IGF-2 mediated hypoglycemia and the paradox of an apparently benign lesion: a case report & review of the literature.

BACKGROUND: Non-islet cell tumour hypoglycemia (NICTH) is rarely encountered in clinical practice. Insulin-like growth factor 2 (IGF2) is the most common cause of NICTH observed in the setting of mesenchymal and epithelial neoplasia. This is a paraneoplastic syndrome caused by IGF2 activation of the insulin receptor. CASE PRESENTATION: An 80 year old female presented with a short history of recurrent episodes of confusion with laboratory confirmed hypoglycemia with a plasma glucose of 2.7 mmol/L on fasting which fulfilled Whipple's triad. Diagnostic clues to the aetiology at presentation include the fasting pattern of hypoglycemia, hypokalaemia and the absence of weight gain. A 72 hour fast with results showed early hypoglycemia and suppression of serum insulin, c-peptide, and proinsulin. Serum insulin antibody was not detected. Subsequent measurement of the serum IGF2:IGF1 ratio was elevated at 22.3 and consistent with IGF-2 mediated hypoglycemia and imaging studies demonstrated a pelvic mass. Dietary intervention and oral prednisolone abated hypoglycemia prior to surgery. Ultimately, hypoglycemia resolved following operative intervention and steroid therapy was successfully withdrawn. Histopathology was remarkable for dual neoplastic processes with uterine solitary fibrous tumour (SFT) confirmed as the source of IGF2 hypersecretion on IGF-2 immunohistochemistry and a coincidental invasive high grade serous carcinoma involving the fimbria of the right fallopian tube. CONCLUSION: The paradox in this case is that the benign solitary fibrous tumour accounted for patient morbidity through secretion of IGF2 and without treatment, posed a mortality risk. This is despite the synchronous presence of a highly malignant fallopian tube neoplasm. This case reinforces the need for thorough clinical evaluation of hypoglycemia to allow prompt and definitive management.

A Multicenter External Validation of a Score Model to Predict Risk of Events in Patients With Brugada Syndrome.

A multivariate risk score model was proposed by Sieira et al in 2017 for sudden death in Brugada syndrome; their validation in 150 patients was highly encouraging, with a C-index of 0.81; however, this score is yet to be validated by an independent group. A total of 192 records of patients with Brugada syndrome were collected from 2 centers in the United Kingdom and retrospectively scored according to a score model by Sieira et al. Data were compiled summatively over follow-up to mimic regular risk re-evaluation as per current guidelines. Sudden cardiac death survivor data were considered perievent to ascertain the utility of the score before cardiac arrest. Scores were compared with actual outcomes. Sensitivity in our cohort was 22.7%, specificity was 57.6%, and C-index was 0.58. In conclusion, up to 75% of cardiac arrest survivors in this cohort would not have been offered a defibrillator if evaluated before their event. This casts doubt on the utility of the score model for primary prevention of sudden death. Inherent issues with modern risk scoring strategies decrease the likelihood of success even in robustly designed tools such as the Sieira score model.

Arrhythmia in an athlete diagnosed by smartphone electrocardiogram: a case report.

BACKGROUND: This is the first case report demonstrating the use of a smartphone device, enabling the diagnosis of an arrhythmia in the sports cardiology literature. CASE SUMMARY: A 17-year-old semi-professional rugby player presented with recurrent episodes of palpitations terminated by vagal manoeuvres. The rugby player's resting 12-lead electrocardiogram (ECG), echocardiogram, and exercise stress test were normal. Due to his suggestive history and an ECG trace from a smartphone device, demonstrating a narrow complex tachycardia, an electrophysiological study was arranged. The study demonstrated a slow-fast atrioventricular nodal re-entrant tachycardia which was successfully ablated. DISCUSSION: The ambulatory use of a smartphone ECG device assisted in the timely diagnosis and management of an undiagnosed paroxysmal arrhythmia in a rugby player. This resulted in an expedited return to play.

The indications and safety of prolonged temporary pacing using active-fixation leads and externalized pulse generator.

BACKGROUND: Temporary cardiac pacing, conventionally achieved using a passive transvenous pacing wire, can be life-saving for unstable arrhythmias. However, they run the risk of complications, the longer they remain in-situ. Externalized prolonged temporary pacing (EPTP), using active-fixation lead and an externalized pulse generator; may be an alternative for transient pacing indications, concurrent illness or sepsis that precludes permanent pacing. METHODS: Sixty-seven patients (mean age 69 ± 14 years; 82% male) underwent EPTP between November 2011 and April 2019. EPTP was performed in a sterile facility, under fluoroscopy, using active-fixation leads anchored to the right ventricle septum. Externalized lead was connected to a re-sterilized pulse generator and secured to anterior chest wall with transparent dressings. EPTP indications and patient outcomes were evaluated. RESULTS: Pacing indications were high-grade atrio-ventricular (AV) block (73.2%), sinus arrest (14.9%), overdrive suppression of VT (5.9%) and pause-dependent VT (4.5%). Reasons for ETPT rather than permanent pacing included: sepsis (38.8%), CIED-related infection (8.9%), transient pacing indication (25%), to allow further investigations prior to decision on CIED type (22%), and over-drive arrhythmia suppression (6%). Sixty three percent patients were severely ill in an ICU. Mean duration of pacing was 16 ± 12 days. Sixty seven percent patients subsequently received a CIED and had no evidence of device-related infection at 1-year post-implant. There were three non-fatal complications during EPTP while no deaths were attributed to EPTP. CONCLUSION: EPTP is a safe and useful method of prolonged temporary pacing for patients who require chronotropic support, but in whom immediate permanent pacemaker implantation is contraindicated.

High-resolution three-dimensional chromatin profiling of the Chinese hamster ovary cell genome.

Chinese hamster ovary (CHO) cell lines are the pillars of a multibillion-dollar biopharmaceutical industry producing recombinant therapeutic proteins. The effects of local chromatin organization and epigenetic repression within these cell lines result in unpredictable and unstable transgene expression following random integration. Limited knowledge of the CHO genome and its higher order chromatin organization has thus far impeded functional genomics approaches required to tackle these issues. Here, we present an integrative three-dimensional (3D) map of genome organization within the CHOK1SV® 10E9 cell line in conjunction with an improved, less fragmented CHOK1SV 10E9 genome assembly. Using our high-resolution chromatin conformation datasets, we have assigned ≈90% of sequence to a chromosome-scale genome assembly. Our genome-wide 3D map identifies higher order chromatin structures such as topologically associated domains, incorporates our chromatin accessibility data to enhance the identification of active cis-regulatory elements, and importantly links these cis-regulatory elements to target promoters in a 3D promoter interactome. We demonstrate the power of our improved functional annotation by evaluating the 3D landscape of a transgene integration site and two phenotypically different cell lines. Our work opens up further novel genome engineering targets, has the potential to inform vital improvements for industrial biotherapeutic production, and represents a significant advancement for CHO cell line development.

Altered cerebrovascular response to acute exercise in patients with Huntington's disease.

The objective of this study was to determine whether a single session of exercise was sufficient to induce cerebral adaptations in individuals with Huntington's disease and to explore the time dynamics of any acute cerebrovascular response. In this case-control study, we employed arterial-spin labelling MRI in 19 Huntington's disease gene-positive participants (32-65 years, 13 males) and 19 controls (29-63 years, 10 males) matched for age, gender, body mass index and self-reported activity levels, to measure global and regional perfusion in response to 20 min of moderate-intensity cycling. Cerebral perfusion was measured at baseline and 15, 40 and 60 min after exercise cessation. Relative to baseline, we found that cerebral perfusion increased in patients with Huntington's disease yet was unchanged in control participants in the precentral gyrus (P = 0.016), middle frontal gyrus (P = 0.046) and hippocampus (P = 0.048) 40 min after exercise cessation (+15 to +32.5% change in Huntington's disease participants, -7.7 to 0.8% change in controls). The length of the disease-causing trinucleotide repeat expansion in the huntingtin gene predicted the change in the precentral gyrus (P = 0.03) and the intensity of the exercise intervention predicted hippocampal perfusion change in Huntington's disease participants (P < 0.001). In both groups, exercise increased hippocampal blood flow 60 min after exercise cessation (P = 0.039). These findings demonstrate the utility of acute exercise as a clinically sensitive experimental paradigm to modulate the cerebrovasculature. Twenty minutes of aerobic exercise induced transient cerebrovascular adaptations in the hippocampus and cortex selectively in Huntington's disease participants and likely represents latent neuropathology not evident at rest.

Availability of automated external defibrillators in Hamilton, New Zealand.

BACKGROUND: Last year, there were 2,000 out-of-hospital cardiac arrests (OHCA) in New Zealand, 74% received CPR but only 5.1% accessed an automated external defibrillator (AED). The average survival rate of OHCA is 13%. The aim of this study was to visit all 50 AED locations shown on www.hamiltoncentral.co.nz to assess their true availability and visibility to the public in the event of an OHCA. METHOD: All premises were visited and the first staff member encountered was asked if they were aware an AED was onsite, its location, hours of availability, if restricted access applied and whether it had been used. RESULTS: Of the 50 locations, three sites no longer exist and two AEDs were listed twice. Therefore, only 45 AEDs exist. Two sites had grossly inaccurate locations. Three AEDs (7%) were continuously available. Nine AEDs were accessible after 6pm at least one day of the week. Thirteen AEDs were available on weekends; however, five required swipe card access. None of the AEDs were located outdoors. CONCLUSION: Far fewer than 50 listed AEDs are freely available to the public, especially after 6pm and on weekends. Lack of signposting and restrictions to access would lead to delayed defibrillation. This important health issue needs addressing.

Diversity in host clone performance within a Chinese hamster ovary cell line.

Much effort has been expended to improve the capabilities of individual Chinese hamster ovary (CHO) host cell lines to synthesize recombinant therapeutic proteins (rPs). However, given the increasing variety in rP molecular types and formats it may be advantageous to employ a toolbox of CHO host cell lines in biomanufacturing. Such a toolbox would contain a panel of hosts with specific capabilities to synthesize certain molecular types at high volumetric concentrations and with the correct product quality (PQ). In this work, we examine a panel of clonally derived host cell lines isolated from CHOK1SV for the ability to manufacture two model proteins, an IgG4 monoclonal antibody (Mab) and an Fc-fusion protein (etanercept). We show that these host cell lines vary in their relative ability to synthesize these proteins in transient and stable pool production format. Furthermore, we examined the PQ attributes of the stable pool-produced Mab and etanercept (by N-glycan ultra performance liquid chromatography (UPLC) and liquid chromatography - tandem mass spectrometry (LC-MS/MS), respectively), and uncovered substantial variation between the host cell lines in Mab N-glycan micro-heterogeneity and etanercept N and O-linked macro-heterogeneity. To further investigate the capabilities of these hosts to act as cell factories, we examined the glycosylation pathway gene expression profiles as well as the levels of endoplasmic reticulum (ER) and mitochondria in the untransfected hosts. We uncovered a moderate correlation between ER mass and the volumetric product concentration in transient and stable pool Mab production. This work demonstrates the utility of leveraging diversity within the CHOK1SV pool to identify new host cell lines with different performance characteristics.

Patient reported outcome measures for cardiac ablation procedures: a multicentre pilot to develop a new questionnaire.

AIM: To assess the feasibility of administering Patient Reported Outcomes Measures (PROMs) in patients treated with ablation for cardiac arrhythmias, and to conduct the first stage of development and testing of a new PROM tool. METHODS AND RESULTS: A new tool was developed by a multidisciplinary team and tested alongside an adaptation of the patient perception of arrhythmia questionnaire (PPAQ) and EQ-5D-5L in a multicentre retrospective audit involving 791 consecutive cardiac arrhythmia patients treated with catheter ablation at three UK centres over 13 months. Data were recorded in the National Cardiac Rhythm Management Database, part of the National Institute for Cardiovascular Outcomes Research. The response rate was 71.9% (n = 569). Patients reported significant improvements across all outcomes and impacts, with reductions in symptoms of 51.7% (heart racing), 33.9% (fatigue) 31.8% (heart flutters), 43.5% (dizziness), 38.6% (breathlessness), 44.2% (chest pressure), 33.1% (trouble concentrating), 15.9% (headache), 28.3% (neck pressure), and 23.4% (fainting) (P < 0.001). The mean number of social days affected reduced by 7.49 days/month (P < 0.001); mean work/school days affected/month reduced by 6.26 (P < 0.001); mean GP/hospital visits reduced by 1.36 days/month (P < 0.001). The procedure met patient expectations in 72% of responders. CONCLUSIONS: The high response rate suggests that the use of PROMs in this patient group is feasible, with rates equalling those of the National PROMs Programme. The results showed that patients experienced significant improvements in their quality of life following ablation, while feedback allowed the tools to be improved. Further work is required to validate these tools; however, the findings suggest that PROMs could be useful in the audit of ablation techniques.

Cardiac syncope.

Clinicians who diagnose and manage epilepsy frequently encounter diagnoses of a nonneurological nature, particularly when assessing patients with transient loss of consciousness (T-LOC). Among these, and perhaps the most important, is cardiac syncope. As a group, patients with cardiac syncope have the highest likelihood of subsequent sudden death, and yet, unlike sudden unexpected death in epilepsy (SUDEP) for example, it is the norm for these tragic occurrences to be both easily predictable and preventable. In the 12 months following initial presentation with cardiac syncope, sudden death has been found to be 6 times more common than in those with noncardiac syncope (N Engl J Med 309, 1983, 197). In short, for every patient seen with T-LOC, two fundamental aims of the consultation are to assess the likelihood of cardiac syncope as the cause, and to estimate the risk of future sudden death for the individual. This article aims to outline for the noncardiologist how to recognize cardiac syncope, how to tell it apart from more benign cardiovascular forms of syncope as well as from seizures and epilepsy, and what can be done to predict and prevent sudden death in these patients. This is achieved through the assessment triad of a clinical history and examination, risk stratification, and 12-lead electrocardiography (ECG).

Impact of gene vector design on the control of recombinant monoclonal antibody production by Chinese hamster ovary cells.

In this study, we systematically compare two vector design strategies for recombinant monoclonal antibody (Mab) synthesis by Chinese hamster ovary (CHO) cells; a dual open reading frame (ORF) expression vector utilizing separate cytomegalovirus (CMV) promoters to drive heavy chain (HC) and light chain (LC) expression independently, and a single ORF vector design employing a single CMV promoter to drive HC and LC polypeptide expression joined by a foot and mouth disease virus F2A polypeptide self-cleaving linker sequence. Initial analysis of stable transfectants showed that transfectants utilizing the single ORF vector designs exhibited significantly reduced Mab production. We employed an empirical modeling strategy to quantitatively describe the cellular constraints on recombinant Mab synthesis in all stable transfectants. In all transfectants, an intracellular molar excess of LC polypeptide over HC polypeptide was observed. For CHO cells transfected with the single ORF vectors, model-predicted, and empirical intracellular intermediate levels could only be reconciled by inclusion of nascent HC polypeptide degradation. Whilst a local sensitivity analysis showed that qMab of all transfectants was primarily constrained by recombinant mRNA translation rate, our data indicated that all single ORF transfectants exhibited a reduced level of recombinant gene transcription and that Mab folding and assembly reactions generically exerted greater control over qMab. We infer that the productivity of single ORF transfectants is limited by ER processing/degradation "capacity" which sets a limit on transcriptional input. We conclude that gene vector design for oligomeric recombinant proteins should be based on an understanding of protein-specific synthetic kinetics rather than polypeptide stoichiometry.

A mechanistic understanding of production instability in CHO cell lines expressing recombinant monoclonal antibodies.

One of the most significant problems in industrial bioprocessing of recombinant proteins using engineered mammalian cells is the phenomenon of cell line instability, where a production cell line suffers a loss of specific productivity (qP). This phenomenon occurs with unpredictable kinetics and has been widely observed in Chinese hamster ovary (CHO) cell lines and with all commonly used gene expression systems. The underlying causes (both genetic and physiological) and the precise molecular mechanisms underpinning cell line instability have yet to be fully elucidated, although recombinant gene silencing and loss of recombinant gene copies have been shown to cause qP loss. In this work we have investigated the molecular mechanisms underpinning qP instability over long-term sub-culture in CHO cell lines producing recombinant IgG1 and IgG2 monoclonal antibodies (Mab's). We demonstrate that production instability derives from two primary mechanisms: (i) epigenetic--methylation-induced transcriptional silencing of the CMV promoter driving Mab gene transcription and (ii) genetic--progressive loss of recombinant Mab gene copies in a proliferating CHO cell population. We suggest that qP decline resulting from loss of recombinant genes is a consequence of the inherent genetic instability of recombinant CHO cell lines.

An empirical modeling platform to evaluate the relative control discrete CHO cell synthetic processes exert over recombinant monoclonal antibody production process titer.

In this study we have combined empirically derived mathematical models of intracellular Mab synthesis to quantitatively compare the degree to which individual cellular processes limit recombinant IgG(4) monoclonal antibody production by GS-CHO cells throughout a state-of-the-art industrial fed-batch culture process. Based on the calculation of a production process control coefficient for each stage of the intracellular Mab synthesis and secretion pathway, we identified the major cellular restrictions on Mab production throughout the entire culture process to be recombinant heavy chain gene transcription and heavy chain mRNA translation. Surprisingly, despite a substantial decline in the rate of cellular biomass synthesis during culture, with a concomitant decline in the calculated rate constants for energy-intensive Mab synthetic processes (Mab folding/assembly and secretion), these did not exert significant control of Mab synthesis at any stage of production. Instead, cell-specific Mab production was maintained by increased Mab gene transcription which offset the decline in cellular biosynthetic rates. Importantly, this study shows that application of this whole-process predictive modeling strategy should rationally precede and inform cell engineering approaches to increase production of a recombinant protein by a mammalian host cell--where control of productivity is inherently protein product and cell line specific.

Cell line-specific control of recombinant monoclonal antibody production by CHO cells.

In this study we compare the cellular control of recombinant human IgG(4) monoclonal antibody (Mab) synthesis in different CHO cell lines. Based on comprehensive empirical analyses of mRNA and polypeptide synthetic intermediates we constructed cell line-specific mathematical models of recombinant Mab manufacture in seven GS-CHO cell lines varying in specific production rate (qMab) over 350-fold. This comparative analysis revealed that control of qMab involved both genetic construct and cell line-specific factors. With respect to the former, all cell lines exhibited excess production of light chain (LC) mRNA and polypeptide relative to heavy chain (HC) mediated by more rapid LC transcription and enhanced LC mRNA stability. Downstream of this, cell lines differed markedly in their relative rates of recombinant mRNA translation, Mab assembly and secretion although HC mRNA abundance and the rate of HC translation generally exerted most control over qMab--the latter being directly proportional to qMab. This study shows that (i) cell lines capable of high qMab exceed a threshold functional competency in all synthetic processes, (ii) the majority of cells in parental and transfected cell populations are functionally limited and (iii) cell engineering strategies to increase Mab production should be cell line specific.

The complete array of electrocardiogram abnormalities secondary to myocardial contusion in a single case.

Myocardial contusion is a complication of blunt thoracic injuries. Transthoracic echocardiography and electrocardiography (ECG) monitoring are important in suspected cases. We report a 54-year-old man, who sustained a number of injuries including blunt chest injury as a consequence of a road traffic accident. Electrocardiography monitoring over a 48 h period demonstrated sequential degrees of conduction system block coupled with a temporary cardio-version from persistent atrial fibrillation to sinus rhythm, suggesting coincident pulmonary vein contusion.

Mitral valve surgery plus concomitant atrial fibrillation ablation is superior to mitral valve surgery alone with an intensive rhythm control strategy.

OBJECTIVE: Prospective randomised study comparing patients with atrial fibrillation (AF) of more than 6 months duration after mitral valve surgery plus biatrial modified radiofrequency Maze procedure using Medtronic Cardioblate System (Cardioblate group, n=24) vs mitral valve surgery plus intensive rhythm control strategy (control group, n=25). METHODS: Patients were blinded to randomisation. Preoperatively, at discharge, and at 3-month and 1-year follow-up, echocardiography, quality of life assessments and ECGs were done. In both groups, sinus rhythm (SR) restoration was attempted by intra- and postoperative DC cardioversion and class III antiarrhythmic medication. All patients received warfarin. Amiodarone and warfarin was considered for discontinuation after 3 months in SR, 24-h Holter or event monitor excluding AF. RESULTS: Both groups underwent mitral valve replacement or repair (Cardioblate vs control: 16:8 vs 10:15), had similar gender (male: 33% vs 56%), age (66+/-8 years vs 68+/-9 years), additional aortic valve replacement (7 vs 6 patients), tricuspid annuloplasty (13 vs 13 patients), and CABG (10 vs 16 patients). There was 0% operative mortality, 0% postoperative cerebrovascular accidents, but 2 late deaths in the control group. At discharge, 3- and 12-month follow-up, more patients in the Cardioblate group returned to normal SR compared to control (29%, 57% and 75% vs 20%, 43% and 39%; p=0.030). Return of functional atrial contraction in patients in SR at 1 year was comparable between groups (63% vs 89%, NS), and more likely in non-rheumatic pathology and preoperative AF of shorter duration. The effectiveness of atrial contraction was 36+/-14% vs 43+/-18% of transmitral flow and there was no difference between groups. Amiodarone treatment decreased more in Cardioblate group over time (92%, 55% and 29% vs 52%, 52% and 21%; p=0.003), whereas warfarin decrease was comparable (100%, 100% and 71% vs 100%, 95% and 82%; NS). CONCLUSIONS: Radiofrequency Maze ablation additional to mitral valve surgery resulted in a higher SR conversion rate (75%), despite control group treatment with intensive rhythm control strategy having a higher SR conversion rate (39%) compared to literature (approximately 25%). Maze ablation resulted in normalisation of atrial function in 63% of patients converted to SR.

Systems biotechnology of mammalian cell factories.

The increasing demand for recombinant therapeutic proteins has placed significant pressure on the biopharmaceutical industry to develop high-yielding, mammalian cell-based production systems. Current efforts to increase the production of recombinant proteins by mammalian host cells largely proceed by the lengthy screening of clonal derivatives rather than by directed genetic or metabolic engineering. However, the advent of systems biology has created a new set of tools that will ensure that future engineering strategies will be informed by an understanding of the genetic/regulatory and metabolic networks that determine the functional competence of mammalian cell factories in vitro. In this review we summarize recent systems-level studies that utilize genome-scale analytical tools to analyse the functional basis for key production process characteristics such as high cell-specific productivity, correct product processing and rapid cell proliferation in the in vitro environment. We also describe the use of high-throughput -omic technologies to investigate how mammalian cell factories respond to environmental and metabolic perturbation.

The humble electrocardiogram.

Many patients with potentially life-threatening cardiac conditions presenting with transient loss of consciousness are referred first to neurology clinics. Therefore, neurologists must remain competent to interpret electrocardiograms (ECGs) and in particular be able to identify those rare conditions that predict sudden cardiac death. A 12-lead ECG is cheap and readily available, and can give essential diagnostic information. Here the authors review abnormalities in, and indications for, the ECG in neurological practice.

The investigation of syncope.

Patients with syncope are usually referred to either neurology or cardiology clinics, yet the facilities for detailed syncope investigation are mostly in cardiac units. The diagnosis rests principally upon the history, but investigations may be required to support the clinical diagnosis. Close collaboration between the epilepsy clinician and a cardiologist is essential for effective investigation and safe management of syncope. It is frequently misdiagnosed and often erroneously treated as epilepsy. Furthermore, it is potentially a marker of sudden death when associated with certain cardiac disorders. Here we review the main syncope types and explore diagnostic approaches.