Antidepressant Exposure and DNA Methylation: Insights from a Methylome-Wide Association Study.

Importance: Understanding antidepressant mechanisms could help design more effective and tolerated treatments. Objective: Identify DNA methylation (DNAm) changes associated with antidepressant exposure. Design: Case-control methylome-wide association studies (MWAS) of antidepressant exposure were performed from blood samples collected between 2006-2011 in Generation Scotland (GS). The summary statistics were tested for enrichment in specific tissues, gene ontologies and an independent MWAS in the Netherlands Study of Depression and Anxiety (NESDA). A methylation profile score (MPS) was derived and tested for its association with antidepressant exposure in eight independent cohorts, alongside prospective data from GS. Setting: Cohorts; GS, NESDA, FTC, SHIP-Trend, FOR2107, LBC1936, MARS-UniDep, ALSPAC, E-Risk, and NTR. Participants: Participants with DNAm data and self-report/prescription derived antidepressant exposure. Main Outcomes and Measures: Whole-blood DNAm levels were assayed by the EPIC/450K Illumina array (9 studies, N exposed = 661, N unexposed = 9,575) alongside MBD-Seq in NESDA (N exposed = 398, N unexposed = 414). Antidepressant exposure was measured by self- report and/or antidepressant prescriptions. Results: The self-report MWAS (N = 16,536, N exposed = 1,508, mean age = 48, 59% female) and the prescription-derived MWAS (N = 7,951, N exposed = 861, mean age = 47, 59% female), found hypermethylation at seven and four DNAm sites (p < 9.42x10 -8 ), respectively. The top locus was cg26277237 ( KANK1, p self-report = 9.3x10 -13 , p prescription = 6.1x10 -3 ). The self-report MWAS found a differentially methylated region, mapping to DGUOK-AS1 ( p adj = 5.0x10 -3 ) alongside significant enrichment for genes expressed in the amygdala, the "synaptic vesicle membrane" gene ontology and the top 1% of CpGs from the NESDA MWAS (OR = 1.39, p < 0.042). The MPS was associated with antidepressant exposure in meta-analysed data from external cohorts (N studies = 9, N = 10,236, N exposed = 661, f3 = 0.196, p < 1x10 -4 ). Conclusions and Relevance: Antidepressant exposure is associated with changes in DNAm across different cohorts. Further investigation into these changes could inform on new targets for antidepressant treatments. 3 Key Points: Question: Is antidepressant exposure associated with differential whole blood DNA methylation?Findings: In this methylome-wide association study of 16,536 adults across Scotland, antidepressant exposure was significantly associated with hypermethylation at CpGs mapping to KANK1 and DGUOK-AS1. A methylation profile score trained on this sample was significantly associated with antidepressant exposure (pooled f3 [95%CI]=0.196 [0.105, 0.288], p < 1x10 -4 ) in a meta-analysis of external datasets. Meaning: Antidepressant exposure is associated with hypermethylation at KANK1 and DGUOK-AS1 , which have roles in mitochondrial metabolism and neurite outgrowth. If replicated in future studies, targeting these genes could inform the design of more effective and better tolerated treatments for depression.

Chromatin modifier developmental pluripotency associated factor 4 (DPPA4) is a candidate gene for alcohol-induced developmental disorders.

BACKGROUND: Prenatal alcohol exposure (PAE) affects embryonic development, causing a variable fetal alcohol spectrum disorder (FASD) phenotype with neuronal disorders and birth defects. We hypothesize that early alcohol-induced epigenetic changes disrupt the accurate developmental programming of embryo and consequently cause the complex phenotype of developmental disorders. To explore the etiology of FASD, we collected unique biological samples of 80 severely alcohol-exposed and 100 control newborns at birth. METHODS: We performed genome-wide DNA methylation (DNAm) and gene expression analyses of placentas by using microarrays (EPIC, Illumina) and mRNA sequencing, respectively. To test the manifestation of observed PAE-associated DNAm changes in embryonic tissues as well as potential biomarkers for PAE, we examined if the changes can be detected also in white blood cells or buccal epithelial cells of the same newborns by EpiTYPER. To explore the early effects of alcohol on extraembryonic placental tissue, we selected 27 newborns whose mothers had consumed alcohol up to gestational week 7 at maximum to the separate analyses. Furthermore, to explore the effects of early alcohol exposure on embryonic cells, human embryonic stem cells (hESCs) as well as hESCs during differentiation into endodermal, mesodermal, and ectodermal cells were exposed to alcohol in vitro. RESULTS: DPPA4, FOXP2, and TACR3 with significantly decreased DNAm were discovered-particularly the regulatory region of DPPA4 in the early alcohol-exposed placentas. When hESCs were exposed to alcohol in vitro, significantly altered regulation of DPPA2, a closely linked heterodimer of DPPA4, was observed. While the regulatory region of DPPA4 was unmethylated in both control and alcohol-exposed hESCs, alcohol-induced decreased DNAm similar to placenta was seen in in vitro differentiated mesodermal and ectodermal cells. Furthermore, common genes with alcohol-associated DNAm changes in placenta and hESCs were linked exclusively to the neurodevelopmental pathways in the enrichment analysis, which emphasizes the value of placental tissue when analyzing the effects of prenatal environment on human development. CONCLUSIONS: Our study shows the effects of early alcohol exposure on human embryonic and extraembryonic cells, introduces candidate genes for alcohol-induced developmental disorders, and reveals potential biomarkers for prenatal alcohol exposure.

Accuracy of self-reported anthropometric measures - Findings from the Finnish Twin Study.

OBJECTIVE: To determine the accuracy of self-reported height, weight, body mass index (BMI) and waist circumference (WC) compared to the measured values, and to assess the similarity between self-reported and measured values within dizygotic (DZ) and monozygotic (MZ) twin pairs. METHODS: The data on self-reported and measured height, weight and WC values as well as measured hip circumference (HC) were collected from 444 twin individuals (53-67 years old, 60% women). Accuracies between self-reported and measured values were assessed by Pearson's correlation coefficients, Cohen's kappa coefficients and Bland-Altman 95% limits of agreement. Intra-class correlation was used in within-pair analyses. RESULTS: The correlations between self-reported and measured values were high for all variables (r=0.86-0.98), although the agreement assessed by Bland-Altman 95% limits had relatively wide variation. The degree of overestimating height was similar in both sexes, whereas women tended to underestimate and men overestimate their weight. Cohen's kappa coefficients between self-reported and measured BMI categories were high: 0.71 in men and 0.70 in women. Further, the mean self-reported WC was less than the mean measured WC (difference in men 2.5cm and women 2.6cm). The within-pair correlations indicated a tendency of MZ co-twins to report anthropometric measures more similarly than DZ co-twins. CONCLUSIONS: Self-reported anthropometric measures are reasonably accurate indicators for obesity in large cohort studies. However, the possibility of more similar reporting among MZ pairs should be taken into account in twin studies exploring the heritability of different phenotypes.

Subcutaneous adipose tissue gene expression and DNA methylation respond to both short- and long-term weight loss.

BACKGROUND: Few studies have examined both gene expression and DNA methylation profiles in subcutaneous adipose tissue (SAT) during long-term weight loss. Thus, molecular mechanisms in weight loss and regain remain elusive. PARTICIPANTS/METHODS: We performed a 1-year weight loss intervention on 19 healthy obese participants (mean body mass index (BMI) 34.6 kg m-2) and studied longitudinal gene expression (Affymetrix Human Genome U133 Plus 2.0) and DNA methylation (Infinium HumanMethylation450 BeadChip) in SAT at 0, 5 and 12 months. To examine whether weight loss and acquired obesity produce reciprocal profiles, we verified our findings in 26 BMI-discordant monozygotic twin pairs. RESULTS: We found altered expression of 69 genes from 0 to 5' months (short-term) weight loss. Sixty of these genes showed reversed expression in acquired obesity (twins). Altogether 21/69 genes showed significant expression-DNA methylation correlations. Pathway analyses revealed increased high-density lipoprotein-mediated lipid transport characteristic to short-term weight loss. After the fifth month, two groups of participants evolved: weight losers (WLs) and weight regainers (WRs). In WLs five genes were differentially expressed in 5 vs 12 months, three of which significantly correlated with methylation. Signaling by insulin receptor pathway showed increased expression. We further identified 35 genes with differential expression in WLs from 0 to 12 months (long-term) weight loss, with 20 showing opposite expression patterns in acquired obesity, and 16/35 genes with significant expression-DNA methylation correlations. Pathway analyses demonstrated changes in signal transduction, metabolism, immune system and cell cycle. Notably, seven genes (UCHL1, BAG3, TNMD, LEP, BHMT2, EPDR1 and OSTM1) were found to be downregulated during both short- and long-term weight loss. CONCLUSIONS: Our study indicates short- and long-term weight loss influences in transcription and DNA methylation in SAT of healthy participants. Moreover, we demonstrate that same genes react in an opposite manner in weight loss and acquired obesity.

Neuregulin signaling pathway in smoking behavior.

Understanding molecular processes that link comorbid traits such as addictions and mental disorders can provide novel therapeutic targets. Neuregulin signaling pathway (NSP) has previously been implicated in schizophrenia, a neurodevelopmental disorder with high comorbidity to smoking. Using a Finnish twin family sample, we have previously detected association between nicotine dependence and ERBB4 (a neuregulin receptor), and linkage for smoking initiation at the ERBB4 locus on 2q33. Further, Neuregulin3 has recently been shown to associate with nicotine withdrawal in a behavioral mouse model. In this study, we scrutinized association and linkage between 15 036 common, low frequency and rare genetic variants in 10 NSP genes and phenotypes encompassing smoking and alcohol use. Using the Finnish twin family sample (N=1998 from 740 families), we detected 66 variants (representing 23 LD blocks) significantly associated (false discovery rate P<0.05) with smoking initiation, nicotine dependence and nicotine withdrawal. We comprehensively annotated the associated variants using expression (eQTL) and methylation quantitative trait loci (meQTL) analyses in a Finnish population sample. Among the 66 variants, we identified 25 eQTLs (in NRG1 and ERBB4), 22 meQTLs (in NRG3, ERBB4 and PSENEN), a missense variant in NRG1 (rs113317778) and a splicing disruption variant in ERBB4 (rs13385826). Majority of the QTLs in blood were replicated in silico using publicly available databases, with additional QTLs observed in brain. In conclusion, our results support the involvement of NSP in smoking behavior but not in alcohol use and abuse, and disclose functional potential for 56 of the 66 associated single-nucleotide polymorphism.

Gene expression profile of subcutaneous adipose tissue in BMI-discordant monozygotic twin pairs unravels molecular and clinical changes associated with sub-types of obesity.

BACKGROUND: Subcutaneous adipose tissue (SAT) undergoes major changes in obesity, but little is known about the whole-genome scale patterns of these changes or about their variation between different obesity sub-groups. We sought to compare how transcriptomics profiles in SAT differ between monozygotic (MZ) co-twins who are discordant for body mass index (BMI), whether the profiles vary between twin pairs and whether the variation can be linked to clinical characteristics. METHODS: We analysed the transcriptomics (Affymetrix U133 Plus 2.0) patterns of SAT in young MZ twin pairs (n=26, intra-pair difference in BMI >3 kg m-2, aged 23-36), from 10 birth cohorts of adult Finnish twins. The clinical data included measurements of body composition, insulin resistance, lipids and adipokines. RESULTS: We found 2108 genes differentially expressed (false discovery rate (FDR)<0.05) in SAT of the BMI-discordant pairs. Pathway analyses of these genes revealed a significant downregulation of mitochondrial oxidative pathways (P<0.05) and upregulation of inflammation pathways (P<0.05). Hierarchical clustering of heavy/lean twin ratios, representing effects of acquired obesity in the transcriptomics data, revealed three sub-groups with different molecular profiles (FDR<0.05). Analyses comparing these sub-groups showed that, in the heavy co-twins, downregulation of the mitochondrial pathways, especially that of branched chain amino acid degradation was more evident in two clusters while and upregulation of the inflammatory response was most evident in the last, presumably the unhealthiest cluster. High-fasting insulin levels and large adipocyte diameter were the predominant clinical characteristic of the heavy co-twins in this cluster (Bonferroni-adjusted P<0.05). CONCLUSIONS: This is the first study in BMI-discordant MZ twin pairs reporting sub-types of obesity based on both SAT gene expression profiles and clinical traits. We conclude that a decrease in mitochondrial BCAA degradation and an increase in inflammation in SAT co-occur and associate with hyperinsulinemia and large adipocyte size in unhealthy obesity.

DNA methylation and gene expression patterns in adipose tissue differ significantly within young adult monozygotic BMI-discordant twin pairs.

BACKGROUND: Little is known about epigenetic alterations associated with subcutaneous adipose tissue (SAT) in obesity. Our aim was to study genome-wide DNA methylation and gene expression differences in SAT in monozygotic (MZ) twin pairs who are discordant for body mass index (BMI). This design completely matches lean and obese groups for genetic background, age, gender and shared environment. METHODS: 14We analyzed DNA methylome and gene expression from SAT, together with body composition (magnetic resonance imaging/spectroscopy) and glucose tolerance test, lipids and C-reactive protein from 26 rare BMI-discordant (intrapair difference in BMI ⩾3 kg m(-2)) MZ twin pairs identified from 10 birth cohorts of young adult Finnish twins. RESULTS: We found 17 novel obesity-associated genes that were differentially methylated across the genome between heavy and lean co-twins. Nine of them were also differentially expressed. Pathway analyses indicated that dysregulation of SAT in obesity includes a paradoxical downregulation of lipo/adipogenesis and upregulation of inflammation and extracellular matrix remodeling. Furthermore, CpG sites whose methylation correlated with metabolically harmful fat depots (intra-abdominal and liver fat) also correlated with measures of insulin resistance, dyslipidemia and low-grade inflammation, thus suggesting that epigenetic alterations in SAT are associated with the development of unhealthy obesity. CONCLUSION: This is the first study in BMI-discordant MZ twin pairs reporting genome-wide DNA methylation and expression profiles in SAT. We found a number of novel genes and pathways whose methylation and expression patterns differ within the twin pairs, suggesting that the pathological adaptation of SAT to obesity is, at least in part, epigenetically regulated.

Biotin-dependent functions in adiposity: a study of monozygotic twin pairs.

BACKGROUND: Biotin acts as a coenzyme for carboxylases regulating lipid and amino-acid metabolism. We investigated alterations of the biotin-dependent functions in obesity and the downstream effects of biotin restriction in adipocytes in vitro. SUBJECTS: Twenty-four monozygotic twin pairs discordant for body mass index (BMI). Mean within-pair difference (heavy-lean co-twin, Δ) of BMI was 6.0 kg m(-2) (range 3.1-15.2 kg m(-)(2)). METHODS: Adipose tissue (AT) DNA methylation, gene expression of AT and adipocytes, and leukocytes (real-time quantitative PCR), serum biotin, C-reactive protein (CRP) and triglycerides were measured in the twins. Human adipocytes were cultured in low and control biotin concentrations and analyzed for lipid droplet content, mitochondrial morphology and mitochondrial respiration. RESULTS: The gene expression levels of carboxylases, PCCB and MCCC1, were upregulated in the heavier co-twins' leukocytes. ΔPCCB (r=0.91, P=0.0046) and ΔMCCC1 (r=0.79, P=0.036) correlated with ΔCRP within-pairs. Serum biotin levels were lower in the heavier (274 ng l(-1)) than in the lean co-twins (390 ng l(-1), P=0.034). ΔBiotin correlated negatively with Δtriglycerides (r=-0.56, P=0.045) within-pairs. In AT, HLCS and ACACB were hypermethylated and biotin cycle genes HLCS and BTD were downregulated (P<0.05). Biotin-dependent carboxylases were downregulated (ACACA, ACACB, PCCB, MCCC2 and PC; P<0.05) in both AT and adipocytes of the heavier co-twins. Adipocytes cultured in low biotin had decreased lipid accumulation, altered mitochondrial morphology and deficient mitochondrial respiration. CONCLUSIONS: Biotin-dependent functions are modified by adiposity independent of genetic effects, and correlate with inflammation and hypertriglyceridemia. Biotin restriction decreases lipid accumulation and respiration, and alters mitochondrial morphology in adipocytes.

Mechanisms of inactivation of MLH1 in hereditary nonpolyposis colorectal carcinoma: a novel approach.

Mutations in the DNA mismatch repair gene MLH1 are a major cause of hereditary nonpolyposis colorectal cancer (HNPCC). No mutant phenotype is observed before the wild-type (wt) allele is somatically inactivated in target tissue. We addressed the mechanisms of MLH1 inactivation in 25 colorectal (CRC) and 32 endometrial cancers (ECs) from MLH1 mutation carriers (Mut1, in-frame genomic deletion; Mut2, out-of-frame splice site mutation; Mut3, missense mutation). By a quantitative method, matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF), utilizing four intragenic single nucleotide polymorphisms and mutations, loss of heterozygosity (LOH) was present in 31/57 (54.4%) of tumors. The wt allele displayed LOH more often than the mutant allele (23/57 vs 8/57, P=0.006). For Mut1, LOH was more frequent in CRC than EC (10/11 vs 1/13, P<0.0001), whereas Mut2 and Mut3 displayed opposite LOH pattern. Moreover, although wt LOH predominated in CRC irrespective of the predisposing mutation, LOH often affected the mutant allele in EC from Mut2 and Mut3 carriers (6/19, 31.6%). MLH1 promoter methylation, which reflected a more widespread hypermethylation tendency, occurred in 4/55 (7.3%) of tumors and was inversely associated with LOH. In conclusion, the patterns of somatic events (LOH and promoter methylation) differ depending on the tissue and germline mutation, which may in part explain the differential tumor susceptibility of different organs in HNPCC. MALDI-TOF provides a novel approach for the detection and quantification of LOH.

BRAF screening as a low-cost effective strategy for simplifying HNPCC genetic testing.

BACKGROUND: According to the international criteria for hereditary non-polyposis colorectal cancer (HNPCC) diagnostics, cancer patients with a family history or early onset of colorectal tumours showing high microsatellite instability (MSI-H) should receive genetic counselling and be offered testing for germline mutations in DNA repair genes, mainly MLH1 and MSH2. Recently, an oncogenic V600E hotspot mutation within BRAF, a kinase encoding gene from the RAS/RAF/MAPK pathway, has been found to be associated with sporadic MSI-H colon cancer, but its association with HNPCC remains to be further clarified. METHODS: BRAF-V600E mutations were analysed by automatic sequencing in colorectal cancers from 206 sporadic cases with MSI-H and 111 HNPCC cases with known germline mutations in MLH1 and MSH2. In addition, 45 HNPCC cases showing abnormal immunostaining for MSH2 were also analysed. RESULTS: The BRAF-V600E hotspot mutation was found in 40% (82/206) of the sporadic MSI-H tumours analysed but in none of the 111 tested HNPCC tumours or in the 45 cases showing abnormal MSH2 immunostaining. CONCLUSIONS: Detection of the V600E mutation in a colorectal MSI-H tumour argues against the presence of a germline mutation in either the MLH1 or MSH2 gene. Therefore, screening of these mismatch repair (MMR) genes can be avoided in cases positive for V600E if no other significant evidence, such as fulfilment of the strict Amsterdam criteria, suggests MMR associated HNPCC. In this context, mutation analysis of the BRAF hotspot is a reliable, fast, and low cost strategy which simplifies genetic testing for HNPCC.

Towards efficient pollution control in the Baltic Sea: an anatomy of current failure with suggestions for change.

We study the Baltic Sea countries' declaration to reduce nutrient loads by 50% in each country in an ecological-economic model. The model consists of country-based abatement-cost functions, and transfer coefficients describing how phosphorus and nitrogen flow from one country to another, as estimated in a hydrological model of the Baltic Sea. We show that for nitrogen in particular the overall abatement costs of the current policy are much higher and that the benefits are more uneven than under a cost-efficient policy. Consequently, one can expect that countries with high marginal abatement costs have the least incentives to follow the agreement and to invest in nitrogen abatement. This is also confirmed by our data. Therefore, we suggest and outline a joint implementation policy to promote cost efficiency and to increase incentives for investments.

Ethanol modulates cortical activity: direct evidence with combined TMS and EEG.

The motor cortex of 10 healthy subjects was stimulated by transcranial magnetic stimulation (TMS) before and after ethanol challenge (0.8 g/kg resulting in blood concentration of 0.77 +/- 0.14 ml/liter). The electrical brain activity resulting from the brief electromagnetic pulse was recorded with high-resolution electroencephalography (EEG) and located using inversion algorithms. Focal magnetic pulses to the left motor cortex were delivered with a figure-of-eight coil at the random interstimulus interval of 1.5-2.5 s. The stimulation intensity was adjusted to the motor threshold of abductor digiti minimi. Two conditions before and after ethanol ingestion (30 min) were applied: (1) real TMS, with the coil pressed against the scalp; and (2) control condition, with the coil separated from the scalp by a 2-cm-thick piece of plastic. A separate EMG control recording of one subject during TMS was made with two bipolar platinum needle electrodes inserted to the left temporal muscle. In each condition, 120 pulses were delivered. The EEG was recorded from 60 scalp electrodes. A peak in the EEG signals was observed at 43 ms after the TMS pulse in the real-TMS condition but not in the control condition or in the control scalp EMG. Potential maps before and after ethanol ingestion were significantly different from each other (P = 0.01), but no differences were found in the control condition. Ethanol changed the TMS-evoked potentials over right frontal and left parietal areas, the underlying effect appearing to be largest in the right prefrontal area. Our findings suggest that ethanol may have changed the functional connectivity between prefrontal and motor cortices. This new noninvasive method provides direct evidence about the modulation of cortical connectivity after ethanol challenge.

EEG responses to combined somatosensory and transcranial magnetic stimulation.

OBJECTIVES: To investigate a possible interaction between sensory processing and transcranial magnetic stimulation (TMS), an experimental set-up permitting multichannel EEG measurements was used. METHODS: A somatosensory stimulus was delivered to the right wrist, while single-pulse TMS was applied to the contralateral somatosensory cortex, either concurrent with the somatosensory stimulus or 10 ms after it. A control condition served to mimic the sound of TMS without actually resulting in brain stimulation. RESULTS: An enhancement of the P25 component of the somatosensory-evoked potential (SEP) was consistently observed for TMS concurrent with somatosensory stimulus. The effect was topographically specific to the EEG recording sites below the TMS coil, i.e. above the somatosensory cortex contralateral to the stimulated peripheral nerve. CONCLUSIONS: The results can be interpreted (1) as an indication of local interaction between the somatosensory-evoked cortical activity and TMS-evoked activity or (2) as support of a relationship between the background EEG and the evoked potential (EP), this relationship being 'disrupted' by TMS.

Coil design for real and sham transcranial magnetic stimulation.

Transcranial magnetic stimulation (TMS) can be used to excite the human cortex noninvasively. TMS also activates scalp muscles and sensory receptors; additionally, the loud sound from the stimulating coil activates auditory pathways. These side effects complicate the interpretation of the results of TMS studies. For control experiments, we have designed a coil that can produce both real and sham stimulation without moving the coil. The sham TMS is similar to the real TMS, except for the different relative direction of the currents in the two loops of the figure-of-eight coil. While the real TMS elicited activation of hand muscles, sham TMS had no such effect; however, the auditory-evoked potentials were similar.

Separation of contamination caused by coil clicks from responses elicited by transcranial magnetic stimulation.

Transcranial magnetic stimulation (TMS) is accompanied with loud clicks that evoke auditory responses in the brain, confounding several types of TMS studies. We investigated the effects of these clicks with high-resolution EEG by applying TMS pulses at 3 magnitudes, with the coil placed either at 10 or 50 mm over the subjects' vertex and recording event-related potentials (ERPs). The clicks were found to elicit a positively displaced response at 150-250 ms post-TMS. Furthermore, clicks were found to interact with simultaneously presented auditory sinewave stimuli, resulting in an amplitude decrease in the auditory N1 response.

C-reactive protein in uncomplicated parturients delivered by cesarean section.

BACKGROUND: The purpose of this study was to determine normal values and evaluate clinical variables associated with postoperative C-reactive protein serum concentration after uncomplicated abdominal delivery. METHODS: C-reactive protein serum concentrations were determined serially by a quantitative immunoturbidimetric assay in 479 clinically non-infected parturients undergoing uncomplicated Cesarean section, before operation and during the first week of puerperium. Associations between clinical variables and maximal postoperative C-reactive protein levels were examined by variance analysis. RESULTS: The preoperative values were significantly higher in parturients from whom samples were obtained after the onset of labor and/or rupture of the membranes (16 +/- 10 (s.d.) mg/l) compared with the preoperative values obtained from parturients operated with no labor and intact membranes (12 +/- 7 (s.d.) mg/l) (p < 0.0001). An overall significant increase was observed in C-reactive protein concentrations after delivery, peaking on the second postoperative day, compared with the preoperative values. In the parturients operated upon after the onset of labor or ruptured membranes, postoperative values were 24% higher up to the sixth postoperative day than in parturients operated upon with intact membranes and no labor. The onset of labor, duration of operation, the number of vaginal examinations and the duration of internal monitoring before operation were associated with the maximal postoperative C-reactive protein concentrations, but they explained only 9% of postoperative CRP level variation. CONCLUSIONS: The wide range of serum C-reactive protein concentrations during puerperium complicates its use as a simple marker in obstetric postoperative complications. However, after normal uncomplicated operative delivery, C-reactive protein concentrations rapidly decrease towards the baseline after the second to third postoperative day.

Amniotic fluid U. urealyticum colonization: significance for maternal peripartal infections at term.

The aim of this study was to examine the relationship between maternal peripartal infectious morbidity and amniotic fluid colonization by U. urealyticum. Amniotic fluid specimens for bacterial and mycoplasmal cultures were obtained by aspiration at nonelective cesarean section from 98 pregnant women. Amniotic fluid cultures revealed micro-organisms in 70 (71%) parturients and U. urealyticum was the most frequently isolated species, detected in the specimens from 38 (39%) women. In the total study population, the prevalence of clinically defined infectious morbidity was 26%, including 8 (8%) patients with chorioamnionitis, 11 (11%) with endometritis, 5 (5%) with urinary tract infection, and 4 (4%) with wound infection. A significant association between the amniotic fluid U. urealyticum colonization and increased maternal peripartal infectious morbidity was observed (p < 0.0027; relative risk 4.1; 95% confidence limits 1.6 to 10.7). Amniotic fluids positive for U. urealyticum were significantly more often positive for other potentially pathogenic bacteria (p < 0.0024) and particularly for Bacteroides spp. (p < 0.0074). Our results support the existence of an association between amniotic fluid U. urealyticum invasion and maternal infectious morbidity. However, U. urealyticum was not usually isolated alone from amniotic fluids but combined with other pathogenic bacteria, the severity of infections were not enhanced when U. urealyticum was present and parturients with diagnosed infections managed well without specific antibiotic against U. urealyticum. Hence, it appears that the significance of U. urealyticum in maternal infections in this study population was mainly based on its interactions with other bacteria.

Cesarean delivery. Microbial colonization in amniotic fluid.

OBJECTIVE: To determine the frequency, clinical significance and causative factors behind intraamniotic microbial colonization in uninfected parturients at the time of cesarean delivery. STUDY DESIGN: Amniotic fluid specimens for bacterial and mycoplasmal cultures were obtained by direct aspiration at cesarean section from 251 pregnant women (24-43 completed weeks) who had no clinical infection at the time of the operation. The symptoms of maternal infection were followed postoperatively for the first week of the puerperium. RESULTS: The prevalence of amniotic fluid microbial invasion was 29% (72/251). In patients not in labor and with intact membranes, it was 13% (20/158); in patients in labor and with intact membranes, 23% (5/22); and in those with ruptured membranes, 66% (47/71). The most common species isolated were Ureaplasma urealyticum, Lactobacillus species and coagulase-negative staphylococci. In the total 251 patients, clinically evident postoperative endometritis was observed in 6 (2%) and wound infection in 10 (4%). In patients operated on and with intact membranes, no risk factors were found as regards amniotic fluid microbial colonization. In patients operated on after rupture of the membranes, the only significant risk factor as regards amniotic fluid microbial invasion was use of an internal monitor before the operation (P < .0003) (relative risk 10.7, 95% confidence limit 2.9-39.4). The relative risk of postoperative endometritis was 2.3 (95% confidence limit 1.3-4.3) in patients with microbial invasion of the amniotic cavity as compared to patients without invasion. The corresponding risk value for post-operative wound infection was 1.4 (95% confidence limit 0.6-3.1). CONCLUSION: Though the incidence of microbial invasion of the amniotic fluid before surgery was unexpectedly high, its clinical significance as regards maternal puerperal morbidity appeared to be low. The use of internal monitoring during labor was the only significant risk factor as regards amniotic fluid microbial colonization in patients operated on after membrane rupture.

Amniotic fluid leukocytes and leukocyte esterase activity in parturients delivered by caesarean section.

Amniotic fluid specimens from parturients undergoing caesarean delivery were examined for leukocytes and leukocyte esterase activity, as well as for amniotic fluid bacteria and mycoplasmas by cultivation. The aim of this study was to evaluate the intrauterine environment in noninfected parturients and the associations between leukocyte test results, amniotic fluid microbial colonization and postcaesarean endometritis. Samples were obtained by direct aspiration at operation from 289 parturients with no clinical intrauterine infection. Among the total study population, leukocytes were found in 41% of the amniotic fluid samples by Gram staining and in 39% of the samples by the leukocyte esterase activity test. Leukocytes and leukocyte esterase activity were observed significantly more often in the amniotic fluids of parturients undergoing operation after onset of labour or ruptured membranes compared with those operated upon with intact membranes and no labour (p < 0.0001 and p < 0.0001). Of 255 amniotic fluid cultures, microbial colonization was observed in 82 (32%) parturients. Positive results in the leukocyte tests were associated significantly with amniotic fluid microbial colonization among parturients who underwent operation with intact membranes, or who underwent operation after rupture of the membranes and had cervical dilatation of < 5 cm at the operation (Gram stain: p < 0.0001; leukocyte esterase: p < 0.003). If cervical dilatation was > or = 5 cm, no such association was observed. In the population studied, endometritis developed in 2% and 4% of the parturients with positive test results. Thus, neither the presence of leukocytes nor detected leukocyte esterase activity were predictive of subsequent postoperative endometritis. In the detection of amniotic fluid microbial colonization, the tests functioned best in non-laboring parturients with intact membranes and in those operated on at the early stage of labour.