A novel protocol for three-dimensional mapping of sand tiger shark (Carcharias taurus) enclosure use in aquaria: Implications for management.

This study investigated sand tiger shark (STS; Carcharias taurus) spatial use and exclusion in public aquarium enclosures using a novel protocol for three-dimensional mapping. Fifty-one STS were observed in 14 enclosures, and swimming pattern, depth, and location were recorded in ZooMonitor. Data were converted into quantitative, three-dimensional representations using ArcGIS® Pro v. 2.9. All observed STS except one swam in circular patterns, and 80% (n = 41) showed a directional swimming bias. Most STS (80%; n = 41) predominantly utilized the top two-thirds of the enclosures, though 83% (n = 34) of those had swimming obstructions in the bottom of the enclosure. Avoidance of obstructed areas, sections <7 m wide, as well as behavioral spatial separation, resulted in utilization of between 27% and 66% of available enclosure space. STS underutilized corners, pinch-points, and obstructed areas requiring abrupt directional changes and instead exhibited continual, unimpeded swimming patterns. In addition, this study found no relationship between directional swimming bias or use of smaller enclosure volumes and spinal deformity, a health issue affecting 26% of STS 10 years ago but now with an incidence of 6%. Using novel protocols for three-dimensional mapping and volume estimation, this study demonstrated that enclosures facilitating unimpeded, continuous swimming are most usable for STS and provides important information that will be useful for future enclosure design.

Evaluating the Utilization of a Perioperative Hyperglycemic Protocol: A Quality Improvement Project.

PURPOSE: The purpose of this project to evaluate adherence to the perioperative hyperglycemic protocol among Certified Registered Nurse Anesthetists (CRNAs) at a large academic hospital. A secondary objective of this project is CRNAs' perceptions of barriers to point-of-care (POC) testing and the protocol. DESIGN: A quality improvement project. METHODS: Using Donabedian's conceptual framework, a Phase 1 retrospective chart analysis of 297 patients with diabetes undergoing noncardiac surgery before and after implementing POC testing for intraoperative glucose control was performed. Only patients with preoperative BG ≥ 180 mg/dL were included in this phase of the project, which involved a comparison of the protocol utilization before and after implementation of POC testing. Phase 2 included an assessment of CRNA's perceptions of the protocol. FINDINGS: The final sample included 91 (37 preimplementation; 54 postimplementation) participants. There were no significant demographic differences between the groups. Overall, 52.7% of patients had intraoperative glucose checks, and only 16.5% received insulin. Preoperative BG levels decreased 11.4-points, and postoperative BG levels increased 20.4 points when comparing pre- and postimplementation groups. However, there were significant differences in postoperative glucose levels, pre- and postimplementation. The survey showed that the majority (65.5%) of CRNAs identified difficulty locating the protocol as the primary barrier to utilization. CONCLUSIONS: Although all patients included in this project qualified for an intraoperative glucose check, findings revealed that only half of the patients had a glucose check and less than one fifth of the patients received insulin treatment, indicating poor adherence to the protocol. Thus, while implementing protocols is essential, utilization and adherence to the protocol are critical to improving patient outcomes. Recommendations for continued improvement include increasing protocol accessibility, staff training, compliance monitoring, and a more simple protocol structure.

Use of embedded and patterned dichroic surfaces with reflective optical power to enable multiple optical paths in a micro-objective.

We demonstrate the use of patterned dichroic surfaces with reflective optical power to create multiple optical paths in a single lens system. The application of these surfaces enables a micro-endoscope to accommodate multiple imaging technologies with only one optical system, making the packaging more compact and reliable. The optical paths are spectrally separated using different wavelengths for each path. The dichroic surfaces are designed such that the visible wavelengths transmit through the surfaces optically unaffected, but the near-infrared wavelengths are reflected in a telescope-like configuration with the curved dichroic surfaces providing reflective optical power. We demonstrate wide-field visible monochromatic imaging and microscopic near-infrared imaging using the same set of lenses. The on-axis measured resolution of the wide-field imaging configuration is approximately 14 µm, and the measured resolution of the microscopic imaging configuration is approximately 2 µm. Wide-field white-light imaging of an object is also demonstrated for a qualitative perspective on the imaging capabilities. Other configurations and applications in fields such as optical metrology are discussed to expand on the versatility of the demonstrated optical system.

A Comprehensive Review of Cross-Sectional Imaging of the Nasolacrimal Drainage Apparatus: What Radiologists Need to Know.

OBJECTIVE. The purpose of this study is to provide a comprehensive review of the radiographic anatomy and cross-sectional imaging findings of the full gamut of nasolacrimal drainage apparatus diseases, highlighting imaging findings from the different nasolacrimal drainage apparatus surgeries, posttreatment complications, and potential imaging pitfalls. CONCLUSION. Radiologists play a critical role in guiding the management of nasolacrimal drainage apparatus diseases and should be familiar with the anatomy and characteristic imaging findings of commonly encountered nasolacrimal drainage apparatus abnormalities and surgeries.

Identification of quinones as novel PIM1 kinase inhibitors.

PIM1 is a proto-oncogene encoding the serine/threonine PIM1 kinase. PIM1 kinase plays important roles in regulating aspects of cell cycle progression, apoptosis resistance, and has been implicated in the development of such malignancies as prostate cancer and acute myeloid leukemia among others. Knockout of PIM1 kinase in mice has been shown to be non-lethal without any obvious phenotypic changes, making it an attractive therapeutic target. Our investigation of anthraquinones as kinase inhibitors revealed a series of quinone analogs showing high selectivity for inhibition of the PIM kinases. Molecular modeling studies were used to identify key interactions and binding poses of these compounds within the PIM1 binding pocket. Compounds 1, 4, 7 and 9 inhibited the growth of DU-145 prostate cancer cell lines with a potency of 8.21µM, 4.06µM, 3.21µM and 2.02µM.

Performance of high-resolution CT for detection and discrimination tasks related to stenotic lesions - A phantom study using model observers.

BACKGROUND: Accurate detection and grading of atheromatous stenotic lesions within the cardiac, renal, and intracranial vasculature is imperative for early recognition of disease and guiding treatment strategies. PURPOSE: In this work, a stenotic lesion phantom was used to compare high resolution and normal resolution modes on the same CT scanner in terms of detection and size discrimination performance. MATERIALS AND METHODS: The phantom is comprised of three acrylic cylinders (each 15.0 cm in diameter and 1.3 cm thick) with a matching array of holes in each module. The outer two modules contain holes that are slightly larger than the corresponding hole in the central module to simulate stenotic narrowing in vasculature. The stack of modules was submerged in an iodine solution simulating contrast-enhanced stenotic lesions with a range of lumen diameters (1.32-10.08 mm) and stenosis severity (0%, 50%, 60%, 70%, and 80%). The phantom was imaged on the Canon Aquilion Precision high-resolution CT scanner in high-resolution (HR) mode (0.25 mm × 0.50 mm detector element size) and normal-resolution (NR) mode (0.50 mm × 0.50 mm) using 120 kV and two dose levels (14 and 21 mGy SSDE) with 30 repeat scans acquired for each combination. Filtered back-projection (FBP) and a hybrid-iterative reconstruction (AIDR) were used with the FC18 kernel, as well as a deep learning algorithm (AiCE) which is only available for HR. A non-prewhitening model observer with an eye filter was implemented to quantify performance for detection and size discrimination tasks in the axial plane. RESULTS: Detection performance improved with increasing diameter, dose, and for AIDR in comparison to FBP for a fixed resolution mode. Performance in the HR mode was generally higher than NR for the smaller lumen diameters (1-5 mm) with decreasing differences as the diameter increased. Performance in NR mode surpassed HR mode for lumen diameters greater than ∼4 mm and ∼5 mm for 14 mGy and 21 mGy, respectively. AiCE provided consistently higher detection performance compared with AIDR-FC18 (48% higher for a 6 mm lumen diameter). Discrimination performance increased with increasing nominal diameter, dose, and for larger differences in stenosis severity. When comparing discrimination performance in HR to NR modes, the largest relative differences occur at the smallest nominal diameters and smallest differences in stenosis severity. The AiCE reconstruction algorithm produced the highest overall discrimination performance values, and these were significantly higher than AIDR-FC18 for nominal diameters of 7.14 and 10.08 mm. CONCLUSIONS: HR mode outperforms NR for detection up to a specific diameter and the results improve with AiCE and for higher dose levels. For the task of size discrimination, HR mode consistently outperforms NR if AIDR-FC18 is used for dose levels of at least 21 mGy, and the results improve with AiCE and for the smallest differences in stenosis severity investigated (50% vs. 60%). High-resolution CT appears to be beneficial for detecting smaller simulated lumen diameters (<5 mm) and is generally advantageous for discrimination tasks related to stenotic lesions, which inherently contain information at higher frequencies, given the right reconstruction algorithm and dose level.

Multi-Parametric Radiomic Model to Predict 1p/19q Co-Deletion in Patients with IDH-1 Mutant Glioma: Added Value to the T2-FLAIR Mismatch Sign.

PURPOSE: The T2-FLAIR mismatch sign has shown promise in determining IDH mutant 1p/19q non-co-deleted gliomas with a high specificity and modest sensitivity. To develop a multi-parametric radiomic model using MRI to predict 1p/19q co-deletion status in patients with newly diagnosed IDH1 mutant glioma and to perform a comparative analysis to T2-FLAIR mismatch sign+. METHODS: In this retrospective study, patients with diagnosis of IDH1 mutant gliomas with known 1p/19q status who had preoperative MRI were included. T2-FLAIR mismatch was evaluated independently by two board-certified neuroradiologists. Texture features were extracted from glioma segmentation of FLAIR images. eXtremeGradient Boosting (XGboost) classifiers were used for model development. Leave-one-out-cross-validation (LOOCV) and external validation performances were reported for both the training and external validation sets. RESULTS: A total of 103 patients were included for model development and 18 patients for external testing validation. The diagnostic performance (sensitivity/specificity/accuracy) in the determination of the 1p/19q co-deletion status was 59%/83%/67% (training) and 62.5%/70.0%/66.3% (testing) for the T2-FLAIR mismatch sign. This was significantly improved (p = 0.04) using the radiomics model to 77.9%/82.8%/80.3% (training) and 87.5%/89.9%/88.8% (testing), respectively. The addition of radiomics as a computer-assisted tool resulted in significant (p = 0.02) improvement in the performance of the neuroradiologist with 13 additional corrected cases in comparison to just using the T2-FLAIR mismatch sign. CONCLUSION: The proposed radiomic model provides much needed sensitivity to the highly specific T2-FLAIR mismatch sign in the determination of the 1p/19q non-co-deletion status and improves the overall diagnostic performance of neuroradiologists when used as an assistive tool.

Complications of aortic valve surgery: manifestations at CT and MR imaging.

Aortic valve replacement accounts for a significant portion of cardiac surgeries in the United States. Despite advances in prosthetic heart valve design, surgical technique, and postoperative care, complications after aortic valve replacement remain a leading cause of morbidity and mortality. Routine surveillance of prosthetic heart valves with transthoracic echocardiography (TTE), transesophageal echocardiography (TEE), and fluoroscopy is important, as these techniques allow accurate detection of prosthetic valve dysfunction. However, echocardiography and fluoroscopy may not allow identification of the specific underlying cause, including paravalvular leak, dehiscence, endocarditis, obstruction, structural failure, pseudoaneurysm formation, aortic dissection, and hemolysis. Magnetic resonance (MR) imaging and computed tomography (CT) have an emerging role as diagnostic tools complementary to conventional imaging for detection and monitoring of complications after aortic valve replacement. The choice between CT and MR imaging depends on individual patient characteristics, the type of prosthetic valve, and the acuity of the clinical situation. In general, screening with TTE followed by TEE is recommended. When results of TTE and TEE are inconclusive, cardiac CT and MR imaging should be considered. The choice between these imaging techniques depends on the presence of patient-specific contraindications to CT or MR imaging.

Artificial Intelligence in Head and Neck Imaging.

Artificial intelligence (AI) can be applied to head and neck imaging to augment image quality and various clinical tasks including segmentation of tumor volumes, tumor characterization, tumor prognostication and treatment response, and prediction of metastatic lymph node disease. Head and neck oncology care is well positioned for the application of AI since treatment is guided by a wealth of information derived from CT, MRI, and PET imaging data. AI-based methods can integrate complex imaging, histologic, molecular, and clinical data to model tumor biology and behavior, and potentially identify associations, far beyond what conventional qualitative imaging can provide alone.

High-Resolution CT Imaging of the Temporal Bone: A Cadaveric Specimen Study.

Objective Super-high and ultra-high spatial resolution computed tomography (CT) imaging can be advantageous for detecting temporal bone pathology and guiding treatment strategies. Methods Six temporal bone cadaveric specimens were used to evaluate the temporal bone microanatomic structures utilizing the following CT reconstruction modes: normal resolution (NR, 0.5-mm slice thickness, 512 2 matrix), high resolution (HR, 0.5-mm slice thickness, 1,024 2 matrix), super-high resolution (SHR, 0.25-mm slice thickness, 1,024 2 matrix), and ultra-high resolution (UHR, 0.25-mm slice thickness, 2,048 2 matrix). Noise and signal-to-noise ratio (SNR) for bone and air were measured at each reconstruction mode. Two observers assessed visualization of seven small anatomic structures using a 4-point scale at each reconstruction mode. Results Noise was significantly higher and SNR significantly lower with increases in spatial resolution (NR, HR, and SHR). There was no statistical difference between SHR and UHR imaging with regard to noise and SNR. There was significantly improved visibility of all temporal bone osseous structures of interest with SHR and UHR imaging relative to NR imaging ( p < 0.001) and most of the temporal bone osseous structures relative to HR imaging. There was no statistical difference in the subjective image quality between SHR and UHR imaging of the temporal bone ( p ≥ 0.085). Conclusion Super-high-resolution and ultra-high-resolution CT imaging results in significant improvement in image quality compared with normal-resolution and high-resolution CT imaging of the temporal bone. This preliminary study also demonstrates equivalency between super-high and ultra-high spatial resolution temporal bone CT imaging protocols for clinical use.

Neuroimaging findings and pathophysiology of dorsal spinal arachnoid webs: illustrative case.

BACKGROUND: Spinal arachnoid webs are uncommon and difficult to diagnose, especially because causative intradural transverse bands of arachnoid tissue are radiographically occult. Left untreated, arachnoid webs may cause progressive, debilitating, and permanent neurological dysfunction. Conversely, more than 90% of patients may experience rapid neurological recovery after resection, even with a prolonged duration of presenting symptoms. Indirect imaging signs such as spinal cord indentation and compression with cerebrospinal fluid (CSF) flow alteration provide crucial diagnostic clues that are critical in guiding appropriate management of such patients. OBSERVATIONS: The authors reported a patient with no significant medical history who presented with back pain, progressive lower extremity weakness, gait ataxia, and bowel and bladder incontinence. They discussed multimodality imaging for determining the presence of arachnoid webs, including magnetic resonance imaging, phase-contrast CSF flow study, computed tomography myelography, and intraoperative ultrasound. They also discussed the detailed anatomy of the spinal subarachnoid space and a plausible pathophysiological mechanism for dorsal arachnoid webs. LESSONS: The authors report on a patient who underwent comprehensive imaging evaluation detailing the arachnoid web and whose subsequent anatomical localization and surgical treatment resulted in a full neurological recovery.

Triple-modality co-registered endoscope featuring wide-field reflectance imaging, and high-resolution multiphoton and optical coherence microscopy.

We present the design and feasibility testing of a multimodal co-registered endoscope based on a dual-path optical system integrated with a scanning piezo. This endoscope incorporates three different imaging modalities. A large field of view reflectance imaging system enables visualization of objects several millimeters in front of the endoscope, while optical coherence microscopy and multiphoton microscopy are employed in contact with tissue to further analyze suspicious areas. The optical system allows multiple different imaging modalities by employing a dual optical path. One path features a low numerical aperture and wide field of view to allow reflectance imaging of distant objects. The other path features a high numerical aperture and short working distance to allow microscopy techniques such as optical coherence microscopy and multiphoton microscopy. Images of test targets were obtained with each imaging modality to verify and characterize the imaging capabilities of the endoscope. The reflectance modality was demonstrated with a 561 nm laser to allow high contrast with blood vessels. It achieved a lateral resolution of 24.8 µm at 5 mm and a working distance from 5 mm to 30 mm. Optical coherence microscopy (OCM) was performed with a 1300 nm super-luminescent diode since this wavelength experiences low relative scattering to allow for deeper tissue imaging. Measured OCM lateral and axial resolution was 4.0 µm and 14.2 µm, respectively. Multiphoton microscopy (MPM) was performed with a custom 1400 nm femtosecond fiber laser, a wavelength suitable for exciting multiple exogenous and some endogenous fluorophores, as well as providing information on tissue composition through harmonic generation processes. A 4.0 µm MPM lateral resolution was measured.

Cochlear Meniere's: A Distinct Clinical Entity With Isolated Cochlear Hydrops on High-Resolution MRI?

Objective: Describe the clinical characteristics of patients with isolated cochlear endolymphatic hydrops (EH). Study design: Clinical case series. Setting: Tertiary Neurotology referral clinic. Patients: All subjects presenting to a University Neurotology clinic during a 1-year period from July 2015 until August 2016 who had isolated cochlear EH on MRI. Patients with a history of temporal bone surgery prior to the MRI were excluded. Intervention: High-resolution delayed-intravenous contrast MRI. Main outcome measures: Audiometric and vestibular testing, clinical history analysis. Results: 10 subjects demonstrated isolated, unilateral cochlear hydrops on MRI. None of these patients met the criteria for Meniere's disease. Mean age of the group was 66.4 years and most were males (70%). Unilateral aural fullness (70%), tinnitus (80%), and hearing loss (90%) were frequently observed. Only one patient presented with unsteadiness (10%) and one patient had a single isolated spell of positional vertigo 1 month prior to the MRI (10%) but no further vertigo spells in the 4 years following the MRI. The mean PTA was 37.8 dB which was significantly decreased from the non-affected ear with PTA of 17.9 (p < 0.001). One patient developed vertiginous spells and unsteadiness 4 years after initial presentation and a repeat MRI revealed progression to utricular, saccular and cochlear hydrops. Vestibular testing was obtained in five patients with one patient presenting with 50% caloric paresis and all others normal. The most common treatment tried was acetazolamide in seven patients with 86% reporting subjective clinical improvement. Two out of the 10 patients had a history of migraine (20%). Conclusions: Patients with MRI exhibiting isolated cochlear EH present with predominantly auditory symptoms: mild to moderate low-frequency hearing loss, aural fullness, tinnitus without significant vertigo. Isolated cochlear hydrops is more common in males, average age in mid-60's and there is a low comorbidity of migraine headaches. This contrasts significantly with patients with isolated saccular hydrops on MRI from our prior studies. We believe that isolated cochlear EH with hearing loss but no vertigo is distinct from Meniere's disease or its variant delayed endolymphatic hydrops. We propose that cochlear Meniere's disease represents a distinct clinical entity that could be a variant of Meniere's disease.

A modulator of the low-voltage-activated T-type calcium channel that reverses HIV glycoprotein 120-, paclitaxel-, and spinal nerve ligation-induced peripheral neuropathies.

The voltage-gated calcium channels CaV3.1-3.3 constitute the T-type subfamily, whose dysfunctions are associated with epilepsy, psychiatric disorders, and chronic pain. The unique properties of low-voltage-activation, faster inactivation, and slower deactivation of these channels support their role in modulation of cellular excitability and low-threshold firing. Thus, selective T-type calcium channel antagonists are highly sought after. Here, we explored Ugi-azide multicomponent reaction products to identify compounds targeting T-type calcium channel. Of the 46 compounds tested, an analog of benzimidazolonepiperidine-5bk (1-{1-[(R)-{1-[(1S)-1-phenylethyl]-1H-1,2,3,4-tetrazol-5-yl}(thiophen-3-yl)methyl]piperidin-4-yl}-2,3-dihydro-1H-1,3-benzodiazol-2-one) modulated depolarization-induced calcium influx in rat sensory neurons. Modulation of T-type calcium channels by 5bk was further confirmed in whole-cell patch clamp assays in dorsal root ganglion (DRG) neurons, where pharmacological isolation of T-type currents led to a time- and concentration-dependent regulation with a low micromolar IC50. Lack of an acute effect of 5bk argues against a direct action on T-type channels. Genetic knockdown revealed CaV3.2 to be the isoform preferentially modulated by 5bk. High voltage-gated calcium, as well as tetrodotoxin-sensitive and -resistant sodium, channels were unaffected by 5bk. 5bk inhibited spontaneous excitatory postsynaptic currents and depolarization-evoked release of calcitonin gene-related peptide from lumbar spinal cord slices. Notably, 5bk did not bind human mu, delta, or kappa opioid receptors. 5bk reversed mechanical allodynia in rat models of HIV-associated neuropathy, chemotherapy-induced peripheral neuropathy, and spinal nerve ligation-induced neuropathy, without effects on locomotion or anxiety. Thus, 5bk represents a novel T-type modulator that could be used to develop nonaddictive pain therapeutics.

Phosphorylated CRMP2 Regulates Spinal Nociceptive Neurotransmission.

The collapsin response mediator protein 2 (CRMP2) has emerged as a central node in assembling nociceptive signaling complexes involving voltage-gated ion channels. Concerted actions of post-translational modifications, phosphorylation and SUMOylation, of CRMP2 contribute to regulation of pathological pain states. In the present study, we demonstrate a novel role for CRMP2 in spinal nociceptive transmission. We found that, of six possible post-translational modifications, three phosphorylation sites on CRMP2 were critical for regulating calcium influx in dorsal root ganglion sensory neurons. Of these, only CRMP2 phosphorylated at serine 522 by cyclin-dependent kinase 5 (Cdk5) contributed to spinal neurotransmission in a bidirectional manner. Accordingly, expression of a non-phosphorylatable CRMP2 (S522A) decreased the frequency of spontaneous excitatory postsynaptic currents (sEPSCs), whereas expression of a constitutively phosphorylated CRMP2 (S522D) increased the frequency of sEPSCs. The presynaptic nature of CRMP2's actions was further confirmed by pharmacological antagonism of Cdk5-mediated CRMP2 phosphorylation with S-N-benzy-2-acetamido-3-methoxypropionamide ((S)-lacosamide; (S)-LCM) which (i) decreased sEPSC frequency, (ii) increased paired-pulse ratio, and (iii) reduced the presynaptic distribution of CaV2.2 and NaV1.7, two voltage-gated ion channels implicated in nociceptive signaling. (S)-LCM also inhibited depolarization-evoked release of the pro-nociceptive neurotransmitter calcitonin gene-related peptide (CGRP) in the spinal cord. Increased CRMP2 phosphorylation in rats with spared nerve injury (SNI) was decreased by intrathecal administration of (S)-LCM resulting in a loss of presynaptic localization of CaV2.2 and NaV1.7. Together, these findings indicate that CRMP2 regulates presynaptic excitatory neurotransmission in spinal cord and may play an important role in regulating pathological pain. Novel targeting strategies to inhibit CRMP2 phosphorylation by Cdk5 may have great potential for the treatment of chronic pain.

The Natural Flavonoid Naringenin Elicits Analgesia through Inhibition of NaV1.8 Voltage-Gated Sodium Channels.

Naringenin (2S)-5,7-dihydroxy-2-(4-hydroxyphenyl)-3,4-dihydro-2H-1-benzopyran-4-one is a natural flavonoid found in fruits from the citrus family. Because (2S)-naringenin is known to racemize, its bioactivity might be related to one or both enantiomers. Computational studies predicted that (2R)-naringenin may act on voltage-gated ion channels, particularly the N-type calcium channel (CaV2.2) and the NaV1.7 sodium channel-both of which are key for pain signaling. Here we set out to identify the possible mechanism of action of naringenin. Naringenin inhibited depolarization-evoked Ca2+ influx in acetylcholine-, ATP-, and capsaicin-responding rat dorsal root ganglion (DRG) neurons. This was corroborated in electrophysiological recordings from DRG neurons. Pharmacological dissection of each of the voltage-gated Ca2+ channels subtypes could not pinpoint any selectivity of naringenin. Instead, naringenin inhibited NaV1.8-dependent and tetrodotoxin (TTX)-resistant while sparing tetrodotoxin sensitive (TTX-S) voltage-gated Na+ channels as evidenced by the lack of further inhibition by the NaV1.8 blocker A-803467. The effects of the natural flavonoid were validated ex vivo in spinal cord slices where naringenin decreased both the frequency and amplitude of sEPSC recorded in neurons within the substantia gelatinosa. The antinociceptive potential of naringenin was evaluated in male and female mice. Naringenin had no effect on the nociceptive thresholds evoked by heat. Naringenin's reversed allodynia was in mouse models of postsurgical and neuropathic pain. Here, driven by a call by the National Center for Complementary and Integrative Health's strategic plan to advance fundamental research into basic biological mechanisms of the action of natural products, we advance the antinociceptive potential of the flavonoid naringenin.

Reversal of Peripheral Neuropathic Pain by the Small-Molecule Natural Product Physalin F via Block of CaV2.3 (R-Type) and CaV2.2 (N-Type) Voltage-Gated Calcium Channels.

No universally efficacious therapy exists for chronic pain, a disease affecting one-fifth of the global population. An overreliance on the prescription of opioids for chronic pain despite their poor ability to improve function has led to a national opioid crisis. In 2018, the NIH launched a Helping to End Addiction Long-term plan to spur discovery and validation of novel targets and mechanisms to develop alternative nonaddictive treatment options. Phytochemicals with medicinal properties have long been used for various treatments worldwide. The natural product physalin F, isolated from the Physalis acutifolia (family: Solanaceae) herb, demonstrated antinociceptive effects in models of inflammatory pain, consistent with earlier reports of its anti-inflammatory and immunomodulatory activities. However, the target of action of physalin F remained unknown. Here, using whole-cell and slice electrophysiology, competition binding assays, and experimental models of neuropathic pain, we uncovered a molecular target for physalin F's antinociceptive actions. We found that physalin F (i) blocks CaV2.3 (R-type) and CaV2.2 (N-type) voltage-gated calcium channels in dorsal root ganglion (DRG) neurons, (ii) does not affect CaV3 (T-type) voltage-gated calcium channels or voltage-gated sodium or potassium channels, (iii) does not bind G-protein coupled opioid receptors, (iv) inhibits the frequency of spontaneous excitatory postsynaptic currents (EPSCs) in spinal cord slices, and (v) reverses tactile hypersensitivity in models of paclitaxel-induced peripheral neuropathy and spinal nerve ligation. Identifying CaV2.2 as a molecular target of physalin F may spur its use as a tool for mechanistic studies and position it as a structural template for future synthetic compounds.

Normal cellular prion protein with a methionine at position 129 has a more exposed helix 1 and is more prone to aggregate.

The human prion gene, PRNP, has two allelic forms that encode either a methionine or valine at codon 129. This polymorphism strongly influences the pathogenesis of prion disease. However, the underlying mechanism remains unclear. We compared the conformation between wild-type human prion protein (rPrP(C)) with either a valine or methionine at position 129, using a panel of monoclonal antibodies that are specific for epitopes along the entire protein. We found that rPrP(C(129M)) has a more exposed helix 1 region compared to rPrP(C(129V)). Helix 1 is important in the aggregation process. Accordingly, rPrP(C(129M)) aggregates at a faster rate and forms more aggregate than rPrP(C(129V)). In addition, by using a rPrP with a pathogenic mutation of five additional octapeptide repeat insertions, rPrP((129M)/10OR), as "seeds", we showed that rPrP((129M)/10OR) promotes the aggregation of rPrP(C(129M)) more efficiently than rPrP(C(129V)). These findings provide a possible mechanism underlying the influence of residue 129 on human prion disease.

HLA-DR3 transgenic mice immunized with adenovirus encoding the thyrotropin receptor: T cell epitopes and functional analysis of the CD40 Graves' polymorphism.

The major histocompatibility (MHC) molecule HLA-DR3 is a susceptibility gene for Graves' disease (GD) in Caucasians. Mice lacking murine MHC and expressing human HLA-DR3 develop thyrotropin receptor (TSHR) antibodies and sometimes hyperthyroidism after vaccination with TSHR-DNA. MHC molecules present peptides processed from antigens to T cells. Therefore, we used DR3-transgenic mice to investigate recognition of TSHR ectodomain peptides. After immunization with TSHR A-subunit adenovirus (A-subunit-Ad) but not control-adenovirus (Control-Ad), splenocytes from DR3 mice responded to A-subunit protein in culture by producing interferon-gamma (IFN-gamma). When challenged with 29 overlapping TSHR peptides, splenocytes from A-subunit-Ad- or Control-Ad-immunized mice responded to several peptides. However, in splenocytes from A-subunit-Ad but not Control-Ad mice, a peptide containing TSHR residues 142-161 induced significantly more IFN-gamma than the same splenocytes in medium alone. Immunized DR3 mice also permitted testing the TSHR-specific function of the CD40 single nucleotide polymorphism (C vs. T) associated with GD. Of three human DR3 human Epstein-Barr virus lines (EBVL), two had C in both alleles (CC) and one was CT. However, these EBVL presented peptides poorly and there was no difference between CC vs. CT EBVL in peptide presentation to splenocytes from immunized mice. A peptide corresponding to residues 145-163 is one of seven suggested to be important in GD based on HLA-binding affinities, T-epitope algorithms, and human studies. Consequently, as in human GD, TSHR amino acids 142-161 appear to include a major T cell epitope in HLA-DR3 transgenic mice immunized with A-subunit-Ad.

Relationship between thyroid peroxidase T cell epitope restriction and antibody recognition of the autoantibody immunodominant region in human leukocyte antigen DR3 transgenic mice.

We investigated the relationship between thyroid peroxidase (TPO) antibody and T lymphocyte epitopes in TPO-adenovirus (TPO-Ad) immunized BALB/c mice and mice transgenic for the human class II molecule DR3 associated with human thyroid autoimmunity. TPO autoantibodies are largely restricted to an immunodominant region (IDR). BALB/c mice immunized with fewer (10(7) vs. 10(9)) TPO-Ad particles developed TPO antibodies with lower titers that displayed greater restriction to the IDR. However, as with higher-dose TPO-Ad immunization, T cell epitopes (assessed by splenocyte interferon-gamma response to TPO in vitro) were highly diverse and variable in different animals. In contrast, DR3 mice immunized the higher TPO-Ad dose (10(9) particles) had high TPO antibody levels that showed relative focus on the IDR. Moreover, T cell epitopes recognized by splenocytes from DR3 mice showed greater restriction than BALB/c mice. Antibody affinities for TPO were higher in DR3 than in BALB/c mice. The present study indicates that weak TPO-Ad immunization of BALB/c mice (with consequent low TPO antibody titers) is required for enhanced IDR focus yet is not associated with T cell epitopic restriction. Humanized DR3 transgenic mice, despite stronger TPO-Ad immunization, develop higher titer TPO antibodies that do focus on the autoantibody IDR with T cells that recognize a more limited range of TPO peptides. These data suggest a relationship between major histocompatibility complex class II molecules and the development of antibodies to the IDR, a feature of human thyroid autoimmunity.