Nevertheless, assembly of complex genomes remains challenging, in part due to the presence of dispersed repeats which introduce ambiguity during genome reconstruction. Transposable elements (TEs) can be particularly problematic, especially for TE families exhibiting high sequence identity, high copy number, or complex genomic arrangements. While TEs strongly affect genome function and evolution, most current de novo assembly approaches cannot resolve long, identical, and abundant families of TEs. Here, we applied a novel Illumina technology called TruSeq synthetic long-reads, which are generated through highly-parallel library preparation and local assembly of short read data and which achieve lengths of 1.5–18.5 Kbp with an extremely low error rate  achieving an N50 contig size of 69.7 Kbp and covering 96.9% of the euchromatic chromosome arms of the current reference genome. TruSeq synthetic long-read technology enables placement of individual TE copies in their proper genomic locations as well as accurate reconstruction of TE sequences. We entirely recovered and accurately placed 4,229 (77.8%) of the 5,434 annotated transposable elements with perfect identity to the current reference genome. As TEs are ubiquitous features of genomes of many species, TruSeq synthetic long-reads, and likely other methods that generate long-reads, offer a powerful approach to improve de novo assemblies of whole genomes.High-throughput DNA sequencing technologies have revolutionized genomic analysis, including the  Tremendous advances in DNA sequencing technology, computing power, and assembly approaches, have enabled the assembly of genomes of thousands of species from the sequences of DNA fragments, but several challenges still remain. All assembly approaches are based on the assumption that similar sequence reads originate from the same genomic region, thereby allowing the reads to be overlapped and merged to reconstruct the underlying genome sequence de novo assembly of the genome of the colonial tunicate, Botryllus schlosseriDrosophila melanogaster genome, our study uses comparison to a high-quality reference to evaluate the application of synthetic long-read technology for de novo assembly. While future work will be required to investigate the use of the technology for resolving polymorphism in outbred species, our work specifically focuses on the accuracy of assembly of repetitive DNA sequences.By spanning regions of high diversity and regions of high identity, the use of longer input sequences can also help overcome problems posed by both polymorphism and repeats. The recent application of Pacific Biosciences' (PacBio) long-read technology to resolve complex segmental duplications In some species, repetitive DNA accounts for a large proportion of the total genome size, for example comprising more than half of the human genome Drosophila 12 Genomes Consortium Drosophila simulans reported that the majority of TE sequences (identified by homology to D. melanogaster TEs) were contained in fragmented contigs less than 500 bp in length Despite their biological importance, knowledge of TE dynamics is hindered by technical limitations resulting in the absence of certain TE families from genome assemblies. Many software packages for whole genome assembly use coverage-based heuristics, distinguishing putative unique regions from putative repetitive regions based on deviation from average coverage  represents the only high-throughput long-read . Polymerase chain reaction (PCR) is used to amplify molecules within wells, followed by highly-parallel Nextera-based fragmentation and barcoding of individual wells. DNA from all wells is then pooled and sequenced on the Illumina HiSeq 2000 platform. Data from individual wells are demultiplexed in silico according to the barcode sequences. Synthetic long-reads are then assembled from the short reads using an assembly pipeline that accounts for properties of the molecular biology steps used in the library preparation  were successfully mapped to the reference genome, with 90.88%  mapping uniquely and 96.36%  having at least one alignment with a MAPQ score In order to evaluate the accuracy of TruSeq synthetic long-reads, we mapped sequences to the reference genome of base see . Along wbase see , & 1D. MWe quantified the average depth of coverage of the mapped synthetic long-reads for each reference chromosome arm. We observed 33.3–35.2× coverage averages of the euchromatic chromosome arms of each major autosome  D. melanogasterTo perform We used the program QUAST To gain more insight about the alignment on a per-chromosome basis, we further investigated the NUCmer alignment of the 5,598 assembled contigs to the reference genome. Upon requiring high stringency alignment  overlapping gap coordinates  and recently diverged (mean of 0.0086 substitutions per base), and are therefore difficult to assemble In order to gain further insight about the content of alignment gaps, we applied RepeatMasker Manual curation of the alignment also revealed that assembly is particularly poor in regions of tandem arrangement of TE copies from the same family, a result that is expected because repeats will be present within individual wells during library preparation  had both boundaries contained in a single contig of our assembly aligned to the reference genome, with 4,229 (77.8%) perfectly reconstructed based on length and sequence identity.Repeats can induce three common classes of mis-assembly. First, tandem repeats may be erroneously collapsed into a single copy. While the accuracy of TruSeq synthetic long-reads are advantageous in this case, such elements may still complicate assembly because they are likely to be present within a single molecule (and therefore a single well) during library preparation. Second, large repeats may fail to be assembled because reads do not span the repeat anchored in unique sequence, a situation where TruSeq synthetic long-reads are clearly beneficial. Finally, highly identical repeat copies introduce ambiguity into the assembly graph, which can result in breaks or repeat copies placed in the wrong location in the assembly. As TEs are diverse in their organization, length, copy number, GC content, and divergence, we decided to assess the accuracy of TE assembly with respect to each of these factors. We therefore compared reference TE sequences to the corresponding sequences in our assembly. Because a naive mapping approach could result in multiple reference TE copies mapping to the same location in the assembly, our approach was specifically designed to restrict the search space within the assembly based on the NUCmer global alignment see . Of the In order to test which properties of TE copies affected faithful reconstruction, we fit a generalized linear mixed model (GLMM) with a binary response variable indicating whether or not each TE copy was perfectly assembled. We included TE length as a fixed effect because we expected assembly to be less likely in cases where individual synthetic reads do not span the length of the entire TE copy. We also included GC content of the interval, including each TE copy and 1 Kbp flanking sequence on each side, as a fixed effect to capture library preparation biases as well as correlated aspects of genomic context (e.g. gene rich vs. gene poor). TE divergence estimates assembly . Longer assembly . We foune family .Tc1 family have an average of 0.039 substitutions per base with respect to the 947 bp canonical sequence, yet 25 of 26 annotated copies were assembled with 100% accuracy  increases rapidly with coverage up to approximately 10×, increasing only marginally at higher synthetic long-read coverage  of the reference genome, and containing 84.2% of annotated genes with perfect sequence identity. Using standard assembly size  and correctness metrics based on alignment to the reference genome, we demonstrated that our assembly is comparable to other D. melanogaster. This was beneficial to our study because it allowed us to attribute TE sequence differences to divergence among TE copies. For an outbred species, distinguishing between divergence among TE copies and polymorphism within TE copies complicates this analysis. For the same reasons, polymorphism in general is a key feature limiting non-haploid genome assemblies, as algorithms must strike a balance between merging polymorphic haplotypes and splitting slightly diverged repeat copies to produce a haploid representation of the genome sequence. Assemblies of other Drosophila from the modENCODE project (using 454 reads) demonstrate the importance of polymorphism, achieving N50 contig lengths up to 436 Kbp for inbred species, but only 19 Kbp for the species that could not be inbred Botryllus schlosseri, but assessment of assembly quality was difficult as no high-quality reference genome exists for comparison. Likewise, recent work demonstrated the utility of the same technology for assigning polymorphisms to individual haplotypes de novo resolution of polymorphism in the absence of a reference genome. Future work will be required to systematically evaluate the ability of synthetic long-read data to help resolve polymorphism in outbred species.An important caveat in the interpretation of our results is the fact that the assembly was performed on a highly-inbred strain of de novo assembly generally fail to assemble and place long, abundant, and identical TE copies with respect to the rest of the assembly. For example, the majority of TE-containing contigs in the improved draft assembly of Drosophila simulans (which combined Illumina short read and Sanger data) were shorter than 500 bp Drosophila 12 Genomes Consortium D. melanogaster, a species with particularly active, abundant, and identical TEs, our results suggest that synthetic long-read technology can empower studies of TE dynamics for many non-model species.Our study demonstrates that TruSeq synthetic long-reads enable accurate assembly of complex, highly-repetitive TE sequences. Previous approaches to y; cn, bw, sp substrain of D. melanogaster using corrected PacBio data achieved an N50 contig length of 15.3 Mbp and closed two of the remaining gaps in the euchromatin of the Release 5 reference sequence Alongside this synthetic long-read technology, several third-generation sequencing platforms have been developed to sequence long molecules directly. One such technology, Oxford Nanopore  sequencing de novo assembly fare poorly on long, abundant, and recently diverged repetitive elements, including some families of TEs. The resulting assemblies offer a biased perspective of evolution of complex genomes. In addition to accurately recovering 96.9% of the euchromatic portion of the high-quality reference genome, our assembly using TruSeq synthetic long-reads accurately placed and perfectly reconstructed the sequence of 84.2% of genes and 77.8% of TEs. Improvements to de novo assembly, facilitated by TruSeq synthetic long-reads and other long-read technologies, will empower comparative analyses that will enlighten the understanding of the dynamics of repeat elements and genome evolution in general.Most current approaches to D. melanogaster genome sequence at the time of the preparation of this manuscript (Release 5.56) and corresponding TE annotations were downloaded from FlyBase The latest release of the y; cn, bw, sp strain of D. melanogaster using a standard ethanol precipitation-based protocol. Approximately 50 adult individuals, both males and females, were pooled for the extraction to achieve sufficient gDNA quantity for preparation of multiple TruSeq synthetic long-read libraries.High molecular weight DNA was separately isolated from pooled samples of the Six libraries were prepared by Illumina's FastTrack Service using the TruSeq synthetic long-read technology, previously known as Moleculo. To produce each library, extracted gDNA is sheared into approximately 10 Kbp fragments, ligated to amplification adapters, and then diluted to the point that each well on a 384-well plate contains approximately 200 molecules, representing approximately 1.5% of the entire genome. These pools of DNA are then amplified by long-range PCR. Barcoded libraries are prepared within each well using Nextera-based fragmentation and PCR-mediated barcode and sequencing adapter addition. The libraries undergo additional PCR amplification if necessary, followed by paired-end sequencing on the Illumina HiSeq 2000 platform.Based on the unique barcodes, assembly is performed among molecules originating from a single well, which means that the likelihood of individual assemblies containing multiple members of gene families  is greatly reduced. The assembly process, which is described in detail in the Supplemental Materials , consistD. melanogaster libraries prepared by Illumina, we used BLASTN (version 2.2.28To estimate the degree of contamination of the D. melanogaster reference genome as single-end reads using BWA-MEM Drosophila Genetic Reference Panel (DGRP) D. melanogaster genome sequences, estimated as 0.141 from DGRP. Meanwhile, the number of mismatches that overlap with SNP sites in DGRP  approaches when applied to large datasets. Nevertheless, for datasets with moderate sequencing depth (such as TruSeq synthetic long-read libraries), OLC approaches can be computationally tractable and tend to be less affected by both repeats and sequencing errors than de Bruijn graph-based algorithms. Likewise, many modern Bruijn graph-based assemblers simply do not permit reads exceeding arbitrary length cutoffs. We therefore elected to use the Celera Assembler D. melanogasterMost recent approaches to As TruSeq synthetic long-reads share some characteristics with consensus-corrected PacBio reads, we applied Celera Assembler parameters recommended for these PacBio data to take advantage of the read length and low error rate For the down-sampled assemblies with lower coverage, we based the expected coverage on the average euchromatic autosomal depth of coverage of 34We aligned the contigs produced by the Celera Assembler to the reference genome sequence using the NUCmer pipeline (version 3.23) lmeY family of TEs, which was recently annotated (Release 5.56). The response variable was represented by a binary indicator denoting whether or not the entire length of the TE was accurately assembled. This model assumed a binomial error distribution with a logit link function. TE copy length, GC content (including 1 Kbp flanking regions on each side), divergence , number of high-identity Click here for additional data file.


Pulmonary hyalinizing granuloma is a very rare benign condition, which usually manifests as solitary and sometimes as multiple pulmonary nodules. Deposition of immune complexes in the lung parenchyma due to hypersensitivity reactions is implicated in the etiology of pulmonary hyalinizing granuloma. A 59-year-old female patient who presented to our clinic with complaints of chest pain and cough had bilateral, multiple, and rounded lesions with regular margins suggesting metastatic lung disease. A transthoracic needle biopsy of the nodule was performed in the left pulmonary anterior segment. Biopsy showed no malignancy. Since no diagnosis was made by the biopsy, the patient underwent a video-assisted thoracic surgery. The wedge biopsy reported pulmonary hyalinizing granuloma. We aimed to present the diagnosis and treatment stages of our patient who was diagnosed with pulmonary hyalinizing granuloma in the light of literature review. Pulmonary hyalinizing granuloma (PHG), which was first described in 1977 by Engleman et al., has been usually reported as individual cases in the world literature [A 59-year-old female patient presented to our clinic with complaints of chest pain and cough. The physical examination and blood tests showed no pathological finding. The patient had no history of tuberculosis or prior lung disease. Additionally, she had well-regulated type 2 diabetes. Computed tomography (CT) showed pulmonary nodules with regular margins and lobulated contours scattered throughout both lungs, the largest measuring 14 × 12 mm in size located in the laterobasal segment of the lower lobe, which suggested metastatic lung disease . Due to PHG is a rare benign lung disease. Generally, regardless of race or gender, the age range of PHG is from 19 to 77 years, with a mean age of 43 years at presentation . Twenty-In conclusion, PHG can be misdiagnosed as several benign and malignant diseases. Therefore, pulmonary hyalinizing granuloma should be considered in differential diagnosis of lesions suggesting metastatic lung carcinoma, particularly without any primary focus as in our case.


Small wireless trunk accelerometers have become a popular approach to unobtrusively quantify human locomotion and provide insights into both gait rehabilitation and sports performance. However, limited evidence exists as to which trunk accelerometry measures are suitable for the purpose of detecting movement compensations while running, and specifically in response to fatigue. The aim of this study was therefore to detect deviations in the dynamic center of mass (CoM) motion due to running-induced fatigue using tri-axial trunk accelerometry. Twenty runners aged 18–25 years completed an indoor treadmill running protocol to volitional exhaustion at speeds equivalent to their 3.2 km time trial performance. The following dependent measures were extracted from tri-axial trunk accelerations of 20 running steps before and after the treadmill fatigue protocol: the tri-axial ratio of acceleration root mean square (RMS) to the resultant vector RMS, step and stride regularity (autocorrelation procedure), and sample entropy. Running-induced fatigue increased mediolateral and anteroposterior ratios of acceleration RMS (p < .05), decreased the anteroposterior step regularity (p < .05), and increased the anteroposterior sample entropy (p < .05) of trunk accelerometry patterns. Our findings indicate that treadmill running-induced fatigue might reveal itself in a greater contribution of variability in horizontal plane trunk accelerations, with anteroposterior trunk accelerations that are less regular from step-to-step and are less predictable. It appears that trunk accelerometry parameters can be used to detect deviations in dynamic CoM motion induced by treadmill running fatigue, yet it is unknown how robust or generalizable these parameters are to outdoor running environments. Given the repetitive nature of running, it is generally recommended that a running bout be stopped at a point before an individual’s typical level of fatigue causes adverse biomechanical effects and subsequent injury . Indeed,In contrast to the lower extremity which has been well studied ,7,8, theIn comparison to traditional 3D motion capture methods, a single tri-axial trunk accelerometer offers a valid –17 and r2) during an incremental running protocol to exhaustion. However, they  of participant characteristics from the remaining 20 participants are represented in Table 2. In the fatigued condition, runners significantly increased their CoM displacements and range for mediolateral and anteroposterior directions, but not for the vertical direction. An exemplary stabilogram of changes in horizontal sacral marker trajectory for one participant can be seen in Fig 2A (pre-fatigue) and Fig 2B (fatigued).The effect of running fatigue on sacral marker trajectory variables can be seen in Table 3. RMS values increased significantly for all three axes with fatigue. In contrast, the RATIO of acceleration RMS increased in the mediolateral and anteroposterior directions but did not change in the vertical direction. A typical example of horizontal plane accelerations can be seen for the same aforementioned participant in Fig 2C (pre-fatigue) and Fig 2D (fatigued). Step regularity of accelerations decreased in the anteroposterior direction. No significant changes were detected for stride regularity of accelerations in any axis, or step frequency. Sample entropy of accelerations increased significantly in the anteroposterior direction.The effect of running fatigue on trunk acceleration variables can be seen in The purpose of the current study was to detect directional changes in dynamic CoM motion in relation to running-induced fatigue based on tri-axial accelerometry. We focused on extracting a select number of measures from a single trunk-mounted accelerometer during speed controlled running until volitional fatigue. Furthermore, the trunk accelerometry measures used in this study were chosen on the basis that 1) they have previously been used to successfully detect limitations in walking gait function, and 2) were calculated relatively easily that avoided algorithms for accurate peak detection or stride-to-stride partitioning. In support of our hypothesis, our main findings indicate that running-induced fatigue resulted in a higher contribution of variability in horizontal plane trunk accelerations, as evidenced by higher mediolateral and anteroposterior ratios of acceleration RMS. In partial support of our hypothesis, only anteroposterior acceleration patterns became less regular and less predictable once running fatigue was induced, as supported by lower step regularity and larger sample entropy values.Running-induced fatigue greatly influenced the variability of horizontal plane trunk accelerations. This increase was expected, since running-induced fatigue may introduce horizontal compensations in trunk kinematics  that mayRunning-induced fatigue did not influence the step or stride regularity measures except for the anteroposterior step regularity. Step regularity has more recently been used as a symmetry index  that repSample entropy of trunk accelerations was assessed to gain insight in the non-linearity of the acceleration waveforms. Sample entropy is a useful measure, since it examines every time point of the time-series being analyzed. To the best of our knowledge, sample entropy has never been used to elucidate on running movement patterns. We found that sample entropy values for trunk acceleration patterns were higher and thus less predictable in all three axes when runners were fatigued, although only significant for the anteroposterior direction. This may be partially substantiated by the findings of Meardon et al.,  who founThe indoor treadmill protocol we selected was chosen on the basis of selecting a controlled steady state environment. An important question to ask is whether certain measures that changed with fatigue, such as anteroposterior step regularity for example would change even more in an overground situation. Firstly, the speed of running is naturally more variable overground than compared to treadmill running, even under controlled conditions . Secondl


This study investigated whether neuroticism impacts neuronal, physiological, and emotional responses to infant crying by investigating blood-oxygenation-level dependent (BOLD) responses using functional magnetic resonance imaging (fMRI) in a large sample of healthy women (N = 102) with simultaneous skin conductance recordings. Participants were repeatedly exposed to a video clip that showed crying infants and emotional responses  were assessed after every video clip presentation. Increased BOLD signal during the perception of crying infants was found in brain regions that are associated with emotional responding, the amygdala and anterior insula. Significant BOLD signal decrements  were found in the fusiform gyrus, middle temporal gyrus, superior temporal gyrus, Broca’s homologue on the right hemisphere,  amygdala, and hippocampus. Individuals with high neuroticism showed stronger activation in the amygdala and subgenual anterior cingulate cortex (sgACC) when exposed to infant crying compared to individuals with low neuroticism. In contrast to our prediction we found no evidence that neuroticism impacts fMRI-based measures of habituation. Individuals with high neuroticism showed elevated skin conductance responses, experienced more irritation, and perceived infant crying as more unpleasant. The results support the hypothesis that individuals high in neuroticism are more emotionally responsive, experience more negative emotions, and may show enhanced cognitive control during the exposure to infant distress, which may impact infant-directed behavior.Newborns and infants communicate their needs and physiological states through crying and emotional facial expressions. Little is known about individual differences in responding to infant crying. Several theories suggest that people vary in their environmental sensitivity with some responding generally more and some generally less to environmental stimuli. Such differences in environmental sensitivity have been associated with personality traits, including  Sensory-Processing Sensitivity [Infant crying is considered to be a powerful communication signal ensuring infants`survival  and it isitivity  that corsitivity ), one ofsitivity  and hencsitivity . High nesitivity . Neurotisitivity . Neurotisitivity . Researcsitivity  and thatsitivity . More resitivity  and may habituate to repeated stimuli presentations—that is brain responses decline upon repeated stimulus presentations. Research has shown that habituation in response to repeatedly presented emotional stimuli include the  amygdala [Previous studies underscores the importance of investigating temporal dynamics when studying affective processing using neuroimaging methods ,22. Studamygdala ,23,25no experience in professional child care, and investigated the following hypotheses: First, we expected that the exposure to infants crying evokes activation in brain regions that are associated with emotional responding. Second, we expected habituation of the blood-oxygenation-level dependent (BOLD) signal during the repeated exposure to infant crying in cortical and subcortical brain regions such as in the laterobasal amygdala. Third, we hypothesized that high-neuroticism individuals show during the repeated exposure to infant crying more activation in sgACC, which is involved in cognitive control during distressing emotions. Fourth, we expected that individuals with high neuroticism experience more negative emotions , exhibit greater skin conductance responses, and show greater amygdala activation during infant crying, because they are emotionally more responsive. Finally, we hypothesized that women high in neuroticism demonstrate less habituation on the neuronal, the physiological (measured by skin conductance recordings), as well as on the emotional level  in response to repeated infant crying.This study investigated whether neuroticism impacts neuronal, physiological and emotional responses to infant crying in healthy women using fMRI with simultaneous skin conductance recordings. This study repeatedly presented a video clip that showed crying infants. Participants`emotional responses were assessed by immediate ratings of valence, arousal, and irritation after every video clip presentation. Valence and arousal are considered to be two basic dimensions of emotional experience . A studyexclusion criteria using a checklist, comprising the following criteria: (1) Participants with children and experience in professional childcare at the time of investigation, (2) a history of seizure or head injury, (3) the use of medication that can influence the test results, (3) visual or auditory problems that cannot be corrected, (4) MRI incompatible implants, (5) pregnancy, (6) claustrophobia, (7) a history of psychiatric or neurological disorders. The ethics committee for medical research in Basel, Switzerland approved the study. Before participation, subjects gave their written informed consent.One-hundred-and-two healthy women were recruited from the University of Basel, Switzerland and participated in the fMRI experiment . Five participants were excluded from fMRI data analysis because of excessive motion . Three participants were excluded from the analysis of skin conductance responses because of technical difficulties during recoding. Psychological measures and subjective affective ratings were analyzed from all individuals. Participants were screened for familiarized with the experimental setting with a video clip that showed laughing infants (LI). That video clip was shown five times. Subsequently, a video clip with crying infants (CI) was presented 33 times including three video clip presentations that were varied for an attention-control task . This video clip was shown five times. This phase was used to distinguish habituation from sensory and motor fatigue [Participants were given 30 min to adjust to the laboratory setting. During this time they received instructions for the experiment. Subsequently, subjects were placed in the MR scanner and were asked to complete the state anxiety scale with 20 items ). Subseqlsewhere ). This p fatigue . ParticiNEO Five Factor Inventory [Center for Epidemiologic Studies Depression Scale (CES-D) [To asses neuroticism, all participants completed the German version of the nventory . This qu (CES-D)  which as (CES-D) .Skin conductance was continuously recorded from the thenar and hypothenar palm of the immobilized, left hand in parallel to the fMRI recording. Data were saved at a sampling rate of 200 Hz using the BIOPAC MP150 system skin conductance module  with MR-conditional, disposable electrodes (EL509) filled with isotonic gel . Electrode cables were grounded and passed through an RF filter panel and stimulus timing information was recorded via TTL pulses from the MR scanner to synchronize fMRI and physiological data analysis.Functional images were acquired on a 3T scanner . Image acquisition started with a localizer, a reference scan for the distortion correction, and a magnetization-prepared rapid-acquisition gradient echo (MPRAGE) sequence of 7-min duration . Functional images were obtained using a multislice gradient echo planar imaging (EPI) method. Each volume consisted of 44 transversal slices  3000 ms, echo time (TE) 35 ms, 90° flip angle). An accurate registration of the functional images was accomplished by online correction of the functional image data for geometric distortions .http://www.fil.ion.ucl.ac.uk/spm/). Preprocessing consisted of realignment, and normalization, and smoothing. All functional images were normalized into standard stereotaxic space of the Montreal Neurological Institute (MNI) template. The images were smoothed using a 9 mm full-width-at-half-maximum (FWHM) Gaussian kernel to minimize the effects of individual variations in anatomy and to improve the signal-to-noise ratio. A high-pass filter with a cut-off at 1/128 Hz was applied before parameter estimation. First level modeling: Regressors were the timing information of the video clips and the timing information of the evaluation period after every video clip presentation. Regressors were modeled with a boxcar function convolved with a canonical hemodynamic response function. The first model included the following regressors: 9 for the video clips  and one for the evaluation period after each film clip. Additional regressors were included to model the six head-movement parameters obtained during realignment. For each individual a contrast image crying infants>baseline (that is the time periods during which subjects passively viewed the fixation cross without stimulus presentation) was analyzed. To analyze response decrements the 30 crying-infant video clip presentations were grouped into 6 consecutive blocks and habituation was modeled as a linear change across these groups (duration 35 min). The attention-control variant of the CI video was not included in the statistical analyses. Second level group analyses: Brain areas with significant response decrements (habituation) during crying-infant presentations were tested at p < 0.05, family wise error (FWE) corrected for multiple comparisons, cluster size ≥ 15 voxels. The contrast perception of crying infants > baseline was tested at p < 0.05, FWE corrected, cluster size ≥ 15 voxels. To test whether individuals with high neuroticism differ in comparison to individuals with low neuroticism in terms of amygdala and sgACC activation a two-sample t-test was applied and tested at p < 0.05, FWE small volume correction. A small volume correction was performed because the amygdala and sgACC were included in our a priori hypothesis based on previous research showing that both brain regions are associated with neuroticism [Data analysis was performed using SPM5/12 (roticism . The medroticism .skin conductance responses (SCR) waveforms was conducted using ANSLAB software (http://www.sprweb.org). Recordings were visually inspected and periods of signal loss were manually excluded (approx. 1% of video epochs). For each video presentation, the level of SCR was assessed as the pre-film baseline (2 s) to peak difference for the largest deflection in the 0–15 s video window [Offline data analysis of o window ; values o window ,39 resulLaugh_1; Familiarization), six mean values (thirty video clips averaged across each of five) were obtained with crying children , and finally one mean value was obtained by averaging across five video clip presentations with laughing children presented at the end of the experiment (Laugh_2). The attention-control variant of the CI video was not included in the statistical analysis. All four outcome measures were analyzed using linear mixed models and each outcome measure was checked for normality using QQ-plots. SCR data were transformed using reciprocal transformation while the three ratings were left untransformed. The mixed model for testing habituation contained neuroticism as continuous between-subjects variable and time (Cry_1 to Cry_6) as within-subjects variable. The test for fatigue at the end of the experiment included neuroticism as continuous between-subjects variable and time with the levels Cry_6 versus Laugh_2. Interaction effects between neuroticism and time were also tested but were not significantly different from 0 for any outcome measure and therefore not considered further in the analysis of the SCR and rating data. The models also included a random intercept and, if significantly improving model fit, a random linear slope for time. To test whether at the end of the experiment (block Laugh_2) psychological  and physiological (SCR) data were related to subjects’ neuroticism scores, we performed linear regression models with neuroticism as predictor and the respective psychological or physiological variable as outcome.Statistical analyses were conducted using SPSS . For analysis of the time-course of SCR levels, as well as valence, arousal, and irritation ratings, values for five subsequent video clip presentations were averaged into one value. Thus, one mean value was obtained by averaging across the five video clip presentations with laughing infants at the beginning of the experiment  in the Heschl’s gyrus, visual cortex, superior temporal gyrus (STG), insular cortex, fusiform gyrus, temporal pole, inferior frontal cortex (including Broca’s area and Broca’s homologue on the right hemisphere), supplementary motor area (SMA), cerebellum, amygdala, and hippocampus . All brain regions, MNI coordinates, and T-scores are provided in FMRI results revealed increased BOLD signal during the p<0.05, FWE corrected, clustersize ≥ 15 voxels, see Significant fMRI BOLD signal decrements during crying infant presentations (35 min) were found in the fusiform gyrus, middle temporal gyrus, superior temporal gyrus (TE3 ), Broca’Individuals with high neuroticism levels showed stronger activation in the amygdala and sgACC when exposed to infant crying compared to individuals with lower neuroticism levels  were in a normal range [p<0.001), and irritation increased , whereas no significant trend was observed for valence . Individuals with increased neuroticism experienced more irritation  and perceived infant crying as more unpleasant  in comparison to individuals with lower neuroticism scores. Arousal was not related to neuroticism . We did not detect any interaction effects between time and neuroticism . To test for fatigue we presented at the end of the experiment a video clip that showed laughing infants . We compared Cry_6 with Laugh_2 and found significant differences for all three psychological variables, with arousal and irritation exhibiting strongly reduced values , and valence exhibiting strongly increased values  when showing a video clip with laughing rather than crying children. At the end of the experiment (block Laugh_2) psychological values for arousal and valence were not related to neuroticism . Psychological values for irritation were significantly higher in subjects with higher neuroticism scores .Neuroticism scores . Participants with the highest neuroticism values also displayed high CES-D values . CES-D values themselves had little effect on SCR values (p = 0.063 for linear relationship). The test for fatigue at the end of the experiment (comparing block Cry_6 with block Laugh_2) showed that SCRs were strongly increased when displaying a video clip with laughing rather than crying children at the end of the experiment , the left anterior insula, left and right fusiform gyrus, hippocampus, amygdala, and superior temporal gyrus . This study found highly significant differences for all three variables of emotional reactivity  as well as for the level of SCRs between the laughing and crying conditions. These findings are in agreement with the definition of habituation according to which response decrement results from repeated stimulation and does not involve sensory and motor fatigue . A futurTo reduce sources of variance not directly related to the study aims this sample was confined to healthy women. Women have been found to score higher than men on neuroticism  and showIn conclusion, the findings of this study show that individuals high in neuroticism are more emotionally responsive, experience more negative emotions, and may show enhanced cognitive control during the exposure to infant distress, which may impact infant-directed behavior. This study presents a valuable approach to investigate neuronal, physiological and emotional responses to infant crying by using simultaneous fMRI and SCR recordings and the assessment of emotional experience.S1 FileFig A. Illustration of the experiment design. (A) State-anxiety was assessed in the scanner before (STAI before) and after the experiment (STAI after). (B) Females were familiarized with the experimental setting and a video clip showing laughing infants (LI) was presented five times. Subsequently, in order to assess habituation, a video clip that showed crying infants (CI) was presented 30 times. A crontrol movie was presented three times. In order to distinguish habituation from sensory and motor fatigue we tested for fatigue at the end of the experiment by presenting the video clip with laughing infants five times. Individuals rated valence, arousal, and irritation after every video clip presentation. (C) Each video clip consisted of a sequence of five laughing or five crying children with a total length of 15 s. Fig B. Some participants with high neuroticism scores who obtained high scores on the Center for Epidemiologic Studies Depression Scale displayed low skin conductance responses. The highest neuroticism scores are shown in squares. X-axis: Shows scores on the Center for Epidemiologic Studies Depression Scale (CES-D). Y-axis: Shows skin conductance responses  during the exposure to infant crying.(DOCX)Click here for additional data file.


Aim. The aim of this comprehensive review is to systematicallyorganize the current knowledge regarding the cementation of glass-ceramicmaterials and restorations, with an additional focus on the benefits of ImmediateDentin Sealing (IDS). Materials and Methods. An extensive literaturesearch concerning the cementation of single-unit glass-ceramic posterior restorationswas conducted in the databases of MEDLINE (Pubmed), CENTRAL , and EMBASE. To be considered for inclusion,in vitro and in vivo studies should compare differentcementation regimes involving a “glass-ceramic/cement/human tooth” complex.Results and Conclusions. 88 studies were included in total.The in vitro data were organized according to the following topics:(micro)shear and (micro)tensile bond strength, fracture strength, and marginal gapand integrity. For in vivo studies survival and quality of survivalwere considered. In vitro studies showed that adhesive systems result in the best (micro)shear bond strength values comparedto self-adhesive and self-etch systems when luting glass-ceramic substratesto human dentin. The highest fracture strength is obtained with adhesive cementsin particular. No marked clinical preference for one specific procedure could bedemonstrated on the basis of the reviewed literature. The possible merits of IDSare most convincingly illustrated by the favorable microtensile bond strengths.No clinical studies regarding IDS were found. Bonded glass-ceramic restorations have gained popularity, particularly after new materials, bonding systems, cements, and cementation techniques became available in recent years. Nowadays different ceramics are introduced for the use of posterior restorations, being either an oxide-ceramic or a glass-ceramic. Glass-ceramics are of special interest in this review because their silica content and micromechanical interlocking structure allow adhesive cementation to enamel and dentin. Consequently, glass-ceramic restorations can withstand tensile forces without cement failure, even if the preparation of the tooth is nonretentive. Since the surface treatment of feldspathic porcelain in 1983 . ContempTo ensure proper attachment of an indirect restoration, basically two aspects have to be taken into consideration: conditioning of the ceramic material and conditioning of the tooth substrate followed by cementation. The most commonly used conditioning method for the glass-ceramic surface these days is application of hydrofluoric acid and silanisation, as reviewed by Tian et al. . Cements in vitro by Pashley et al.  OR ceramic∗[tw]) AND ) NOT  NOT (“Case Reports”[Publication Type] OR “Review”[Publication type]) AND English[lang].Run data search: January 1, 2015 (1868 results).EMBASE. “dental ceramics”/exp OR ceramic∗:ab,ti AND .NOT  NOT (“case report”/exp OR “review”/exp) AND[english]/lim.Run data search: January 1, 2015 (806 results).COCHRANE Library  . ceramic∗ AND .Run data search: January 1, 2015 (332 results).Titles and abstracts of the identified publications were screened by one of the authors. Full text documents were obtained for all articles meeting the inclusion criteria. Additional hand searching was performed by following up on the reference lists from included articles. Full text analysis to decide on inclusion/exclusion was subsequently performed by two reviewers and Cohen's Kappa was used as the measure of agreement. Disagreements were resolved by manner of discussion.Methodological quality regarding the risk of bias in selected articles was assessed by one of the authors according to the criteria as set by the Cochrane Collaboration Tables , and 5. Only articles about glass-ceramic materials were considered. Clinically, the focus was on single-unit posterior restorations. Included studies should compare different cementation regimes and involve a “glass-ceramic/cement/human tooth” complex. Studies regarding the benefits of IDS attracted special attention. Descriptive studies , systematic reviews, case reports, or studies with less than ten patients were excluded . Descrip in vitro and in vivo studies. For in vitro studies the data were organized according to the following topics: (micro)shear and (micro)tensile bond strength, fracture strength, and finally marginal gap and integrity. For in vivo studies survival and quality of survival were considered.The included studies were divided intoThe searches of MEDLINE (Pubmed), CENTRAL , and EMBASE resulted in 3008 publications. After exclusion of double publications, 2117 publications remained for title and abstract analysis. 1121 articles were hereafter included for full text analysis. Only a limited additional number of publications were found after checking the references of the included studies. Application of specified exclusion criteria resulted in 88 publications that could be included in the review. The exclusion criteria are described in Interobserver agreement (Cohen's Kappa) regarding final inclusion or exclusion of studies that were proposed after full text analysis was 0.80 (IBM SPSS 22), which is generally considered to be a strong level of agreement . Initial in vitro studies proved difficult and hardly ever applicable. Sequence generation and incomplete outcome data for in vitro studies are not explained in most cases but just named. Assessment “unclear” on incomplete outcome data generally implies that no missing data were reported. Most studies in this review did not report sequence generation; for in vitro studies the relevance of this can be subject of debate. For in vivo studies sequence generation, allocation concealment, and blinding were often assessed as “unclear,” because studies often did not describe these procedures. Overall the included studies had a low risk of bias. More specifically, a low risk of bias was assessed for shear bond strength studies, tensile strength studies, and marginal gap studies. An unclear risk of bias was assessed for fracture strength studies and in vivo studies.The included studies were assessed for their risk of bias according to the Cochrane library Tables , and 5. Because of their great variety it is important to divide contemporary resin cements into subgroups regarding their curing type, their viscosity, and whether they are either adhesive (with a 3-step adhesive), self-etching (with a 2-step or 1-step adhesive), or self-adhesive. This terminology is not used consistently in literature. An overview is presented in Seventeen studies could be identified that met the inclusion criteria; their risk of bias is overviewed in In only one study different groups of luting agents were used and the authors concluded that zinc phosphate cement and glass ionomer cements produced the lowest shear bond strengths, whereas the highest shear bond strengths were found with two self-etching cements (Panavia F2.0 and Multilink) and one self-adhesive resin cement (RelyX Unicem) .n = 7, [Several studies n = 7, –22) comp compn = n = 7, , 17. When = 7, . To enamn = 7, . When din = 7, . To dentn = 7, . In anotOthers, using a push-out test, concluded that an adhesive cement (Variolink II/Syntac) did not perform better than three self-adhesive cements .To enamel three different self-etching resin cements with different setting modes  were compared in a microshear bond strength test; no significant differences were seen .Four studies –28 focusThe application of fluoride or triclosan based desensitizing agents prior to adhesive cementation did not influence the shear bond strength , nor didIn summary, some evidence supports the use of adhesive cement with respect to the shear bond strength compared to self-adhesive and self-etch systems when luting all ceramic materials to human dentin. There is little evidence to support the assumption that IDS improves the shear bond strength especially when Clearfil SE Bond was used.Fifteen articles could be included investigating the effect of different cements on glass-ceramic restorative materials with a (micro)tensile bond strength test; their risk of bias is overviewed in When comparing different cement groups, glass ionomer cement (Aquacem) yielded far lower tensile bonding strengths (2-3 times) compared to a self-etch resin cement (Dicor LAC) .In studies comparing different resin cements results were opposite or similar about which cement, self-etching or self-adhesive, resulted in the highest tensile bond strength –35 or obIt was hypothesized that the tensile bonding strength is not so much dependent on the type of adhesive approach but more so on the chemical composition and viscosity of the cement used. Interestingly, the use of self-etch adhesive combined with a restorative composite (Clearfil SE Bond with Clearfil APX) yielded higher tensile bond stresses to dentin than dedicated self-adhesive, self-etch, and adhesive cements . But no Overall, autocure leads to a lower microtensile bond strength when compared to dual-cure cement modes , 43. PreAs before, tensile bond strengths were also higher for enamel than for dentin, that is, in a study by Habekost et al. .The effect of IDS on microtensile bond strength was tested in two studies. An IDS layer (one or two resin coatings) applied directly after preparation yielded higher values compared to applying it just prior to cementation or not at all. No temporary restorations were made , 46.In summary, no one particular cement or adhesive system, be it self-etching, self-adhesive, or adhesive, showed overall superior results with respect to (micro)tensile bond strength. IDS improved microtensile bond strength in both included studies.Fifteen studies could be identified that met the inclusion criteria; their risk of bias is overviewed in Seven studies , 54–59 wOne study described the effect of the thickness of IDS materials (Clearfil SE Bond and Protect Liner F) on the fracture strength of IPS Empress II crowns cemented with Panavia F. The film thickness formed by Clearfil SE Bond and Protect Liner F increased the fracture load of IPS Empress II crowns . in vitro fracture strength of posterior adhesively cemented specimen, exhibit higher fracture strength with adhesive cements. Literature is inconclusive about the type of resin cement used. The modulus of elasticity is considered more important than the type of resin cement. There are no data found in the literature on fracture strength using contemporary glass-ceramics, such as lithium disilicate. So extrapolation of the findings to current materials and cementation protocols should only be done with great reservations. Little evidence supports the use of IDS in increasing the fracture load [In summary, teeth that are restored with an indirect glass-ceramic restoration, with respect toure load . in vitro studies focusing on marginal gap and marginal integrity are too numerous to allow for individual discussion. Therefore the relevant findings evolving from these studies are outlined below.Twenty-six studies could be identified that met the inclusion criteria; their risk of bias is overviewed in A consistent finding is that the least microleakage and the best marginal adaptation are obtained when using a resin cement , 61–64. μm can be partially compensated by a resin cement. In such cases highly viscous, filled composite cements are recommended when considering the quality of postcementation marginal integrity [Four studies –68 focusWhen applying resin cements, the degree of microleakage is generally higher on dentin margins than on enamel margins , 69–75. In a study involving the cementation of partial crowns, preparation design was of no influence with respect to the size of the marginal gap .Five studies , 80–82 iIn summary, adhesive resin cements showed the least microleakage and are least affected by artificial aging. With a large marginal gap a highly viscous cement is recommended, when the gap is smaller there is no advantage but also no disadvantage of using a highly viscous cement. “Small” and “Large” are not further specified. Compared to enamel, there was generally more microleakage in dentin. There was little proof that with etch-and-rinse systems a higher percentage of gap-free margins could be obtained in enamel, compared to dentin. With self-etching systems and self-adhesive systems equivalent or even more gap-free margins were reached in dentin. IDS was generally considered of merit in reducing microleakage.There were twenty clinical studies on glass-ceramic restorations comparing different cementation protocols, but protocols and materials were seldom similar among different studies. Their risk of bias is overviewed in Mirage fired feldspathic restorations were luted with either a dual-cure composite (Mirage) or a glass ionomer luting cement (Fuji I), resulting in 2% and 15% lost or fractured restorations, respectively, after a maximum observation period of 3 years. The predominant complication was adhesive bond failure at the cement-porcelain interface  as also In another similar study restorations could be evaluated after 6 years with 12% and 26% failures, respectively. The difference was already obvious at the 3-year recall period . In contIn another split mouth study, Cerec (Vita Mark II) inlays were cemented with either a dual-cured  or chemically cured resin cement (Cavex Clearfil F2) and evaluated according to the criteria of the California Dental Association. Twenty-three percent of the restorations were replaced, all from the dual-cured resin cement group within a 10-year period. Possibly, the self-curing capacity of the dual-cured resin cement was insufficient to achieve adequate hardening in order to withstand the stresses and strains that can arise in posterior regions. Although no differences in qualitative parameters were reported between baseline and the period after 10 years, acceptable scores for marginal discoloration after 10 years were seen more frequently in the dual-cured than in the chemically cured cement group (58% versus 78%) .Klink and colleagues also used Vitablocs Mark II full crowns, partial crowns, and inlays luted with either Variolink II or RelyX Unicem. According to the CDA-criteria inlays and partial crowns performed well. Prevalence of complications or failure was highest for crowns. They concluded that success was related to patient factors and restoration type, not luting protocol . Others In a study by Gemalmaz and colleagues two adhesive cements (Variolink Ultra and Enforce) and a glass ionomer cement (Geristore) were used to lute Ducere LFC ceramic inlays resulting in 13%, 13%, and 33% failures, respectively, after a little more than 2 years. Margins were evaluated by SEM on gypsum models. Deterioration of marginal adaptation, rate of submargination, and marginal discoloration of surviving restorations luted with the glass ionomer cement were markedly inferior to those luted with the other two cements, with the restorations cemented with Variolink Ultra performing the best .In a prospective dual-center study, the clinical behavior of adhesively luted pressed glass-ceramic restorations (Cergogold) was evaluated using two cementation regimens . One group of restorations was luted with Definite Multibond primer with corresponding adhesive and definite cement and the other with Syntac classic (3-step) with Variolink Ultra cement. Survival rates were 93% and 95%, respectively, after 4 years, with the first group exhibiting more hypersensitivity shortly after cementation of the restoration (27% versus 0%). Hence both luting protocols provided similar results when compared according to USPHS criteria and by SEM . A similLithium disilicate restorations were cemented with either a commercially available self-etching dual-curing cement  or a self-adhesive dual-curing “experimental” cement originating from the same company . Both cements had qualitatively similar results after 2 years of function as assessed by the modified USPHS criteria. All restorations functioned for 2 years without crown fracture or surface chipping. The undisclosed nature of the experimental cement leaves little room for practical comparison or interpretation. The publication did not mention the type of restoration that was provided  . For thiIn conclusion, most included, rather heterogeneous clinical studies involve relatively old, no longer available restoration types or systems. The use of lining cements in several older protocols challenges external validity. Cementation protocols involving glass ionomer cements generally  result in more fracture and loss of restorations as well as poorer qualitative performance of surviving restorations compared to protocols involving adhesive resin cements. Studies comparing cementation protocols for more contemporary restorative materials (lithium disilicate) are rare and involve self-etching, self-adhesive, or adhesive procedures. None of these cementation protocols can be considered clearly superior in clinical performance on the basis of the reviewed literature.There is limited evidence that light-cured resin cements perform worse than dual-cured cements, whereas solely chemically cured resin cements perform the best. Results obtained with technically challenging adhesive cementation procedures may be operator-dependent. Marginal deterioration is frequently reported, also when using adhesive cements.No clinical studies evaluated the potential benefits of IDS protocols that were identified.This review is aimed at organizing knowledge regarding the cementation of glass-ceramic restorations, particularly posterior, single-unit ones, with a special emphasis on the possible merits of IDS. The topic is of interest to the clinician because of the growing number of all ceramic restorations that are being placed. They substitute metal and metal-ceramic crowns and are advantageous because they are relatively cheap in light of the current gold price and their manufacturing price and because of their superior esthetics. In early years, glass-ceramics were cemented with conventional cements like glass ionomers, with limited adhesive properties. This reflects on the results, as demonstrated in this comprehensive review, and consequently challenges the external validity of data subtracted from these studies to contemporary, strengthened glass-ceramics (leucite reinforced glass-ceramic and lithium disilicate). By removing superficial glass content by etching, glass-ceramics can be cemented adhesively and as a result allow nonretentive preparation forms, maintaining sound tooth tissue. This may help in avoiding endodontic complications.Bonding to dentin has traditionally been considered to be more challenging than to enamel. IDS may provide better results with respect to the bonding capacity and it is possibly also more friendly to the pulp.Over 3000 studies were initially identified for this review, but many were discarded, predominantly because they did not compare different cementation protocols or evaluated a “glass-ceramic/cement/human tooth” complex. The selection on articles in the English language only may have introduced some bias. in vitro and in vivo studies that were included proved dramatically heterogeneous. Consequently, they do not allow meta-analysis or relevant grouping because of different test methods , testing machines, standardization of the test method, crosshead speed of the testing device and the size of the steel ball during instrumentation, the use of a “stress breaker” such as a rubber dam, film thickness of luting cements, or (lack of) definition of outcome parameters, particularly the mode of failure). It was decided to include studies only if they compared cements or cementation procedures, thus correcting for the heterogeneity in some manner. Often it was complicated to categorize the cementation procedures into “adhesive,” “self-etching,” or “self-adhesive” because of the chosen bonding agents and the confusing way that they were applied and described.TheWith respect to the application of IDS, terminology and the clinical application in the literature regarding this procedure are different. The present authors regard IDS as a procedure in which a resin layer is applied immediately after preparation, followed by impression taking and the provision of a temporary restoration in combination with a temporary cement. Eventually, this restoration is replaced by a glass-ceramic one, which is luted to the reactivated IDS layer and the uncovered tooth structure by means of a resin cement. In the current review, when no temporary restoration was provided in an evaluated study, it is referred to as a “resin coating,” which is fundamentally different. The manner in which such an intermediate layer is applied and conditioned is also expected to be of influence and often different among studies that were included.Nevertheless and possibly as a result of the rather rigorous inclusion and exclusion criteria, the included studies in the review are generally considered of good methodological quality as evaluated by Cochrane's collaboration tool of bias.In vitro studies identify some differences in outcome resulting from the tested protocols or variables. These are generally not reflected in rather more crude, clinical outcome measures, such as survival of a restoration, presented in in vitro studies. Therefore it is tentatively suggested that when luting modern glass-ceramics to posterior teeth, adhesive protocols that are the most operator and patient friendly may be preferred.Bearing in mind the shortcomings and limitations of this review as described above, the following conclusions are drawn. in vitro studies it can be concluded that adhesive systems  show the best (micro)shear bond strength values compared to self-adhesive and self-etch systems when luting to human dentin. For (micro)tensile strength values or evaluation of the marginal gap no such preference can be identified on the basis of the reviewed literature. The highest fracture strength is obtained using adhesive cements, rather than water based cements like glass ionomer.FromClinical studies comparing cementation protocols for contemporary restorative glass-ceramic materials (lithium disilicate) are rare and involve self-etching, self-adhesive, and adhesive procedures. No marked clinical preference for one specific procedure could be demonstrated on the basis of the reviewed literature. in vitro. No clinical trials have been performed and deleterious clinical consequences, be it objective or subjective, were not reported.Few studies focus on the possible merits of IDS. The benefits are most convincingly illustrated by the favorable microtensile bond strengths when compared to negative or positive controls


During tissue homeostasis, normal stem cells self-renew and repopulate the diverse cell types found within the tissue via a series of carefully controlled symmetric and asymmetric cell divisions (ACDs). The notion that solid tumors comprise a subset of cancer stem cells (CSCs) with dysregulated self-renewal and excessive symmetric cell divisions has led to numerous studies aimed to elucidate the mechanisms regulating ACD under steady-state conditions, during stem-cell expansion and in cancer. In this perspective, we focus on a type of asymmetry that can be established during ACD, called non-random co-segregation of template DNA, which has been identified across numerous species, cell types, and cancers. We discuss the role of p53 loss in maintaining self-renewal in both normal and malignant cells. We then review our current knowledge of the mechanisms underlying co-segregation of template DNA strands and the stem-cell pathways associated with it in normal and CSCs. The continual maintenance of a reservoir of tissue-specific stem cells affords an organism with the ability to generate all the differentiated cells needed for tissue homeostasis and repair throughout its lifespan. During homeostasis or repair, a stem cell can undergo asymmetric cell division (ACD) to simultaneously generate daughters with differing cell fates. One daughter remains a stem cell and the other gives rise to differentiated progeny that carry out the functions of the mature tissue. When a stem cell asymmetrically divides, it does so by actively segregating one or more intrinsic cell fate-determining constituents, or by polarizing the cell cortex and aligning the mitotic spindle so that the two daughter cells are exposed to differing external stimuli directing their cell fate potential , 2. In cIt is necessary to maintain a tightly controlled balance between symmetric and asymmetric stem cell divisions in order to preserve an optimal number of stem cells within a tissue or organ. When the balance is shifted to favor excessive symmetric self-renewing divisions, it can lead to a hyperplastic state and cancer development [reviewed in Ref. ]. The tuA debated question in CSC biology is whether neoplastic transformation emerges from normal stem/progenitor cells or from more differentiated cells that commandeer stem cell properties during or after the oncogenic process. Since normal stem cells inherently possess many of the properties that CSCs exploit, it seems more likely that a tumor would arise from a normal stem cell. Lineage tracing experiments and genetically engineered mouse models in which oncogenic events were restricted to specific cell types have confirmed that transformation can indeed arise from the normal stem cell population . Howeverin vivo lineage tracing experiments have proved that differentiated cells within a tissue can replenish the stem cell population  undergo repair double strand breaks much faster than differentiated cells, primarily using homologous recombination [reviewed in Ref. ]. Given in vitro. Furthermore, p53 isoform switching was associated with tumor progression in colon cancer  regulating this process has remained elusive. In order to decipher how template chromosomes are asymmetrically segregated during cell division, one must identify what “marks” the newer and older DNA strands, what cellular machinery recognizes those marks and finally, what process actively recruits the chromosomes bearing those marks to one daughter cell. The Tajbakhsh group speculated that the mother centrosome might anchor certain sister chromatids to the polarized cortex . How theMuch of the effort to identify the molecular mechanisms responsible for a cell’s decision to co-segregate its template DNA strands has been pioneered by the Sherley group. Rambhatla et al. demonstrated that template DNA co-segregation was induced by p53 and required the down-regulation of an enzyme essential for guanine nucleotide biosynthesis, IMP dehydrogenase . PerhapsS. pombe and Schizosaccharomyces japonicas yeast that a DNA strand-specific epigenetic imprint at the mating locus initiates a recombination event, which is required for cellular differentiation (Sherley’s group later discovered that the template DNA strands harbor more of the histone H2A variant H2A.Z that is “uncloaked”, meaning that it is more detectable by immunofluorescence . They montiation , 39. In ntiation . An intrAdditional clues about the mechanisms driving template DNA strand co-segregation have emerged from studies of stem-cell signaling pathways that are either associated with or actively affect the balance of ACD and template DNA strand co-segregation. Wnt signaling is a key stem cell signaling pathway implicated in self-renewal of both normal stem cells and CSCs, as well as epithelial–mesenchymal transition in cancer . InhibitDrosopila and C. elegans, as well as in mice and human beings (Drosophila neuroblasts the Notch inhibitor Numb is inherited by the daughter cell that is destined to undergo differentiation (The most extensively studied pathway associated with template DNA co-segregation is the Notch signaling pathway. Notch is a developmental and adult stem/progenitor cell transcription factor that regulates self-renewal and differentiation in a highly cell-type and context-specific manner . Notch sn beings . For exantiation , 44. Whentiation , 46. Notntiation –49, thouNotch signaling has been linked to template DNA co-segregation. One example has been demonstrated in muscle satellite stem cells. Satellite cells self-renew and provide the generation of myoblasts that are needed for skeletal muscle homeostasis and repair. Muscle satellite stem cells co-segregate their template DNA strands , 51 and It has been nearly 50 years since the first discovery of template DNA co-segregation. Though it has been observed across numerous species, normal tissue, and cancer, we have still only scratched the surface toward elucidating the purpose for conserving template DNA strands in one daughter cell and the mechanisms regulating the process. Consistent with the notion that the immortal DNA strand hypothesis does not fully capture why stem cells asymmetrically segregate their template DNA, there has been increasing evidence that the mother cell controls gene expression in the daughter cells through asymmetric segregation of epigenetic marks. With recent advances in genomics, chromosome-imaging technologies, and genetically modified model organisms [reviewed in Ref. ], the ouThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Smoking is the second leading cause of preventable death in the United States. Cohort epidemiological studies have demonstrated that women are more vulnerable to cigarette-smoking induced diseases than their male counterparts, however, the molecular basis of these differences has remained unknown. In this study, we explored if there were differences in the gene expression patterns between male and female smokers, and how these patterns might reflect different sex-specific responses to the stress of smoking. Using whole genome microarray gene expression profiling, we found that a substantial number of oxidant related genes were expressed in both male and female smokers, however, smoking-responsive genes did indeed differ greatly between male and female smokers. Gene set enrichment analysis (GSEA) against reference oncogenic signature gene sets identified a large number of oncogenic pathway gene-sets that were significantly altered in female smokers compared to male smokers. In addition, functional annotation with Ingenuity Pathway Analysis (IPA) identified smoking-correlated genes associated with biological functions in male and female smokers that are directly relevant to well-known smoking related pathologies. However, these relevant biological functions were strikingly overrepresented in female smokers compared to male smokers. IPA network analysis with the functional categories of immune and inflammatory response gene products suggested potential interactions between smoking response and female hormones. Our results demonstrate a striking dichotomy between male and female gene expression responses to smoking. This is the first genome-wide expression study to compare the sex-specific impacts of smoking at a molecular level and suggests a novel potential connection between sex hormone signaling and smoking-induced diseases in female smokers. Cigarette smoking is the largest single risk factor for premature death in the United States and is responsible for 440,000 deaths every year . SmokingIncreasing evidence suggests respiratory symptoms vary by sex, smoking habits and age. It has been reported that the health effects of smoking are more serious for women than for men . For insSex differences in rates of survival following diagnosis of lung cancer have also been reported. Interestingly, women have been found to have higher survival rates regardless of lung cancer type, stage and therapy ,20–23. IThe toxic components of cigarette smoke enter the blood stream through the pulmonary alveoli and are distributed throughout the body. Thus, blood may provide an appropriate biological material in which to study the systemic effects of cigarette smoke exposure. Several studies have investigated gene expression in smokers’ peripheral blood lymphocytes and reported substantial numbers of genes differentially expressed in smokers and non-smokers –29. WhilMoreover, although many clinical and epidemiological studies have indicated that women are more vulnerable to smoking associated diseases, no genome-wide expression study has investigated the effect of sex on the smoking response. Very recently, Pan et al.,  comparedIn the present study, we compared gene expression profiles of peripheral blood cells between smokers and non-smokers using Agilent whole-genome microarrays, and distinguished patterns of smoking-related gene expression that differ dramatically between males and females. A large number of the smoking-associated genes identified were directly relevant to well-known smoking-related pathologies, and in females were connected to signaling by estrogen and progesterone. This is the first genome-wide expression study on the sex-specific impact of smoking as reflected in peripheral blood cells, and our findings may begin to suggest molecular connections underlying some of the differences in smoking-related cancer risks and outcomes previously found in epidemiological studies.Healthy smokers and non-smokers were recruited with written informed consent. The study was approved by the Institutional Review Board of Columbia University. A total of 24 males and 24 females consisting of an equal number of smokers (≥1 pack per day) and non-smokers (never smokers) of each sex participated in this study. Characteristics including age, sex, and details of smoking habit were provided by the subjects at the time of enrollment. All participants also reported were not under any therapeutic drug or no record of recent radio diagnostic examination. Characteristics of the study population are summarized in Peripheral blood from smoker and non-smoker volunteers was drawn into 0.105 mol/l sodium citrate Vacutainer tubes . RNA was prepared using the PerfectPure RNA blood kit  as recommended by the manufacturer, followed by GLOBINclear  treatment to further reduce levels of both α- and β-globin. The RNA was quantified using a NanoDrop-1000 spectrophotometer, and quality was monitored with the Agilent 2100 Bioanalyzer . All RNA samples had RNA integrity numbers between 7.8 and 9.1 . RNA was stored at −80°C until use.Cyanine-3 (Cy3)-labeled cRNA was prepared from 0.5 μg input RNA using Agilent Technologies’ One-Color Quick Amp labeling kit according to the manufacturer’s instructions, followed by purification of cRNA by RNAeasy column . Cy3-dye incorporation and yield of cRNA was checked with a NanoDrop ND-1000 spectrophotometer. The specific activity of all cRNA samples ranged from 10.98 to 20.02 (mean 14.94). cRNA (1.65 mg) was fragmented and hybridized to Agilent’s whole genome microarrays (G4112A) at 65°C for 17 hr with rotation, followed by washing according to the manufacturer’s recommendations. Microarray slides were scanned immediately with the Agilent scanner (G2404B) using the recommended settings and the images were analyzed with Feature Extraction 9.1 (Agilent Technologies) using default parameters.Background corrected hybridization intensities were imported into BRB-ArrayTools Version 3.8.0 beta , log2 trBRB class comparison was applied to identify genes that were differentially expressed between smokers and non-smokers using a two-sample Student t-test. Genes with p-values of p<0.005 were considered statistically significant. The false discovery rate (FDR) was also estimated for each gene using the method of Benjamini and Hochberg  to contrMultidimensional scaling (MDS) was used in BRB-ArrayTools to visualize differences between groups at the transcriptional level based on differentially expressed sets of genes using the Euclidian distance metric to compute a distance matrix and the principal components of the gene expression signature. In the output, each sample is represented by a single point and the distance between any two points indicates the overall similarity of the two represented samples. In addition, we executed hierarchical clustering analysis to generate a heat map image of the expression of genes differentially expressed between smokers and non-smokers using the Euclidean distance matrix.® Systems, www.ingenuity.com). Following collation, there were 131 and 191 transcripts in males and females, respectively that mapped to known genes in IPA. We first used the IPA biomarker analysis workflow to identify promising molecular biomarker candidates from the dataset. The IPA-Biomarker filter mapped 124 and 175 biomarker candidate genes in male and female smokers, respectively. We next performed IPA core-analysis in the context of pathways and networks, biological function and/or diseases. The right-tailed Fisher’s exact test was applied to calculate the p-value ascertaining the probability that each biological function and/or disease assigned to that dataset was due to chance alone.The significantly differentially expressed genes in male and female smokers were imported into IPA version 7.6  for interpreting smoking induced microarray data using the Broad Institute’s GSEA software . GSEA, uGlobal gene expression was measured in peripheral white blood cells of 48 donors, comprising equal numbers of smokers and non-smokers from each sex. Agilent whole genome microarrays were hybridized using the one-color protocol to identify genes differentially expressed between smokers and non-smokers. We used the class comparison feature of BRB-ArrayTools to identify genes that were differentially expressed between smokers and non-smokers. We identified 300 genes with significantly different expression (p<0.005) , of whicWe next investigated the possibility of sex specificity of smoking signatures in males and females. We first performed a class comparison for differentially expressed genes between smokers and non-smokers using only the 24 male donors. This analysis identified 175 genes significantly differentially expressed between male smokers and non-smokers (p<0.005) . Of the We next used the same analyses to look for altered gene expression between female smokers and non-smokers. In this case, class comparison identified 237 genes as differentially expressed (p<0.005) between female smokers and non-smokers . Among tAlthough a substantial number of genes were found to be differentially expressed in both male and female smokers, a divergent pattern of expression was apparent on the basis of the number of genes up- or down-regulated in male and female smokers. The intersection of male and female gene sets yielded only 13 genes (3%) that responded significantly in both sexes .To investigate further if cigarette smoking has any significant interaction with sex, we applied BRB-ArrayTools’ 2-way mixed model ANOVA. This analysis identified eighty genes with smoking responses modified by sex (p<0.005) . InteresTo further assess the sex specific smoking response, we performed pathway analysis with the gene sets differentially expressed in male and female smokers using Ingenuity Pathway Analysis (IPA), version 7.6. First, we identified overrepresentation of smoking-affected genes within known functional categories . AlthougThe over-represented biological function categories also differed extensively between male and female smokers . FurtherThere was also high over-representation of canonical pathway categories of smoking correlated genes in female smokers that were related well to known smoking pathologies, shown in The functional categories of immune and inflammatory responses/diseases differed widely between smoking-responsive genes in male and female smokers . We addeSmoking undeniably causes a different pattern of elevated cancer risk in females and many clinical studies have reported that women have a higher absolute risk of smoking induced cancers –16,34, aThe effects of cigarette smoking on human health are serious and in many cases, deadly. Cohort epidemiological studies have shown that women are more vulnerable to cigarette-smoking induced diseases; however, the molecular basis of these differences has remained unclear. In this study human peripheral blood was used to explore sex-specific smoking induced differential gene expression between smokers and non-smokers. Cigarette smoke toxicants enter the body through the pulmonary alveoli and are directly absorbed and distributed throughout all tissues. Blood cells are not only one of the most accessible tissues for gene expression analysis, they also have been shown to be an excellent tissue type to study environmental exposures, like cigarette smoke  and radiThe male and female smoking-correlated genes associated with biological function and/or diseases identified in this study are directly relevant to well-known smoking related pathologies. The results include strong involvement of a range of smoking-correlated diseases such as cancer, neurological disease, immune response, inflammatory diseases, cardiovascular disease, respiratory disease, hematological disease, cell death and proliferation, cellular development and cell-to-cell signaling, and xenobiotic metabolism. However, the extent of these relevant biological function and/or diseases, the number of correlated genes and their expression patterns were strikingly over-represented in female smokers . Our obsCigarette smoking is an established risk factor for many cancers, not only at the site of contact but also throughout the body. We identified sets of smoking correlated genes in both male and female smokers with documented associations with various cancers, consistent with risks observed in epidemiological studies. One remarkable finding of this study was the altered expression of genes corresponding to sex specific cancers, hyperplasia of prostate in male smokers and epithelial ovarian cancer in female smokers. Epidemiological studies have suggested that smoking increases the risk of ovarian  and prosGene set enrichment analysis (GSEA) with oncogenic signature gene sets further substantiated the gene expression differences underlying the female smokers’ heightened susceptibility to tobacco carcinogens. GSEA identified a large number of oncogenic pathway gene-sets that were significantly altered in female smokers compared to male smokers. Our data corroborate the findings of other investigators suggesting that women are more susceptible than men to the ill effects of the carcinogens in tobacco and tobacco smoke ,12,15,16Although female smokers have a higher absolute risk of developing cancers than their male counterparts, women’s better survival rate from smoking-associated cancers ,14,24 haExpression of genes within the functional categories of immune and inflammatory responses/diseases also differed widely between male and female smokers. Compared to male smokers, a large number of genes corresponding to immune alteration and suppression were altered in female smokers. In a recent study, Pan et al.,  demonstrSince all these studies suggest the importance of immune and inflammatory response in the female smoking response, we constructed a network of immune and inflammatory response affected by smoking in females based on connectivity information of gene/gene product interactions from the IPA Knowledge Base . This neThe present study was carried out with a modest number of smokers and non-smokers with a broad range of ages using peripheral blood leukocytes. Future investigation of smoking effects on different lymphocyte populations in both sexes may provide more precise information of the impact of sex on the physiological response to smoking. The present study suggests genes associated with immune functions are severely affected in female smokers. Thus, it may be informative to examine smoking impacts specifically in B cells in both male and female smokers. To further examine the impact of sex on the response to smoking, our results suggest that it will be essential to study sex-specific gene expression changes in female smokers in different age groups. If the gene expression response to smoking is, indeed, influenced greatly by interactions with female hormones, the smoking-induced gene expression responses seen in pre- and post- menopausal women should differ significantly. Some recent studies have indicated a link between menopausal hormone treatment therapy and increased lung cancer risk in females, and female smokers having hormone therapy possessed a particular high risk of mortality from lung cancer ,46. ThisThis is the first genome-wide expression study to compare the sex-specific impact of active smoking in vivo. Using defined functional network analyses we have identified sets of altered genes related to a large number of smoking-induced pathologies in both sexes. Based on defined functional relationships, we suggested a potential novel connection between sex hormone signaling and smoking-induced diseases in female smokers . HoweverSI Table 1SI Table 2SI Table 3SI Table 4SI Table 5SI Table 6SI Table 7SI Table 8


Background. Thyroid hormones (THs) are essential for normal human fetal development and play a major role in the regulation of glucose and lipid metabolism. Delivery of TH to target tissues is dependent on processes including TH synthesis, transport, and metabolism. Thyroid hormone endocrine disruptors (TH-EDCs) are chemical substances that interfere with these processes, potentially leading to adverse pregnancy outcomes. Objectives. This review focuses on the effects of prenatal exposures to combinations of TH-EDCs on fetal and neonatal glucose and lipid metabolism and also discusses the various mechanisms by which TH-EDCs interfere with other hormonal pathways. Methods. We conducted a comprehensive narrative review on the effects of TH-EDCs with particular emphasis on exposure during pregnancy. Discussion. TH imbalance has been linked to many metabolic processes and the effects of TH imbalance are particularly pronounced in early fetal development due to fetal dependence on maternal TH for proper growth and development. The pervasive presence of EDCs in the environment results in ubiquitous exposure to either single or mixtures of EDCs with deleterious effects on metabolism. Conclusions. Further evaluation of combined effects of TH-EDCs on fetal metabolic endpoints could improve advice provided to expectant mothers. Pregnancy results in many physiological changes that have significant effects on thyroid status . The fetCaloric intake and personal lifestyle are strongly associated with obesity and metabolic syndrome; however, there is a growing concern that a subgroup of endocrine disrupting chemicals (EDCs) known to specifically alter TH regulation (TH-EDCs) may contribute to the prevalence of metabolic syndromes by altering signalling pathways involved in glucose and lipid homeostasis during pregnancy. The risk of exposure to TH-EDCs is rising with exposure to multiple TH-EDCs more common than exposure to single chemicals. Several studies have reported the effects of exposure to individual EDCs and this has been reviewed elsewhere . This reFor this narrative review conducted between 2014 and 2015, the PubMed  and Google Scholar databases were interrogated with the following key words and phrases: TH endocrine disruptors and pregnancy/early or late pregnancy; prenatal effect of TH endocrine disruptors on glucose/lipid metabolism; mechanism of action of TH endocrine disruptors; placental transfer/biotransformation of TH endocrine disruptors. All study types including randomized controlled trials, case-control, human, and animal studies were considered and results are restricted to English only. The articles were grouped according to the effects of the TH-EDCs on related endpoints.The THs thyroxine (T4) and triiodothyronine (T3) are synthesized and secreted by the thyroid gland. Thyroid epithelial cells synthesize thyroglobulin, which provides tyrosine residues that are iodinated to iodotyrosine by thyroid peroxidase (TPO) to form T4 and T3  . MaintenTHs are highly lipophilic and are secreted into the blood stream where they are bound by the TH distribution proteins thyroxine binding globulin (TBG), transthyretin (TTR), or albumin . TH cellOnce inside the cell, the deiodinase enzymes (Type 1 (D1), Type 2 (D2), and Type 3 (D3)) regulate the conversion of T4 to either active T3 (D1 and D2) or inactive reverse T3 (rT3) (D1 and D3) by the removal of a specific iodine atom . MetabolTHs converted to T3 in the cell can either activate or repress gene transcription by binding to nuclear TH receptors (TRs) which are DNA-binding transcription factors that bind specific thyroid hormone response elements (TREs) in the regulatory region of target genes . TRs mayA person is described as euthyroid when the thyroid gland is functioning normally resulting in normal steady state levels of serum THs and TSH. Hypothyroidism results when the thyroid gland does not produce adequate amounts of TH resulting in low serum TH levels and elevated TSH. Hyperthyroidism results from the thyroid gland producing too much TH resulting in high TH and suppressed TSH in the serum. Both hypo- and hyperthyroidism commonly result from autoimmune disease attack of TPO (hypo- and hyper-), thyroglobulin (hypo-), TSH receptor antibodies, and thyroid stimulating immunoglobulin (hyper-). When serum TH levels fall within the normal range but TSH levels are not normal , this signifies the early stages of thyroid disease and is described as a subclinical state . TH disrTH levels are markedly altered by the hormonal and metabolic demands of normal pregnancy . The plaThe maternal thyroid gland accommodates the demands of pregnancy and a growing fetus by increasing hormonal output. The human fetal thyroid can secrete TH from 16 weeks of gestation , whereasGlucose metabolism is altered during pregnancy to guarantee adequate delivery of nutrients to the growing placenta and fetus. In pregnancy, maternofetal glucose transport is largely dependent on the expression of one or more member of the facilitative family of glucose transporters (GLUTs) in the placenta. Increased fat mass in overweight and obesity is associated with prepregnancy insulin resistance, which is compounded by insulin resistance of pregnancy , 48. In TH affects many aspects of glucose metabolism: circulating insulin levels and counter-regulatory hormones; hepatic gluconeogenesis and glycogenolysis; and intestinal glucose uptake. TH regulates the transcription of several genes involved in glucose metabolism. At the tissue level, TH actions on glucose metabolism are regulated by transmembrane transporters, ligand-dependent TH receptors, and deiodinases .β cells continuously secrete insulin even in the absence of stimulatory glucose concentrations. This results in hyperinsulinemia and subsequently insulin resistance [Deranged TH levels in pregnancy are associated with adverse maternal and fetal metabolic outcomes , 56, 57.sistance . Untreatsistance , 61. FurIn contrast, evidence of decreased glucose production and utilization has been observed in overt and subclinical hypothyroidism . TH replMaternal serum lipid concentrations increase as pregnancy progresses . The dynMaternal lipid metabolism in normal and complicated pregnancies has recently been reviewed . In womeTH is a main regulator of lipid metabolism. The expression of TH receptor isoforms is an important determinant of its actions on lipid metabolism . The oveThyroid dysfunction has been associated with abnormal lipid profiles. During normal pregnancy, altered TH levels have been associated with a less favorable metabolic phenotype in pregnant women with normal thyroid function prepregnancy , 72. HypExogenous substances capable of interfering with the structure or function of the endocrine system are known as endocrine disrupting chemicals (EDCs). As with other hormones, TH is a target of EDCs and many studies have identified chemicals that alter TH homeostasis. TH-EDCs may be clustered into two main groups based on their biodegradability and bioaccumulation in the environment. Nonpersistent organic chemicals (N-POCs) are widespread in the environment but are nonlipophilic and do not bioaccumulate. Persistent organic chemicals (POCs) are highly stable lipophilic compounds that bind to adipose tissue in living organisms and bioaccumulate up the food chain. N-POCs and some POCs are rapidly metabolized by enzymes and eliminated from the body. Organochlorine (OC) and organophosphate (OP) pesticides are highly lipophilic; OPs are unstable and therefore more readily metabolized than OCs which have been found to accumulate in adipose tissue .TH-EDCs are ubiquitous and originate from chemical additives used as flame retardants; synthetic plasticizers and solvents used in food packaging, polyvinyl chloride tubing, medical equipments, pesticides, toys, personal products, adhesives, powder paints, and dental sealants; antimicrobial compounds used in household detergents; toxic by-products of combustion processes; insulating materials for electrical equipment such as transformers and capacitors, heat transfer systems, hydraulic fluids, and lubricants; airbag inflation systems, fireworks, nitrate fertilizers, matches, and oxidants in propelling rockets and missiles; and synthetic and naturally predominant compounds in soy rich foods , 16, 76.TH-EDCs interfere with TH and TSH signalling through many pathways in many different species: altering deiodinase activity , 28, 77, In vitro toxicology studies of non-dioxin-like PCBs on humans revealed that PCB related compounds (congener) 168 and 125 completely inhibited T4-TTR binding because of their structural similarity to T4, but they also had very high androgen-inhibitory potencies. PCB-168 exhibited weak antiestrogenic activities whereas PCB-125 exhibited ER-mediated activity. On the other hand, PCB-104 had high AR-antagonistic potencies and was the most effective congener with estrogenic properties of all congeners studied [Certain TH-EDCs such as brominated flame retardants, hydroxylated polychlorinated biphenyls (PCBs) metabolites, and dioxin (PCDD) share structural similarities with TH and bind with the high affinity TH transport protein TTR , consequ studied . Exposur in vivo [The effects of TH-EDCs on the levels of TH or its receptor will affect downstream signalling including metabolic pathways. Bisphenol A functions as a selective TR-beta antagonist in vivo . TR-beta in vivo . It may In modern society, most individuals will have been exposed to mixtures of EDCs rather than single EDC. Therefore, the study of the effects of combined chemicals is critical to reach meaningful conclusions on the plausible role of these chemicals on human health. Effects of EDCs can either be additive, synergistic, or antagonistic since chemicals may interact with one another to modify the nature of the toxic effect. For instance, human studies  revealedPrenatal exposure to EDCs continues to pose serious health risks to developing fetus and children as evidence of adverse effect on birth outcomes, childhood obesity, and intellectual disability are increasing , 84, 85.The placental barrier is not impervious to TH-EDCs as many of them have been measured in human fetal cord blood , neonataEx vivo studies on human term placentas demonstrated that environmentally relevant levels of BPA in mothers freely diffuse across the placenta in an unconjugated form suggesting that SULTs and UGTs play a minor role in the transplacental transport of BPA. This was attributed to the low expression of placental enzymes involved in metabolizing the chemicals towards the end of pregnancy [β-glucuronidase enzymes in the rat fetus [regnancy . A similregnancy . BPA andregnancy . UGT actregnancy  and to dregnancy . This imat fetus . In addiat fetus  leading  in vitro [γ) signalling pathway.Prenatal exposures to mixtures of TH-EDCs have revealed diverse ways by which chemicals alter TH homeostasis. Perinatal exposures to low doses of BPA increase abdominal adipocyte tissue mass and correlate with hyperlipidemia in a dose-response manner in mice . Howeverin vitro . Althougin vitro . Howeverin vitro . This suin vitro . On the Altered TH levels and postnatal thyroid function have been reported in preterm infants . EpidemiHigher maternal prepregnancy BMI and higher gestational weight gain are correlated with higher birth weight and fat mass at birth and increased BMI in young and adult offspring , 110. Inγ [γ [γ activity and lipid metabolism.Fetal exposures to phthalates and dichlorodiphenyldichloroethylene (DDE) have been associated with increased BMI and head circumference (HC) during the first year after birth in a nonmonotonic manner . Animal γ , but MEHγ [γ . Hence, Perinatal exposure to polychlorinated biphenyls (PCBs) and dioxins and their effects on puberty have been reviewed . ExposurMany studies have associated exposures of persistent organic pollutants with TH dysregulation , 28, 30.A negative association was recorded between PBDE congeners and birth weight; however, when adjusted for maternal weight gain, these findings were no longer statistically significant. Also, no association was observed between PBDEs and birth length, head circumference, or gestational duration . ExposurThe role of TH-EDCs on metabolic risk factors such as insulin resistance, glucose tolerance, and triglycerides and cholesterol levels has only recently become the subject of research. Early studies suggest that TH-EDCs may act via a variety of mechanisms and that studies using high doses of these chemicals are probably not good predictors of effects at low doses. Additionally, low doses of combinations of chemicals, which would be a better reflection of the current situation, have shown conflicting results on the endpoints investigated. Variations in the effects of TH-EDCs on fetal growth have been reported with respect to gender and gestational age. In order to provide better prenatal care and improved health outcomes, it is important that studies be carried out to evaluate the effects of TH-EDCs on lipid levels, insulin sensitivity, and glucose tolerance as they are significant underlying factors in the development of metabolic syndrome.