Genetic polymorphisms to identify patients with an optimal response to tildrakizumab in psoriasis patients from real-life clinical practice.

Detecting the association of genetic variants to the response of biological therapy represents an important advance in developing a personalized therapy. The aim of this work was to study the association of polymorphisms with an optimal response to tildrakizumab in patients with psoriasis in a real-life clinical practice. Ninety patients with plaque psoriasis recruited from-Spanish hospitals receiving tildrakizumab for at least 24 weeks were genotyped for 180 polymorphisms. Optimal response to tildrakizumab was evaluated by absolute PASI ≤1 at 6 and 12 months. Polymorphisms corrected for weight and disease duration with an FDR <0.15 were included in a multiple regression model. Sixty three percent of patients achieved an absolute PASI ≤1 at 6 months, while 71% did so after 12 months. Disease duration (>27 years) and weight (>76 kg) were associated with treatment response; after correcting by these factors, no association (FDR >0.15) was found for any polymorphism and response to tildrakizumab at 6 months. The analysis at 12 months identified the genotype GG for rs610604 (TNFAIP3), CT for rs9373839 (ATG5), and delCTGT/delCTGT for rs72167053 (PDE4D) as risk factors to not achieve an optimal response (PASI ≤1), while CT for rs708567 (IL17RC) was protective, independently of weight and disease duration (FDR <0.15). The final multivariable model at 12 months showed an AUC of 0.90 (95% CI 0.82 to -0.98). We identified a set of polymorphisms that could be helpful to identify psoriatic patients with an optimal response to tildrakizumab at 12 months in real-world practice conditions.

The upside of cumulative conceptual interference on exemplar-level mnemonic discrimination.

Although long-term visual memory (LTVM) has a remarkable capacity, the fidelity of its episodic representations can be influenced by at least two intertwined interference mechanisms during the encoding of objects belonging to the same category: the capacity to hold similar episodic traces (e.g., different birds) and the conceptual similarity of the encoded traces (e.g., a sparrow shares more features with a robin than with a penguin). The precision of episodic traces can be tested by having participants discriminate lures (unseen objects) from targets (seen objects) representing different exemplars of the same concept (e.g., two visually similar penguins), which generates interference at retrieval that can be solved if efficient pattern separation happened during encoding. The present study examines the impact of within-category encoding interference on the fidelity of mnemonic object representations, by manipulating an index of cumulative conceptual interference that represents the concurrent impact of capacity and similarity. The precision of mnemonic discrimination was further assessed by measuring the impact of visual similarity between targets and lures in a recognition task. Our results show a significant decrement in the correct identification of targets for increasing interference. Correct rejections of lures were also negatively impacted by cumulative interference as well as by the visual similarity with the target. Most interestingly though, mnemonic discrimination for targets presented with a visually similar lure was more difficult when objects were encoded under lower, not higher, interference. These findings counter a simply additive impact of interference on the fidelity of object representations providing a finer-grained, multi-factorial, understanding of interference in LTVM.

Pharmacogenetic biomarkers for secukinumab response in psoriasis patients in real-life clinical practice.

BACKGROUND: Prediction of the response to a biological treatment in psoriasis patients would allow efficient treatment allocation. OBJECTIVE: To identify polymorphisms associated with secukinumab response in psoriasis patients in a daily practice setting. METHODS: We studied 180 SNPs in patients with moderate-to-severe plaque psoriasis recruited from 15 Spanish hospitals. Treatment effectiveness was evaluated by absolute PASI ≤3 and ≤1 at 6 and 12 months. Individuals were genotyped using a custom Taqman array. Multiple logistic regression models were generated. Sensitivity, specificity and area under the curve (AUC) were analysed. RESULTS: A total of 173 patients were studied at 6 months, (67% achieved absolute PASI ≤ 3 and 65% PASI ≤ 1) and 162 at 12 months (75% achieved absolute PASI ≤ 3 and 64% PASI ≤ 1). Multivariable analysis showed the association of different sets of SNPs with the response to secukinumab. The model of absolute PASI≤3 at 6 months showed best values of sensitivity and specificity. Four SNPs were associated with the capability of achieving absolute PASI ≤ 3 at 6 months. rs1801274 (FCGR2A), rs2431697 (miR-146a) and rs10484554 (HLCw6) were identified as risk factors for failure to achieve absolute PASI≤3, while rs1051738 (PDE4A) was protective. AUC including these genotypes, weight of patients and history of biological therapy was 0.88 (95% CI 0.83-0.94), with a sensitivity of 48.6% and specificity of 95.7% to discriminate between both phenotypes. CONCLUSION: We have identified a series of polymorphisms associated with the response to secukinumab capable of predicting the potential response/non-response to this drug in patients with plaque psoriasis.

Influence of abiotic factors on diapause termination and temperature requirements for postdiapause development in the European red mite, Panonychus ulmi (Acari: Tetranychidae).

The European red mite Panonychus ulmi (Koch) is widely distributed and it can severely affect pome fruit crops, particularly apple. Pest outbreaks are related to an overuse of non-selective pesticide treatments that lead to the development of resistance and the absence of natural enemies in the orchard. A key aspect to optimize the use of pesticide treatments in the context of IPM is to increase the knowledge on the biology and ecology of the pest to better predict population dynamics and outbreaks. For the European red mite, knowledge on the conditions that lead to diapause breaking by winter eggs is essential to model population dynamics. To increase this knowledge, winter eggs were collected during field surveys in northen Spain during three years and egg hatching was monitored under controlled temperature and photoperiod conditions in the laboratory. The "number of days exposed to cold temperatures" was the most significant factor that positively affected hatching of overwintering eggs. The time required for 50% of the egg population to hatch (T50%) was also negatively modulated by the duration of exposure to cold temperature. The temperature threshold for postdiapause eggs development collected from the field was estimated between 5 and 6 ºC in 2005 and 2007, respectively. Moreover, the degree-days required for post diapause development were estimated between 263.2 and 270.3, depending on the year of collection. Collectively, we provide additional information on the diapause termination and postdiapause development of the European red mite that may effectively contribute to optimize pest population models.

Differential effects of intrinsic properties of natural scenes and interference mechanisms on recognition processes in long-term visual memory.

Humans display remarkable long-term visual memory (LTVM) processes. Even though images may be intrinsically memorable, the fidelity of their visual representations, and consequently the likelihood of successfully retrieving them, hinges on their similarity when concurrently held in LTVM. In this debate, it is still unclear whether intrinsic features of images (perceptual and semantic) may be mediated by mechanisms of interference generated at encoding, or during retrieval, and how these factors impinge on recognition processes. In the current study, participants (32) studied a stream of 120 natural scenes from 8 semantic categories, which varied in frequencies (4, 8, 16 or 32 exemplars per category) to generate different levels of category interference, in preparation for a recognition test. Then they were asked to indicate which of two images, presented side by side (i.e. two-alternative forced-choice), they remembered. The two images belonged to the same semantic category but varied in their perceptual similarity (similar or dissimilar). Participants also expressed their confidence (sure/not sure) about their recognition response, enabling us to tap into their metacognitive efficacy (meta-d'). Additionally, we extracted the activation of perceptual and semantic features in images (i.e. their informational richness) through deep neural network modelling and examined their impact on recognition processes. Corroborating previous literature, we found that category interference and perceptual similarity negatively impact recognition processes, as well as response times and metacognitive efficacy. Moreover, images semantically rich were less likely remembered, an effect that trumped a positive memorability boost coming from perceptual information. Critically, we did not observe any significant interaction between intrinsic features of images and interference generated either at encoding or during retrieval. All in all, our study calls for a more integrative understanding of the representational dynamics during encoding and recognition enabling us to form, maintain and access visual information.

[Advanced Surgical Techniques in Forehead Feminization and Hairline Redefinition]. / Techniques chirurgicales avancées dans la féminisation du front et la redéfinition de la ligne des cheveux chez les patientes transgenres.

Facial Gender-Affirming Surgery (FGAS) has emerged as a transformative option for individuals who wish to align their external appearance with their asserted gender identity. This article delves into the surgical techniques employed in forehead feminization and hairline redefinition, highlighting the nuanced approaches used to modify specific facial characteristics to achieve the desired feminizing outcomes. Our extensive experience, encompassing over 2300 forehead feminization surgeries conducted over the past 16 years, provides a robust foundation for understanding the complexities and intricacies of these procedures. This knowledge is crucial for maxillofacial and plastic surgeons, as well as other healthcare professionals involved in comprehensive gender-affirming care, ensuring they are well-equipped to deliver optimal results for their patients.

Preserved Extra-Foveal Processing of Object Semantics in Alzheimer's Disease.

Alzheimer's disease (AD) patients underperform on a range of tasks requiring semantic processing, but it is unclear whether this impairment is due to a generalised loss of semantic knowledge or to issues in accessing and selecting such information from memory. The objective of this eye-tracking visual search study was to determine whether semantic expectancy mechanisms known to support object recognition in healthy adults are preserved in AD patients. Furthermore, as AD patients are often reported to be impaired in accessing information in extra-foveal vision, we investigated whether that was also the case in our study. Twenty AD patients and 20 age-matched controls searched for a target object among an array of distractors presented extra-foveally. The distractors were either semantically related or unrelated to the target (e.g., a car in an array with other vehicles or kitchen items). Results showed that semantically related objects were detected with more difficulty than semantically unrelated objects by both groups, but more markedly by the AD group. Participants looked earlier and for longer at the critical objects when these were semantically unrelated to the distractors. Our findings show that AD patients can process the semantics of objects and access it in extra-foveal vision. This suggests that their impairments in semantic processing may reflect difficulties in accessing semantic information rather than a generalised loss of semantic memory.

Can physiological stimulation prior to ileostomy closure reduce postoperative ileus? A prospective multicenter pilot study.

BACKGROUND: The aim of this study was to assess the impact of ileostomy closure following preoperative physiological stimulation (PPS) on postoperative ileus (POI) in patients with loop ileostomy after low anterior resection for rectal cancer. METHODS: Patients who underwent ileostomy closure between January 2017 and February 2020 in two tertiary referral centers were prospectively included. PPS stimulation was compared to standard treatment. Stimulation was carried out daily during the 15 days prior to ileostomy closure by the patient's self-instillation of 200 ml of fecal contents from the ileostomy bag via the efferent loop, using a rectal catheter. Standard treatment (ST) consisted of observation. Outcomes measures were POI, morbidity, stimulation feasibility, and predictors to ileus. RESULTS: A total of 58 patients were included [42 males and 16 females, median age 67 (43-85) years]. PPS was used in 24 patients, who completed the entire stimulation process, and ST in 34 patients. No differences in preoperative factors were found between the two groups. POI was significantly lower in the PPS group (4.2%) vs the ST group (32.4%); p < 0.01, OR: 0.05 (CI 95% 0.01-0.65). The PPS group had a shorter time to restoration of bowel function (1 day vs 3 days) p = 0.02 and a shorter time to tolerance of liquids (1 day vs 2 days), p = 0.04. Age (p = 0.01), open approach at index surgery, p = 0.03, adjuvant capecitabine (p = 0.01). and previous abdominal surgeries (p = 0.02) were associated with POI in the multivariate analysis. C-reactive-protein values on the 3rd (p = 0.02) and 5th (p < 0.01) postoperative day were also associated with POI. CONCLUSIONS: PPS for patients who underwent ileostomy closure after low anterior resection for rectal cancer is feasible and might reduce POI.

[Hand hygiene and pandemic. Controversies]. / Higiene de manos y pandemia. Controversias.

During the COVID-19 pandemic, the world's healthcare systems were extremely strained. Intensive care units were stretched to capacity and healthcare facilities were forced to set up spaces to care for critically ill patients. Professionals were required to work in strenuous conditions, completely disrupting their work routines.In this scenario, hand hygiene and the use of gloves by healthcare professionals became a critical point of transmission risk.The results of the ENVIN study in 2020 and 2021, corresponding to the pandemic period, showed worrying data on the increase in infection rates, with rates rising by 250% at the worst moments of the pandemic. This suggested that excessive risk situations were occurring for the patient. Any preventive strategy must place correct hand hygiene and proper use of gloves among its priority objectives. For this reason, the Project Zero Advisory Board made a series of adaptations and recommendations based on available evidence and expert opinion related to hand hygiene and glove use during the pandemic situation to promote best practice in extreme situations. This article reviews the key aspects of hand hygiene as part of the WHO safety strategy, the main barriers to compliance and the main adaptations proposed by the Advisory Board of the Zero projects.

Single-cell RNA sequencing of SARS-CoV-2 cell entry factors in the preconceptional human endometrium.

STUDY QUESTION: Are SARS-CoV-2 canonical cell entry machinery, consisting of ACE2, TMPRSS2, NRP1 and LY6E, or alternative potential cell entry machinery, consisting of BSG, ANPEP, CD209, CLEC4G, TMPRSS4, TMPRSS11A, FURIN, CTSB, CTSL and IFITM1, expressed in the human endometrium across the menstrual cycle? SUMMARY ANSWER: Analysis of cell entry factors for SARS-CoV-2 by single-cell RNA-sequencing (scRNAseq) in the preconceptional human endometrium reveals low risk of infection. WHAT IS KNOWN ALREADY: Gene expression datasets from bulk endometrial tissue show no significant expression of the SARS-CoV-2 receptor ACE2 and TMPRSS2. This is in contrast to reported expression of ACE2 at the single-cell level in the decidua and trophoblast cells at the maternal-fetal interface in early pregnancy, as well as vertical transmission of SARS-CoV-2 during pregnancy. STUDY DESIGN, SIZE, DURATION: This analysis of SARS-CoV-2 cell entry machinery gene expression was conducted by scRNAseq in 73 181 human endometrial cells isolated from endometrial biopsies obtained from 27 donors across the menstrual cycle. PARTICIPANTS/MATERIALS, SETTING, METHODS: ScRNAseq examined the expression of genes encoding cell entry machinery for SARS-CoV-2. The raw data were from a previously published dataset. MAIN RESULTS AND THE ROLE OF CHANCE: ScRNAseq analysis showed no significant expression of ACE2 in stromal or unciliated epithelial cells in any phase of the menstrual cycle. TMPRSS2 was expressed in epithelial cells during the early proliferative and mid-secretory phases. Interestingly, the expression of NRP1 was observed in both stromal and epithelial cells across all phases of the menstrual cycle, and LY6E was highly expressed in stromal cells. In the mid-secretory phase, coexpression of ACE2 and TMPRSS2 was detected in 0.07% of luminal epithelial cells. No cells simultaneously expressed ACE2, NRP1 and TMPRSS2 at the time of embryo implantation. Focusing on non-canonical cell entry machinery, BSG was highly expressed in all cell types across the menstrual cycle and may interact with CTSB or CTSL proteases, but viral infection using this machinery has not yet been confirmed. LARGE SCALE DATA: All raw data in this study can be found at NCBI's Gene Expression Omnibus (series accession code GSE111976) and Sequence Read Archive (accession code SRP135922). LIMITATIONS, REASONS FOR CAUTION: Our findings at the single-cell level imply low efficiency of SARS-CoV-2 endometrial infection using canonical receptors in a cohort of healthy reproductive-age women; however, infection of endometrial cells can only be assessed in the presence of the virus. All samples were processed for scRNAseq, so no samples are remaining to analyze protein expression or spatial transcriptomics. WIDER IMPLICATIONS OF THE FINDINGS: Our results offer a useful resource to guide reproductive decisions when assessing risk of endometrial infection by SARS-CoV-2 during the preconceptional period in asymptomatic COVID-19 carriers. STUDY FUNDING/COMPETING INTEREST(S): This study was jointly supported by the March of Dimes, Chan Zuckerberg Biohub and MINECO/FEDER (SAF-2015-67164-R, to C.S.) (Spanish Government), and the European Union's Horizon 2020 Framework Programme for Research and Innovation (Grant agreement 874867). W.W. was supported by the Stanford Bio-X Graduate Bowes Fellowship and Chan Zuckerberg Biohub. F.V. was supported by the Miguel Servet Program Type II of ISCIII (CPII18/00020) and the FIS project (PI18/00957). A patent disclosure has been filed for the study with the title 'Methods for assessing endometrial transformation' and the global patent number 'EP 3807648 A2' under the inventors S.R.Q., C.S., W.W. and F.V. C.S. is the Founder and Head of the Scientific Advisory Board of Igenomix SL. S.R.Q is the Director of Mirvie. I.M. is partially employed by Igenomix SL. B.R. has no interests to declare.

Characterization of Globodera ellingtonae Populations from Chile Utilizing Whole Genome Sequencing.

Globodera ellingtonae was originally described from populations collected in the United States. In the original description, ribosomal DNA loci from Globodera sp. collected in Chile and Argentina were similar to G. ellingtonae, suggesting this nematode originated in this region of South America. In an effort to find additional populations of G. elllingtonae, collection trips were conducted in 2017 and 2020 in the Antofagasta and Arica y Parinacota Regions in Northern Chile, respectively. Globodera sp. were more prevalent in Antofagasta (17 samples collected, 53% positive for Globodera sp.) than in Arica y Parincota (16 samples collected, 13% positive for Globodera sp.). The genomes of single cysts (N ≥ 3) from four fields were sequenced. Additionally, the genomes of the G. ellingtonae population from Oregon and a Globodera sp. population originally collected in Antofagasta Region but maintained in culture in France were also sequenced. Based upon a HSP90 sequenced data mined from WSG data, all of the populations from the Antofagasta Region were G. ellingtonae and grouped in a monophyletic clade. A population collected from the Arica y Parincota Region was identified as G. rostochiensis based upon HSP90 data. Genome-wide SNP patterns of the G. ellingtonae populations showed strong clustering based on geographic location indicating that G. ellingtonae has high genetic diversity within Chile. A phylogenetic tree derived from 168,354 binary SNPs in the nuclear genome showed separate but distinct clustering of the Oregon population and the population from Antofagasta maintained in France. The Oregon G. ellingtonae population subtended the Chilean clades and placed on a long branch representing approximately twice the genetic variation observed among all Chilean G. ellingtonae populations. The possibility remains that G. ellingtonae from Chile may be sufficiently diverged to constitute a new species from G. ellingtonae originally described from a population collected in Oregon.

Fixation-related Brain Potentials during Semantic Integration of Object-Scene Information.

In vision science, a particularly controversial topic is whether and how quickly the semantic information about objects is available outside foveal vision. Here, we aimed at contributing to this debate by coregistering eye movements and EEG while participants viewed photographs of indoor scenes that contained a semantically consistent or inconsistent target object. Linear deconvolution modeling was used to analyze the ERPs evoked by scene onset as well as the fixation-related potentials (FRPs) elicited by the fixation on the target object (t) and by the preceding fixation (t - 1). Object-scene consistency did not influence the probability of immediate target fixation or the ERP evoked by scene onset, which suggests that object-scene semantics was not accessed immediately. However, during the subsequent scene exploration, inconsistent objects were prioritized over consistent objects in extrafoveal vision (i.e., looked at earlier) and were more effortful to process in foveal vision (i.e., looked at longer). In FRPs, we demonstrate a fixation-related N300/N400 effect, whereby inconsistent objects elicit a larger frontocentral negativity than consistent objects. In line with the behavioral findings, this effect was already seen in FRPs aligned to the pretarget fixation t - 1 and persisted throughout fixation t, indicating that the extraction of object semantics can already begin in extrafoveal vision. Taken together, the results emphasize the usefulness of combined EEG/eye movement recordings for understanding the mechanisms of object-scene integration during natural viewing.

Roles of the multiplex real-time PCR assay and ß-D-glucan in a high-risk population for intra-abdominal candidiasis (IAC).

Multiplex quantitative real-time PCR (MRT-PCR) using blood can improve the diagnosis of intra-abdominal candidiasis (IAC). We prospectively studied 39 patients with suspected IAC in the absence of previous antifungal therapy. Blood cultures, MRT-PCR, and ß-D-glucan (BDG) in serum were performed in all patients. IAC was defined according to the 2013 European Consensus criteria. For MRT-PCR, the probes targeted the ITS1 or ITS2 regions of ribosomal DNA. Candidaemia was confirmed only in four patients (10%), and IAC criteria were present in 17 patients (43.6%). The sensitivity of MRT-PCR was 25% but increased to 63.6% (P = .06) in plasma obtained prior to volume overload and transfusion; specificity was above 85% in all cases. BDG performance was improved using a cutoff > 260 pg/ml, and improvement was not observed in samples obtained before transfusion. In this cohort of high risk of IAC and low rate of bloodstream infection, the performance of non-culture-based methods (MRT-PCR or BDG) was moderate but may be a complementary tool given the limitations of diagnostic methods available in clinical practice. Volume overload requirements, in combination with other factors, decrease the accuracy of MRT-PCR in patients with IAC.

Tuberculosis caused by Mycobacterium caprae in a camel (Camelus dromedarius).

BACKGROUND: Animal tuberculosis (TB) is distributed worldwide and has a wide range of wild and domestic reservoirs. Few studies concerning TB in camelids have been published in the last decade, particularly as regards Old World Camelids (OWC), but the increase in reports of TB outbreaks in these species in recent years suggests a high susceptibility to the infection. CASE PRESENTATION: We studied a dromedary camel (Camelus dromedarius) herd (n = 24) in which a Mycobacterium caprae infection was detected. The TB infection was confirmed in one animal at necropsy through the detection of TB lesions, mainly in the abdominal organs, and the subsequent isolation of M. caprae (SB0157 spoligotype). The whole herd was additionally tested using cellular and humoral based diagnostic techniques. The intradermal tuberculin test results were compared with those obtained using P22 ELISA for the detection of specific antibodies against the M. tuberculosis complex. The TB infected animal was a positive reactor to both the intradermal tuberculin tests and P22 ELISA, while the others were negative to all the diagnostic tests. CONCLUSION: The present study found M. caprae infection in OWC. This is the first report of M. caprae infection in an OWC not living in a zoo. Since the animal was born in the herd and fed with goat's milk, this practice was suspected to be the potential source of TB infection, which was not confirmed in the other animals present in the herd. Moreover, our results highlight that the intradermal tuberculin test and the P22 ELISA could be valuable tools for the diagnosis of TB in OWC.

Specificity of serological test for detection of tuberculosis in cattle, goats, sheep and pigs under different epidemiological situations.

BACKGROUND: Serum antibody detection has potential as a complementary diagnostic tool in animal tuberculosis (TB) control, particularly in multi-host systems. The objective of the present study was to assess the specificity (Sp) of an enzyme-linked immunosorbent assay (ELISA) based on the new multiprotein complex P22 for the detection of specific antibodies against the Mycobacterium tuberculosis complex (MTC) in the four most relevant domestic animals acting as MTC hosts: cattle, goat, sheep and pig. We used sera from an officially TB-free (OTF) country, Norway, and from a non-OTF one, Spain. The samples included sera from goats that had been vaccinated against M. avium subsp. paratuberculosis (MAP) and sheep from a herd in which Corynebacterium pseudotuberculosis had been isolated. RESULTS: In cattle, the Sp ranged from 92.5 (IC95% 90.7-94) to 99.4% (IC95% 98.3-99.8) depending on the cut-off used and the origin of the samples (Spain or Norway). Sp in cattle (cut-off point 100) was significantly higher (P < 0.05) for Norwegian samples. By contrast, Sp in goats was consistently low at the 100 cut-off [30.9 (CI95%23.4-39.5)-78% (CI95% 68.9-85)]. A higher cut-off of 150 improved Sp in Norwegian goats [97% (CI95% 91.6-99)], but still yielded a poor Sp of 56.1% (CI95% 47.3-64.6) in Spanish goats. In Norway at the 100 cut-off the Sp was 58.3 (CI95% 42.2-72.9) and 90.6% (CI95% 81-95.6) in MAP vaccinated and non-vaccinated goats, respectively, indicating interference due to MAP vaccination. Sp in sheep was between 94.4 (CI95% 91.7-96.3) and 100% (CI95% 96.3-100) depending on the cut-off and country, and no diagnostic interference due to infection with C. pseudotuberculosis was recorded. Sp in pigs was 100%, regardless the cut-off point applied, and no significant differences were observed between pigs from Norway and from Spain. CONCLUSIONS: Due to its excellent Sp in pigs and acceptable Sp in cattle and sheep, this ELISA may constitute a suitable option for TB screening at herd level, particularly in OTF-countries.

BOPHYs versus BODIPYs: A comparison of their performance as effective multi-function organic dyes.

The computationally-aided photophysical and lasing properties of a selected battery of BOPHYs are described and compared to those of related BODIPY counterparts. The present joined theoretical-experimental study helps to put into context the weaknesses and strengths of both dye families under different irradiation conditions. The chemical versatility of the BOPHY scaffold has been also comparatively explored to modulate key photonic properties towards the development of red-emitting dyes, chiroptical dyes and singlet oxygen photosensitizers. Thus, BOPHY BINOLation by fluorine substitution with enantiopure BINOLs endows the BOPHY chromophore with chiroptical activity, as supporting by the simulated circular dichroism, decreasing deeply its fluorescent response due to the promotion of fluorescence-quenching intramolecular charge transfer (ICT). Interestingly, the sole alkylation of the BOPHY core strongly modulates the promotion of ICT, allowing the generation of highly bright BINOL-based BOPHY dyes. Moreover, 3,3'-dibromoBINOLating BOPHYs can easily achieve singlet-oxygen photogeneration, owing to spin-orbit coupling mediated by heavy-atom effect feasible in view of the theoretically predicted disposition of the bromines surrounding the chromophore. From this background, we have established the master guidelines to design bright fluorophores and laser dyes, photosensitizers for singlet oxygen production and chiroptical dyes based on BOPHYs. The possibility to finely mix and balance such properties in a given molecular scaffold outstands BOPHYs as promising dyes competing with the well-settled BODIPY dyes.

Heterogeneous nuclear ribonucleoprotein C1 may control miR-30d levels in endometrial exosomes affecting early embryo implantation.

STUDY QUESTION: Is there a specific mechanism to load the microRNA (miRNA), hsa-miR-30d, into exosomes to facilitate maternal communication with preimplantation embryos? SUMMARY ANSWER: The heterogeneous nuclear ribonucleoprotein C1 (hnRNPC1) is involved in the internalization of endometrial miR-30d into exosomes to prepare for its subsequent incorporation into trophectoderm cells. WHAT IS KNOWN ALREADY: Our group previously described a novel cell-to-cell communication mechanism involving the delivery of endometrial miRNAs from the maternal endometrium to the trophectoderm cells of preimplantation embryos. Specifically, human endometrial miR-30d is taken up by murine blastocysts causing the overexpression of certain genes involved in embryonic adhesion (Itb3, Itga7 and Cdh5) increasing embryo adhesion rates. STUDY DESIGN, SIZE, DURATION: Transfer of maternal miR-30d to preimplantation embryos was confirmed by co-culture of wild-type (WT) and miR-30d knockout (KO) murine embryos with primary cultures of human endometrial epithelial cells (hEECs) in which mir-30d was labeled with specific Molecular Beacon (MB) or SmartFlare probes. Potential molecules responsible for the miR-30d loading into exosomes were purified by pull-down analysis with a biotinylated form of miR-30d on protein lysates from human endometrial exosomes, identified using mass spectrometry and assessed by flow cytometry, western blotting and co-localization studies. The role of hnRNPC1 in the miR-30d loading and transportation was interrogated by quantification of this miRNA in exosomes isolated from endometrial cells in which hnRNPC1 was transiently silenced using small interference RNA. Finally, the transfer of miR-30d to WT and KO embryos was assessed upon co-culture with sihnRNPC1 transfected cells. PARTICIPANTS/MATERIALS, SETTING, METHODS: Murine embryos from miR-30d WT and KO mice, (strain MirC26tm1Mtm/Mmjax), were obtained by oviduct flushing of superovulated females. Endometrial Exosomes were purified by ultracentrifugation of supernatants from primary cultures of hEECs or Ishikawa cells. MB and Smartflare miR-30d probes were detected by confocal and/or transmission electron microscopy (TEM). hEECs and exosomes derived from them were subjected to pull-down with a biotinylated form of miR-30d. Captured proteins were identified by mass spectrometry (MS/MS). Western blotting was performed to detect hnRNPC1 and CYR61 in whole lysates, subcellular fractions and secreted vesicles from hEECs. Co-localization studies of the selected proteins with the exosomal marker CD63 were performed. FACS analysis was carried out to determine the presence of hnRNPC1 inside exosomes. Silencing of hnRNPC1 was conducted in the Ishikawa Cell Line with the Smart Pool Accell HNRNPC siRNA at a final concentration of 50 nM. RT-qPCRs were done to determine the messenger levels of miR-30d in cells and exosomes. Co-cultures of WT and KO embryos were established with Ishikawa cells double-transfected with sihnRPNC1 and MB probes. MAIN RESULTS AND THE ROLE OF CHANCE: MS/MS analysis allowed us to identify hnRNPC1 as a possible protein to influence miR-30d loading into exosomes. Co-localization studies of hnRNPC1 with CD63 and FACS analyses suggested the presence of hnRNPC1 inside exosomes. Silencing of hnRNPC1 in Ishikawa cells resulted in a sharp decrease of the levels of miR-30d in both epithelial-like cells (P = 0.0001) and exosomes (P = 0.0152), suggesting its potential role in miR-30d biogenesis and transfer. Co-culture assays of miR-30d KO embryos with sihnRNPC1 hEECs revealed a decrease in embryo-miR-30d acquisition during the adhesion and invasion stages. In turn, transient silencing of hnRNPC1 results in a significant decrease of blastocyst adhesion compared to mock transfection conditions using Block-it, in both WT [Mean ± SD; 67 ± 10.0% vs. 38 ± 8.5%(P = 0.0006)] and miR-30d KO embryos [Mean ± SD; 50 ± 11.5% vs. 26 ± 8.8% (P = 0.0029) (n = 2); 14 embryos transferred per condition tested]. LARGE-SCALE DATA: MS/MS data are available via ProteomeXchange with identifier PXD008773. LIMITATIONS, REASONS FOR CAUTION: The Ishikawa Cell Line was used as a model of hEECs in silencing experiments due to the low survival rates of primary hEECs after transfection. WIDER IMPLICATIONS OF THE FINDINGS: The data show that hnRNPC1 may be involved in the internalization of miR-30d inside exosomes. The decreased rates of embryo adhesion in endometrial epithelial-like cells transiently silenced with sihnRNPC1evidence that hnRNPC1 could be an important player in the maternal-embryo communication established in the early stages of implantation. STUDY FUNDING AND COMPETING INTEREST(S): This work was supported by the Miguel Servet Program Type I of Instituto de Salud Carlos III [CP13/00038]; FIS project [PI14/00545] to F.V.; the 'Atracció de Talent' Program from VLC-CAMPUS [UV-INV-PREDOC14-178329 to NB]; a Torres-Quevedo grant (PTQ-13-06133) by the Spanish Ministry of Economy and Competitiveness to IM and MINECO/FEDER Grant [SAF2015-67154-R] to C.S. The authors declare there is no conflict of interest.

Origin and composition of cell-free DNA in spent medium from human embryo culture during preimplantation development.

STUDY QUESTION: What is the origin and composition of cell-free DNA in human embryo spent culture media? SUMMARY ANSWER: Cell-free DNA from human embryo spent culture media represents a mix of maternal and embryonic DNA, and the mixture can be more complex for mosaic embryos. WHAT IS KNOWN ALREADY: In 2016, ~300 000 human embryos were chromosomally and/or genetically analyzed using preimplantation genetic testing for aneuploidies (PGT-A) or monogenic disorders (PGT-M) before transfer into the uterus. While progress in genetic techniques has enabled analysis of the full karyotype in a single cell with high sensitivity and specificity, these approaches still require an embryo biopsy. Thus, non-invasive techniques are sought as an alternative. STUDY DESIGN, SIZE, DURATION: This study was based on a total of 113 human embryos undergoing trophectoderm biopsy as part of PGT-A analysis. For each embryo, the spent culture media used between Day 3 and Day 5 of development were collected for cell-free DNA analysis. In addition to the 113 spent culture media samples, 28 media drops without embryo contact were cultured in parallel under the same conditions to use as controls. In total, 141 media samples were collected and divided into two groups: one for direct DNA quantification (53 spent culture media and 17 controls), the other for whole-genome amplification (60 spent culture media and 11 controls) and subsequent quantification. Some samples with amplified DNA (N = 56) were used for aneuploidy testing by next-generation sequencing; of those, 35 samples underwent single-nucleotide polymorphism (SNP) sequencing to detect maternal contamination. Finally, from the 35 spent culture media analyzed by SNP sequencing, 12 whole blastocysts were analyzed by fluorescence in situ hybridization (FISH) to determine the level of mosaicism in each embryo, as a possible origin for discordance between sample types. PARTICIPANTS/MATERIALS, SETTING, METHODS: Trophectoderm biopsies and culture media samples (20 µl) underwent whole-genome amplification, then libraries were generated and sequenced for an aneuploidy study. For SNP sequencing, triads including trophectoderm DNA, cell-free DNA, and follicular fluid DNA were analyzed. In total, 124 SNPs were included with 90 SNPs distributed among all autosomes and 34 SNPs located on chromosome Y. Finally, 12 whole blastocysts were fixed and individual cells were analyzed by FISH using telomeric/centromeric probes for the affected chromosomes. MAIN RESULTS AND THE ROLE OF CHANCE: We found a higher quantity of cell-free DNA in spent culture media co-cultured with embryos versus control media samples (P ≤ 0.001). The presence of cell-free DNA in the spent culture media enabled a chromosomal diagnosis, although results differed from those of trophectoderm biopsy analysis in most cases (67%). Discordant results were mainly attributable to a high percentage of maternal DNA in the spent culture media, with a median percentage of embryonic DNA estimated at 8%. Finally, from the discordant cases, 91.7% of whole blastocysts analyzed by FISH were mosaic and 75% of the analyzed chromosomes were concordant with the trophectoderm DNA diagnosis instead of the cell-free DNA result. LIMITATIONS, REASONS FOR CAUTION: This study was limited by the sample size and the number of cells analyzed by FISH. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study to combine chromosomal analysis of cell-free DNA, SNP sequencing to identify maternal contamination, and whole-blastocyst analysis for detecting mosaicism. Our results provide a better understanding of the origin of cell-free DNA in spent culture media, offering an important step toward developing future non-invasive karyotyping that must rely on the specific identification of DNA released from human embryos. STUDY FUNDING/ COMPETING INTEREST: This work was funded by Igenomix S.L. There are no competing interests.

The potential predictive value of DEK expression for neoadjuvant chemoradiotherapy response in locally advanced rectal cancer.

BACKGROUND: Limited data are available regarding the ability of biomarkers to predict complete pathological response to neoadjuvant chemoradiotherapy in locally advanced rectal cancer. Complete response translates to better patient survival. DEK is a transcription factor involved not only in development and progression of different types of cancer, but is also associated with treatment response. This study aims to analyze the role of DEK in complete pathological response following chemoradiotherapy for locally advanced rectal cancer. METHODS: Pre-treated tumour samples from 74 locally advanced rectal-cancer patients who received chemoradiation therapy prior to total mesorectal excision were recruited for construction of a tissue microarray. DEK immunoreactivity from all samples was quantified by immunohistochemistry. Then, association between positive stained tumour cells and pathologic response to neoadjuvant treatment was measured to determine optimal predictive power. RESULTS: DEK expression was limited to tumour cells located in the rectum. Interestingly, high percentage of tumour cells with DEK positiveness was statistically associated with complete pathological response to neoadjuvant treatment based on radiotherapy and fluoropyrimidine-based chemotherapy and a marked trend toward significance between DEK positiveness and absence of treatment toxicity. Further analysis revealed an association between DEK and the pro-apoptotic factor P38 in the pre-treated rectal cancer biopsies. CONCLUSIONS: These data suggest DEK as a potential biomarker of complete pathological response to treatment in locally advanced rectal cancer.