VAMP2 regulates phase separation of α-synuclein.

α-Synuclein (αSYN), a pivotal synaptic protein implicated in synucleinopathies such as Parkinson's disease and Lewy body dementia, undergoes protein phase separation. We reveal that vesicle-associated membrane protein 2 (VAMP2) orchestrates αSYN phase separation both in vitro and in cells. Electrostatic interactions, specifically mediated by VAMP2 via its juxtamembrane domain and the αSYN C-terminal region, drive phase separation. Condensate formation is specific for R-SNARE VAMP2 and dependent on αSYN lipid membrane binding. Our results delineate a regulatory mechanism for αSYN phase separation in cells. Furthermore, we show that αSYN condensates sequester vesicles and attract complexin-1 and -2, thus supporting a role in synaptic physiology and pathophysiology.

Defining Normal Reference Range For The Cross Sectional Area Of The Median Nerve At The Wrist And Forearm Using High-Resolution Ultrasonography In Asymptomatic Pakistani Adults.

BACKGROUND: Carpal tunnel syndrome (CTS) can be diagnosed easily on ultrasonography (USG); which is a cheap, non-invasive and readily available modality. However, there is wide normal variation in the normal values of cross-sectional area (CSA) of median nerve among different populations; therefore, its necessary to establish a normal range of variability in median nerve dimensions in different populations. METHODS: A total of 500 asymptomatic patients i.e., 1000 median nerves were evaluated at the distal wrist crease and mid-forearm by 3 expert radiologists independently. All patients having a positive nerve conduction study or history of carpal tunnel syndrome and wrist trauma were excluded. Ultrasound was performed with a 7.5-15 MHz high-frequency linear probe. SPSS v 20 was used to analyze data. RESULTS: The study population had a mean age of 31.40±10.11 years with a female-to-male ratio of 1.36:1. Mean BMI was 22.15±4.34 Kg/m2 . The mean cross section area of the median nerve at the right wrist was calculated to be 6.8±1.96 mm2 and the left wrist was 6.6±1.96 mm2 . The mean median nerve cross-section area at the right mid-forearm was 5.3±1.46 mm2 and the left mid-forearm was 5.2±1.50 mm2 . A decrease in mean median nerve cross-section areas was noted by moving from wrist to forearm. Similarly, males showed higher median nerve CSA than females. CONCLUSIONS: Mean median nerve cross-section area was found to be different from Western countries. This warrants the utilization of the data of the Pakistani population to establish our own normal reference range for median nerve cross-sectional area to avoid misdiagnoses.

The Essential Role Of Conventional Radiography In Covid-19; Perspective Of A Developing Country.

BACKGROUND: The Coronavirus disease (COVID-19) pandemic has shaken the world. So far, CT has emerged as main stay of imaging whereas the local data on radiographic features of COVID-19 is sparse. METHODS: Prospective study includes 402 chest X rays (CXRs) of 105 patients presenting with symptoms of COVID-19. The nature of abnormality, distribution and lung zone involvement was documented. Following British Society of Thoracic Imaging (BSTI) guidelines, CXRs were grouped into classic/ probable COVID-19, indeterminate, non-COVID-19 and normal categories. The lung involvement was scored according to modified Radiographic Assessment of Lung Edema (RALE) scoring. The follow up radiographs were assessed for disease progression and improvement. RESULTS: Seventy-six males and 29 females with mean age of 50 years were included in our study. 47 out of 105 baseline radiographs were categorized as classic/ probable COVID-19, 26 as indeterminate, 7 as Non-COVID-19 and 25 as normal. 75 patients were positive and 30 were negative on RT-PCR testing. The sensitivity of CXR in diagnosing COVID-19 is 84%. The worsening radiographic features and higher RALE score correlates with longer hospital stay, ICU admissions and mortality. The ground glass opacities and consolidations in peripheral distribution involving bilateral mid and lower zones are the predominant findings of COVID-19 in Pakistani population. CONCLUSIONS: Combination of bilateral peripheral ground glass opacities and consolidations are the cardinal feature of COVID-19 on CXRs. The diagnostic categories described by BSTI correlates with PCR results in Pakistani population. The worsening radiographic findings correspond to poor prognosis; hence serial radiographs can be used for assessing disease course.

Non-diabetic ketoacidosis: a case series and literature review.

The genesis of ketone bodies by organisms is a protective mechanism. This metabolic process helps organisms to survive acute metabolic derangements in times of nutrient deficiency. When prolonged, ketogenesis leads to ketoacidosis, which is a potentially life-threatening metabolic disorder due to the accumulation of keto-acids in the body. The most common cause is diabetic ketoacidosis, though starvation ketoacidosis and alcoholic ketoacidosis are not uncommon. The presentation of all ketoacidotic states is similar-being generally unwell, abdominal pain, rapid and shallow breathing, vomiting and dehydration. Non-diabetic ketoacidotic states are very commonly overlooked due to relative unawareness among the clinicians, leading to misdiagnosis and thereby inappropriate management culminating in added mortality and morbidity. We describe here six cases of alcoholic and starvation ketoacidosis, review the literature currently available and discuss the common pitfalls in managing such cases.

Segment IV hepatic artery in potential donors for living related liver transplantation: Evaluation with multi-detector CT.

OBJECTIVE: To assess segment IV hepatic arterial anatomy and its variation on multi-detector computed tomography in potential liver donors. METHODS: The retrospective study was conducted at Shifa International Hospital, Islamabad and comprised data of potential liver transplant donors related to the period between January 2012 and June 2017. Computed tomography scans were performed using multi-detector scanners. Images were transferred to work station for postprocessing and were analysed regarding the origination and variation of the arteries by two independent experienced radiologists. RESULTS: Of the 455 patients whose records were evaluated, 299(65.7%) were males and 156(34.3%) were females. Six types of segment IV artery were defined based on their points of origin: left hepatic artery 285(62.6%), right hepatic artery 111(24.4 %), proper hepatic artery 9(1.8 %), common hepatic artery 29(6.4%), gastro duodenal artery 3(0.7 %), and dual 18(4.1 %).313 of total cases (68.8%) had normal anatomy with no variation. Those with aberrant/variant anatomy constituted 142(31.2%) of the total. CONCLUSIONS: Multi-detector computed tomography angiography was found to be a fast, reliable and non-invasive technique that could evaluate normal as well as anatomical variants of segment IV arteries.

Detection of Transition Zone in Bowel Obstruction via Curved Multiplanar Reformations with Multidetector Computed Tomography.

Objective We conducted this study to determine the added value of curved multiplanar reformations (CMPR) and multiplanar reformations (MPR) of multidetector computed tomography (MDCT) scan in the visualization and localization of the zone of transition in patients with intestinal obstruction. Materials and methods A total of 100 patients with suspected bowel obstruction were evaluated in a retrospective cross-sectional study from September 2016 to September 2018 at Dr. Ziauddin University Hospital, Clifton Campus. All patients underwent multidetector computed tomography (CT) scans with oral and intravenous contrast before surgical exploration. CMPR and MPR were acquired at the time of examination in each patient in addition to routine axial images. The CT scans were analyzed by two independent, experienced radiologists skilled at detecting the zones of transition in patients with bowel obstruction using the axial images alone, followed by axial images along with MPR, and then MPR plus CMPR. Patient data were masked to the radiologists. The CT scan findings were compared with surgical findings to determine the accuracy of CMPR in detecting the zone of transition between distended and collapsed bowel loops. The added CMPR showed high accuracy in the diagnosis of intestinal obstruction with a remarkable advantage over the conventional axial images. Data analysis was done on IBM SPSS Statistics for Windows, Version 20.0 (IBM Corp., Armonk, NY). Cohen's kappa statistics were obtained to show the measure of agreement between the two readers. McNemar's test was also applied to determine the homogeneity. Results Two radiologists, one with two years of experience and the other with five years of experience were 80% and 81% accurate, respectively, in identifying the zones of transition using axial images alone. Using axial images plus MPR, their accuracy was 88% and 92%, respectively. Using MPR plus CMPR, their accuracy was 96% and 98%, respectively. The accuracy of MPR plus CMPR views was significantly increased when compared to the accuracy using axial images alone. CT findings were compared to surgical findings in terms of diagnostic performance. The kappa value of 0.6 indicates moderate association and substantial agreement between two radiologists. McNemar's test showed homogeneity in the number of valid cases. Conclusion CMPR is an important and accurate technique for evaluating intestinal obstruction in addition to MPR as it helps in better localization of the zone of transition and in determining the cause of obstruction. This insight provides guidance for the appropriate treatment.

Tension Pneumoperitoneum: A Rare Presentation Of Megacystis Microcolon Intestinal Hypoperistalsis Syndrome.

Megacystis microcolon intestinal hypoperistalsis syndrome also known as Berdon syndrome is characterized by enlarged urinary bladder, small colon and reduced or absent intestinal peristalsis. We report a case of 4 days old female suffering from MMIHS presenting with tension pneumoperitoneum. To the best of our knowledge, this is the first reported case of MMIHS, having this unusual presentation.

eIF4A alleviates the translational repression mediated by classical secondary structures more than by G-quadruplexes.

Increased activity of the mRNA helicase eIF4A drives cellular malignancy by reprogramming cellular translation, and eIF4A activity is the direct or indirect target of many emerging cancer therapeutics. The enriched presence of (GGC)4 motifs, which have the potential to fold into two-layered G-quadruplexes, within the 5'UTRs of eIF4A-dependent mRNAs suggests that eIF4A is required for the unwinding of these structures within these eIF4A-controlled mRNAs. However, the existence of folded G-quadruplexes within cells remains controversial, and G-quadruplex folding is in direct competition with classical Watson-Crick based secondary structures. Using a combination of reverse transcription stalling assays and 7-deazaguanine incorporation experiments we find that (GGC)4 motifs preferentially form classical secondary structures rather than G-quadruplexes in full-length mRNAs. Furthermore, using translation assays with the eIF4A inhibitor hippuristanol, both in vitro and in cells, we find that eIF4A activity alleviates translational repression of mRNAs with 5'UTR classical secondary structures significantly more than those with folded G-quadruplexes. This was particularly evident in experiments using a G-quadruplex stabilizing ligand, where shifting the structural equilibrium in favour of G-quadruplex formation diminishes eIF4A-dependency. This suggests that enrichment of (GGC)4 motifs in the 5'UTRs of eIF4A-dependent mRNAs is due to the formation of stable hairpin structures rather than G-quadruplexes.

Phosphorylation Induces Structural Changes in the Autographa californica Nucleopolyhedrovirus P10 Protein.

Baculoviruses encode a variety of auxiliary proteins that are not essential for viral replication but provide them with a selective advantage in nature. P10 is a 10-kDa auxiliary protein produced in the very late phase of gene transcription by Autographa californica multiple nucleopolyhedrovirus (AcMNPV). The P10 protein forms cytoskeleton-like structures in the host cell that associate with microtubules varying from filamentous forms in the cytoplasm to aggregated perinuclear tubules that form a cage-like structure around the nucleus. These P10 structures may have a role in the release of occlusion bodies (OBs) and thus mediate the horizontal transmission of the virus between insect hosts. Here, using mass spectrometric analysis, it is demonstrated that the C terminus of P10 is phosphorylated during virus infection of cells in culture. Analysis of P10 mutants encoded by recombinant baculoviruses in which putative phosphorylation residues were mutated to alanine showed that serine 93 is a site of phosphorylation. Confocal microscopy examination of the serine 93 mutant structures revealed aberrant formation of the perinuclear tubules. Thus, the phosphorylation of serine 93 may induce the aggregation of filaments to form tubules. Together, these data suggest that the phosphorylation of serine 93 affects the structural conformation of P10.IMPORTANCE The baculovirus P10 protein has been researched intensively since it was first observed in 1969, but its role during viral infection remains unclear. It is conserved in the alphabaculoviruses and expressed at high levels during virus infection. Producing large amounts of a protein is wasteful for the virus unless it is advantageous for the survival of its progeny, and therefore, P10 presents an enigma. As P10 polymerizes to form organized cytoskeletal structures that colocalize with host cell microtubules, the structural relationship of the protein with the host cell may present a key to help understand the function and importance of this protein. This study addresses the importance of the structural changes in P10 during infection and how they may be governed by phosphorylation. The P10 structures affected by phosphorylation are closely associated with the viral progeny and thus may potentially be responsible for its dissemination and survival.

Translational dysregulation in cancer: eIF4A isoforms and sequence determinants of eIF4A dependence.

The malignant phenotype is largely the consequence of dysregulated gene expression. Transformed cells depend upon not just a global increase in protein synthesis but an altered translational landscape in which pro-oncogenic mRNAs are translationally up-regulated. Such mRNAs have been shown to possess longer and more structured 5'-UTRs requiring high levels of eukaryotic initiation factor 4A (eIF4A) helicase activity for efficient translation. As such there is a developing focus on targeting eIF4A as a cancer therapy. In order for such treatments to be successful, we must develop a detailed understanding of the mechanisms which make specific mRNAs more dependent on eIF4A activity than others. It is also crucial to fully characterize the potentially distinct roles of eIF4A1 and eIF4A2, which until recently were thought to be functionally interchangeable. This review will highlight the recent advances made in this field that address these issues.

Activin-like kinase 5 (ALK5) mediates abnormal proliferation of vascular smooth muscle cells from patients with familial pulmonary arterial hypertension and is involved in the progression of experimental pulmonary arterial hypertension induced by monocrotaline.

Mutations in the gene for the transforming growth factor (TGF)-beta superfamily receptor, bone morphogenetic protein receptor II, underlie heritable forms of pulmonary arterial hypertension (PAH). Aberrant signaling via TGF-beta receptor I/activin receptor-like kinase 5 may be important for both the development and progression of PAH. We investigated the therapeutic potential of a well-characterized and potent activin receptor-like kinase 5 inhibitor, SB525334 [6-(2-tert-butyl-5-{6-methyl-pyridin-2-yl}-1H-imidazol-4-yl)-quinoxaline] for the treatment of PAH. In this study, we demonstrate that pulmonary artery smooth muscle cells from patients with familial forms of idiopathic PAH exhibit heightened sensitivity to TGF-beta1 in vitro, which can be attenuated after the administration of SB525334. We further demonstrate that SB525334 significantly reverses pulmonary arterial pressure and inhibits right ventricular hypertrophy in a rat model of PAH. Immunohistochemical studies confirmed a significant reduction in pulmonary arteriole muscularization induced by monocrotaline (used experimentally to induce PAH) after treatment of rats with SB525334. Collectively, these data are consistent with a role for the activin receptor-like kinase 5 in the progression of idiopathic PAH and imply that strategies to inhibit activin receptor-like kinase 5 signaling may have therapeutic benefit.