupture of the membranes. In this study, we verified the proteomics findings by assessing the amniotic fluid NGAL by ELISA in the original exploratory cohort. The NGAL level was significantly higher in women positive for both MIAC and HCA compared to women with both conditions ruled out (median 75.1?ng/ml versus 27.9?ng/ml; p?<?0.0001). For independent validation and to assess NGALs potential to stratify women positive for both MIAC and HCA from women in whom at least one of these conditions was absent, we subsequently designed a retrospective replication cohort. Significantly higher NGAL levels were found in women positive for both MIAC and HCA (median 65.9?ng/ml versus 34.2?ng/ml; p?=?0.0061). Significantly higher levels of NGAL were confirmed only in strata below 32 weeks of gestation. Based on the observed likelihood ratio, the best predictive cutoff level (47.1?ng/ml) was evaluated in both cohorts. Data from the verification cohort implied that NGAL is a valuable clinical marker for revealing MIAC leading to HCA; however, this potential was not replicated in the replication cohort.     

Evidence of participation of soluble CD14 in the host response to microbial invasion of the amniotic cavity and intra-amniotic inflammation in term and preterm gestations

Objective: Endotoxin has been implicated in the mechanism responsible for the setting of infection in preterm labor. To exert its biological effects, endotoxin binds to a circulating protein known as lipopolysaccharide binding protein (LBP) and presents endotoxin monomers to CD14, which may be a membrane-bound receptor or a soluble molecule. The endotoxin- LBP-CD14 complex interacts with Toll-like receptor 4 and other regulatory proteins leading to cellular activation and an inflammatory response. The purpose of this study was to determine whether microbial invasion of the amniotic cavity (MIAC)/intra-amniotic inflammation (both preterm and term) and parturition at term are associated with changes in the amniotic fluid and umbilical plasma soluble concentrations of CD14 (sCD14). Study design: Amniotic fluid was retrieved by amniocentesis from 88 patients in the following groups: group 1, preterm labor with intact membranes with MIAC/intra-amniotic inflammation (n = 18) and without these conditions (n = 26); group 2, term gestations not in labor without MIAC/intra-amniotic inflammation (n = 11), in labor without MIAC/intra-amniotic inflammation (n = 12) and in labor with MIAC/intra-amniotic inflammation (n = 13); and group 3, patients who underwent genetic amniocentesis at mid-trimester (n = 8). A sample of cord blood was obtained after delivery in all patients except those in group 3. sCD14 was assayed with a sensitive and specific immunoassay. Non-parametric statistics were used for analysis. A p value of < 0.05 was considered significant. Results: sCD14 was detectable in 97% (85/88) of the amniotic fluid samples. Amniotic fluid sCD14 concentrations were lower in patients at term than in the mid-trimester of pregnancy (mid-trimester: median 482 ng/ml, range 258-838 ng/ml vs. term no labor: median 7 ng/ml, range 2-274 ng/ml, p = 0.01). Among patients with preterm labor with intact membranes, the median amniotic fluid sCD14 level of patients with MIAC/intra-amniotic inflammation was higher than in patients without these conditions (median 1568 ng/ml, range 98-5887 ng/ml vs. median 645 ng/ml, range 0-3961 ng/ml, respectively; p = 0.01). Among women at term in labor, those with MIAC/intra-amniotic inflammation had a higher median amniotic fluid sCD14 concentration than those without these conditions (median 85 ng/ml, range 2-1113 ng/ml vs. median 17 ng/ml, range 0-186 ng/ml; p = 0.01). MIAC/ intra-amniotic inflammation in women with preterm labor with intact membranes was associated with a higher median umbilical venous plasma sCD14 concentration (median 744 ng/ml, range 0-3620 ng/ml vs. median 0 ng/ml, range 0-2060 ng/ml; p = 0.04). sCD14 was undetectable in plasma from umbilical cords of all neonates born to women at term. An increase in amniotic fluid concentration of sCD14 was observed in cases of intrauterine infection, not only by Gram-negative bacteria, but also Gram-positive bacteria and Ureaplasma spp. Conclusion: sCD14 is a physiological constituent of amniotic fluid, and its concentrations at term are lower than in the mid-trimester. Intrauterine infection/inflammation is associated with a higher median amniotic fluid sCD14 concentration in both preterm and term parturition. Neonates born from mothers with preterm labor with intact membranes and MIAC/intra-amniotic inflammation had a higher median concentration of sCD14 in umbilical cord plasma than those without these conditions. sCD14 concentrations are increased in the amniotic fluid and umbilical cord blood even in the absence of a microbiologically proven Gram-negative infection. CD14 appears to participate in the host response to intrauterine infection even in cases involving genital mycoplasmas.     

Vaginal fluid IL-6 and IL-8 levels in pregnancies complicated by preterm prelabor membrane ruptures

Objective: To determine the vaginal fluid interleukin (IL)-6 and IL-8 concentrations in pregnancies complicated by preterm prelabor rupture of membranes and their correlation to microbial invasion of the amniotic cavity (MIAC) as well as histological chorioamnionitis (HCA).

Methods: Sixty-eight women with singleton pregnancies were included in this study. Vaginal fluid was collected at the time of admission. IL-6 and IL-8 concentrations in the vaginal fluid were determined using ELISA.

Result: Women with MIAC had higher vaginal fluid IL-6 levels compared to those without MIAC (with MIAC: median 374?pg/mL versus without MIAC: median 174?pg/mL; p?=?0.03). IL-8 levels were higher in women with MIAC only in the crude analysis but not after adjustment for gestational age. There was no difference in the IL-6 and IL-8 concentrations between those with and without HCA. Women with both MIAC and HCA had higher IL-6 vaginal fluid levels than those without both MIAC and HCA (with MIAC and HCA: median 466?pg/mL versus without MIAC and HCA: median 178?pg/mL; p?=?0.02). IL-8 levels were higher in women with MIAC and HCA only in the crude analysis but not after adjustment for gestational age.

Conclusions: Vaginal fluid IL-6 but not IL-8 levels reflect the presence of MIAC and both MIAC and HCA.     

Comparison of sVCAM-1 and sICAM-1 levels in maternal serum and vaginal secretion between pregnant women with preterm prelabour ruptures of membranes and healthy pregnant women

Objective: The study aims to evaluate the maternal serum and the vaginal fluid levels of soluble vascular cell adhesion molecule-1 (sVCAM-1) and soluble intercellular adhesion molecular (sICAM-1) in pregnant women complicated by preterm prelabour ruptures of membranes (PPROM).

Materials and methods: The prospective case control study included 34 pregnant women with PPROM and 34 healthy pregnant women. Patients with additional diseases, a smoking habit and vaginal bleeding, as well as those using antibiotics, during the study period were not included in the study. Cervicovaginal fluid and serum samples were taken during the patients’ admission. The demographic data, maternal serum and vaginal fluid sVCAM-1 and sICAM-1, C reactive protein (CRP) and leukocyte counts were noted for all pregnant women included in the study. The sVCAM-1 and sICAM-1 levels were measured by enzyme-linked immunosorbent assay kits.

Results: In pregnant women with PPROM, the serum leukocyte (mean?±?SD =11.41?±?1.067 versus 9.18?±?1.56, p?p?p?=?.06), vaginal sVCAM-1 (median 208.00 versus 140.20?ng/ml, p?=?.014) and sICAM-1 (mean?±?SD 32.32?±?6.49?ng/ml versus 24.87?±?6.79?ng/ml, p?r?=?0.850; p?Conclusion: To the best of our knowledge, this is the first study evaluating the levels of sICAM-1 in maternal serum in pregnant women with PPROM. The maternal serum and vaginal fluid sVCAM-1 and sICAM-1 levels can be used as biochemical markers supporting the PPROM diagnosis because of the increase in both maternal serum and vaginal fluid sVCAM-1 and sICAM-1 levels in pregnant women with PPROM.     

Soluble Toll-like receptor 1 family members in the amniotic fluid of women with preterm prelabor rupture of the membranes

Objective: To determine soluble Toll-like receptor (sTLR) 1, sTLR2 and sTLR6 concentrations in amniotic fluid (AF) of women with preterm prelabor rupture of membranes (PPROM) and if there is an association with microbial invasion of the amniotic cavity and histological chorioamnionitis (HCA). Methods: Cross-sectional study was performed. Forty-two women with singleton PPROM pregnancies at a gestational age between 24 + 0 and 36 + 6 weeks were included in the study (twenty-two women with presence of both microbial invasion of the amniotic cavity and HCA, and 20 women without microbial invasion of the amniotic cavity and HCA). Amniocenteses were performed, and the concentrations of sTLRs were determined by sandwich enzyme-linked immunosorbent assays. Results: Women with microbial invasions of the amniotic cavity and HCA (n = 22) had significantly higher median sTLR1, sTLR2 and sTLR6 levels than those without (n = 20). (20.4 ng/mL vs. 0.44 ng/mL; p < 0.0001, 577.6 ng/mL vs. 60.7 ng/mL; p < 0.0001 and 0.44 ng/mL vs. 0.26 ng/mL; p = 0.02, respectively). Conclusions: Women with microbial invasion of the amniotic cavity and HCA had higher AF sTLR1, 2 and 6 levels.     

Amniotic fluid heat shock protein 70 concentration in histologic chorioamnionitis,term and preterm parturition

Objective. Heat shock protein (HSP) 70, a conserved member of the stress protein family, is produced in almost all cell types in response to a wide range of stressful stimuli, and its production has a survival value. Evidence suggests that extracellular HSP70 is involved in the activation of the innate and adaptive immune response. Furthermore, increased mRNA expression of HSP70 has been observed in human fetal membranes following endotoxin stimulation. This study was conducted to determine the changes in amniotic fluid HSP70 concentrations during pregnancy, term and preterm parturition, intra-amniotic infection (IAI), and histologic chorioamnionitis.

Study design. A cross-sectional study was conducted in 376 pregnant women in the following groups: (1) women with a normal pregnancy who were classified into the following categories: (a) women in the mid-trimester (14–18 weeks) who underwent amniocentesis for genetic indications and delivered normal infants at term (n=72); (b) women at term not in labor (n = 23); and (c) those at term in labor (n = 48). (2) Women with spontaneous preterm labor and intact membranes who were subdivided into the following categories: (a) preterm labor who delivered at term without IAI (n = 42); (b) preterm labor who delivered preterm without IAI (n = 57); and (c) preterm labor and delivery with IAI (n = 30). (3) Women with preterm prelabor rupture of membranes (PROM) with (n = 50) and without (n = 54) IAI. Among patients with preterm labor with intact membranes and preterm PROM who delivered within 72 hours of amniocentesis, placenta, umbilical cord, and chorioamniotic membranes were collected and assessed for the presence or absence of acute inflammatory lesions in the extraplacental membranes (histologic chorioamnionitis) and/or umbilical cords (funisitis). HSP70 concentrations in amniotic fluid were determined using a sensitive and specific immunoassay. Non-parametric statistics were used for analysis. A p value of <0.05 was considered statistically significant.

Results. Immunoreactive HSP70 was detected in 88% (332/376) of amniotic fluid samples. The median amniotic fluid HSP70 concentration was significantly higher in women at term without labor than in those in the mid-trimester (term no labor: median 34.9 ng/mL, range 0–78.1 ng/mL vs. mid-trimester; median 6.6 ng/mL, range 0–20.8 ng/mL; p<0.001). Among patients with spontaneous preterm labor and preterm PROM, those with IAI had a significantly higher median amniotic fluid HSP70 concentration than those without IAI (preterm labor with IAI: median 82.9 ng/mL, range 0–500 ng/mL vs. preterm labor without IAI: median 41.7 ng/mL, range 0–244 ng/mL; p = 0.001; preterm PROM with IAI: median 86.5 ng/mL, range 0–428 ng/mL vs. preterm PROM without IAI: median 55.9 ng/mL, range 14.9–299.9 ng/mL; p = 0.007). There was no significant difference in the median amniotic fluid HSP70 concentration between patients with preterm labor who delivered preterm without IAI and those who delivered at term (p = 0.6). However, among patients with preterm labor without IAI, there was an inverse relationship between amniotic fluid concentration of HSP70 and the amniocentesis-to-spontaneous delivery interval (Spearman's Rho = ?0.26; p = 0.02). Patients with histologic chorioamnionitis/funisitis had a significantly higher median amniotic fluid HSP70 concentration than those without inflammation (inflammation: median 108.7 ng/mL, range 0–500 ng/mL vs. without inflammation: median 67.9 ng/mL, range 7.1–299.9 ng/mL; p = 0.02). Women at term in labor had a median amniotic fluid concentration of HSP70 significantly higher than those not in labor (term in labor: median 60.7 ng/mL, range 0–359.9 ng/mL vs. term not in labor: median 34.9 ng/mL, range 0–78.1 ng/mL; p = 0.02).

Conclusions. Intra-amniotic infection, histologic chorioamnionitis, and term parturition are associated with elevated amniotic fluid HSP70 concentrations. HSP70 plays a role in the host defense mechanism by activating the innate arm of the immune response in women with intrauterine infection. The mechanisms of preterm and term parturition in humans may involve extracellular HSP70.     

Adiponectin in amniotic fluid in normal pregnancy,spontaneous labor at term,and preterm labor: A novel association with intra-amniotic infection/inflammation

Objective.?Adiponectin, an anti-inflammatory and anti-diabetogenic adipokine, has an important regulatory effect on both the innate and adaptive limbs of the immune response. The objective of this study was to determine whether adiponectin is present in amniotic fluid (AF) and if its concentration changes with gestational age, in the presence of labor, and in the presence of intra-amniotic infection (IAI) in patients with spontaneous preterm labor (PTL) and intact membranes.

Study design.?This cross-sectional study included 468 patients in the following groups: (1) women in the mid-trimester of pregnancy (14–18 weeks) who underwent amniocentesis for genetic indications and delivered a normal neonate at term (n?=?52); (2) normal pregnant women at term with (n?=?49) and without (n?=?41) spontaneous labor; (3) patients with an episode of PTL and intact membranes who were classified into: (a) PTL who delivered at term (n?=?149); (b) PTL who delivered preterm (<37 weeks gestation) without IAI (n?=?108); and (c) PTL with IAI (n?=?69). Adiponectin concentration in AF was determined by ELISA.

Results.?(1) The median AF adiponectin concentration at term was significantly higher than in the mid-trimester (35.6?ng/ml, interquartile range [IQR] 26.4–52.7 vs. 29.9?ng/ml, IQR 19.9–35.2; p?=?0.01); (2) among women with PTL and intact membranes, the median AF adiponectin concentration was significantly higher in patients with IAI than in those without IAI who delivered either at term (54.3?ng/ml, 39.0–91.8 vs. 50.1?ng/ml, 33.2–72.8; p?=?0.02) or preterm (47.6?ng/ml, 32.6–74.6; p?=?0.01); and (3) among women at term, there was no significant difference in the median AF adiponectin concentration between those with and without labor (33.7?ng/ml, IQR 21.7–53.9 vs. 35.6?ng/ml, IQR 26.4–52.7; respectively p?=?0.5).

Conclusions.?(1) Adiponectin is a physiologic constituent of AF; and (2) adiponectin concentrations in AF are increased significantly with advancing gestation and in the presence of IAI. Collectively, these findings suggest that adiponectin plays a dynamic role in normal gestation and in the presence of IAI.     

Secreted phospholipase A2 is increased in meconium-stained amniotic fluid of term gestations: potential implications for the genesis of meconium aspiration syndrome

Abstract

Background: Meconium-stained amniotic fluid (MSAF) represents the passage of fetal colonic content into the amniotic cavity. Meconium aspiration syndrome (MAS) is a complication that occurs in a subset of infants with MSAF. Secreted phospholipase A₂ (sPLA₂) is detected in meconium and is implicated in the development of MAS. The purpose of this study was to determine if sPLA₂ concentrations are increased in the amniotic fluid of women in spontaneous labor at term with MSAF.

Materials and methods: This was a cross-sectional study of patients in spontaneous term labor who underwent amniocentesis (n?=?101). The patients were divided into two study groups: (1) MSAF (n?=?61) and (2) clear fluid (n?=?40). The presence of bacteria and endotoxin as well as interleukin-6 (IL-6) and sPLA₂ concentrations in the amniotic fluid were determined. Statistical analyses were performed to test for normality and bivariate analysis. The Spearman correlation coefficient was used to study the relationship between sPLA₂ and IL-6 concentrations in the amniotic fluid.

Results: Patients with MSAF have a higher median sPLA₂ concentration (ng/mL) in amniotic fluid than those with clear fluid [1.7 (0.98–2.89) versus 0.3 (0–0.6), p?<?0.001]. Among patients with MSAF, those with either microbial invasion of the amniotic cavity (MIAC, defined as presence of bacteria in the amniotic cavity), or bacterial endotoxin had a significantly higher median sPLA₂ concentration (ng/mL) in amniotic fluid than those without MIAC or endotoxin [2.4 (1.7–6.0) versus 1.7 (1.3–2.5), p?<?0.05]. There was a positive correlation between sPLA₂ and IL-6 concentrations in the amniotic fluid (Spearman Rho?=?0.3, p?<?0.05).

Conclusion: MSAF that contains bacteria or endotoxin has a higher concentration of sPLA₂, and this may contribute to induce lung inflammation when meconium is aspirated before birth.     

Idiopathic vaginal bleeding during pregnancy as the only clinical manifestation of intrauterine infection

Objective.?To determine the frequency and clinical significance of microbial invasion of the amniotic cavity (MIAC) in patients with vaginal bleeding in the absence of placenta previa, preterm labor or preterm premature rupture of membranes (PROM).

Study design.?This retrospective cohort study included patients who presented with vaginal bleeding between 18 and 35 weeks, and underwent an amniocentesis shortly after admission for the assessment of the microbiologic status of the amniotic cavity and/or fetal lung maturity. Amniotic fluid was cultured for aerobic and anaerobic bacteria, as well as genital mycoplasmas. Patients presenting with preterm labor, preterm PROM, placenta previa, overt placental abruption, and an intrauterine device in situ were excluded, as well as those with local cervical bleeding. MIAC was defined as a positive amniotic fluid culture. Analysis was conducted with non-parametric statistics.

Results.?One hundred and fourteen patients met the entry criteria. MIAC was detected in 14% of cases (16/114). Patients with vaginal bleeding and a gestational age <?28 weeks at the time of amniocentesis had a significantly higher frequency of MIAC than those with a gestational age ??28 weeks [25% (13/52) vs. 4.8% (3/62), respectively; p?<?0.01]. Ureaplasma urealyticum was the microorganism most frequently isolated from the amniotic fluid. Except for one case admitted at 33 weeks, all patients with MIAC?had an early preterm delivery ??32 weeks. Patients with vaginal bleeding and MIAC?had a shorter procedure-to-delivery interval than those without MIAC [MIAC, median survival 19 days (95% CI 10–27 days) vs. no MIAC, median survival 50 days (95% CI 37–62 days); p?<?0.0001]. Patients with vaginal bleeding and MIAC?had a significantly lower gestational age at delivery and lower birth weight than those with vaginal bleeding and negative amniotic fluid cultures (for gestational age, median 25 weeks, range 21–33 weeks vs. median 37 weeks, range 19–42 weeks, respectively; p?<?0.01, and for birth weight, median 750?grams, range 520–1820?grams vs. 2800?grams, range 520–4880?grams, respectively; p?<?0.01), as well as a higher frequency of subsequent preterm PROM [81.3% (13/16) vs. 9.2% (9/98); p?<?0.01].

Conclusions.?MIAC was detected in 14% of patients with ‘idiopathic’ vaginal bleeding and was associated with subsequent preterm PROM and early preterm delivery. Vaginal bleeding may be the only clinical manifestation of MIAC, and it predisposes to adverse outcome.