Next-generation gene panel testing in adolescents and adults in a medical neuropsychiatric genetics clinic.

Intellectual disability (ID) encompasses a clinically and genetically heterogeneous group of neurodevelopmental disorders that may present with psychiatric illness in up to 40% of cases. Despite the evidence for clinical utility of genetic panels in pediatrics, there are no published studies in adolescents/adults with ID or autism spectrum disorder (ASD). This study was approved by our institutional research ethics board. We retrospectively reviewed the medical charts of all patients evaluated between January 2017 and December 2019 in our adult neuropsychiatric genetics clinic at the McGill University Health Centre (MUHC), who had undergone a comprehensive ID/ASD gene panel. Thirty-four patients aged > 16 years, affected by ID/ASD and/or other neuropsychiatric/behavioral disorders, were identified. Pathogenic or likely pathogenic variants were identified in one-third of our cohort (32%): 8 single-nucleotide variants in 8 genes (CASK, SHANK3, IQSEC2, CHD2, ZBTB20, TREX1, SON, and TUBB2A) and 3 copy number variants (17p13.3, 16p13.12p13.11, and 9p24.3p24.1). The presence of psychiatric/behavioral disorders, regardless of the co-occurrence of ID, and, at a borderline level, the presence of ID alone were associated with positive genetic findings (p = 0.024 and p = 0.054, respectively). Moreover, seizures were associated with positive genetic results (p = 0.024). One-third of individuals presenting with psychiatric illness who met our red flags for Mendelian diseases have pathogenic or likely pathogenic variants which can be identified using a comprehensive ID/ASD gene panel (~ 2500 genes) performed on an exome backbone.

Two-Step Fowler-Stephens orchiopexy for intra-abdominal testes: a 28-year single institution experience.

PURPOSE: Two-Step Fowler-Stephens orchiopexy for high undescended testes allows for mobilization of the testicle to the scrotal position while preserving perfusion by collateral circulation after gonadal vessel ligation. Although used for decades, the long-term efficacy of this procedure has not been reported. We present our 28-year clinical experience with this technique. MATERIALS AND METHODS: We retrospectively studied a cohort of patients who underwent 2-step Fowler-Stephens orchiopexy at our institution between 1982 and 2009. Patients were excluded if either step was performed elsewhere or if followup was less than 6 months. Bivariate and multivariate analyses were performed to determine associations between clinical, surgical and anatomical factors and testicular viability at last followup. RESULTS: A total of 62 patients (79 testes) met inclusion criteria. Median followup was 3.1 years (range 0.6 to 20). Based on the most recent examination/ultrasound, 70.9% of testicles were considered to be normal, with the remainder exhibiting relative (15.2%) or complete (14.9%) atrophy. Of the 10 testes assessed at or after puberty 6 were normal and 4 showed relative atrophy. On bivariate and multivariate analyses only an open second stage approach was associated with normal testicular viability, with 69.9% of normal vs 18.9% of completely atrophic testes being managed by an open approach (p = 0.0084). CONCLUSIONS: Thought to be highly effective in short-term followup, our data suggest that 2-step Fowler-Stephens orchiopexy leads to complete testicular viability in 70.9% of cases. This viability is strongly associated with an open second step.

Neurocognitive deficits and neuroimaging abnormalities are prevalent in children with lupus: clinical and research experiences at a US pediatric institution.

Neurocognitive impairments and neuroimaging abnormalities are frequently observed in adults with systemic lupus erythematosus. There is a paucity of similar data in childhood-onset disease. We hypothesized that neurocognitive and neuroimaging abnormalities would be prevalent in children undergoing neuropsychological evaluations. We reviewed patient neurocognitive evaluations performed at a large United States pediatric institution during the period 2001 to 2008. Records were retrieved from 24 children referred to neuropsychology due to clinical indications. Data from 15 children enrolled in a prospective structure-function association study were also analyzed. Subjects were predominantly African-American and Hispanic adolescent girls of average intelligence. aPL positivity and aspirin use was prevalent. Neurocognitive impairment was designated in 70.8% of retrospective, and 46.7% of prospective cohort patients. Deficits were seen at times of wellness, without previous neuropsychiatric lupus, and early in disease courses. Scores >1.5 standard deviations below published age-matched norms were common in tests of executive functioning, visual memory and visual-spatial planning. Features of depression were seen in 33.3% of the children in the retrospective cohort (clinical referrals). Cerebral and cerebellar volume loss was observed in a majority of blinded prospective cohort research magnetic resonance images (73.3% and 67.7% respectively). White matter hyperintensities were observed in retrospective and prospective cohort magnetic resonance images (36.6% and 46.7% respectively). Larger prospective studies that elucidate structure-function associations in children with systemic lupus erythematosus are planned.

Fragile X mental retardation protein replacement restores hippocampal synaptic function in a mouse model of fragile X syndrome.

Fragile X syndrome (FXS) is caused by a mutation that silences the fragile X mental retardation gene (FMR1), which encodes the fragile X mental retardation protein (FMRP). To determine whether FMRP replacement can rescue phenotypic deficits in a fmr1-knockout (KO) mouse model of FXS, we constructed an adeno-associated virus-based viral vector that expresses the major central nervous system (CNS) isoform of FMRP. Using this vector, we tested whether FMRP replacement could rescue the fmr1-KO phenotype of enhanced long-term depression (LTD), a form of synaptic plasticity that may be linked to cognitive impairments associated with FXS. Extracellular excitatory postsynaptic field potentials were recorded from CA3-CA1 synaptic contacts in hippocampal slices from wild-type (WT) and fmr1-KO mice in the presence of AP-5 and anisomycin. Paired-pulse low-frequency stimulation (PP-LFS)-induced LTD is enhanced in slices obtained from fmr1 KO compared with WT mice. Analyses of hippocampal synaptic function in fmr1-KO mice that received hippocampal injections of vector showed that the PP-LFS-induced LTD was restored to WT levels. These results indicate that expression of the major CNS isoform of FMRP alone is sufficient to rescue this phenotype and suggest that post-developmental protein replacement may have the potential to improve cognitive function in FXS.

BAFF is increased in renal transplant patients following treatment with alemtuzumab.

Alemtuzumab is a monoclonal antibody that depletes T and B cells and is used as induction therapy for renal transplant recipients. Without long-term calcineurin inhibitor (CNI) therapy, alemtuzumab-treated patients have a propensity to develop alloantibody and may undergo antibody-mediated rejection (AMR). In pursuit of a mechanistic explanation, we analyzed peripheral B cells and serum of these patients for BAFF (Blys) and BAFF-R, factors known to be integral for B-cell activation, survival, and homeostasis. Serum BAFF levels of 22/24 alemtuzumab-treated patients were above normal range, with average levels of 1967 pg/mL compared to 775 pg/mL in healthy controls (p = 0.006). BAFF remained elevated 2 years posttransplant in 78% of these patients. BAFF-R on CD19(+) B cells was significantly downregulated, suggesting ligand/receptor engagement. BAFF mRNA expression was increased 2-7-fold in CD14(+) cells of depleted patients, possibly linking monocytes to the BAFF dysregulation. Addition of recombinant BAFF to mixed lymphocyte cultures increased B-cell activation to alloantigen, as measured by CD25 and CD69 coexpression on CD19(+) cells. Of note, addition of sirolimus (SRL) augmented BAFF-enhanced B-cell activation whereas CNIs blocked it. These data suggest associations between BAFF/BAFF-R and AMR in alemtuzumab-treated patients.

Early and limited use of tacrolimus to avoid rejection in an alemtuzumab and sirolimus regimen for kidney transplantation: clinical results and immune monitoring.

Alemtuzumab induction with 60 days of tacrolimus treatment and continuous sirolimus treatment prevented acute rejection in nine of 10 consecutive renal allograft recipients. All patients are alive with a functioning kidney graft at 27-39 months of follow-up. Extensive immune monitoring was performed in all patients. Alloantibody detection, cytokine kinetics assay (CKA), and trans vivo delayed-type hypersensitivity (DTH) assay were performed every 6 months showing correlation with clinical evolution. Despite alloantibody presence in five patients, eight patients remain without the need for specific treatment and only sirolimus monotherapy in decreasing dosage. Four patients take only 1 mg sirolimus daily with levels of 3-4 ng/mL. One patient showed clinical signs of rejection at month 9 post-transplant, with slow increase in serum creatinine and histological signs of mixed cellular (endarteritis) and humoral rejection (C4d positivity in peritubular capillaries and donor-specific antibody (DSA)). In summary, the addition of tacrolimus therapy for 2 months to a steroid-free, alemtuzumab induction and sirolimus maintenance protocol limited the previously shown acute rejection development. Nevertheless, alloantibody was present in serum and/or C4d present on 1-year biopsy in half the patients. The combination of CKA and DSA monitoring or the performance of transvivo DTH correlated with immune status of the patients.

International public opinion on the environment.

This article analyzes public opinion data on environmental issues collected in two major surveys. The data reveal substantial concern about the environment in both developing and industrial countries along with perceptions that the quality of the environment has declined and will continue to decline. Developing country respondents rate their local and national environmental quality lower than do industrial country respondents, whereas both groups rate global environmental quality about the same. The data also reveal considerable willingness among the developing and industrial countries to accept responsibility for the world's environmental problems and recognition of the importance of governments in addressing local and national environmental issues and of strong international agencies in addressing transnational issues.

Something to be done: treating HIV/AIDS. River Path Associates.

Largely because of disparities in access to drug treatment and care, AIDS morbidity and mortality have fallen in the developed world but continue to rise among developing countries. Achieving more equitable access to AIDS drugs is hindered by high drug prices, technical complexities related to the provision of health care, and conflict among stakeholders. Recognition that health is vital to the prospects of the emerging global society must be combined with new mechanisms to help all stakeholders work together cooperatively. Tiered drugs pricing should be coupled with investment in health services. An independent "Global Task Force," able to act as an "active think tank," could build consensus about the way forward.

Benefits and costs of HIV testing.

The benefits and costs of human immunodeficiency virus (HIV) testing in employment settings are examined from two points of view: that of private employers whose profitability may be affected by their testing policies and that of public policy-makers who may affect social welfare through their design of regulations related to HIV testing. The results reveal that HIV testing is clearly not cost-beneficial for most firms, although the benefits of HIV testing may outweigh the costs for some large firms that offer generous fringe-benefit packages and that recruit workers from populations in which the prevalence of HIV infection is high. The analysis also indicates that the testing decisions of unregulated employers are not likely to yield socially optimal economic outcomes and that existing state and federal legislation related to HIV testing in employment settings has been motivated primarily by concerns over social equity.

The economic impact of AIDS in the United States.

This analysis of several previous studies of the cost of AIDS suggests that the lifetime cost of medical care per patient will not exceed +80,000, an amount similar to the cost of treating other serious illnesses. If current projections of future AIDS cases are accurate, the cumulative lifetime costs of 270,000 cases diagnosed between 1981 and the end of 1991 will not exceed +22 billion. This amount is small compared with total U.S. medical spending. The economic impact of AIDS on San Francisco, New York, and some other cities, however, is likely to be more serious. The AIDS epidemic will also highlight the financial problems of Americans who face large medical bills without adequate insurance.

Prevalence estimate of intimate partner violence in Jordan.

This study examined the prevalence of intimate partner violence in Jordan among a sample of 517 reproductive health clinic attendees. Intimate partner violence was measured using the World Health Organization's domestic violence questionnaire which was modified by the results of focus group discussions conducted in Amman. The percentages of women experiencing at least 1 form of control or violence since marriage were: control, 97.2%; psychological violence, 73.4%; physical violence, 31.2%; and sexual violence, 18.8%. Modifications of the WHO questionnaire were needed to measure control and psychological violence in Jordan. Similar modifications might be required when conducting research in the Region.

Calcineurin inhibitor withdrawal after renal transplantation with alemtuzumab: clinical outcomes and effect on T-regulatory cells.

To address the results of calcineurin inhibitor (CNI) withdrawal after alemtuzumab induction relative to CNI continuation, we performed a pilot randomized clinical trial in renal allograft recipients on CNI, a mycophenolic acid derivative and steroids after the first 2 months posttransplantation. Forty patients were randomized to taper off CNI or to maintain it, and followed for at least 1 year. Four patients in the withdrawal group were treated for acute rejection while no patient received antirejection treatment in the control group. Two control patients withdrew CNI due to nephrotoxicity. Estimated GFR was similar in both groups after 1 year. Flow cytometry of CD4(+)CD25(+)CTLA-4(+)FoxP3(+) regulatory T cells (Treg) demonstrated a significant increase in Treg percentages in the peripheral blood of alemtuzumab-treated patients on CNI early postransplant. Furthermore, the increased Treg percentages in the withdrawal cohort were unchanged at month 6 postenrollment, whereas they decreased significantly in those patients maintained on CNI. Patients withdrawn from CNI after alemtuzumab trend toward a higher rejection rate, but most patients can be weaned from a CNI using this regimen. With the exception of maintaining increased Treg levels, the benefits are not appreciable in this short follow-up, and a larger randomized trial is justified.

CD4+ CD25+ FOXP3+ regulatory T cells increase de novo in kidney transplant patients after immunodepletion with Campath-1H.

Campath-1H (Alemtuzumab) is an effective immunodepletion agent used in renal transplantation. To evaluate its influence on T lymphocytes during repletion, we analyzed peripheral blood from Campath-1H-treated renal allograft recipients for the presence of FOXP3(+) regulatory T (Treg) cells. Flow cytometry demonstrated that CD4(+)CD25(+)FOXP3(+) lymphocytes increased significantly within the CD4(+) T-cell population, skewing Treg/Teff (T effector) ratios for up to several years. In contrast, Treg levels in patients treated with anti-CD25 (Basiliximab) and maintained on CsA demonstrated a sustained decrease. The increase in Tregs in Campath-1H treated patients developed independent of maintenance immunosuppression. Importantly, the increase in Tregs was not fully explained by their homeostatic proliferation, increased thymic output, or Treg sparing, suggesting de novo generation/expansion. Consistent with this, in vitro stimulation of PBMCs with Campath-1H, with or without anti-CD3, activation led to an increase in CD4(+)CD25(+)FOXP3(+) cells that had suppressive capabilities. Together, these data suggest that Campath-1H promotes an increase in peripheral Tregs and may act as an intrinsic generator of Tregs in vivo.

Different tropism of adenoviruses and adeno-associated viruses to corneal cells: implications for corneal gene therapy.

PURPOSE: Diseased corneas are potential targets for viral-based gene therapy to normalize (stimulate or inhibit) the expression of specific proteins. The choice of viral vectors is important to achieve optimal effect. The purpose of this study was to compare the tropism to different corneal cells of recombinant adenovirus (rAV) and recombinant adeno-associated virus (rAAV) constructs using live rabbit and organ-cultured human corneas. METHODS: rAV constructs harbored the green fluorescent protein (GFP) gene under the control of major immediate early cytomegalovirus (CMV) promoter. rAAV constructs from virus serotypes 1, 2 5, 7, and 8 had GFP under the chicken beta-actin promoter and CMV enhancer. For organ culture, 16 healthy and diabetic postmortem human corneas were used. Five or fifteen microl rAV at 10(7) plaque forming units per 1 microl were added for 2 days to culture medium of uninjured corneas that were further cultured for 5-32 days. rAAV were added at 1.2-7.8x10(10) vector genomes per cornea for 3 days to each cornea; the culture then continued for another 14-23 days. Corneal cryostat sections were examined by immunohistochemistry. Live rabbit corneas were used following excimer laser ablation of the corneal epithelium with preservation of the basal cell layer. Equal numbers of rAAV particles (2x10(11) vector genomes) were applied to the cornea for 10 min. After seven days to allow for corneal healing and gene expression the animals were euthanized, the corneas were excised, and sections analyzed by immunohistochemistry. RESULTS: By direct fluorescence microscopy of live organ-cultured human corneas GFP signal after rAV transduction was strong in the epithelium with dose-dependent intensity. On corneal sections, GFP was seen in all epithelial layers and some endothelial cells but most keratocytes were negative. In rAAV-transduced organ-cultured human corneas GFP signal could only be detected with anti-GFP antibody immunohistochemistry. GFP was observed in the epithelium, keratocytes, and endothelium, with more pronounced basal epithelial cell staining with rAAV1 than with other serotypes. No difference in the GFP expression patterns or levels between normal and diabetic corneas was noted. The rabbit corneas showed very similar patterns of GFP distribution to human corneas. With all rAAV serotype vectors, GFP staining in the epithelium was significantly (p=0.007) higher than the background staining in non-transduced corneas, with a trend for rAAV1 and rAAV8 to produce higher staining intensities than for rAAV2, rAAV5 (p=0.03; rAAV5 versus rAAV1), and rAAV7. rAAV serotype vectors also transduced stromal and endothelial cells in rabbit corneas to a different extent. CONCLUSIONS: rAAV appears to reach many more corneal cells than rAV, especially keratocytes, although GFP expression levels were lower compared to rAV. rAV may be more useful than rAAV for gene therapy applications requiring high protein expression levels, but rAAV may be superior for keratocyte targeting.

Novel nuclear signaling pathway mediates activation of fibroblast growth factor-2 gene by type 1 and type 2 angiotensin II receptors.

In bovine adrenal medullary cells synergistically acting type 1 and type 2 angiotensin II (AII) receptors activate the fibroblast growth factor-2 (FGF-2) gene through a unique AII-responsive promoter element. Both the type 1 and type 2 AII receptors and the downstream cyclic adenosine 1',3'-monophosphate- and protein kinase C-dependent signaling pathways activate the FGF-2 promoter through a novel signal-transducing mechanism. This mechanism, which we have named integrative nuclear FGF receptor-1 signaling, involves the nuclear translocation of FGF receptor-1 and its subsequent transactivation of the AII-responsive element in the FGF-2 promoter.

Smile lifts - a functional and aesthetic perspective.

Cosmetic dentistry has evolved with the advent of more robust porcelain materials and ever-stronger bonding agents. This series of three articles aims to provide a practical overview of what is now possible both functionally and cosmetically from the preparation of a small number of teeth, through a whole smile, to full mouth rehabilitation. A complete diagnosis is the starting point to planning any cosmetic or functional changes. Guidance is given on the techniques used but adequate training must be considered essential before embarking upon modification in occlusal schemes or even minor adjustments in smile design. Porcelain laminate veneers have a role in the restoration and rehabilitation of a wearing and functionally compromised dentition.

Aesthetic changes with four anterior units.

Cosmetic dentistry has evolved with the advent of more robust porcelain materials and ever-stronger bonding agents. This series of three articles aims to provide a practical overview of what is now possible both functionally and cosmetically from the preparation of a small number of teeth, through a whole smile, to full mouth rehabilitation. A complete diagnosis is the starting point to planning any cosmetic or functional changes. Guidance is given on the techniques used but adequate training must be considered essential before embarking upon modification in occlusal schemes or even minor adjustments in smile design. Conservative use of porcelain laminate veneers can result in significant aesthetic improvements to a patient's smile and it is not always necessary to prepare all the teeth visible in a smile.

Increasing occlusal vertical dimension--why, when and how.

Cosmetic dentistry has evolved with the advent of more robust porcelain materials and ever-stronger bonding agents. This series of three articles aims to provide a practical overview of what is now possible both functionally and cosmetically from the preparation of a small number of teeth, through a whole smile, to full mouth rehabilitation. A complete diagnosis is the starting point to planning any cosmetic or functional changes. Guidance is given on the techniques used but adequate training must be considered essential before embarking upon modification in occlusal schemes or even minor adjustments in smile design. Understanding vertical dimension and how and when it can be changed has always been a challenging prospect for the general dental practitioner. This article aims to discuss the rationale behind changes in vertical dimension and demonstrate how it can be achieved in general practice assuming adequate hands-on postgraduate training has been completed.

The role of intratumour therapy with electroporation and bleomycin in the management of advanced squamous cell carcinoma of the head and neck.

OBJECTIVES: To determine the safety and efficacy of electroporation with bleomycin in patients with advanced squamous cell carcinoma of the head and neck. METHODS: Two open-label, multicenter, single-arm Phase II studies of intratumour electroporation therapy. Sixty-two patients with 86 squamous cell carcinoma tumours of the head and neck were enrolled. Twenty-five patients were treated with bleomycin alone. Fifty-four patients (17 initially treated with bleomycin alone) were treated with electroporation and bleomycin therapy. Local tumour response was measured. RESULTS: In the bleomycin alone group, one tumour showed a partial response and 36 tumours showed no response to treatment. In the bleomycin with electroporation groups, 17 tumours showed complete response, 22 tumours showed partial response and 30 failed to achieve more than a 50% reduction in tumour size (no response). Bleomycin with electroporation had a significantly (p<0.001) greater number of patients showing a partial or complete response to the therapy when compared to bleomycin alone. Thirteen adverse events were reported which included five episodes of local bleeding, six local infections, one local tongue swelling and one cardiac arrhythmia. CONCLUSIONS: Fifty-seven percent of squamous cell carcinomas of the head and neck demonstrated a partial or complete response to intratumour electroporation with bleomycin suggesting that further work investigating its use as a treatment for local control of these lesions should be pursued.

In vivo gene transfer to the brain cortex using a single injection of HSV-1 vector into the medial septum.

This study shows that an ICP4-replication-deficient herpes simplex virus containing the Moloney murine leukaemia virus LTR fused with the coding sequence for the beta-galactosidase gene can be used as a very effective vector for delivering the beta-galactosidase reporter gene into the rat brain septum. F344 rats received bilateral stereotaxic injections into the nucleus of the diagonal band and into the medial septum. The X-gal stain was used to detect the activity of the expressed beta-galactosidase enzyme. The delivered reporter gene was expressed successfully not only in the neuronal cells of the injected areas but also in cells that project to the injection area such as cortex cells about 6 mm away from the injection sites. Expression was visible at 1, 3 and 9 weeks following injection. We conclude that this vector can effectively deliver genes into different regions of the mature mammalian brain and also to areas distant from the injection site.