Alpha-mannosidosis: a case with novel ultrastructural and light microscopy findings.

OBJECTIVES: Alpha-mannosidosis is a rare genetic lysosomal storage condition leading to the systemic buildup of oligomannoside. Clinical presentation and associated conditions, as well as the full extent of histopathologic changes associated with this disease process, are not fully understood. CASE PRESENTATION: We present the case of an 8-year-1-month old patient with persistent anemia and who was initially diagnosed with Celiac disease before ultimately being diagnosed with alpha-mannosidosis. As part of his diagnostic work-up, duodenal and bone marrow biopsies were examined by pathology. Duodenal biopsies showed foamy plasma cells expanding the lamina propria which triggered a workup for a genetic storage disease; features suggestive of Celiac disease which resolved on gluten-free diet were also noted by pathology. Bone marrow analysis via electron microscopy showed cytoplasmic granules and inclusions in multiple immune cell lines. CONCLUSIONS: Alpha-mannosidosis can occur with Celiac disease and milder forms may only be suspected from incidental pathology findings. The ultrastructural bone marrow findings from this case, the first to be reported from human, show numerous disease-associated changes in multiple immune cell lines whose contribution to disease-associated immunodeficiency is unclear.

The Genetic Landscape of Fibroepithelial Lesions of the Breast.

Fibroepithelial lesions of the breast encompass a broad spectrum of lesions from fibroadenomas and their variants to phyllodes tumors, including their clinical range of benign, borderline, and malignant. Classification of this spectrum of neoplasms has historically and currently been based purely on morphology, although the nomenclature has shifted over the years largely due to the significant histologic overlap that exists primarily within the cellular fibroadenomas to borderline malignant phyllodes tumor categories. A review of the current diagnostic challenge, proposed ancillary studied and their value in prognostic significance, is provided. This article highlights the most recent molecular and genetic findings as well as the limitations of the studies, in the context of practical and available applications for the diagnostician and managerial implications for the clinician.

Gene Expression Profiling of Fibroepithelial Lesions of the Breast.

Fibroepithelial lesions of the breast (FELs) are a heterogeneous group of neoplasms exhibiting a histologic spectrum ranging from fibroadenomas (FAs) to malignant phyllodes tumors (PTs). Despite published histologic criteria for their classification, it is common for such lesions to exhibit overlapping features, leading to subjective interpretation and interobserver disagreements in histologic diagnosis. Therefore, there is a need for a more objective diagnostic modality to aid in the accurate classification of these lesions and to guide appropriate clinical management. In this study, the expression of 750 tumor-related genes was measured in a cohort of 34 FELs (5 FAs, 9 cellular FAs, 9 benign PTs, 7 borderline PTs, and 4 malignant PTs). Differentially expressed gene analysis, gene set analysis, pathway analysis, and cell type analysis were performed. Genes involved in matrix remodeling and metastasis (e.g., MMP9, SPP1, COL11A1), angiogenesis (VEGFA, ITGAV, NFIL3, FDFR1, CCND2), hypoxia (ENO1, HK1, CYBB, HK2), metabolic stress (e.g., UBE2C, CDKN2A, FBP1), cell proliferation (e.g., CENPF, CCNB1), and the PI3K-Akt pathway (e.g., ITGB3, NRAS) were highly expressed in malignant PTs and less expressed in borderline PTs, benign PTs, cellular FAs, and FAs. The overall gene expression profiles of benign PTs, cellular FAs, and FAs were very similar. Although a slight difference was observed between borderline and benign PTs, a higher degree of difference was observed between borderline and malignant PTs. Additionally, the macrophage cell abundance scores and CCL5 were significantly higher in malignant PTs compared with all other groups. Our results suggest that the gene-expression-profiling-based approach could lead to further stratification of FELs and may provide clinically useful biological and pathophysiological information to improve the existing histologic diagnostic algorithm.

Pocket ACEs: Discovering new function within an old player.

Angiotensin-converting enzyme (ACE) is canonically known for its role in the renin-angiotensin system (RAS) where its conversion of angiotensin I (Ang I) to the bioactive peptide angiotensin II (Ang II) helps to regulate blood pressure, electrolyte, and volume homeostasis. Further studies on ACE have shown that its enzymatic activity is relatively non-specific and functions outside of the RAS axis. Of the multiple systems it has been implicated in, ACE has been found to play an important role in the development and modulation of hematopoiesis and the immune system, both through the RAS and independently of the RAS axis.

Comparative Upper Respiratory Tract Transcriptomic Profiling Reveals a Potential Role of Early Activation of Interferon Pathway in Severe COVID-19.

Infection with SARS-CoV-2 results in Coronavirus disease 2019 (COVID-19) is known to cause mild to acute respiratory infection and sometimes progress towards respiratory failure and death. The mechanisms driving the progression of the disease and accumulation of high viral load in the lungs without initial symptoms remain elusive. In this study, we evaluated the upper respiratory tract host transcriptional response in COVID-19 patients with mild to severe symptoms and compared it with the control COVID-19 negative group using RNA-sequencing (RNA-Seq). Our results reveal an upregulated early type I interferon response in severe COVID-19 patients as compared to mild or negative COVID-19 patients. Moreover, severely symptomatic patients have pronounced induction of interferon stimulated genes (ISGs), particularly the oligoadenylate synthetase (OAS) family of genes. Our results are in concurrence with other studies depicting the early induction of IFN-I response in severe COVID-19 patients, providing novel insights about the ISGs involved.

Amyloid Deposition: An Unusual Case of Deep Gluteal Syndrome and Sciatic Nerve Compression: A Case Report.

CASE: Soft-tissue amyloidomas are exceedingly rare, with only a few cases reported in the literature. There are no reports of sciatic nerve compression secondary to a soft-tissue amyloidoma. We report a unique case of a 71-year-old man with an incidentally found amyloidoma who was initially believed to have deep gluteal syndrome. He had a favorable outcome after surgical decompression. CONCLUSION: For patients who do not have classic examination and electromyography/nerve conduction findings of piriformis syndrome, providers should explore other etiologies of peripheral nerve compression including soft-tissue amyloidoma.

Temperature rhythms and ICU sleep: the TRIS study.

BACKGROUND: Core body temperature (CBT) patterns associated with sleep have not been described in the critically ill. This study aimed to characterize night-time sleep and its relationship to CBT in ICU patients. METHODS: A prospective study was performed in a 27-bed tertiary adult intensive care unit of 20 mechanically ventilated patients in the weaning stage of their critical illness. The study assessed sleep by polysomnography (PSG) during the evening between 21:00-7:00 hours, nursing interventions using the Therapeutic Intervention Scoring System (TISS), illness severity using SOFA and APACHE II scores and CBT 24-hour pattern. RESULTS: Patients were awake for approximately half the study period (45.04%, IQR 13.81-77-17) with no REM (0%, IQR 0-0.04%) and median arousals of 19.5/hour (IQR 7.1-40.9). The 24-hour CBT had a rhythmic pattern in 13 (65%) patients with a highly variable phase of median peak time at 17:35 hours (IQR 12:40-19:39). No significant associations were found between CBT rhythmicity, sleep stages, sleep EEG frequency density, illness severity scores or TISS on the day of PSG. There was no relationship between time awake and CBT rhythmicity (P=0.48) or CBT peak time (P=0.82). The relationship between circadian rhythms and sleep patterns in the critically ill is complex. CONCLUSIONS: Patients recovering in ICU commonly have CBT loss of rhythmicity or a significant phase shift with loss of normal night-time patterns of sleep architecture. Appropriate care plans to promote sleep and circadian rhythm require further investigation of contributing factors such as environment, clinical care routines, illness type and severity.

The plant pathogen enzyme AldC is a long-chain aliphatic aldehyde dehydrogenase.

Aldehyde dehydrogenases are versatile enzymes that serve a range of biochemical functions. Although traditionally considered metabolic housekeeping enzymes because of their ability to detoxify reactive aldehydes, like those generated from lipid peroxidation damage, the contributions of these enzymes to other biological processes are widespread. For example, the plant pathogen Pseudomonas syringae strain PtoDC3000 uses an indole-3-acetaldehyde dehydrogenase to synthesize the phytohormone indole-3-acetic acid to elude host responses. Here we investigate the biochemical function of AldC from PtoDC3000. Analysis of the substrate profile of AldC suggests that this enzyme functions as a long-chain aliphatic aldehyde dehydrogenase. The 2.5 Å resolution X-ray crystal of the AldC C291A mutant in a dead-end complex with octanal and NAD+ reveals an apolar binding site primed for aliphatic aldehyde substrate recognition. Functional characterization of site-directed mutants targeting the substrate- and NAD(H)-binding sites identifies key residues in the active site for ligand interactions, including those in the "aromatic box" that define the aldehyde-binding site. Overall, this study provides molecular insight for understanding the evolution of the prokaryotic aldehyde dehydrogenase superfamily and their diversity of function.

Novel Immunoglobulin Domain Proteins Provide Insights into Evolution and Pathogenesis Mechanisms of SARS-Related Coronaviruses.

A novel coronavirus (SARS-CoV-2) is the causative agent of an emergent severe respiratory disease (COVID-19) in humans that is threatening to result in a global health crisis. By using genomic, sequence, structural and evolutionary analysis, we show that Alpha- and Beta-CoVs possess several novel families of immunoglobulin (Ig) domain proteins, including ORF8 and ORF7a from SARS-related coronaviruses and two protein groups from certain Alpha-CoVs. Among them, ORF8 is distinguished in being rapidly evolving, possessing a unique insert and a hypervariable position among SARS-CoV-2 genomes in its predicted ligand-binding groove. We also uncover many Ig proteins from several metazoan viruses which are distinct in sequence and structure but share an architecture comparable to that of CoV Ig domain proteins. Hence, we propose that deployment of Ig domain proteins is a widely-used strategy by viruses, and SARS-CoV-2 ORF8 is a potential pathogenicity factor which evolves rapidly to counter the immune response and facilitate the transmission between hosts.

Novel Immunoglobulin Domain Proteins Provide Insights into Evolution and Pathogenesis of SARS-CoV-2-Related Viruses.

A novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was recently identified as the causative agent for the coronavirus disease 2019 (COVID-19) outbreak that has generated a global health crisis. We use a combination of genomic analysis and sensitive profile-based sequence and structure analysis to understand the potential pathogenesis determinants of this virus. As a result, we identify several fast-evolving genomic regions that might be at the interface of virus-host interactions, corresponding to the receptor binding domain of the Spike protein, the three tandem Macro fold domains in ORF1a, and the uncharacterized protein ORF8. Further, we show that ORF8 and several other proteins from alpha- and beta-CoVs belong to novel families of immunoglobulin (Ig) proteins. Among them, ORF8 is distinguished by being rapidly evolving, possessing a unique insert, and having a hypervariable position among SARS-CoV-2 genomes in its predicted ligand-binding groove. We also uncover numerous Ig domain proteins from several unrelated metazoan viruses, which are distinct in sequence and structure but share comparable architectures to those of the CoV Ig domain proteins. Hence, we propose that SARS-CoV-2 ORF8 and other previously unidentified CoV Ig domain proteins fall under the umbrella of a widespread strategy of deployment of Ig domain proteins in animal viruses as pathogenicity factors that modulate host immunity. The rapid evolution of the ORF8 Ig domain proteins points to a potential evolutionary arms race between viruses and hosts, likely arising from immune pressure, and suggests a role in transmission between distinct host species.IMPORTANCE The ongoing COVID-19 pandemic strongly emphasizes the need for a more complete understanding of the biology and pathogenesis of its causative agent SARS-CoV-2. Despite intense scrutiny, several proteins encoded by the genomes of SARS-CoV-2 and other SARS-like coronaviruses remain enigmatic. Moreover, the high infectivity and severity of SARS-CoV-2 in certain individuals make wet-lab studies currently challenging. In this study, we used a series of computational strategies to identify several fast-evolving regions of SARS-CoV-2 proteins which are potentially under host immune pressure. Most notably, the hitherto-uncharacterized protein encoded by ORF8 is one of them. Using sensitive sequence and structural analysis methods, we show that ORF8 and several other proteins from alpha- and beta-coronavirus comprise novel families of immunoglobulin domain proteins, which might function as potential immune modulators to delay or attenuate the host immune response against the viruses.

Use of a limited-channel device for obstructive sleep apnoea diagnosis in a tertiary sleep disorders centre.

BACKGROUND: A major impediment to the provision of obstructive sleep apnoea (OSA) treatment is reliance on labour-intensive and costly laboratory-based polysomnography (PSG). AIMS: To investigate if measurement of oximetry and nasal flow through the ApneaLink device (AL) could identify patients with moderate-severe OSA among those referred for PSG to a tertiary sleep service. METHODS: New referrals to The Prince Charles Hospital Sleep Disorders Centre were assessed for suitability. Demographics, anthropometrics, Epworth Sleepiness and OSA50 scores were collected. Exclusion criteria included age <18 years, pregnancy, significant cognitive impairment, poorly controlled psychiatric disorder, domiciliary oxygen and prior OSA treatment. Participants underwent concurrent type 1 PSG and AL assessments. RESULTS: One hundred participants had a mean age of 55 years (standard deviation 17) and were 49% male. Forty-eight (48%) had moderate-severe OSA on PSG. Composite variable AL 3% oxygen desaturation index ≥16 and AL apnoea-hypopnoea index (AHI) ≥15 had receiver operator characteristic area under the curve of 0.87, sensitivity of 80% and specificity of 94% for PSG AHI ≥15. The three false-positives seen with this composite variable had PSG AHI 11-14 and Epworth Sleepiness Score 6-17. The various composites of AL, anthropometric and questionnaire variables did not improve the AUC or specificity but did improve sensitivity. CONCLUSIONS: AL is useful in the diagnosis of moderate-severe OSA in patients referred to a tertiary sleep disorders centre. This could lessen reliance on PSG, expedite OSA care, lead to significant cost savings and make diagnosis of OSA more available in non-urban areas.

Modulating Responses of Toehold Switches by an Inhibitory Hairpin.

The toehold switch consists of a cis-repressing switch RNA hairpin and a trans-acting trigger RNA. The binding of the trigger RNA to an unpaired toehold sequence of the switch hairpin allows for a branch migration process, exposing the start codon and ribosome binding site for translation initiation. In this work, we demonstrate that responses of toehold switches can be modulated by introducing an inhibitory hairpin that shortens the length of the unpaired toehold region. First, we investigated the effect of the toehold region length on output gene expression and showed that the second trigger RNA, which binds to the inhibitory hairpin, is necessary for output gene activation when the hairpin-to-hairpin spacing is short. Second, the apparent Hill coefficient was found generally to increase with decreasing hairpin-to-hairpin spacing or increasing hairpin number. This work expands the utility of toehold switches by providing a new way to modulate their response.

Acute esophageal necrosis: a case series and its management.

'Black Esophagus' or acute esophageal syndrome (AEN) is extremely rare entity characterized by patchy or diffuse circumferential necrosis of esophagus. To date, there is minimal number of cases described with this phenomenon. The main objective of this case series is to discuss the diagnosis of extremely rare condition seen in two patients within 1 year and its effective management by adopting conservative measures in a tertiary hospital. The objective of our case series is to discuss the diagnosis of an extremely rare condition seen in two patients within a short span of 6 months and its effective management by adopting conservative measures in a tertiary care hospital. An overall conservative approach, serial endoscopic monitoring and effective follow-up resulted in zero mortality with successful outcomes.

Massive symptomatic right-sided Bochdalek hernia in an adult man.

A 22-year-old man presented to a rural hospital in Australia with right-sided pleuritic chest pain, right shoulder pain and dyspnoea. The patient had been receiving chronic asthma therapy without improvement. CT of the chest was performed after an abnormal X-ray, incidentally revealing one of the largest documented right-sided diaphragmatic hernias, with left lung compression due to mediastinal shift. The patient was definitively managed with thoracotomy alone. The contents of the hernia sac included colon and multiple loops of small bowel with a 10 cm neck. Definitive treatment was achieved with significant reduction in hernia size and formation of a neo-diaphragm with composite mesh. The postoperative period was complicated only by a wound infection. Two weeks after discharge the patient remained clinically well. Repeat chest X-ray showed no recurrence of the hernia. Congenital diaphragmatic hernias should be considered in patients with ongoing respiratory symptoms. Thoracotomy provides a safe approach.

Programmable control of bacterial gene expression with the combined CRISPR and antisense RNA system.

A central goal of synthetic biology is to implement diverse cellular functions by predictably controlling gene expression. Though research has focused more on protein regulators than RNA regulators, recent advances in our understanding of RNA folding and functions have motivated the use of RNA regulators. RNA regulators provide an advantage because they are easier to design and engineer than protein regulators, potentially have a lower burden on the cell and are highly orthogonal. Here, we combine the CRISPR system from Streptococcus pyogenes and synthetic antisense RNAs (asRNAs) in Escherichia coli strains to repress or derepress a target gene in a programmable manner. Specifically, we demonstrate for the first time that the gene target repressed by the CRISPR system can be derepressed by expressing an asRNA that sequesters a small guide RNA (sgRNA). Furthermore, we demonstrate that tunable levels of derepression can be achieved (up to 95%) by designing asRNAs that target different regions of a sgRNA and by altering the hybridization free energy of the sgRNA-asRNA complex. This new system, which we call the combined CRISPR and asRNA system, can be used to reversibly repress or derepress multiple target genes simultaneously, allowing for rational reprogramming of cellular functions.

Oesophagectomy is a safe option for early adenocarcinoma arising from Barrett's oesophagus.

BACKGROUND: Over the last decade, there has been a shift towards endoscopic treatment of high-grade dysplasia (HGD) and T1 stage adenocarcinoma arising in Barrett's oesophagus. Although short-term outcomes are promising, longer-term outcomes remain uncertain and the role of these therapies versus surgery is debated, with surgical mortality rates assumed. However, few studies have specifically determined the outcome for oesophagectomy in the subgroup with HGD or T1 adenocarcinoma. To determine this, we evaluated experience with oesophagectomy for HGD and T1 adenocarcinoma in Barrett's oesophagus. METHODS: Data were analysed from a prospective audit database for oesophagectomy performed at two public and four associated private hospitals in Adelaide, South Australia. Patients with HGD, T1a and T1b adenocarcinoma who underwent oesophagectomy from 20 February 1998 to 17 February 2012 were identified, and their perioperative, post-operative and survival outcomes were determined. RESULTS: From 452 oesophagectomy procedures, 63 (13.9%) individuals who underwent surgery for HGD or T1 adenocarcinoma were identified; HGD - 19 (30.1%), T1a - 18 (28.5 %), T1b - 26 (41.3%). Major complications occurred in eight (12.7%) patients including one (1.6%) death following surgery. Five-year survival for HGD and T1a cancers using Kaplan-Meier analysis was not significantly different from a matched general population without cancer. CONCLUSION: Oesophagectomy for HGD and T1 stage adenocarcinoma in Barrett's oesophagus is associated with favourable outcomes. Outcomes following endoscopic treatments should be benchmarked against these outcomes, not those following oesophagectomy for advanced cancer.

Measuring leg movements during sleep using accelerometry: comparison with EMG and piezo-electric scored events.

Periodic Limb Movements during Sleep (PLMS) can cause significant disturbance to sleep, resulting in daytime sleepiness and reduced quality of life. In conventional clinical practice, PLMS are measured using overnight electromyogram (EMG) of the tibialis anterior muscle, although historically they have also been measured using piezo-electric gauges placed over the muscle. However, PLMS counts (PLM index) do not correlate well with clinical symptomology. In this study, we propose that because EMG and piezo derived signals measure muscle activation rather than actual movement, they may count events with no appreciable movement of the limb and therefore no contribution to sleep disturbance. The aim of this study is thus to determine the percentage of clinically scored limb movements which are not associated with movement of the great toe measured using accelerometry. 9 participants were studied simultaneously with an overnight diagnostic polysomnogram (including EMG and piezo instrumentation of the right leg) and high temporal resolution accelerometry of the right great toe. Limb movements were scored, and peak acceleration during each scored movement was quantified. Across the participant population, 54.9% (range: 26.7-76.3) and 39.0% (range: 4.8-69.6) of limb movements scored using piezo and EMG instrumentation respectively, were not associated with toe movement measured with accelerometry. If sleep disturbance is the consequence of the limb movements, these results may explain why conventional piezo or EMG derived PLMI is poorly correlated with clinical symptomology.