Characterisation of C101248: A novel selective THIK-1 channel inhibitor for the modulation of microglial NLRP3-inflammasome.

Neuroinflammation, specifically the NLRP3 inflammasome cascade, is a common underlying pathological feature of many neurodegenerative diseases. Evidence suggests that NLRP3 activation involves changes in intracellular K+. Nuclear Enriched Transcript Sort Sequencing (NETSseq), which allows for deep sequencing of purified cell types from human post-mortem brain tissue, demonstrated a highly specific expression of the tandem pore domain halothane-inhibited K+ channel 1 (THIK-1) in microglia compared to other glial and neuronal cell types in the human brain. NETSseq also showed a significant increase of THIK-1 in microglia isolated from cortical regions of brains with Alzheimer's disease (AD) relative to control donors. Herein, we report the discovery and pharmacological characterisation of C101248, the first selective small-molecule inhibitor of THIK-1. C101248 showed a concentration-dependent inhibition of both mouse and human THIK-1 (IC50: ∼50 nM) and was inactive against K2P family members TREK-1 and TWIK-2, and Kv2.1. Whole-cell patch-clamp recordings of microglia from mouse hippocampal slices showed that C101248 potently blocked both tonic and ATP-evoked THIK-1 K+ currents. Notably, C101248 had no effect on other constitutively active resting conductance in slices from THIK-1-depleted mice. In isolated microglia, C101248 prevented NLRP3-dependent release of IL-1ß, an effect not seen in THIK-1-depleted microglia. In conclusion, we demonstrated that inhibiting THIK-1 (a microglia specific gene that is upregulated in brains from donors with AD) using a novel selective modulator attenuates the NLRP3-dependent release of IL-1ß from microglia, which suggests that this channel may be a potential therapeutic target for the modulation of neuroinflammation in AD.

Negative-pressure wound therapy as an adjunct to treating pyoderma gangrenosum.

Negative-pressure wound therapy has been shown to be effective in re-epithelialisation in recalcitrant pyoderma gangrenosum. This case documents a patient whose co-morbidities limited the standard treatment options and required an alternative management plan. The STOP GAP randomised control trial found that both oral prednisolone and ciclosporin were equally effective in the treatment of pyoderma gangrenosum. However, in this case, the patient had type 2 diabetes and prednisolone resulted in persistently elevated blood glucose levels. Lower doses were ineffective and it was subsequently stopped once other treatments showed a clinical improvement. A recent diagnosis of breast cancer prevented the use of ciclosporin and other immunosuppressive treatments. After multidisciplinary discussion, involving breast surgeons, oncology, dermatology and tissue viability, doxycycline was commenced alongside a portable negative-pressure device with twice weekly tissue viability input. The introduction of this device resulted in a rapid reduction in the wound size and facilitated healing with an excellent outcome.

Endometrial scratch to increase live birth rates in women undergoing first-time in vitro fertilisation: RCT and systematic review.

BACKGROUND: In vitro fertilisation is a widely used reproductive technique that can be undertaken with or without intracytoplasmic sperm injection. The endometrial scratch procedure is an in vitro fertilisation 'add-on' that is sometimes provided prior to the first in vitro fertilisation cycle, but there is a lack of evidence to support its use. OBJECTIVES: (1) To assess the clinical effectiveness, safety and cost-effectiveness of endometrial scratch compared with treatment as usual in women undergoing their first in vitro fertilisation cycle (the 'Endometrial Scratch Trial') and (2) to undertake a systematic review to combine the results of the Endometrial Scratch Trial with those of previous trials in which endometrial scratch was provided prior to the first in vitro fertilisation cycle. DESIGN: A pragmatic, multicentre, superiority, open-label, parallel-group, individually randomised controlled trial. Participants were randomised (1 : 1) via a web-based system to receive endometrial scratch or treatment as usual using stratified block randomisation. The systematic review involved searching electronic databases (undertaken in January 2020) and clinicaltrials.gov (undertaken in September 2020) for relevant trials. SETTING: Sixteen UK fertility units. PARTICIPANTS: Women aged 18-37 years, inclusive, undergoing their first in vitro fertilisation cycle. The exclusion criteria included severe endometriosis, body mass index ≥ 35 kg/m2 and previous trauma to the endometrium. INTERVENTIONS: Endometrial scratch was undertaken in the mid-luteal phase of the menstrual cycle prior to in vitro fertilisation, and involved inserting a pipelle into the cavity of the uterus and rotating and withdrawing it three or four times. The endometrial scratch group then received usual in vitro fertilisation treatment. The treatment-as-usual group received usual in vitro fertilisation only. MAIN OUTCOME MEASURES: The primary outcome was live birth after completion of 24 weeks' gestation within 10.5 months of egg collection. Secondary outcomes included implantation, pregnancy, ectopic pregnancy, miscarriage, pain and tolerability of the procedure, adverse events and treatment costs. RESULTS: One thousand and forty-eight (30.3%) women were randomised to treatment as usual (n = 525) or endometrial scratch (n = 523) and were followed up between July 2016 and October 2019 and included in the intention-to-treat analysis. In the endometrial scratch group, 453 (86.6%) women received the endometrial scratch procedure. A total of 494 (94.1%) women in the treatment-as-usual group and 497 (95.0%) women in the endometrial scratch group underwent in vitro fertilisation. The live birth rate was 37.1% (195/525) in the treatment-as-usual group and 38.6% (202/523) in the endometrial scratch group: an unadjusted absolute difference of 1.5% (95% confidence interval -4.4% to 7.4%; p = 0.621). There were no statistically significant differences in secondary outcomes. Safety events were comparable across groups. No neonatal deaths were recorded. The cost per successful live birth was £11.90 per woman (95% confidence interval -£134 to £127). The pooled results of this trial and of eight similar trials found no evidence of a significant effect of endometrial scratch in increasing live birth rate (odds ratio 1.03, 95% confidence interval 0.87 to 1.22). LIMITATIONS: A sham endometrial scratch procedure was not undertaken, but it is unlikely that doing so would have influenced the results, as objective fertility outcomes were used. A total of 9.2% of women randomised to receive endometrial scratch did not undergo the procedure, which may have slightly diluted the treatment effect. CONCLUSIONS: We found no evidence to support the theory that performing endometrial scratch in the mid-luteal phase in women undergoing their first in vitro fertilisation cycle significantly improves live birth rate, although the procedure was well tolerated and safe. We recommend that endometrial scratch is not undertaken in this population. TRIAL REGISTRATION: This trial is registered as ISRCTN23800982. FUNDING: This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 26, No. 10. See the NIHR Journals Library website for further project information.

The endometrial scratch is a simple procedure that involves 'scratching' the lining of the womb (the endometrium). Several small studies have shown that undertaking this before the first in vitro fertilisation cycle may improve live birth rates; however, other studies have contradicted this. This large study was carried out to confirm whether or not having an endometrial scratch before the first in vitro fertilisation cycle would increase the number of women having a live birth compared with those having 'usual' in vitro fertilisation treatment (known as the 'control' group). We collected information about pregnancy, miscarriage, stillbirth, pain during the procedure and costs of treatment to find out if there were any meaningful differences. A total of 1048 women aged between 18 and 37 years were randomly allocated to the two groups, so participants had a 50% chance of having the endometrial scratch. Women were followed up throughout their pregnancy to ascertain the outcome of their in vitro fertilisation cycle. Although the live birth rate was 1.5% higher in the endometrial scratch group (38.6%) than in the control group (37.1%), the difference was not large enough to show any benefit of having the procedure. Other outcomes did not differ significantly between the two groups. However, the procedure was safe and tolerable. We found that the cost of treatment was, on average, £316 per participant higher in the group that received endometrial scratch than in the control group; the difference was not large enough to show that receiving endometrial scratch was more cost-effective. We combined the results of this trial with those of previous trials that looked to answer a similar question, and found that, overall, the endometrial scratch procedure does not enhance the chances of achieving a live birth. We conclude that endometrial scratch before first-time in vitro fertilisation does not improve the outcome of treatment, and we recommend that this procedure is not undertaken prior to a first cycle of in vitro fertilisation.

The expression and activity of Toll-like receptors in the preimplantation human embryo suggest a new role for innate immunity.

STUDY QUESTION: Is the innate immunity system active in early human embryo development? SUMMARY ANSWER: The pattern recognition receptors and innate immunity Toll-like receptor (TLR) genes are widely expressed in preimplantation human embryos and the pathway appears to be active in response to TLR ligands. WHAT IS KNOWN ALREADY: Early human embryos are highly sensitive to their local environment, however relatively little is known about how embryos detect and respond to specific environmental cues. While the maternal immune response is known to be key to the establishment of pregnancy at implantation, the ability of human embryos to detect and signal the presence of pathogens is unknown. STUDY DESIGN, SIZE, DURATION: Expression of TLR family and related genes in human embryos was assessed by analysis of published transcriptome data (n = 40). Day 5 (D-5) human embryos (n = 25) were cultured in the presence of known TLR ligands and gene expression and cytokine production measured compared to controls. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human embryos surplus to treatment requirements were donated with informed consent from several ART centres. Embryos were cultured to Day 6 (D-6) in the presence of the TLR3 and TLR5 ligands Poly (I: C) and flagellin, with gene expression measured by quantitative PCR and cytokine release into medium measured using cytometric bead arrays. MAIN RESULTS AND THE ROLE OF CHANCE: TLR and related genes, including downstream signalling molecules, were expressed variably at all human embryo developmental stages. Results showed the strongest expression in the blastocyst for TLRs 9 and 5, and throughout development for TLRs 9, 5, 2, 6 and 7. Stimulation of Day 5 blastocysts with TLR3 and TLR5 ligands Poly (I: C) and flagellin produced changes in mRNA expression levels of TLR genes, including the hyaluronan-mediated motility receptor (HMMR), TLR5, TLR7, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and monocyte chemoattractant Protein-1 (MCP-1) (P < 0.05, P < 0.001 compared to unstimulated controls), and release into culture medium of cytokines and chemokines, notably IL8 (P = 0.00005 and 0.01277 for flagellin and Poly (I: C), respectively). LIMITATIONS, REASONS FOR CAUTION: This was a descriptive and experimental study which suggests that the TLR system is active in human embryos and capable of function, but does not confirm any particular role. Although we identified embryonic transcripts for a range of TLR genes, the expression patterns were not always consistent across published studies and expression levels of some genes were low, leaving open the possibility that these were expressed from the maternal rather than embryonic genome. WIDER IMPLICATIONS OF THE FINDINGS: This is the first report of the expression and activity of a number of components of the innate immunity TLR system in human embryos. Understanding the role of TLRs during preimplantation human development may be important to reveal immunological mechanisms and potential clinical markers of embryo quality and pregnancy initiation during natural conception and in ART. STUDY FUNDING/COMPETING INTEREST(S): This work was funded by the Ministry of Higher Education, The State of Libya, the UK Medical Research Council, and the NIHR Local Comprehensive Research Network and NIHR Manchester Clinical Research Facility and the European Union's Horizon 2020 Research and Innovation Programmes under the Marie Sklodowska-Curie Grant Agreement No. 812660 (DohART-NET). In accordance with H2020 rules, no new human embryos were sacrificed for research activities performed from the EU funding, which concerned only in silico analyses of recorded time-lapse and transcriptomics datasets. None of the authors has any conflict of interest to declare. TRIAL REGISTRATION NUMBER: n/a.

Chemical signals from eggs facilitate cryptic female choice in humans.

Mate choice can continue after mating via chemical communication between the female reproductive system and sperm. While there is a growing appreciation that females can bias sperm use and paternity by exerting cryptic female choice for preferred males, we know surprisingly little about the mechanisms underlying these post-mating choices. In particular, whether chemical signals released from eggs (chemoattractants) allow females to exert cryptic female choice to favour sperm from specific males remains an open question, particularly in species (including humans) where adults exercise pre-mating mate choice. Here, we adapt a classic dichotomous mate choice assay to the microscopic scale to assess gamete-mediated mate choice in humans. We examined how sperm respond to follicular fluid, a source of human sperm chemoattractants, from either their partner or a non-partner female when experiencing a simultaneous or non-simultaneous choice between follicular fluids. We report robust evidence under these two distinct experimental conditions that follicular fluid from different females consistently and differentially attracts sperm from specific males. This chemoattractant-moderated choice of sperm offers eggs an avenue to exercise independent mate preference. Indeed, gamete-mediated mate choice did not reinforce pre-mating human mate choice decisions. Our results demonstrate that chemoattractants facilitate gamete-mediated mate choice in humans, which offers females the opportunity to exert cryptic female choice for sperm from specific males.

Schizophrenia spectrum personality disorders in psychometrically identified schizotypes at two-year follow-up.

Earlier (Bolinskey et al., 2015), we reported that psychometrically identified schizotypes displayed greater symptom levels and higher incidences of schizophrenia spectrum (schizotypal, schizoid, paranoid, and avoidant) personality disorders (PDs). In this study, 49 schizotypes and 39 matched controls participated in follow-up assessments after two years. Participants were previously identified as schizotypes or controls based on scores on the Chapman Psychosis Proneness Scales (CPPS), and were interviewed at baseline and follow-up with the Personality Disorder Interview for DSM-IV (PDI-IV). At follow-up, schizotypes displayed significantly higher symptom levels compared to controls, with medium to large effects, and appeared to meet criteria for diagnosis of each PD more often than controls, although significant differences were only observed for paranoid PD. Overall, schizotypes were more likely to have met criteria for a diagnosis at either baseline or follow-up. Finally, we observed a widening disparity over time between schizotypes and controls in avoidant and schizoid PDs. These results suggest that schizophrenia spectrum PDs, as well as subthreshold symptoms of these disorders, can represent a greater liability for schizophrenia in individuals identified as at-risk on the basis of psychometric means only. Furthermore, these findings demonstrate that such differences persist, and in some cases increase, over time.

Revisiting the blurry boundaries of schizophrenia: spectrum disorders in psychometrically identified schizotypes.

Certain Personality Disorders (PDs) have been found to be present in the prodromal phase of schizophrenia at a higher rate than other personality disorders. Although schizotypal, paranoid, and schizoid PDs are traditionally viewed as spectra for schizophrenia, research suggests that avoidant PD should be included in this group (e.g., Fogelson et al., 2007). The present study examines whether a sample of psychometrically identified schizotypes (SZT) have higher incidence of schizophrenia-spectrum PDs, as well as more symptoms of these PDs, in general, than does a matched comparison (MC) sample. Eighty-five SZT and 78 MC participants were administered the Personality Disorder Interview for DSM-IV (PDI-IV) to assess PD symptoms and diagnoses. Results indicate that the SZT group evidenced significantly more symptoms of avoidant, schizoid, paranoid, and schizotypal PDs than did the MC group. Further, there were significant differences in the incidence of these PDs between the groups.

Using the MMPI-2-RF to discriminate psychometrically identified schizotypic college students from a matched comparison sample.

This study investigates the extent to which the Minnesota Multiphasic Personality Inventory-2 Restructured Form (MMPI-2-RF) profiles of 52 individuals making up a psychometrically identified schizotypes (SZT) sample could be successfully discriminated from the protocols of 52 individuals in a matched comparison (MC) sample. Replication analyses were performed with an additional 53 pairs of SZT and MC participants. Results showed significant differences in mean T-score values between these 2 groups across a variety of MMPI-2-RF scales. Results from discriminant function analyses indicate that schizotypy can be predicted effectively using 4 MMPI-2-RF scales and that this method of classification held up on replication. Additional results demonstrated that these MMPI-2-RF scales nominally outperformed MMPI-2 scales suggested by previous research as being indicative of schizophrenia liability. Directions for future research with the MMPI-2-RF are suggested.

Season of birth, mixed-handedness, and psychometric schizotypy: preliminary results from a prospective study.

Season of birth and hand preference were examined in a sample of 42 (7 males, 35 females) individuals who were identified as schizotypic based on their scores on selected scales of the Chapman Psychosis Proneness Scales (CPPS) and a matched comparison sample of 42 individuals with non-deviant CPPS scores. Presence or absence of schizotypy was analyzed using chi square tests of independence with the presence or absence of each risk factor serving as the independent variable. Further analyses incorporated independent means t tests to examine mean scores on the CPPS with the presence or absence of each risk factor again serving as the independent variable. Results supported the hypothesis that winter/early-spring birth would be associated with psychometric schizotypy, although the results for mixed-handedness fell just short of statistical significance. However, mixed hand preference was associated with higher scores on MagId and PerAb scales of the CPPS, but not the RSA scale, which suggests that mixed laterality is associated with the more cognitive-perceptual aspects of schizotypy. Results are discussed in relation to previous literature and their relevance to the prediction of schizophrenia-related psychosis.

Cryopreserved-thawed embryo transfer in natural or down-regulated hormonally controlled cycles: a retrospective study.

OBJECTIVE: To assess the implantation, pregnancy, and live birth rates after the transfer of frozen-thawed embryos (FET) in a natural or hormonal control cycle. DESIGN: Retrospective study. SETTING: National Health Service tertiary referral center for reproductive medicine in Manchester, United Kingdom. PATIENT(S): Two comparable groups of women with regular menstrual cycles: Group A (n = 212) had FET in a natural cycle after spontaneous ovulation; group B (n = 205) had FET in a pituitary-desensitized hormonally controlled cycle. INTERVENTION(S): In group B, GnRH agonist was commenced in the midluteal phase of the previous cycle and discontinued 3 days before embryo transfer. Oral estradiol valerate and vaginal progesterone pessary were used to prepare the endometrium. Embryo transfer was carried out 3 days after detection of the endogenous LH surge in group A and on day 3 of progesterone administration in group B. MAIN OUTCOME MEASURE(S): Implantation, pregnancy, and live birth rates per cycle and per embryo transfer (ET). RESULT(S): In the 212 women who had natural-cycle FET, 172 ETs were performed and 247 embryos replaced. The implantation rate was 14.1% (35/247). Twenty clinical pregnancies (20/172, 11.6%) were achieved. In the 205 women who had down-regulated hormone replacement-cycle FET, 173 embryo transfers were performed and 243 embryos replaced. The implantation rate was 13.5% (33/243). Eighteen clinical pregnancies (18/173, 10.2%) were achieved. There were no significant differences between the two groups with regard to the implantation, clinical pregnancy, or live birth rates per cycle or per ET. CONCLUSION(S): These findings suggest that both FET protocols are equally effective in terms of implantation rate and pregnancy outcome in women with regular menstrual cycles.

Case study: managing and caring for a patient undergoing stoma formation.

A 75-year-old man had attended his GP, as he was worried that his altered bowel habits and slight rectal bleeding was more sinister than haemorrhoids. This article describes the subsequent investigations, care and treatment that he received. It highlights how early detection, prompt treatment, appropriate care and management can enable a person to cope with the devastating diagnosis of bowel cancer, surgery and the need for a stoma. This patient was otherwise well which contributed both to the treatment decision and successful outcome. Unfortunately, not all patients have such an experience, and the measures taken to prevent complications are described.

Expression of 11 members of the BCL-2 family of apoptosis regulatory molecules during human preimplantation embryo development and fragmentation.

Apoptosis during preimplantation development has received much interest because of its potential role in eliminating defective cells. Although development in humans is characterised by a high degree of genetic abnormality, little is known of the regulation of apoptosis in embryos. By PolyA PCR we analysed expression of 11 BCL-2 genes in individual human embryos representative of normal development and in severely fragmented embryos. We demonstrate constitutive expression of BAX in virtually all embryos at all stages of development, and variable expression of BCL2, BCL-XL, BCL-W, MCL-1 BAK, BAD, BOKL, BID, BIK, and BCL-XS. The frequency of expression of pro- and anti-apoptotic BCL-2 members was similar throughout development, except at the two-cell stage where pro-apoptotic genes predominated. Protein expression was confirmed for BCL-2, MCL-1, BCL-X, BAX, BAD, and activated caspase 3. BCL-2 protein was associated with mitochondria but expressed inconsistently in the blastocyst inner cell mass. Consistent differences between morphologically intact and fragmented embryos included the expression of BAK in fragmented but not intact four-cell embryos. Our study addresses the importance of examining single human embryos representative of the viable population for a large number of genes, in order to establish meaningful expression profiles and provide information on overlapping function in a large gene family.