Microanatomy of the human tunnel of Corti structures and cochlear partition-tonotopic variations and transcellular signaling.

Auditory sensitivity and frequency resolution depend on the optimal transfer of sound-induced vibrations from the basilar membrane (BM) to the inner hair cells (IHCs), the principal auditory receptors. There remains a paucity of information on how this is accomplished along the frequency range in the human cochlea. Most of the current knowledge is derived either from animal experiments or human tissue processed after death, offering limited structural preservation and optical resolution. In our study, we analyzed the cytoarchitecture of the human cochlear partition at different frequency locations using high-resolution microscopy of uniquely preserved normal human tissue. The results may have clinical implications and increase our understanding of how frequency-dependent acoustic vibrations are carried to human IHCs. A 1-micron-thick plastic-embedded section (mid-modiolar) from a normal human cochlea uniquely preserved at lateral skull base surgery was analyzed using light and transmission electron microscopy (LM, TEM). Frequency locations were estimated using synchrotron radiation phase-contrast imaging (SR-PCI). Archival human tissue prepared for scanning electron microscopy (SEM) and super-resolution structured illumination microscopy (SR-SIM) were also used and compared in this study. Microscopy demonstrated great variations in the dimension and architecture of the human cochlear partition along the frequency range. Pillar cell geometry was closely regulated and depended on the reticular lamina slope and tympanic lip angle. A type II collagen-expressing lamina extended medially from the tympanic lip under the inner sulcus, here named "accessory basilar membrane." It was linked to the tympanic lip and inner pillar foot, and it may contribute to the overall compliance of the cochlear partition. Based on the findings, we speculate on the remarkable microanatomic inflections and geometric relationships which relay different sound-induced vibrations to the IHCs, including their relevance for the evolution of human speech reception and electric stimulation with auditory implants. The inner pillar transcellular microtubule/actin system's role of directly converting vibration energy to the IHC cuticular plate and ciliary bundle is highlighted.

Development and validation of a surgical planning tool for bone-conduction implants.

Background: The BONEBRIDGE® (Med-El GmbH) is a bone-conduction device comprising an external audio processor and an internal Bone Conduction-Floating Mass Transducer (BC-FMT) surgically anchored to the temporal bone. Due to the implant's size, its placement may be challenging in certain anatomies, necessitating thorough surgical planning. Manual planning methods are laborious, time-intensive, and prone to errors. This study aimed to develop and validate an automated algorithm for determining skull thickness, aiding in the surgical planning of the BONEBRIDGE and other devices requiring similar bone thickness estimations. Materials and methods: Twelve cadaveric temporal bones underwent clinical computed tomography (CT). A custom Python algorithm was developed to automatically segment bone from soft tissue, generate 3D models, and perform ray-tracing to estimate bone thickness. Two thickness colormaps were generated for each sample: the cortical thickness to the first air cell and the total thickness down to the dura. The algorithm was validated against expert manual measurements to achieve consensus interpretation. Results: The algorithm estimated bone-to-air thicknesses (mean = 4.7 mm, 95% Confidence Interval [CI] of 4.3-5.0 mm) that closely matched the expert measurements (mean = 4.7 mm, CI of 4.4-5.0 mm), with a mean absolute difference (MAD) of 0.3 mm. Similarly, the algorithm's estimations to the dura (6.0 mm, CI of 5.4-6.5 mm) were comparable to the expert markings (5.9 mm, CI of 5.4-6.5 mm), with a MAD of 0.3 mm. Conclusions: The first automated algorithm to calculate skull thickness to both the air cells and dura in the temporal bone was developed. Colormaps were optimized to aid with the surgical planning of BONEBRIDGE implantation, however the tool can be generalized to aid in the surgical planning of any bone thickness application. The tool was published as a freely available extension to the open-source 3D Slicer software program (www.slicer.org).

Morphologic Analysis of the Scala Tympani Using Synchrotron: Implications for Cochlear Implantation.

OBJECTIVES: To use synchrotron radiation phase-contrast imaging (SR-PCI) to visualize and measure the morphology of the entire cochlear scala tympani (ST) and assess cochlear implant (CI) electrode trajectories. METHODS: SR-PCI images were used to obtain geometric measurements of the cochlear scalar diameter and area at 5-degree increments in 35 unimplanted and three implanted fixed human cadaveric cochleae. RESULTS: The cross-sectional diameter and area of the cochlea were found to decrease from the base to the apex. This study represents a wide variability in cochlear morphology and suggests that even in the smallest cochlea, the ST can accommodate a 0.4 mm diameter electrode up to 720°. Additionally, all lateral wall array trajectories were within the anatomically accommodating insertion zone. CONCLUSION: This is the first study to use SR-PCI to visualize and quantify the entire ST morphology, from the round window to the apical tip, and assess the post-operative trajectory of electrodes. These high-resolution anatomical measurements can be used to inform the angular insertion depth that can be accommodated in CI patients, accounting for anatomical variability. LEVEL OF EVIDENCE: N/A. Laryngoscope, 134:2889-2897, 2024.

Virtual Reality Simulation for the Middle Cranial Fossa Approach: A Validation Study.

BACKGROUND AND OBJECTIVES: Virtual reality (VR) surgical rehearsal is an educational tool that exists in a safe environment. Validation is necessary to establish the educational value of this platform. The middle cranial fossa (MCF) is ideal for simulation because trainees have limited exposure to this approach and it has considerable complication risk. Our objectives were to assess the face, content, and construct validities of an MCF VR simulation, as well as the change in performance across serial simulations. METHODS: Using high-resolution volumetric data sets of human cadavers, the authors generated a high-fidelity visual and haptic rendering of the MCF approach using CardinalSim software. Trainees from Neurosurgery and Otolaryngology-Head and Neck Surgery at two Canadian academic centers performed MCF dissections on this VR platform. Randomization was used to assess the effect of enhanced VR interaction. Likert scales were used to assess the face and content validities. Performance metrics and pre- and postsimulation test scores were evaluated. Construct validity was evaluated by examining the effect of the training level on simulation performance. RESULTS: Twenty trainees were enrolled. Face and content validities were achieved in all domains. Construct validity, however, was not demonstrated. Postsimulation test scores were significantly higher than presimulation test scores ( P < .001 ). Trainees demonstrated statistically significant improvement in the time to complete dissections ( P < .001 ), internal auditory canal skeletonization ( P < .001 ), completeness of the anterior petrosectomy ( P < .001 ), and reduced number of injuries to critical structures ( P = .001 ). CONCLUSION: This MCF VR simulation created using CardinalSim demonstrated face and content validities. Construct validity was not established because no trainee included in the study had previous MCF approach experience, which further emphasizes the importance of simulation. When used as a formative educational adjunct in both Neurosurgery and Otolaryngology-Head and Neck Surgery, this simulation has the potential to enhance understanding of the complex anatomic relationships of critical neurovascular structures.

Correlating clinical breakpoint concentration of moxifloxacin with gyrA mutations using the GenoType MTBDRsl assay Version 2.0.

INTRODUCTION: Widespread use of Fluoroquinolones (FQs) has led to the development of its resistance in clinical isolates of Mycobacterium tuberculosis. However, in Mycobacterium tuberculosis, phenotypic resistance to FQs has been shown to be heterogeneous, ranging from low-level resistance to high-level resistance. This stratification in resistance has important implications for the inclusion of moxifloxacin (Mfx) in the treatment regimen. The World Health Organization recommends the use of GenoType MTBDRsl assay as the initial test for detecting resistance conferring mutations (both high and low) to FQs in patients with confirmed MDR-RR TB. The present study was conducted to explore the relationship of MTBDRsl Version 2.0 detected mutations in gyrA gene and genotypic DST of Mfx at WHO defined Clinical Breakpoint (CB). MATERIALS AND METHODS: A total of 200 sputum samples from Confirmed MDR/RR TB patients were included in this study. All of these samples had mutations conferring resistance to FQ confirmed by GenoType MTBDRsl assay. These samples were further subjected to Phenotypic DST against moxifloxacin using the Bactec MGIT-960 system. RESULTS: All of the 200 representative FQ resistant isolates had mutations in gyrA gene only with no detectable mutation in gyrB gene. 109 (54.5%) of the isolates had mutations associated with high-level increase in MIC while 91 (45.5%) isolates had mutations associated with low-level increase in MIC. Phenotypic DST of these 200 isolates against Mfx at CB (1.0µg/ml) revealed that of the 109 isolates with mutations associated with high-level increase in MIC and expected to be resistant at CB, only 34 (31.2%) were resistant and the remaining 75 (68.8%) were sensitive. CONCLUSION: Moxifloxacin is an important drug in the regimen for treating Drug-resistant TB and the decision to exclude this drug from the regimen should not be taken merely on the basis of mutational patterns. It should rather be taken after considering the combined results of mutational analysis and phenotypic DST.

Influence of the Frequency-to-Place Function on Recognition with Place-Based Cochlear Implant Maps.

OBJECTIVE: Comparison of acute speech recognition for cochlear implant (CI) alone and electric-acoustic stimulation (EAS) users listening with default maps or place-based maps using either a spiral ganglion (SG) or a new Synchrotron Radiation-Artificial Intelligence (SR-AI) frequency-to-place function. METHODS: Thirteen adult CI-alone or EAS users completed a task of speech recognition at initial device activation with maps that differed in the electric filter frequency assignments. The three map conditions were: (1) maps with the default filter settings (default map), (2) place-based maps with filters aligned to cochlear SG tonotopicity using the SG function (SG place-based map), and (3) place-based maps with filters aligned to cochlear Organ of Corti (OC) tonotopicity using the SR-AI function (SR-AI place-based map). Speech recognition was evaluated using a vowel recognition task. Performance was scored as the percent correct for formant 1 recognition due to the rationale that the maps would deviate the most in the estimated cochlear place frequency for low frequencies. RESULTS: On average, participants had better performance with the OC SR-AI place-based map as compared to the SG place-based map and the default map. A larger performance benefit was observed for EAS users than for CI-alone users. CONCLUSION: These pilot data suggest that EAS and CI-alone users may experience better performance with a patient-centered mapping approach that accounts for the variability in cochlear morphology (OC SR-AI frequency-to-place function) in the individualization of the electric filter frequencies (place-based mapping procedure). LEVEL OF EVIDENCE: 3 Laryngoscope, 133:3540-3547, 2023.

Factors Associated with 30-day Major Adverse Cardiovascular Event in Acute Coronary Syndrome Patients with Non-Dialysis Chronic Kidney Disease: A Retrospective Cohort Study.

BACKGROUND: Acute coronary syndrome (ACS) accounts for the majority of ischemic heart disease-related deaths. It is known that ACS patients with chronic kidney disease (CKD) tend to have worse clinical outcomes, including major adverse coronary events (MACE) compared to patients without CKD.  Some studies suggested that several determinant factors may be involved in this condition. Until now, research on determinant factors of MACE in ACS patients with CKD in Indonesia is still limited. Thus, we aimed to investigate the relationship of various factors to MACE in ACS patients with non-dialysis CKD who underwent percutaneous coronary intervention (PCI), in the form of neutrophile leukocyte ratio (NLR) as a factor describing chronic inflammation, left ventricular hypertrophy (LVH) as a factor describing cardiac remodeling, Gensini score may represent coronary severity, whereas GRACE was used to evaluate the severity and clinical risk of ACS patients. METHODS: This study is a retrospective cohort study using secondary data from the medical records of 117 ACS patients who underwent percutaneous coronary intervention (PCI) at Cipto Mangunkusumo General Hospital Jakarta from January 2018 to June 2018 . Patients were classified based on the stage of CKD and assessed for 30-day MACE. Data were recorded on GRACE score, Gensini score, LVH, and neutrophil-lymphocyte ratio (NLR). Analysis of the relationship between these factors was carried out using the chi-square test. RESULTS: Of the 117 patients, 62.3% were STEMI. At the end of hospital treatment, 67.5% were in the normal-stage 2 CKD group, 17.1% in the CKD stage 3a-3b group, and 15.4% in the CKD stage 4-5 group. MACE occurred in 47 (40.2%) patients with 17 (14.5%) dying. There was a significant relationship between GRACE scores and MACE (54.8% MACE at high GRACE scores vs. 32% MACE at low-moderate GRACE scores, p = 0.016, OR: 2,57 CI 95%, 1,18-5,59), while no significant relationship was found for the Gensini score, LVH, and NLR scores even though there was an increase in the proportion of MACE. CONCLUSION: The incidence of MACE is higher than in the previous studies conducted in the same place, i.e. Cipto Mangunkusumo General Hospital, no significant relationship is found in NLR, LVH, and Gensini score with the 30-day MACE of ACS patients with non-dialysis CKD, meanwhile the GRACE score correlates with the 30-day MACE of ACS in non-dialysis CKD patients as is the known theory regarding this score.

Finite element modelling of the human middle ear using synchrotron-radiation phase-contrast imaging.

Finite element (FE) models of the middle ear often lack accurate geometry of soft tissue structures, such as the suspensory ligaments, as they can be difficult to discern using conventional imaging modalities, such as computed tomography. Synchrotron-radiation phase-contrast imaging (SR-PCI) is a non-destructive imaging modality that has been shown to produce excellent visualization of soft tissue structures without the need for extensive sample preparation. The objectives of the investigation were to firstly use SR-PCI to create and evaluate a biomechanical FE model of the human middle ear that includes all soft tissue structures, and secondly, to investigate how modelling assumptions and simplifications of ligament representations affect the simulated biomechanical response of the FE model. The FE model included the suspensory ligaments, ossicular chain, tympanic membrane, the incudostapedial and incudomalleal joints, and the ear canal. Frequency responses obtained from the SR-PCI-based FE model agreed well with published laser doppler vibrometer measurements on cadaveric samples. Revised models with exclusion of the superior malleal ligament (SML), simplification of the SML, and modification of the stapedial annular ligament were studied, as these revised models represented modelling assumptions that have been made in literature.

Functional and Speech Outcome of Unilaterally Cochlear Implanted Children after 1 year of Implantation in NIENT: A Retrospective Study.

Hearing loss can be congenital or acquired. Most of the time congenital hearing loss means profound sensorineural hearing loss. The invention of cochlear implantation has already an established role in improving the hearing capability in congenital sensorineural hearing loss. To speak properly after cochlear implant, one needs to go for speech therapy or audio-verbal therapy. Speech therapy has become an integral part of the cochlear implantation program. In Bangladesh cochlear implantation (CI) programs started around 15 years ago, which was limited to Bangabandhu Sheikh Mujib Medical University (BSMMU) and Combined Military Hospital (CMH). Our government takes initiative to provide this service to poor and lower income people. National Institute of ENT (NIENT) was the first government hospital where CI started in 2018 and the project is now ongoing. After CI auditory, verbal therapy (AVT) was given at the same institute. No study has not been done to find out the outcome of AVT. NIENT has taken steps to find out the functional and speech outcome of cochlear implanted children considering the two standard criteria: CAP and SIR score. This retrospective study was done to find out the auditory verbal outcome of unilaterally CI children in different ages and conducted at department of Audiology, National Institute of ENT, Tejgaon, Dhaka, Bangladesh from January 2021 to June 2021. The children who underwent cochlear implants and took the auditory verbal therapy were included in this study. Though it is a retrospective study, a written informed consent has been taken from the patient's parents as the cases are minor in age. From the record review, CAP and SIR scores were recorded four times: before implantation 1 time, after cochlear implantation 3 times (At the end of 3, 6 and 12 months after starting of AVT of Cochlear implantees). From the study it has been observed that out of 50 implantees, 48.0% of children's age range was in between >3 to ≤5 years. Most of the implantees were from rural areas and also from lower income group. The study showed significant association with CAP and SIR score at the end of one year of AVT. No significant association was found in different age groups with scores of CAP and SIR. But the parent's education showed significant association with CAP score at the end one year of AVT. Though the age of the recipient is not significantly associated with the AVT outcome but maternal education plays an important role in improving CAP and SIR score. So, maternal education or understanding along with professional speech therapy is vital for better AVT outcome.

Effectiveness of Xpert MTB/RIF for the diagnosis of extrapulmonary tuberculosis at various stand-alone laboratories in Delhi.

BACKGROUND: Globally, EPTB accounts for 15% of the notified incident TB cases. Laboratory confirmation of EPTB is challenging and majority of the cases remain undetected for a longer time. A major breakthrough in the diagnosis of EPTB was the introduction of nucleic acid amplification tests (NAAT). One such test-the Xpert MTB/RIF assay also known as Cartridge based nucleic acid amplification test (CBNAAT) was endorsed by the Scientific and Technical Advisory Board of the WHO for the diagnosis of Tuberculosis. The present study was conduct to evaluate the outcome of various extrapulmonary samples tested in the year 2019 at different standalone NAAT laboratories in Delhi. MATERIALS AND METHODS: A total of 20,238 samples consisting mainly of Pus (21.77%), Cerebrospinal fluid (CSF) (14.96%), Biopsies (13.87%), Pleural fluid (10.49%), Lymph node aspirations (FNAC aspirates) (6.75%), synovial fluid (0.54%) and gastric aspirates (26.4%) tested at 22 standalone NAAT laboratories were included in this study. RESULTS: Mycobacterium tuberculosis was detected in 3496 samples and resistance to rifampicin was detected in 329 of the samples. The overall yield of all the specimens combined was 17.2%. Highest yield was seen in Lymph nodes aspirates (FNAC) (36.0%), followed by pus (35.4%), tissues (15.7%), synovial fluid (13.5%), Endometrial tissues (10.7%), Pleural fluid (9.5%), Gastric aspirates (9.4%) and CSF (6.5%). The lowest yield was seen in Cavitary fluids (6.2%). CONCLUSION: The results of this study highlight the usefulness of Xpert MTB/RIF assay in the diagnosis of EPTB. In particular, this assay proved to be of great utility while testing pus samples, tissue samples and lymph node FNACs.

Unlocking the human inner ear for therapeutic intervention.

The human inner ear contains minute three-dimensional neurosensory structures that are deeply embedded within the skull base, rendering them relatively inaccessible to regenerative therapies for hearing loss. Here we provide a detailed characterisation of the functional architecture of the space that hosts the cell bodies of the auditory nerve to make them safely accessible for the first time for therapeutic intervention. We used synchrotron phase-contrast imaging which offers the required microscopic soft-tissue contrast definition while simultaneously displaying precise bony anatomic detail. Using volume-rendering software we constructed highly accurate 3-dimensional representations of the inner ear. The cell bodies are arranged in a bony helical canal that spirals from the base of the cochlea to its apex; the canal volume is 1.6 µL but with a diffusion potential of 15 µL. Modelling data from 10 temporal bones enabled definition of a safe trajectory for therapeutic access while preserving the cochlea's internal architecture. We validated the approach through surgical simulation, anatomical dissection and micro-radiographic analysis. These findings will facilitate future clinical trials of novel therapeutic interventions to restore hearing.

High-Fidelity Virtual Reality Simulation for the Middle Cranial Fossa Approach-Modules for Surgical Rehearsal and Education.

BACKGROUND: Virtual reality simulation has gained prominence as a valuable surgical rehearsal and education tool in neurosurgery. Approaches to the internal auditory canal, cerebellopontine angle, and ventral brainstem region using the middle cranial fossa are not well explored by simulation. OBJECTIVE: We hope to contribute to this paucity in simulation tools devoted to the lateral skull base, specifically the middle cranial fossa approach. METHODS: Eight high-resolution microcomputed tomography scans of human cadavers were used as volumetric data sets to construct a high-fidelity visual and haptic rendering of the middle cranial fossa using CardinalSim software. Critical neurovascular structures related to this region of the skull base were segmented and incorporated into the modules. RESULTS: The virtual models illustrate the 3-dimensional anatomic relationships of neurovascular structures in the middle cranial fossa and allow a realistic interactive drilling environment. This is facilitated by the ability to render bone opaque or transparent to reveal the proximity to critical anatomy allowing for practice of the virtual dissection in a graduated fashion. CONCLUSION: We have developed a virtual library of middle cranial fossa approach models, which integrate relevant neurovascular structures with aims to improve surgical training and education. A ready extension is the potential for patient-specific application and pathology.

Machine learning approaches used to analyze auditory evoked responses from the human auditory brainstem: A systematic review.

BACKGROUND: The application of machine learning algorithms for assessing the auditory brainstem response has gained interest over recent years with a considerable number of publications in the literature. In this systematic review, we explore how machine learning has been used to develop algorithms to assess auditory brainstem responses. A clear and comprehensive overview is provided to allow clinicians and researchers to explore the domain and the potential translation to clinical care. METHODS: The systematic review was performed based on PRISMA guidelines. A search was conducted of PubMed, IEEE-Xplore, and Scopus databases focusing on human studies that have used machine learning to assess auditory brainstem responses. The duration of the search was from January 1, 1990, to April 3, 2021. The Covidence systematic review platform (www.covidence.org) was used throughout the process. RESULTS: A total of 5812 studies were found through the database search and 451 duplicates were removed. The title and abstract screening process further reduced the article count to 89 and in the proceeding full-text screening, 34 articles met our full inclusion criteria. CONCLUSION: Three categories of applications were found, namely neurologic diagnosis, hearing threshold estimation, and other (does not relate to neurologic or hearing threshold estimation). Neural networks and support vector machines were the most commonly used machine learning algorithms in all three categories. Only one study had conducted a clinical trial to evaluate the algorithm after development. Challenges remain in the amount of data required to train machine learning models. Suggestions for future research avenues are mentioned with recommended reporting methods for researchers.

Standard Binding Free Energy and Membrane Desorption Mechanism for a Phospholipase C.

Peripheral membrane proteins (PMPs) bind temporarily to cellular membranes and play important roles in signaling, lipid metabolism, and membrane trafficking. Obtaining accurate membrane-PMP affinities using experimental techniques is more challenging than for protein-ligand affinities in an aqueous solution. At the theoretical level, calculation of the standard protein-membrane binding free energy using molecular dynamics simulations remains a daunting challenge owing to the size of the biological objects at play, the slow lipid diffusion, and the large variation in configurational entropy that accompanies the binding process. To overcome these challenges, we used a computational framework relying on a series of potential-of-mean-force (PMF) calculations including a set of geometrical restraints on collective variables. This methodology allowed us to determine the standard binding free energy of a PMP to a phospholipid bilayer using an all-atom force field. Bacillus thuringiensis phosphatidylinositol-specific phospholipase C (BtPI-PLC) was chosen due to its importance as a virulence factor and owing to the host of experimental affinity data available. We computed a standard binding free energy of -8.2 ± 1.4 kcal/mol in reasonable agreement with the reported experimental values (-6.6 ± 0.2 kcal/mol). In light of the 2.3-µs separation PMF calculation, we investigated the mechanism whereby BtPI-PLC disengages from interactions with the lipid bilayer during separation. We describe how a short amphipathic helix engages in transitory interactions to ease the passage of its hydrophobes through the interfacial region upon desorption from the bilayer.

Specificity of Loxosceles α clade phospholipase D enzymes for choline-containing lipids: Role of a conserved aromatic cage.

Spider venom GDPD-like phospholipases D (SicTox) have been identified to be one of the major toxins in recluse spider venom. They are divided into two major clades: the α clade and the ß clade. Most α clade toxins present high activity against lipids with choline head groups such as sphingomyelin, while activities in ß clade toxins vary and include preference for substrates containing ethanolamine headgroups (Sicarius terrosus, St_ßIB1). A structural comparison of available structures of phospholipases D (PLDs) reveals a conserved aromatic cage in the α clade. To test the potential influence of the aromatic cage on membrane-lipid specificity we performed molecular dynamics (MD) simulations of the binding of several PLDs onto lipid bilayers containing choline headgroups; two SicTox from the α clade, Loxosceles intermedia αIA1 (Li_αIA) and Loxosceles laeta αIII1 (Ll_αIII1), and one from the ß clade, St_ßIB1. The simulation results reveal that the aromatic cage captures a choline-headgroup and suggest that the cage plays a major role in lipid specificity. We also simulated an engineered St_ßIB1, where we introduced the aromatic cage, and this led to binding with choline-containing lipids. Moreover, a multiple sequence alignment revealed the conservation of the aromatic cage among the α clade PLDs. Here, we confirmed that the i-face of α and ß clade PLDs is involved in their binding to choline and ethanolamine-containing bilayers, respectively. Furthermore, our results suggest a major role in choline lipid recognition of the aromatic cage of the α clade PLDs. The MD simulation results are supported by in vitro liposome binding assay experiments.

The voltage-sensing domain of a hERG1 mutant is a cation-selective channel.

A cationic leak current known as an "omega current" may arise from mutations of the first charged residue in the S4 of the voltage sensor domains of sodium and potassium voltage-gated channels. The voltage-sensing domains (VSDs) in these mutated channels act as pores allowing nonspecific passage of cations, such as Li+, K+, Cs+, and guanidinium. Interestingly, no omega currents have been previously detected in the nonswapped voltage-gated potassium channels such as the human-ether-a-go-go-related (hERG1), hyperpolarization-activated cyclic nucleotide-gated, and ether-a-go-go channels. In this work, we discovered a novel omega current by mutating the first charged residue of the S4 of the hERG1, K525 to serine. To characterize this omega current, we used various probes, including the hERG1 pore domain blocker, dofetilide, to show that the omega current does not require cation flux via the canonical pore domain. In addition, the omega flux does not cross the conventional selectivity filter. We also show that the mutated channel (K525S hERG1) conducts guanidinium. These data are indicative of the formation of an omega current channel within the VSD. Using molecular dynamics simulations with replica-exchange umbrella sampling simulations of the wild-type hERG1 and the K525S hERG1, we explored the molecular underpinnings governing the cation flow in the VSD of the mutant. We also show that the wild-type hERG1 may form water crevices supported by the biophysical surface accessibility data. Overall, our multidisciplinary study demonstrates that the VSD of hERG1 may act as a cation-selective channel wherein a mutation of the first charged residue in the S4 generates an omega current. Our simulation uncovers the atomistic underpinning of this mechanism.

Spectrum of Drug Resistance in Musculoskeletal Tuberculosis.

BACKGROUND: Very few studies report resistance pattern exclusively in musculoskeletal tuberculosis (MSK-TB). METHODS: This study of 100 pus samples from patients of MSK-TB with active disease in whom Mycobacterium tuberculosis (MTB) was detected by cartridge-based nucleic acid amplification test (CBNAAT), revealed the pattern of resistance among newly diagnosed and previously treated cases. Liquid culture and drug susceptibility testing (DST) using MGIT 960 was done for 11 anti-tubercular drugs. RESULTS: Among these 100 cases; 22% were AFB positive; MGIT 960 detected MTB in 58.33% (35/60) new cases and 30.0% (12/40) previously treated cases. Five new and 10 previously treated cases had drug resistance and 12 were detected rifampicin resistance (Rif-R) by CBNAAT. Among new cases MGIT-DST detected mono-INH resistant in 2.86% (1/35), mono-STR resistant in 2.86% (1/35), MDR-TB in 5.7% (2/35) and pre-XDR in 2.9%(1/35).Among previously treated cases Rif-R was found in 10% (4/40) where MTB was not detected by MGIT and MGIT-DST detected mono-INH resistant in 8.33% (1/12); MDR-TB in 8.33% (1/12) and pre-XDR in 33.3%. There were no cases of XDR-TB. CONCLUSION: High disease burden of various type drug resistance were seen more commonly in previously treated cases and was not uncommon in new cases of MSK-TB. Both CBNAAT and DST are essential for detecting resistance pattern in MSK-TB.

Phospholipids in Motion: High-Resolution 31P NMR Field Cycling Studies.

Diverse phospholipid motions are key to membrane function but can be quite difficult to untangle and quantify. High-resolution field cycling 31P NMR spin-lattice relaxometry, where the sample is excited at high field, shuttled in the magnet bore for low-field relaxation, then shuttled back to high field for readout of the residual magnetization, provides data on phospholipid dynamics and structure. This information is encoded in the field dependence of the 31P spin-lattice relaxation rate (R1). In the field range from 11.74 down to 0.003 T, three dipolar nuclear magnetic relaxation dispersions (NMRDs) and one due to 31P chemical shift anisotropy contribute to R1 of phospholipids. Extraction of correlation times and maximum relaxation amplitudes for these NMRDs provides (1) lateral diffusion constants for different phospholipids in the same bilayer, (2) estimates of how additives alter the motion of the phospholipid about its long axis, and (3) an average 31P-1H angle with respect to the bilayer normal, which reveals that polar headgroup motion is not restricted on a microsecond timescale. Relative motions within a phospholipid are also provided by comparing 31P NMRD profiles for specifically deuterated molecules as well as 13C and 1H field dependence profiles to that of 31P. Although this work has dealt exclusively with phospholipids in small unilamellar vesicles, these same NMRDs can be measured for phospholipids in micelles and nanodisks, making this technique useful for monitoring lipid behavior in a variety of structures and assessing how additives alter specific lipid motions.

A Synchrotron and Micro-CT Study of the Human Endolymphatic Duct System: Is Meniere's Disease Caused by an Acute Endolymph Backflow?

Background: The etiology of Meniere's disease (MD) and endolymphatic hydrops believed to underlie its symptoms remain unknown. One reason may be the exceptional complexity of the human inner ear, its vulnerability, and surrounding hard bone. The vestibular organ contains an endolymphatic duct system (EDS) bridging the different fluid reservoirs. It may be essential for monitoring hydraulic equilibrium, and a dysregulation may result in distension of the fluid spaces or endolymphatic hydrops. Material and Methods: We studied the EDS using high-resolution synchrotron phase contrast non-invasive imaging (SR-PCI), and micro-computed tomography (micro-CT). Ten fresh human temporal bones underwent SR-PCI. One bone underwent micro-CT after fixation and staining with Lugol's iodine solution (I2KI) to increase tissue resolution. Data were processed using volume-rendering software to create 3D reconstructions allowing orthogonal sectioning, cropping, and tissue segmentation. Results: Combined imaging techniques with segmentation and tissue modeling demonstrated the 3D anatomy of the human saccule, utricle, endolymphatic duct, and sac together with connecting pathways. The utricular duct (UD) and utriculo-endolymphatic valve (UEV or Bast's valve) were demonstrated three-dimensionally for the first time. The reunion duct was displayed with micro-CT. It may serve as a safety valve to maintain cochlear endolymph homeostasis under certain conditions. Discussion: The thin reunion duct seems to play a minor role in the exchange of endolymph between the cochlea and vestibule under normal conditions. The saccule wall appears highly flexible, which may explain occult hydrops occasionally preceding symptoms in MD on magnetic resonance imaging (MRI). The design of the UEV and connecting ducts suggests that there is a reciprocal exchange of fluid among the utricle, semicircular canals, and the EDS. Based on the anatomic framework and previous experimental data, we speculate that precipitous vestibular symptoms in MD arise from a sudden increase in endolymph pressure caused by an uncontrolled endolymphatic sac secretion. A rapid rise in UD pressure, mediated along the fairly wide UEV, may underlie the acute vertigo attack, refuting the rupture/K+-intoxication theory.