Examining Neurosteroid-Analogue Therapy in the Preterm Neonate For Promoting Hippocampal Neurodevelopment.

Background: Preterm birth can lead to brain injury and currently there are no targeted therapies to promote postnatal brain development and protect these vulnerable neonates. We have previously shown that the neurosteroid-analogue ganaxolone promotes white matter development and improves behavioural outcomes in male juvenile guinea pigs born preterm. Adverse side effects in this previous study necessitated this current follow-up dosing study, where a focus was placed upon physical wellbeing during the treatment administration and markers of neurodevelopment at the completion of the treatment period. Methods: Time-mated guinea pigs delivered preterm (d62) by induction of labour or spontaneously at term (d69). Preterm pups were randomized to receive no treatment (Prem-CON) or ganaxolone at one of three doses [0.5 mg/kg ganaxolone (low dose; LOW-GNX), 1.0 mg/kg ganaxolone (mid dose; MID-GNX), or 2.5 mg/kg ganaxolone (high dose; HIGH-GNX) in vehicle (45% ß-cyclodextrin)] daily until term equivalence age. Physical parameters including weight gain, ponderal index, supplemental feeding, and wellbeing (a score based on respiration, activity, and posture) were recorded throughout the preterm period. At term equivalence, brain tissue was collected, and analysis of hippocampal neurodevelopment was undertaken by immunohistochemistry and RT-PCR. Results: Low and mid dose ganaxolone had some impacts on early weight gain, supplemental feeding, and wellbeing, whereas high dose ganaxolone significantly affected all physical parameters for multiple days during the postnatal period when compared to the preterm control neonates. Deficits in the preterm hippocampus were identified using neurodevelopmental markers including mRNA expression of oligodendrocyte lineage cells (CSPG4, MBP), neuronal growth (INA, VEGFA), and the GABAergic/glutamatergic system (SLC32A1, SLC1A2, GRIN1, GRIN2C, DLG4). These deficits were not affected by ganaxolone at the doses used at the equivalent of normal term. Conclusion: This is the first study to investigate the effects of a range of doses of ganaxolone to improve preterm brain development. We found that of the three doses, only the highest dose of ganaxolone (2.5 mg/kg) impaired key indicators of physical health and wellbeing over extended periods of time. Whilst it may be too early to see improvements in markers of neurodevelopment, further long-term study utilising the lower doses are warranted to assess functional outcomes at ages when preterm birth associated behavioural disorders are observed.

MITO-FIND: A study in 390 patients to determine a diagnostic strategy for mitochondrial disease.

Mitochondrial diseases, due to nuclear or mitochondrial genome mutations causing mitochondrial dysfunction, have a wide range of clinical features involving neurologic, muscular, cardiac, hepatic, visual, and auditory symptoms. Making a diagnosis of a mitochondrial disease is often challenging since there is no gold standard and traditional testing methods have required tissue biopsy which presents technical challenges and most patients prefer a non-invasive approach. Since a diagnosis invariably involves finding a disease-causing DNA variant, new approaches such as next generation sequencing (NGS) have the potential to make it easier to make a diagnosis. We evaluated the ability of our traditional diagnostic pathway (metabolite analysis, tissue neuropathology and respiratory chain enzyme activity) in 390 patients. The traditional diagnostic pathway provided a diagnosis of mitochondrial disease in 115 patients (29.50%). Analysis of mtDNA, tissue neuropathology, skin electron microscopy, respiratory chain enzyme analysis using inhibitor assays, blue native polyacrylamide gel electrophoresis were all statistically significant in distinguishing patients between a mitochondrial and non-mitochondrial diagnosis. From these 390 patients who underwent traditional analysis, we recruited 116 patients for the NGS part of the study (36 patients who had a mitochondrial diagnosis (MITO) and 80 patients who had no diagnosis (No-Dx)). In the group of 36 MITO patients, nuclear whole exome sequencing (nWES) provided a second diagnosis in 2 cases who already had a pathogenic variant in mtDNA, and a revised diagnosis (GLUL) in one case that had abnormal pathology but no pathogenic mtDNA variant. In the 80 NO-Dx patients, nWES found non-mitochondrial diagnosis in 26 patients and a mitochondrial diagnosis in 1 patient. A genetic diagnosis was obtained in 53/116 (45.70%) cases that were recruited for NGS, but not in 11/116 (9.48%) of cases with abnormal mitochondrial neuropathology. Our results show that a non-invasive, bigenomic sequencing (BGS) approach (using both a nWES and optimized mtDNA analysis to include large deletions) should be the first step in investigating for mitochondrial diseases. There may still be a role for tissue biopsy in unsolved cases or when the diagnosis is still not clear after NGS studies.

Advances in Sports Medicine and Care of the Adolescent Athlete.

Over the past several decades there has been a dramatic change in the landscape of youth sports. This article provides an overview of sport-related injuries in the child and adolescent populations, looking at changes over the past 10 to 15 years. A closer look at frequently seen injuries, including assessment, diagnosis, and management in osteochondritis dissecans, sprains, fractures, anterior cruciate ligament, and meniscus tears, is provided. Current protocols and screening tools for this population are discussed, including preparticipation screening. The nursing implications in the clinical and community setting are identified, with ways to incorporate this information into practice.

Chorea and Parkinsonism with Elevated Striational Antibody.

Subacute onset of a mixed movement disorder should alert the clinician to the possibility of an autoimmune or paraneoplastic cause of symptoms. Striational antibodies have been associated with myasthenia gravis but a mixed movement disorder has been rarely reported with this antibody. We report a 90-year-old female who presented with generalized chorea, blepharospasm, and parkinsonism. Extensive evaluation was done which showed an elevation in striational antibody and there was no evidence of malignancy. The patient responded dramatically to intravenous steroids. We suggest that striational antibody should be routinely tested as a part of the work-up for autoimmune or paraneo lastic movement disorder. The presence of chorea in a very elderly patient should not be dismissed as "senile chorea" and a search for treatabl etiology should always be performed.

Trans-inclusive genetic counseling services: Recommendations from members of the transgender and non-binary community.

The term transgender is used to describe individuals whose gender identity does not align with their sex assigned at birth. The term transgender can include individuals who identify as men, as women, as both of the traditional concepts of masculine and feminine gender, or neither masculine or feminine. The latter two gender identities may also be known as non-binary or gender non-conforming identities. Transgender individuals may attend genetic counseling for a variety of reasons, but current pedigree nomenclature does not adequately represent both sex assigned at birth (which is important for determining risk for certain conditions) and gender (which is important for providing trans-inclusive care). We conducted an interpretive description (qualitative, interview based) study with individuals from the transgender community to gather insight on pedigree nomenclature and more broadly, how to provide safe and effective genetic counseling for transgender individuals. We conducted semi-structured telephone interviews with individuals who identified as transgender or gender non-binary, transcribed them verbatim, and checked them for accuracy before coding and inductively identifying themes. Among our eight participants, five identified as trans-masculine, two as trans-feminine, five as non-binary/gender non-conforming (some participants had more than one gender identity). From the interviews, we identified a single key, over-arching theme: participants' felt it is the genetic counselor's responsibility to create safety and provide clarity about the clinical importance of both sex assigned at birth and gender identity for trans patients. Two specific strategies that counselors could use to achieve this safety and clarity were discussed extensively: (a) validating gender identity and (b) using inclusive and well-defined pedigree symbols that denote both sex assigned at birth and gender identity. Our data have important practice implications in terms of the importance of validating gender identity and using respectful pedigree symbols.

Stable Density and Dynamics of Dendritic Spines of Cortical Neurons Across the Estrous Cycle While Expressing Differential Levels of Sensory-Evoked Plasticity.

Periodic oscillations of gonadal hormone levels during the estrous cycle exert effects on the female brain, impacting cognition and behavior. While previous research suggests that changes in hormone levels across the cycle affect dendritic spine dynamics in the hippocampus, little is known about the effects on cortical dendritic spines and previous studies showed contradictory results. In this in vivo imaging study, we investigated the impact of the estrous cycle on the density and dynamics of dendritic spines of pyramidal neurons in the primary somatosensory cortex of mice. We also examined if the induction of synaptic plasticity during proestrus, estrus, and metestrus/diestrus had differential effects on the degree of remodeling of synapses in this brain area. We used chronic two-photon excitation (2PE) microscopy during steady-state conditions and after evoking synaptic plasticity by whisker stimulation at the different stages of the cycle. We imaged apical dendritic tufts of layer 5 pyramidal neurons of naturally cycling virgin young female mice. Spine density, turnover rate (TOR), survival fraction, morphology, and volume of mushroom spines remained unaltered across the estrous cycle, and the values of these parameters were comparable with those of young male mice. However, while whisker stimulation of female mice during proestrus and estrus resulted in increases in the TOR of spines (74.2 ± 14.9% and 75.1 ± 12.7% vs. baseline, respectively), sensory-evoked plasticity was significantly lower during metestrus/diestrus (32.3 ± 12.8%). In males, whisker stimulation produced 46.5 ± 20% increase in TOR compared with baseline-not significantly different from female mice at any stage of the cycle. These results indicate that, while steady-state density and dynamics of dendritic spines of layer 5 pyramidal neurons in the primary somatosensory cortex of female mice are constant during the estrous cycle, the susceptibility of these neurons to sensory-evoked structural plasticity may be dependent on the stage of the cycle. Since dendritic spines are more plastic during proestrus and estrus than during metestrus/diestrus, certain stages of the cycle could be more suitable for forms of memory requiring de novo formation and elimination of spines and other stages for forms of memory where retention and/or repurposing of already existing synaptic connections is more pertinent.

Assessing the performance of a Loop Mediated Isothermal Amplification (LAMP) assay for the detection and subtyping of high-risk suptypes of Human Papilloma Virus (HPV) for Oropharyngeal Squamous Cell Carcinoma (OPSCC) without DNA purification.

BACKGROUND: Oropharyngeal Squamous Cell Carcinoma (OPSCC) is increasing in incidence despite a decline in traditional risk factors. Human Papilloma Virus (HPV), specifically subtypes 16, 18, 31 and 35, has been implicated as the high-risk etiologic agent. HPV positive cancers have a significantly better prognosis than HPV negative cancers of comparable stage, and may benefit from different treatment regimens. Currently, HPV related carcinogenesis is established indirectly through Immunohistochemistry (IHC) staining for p16, a tumour suppressor gene, or polymerase chain reaction (PCR) that directly tests for HPV DNA in biopsied tissue. Loop mediated isothermal amplification (LAMP) is more accurate than IHC, more rapid than PCR and is significantly less costly. In previous work we showed that a subtype specific HPV LAMP assay performed similar to PCR on purified DNA. In this study we examined the performance of this LAMP assay without DNA purification. METHODS: We used LAMP assays using established primers for HPV 16 and 18, and new primers for HPV 31 and 35. LAMP reaction conditions were tested on serial dilutions of plasmid HPV DNA to confirm minimum viral copy number detection thresholds. LAMP was then performed directly on different human cell line samples without DNA purification. RESULTS: Our LAMP assays could detect 105, 103, 104, and 105 copies of plasmid DNA for HPV 16, 18, 31, and 35, respectively. All primer sets were subtype specific, with no cross-amplification. Our LAMP assays also reliably amplified subtype specific HPV DNA from samples without requiring DNA isolation and purification. CONCLUSIONS: The high risk OPSCC HPV subtype specific LAMP primer sets demonstrated, excellent clinically relevant, minimum copy number detection thresholds with an easy readout system. Amplification directly from samples without purification illustrated the robust nature of the assay, and the primers used. This lends further support HPV type specific LAMP assays, and these specific primer sets and assays can be further developed to test for HPV in OPSCC in resource and lab limited settings, or even bedside testing.

Patients' perceptions of "uncertainty" in nurse practitioner consultations.

The aim of this qualitative study was to explore patients' perceptions of consulting with a nurse practitioner in situations of clinical uncertainty. Uncertainty in this context is defined as one where there is no obvious diagnosis, treatment or where the outcome of the consultation is not definite. Three general practice sites were recruited to participate. 43 patients who consulted with one of three nurse practitioners were interviewed using a semistructured schedule. The nurse practitioners identified uncertainty in 30 of these consultations; only two patients expressed any awareness of uncertainty with the consultation. The results showed that patients appear to accept that there will be a level of uncertainty in some consultations. Recognition of uncertainty within the consultation does not appear to have a negative effect on patients' perception of the nurse practitioner as they feel that the nurse will refer to a doctor if necessary.