Two-Generation Epsilon-Sarcoglycan Gene (SGCE) Mutation-Associated Myoclonus-Dystonia (DYT-SGCE) Misdiagnosed as Tourette's Syndrome: A Case Series.

This case series provides a diagnosis of myoclonus-dystonia syndrome (MDS) in two patients whose original presentation was thought to be Tourette's syndrome. The first patient presented with dystonia and myoclonus, which progressively worsened with age, and was diagnosed with an epsilon-sarcoglycan gene (SGCE) mutation. The patient's father, who was diagnosed in his childhood with Tourette's syndrome, also received genetic testing, which proved that to be a misdiagnosis and confirmed that he was the carrier of the SGCE mutation. Both patients were subjected to a levodopa trial, which proved to be an effective treatment. To our knowledge, these are the first reported cases of heterozygous pathogenic mutation of SGCE in Puerto Rico.

Surgical and Medical Management of Purulent Bacterial Pericarditis With Pericardial Mass in a Young Child.

Purulent bacterial pericarditis is rare and associated with significant short- and long-term morbidity. We report a case of purulent bacterial pericarditis caused by Group A Streptococcus in an immunocompetent young child presenting with a pericardial mass. She was successfully treated with a combined medical and early surgical approach. (Level of Difficulty: Intermediate.).

The Impact of Preoperative Oral Health on Buccal Mucosa Graft Histology.

PURPOSE: Previous studies have elucidated the unique macroscopic and histological properties of buccal mucosa that make it a viable and durable graft for urethral augmentation. However, no prior literature has directly investigated the impact of preoperative oral health on these features. MATERIALS AND METHODS: We analyzed all consenting patients who underwent buccal mucosal graft (BMG) urethroplasty at our institution from 2018 to 2020. Validated oral health surveys, the Oral Health Impact Profile (OHIP-14) and the Kayser-Jones Brief Oral Health Status Examination (BOHSE) were completed preoperatively. A staff pathologist analyzed BMG histology and quantified oral mucositis using a modified Oral Mucosa Rating Scale. RESULTS: We analyzed 51 patients with a median age of 40 years (IQR 31-58). Mean BOHSE score was 1.1 and OHIP-14 score was 1.4. Median epithelial thickness was 530 µm and lamina propria thickness was 150 µm. On age-adjusted analysis, increasing BOHSE and OHIP-14 were associated with decreasing epithelial thickness (p values <0.05). Higher BOHSE scores also correlated with thinner lamina proprias (p=0.05) and increased graft stretch (p=0.03). The 2 patients with postoperative urine leaks and available graft histology had lamina propria thicknesses well below the cohort median, at 50 µm and 60 µm. CONCLUSIONS: This is the first study to demonstrate that oral health conditions impact graft histology and stretch. Although much remains to be learned, our findings shed light on the potential importance of optimizing oral health prior to BMG urethroplasty, and raise the question of if preoperative mucosal biopsy could help inform surgical decision making and discussions regarding surgical success.

Synchronous Pulmonary Adenocarcinomas.

OBJECTIVES: To determine concordance/discordance between morphology and molecular testing (MT) among synchronous pulmonary carcinomas using targeted next generation sequencing (NGS), with and without comprehensive molecular review (CMR), vs analyses of multiple singe genes (non-NGS). METHODS: Results of morphologic and MT assessment were classified as concordant, discordant, or indeterminate. For discordant cases, comprehensive histologic assessment (CHA) was performed. RESULTS: Forty-seven cases with 108 synchronous tumors were identified and underwent MT (NGS, n = 23 and non-NGS, n = 24). Histology and MT were concordant, discordant, and indeterminate in 53% (25/47), 21% (10/47), and 26% (12/47) of cases, respectively. CHA of the 10 discordant cases revised results of three cases. CONCLUSIONS: There is discordance between histology and MT in a subset of cases and MT provides an objective surrogate for staging synchronous tumors. A limited gene panel is sufficient for objectively assessing a relationship if the driver mutations are distinct. Relatedness of mutations require CMR with a larger NGS panel (eg, 50 genes).

Nicotinic Acetylcholine Receptors in HIV: Possible Roles During HAND and Inflammation.

Infection with the human immunodeficiency virus (HIV) remains a threat to global health. Since its discovery, many efforts have been directed at understanding the mechanisms and consequences of infection. Although there have been substantial advances since the advent of antiretroviral therapy, there are still complications that significantly compromise the health of infected patients, particularly, chronic inflammation and HIV-associated neurocognitive disorders (HAND). In this review, a new perspective is addressed in the field of HIV, where the alpha7 nicotinic acetylcholine receptor (α7-nAChR) is the protagonist. We comprehensively discuss the available evidence implicating α7-nAChRs in the context of HIV and provide possible explanations about its role in HAND and inflammation in both the central nervous system and the periphery.

The alpha7-nicotinic receptor contributes to gp120-induced neurotoxicity: implications in HIV-associated neurocognitive disorders.

Currently, there are no specific therapies to treat HIV-1 associated neurocognitive disorders (HAND). The HIV-1 envelope, gp120, induces neuropathological changes similar to those in HAND patients; furthermore, it triggers an upregulation of the α7-nicotinic acetylcholine receptor (α7-nAChR), facilitating intracellular calcium overload and neuronal cell death. Using a gp120IIIB-transgenic mouse (gp120-tgm) model, we demonstrate that α7-nAChRs are upregulated on striatal neurons. Activation of α7-nAChRs leads to an increase in both intracellular calcium and percentage of apoptotic cells, which can be abrogated by antagonizing the receptor, suggesting a role for α7-nAChRs in gp120-induced neurotoxicity. Moreover, we demonstrate for the first time that gp120-tgm have learning deficiencies on a striatum-dependent behavioral task. They also show locomotor deficiencies, which improved with α7-nAChR antagonists, further supporting a role for this receptor in gp120-induced neurotoxicity. Together, these results uncover a new mechanism through which gp120-induced modulation of α7-nAChRs in the striatum can contribute to HAND development.

PD-L1 expression in non-small cell lung carcinoma: Comparison among cytology, small biopsy, and surgical resection specimens.

BACKGROUND: One immunotherapeutic agent for patients with advanced non-small cell lung carcinoma, pembrolizumab, has a companion immunohistochemistry (IHC)-based assay that predicts response by quantifying programmed death-ligand 1 (PD-L1) expression. The current study assessed the feasibility of quantifying PD-L1 expression using cytologic non-small cell lung carcinoma specimens and compared the results with those from small biopsy and surgical resection specimens. METHODS: PD-L1 expression was quantified using the IHC-based 22C3 pharmDx assay, with "positivity" defined as staining in ≥50% viable tumor cells; ≥ 100 tumor cells were required for test adequacy. For cytology specimens, IHC was performed on cell block sections. RESULTS: A total of 214 specimens were collected from 188 patients, 206 of which (96%) were found to be adequately cellular, including 36 of 40 cytology (90%) and 69 of 72 small biopsy (96%) specimens. There was no significant difference noted with regard to the feasibility of PD-L1 IHC on small biopsy specimens compared with surgical resection specimens (P = .99), or between the percentage of PD-L1-positive cytology and histology (including surgical resection and histologic small biopsy) specimens (P = .083). PD-L1 expression was found to be concordant among samples from 21 of 23 patients from whom > 1 specimen was collected (91%). There also was no significant difference observed with regard to rates of PD-L1 positivity when comparing age, sex, diagnosis, and specimen site. CONCLUSIONS: Quantification of PD-L1 expression is feasible on cytology specimens, and the results are comparable to those obtained from surgical resection and small biopsy specimens, including in matched specimens and using a single predictive IHC marker. Future studies will be necessary to determine the comparative value of other antibodies and their ability to predict response to immunotherapy. Cancer Cytopathol 2017;125:896-907. © 2017 American Cancer Society.

A Panel of Slow-Channel Syndrome Mice Reveals a Unique Locomotor Behavioral Signature.

Muscle nicotinic acetylcholine receptor (nAChR) mutations can lead to altered channel kinetics and neuromuscular junction degeneration, a neurodegenerative disorder collectively known as slow-channel syndrome (SCS). A multivariate analysis using running wheels was used to generate activity profiles for a variety of SCS models, uncovering unique locomotor patterns for the different nAChR mutants. Particularly, the αL251T and ɛL269F mutations exhibit decreased event distance, duration, and velocity over a period of 24 hours. Our approach suggests a robust relationship between the pathophysiology of SCS and locomotor activity.

Next-generation sequencing of non-small cell lung cancer using a customized, targeted sequencing panel: Emphasis on small biopsy and cytology.

BACKGROUND: Next-generation sequencing (NGS) with a multi-gene panel is now available for patients with lung adenocarcinoma, but the performance characteristics and clinical utility of this testing are not well-described. We present the results of an extended 467 gene panel in a series of advanced, highly selected nonsmall cell lung cancer (NSCLC) patients using a range of specimens, including predominantly small biopsy and cytology specimens. MATERIALS AND METHODS: A retrospective review of 22 NSCLC biopsies sent for NGS using an extended gene panel from January 2014 to July 2015. The customized NGS panel sequences 467 cancer-associated genes with exonic and intronic sequences obtained from purified tumor DNA. Genomic alterations, patient characteristics, and success of testing were determined. RESULTS: The majority of samples tested were metastatic lung adenocarcinoma on final pathology. Of the 22 specimens tested, 5 (22.7%) were surgical resections and 17 (77.3%) were small biopsy and cytology specimens. Twenty-one (95%) of the specimens were adequate for full sequencing and yielded a total of 204 genomic alterations (average 8.9 per tumor), of which 17 (average 0.81 per tumor) were actionable and/or clinically relevant. Genomic alterations were found most commonly in the TP53, EGFR, EPHB1, MLL3, APC, SETD2, KRAS, DNMT3A, RB1, CDKN2A, ARID1A, EP300, KDM6B, RAD50, STK11, and BRCA2 genes. CONCLUSIONS: NGS using a comprehensive gene panel was performed successfully in 95% of all NSCLC cases in this series, including 94% small biopsy and cytology specimens and 100% surgical resections. This custom assay was performed on a range of tumor specimens and demonstrates that small specimens are able to provide a similar depth of information as larger ones. As many patients present at an advanced stage and only small specimens are obtained, the information these provide has the potential for guiding treatment in highly selected patients with advanced lung adenocarcinoma.

Heterogeneous Inhibition in Macroscopic Current Responses of Four Nicotinic Acetylcholine Receptor Subtypes by Cholesterol Enrichment.

The nicotinic acetylcholine receptor (nAChR), located in the cell membranes of neurons and muscle cells, mediates the transmission of nerve impulses across cholinergic synapses. In addition, the nAChR is also found in the electric organs of electric rays (e.g., the genus Torpedo). Cholesterol, which is a key lipid for maintaining the correct functionality of membrane proteins, has been found to alter the nAChR function. We were thus interested to probe the changes in the functionality of different nAChRs expressed in a model membrane with modified cholesterol to phospholipid ratios (C/P). In this study, we examined the effect of increasing the C/P ratio in Xenopus laevis oocytes expressing the neuronal α7, α4ß2, muscle-type, and Torpedo californica nAChRs in their macroscopic current responses. Using the two-electrode voltage clamp technique, it was found that the neuronal α7 and Torpedo nAChRs are significantly more sensitive to small increases in C/P than the muscle-type nAChR. The peak current versus C/P profiles during enrichment display different behaviors; α7 and Torpedo nAChRs display a hyperbolic decay with two clear components, whereas muscle-type and α4ß2 nAChRs display simple monophasic decays with different slopes. This study clearly illustrates that a physiologically relevant increase in membrane cholesterol concentration produces a remarkable reduction in the macroscopic current responses of the neuronal α7 and Torpedo nAChRs functionality, whereas the muscle nAChR appears to be the most resistant to cholesterol inhibition among all four nAChR subtypes. Overall, the present study demonstrates differential profiles for cholesterol inhibition among the different types of nAChR to physiological cholesterol increments in the plasmatic membrane. This is the first study to report a cross-correlation analysis of cholesterol sensitivity among different nAChR subtypes in a model membrane.

Expression of CHRFAM7A and CHRNA7 in neuronal cells and postmortem brain of HIV-infected patients: considerations for HIV-associated neurocognitive disorder.

Despite the recent advances in antiretroviral therapy, human immunodeficiency virus type 1 (HIV-1) remains a global health threat. HIV-1 affects the central nervous system by releasing viral proteins that trigger neuronal death and neuroinflammation, and promotes alterations known as HIV-associated neurocognitive disorders (HAND). This disorder is not fully understood, and no specific treatments are available. Recently, we demonstrated that the HIV-1 envelope protein gp120IIIB induces a functional upregulation of the α7-nicotinic acetylcholine receptor (α7) in neuronal cells. Furthermore, this upregulation promotes cell death that can be abrogated with receptor antagonists, suggesting that α7 may play an important role in the development of HAND. The partial duplication of the gene coding for the α7, known as CHRFAM7A, negatively regulates α7 expression but its role in HIV infection has not been studied. Hence, we studied both CHRNA7 and CHRFAM7A regulation patterns in various gp120IIIB in vitro conditions. In addition, we measured CHRNA7 and CHRFAM7A expression levels in postmortem brain samples from patients suffering from different stages of HAND. Our results demonstrate the induction of CHRNA7 expression accompanied by a significant downregulation of CHRFAM7A in neuronal cells when exposed to pathophysiological concentrations of gp120IIIB. Our results suggest a dysregulation of CHRFAM7A and CHRNA7 expressions in the basal ganglia from postmortem brain samples of HIV+ subjects and expand the current knowledge about the consequences of HIV infection in the brain.

Lateral diffusion, function, and expression of the slow channel congenital myasthenia syndrome αC418W nicotinic receptor mutation with changes in lipid raft components.

Lipid rafts, specialized membrane microdomains in the plasma membrane rich in cholesterol and sphingolipids, are hot spots for a number of important cellular processes. The novel nicotinic acetylcholine receptor (nAChR) mutation αC418W, the first lipid-exposed mutation identified in a patient that causes slow channel congenital myasthenia syndrome was shown to be cholesterol-sensitive and to accumulate in microdomains rich in the membrane raft marker protein caveolin-1. The objective of this study is to gain insight into the mechanism by which lateral segregation into specialized raft membrane microdomains regulates the activable pool of nAChRs. We performed fluorescent recovery after photobleaching (FRAP), quantitative RT-PCR, and whole cell patch clamp recordings of GFP-encoding Mus musculus nAChRs transfected into HEK 293 cells to assess the role of cholesterol and caveolin-1 (CAV-1) in the diffusion, expression, and functionality of the nAChR (WT and αC418W). Our findings support the hypothesis that a cholesterol-sensitive nAChR might reside in specialized membrane microdomains that upon cholesterol depletion become disrupted and release the cholesterol-sensitive nAChRs to the pool of activable receptors. In addition, our results in HEK 293 cells show an interdependence between CAV-1 and αC418W that could confer end plates rich in αC418W nAChRs to a susceptibility to changes in cholesterol levels that could cause adverse drug reactions to cholesterol-lowering drugs such as statins. The current work suggests that the interplay between cholesterol and CAV-1 provides the molecular basis for modulating the function and dynamics of the cholesterol-sensitive αC418W nAChR.

Engineering α4ß2 nAChRs with reduced or increased nicotine sensitivity via selective disruption of consensus sites in the M3-M4 cytoplasmic loop of the α4 subunit.

The α4ß2 neuronal nicotinic acetylcholine receptor (nAChR) plays a crucial role in nicotine addiction. These receptors are known to desensitize and up-regulate after chronic nicotine exposure, but the mechanism remains unknown. Currently, the structure and functional role of the intracellular domains of the nAChR are obscure. To study the effect of subunit phosphorylation on α4ß2 nAChR function and expression, eleven residues located in the M3-M4 cytoplasmic loop were mutated to alanine and aspartic acid. Two-electrode voltage clamp and 125I-labeled epibatidine binding assays were performed on Xenopus oocytes to assess agonist activation and receptor expression. When ACh was used as an agonist, a decrease in receptor activation was observed for the majority of the mutations. When nicotine was used as an agonist, four mutations exhibited a statistically significant hypersensitivity to nicotine (S438D, S469A, Y576A, and S589A). Additionally, two mutations (S516D and T536A) that displayed normal activation with ACh displayed remarkable reductions in sensitivity to nicotine. Binding assays revealed a constitutive up-regulation in these two nicotine mutations with reduced nicotine sensitivity. These results suggest that consensus phosphorylation residues in the M3-M4 cytoplasmic loop of the α4 subunit play a crucial role in regulating α4ß2 nAChR agonist selectivity and functional expression. Furthermore, these results suggest that disruption of specific interactions at PKC putative consensus sites can render α4ß2 nAChRs almost insensitive to nicotine without substantial effects on normal AChR function. Therefore, these PKC consensus sites in the M3-M4 cytoplasmic loop of the α4 nAChR subunit could be a target for smoking cessation drugs.

Activation of the Macrophage α7 Nicotinic Acetylcholine Receptor and Control of Inflammation.

Inflammatory responses to stimuli are essential body defenses against foreign threats. However, uncontrolled inflammation may result in serious health problems, which can be life-threatening. The α7 nicotinic acetylcholine receptor, a ligand-gated ion channel expressed in the nervous and immune systems, has an essential role in the control of inflammation. Activation of the macrophage α7 receptor by acetylcholine, nicotine, or other agonists, selectively inhibits production of pro-inflammatory cytokines while leaving anti-inflammatory cytokines undisturbed. The neural control of this regulation pathway was discovered recently and it was named the cholinergic anti-inflammatory pathway (CAP). When afferent vagus nerve terminals are activated by cytokines or other pro-inflammatory stimuli, the message travels through the afferent vagus nerve, resulting in action potentials traveling down efferent vagus nerve fibers in a process that eventually leads to macrophage α7 activation by acetylcholine and inhibition of pro-inflammatory cytokines production. The mechanism by which activation of α7 in macrophages regulates pro-inflammatory responses is subject of intense research, and important insights have thus been made. The results suggest that activation of the macrophage α7 controls inflammation by inhibiting NF-κB nuclear translocation, and activating the JAK2/STAT3 pathway among other suggested pathways. While the α7 is well characterized as a ligand-gated ion channel in neurons, whole-cell patch clamp experiments suggest that α7's ion channel activity, defined as the translocation of ions across the membrane in response to ligands, is absent in leukocytes, and therefore, ion channel activity is generally assumed not to be required for the operation of the CAP. In this perspective, we briefly review macrophage α7 activation as it relates to the control of inflammation, and broaden the current view by providing single-channel currents as evidence that the α7 expressed in macrophages retains its ion translocation activity despite the absence of whole-cell currents. Whether this ion-translocating activity is relevant for the proper operation of the CAP or other important physiological processes remains obscure.

The α7-nicotinic receptor is upregulated in immune cells from HIV-seropositive women: consequences to the cholinergic anti-inflammatory response.

Antiretroviral therapy partially restores the immune system and markedly increases life expectancy of HIV-infected patients. However, antiretroviral therapy does not restore full health. These patients suffer from poorly understood chronic inflammation that causes a number of AIDS and non-AIDS complications. Here we show that chronic inflammation in HIV+ patients may be due to the disruption of the cholinergic anti-inflammatory pathway by HIV envelope protein gp120IIIB. Our results demonstrate that HIV gp120IIIB induces α7 nicotinic acetylcholine receptor (α7) upregulation and a paradoxical proinflammatory phenotype in macrophages, as activation of the upregulated α7 is no longer capable of inhibiting the release of proinflammatory cytokines. Our results demonstrate that disruption of the cholinergic-mediated anti-inflammatory response can result from an HIV protein. Collectively, these findings suggest that HIV tampering with a natural strategy to control inflammation could contribute to a crucial, unresolved problem of HIV infection: chronic inflammation.

Fluoxetine is neuroprotective in slow-channel congenital myasthenic syndrome.

The slow-channel congenital myasthenic syndrome (SCS) is an inherited neurodegenerative disease that caused mutations in the acetylcholine receptor (AChR) affecting neuromuscular transmission. Leaky AChRs lead to Ca(2+) overload and degeneration of the neuromuscular junction (NMJ) attributed to activation of cysteine proteases and apoptotic changes of synaptic nuclei. Here we use transgenic mouse models expressing two different mutations found in SCS to demonstrate that inhibition of prolonged opening of mutant AChRs using fluoxetine not only improves motor performance and neuromuscular transmission but also prevents Ca(2+) overload, the activation of cysteine proteases, calpain, caspase-3 and 9 at endplates, and as a consequence, reduces subsynaptic DNA damage at endplates, suggesting a long term benefit to therapy. These studies suggest that prolonged treatment of SCS patients with open ion channel blockers that preferentially block mutant AChRs is neuroprotective.

Excision of Elephantiasis Nostras Verrucosa Lesions in a Patient With Hereditary Lymphedema: Case Report and Review of the Literature.

Elephantiasis nostras verrucosa (ENV) is a rare cutaneous sequela of chronic lymphedema. Treatment of ENV remains poorly elucidated but has historically involved conservative management aimed at relieving the underlying lymphedema, with a few cases managed by surgical intervention. We report a case of a 27-year-old male with primary lymphedema complicated by large painful ENV lesions on his left foot that we excised surgically with good functional and cosmetic results as validated by the patient. To our knowledge, this is the first report of a case of ENV with a pedunculated morphology and the presence of a deep invasive stalk.

SorLA in glia: shared subcellular distribution patterns with caveolin-1.

SorLA is an established sorting and trafficking protein in neurons with demonstrated relevance to Alzheimer's disease (AD). It shares these roles with the caveolins, markers of membrane rafts microdomains. To further our knowledge on sorLA's expression and traffic, we studied sorLA expression in various cultured glia and its relation to caveolin-1 (cav-1), a caveolar microdomain marker. RT-PCR and immunoblots demonstrated sorLA expression in rat C6 glioma, primary cultures of rat astrocytes (PCRA), and human astrocytoma 1321N1 cells. PCRA were determined to express the highest levels of sorLA's message. Induction of differentiation of C6 cells into an astrocyte-like phenotype led to a significant decrease in sorLA's mRNA and protein expression. A set of complementary experimental approaches establish that sorLA and cav-1 directly or indirectly interact in glia: (1) co-fractionation in light-density membrane raft fractions of rat C6 glioma, PCRA, and human 1321N1 astrocytoma cells; (2) a subcellular co-localization distribution pattern in vesicular perinuclear compartments seen via confocal imaging in C6 and PCRA; (3) additional confocal analysis in C6 cells suggesting that the perinuclear compartments correspond to their co-localization in early endosomes and the trans-Golgi; and; (4) co-immunoprecipitation data strongly supporting their direct or indirect physical interaction. These findings further establish that sorLA is expressed in glia and that it shares its subcellular distribution pattern with cav-1. A direct or indirect cav-1/sorLA interaction could modify the trafficking and sorting functions of sorLA in glia and its proposed neuroprotective role in AD.

Effects of lipid-analog detergent solubilization on the functionality and lipidic cubic phase mobility of the Torpedo californica nicotinic acetylcholine receptor.

Over the past three decades, the Torpedo californica nicotinic acetylcholine receptor (nAChR) has been one of the most extensively studied membrane protein systems. However, the effects of detergent solubilization on nAChR stability and function are poorly understood. The use of lipid-analog detergents for nAChR solubilization has been shown to preserve receptor stability and functionality. The present study used lipid-analog detergents from phospholipid-analog and cholesterol-analog detergent families for solubilization and affinity purification of the receptor and probed nAChR ion channel function using planar lipid bilayers (PLBs) and stability using analytical size exclusion chromatography (A-SEC) in the detergent-solubilized state. We also examined receptor mobility on the lipidic cubic phase (LCP) by measuring the nAChR mobile fraction and diffusion coefficient through fluorescence recovery after photobleaching (FRAP) experiments using lipid-analog and non-lipid-analog detergents. Our results show that it is possible to isolate stable and functional nAChRs using lipid-analog detergents, with characteristic ion channel currents in PLBs and minimal aggregation as observed in A-SEC. Furthermore, fractional mobility and diffusion coefficient values observed in FRAP experiments were similar to the values observed for these parameters in the recently LCP-crystallized ß(2)-adrenergic receptor. The overall results show that phospholipid-analog detergents with 16 carbon acyl-chains support nAChR stability, functionality and LCP mobility.