A higher spectral range of beetle bioluminescence with infraluciferin.

Coleopteran bioluminescence is unique in that beetle luciferases emit colors ranging between green (ca.550 nm) and red (ca.600 nm), including intermediate colors such as yellow and orange, allowing up to 3 simultaneous parameters to be resolved in vitro with natural luciferin (D-LH2). Here, we report a more than doubling of the maximum bioluminescence wavelength range using a single synthetic substrate, infraluciferin (iLH2). We report that different luciferases can emit colors ranging from visible green to near-infrared (nIR) with iLH2, including in human cells. iLH2 was designed for dual color far-red to nIR bioluminescence imaging (BLI) in small animals and has been utilized in different mouse models of cancer (including a metastatic hepatic model showing detailed hepatic morphology) and for robust dual parameter imaging in vivo (including in systemic hematological models). Here, we report the properties of different enzymes with iLH2: Lampyrid wild-type (WT) Photinus pyralis (Ppy) firefly luciferase, Ppy-based derivatives previously engineered to be thermostable with D-LH2, and also color-shifted Elaterid-based enzymes: blue-shifted Pyrearinus termitilluminans derivative Eluc (reported D-LH2 λmax = 538 nm) and red-shifted Pyrophorus plagiopthalamus derivative click beetle red (CBR) luciferase (D-LH2 λmax = 618 nm). As purified enzyme, in bacteria or in human cells, Eluc emitted green light (λmax = 536 nm) with DL-iLH2 whereas Ppy Fluc (λmax = 689 nm), x2 Fluc (λmax = 704 nm), x5 Fluc (λmax = 694 nm), x11 Fluc (λmax = 694 nm) and CBR (λmax = 721 nm) produced far-red to nIR peak wavelengths. Therefore, with iLH2, enzyme λmaxes can be separated by ca.185nm, giving almost non-overlapping spectra. This is the first report of single-substrate bioluminescence color emission ranging from visible green to nIR in cells and may help shed light on the color tuning mechanism of beetle luciferases. We also report on the reason for the improvement in activity of x11 Fluc with iLH2 and engineer an improved infraluciferase (iluc) based on this mutant.

Diagnostic and Interventional Radiology Case Volume and Education in the Age of Pandemics: Impact Analysis and Potential Future Directions.

RATIONALE AND OBJECTIVES: To assess the immediate impact of the COVID-19 pandemic on Diagnostic and Interventional Radiology education, and to propose measures to preserve and augment trainee education during future crises. MATERIALS AND METHODS: Diagnostic Radiology (DR) studies and Interventional Radiology (IR) procedures at a single tertiary-care teaching institution between 2015 and 2020 were reviewed. DR was divided by section: body, cardiothoracic, musculoskeletal (MSK), neuroradiology, nuclear medicine, pediatrics, and women's imaging. IR was divided by procedural types: arterial, venous, lymphatic, core, neuro, pediatrics, dialysis, cancer embolization or ablation, noncancer embolization, portal hypertension, and miscellaneous. Impact on didactic education was also assessed. ANOVA, t test, and multiple comparison correction were used for analysis. RESULTS: DR and IR caseloads decreased significantly in April 2020 compared to April of the prior 5 years (both p < 0.0001). Case volumes were reduced in body (49.2%, p < 0.01), MSK (54.2%, p < 0.05), neuro (39.3%, p < 0.05), and women's imaging (75.5%, p < 0.05) in DR, and in arterial (62.6%, p < 0.01), neuro IR (57.6%, p < 0.01) and core IR (42.6%, p < 0.05) in IR. IR trainee average caseload in April 2020 decreased 51.9% compared to April of the prior 5 years (p < 0.01). Utilization of online learning increased in April. Trainees saw significant increases in overall DR didactics (31.3%, p = 0.02) and no reduction in IR didactics, all online. Twelve major national and international DR and IR meetings were canceled or postponed between March and July. CONCLUSION: Decreases in caseload and widespread cancellation of conferences have had significant impact on DR/IR training during COVID-19 restrictions. Remote learning technologies with annotated case recording, boards-style case reviews, procedural simulation and narrated live cases as well as online lectures and virtual journal clubs increased during this time. Whether remote learning can mitigate lost opportunities from in-person interactions remains uncertain. Optimizing these strategies will be important for potential future restricted learning paradigms and can also be extrapolated to augment trainee education during unrestricted times.

Acute Improvement in Intraoperative EMG During Common Fibular Nerve Decompression in Patients with Symptomatic Diabetic Sensorimotor Peripheral Neuropathy: EMG and Clinical Attribute Interrelations.

STUDY AIMS: Electromyographic (EMG) recordings of the fibularis longus and tibialis anterior muscles were performed intraoperatively during nerve decompression (ND) of the common fibular nerve (CFN) in patients with symptomatic diabetic sensorimotor peripheral neuropathy. Patient demographics and clinical attributes were compared against changes in EMG after ND and analyzed for possible correlations. METHODS: Intraoperative changes in CFN EMG were analyzed for correlations against sex, age, body mass index (BMI), hemoglobin A1c (A1c), and type and duration of diabetes. RESULTS: Statistically significant changes were found between EMG changes and patient attributes, but no individual correlations were established. Significant EMG improvement was observed for both men and women (p < 0.0001 and p < 0.05, respectively), age groups (4th decade: p < 0.05; 5th decade: p < 0.05; 6th decade: p < 0.01; 7th decade: p < 0.005), diabetes duration (0-9 years: p = 0.002; 10-19 years: p = 0.002; 20-29 years: p = 0.03), and for type 1 and 2 diabetes (type 1: p < 0.005; type 2: p < 0.001). EMG improvement was greater in patients with the highest BMI levels (30-34.9: p = 0.014; 35-39.9: p = 0.013; > 39.9: p = 0.043), and highest A1c levels (> 6.4%; p < 0.0001). CONCLUSION: Although long-term clinical studies are needed, these results provide insight into which patients might benefit most from this surgery. These results also suggest that surgical ND can produce an acute improvement in nerve function for both men and women, for people with type 1 and 2 diabetes, and across a wide range of ages, BMI, A1c levels, and disease duration.

Near-infrared dual bioluminescence imaging in mouse models of cancer using infraluciferin.

Bioluminescence imaging (BLI) is ubiquitous in scientific research for the sensitive tracking of biological processes in small animal models. However, due to the attenuation of visible light by tissue, and the limited set of near-infrared bioluminescent enzymes, BLI is largely restricted to monitoring single processes in vivo. Here we show, that by combining stabilised colour mutants of firefly luciferase (FLuc) with the luciferin (LH2) analogue infraluciferin (iLH2), near-infrared dual BLI can be achieved in vivo. The X-ray crystal structure of FLuc with a high-energy intermediate analogue, 5'-O-[N-(dehydroinfraluciferyl)sulfamoyl] adenosine (iDLSA) provides insight into the FLuc-iLH2 reaction leading to near-infrared light emission. The spectral characterisation and unmixing validation studies reported here established that iLH2 is superior to LH2 for the spectral unmixing of bioluminescent signals in vivo; which led to this novel near-infrared dual BLI system being applied to monitor both tumour burden and CAR T cell therapy within a systemically induced mouse tumour model.

Nerve Decompression and Restless Legs Syndrome: A Retrospective Analysis.

INTRODUCTION: Restless legs syndrome (RLS) is a prevalent sleep disorder affecting quality of life and is often comorbid with other neurological diseases, including peripheral neuropathy. The mechanisms related to RLS symptoms remain unclear, and treatment options are often aimed at symptom relief rather than etiology. RLS may present in distinct phenotypes often described as "primary" vs. "secondary" RLS. Secondary RLS is often associated with peripheral neuropathy. Nerve decompression surgery of the common and superficial fibular nerves is used to treat peripheral neuropathy. Anecdotally, surgeons sometimes report improved RLS symptoms following nerve decompression for peripheral neuropathy. The purpose of this retrospective analysis was to quantify the change in symptoms commonly associated with RLS using visual analog scales (VAS). METHODS: Forty-two patients completed VAS scales (0-10) for pain, burning, numbness, tingling, weakness, balance, tightness, aching, pulling, cramping, twitchy/jumpy, uneasy, creepy/crawly, and throbbing, both before and 15 weeks after surgical decompression. RESULTS: Subjects reported significant improvement among all VAS categories, except for "pulling" (P = 0.14). The change in VAS following surgery was negatively correlated with the pre-surgery VAS for both the summed VAS (r = -0.58, P < 0.001) and the individual VAS scores (all P < 0.01), such that patients who reported the worst symptoms before surgery exhibited relatively greater reductions in symptoms after surgery. CONCLUSION: This is the first study to suggest improvement in RLS symptoms following surgical decompression of the common and superficial fibular nerves. Further investigation is needed to quantify improvement using RLS-specific metrics and sleep quality assessments.

Acute Improvement in Intraoperative EMG Following Common Fibular Nerve Decompression in Patients with Symptomatic Diabetic Sensorimotor Peripheral Neuropathy: 1. EMG Results.

Background and Study Aims Electromyographic (EMG) recordings of the fibularis longus (FL) and tibialis anterior (TA) muscles were performed intraoperatively during common fibular nerve (CFN) nerve decompression (ND) in patients with symptomatic diabetic sensorimotor peripheral neuropathy (DSPN) and clinical nerve compression. Materials and Methods Forty-six legs in 40 patients underwent surgical ND by external neurolysis; FL and TA muscles were monitored intraoperatively. Evoked EMGs were recorded just prior to and within 1 minute after ND. Results Thirty-eight legs (82.6%) demonstrated EMG improvement 1 minute after ND. Sixty muscles (31 FL, 29 TA) were monitored, with 44 (73.3%) improving in EMG amplitude. Mean change in EMG amplitude represented a 73.6% improvement (p < 0.0001). Changes in EMG amplitudes correlated with visual analog scale pain improvement (p = 0.03). Conclusion This is the first report of acute changes in objective EMG responses during ND of CFN in DSPN patients and demonstrates that patients with symptomatic DSPN and clinical nerve entrapment have latent but functional axons that surgical ND can improve immediately.

Intraoperative Nerve Monitoring During Nerve Decompression Surgery in the Lower Extremity.

This article describes the benefits of intraoperative neurophysiologic monitoring (IONM) and proposes methods for integration into nerve decompression procedures. Standard procedures for intraoperative nerve monitoring (IONM) are illustrated as they would apply to the 3 nerve tunnels that have significant motor components within the lower extremity.

Common Fibular Nerve Compression: Anatomy, Symptoms, Clinical Evaluation, and Surgical Decompression.

This article thoroughly describes the clinical examination and treatment of common fibular (peroneal) nerve compression. Aspects discussed include the anatomy of the nerve, cause of entrapment, symptoms associated with impairment, and a surgical approach to decompress the entrapped nerve. The standard protocol for decompression as it would apply to the common fibular nerve tunnel is illustrated.

Staphylococcal phenotypes induced by naturally occurring and synthetic membrane-interactive polyphenolic ß-lactam resistance modifiers.

Galloyl catechins, in particular (-)-epicatechin gallate (ECg), have the capacity to abrogate ß-lactam resistance in methicillin-resistant strains of Staphylococcus aureus (MRSA); they also prevent biofilm formation, reduce the secretion of a large proportion of the exoproteome and induce profound changes to cell morphology. Current evidence suggests that these reversible phenotypic traits result from their intercalation into the bacterial cytoplasmic membrane. We have endeavoured to potentiate the capacity of ECg to modify the MRSA phenotype by stepwise removal of hydroxyl groups from the B-ring pharmacophore and the A:C fused ring system of the naturally occurring molecule. ECg binds rapidly to the membrane, inducing up-regulation of genes responsible for protection against cell wall stress and maintenance of membrane integrity and function. Studies with artificial membranes modelled on the lipid composition of the staphylococcal bilayer indicated that ECg adopts a position deep within the lipid palisade, eliciting major alterations in the thermotropic behaviour of the bilayer. The non-galloylated homolog (-)-epicatechin enhanced ECg-mediated effects by facilitating entry of ECg molecules into the membrane. ECg analogs with unnatural B-ring hydroxylation patterns induced higher levels of gene expression and more profound changes to MRSA membrane fluidity than ECg but adopted a more superficial location within the bilayer. ECg possessed a high affinity for the positively charged staphylococcal membrane and induced changes to the biophysical properties of the bilayer that are likely to account for its capacity to disperse the cell wall biosynthetic machinery responsible for ß-lactam resistance. The ability to enhance these properties by chemical modification of ECg raises the possibility that more potent analogs could be developed for clinical evaluation.

Central nervous system stem cell transplantation for children with neuronal ceroid lipofuscinosis.

OBJECT: Infantile and late-infantile neuronal ceroid lipofuscinoses (NCLs) are invariably fatal lysosomal storage diseases associated with defects in lysosomal enzyme palmitoyl-protein thioesterase 1 (PPT-1) or tripeptidyl peptidase 1 (TPP1) activity. Previous preclinical studies have demonstrated that human CNS stem cells (HuCNS-SCs) produce both PPT-1 and TPP1 and result in donor cell engraftment and reduced accumulation of storage material in the brain when tested in an NCL mouse model. METHODS: HuCNS-SC transplantation was tested in an open-label dose-escalation Phase I clinical trial as a potential treatment for infantile and late-infantile NCL. Study design included direct neurosurgical transplantation of allogeneic HuCNS-SCs into the cerebral hemispheres and lateral ventricles accompanied by 12 months of immunosuppression. RESULTS: Six children with either the infantile or late-infantile forms of NCL underwent low- (3 patients) and high- (3 patients) dose transplantation of HuCNS-SCs followed by immunosuppression. The surgery, immunosuppression, and cell transplantation were well tolerated. Adverse events following transplantation were consistent with the underlying disease, and none were directly attributed to the donor cells. Observations regarding efficacy of the intervention were limited by the enrollment criteria requiring that patients be in advanced stages of disease. CONCLUSIONS: This study represents the first-in-human clinical trial involving transplantation of a purified population of human neural stem cells for a neurodegenerative disorder. The feasibility of this approach and absence of transplantation-related serious adverse events support further exploration of HuCNS-SC transplantation as a potential treatment for select subtypes of NCL, and possibly for other neurodegenerative disorders.

Traditional threshold for retropharyngeal soft-tissue swelling is poorly sensitive for the detection of cervical spine injury on computed tomography in adult trauma patients.

STUDY DESIGN: Retrospective cohort study. OBJECTIVE: To examine the diagnostic value of prevertebral soft-tissue swelling in the setting of cervical spine trauma. SUMMARY OF BACKGROUND DATA: In adult patients with trauma, an increase in the thickness of the retropharyngeal soft tissues is commonly used as a potential indicator of occult injury, but no studies have examined this parameter using computed tomography (CT) as a screening modality. METHODS: A total of 541 patients with trauma with injuries at any level of the spine underwent CT. Patients with cervical injury were divided into those requiring noninvasive (observation or cervical collar, n = 142) management, and those requiring invasive (surgery or halo, n = 61) treatment. A control group of patients with isolated thoracic or lumbar injuries was used for comparison (n = 542). Retropharyngeal soft tissues were measured at the cranial and caudal endplates of all cervical levels on sagittal and axial CT. Sensitivity and specificity were calculated for +1, +2, and +3 standard deviations from mean values. RESULTS: Sensitivity for detection of injury was found to be universally poor for all measurement groups. This ranged from 14.4% to 21.2% at +1 SD to 5.3% to 8.7% at +2 SD. Positive and negative predictive values for injury were also universally poor, ranging from 38% to 75%. Soft-tissue swelling as a sentinel sign of cervical spine injury demonstrates consistently high specificity and low sensitivity, precisely the opposite of what would be desired in a screening test. This study shows at best a sensitivity of 21.6% when using this parameter for the detection of these injuries in adult patients with trauma. CONCLUSION: On the basis of the results of this study, we recommend against the routine use of measurement of the prevertebral soft tissues on CT as a screening tool for cervical spine injury in adult patients with trauma. LEVEL OF EVIDENCE: 3.

Total synthesis of rapamycin.

For over 30 years, rapamycin has generated a sustained and intense interest from the scientific community as a result of its exceptional pharmacological properties and challenging structural features. In addition to its well known therapeutic value as a potent immunosuppressive agent, rapamycin and its derivatives have recently gained prominence for the treatment of a wide variety of other human malignancies. Herein we disclose full details of our extensive investigation into the synthesis of rapamycin that culminated in a new and convergent preparation featuring a macro-etherification/catechol-templating strategy for construction of the macrocyclic core of this natural product.

A novel tridentate coordination mode for the carbonatonickel system exhibited in an unusual hexanuclear nickel(II) mu3-carbonato-bridged complex.

The fixation of CO(2) at ambient temperature has been achieved by the reaction of Ni(cod)(2) and TMEDA in CO(2) saturated THF that yields a novel hexanuclear nickel(II) mu(3)-carbonato bridged complex [Ni(6)(mu(3)-CO(3))(4)(TMEDA)(6)(H(2)O)(12)](OH)(4) in 59% yield. The complex was characterised by MS analysis and the structure corroborated by single-crystal X-ray crystallography. The complex exhibits a rare carbonato binding mode for Ni(II) complexes and moderately strong antiferromagnetic interactions.

Osteoblastoma of the spine with discordant magnetic resonance imaging and computed tomography imaging features in a child.

STUDY DESIGN: Case report. OBJECTIVE: To describe the magnetic resonance imaging (MRI) and computed tomography (CT) findings of spinal osteoblastoma and illustrate how MRI features can be potentially misleading. SUMMARY OF BACKGROUND DATA: Osteoblastoma is a rare benign tumor of bone that is known to incite a localized inflammatory response. These inflammatory features can simulate malignant behavior on MRI and can lead to misdiagnosis and unnecessarily aggressive resection unless one recognizes the classic benign features on CT. We present a case of osteoblastoma in a child to illustrate this concept. METHODS: The patient's clinical and radiologic history is discussed with a brief review of the relevant literature. RESULTS: A 9-year-old girl with back pain receives an MRI of the lumbar spine demonstrating a sacral tumor with potentially malignant features including extensive marrow edema and enhancement. A CT demonstrates a well-circumscribed lesion without lytic changes or malignant bone formation. A benign lesion was favored given the CT features and conservative resection was performed. Final pathologic diagnosis was osteoblastoma. CONCLUSION: This case illustrates that the MRI findings for osteoblastoma can be misleading and caution should be used when evaluating benign tumors with known inflammatory responses on MRI. CT features seem to more accurately reflect the true nature and extent of the tumor.

Radiographic correlation with neck anatomy.

The anatomy of the head and neck is an important set of knowledge for the oral and maxillofacial surgeon. Current imaging techniques and the exquisite detail that they provide are frequently the first glimpse at disease and important to surgical planning. CT and MRI are the primary modes of neck imaging and are complimentary in the information that they provide. This article reviews the current methods of anatomic imaging and the current methods of analysis of the region by radiologists.

Modulation of beta-lactam resistance in Staphylococcus aureus by catechins and gallates.

Aqueous extracts of Japanese green tea (Camellia sinensis) are able to reverse beta-lactam resistance in methicillin-resistant Staphylococcus aureus (MRSA). We have attributed the capacity to reverse oxacillin resistance in the homogeneous PBP2a producer BB568 and in EMRSA-16 to (-)-epicatechin gallate (ECG) and (-)-catechin gallate (CG). Minimum inhibitory concentration (MIC) values for oxacillin were reduced from 256 and 512 to 1-4 mg/l, respectively, in the presence of these polyphenols. In addition, (-)-epigallocatechin gallate (EGCG) had a moderate capacity to modulate oxacillin activity against S. aureus BB568, but none against EMRSA-16. ECG, CG and EGCG increased the sensitivity of EMRSA-15 to oxacillin. The gallate moiety was essential for the oxacillin-modulating activity of ECG, as both (-)-epicatechin and (-)-epicatechin-3-cyclohexylcarboxylate were unable to reverse resistance to oxacillin. Gallic acid and three alkyl gallates (methyl gallate, propyl gallate, and octyl gallate) did not modulate beta-lactam resistance in MRSA. Octyl gallate exhibited direct antibacterial activity against S. aureus BB568 (16 mg/l). Modulation of beta-lactam resistance by ECG significantly enhanced the activities of flucloxacillin and the carbapenem antibiotics imipenem and meropenem against 40 MRSA isolates, with MIC(90) values for the antibiotics reduced to the susceptibility breakpoint or below. Consequently, EGCG, CG and, particularly, ECG warrant further investigation as agents to combat beta-lactam resistance in S. aureus.

July 2002: 66-year-old female with a one-year history of progressive left proptosis.

The July 2002 Case of the Month (COM). This 66-year-old Caucasian female presented with gradually increasing protrusion of her left eye over a one-year period. She complained of increased tearing and foreign body sensation. The physical examination revealed a visual acuity of 20/20, normal color testing, full vision field with motility of her left eye limited in lateral gaze. Pupils were round, symmetric, with no afferent pupillary defect noted. On external examination, her left eye was grossly proptotic with resistance to retropulsion. She had 4 mm proptosis of the left eye. Computed tomography and MR imaging demonstrated a left retro-orbital mass with gadolinium enhancement and focal remodeling of orbital bones. She underwent surgical resection of tumor with a diagnosis of solitary fibrous tumor, and postoperatively she was symptom free. The histopathological differential diagnoses of spindle cell neoplasms of the orbit are discussed. Five months after surgery, no evidence of tumor recurrence was seen on neuroimaging and her vision was 20/20.