Neuropsychological and behavioural outcomes in epilepsy involving the insula: A scoping review.

Objective: In patients with epilepsy, the insula has been increasingly recognized as a common site of seizures. There is growing interest in understanding the cognitive and psychological consequences of insular epilepsy to help provide clinical recommendations to support patient's cognitive and psychosocial functioning, and to help identify candidates for epilepsy resective surgery. The aim of this scoping review was to describe the cognitive and behavioural characteristics associated with insular epilepsy in children and adults. Method: A systematic search was completed according to the Preferred Reporting Items for Systematic Reviews and Meta-analysis -Extension for Scoping Reviews guidelines. Eligible studies reported on a neuropsychological or behavioural outcome, using standardized or research-based psychological measures, in individuals with insular epilepsy, (i.e. the seizure focus and/or surgical resection included the insula), and a comparison group. After duplicates were removed, 2,423 citations were identified from the search, and 39 studies were included in the scoping review. Results: Across the included studies, intellectual/global cognitive functioning and language were most often evaluated. Lower functioning was found across multiple cognitive and behavioural processes in pediatric and adult patients with insular epilepsy. Following resective surgery involving the insula, behavioural and cognitive outcomes are general stable. Conclusions: The results of this scoping review further neuropsychologists' knowledge of the cognitive and behavioural outcomes of insular seizures prior to and following surgical treatment. These results can aid in counselling patients of the potential cognitive dysfunctions, and aid with treatment planning.

Impact of Discontinuing Both Hypertonic Saline and Dornase Alfa After Elexacaftor/Tezacaftor/Ivacaftor in Cystic Fibrosis.

Rationale Evaluating approaches to reduce treatment burden is a research priority among people with CF (pwCF) on highly effective modulators including elexacaftor/tezacaftor/ivacaftor (ETI). Objective To evaluate the impact of discontinuing both hypertonic saline (HS) and dornase alfa (DA) versus continuing both therapies among a subgroup of participants in the SIMPLIFY study who sequentially participated in trials evaluating the independent clinical effects of discontinuing HS and DA. Methods SIMPLIFY participants ≥12 years old on ETI and comprising a subgroup using both HS and DA at study entry were randomized to the HS or DA trial, and then randomized 1:1 to continue or discontinue the applicable therapy for 6 weeks. After completion of the first trial, eligible participants could enroll in the second trial beginning with a 2-week run in. Study outcomes were compared across the duration of SIMPLIFY participation between a cohort remaining on both therapies during SIMPLIFY versus a cohort that sequentially discontinued both as a result of trial randomizations. Multivariable regression models were used to estimate treatment differences, adjusted for time between trials, trial order, baseline age, sex at birth and percent predicted forced expiratory volume in one second (ppFEV1) at study entry. Results There were 43 participants who discontinued both therapies by the end of SIMPLIFY and 63 who remained on both, with overall average ppFEV1 at study entry 96.7% and average duration of follow up from beginning of the first trial to completion of the second trial 3.9 months, including time between trials. No clinically meaningful difference in the change in ppFEV1 from baseline to completion of the second trial was observed between those who discontinued versus remained on both therapies (difference: 0.22% Off-On, 95% CI: -1.60,2.03). Changes in LCI2.5, patient reported, and safety outcomes were also comparable. Patient reported treatment burden, as measured by a CFQ-R subscale, significantly decreased in those discontinuing both therapies. Conclusions SIMPLIFY participants who sequentially discontinued both HS and DA experienced no meaningful changes in clinical outcomes and reported decreased treatment burden as compared to those who remained on both therapies. These data continue to inform a new era of post-modulator care of pwCF.

Attention and neurodevelopment in young children who are HIV-exposed uninfected.

ABSTRACTChildren HIV-exposed, uninfected (CHEU) are at risk for compromised developmental outcomes. Attention is important for behavioural, cognitive and academic skills, yet has not been thoroughly investigated compared to children HIV-unexposed uninfected (CHUU). Fifty-five CHEU and 51 CHUU children were recruited at 5.5 years of age. Measures of inattention (IA), hyperactivity/impulsivity (HI) and total scores were collected using the parent-reported ADHD-Rating-Scale-IV. Measures of intelligence, visuomotor skills, academics and adaptive functioning were obtained. Analyses of between-group differences were performed as were correlational and multiple regression models, accounting for maternal education, employment and delivery type. Few children met clinical cut-offs for probable ADHD (3.6% CHEU, 2.0% CHUU), and no group differences in measures of IA, HI and combined scores were found. CHEU scored significantly lower than CHUU on intelligence, visuomotor function, academic skills and aspects of adaptive behaviour, though within age expectations. Lower Full-Scale IQ and Processing Speed were associated with higher IA in CHEU and lower adaptive functioning with higher IA in CHUU. Across both groups, children of unemployed mothers had more HI symptoms. CHEU were not at increased risk for attention difficulties at 5.5 years of age. Maternal employment status highlights the contribution of sociodemographic factors in shaping behaviour and neurodevelopment.

Postoperative cerebellar mutism syndrome is an acquired Autism-like network disturbance.

BACKGROUND: Cerebellar mutism syndrome (CMS) is a common and debilitating complication of posterior fossa tumour surgery in children. Affected children exhibit communication and social impairments that overlap phenomenologically with subsets of deficits exhibited by children with Autism spectrum disorder (ASD). Although both CMS and ASD are thought to involve disrupted cerebro-cerebellar circuitry, they are considered independent conditions due to an incomplete understanding of their shared neural substrates. METHODS: In this study, we analyzed post-operative cerebellar lesions from 90 children undergoing posterior fossa resection of medulloblastoma, 30 of whom developed CMS. Lesion locations were mapped to a standard atlas, and the networks functionally connected to each lesion were computed in normative adult and paediatric datasets. Generalizability to ASD was assessed using an independent cohort of children with ASD and matched controls (n=427). RESULTS: Lesions in children who developed CMS involved the vermis and inferomedial cerebellar lobules. They engaged large-scale cerebellothalamocortical circuits with a preponderance for the prefrontal and parietal cortices in the paediatric and adult connectomes, respectively. Moreover, with increasing connectomic age, CMS-associated lesions demonstrated stronger connectivity to the midbrain/red nuclei, thalami and inferior parietal lobules and weaker connectivity to prefrontal cortex. Importantly, the CMS-associated lesion network was independently reproduced in ASD and correlated with communication and social deficits, but not repetitive behaviours. CONCLUSIONS: Our findings indicate that CMS-associated lesions result in an ASD-like network disturbance that occurs during sensitive windows of brain development. A common network disturbance between CMS and ASD may inform improved treatment strategies for affected children.

Labs Do Not Predict Postoperative Intra-abdominal Abscess in Pediatric Perforated Appendicitis.

INTRODUCTION: We aim to evaluate the utility of postoperative labs in predicting the development of an intra-abdominal abscess (IAA) in pediatric patients with perforated appendicitis. We hypothesize that postoperative labs are not predictive of IAA development. METHODS: This was a single-institution retrospective cohort study that included pediatric patients (n = 61) who underwent surgery for perforated appendicitis from January 1, 2019 to December 1, 2020. Patients were stratified into those who developed a postoperative IAA (n = 22) and those who did not (n = 39). Postoperative labs (white blood cell [WBC] count, absolute neutrophil count, platelet count, C-reactive protein) were examined. Mann-Whitney U tests and chi-square tests were used to assess for differences between groups. RESULTS: There was extensive heterogeneity and overlap in postoperative lab values between patients who developed an IAA and those who did not. Almost all patients who developed an IAA had clinical signs that were indicative of abscess formation regardless of their postoperative WBC count or change in WBC count. While patients who developed an IAA had a higher postoperative median WBC count (10.8 versus 8.4, P = 0.003) and a smaller WBC count decrease (-4.9 versus -7.4, P = 0.01), no cutoff value for any of the examined lab values specifically predicted abscess formation. Postoperative median absolute neutrophil count (7.4 versus 4.0, P = 0.15), platelet count (360 versus 353, P = 0.98), and C-reactive protein (8.20 versus 5.32, P = 0.06) did not differ significantly. CONCLUSIONS: We conclude that postoperative labs have limited clinical utility in evaluating IAA development in children with perforated appendicitis.

Discontinuation versus continuation of hypertonic saline or dornase alfa in modulator treated people with cystic fibrosis (SIMPLIFY): results from two parallel, multicentre, open-label, randomised, controlled, non-inferiority trials.

BACKGROUND: Reducing treatment burden is a priority for people with cystic fibrosis, whose health has benefited from using new modulators that substantially increase CFTR protein function. The SIMPLIFY study aimed to assess the effects of discontinuing nebulised hypertonic saline or dornase alfa in individuals using the CFTR modulator elexacaftor plus tezacaftor plus ivacaftor (ETI). METHODS: The SIMPLIFY study included two parallel, multicentre, open-label, randomised, controlled, non-inferiority trials at 80 participating clinics across the USA in the Cystic Fibrosis Therapeutics Development Network. We included individuals with cystic fibrosis aged 12-17 years with percent predicted FEV1 (ppFEV1) of 70% or more, or those aged 18 years or older with ppFEV1 of 60% or more, if they had been taking ETI and either (or both) mucoactive therapies (≥3% hypertonic saline or dornase alfa) for at least 90 days before screening. Participants on both hypertonic saline and dornase alfa were randomly assigned to one of the two trials, and those on a single therapy were assigned to the applicable trial. All participants were then randomly assigned 1:1 to continue or discontinue therapy for 6 weeks using permuted blocks of varying size, stratified by baseline ppFEV1 (week 0; ≥90% or <90%), single or concurrent use of hypertonic saline and dornase alfa, previous SIMPLIFY study participation (yes or no), and age (≥18 or <18 years). For participants randomly assigned to continue their therapy during a given trial, this therapy was instructed to be taken at least once daily according to each participant's pre-existing, clinically prescribed regimen. Hypertonic saline concentration was required to be at least 3%. The primary objective for each trial was to determine whether discontinuing was non-inferior to continuing, measured by the 6-week change in ppFEV1 in the per-protocol population. We established a non-inferiority margin of -3% for the difference between groups in the 6-week change in ppFEV1. Safety outcomes were analysed in the intention-to-treat population. This study is registered with ClinicalTrials.gov, NCT04378153. FINDINGS: From Aug 25, 2020, to May 25, 2022, a total of 672 unique participants were screened for eligibility for one or both trials, resulting in 847 total random assignments across both trials with 594 unique participants. 370 participants were randomly assigned in the hypertonic saline trial and 477 in the dornase alfa trial. Participants across both trials had an average ppFEV1 of 96·9%. Discontinuing treatment was non-inferior to continuing treatment with respect to the absolute 6-week change in ppFEV1 in both the hypertonic saline trial (-0·19% [95% CI -0·85 to 0·48] in the discontinuation group [n=133] vs 0·14% [-0·51 to 0·78] in the continuation group [n=140]; between-group difference -0·32% [-1·25 to 0·60]) and dornase alfa trial (0·18% [-0·38 to 0·74] in the discontinuation group [n=199] vs -0·16% [-0·73 to 0·41] in the continuation group [n=193]; between-group difference 0·35% [-0·45 to 1·14]), with consistent results in the intention-to-treat populations. In the hypertonic saline trial, 64 (35%) of 184 in the discontinuation group versus 44 (24%) of 186 participants in the continuation group and, in the dornase alfa trial, 89 (37%) of 240 in the discontinuation group versus 55 (23%) of 237 in the continuation group had at least one adverse event. INTERPRETATION: In individuals with cystic fibrosis on ETI with relatively well preserved pulmonary function, discontinuing daily hypertonic saline or dornase alfa for 6 weeks did not result in clinically meaningful differences in pulmonary function when compared with continuing treatment.

Connectomic Profiles and Cognitive Trajectories After Epilepsy Surgery in Children.

BACKGROUND AND OBJECTIVES: Neurocognitive outcomes after surgery for temporal lobe epilepsy in childhood are variable. Postoperative changes are not directly predicted by seizure freedom, and associations between epilepsy, neuropsychological function, and developing neural networks are poorly understood. Here, we leveraged whole-brain connectomic profiling in magnetoencephalography (MEG) to retrospectively study associations between brain connectivity and neuropsychological function in children with temporal lobe epilepsy undergoing resective surgery. METHODS: Clinical and MEG data were retrospectively analyzed for children who underwent temporal lobe epilepsy surgery at the Hospital for Sick Children from 2000 to 2021. Resting-state connectomes were constructed from neuromagnetic oscillations via the weighted-phase lag index. Using a partial least-squares (PLS) approach, we assessed multidimensional associations between patient connectomes, neuropsychological scores, and clinical covariates. Bootstrap resampling statistics were performed to assess statistical significance. RESULTS: A total of 133 medical records were reviewed, and 5 PLS analyses were performed. Each PLS analysis probed a particular neuropsychological domain and the associations between its baseline and postoperative scores and the connectomic data. In each PLS analysis, a significant latent variable was identified, representing a specific percentage of the variance in the data and relating neural networks to clinical covariates, which included changes in rote verbal memory (n = 41, p = 0.01, σ2 = 0.38), narrative/verbal memory (n = 57, p = 0.00, σ2 = 0.52), visual memory (n = 51, p = 0.00, σ2 = 0.43), working memory (n = 44, p = 0.00, σ2 = 0.52), and overall intellectual function (n = 59, p = 0.00, σ2 = 0.55). Children with more diffuse, bilateral intrinsic connectivity across several frequency bands showed lower scores on all neuropsychological assessments but demonstrated a greater propensity for gains after resective surgery. DISCUSSION: Here, we report that connectomes characterized by diffuse connectivity, reminiscent of developmentally immature networks, are associated with lower preoperative cognition and postoperative cognitive improvement. These findings provide a potential means to understand neurocognitive function in children with temporal lobe epilepsy and expected changes postoperatively.

WNT signalling in the normal human adult testis and in male germ cell neoplasms.

STUDY QUESTION: Is WNT signalling functional in normal and/or neoplastic human male germ cells? SUMMARY ANSWER: Regulated WNT signalling component synthesis in human testes indicates that WNT pathway function changes during normal spermatogenesis and is active in testicular germ cell tumours (TGCTs), and that WNT pathway blockade may restrict seminoma growth and migration. WHAT IS KNOWN ALREADY: Regulated WNT signalling governs many developmental processes, including those affecting male fertility during early germ cell development at embryonic and adult (spermatogonial) ages in mice. In addition, although many cancers arise from WNT signalling alterations, the functional relevance and WNT pathway components in TGCT, including germ cell neoplasia in situ (GCNIS), are unknown. STUDY DESIGN, SIZE, DURATION: The cellular distribution of transcripts and proteins in WNT signalling pathways was assessed in fixed human testis sections with normal spermatogenesis, GCNIS and seminoma (2-16 individuals per condition). Short-term (1-7 h) ligand activation and long-term (1-5 days) functional outcomes were examined using the well-characterised seminoma cell line, TCam-2. Pathway inhibition used siRNA or chemical exposures over 5 days to assess survival and migration. PARTICIPANTS/MATERIALS, SETTING, METHODS: The cellular localisation of WNT signalling components was determined using in situ hybridisation and immunohistochemistry on Bouin's- and formalin-fixed human testis sections with complete spermatogenesis or germ cell neoplasia, and was also assessed in TCam-2 cells. Pathway function tests included exposure of TCam-2 cells to ligands, small molecules and siRNAs. Outcomes were measured by monitoring beta-catenin (CTNNB1) intracellular localisation, cell counting and gap closure measurements. MAIN RESULTS AND THE ROLE OF CHANCE: Detection of nuclear-localised beta-catenin (CTNNB1), and key WNT signalling components (including WNT3A, AXIN2, TCF7L1 and TCF7L2) indicate dynamic and cell-specific pathway activity in the adult human testis. Their presence in germ cell neoplasia and functional analyses in TCam-2 cells indicate roles for active canonical WNT signalling in TGCT relating to viability and migration. All data were analysed to determine statistical significance. LARGE SCALE DATA: No large-scale datasets were generated in this study. LIMITATIONS, REASONS FOR CAUTION: As TGCTs are rare and morphologically heterogeneous, functional studies in primary cancer cells were not performed. Functional analysis was performed with the only well-characterised, widely accepted seminoma-derived cell line. WIDER IMPLICATIONS OF THE FINDINGS: This study demonstrated the potential sites and involvement of the WNT pathway in human spermatogenesis, revealing similarities with murine testis that suggest the potential for functional conservation during normal spermatogenesis. Evidence that inhibition of canonical WNT signalling leads to loss of viability and migratory activity in seminoma cells suggests that potential treatments using small molecule or siRNA inhibitors may be suitable for patients with metastatic TGCTs. STUDY FUNDING AND COMPETING INTEREST(S): This study was funded by National Health and Medical Research Council of Australia (Project ID 1011340 to K.L.L. and H.E.A., and Fellowship ID 1079646 to K.L.L.) and supported by the Victorian Government's Operational Infrastructure Support Program. None of the authors have any competing interests.

Neuropsychological outcomes following pediatric lung transplantation.

OBJECTIVE: Lung transplantation (LTx) is a treatment option for eligible children with end-stage pulmonary diseases. Improving our understanding of longer-term developmental outcomes in pediatric LTx recipients is important for strategized interventions targeting cognitive difficulties. METHODS: Neuropsychological assessments were completed for children who received LTx at our center (2009-2017). Assessments comprised tasks of general intellect, memory, visual-perception, academics, and executive functioning as well as caregiver questionnaires of adaptive, executive, emotional, and behavioral functioning. Results were compared to age-matched population norms. Between-group nonparametric tests were performed pre-LTx vs post-LTx and for children with a primary diagnosis of cystic fibrosis (CF) vs other diagnoses (non-CF). RESULTS: Neuropsychological outcomes were assessed for 21 children post-LTx, with a median age (interquartile range) at the time of transplant of 11.52 (6.89, 14.12) years. Eleven children completed pre- and post-transplant assessments and within this group, improvements for verbal learning (P = .02), aspects of mood, behavior, and adaptive functioning were observed over time (all P < .05). Post-transplant whole group analysis suggested age-appropriate abilities across most cognitive domains, with a relative weakness for executive functioning. Emotional or behavioral difficulties were not endorsed by caregivers. Across pulmonary diagnoses, higher levels of emotional, behavioral, and executive functioning difficulty were reported in the non-CF group (all P < .05). CONCLUSIONS: Overall, LTx has a positive impact on cognitive functioning, particularly learning, adaptive functioning, mood, and behavior. Children transplanted for non-CF related diseases demonstrated greater challenges, highlighting the need for targeted assessments and interventions across the transplant process to support the complex needs of this population.

Evidence that activin A directly modulates early human male germline differentiation status.

Disrupted fetal germline development underpins testicular germ cell neoplasia, which is increasing worldwide. The complex signaling milieu during normal testis development includes TGFß superfamily ligands; this study tests the hypothesis that, activin A, a TGFß superfamily member, can influence gonocyte development. The human seminoma-derived cell line, TCam-2, a model of fetal gonocytes, was cultured with activin A (1.25-25 ng/mL) for 48 h, or with 5 ng/mL activin A for short- (6, 24, and 48 h) and long-term (13 days) exposures, and downstream targets measured by qRT-PCR. Transcripts that exhibited significant dose-dependent responses to activin A included the early germ cell markers KIT, NODAL, and CRIPTO (NODALl co-receptor and activin inhibitor) which all increased and the differentiation marker DNMT3L which decreased. After 48 h, KIT, NODAL, and CRIPTO levels were significantly higher, while the differentiation marker NANOS2 was significantly lower. Interestingly, activin A exposure also significantly reduced both transcript and protein levels of the PIWI/piRNA pathway component DNMT3L. Because TCam-2 cells produce the activin inhibitor CRIPTO, CRIPTO was reduced using siRNA prior to activin A exposure. This selectively increased KIT in response to activin A. Other ligands present in the fetal testis (BMP4, FGF9, TGFß1, and TGFß2) induced distinct effects on germline marker expression. This study showed that activin A can directly modulate germline markers in this human gonocyte-like cell, promoting a less-differentiated phenotype. Additional findings indicate evidence of signaling crosstalk between activin A and NODAL, leading to target-specific effects on gonocyte differentiation.

'Snail factors in testicular germ cell tumours and their regulation by the BMP4 signalling pathway'.

BACKGROUND: Snail transcription factors mediate key cellular transitions in many developmental processes, including spermatogenesis, and their production can be regulated by TGF-ß superfamily signalling. SNAI1 and SNAI2 support many cancers of epithelial origin. Their functional relevance and potential regulation by TGF-ß superfamily ligands in germ cell neoplasia are unknown. METHODS: SNAI1, SNAI2 and importin 5 (IPO5; nuclear transporter that selectively mediates BMP signalling) cellular localization was examined in fixed normal adult human and/or neoplastic testes using in situ hybridization and/or immunohistochemistry. SNAI1 and SNAI2 functions were assessed using the well-characterized human seminoma cell line, TCam-2. Cell migration, adhesion/proliferation and survival were measured by scratch assay, xCELLigence and flow cytometry following siRNA-induced reduction of SNAI1 and SNAI2 in TCam-2 cells. The potential regulation of SNAI1 and SNAI2 in TCam-2 cells by TGF-ß signalling ligands, activin A and BMP4 was evaluated following 48 hours culture, including with siRNA regulation of IPO5 to selectively restrict BMP4 signalling. RESULTS: In normal testes, SNAI1 transcript was identified in some spermatogonia and in spermatocytes, and SNAI2 protein localized to nuclei of spermatogonia, spermatocytes and round spermatids. In neoplastic testes, both SNAI1 and SNAI2 were detected in GCNIS and in seminoma cells. SNAI1 and SNAI2 reduction in TCam-2 cells by siRNAs significantly inhibited migration and survival, respectively. Exposure to BMP4, but not activin A, significantly increased SNAI2 (~18-fold). IPO5 inhibition by siRNAs decreased BMP4-induced SNAI2 upregulation (~5-fold). Additionally, SNAI2 reduction using siRNAs inhibited BMP4-induced TCam-2 cell survival. CONCLUSIONS: This is the first evidence that SNAI1 and SNAI2 are involved in human spermatogenesis, with independent functions. These outcomes demonstrate that SNAI1 and SNAI2 inhibition leads to loss of migratory and viability capacities in seminoma cells. These findings show the potential for therapeutic treatments targeting SNAIL or BMP4 signalling for patients with metastatic testicular germ cell tumours.

Dynamic paraspeckle component localisation during spermatogenesis.

Expression profiles and subcellular localisations of core Drosophila behaviour/human splicing (DBHS) proteins (PSPC1, SFPQ and NONO) and NEAT1, a long noncoding RNA (lncRNA), are investigated in developing and adult mouse testes. Core DBHS proteins are markers for the distinct subnuclear domain termed paraspeckles, while a long NEAT1 isoform scaffold facilitates paraspeckle nucleation. Paraspeckles contain many proteins (>40) and are broadly involved in RNA metabolism, including transcriptional regulation by protein sequestration, nuclear retention of A-to-I edited RNA transcripts to regulate translation and promoting survival during cellular stress. Immunohistochemistry reveals cell-specific profiles for core DBHS paraspeckle protein expression, indicating their functional diversity. PSPC1 is an androgen receptor co-activator, and it is detected in differentiating Sertoli cell nuclei from day 15 onwards, as they develop androgen responsiveness. PSPC1 is nuclear in the most mature male germ cell type present at each age, from foetal to adult life. In adult mouse testes, PSPC1 and SFPQ are present in Sertoli cells, spermatocytes and round spermatids, while the NEAT1 lncRNA appears in the punctate nuclear foci delineating paraspeckles only within Leydig cells. Identification of NEAT1 in the cytoplasm of spermatogonia and spermatocytes must reflect non-paraspeckle-related functions. NONO was absent from germ cells but nuclear in Sertoli cells. Reciprocal nuclear profiles of PSPC1 and γ-H2AX in spermatogenic cells suggest that each performs developmentally regulated roles in stress responses. These findings demonstrate paraspeckles and paraspeckle-related proteins contribute to diverse functions during testis development and spermatogenesis.

Continual reassessment method for dose escalation clinical trials in oncology: a comparison of prior skeleton approaches using AZD3514 data.

BACKGROUND: The continual reassessment method (CRM) requires an underlying model of the dose-toxicity relationship ("prior skeleton") and there is limited guidance of what this should be when little is known about this association. In this manuscript the impact of applying the CRM with different prior skeleton approaches and the 3 + 3 method are compared in terms of ability to determine the true maximum tolerated dose (MTD) and number of patients allocated to sub-optimal and toxic doses. METHODS: Post-hoc dose-escalation analyses on real-life clinical trial data on an early oncology compound (AZD3514), using the 3 + 3 method and CRM using six different prior skeleton approaches. RESULTS: All methods correctly identified the true MTD. The 3 + 3 method allocated six patients to both sub-optimal and toxic doses. All CRM approaches allocated four patients to sub-optimal doses. No patients were allocated to toxic doses from sigmoidal, two from conservative and five from other approaches. CONCLUSIONS: Prior skeletons for the CRM for phase 1 clinical trials are proposed in this manuscript and applied to a real clinical trial dataset. Highly accurate initial skeleton estimates may not be essential to determine the true MTD, and, as expected, all CRM methods out-performed the 3 + 3 method. There were differences in performance between skeletons. The choice of skeleton should depend on whether minimizing the number of patients allocated to suboptimal or toxic doses is more important. TRIAL REGISTRATION: NCT01162395 , Trial date of first registration: July 13, 2010.

TGF-ß superfamily signaling in testis formation and early male germline development.

The TGF-ß ligand superfamily contains at least 40 members, many of which are produced and act within the mammalian testis to facilitate formation of sperm. Their progressive expression at key stages and in specific cell types determines the fertility of adult males, influencing testis development and controlling germline differentiation. BMPs are essential for the interactive instructions between multiple cell types in the early embryo that drive initial specification of gamete precursors. In the nascent foetal testis, several ligands including Nodal, TGF-ßs, Activins and BMPs, serve as key masculinizing switches by regulating male germline pluripotency, somatic and germline proliferation, and testicular vascularization and architecture. In postnatal life, local production of these factors determine adult testis size by regulating Sertoli cell multiplication and differentiation, in addition to specifying germline differentiation and multiplication. Because TGF-ß superfamily signaling is integral to testis formation, it affects processes that underlie testicular pathologies, including testicular cancer, and its potential to contribute to subfertility is beginning to be understood.

Regulated Wnt/beta-catenin signaling sustains adult spermatogenesis in mice.

The importance of Wnt signaling for postnatal testis function has been previously studied in several mouse models, with chronic pathway disruption addressing its function in Sertoli cells and in postmeiotic germ cells. While chronic beta-catenin deletion in Sertoli cells does not profoundly affect testis development, new data indicate that Wnt signaling is required at multiple stages of spermatogenesis. We used two mouse models that allow acute disruption of Wnt signaling to explore the importance of regulated Wnt pathway activity for normal germ cell development in adult male mice. Short-term induction of mutations in Adenomatous polyposis coli (Apc) and beta-catenin (Ctnnbl), which increase and decrease Wnt signaling levels, were generated in AhCre Apc(fl/fl) and AhCre Ctnnb1(fl/fl) mice, respectively. Each exhibited a distinct phenotype of disrupted spermatogenesis that was evident within 24 h and persisted for up to 4 days. Outcomes included germ cell apoptosis and rapid loss and altered blood-testis barrier protein distribution and morphology. The functional significance of nuclear localized beta-catenin protein in spermatocytes and round spermatids, indicative of active Wnt signaling, was highlighted by the profound loss of postmitotic germ cells in both models. Developmentally regulated Wnt signaling mediators identified through transcriptional profiling of wild-type and AhCre Ctnnb1(fl/fl) mouse testes identified Wnt receptors (e.g., Fzd4) and ligands (e.g., Wnt3, Wnt3a, Wnt5b, Wnt7a, and Wnt8b). This demonstration that Wnt signaling control is essential for adult spermatogenesis supports the growing understanding that its disruption may underpin certain cases of male infertility.

The nuclear import factor importin α4 can protect against oxidative stress.

The importin (IMP) superfamily of nuclear transport proteins is essential to key developmental pathways, including in the murine testis where expression of the 6 distinct IMPα proteins is highly dynamic. Present predominantly from the spermatocyte stage onwards, IMPα4 is unique in showing a striking nuclear localization, a property we previously found to be linked to maintenance of pluripotency in embryonic stem cells and to the cellular stress response in cultured cells. Here we examine the role of IMPα4 in vivo for the first time using a novel transgenic mouse model in which we overexpress an IMPα4-EGFP fusion protein from the protamine 1 promoter to recapitulate endogenous testicular germ cell IMPα4 expression in spermatids. IMPα4 overexpression did not affect overall fertility, testis morphology/weight or spermatogenic progression under normal conditions, but conferred significantly (>30%) increased resistance to oxidative stress specifically in the spermatid subpopulation expressing the transgene. Consistent with a cell-specific role for IMPα4 in protecting against oxidative stress, haploid germ cells from IMPα4 null mice were significantly (c. 30%) less resistant to oxidative stress than wild type controls. These results from two unique and complementary mouse models demonstrate a novel protective role for IMPα4 in stress responses specifically within haploid male germline cells, with implications for male fertility and genetic integrity.

Effects of fish oil supplementation on inflammatory acne.

BACKGROUND: Given that acne is a rare condition in societies with higher consumption of omega-3 (n-3) relative to omega-6 (n-6) fatty acids, supplementation with n-3 may suppress inflammatory cytokine production and thereby reduce acne severity. METHODS: 13 individuals with inflammatory acne were given three grams of fish oil containing 930 mg of EPA to their unchanged diet and existing acne remedies for 12 weeks. Acne was assessed using an overall severity grading scale, total inflammatory lesion counts, and colorimetry. FINDINGS: There was no significant change in acne grading and inflammatory counts at week 12 compared to baseline. However, there was a broad range of response to the intervention on an individual basis. The results showed that acne severity improved in 8 individuals, worsened in 4, and remained unchanged in 1. Interestingly, among the individuals who showed improvement, 7 were classified as having moderate to severe acne at baseline, while 3 of the 4 whose acne deteriorated were classified as having mild acne. CONCLUSION: There is some evidence that fish oil supplementation is associated with an improvement in overall acne severity, especially for individuals with moderate to severe acne. Divergent responses to fish oil in our pilot study indicates that dietary and supplemental lipids are worthy of further investigation in acne.

Enhanced thecal androgen production is prenatally programmed in an ovine model of polycystic ovary syndrome.

One of the hallmarks of polycystic ovary syndrome (PCOS) is increased ovarian androgen secretion that contributes to the ovarian, hormonal, and metabolic features of this condition. Thecal cells from women with PCOS have an enhanced capacity for androgen synthesis. To investigate whether this propensity is a potential cause, rather than a consequence, of PCOS, we used an ovine prenatal androgenization model of PCOS and assessed ewes at 11 months of age. Pregnant Scottish Greyface ewes were administered 100 mg testosterone propionate (TP) or vehicle control twice weekly from d 62 to 102 of gestation, and female offspring (TP = 9, control = 5) were studied. Prenatal TP exposure did not alter ovarian morphology or cyclicity, or plasma androgen, estrogen, and gonadotropin concentrations, at this stage. However, follicle function was reprogrammed in vivo with increased proportions of estrogenic follicles (P < 0.05) in the TP-exposed cohort. Furthermore, in vitro the thecal cells of follicles (>4 mm) secreted more LH-stimulated androstenedione after prenatal androgenization (P < 0.05), associated with increased basal expression of thecal StAR (P < 0.01), CYP11A (P < 0.05), HSD3B1 (P < 0.01), CYP17 (P < 0.05), and LHR (P < 0.05). This provides the first evidence of increased thecal androgenic capacity in the absence of a PCOS phenotype, suggesting a thecal defect induced during fetal life.