Juxtametallic Bipolar Bone Radiofrequency Ablation: Thermal Monitoring in an Ex-Vivo Model with Specimen MRI and Histopathologic Correlation.

PURPOSE: To measure the ablation zone temperature and nontarget tissue temperature during radiofrequency (RF) ablation in bone containing metal instrumentation versus no metal instrumentation (control group). MATERIALS AND METHODS: Ex vivo experiments were performed on 15 swine vertebrae (control, n = 5; titanium screw, n = 5; stainless steel screw, n = 5). Screws and RF ablation probe were inserted identically under fluoroscopy. During RF ablation (3 W, 5 minutes), temperature was measured 10 mm from RF ablation centerpoint and in muscle contacting the screw. Magnetic resonance (MR) imaging, gross pathologic, and histopathologic analyses were performed on 1 specimen from each group. RESULTS: Ablation zone temperatures at 2.5 and 5 minutes increased by 12.2 °C ± 2.6 °C and 21.5 °C ± 2.1 °C (control); 11.0 °C ± 4.1 °C and 20.0 °C ± 2.9 °C (juxta-titanium screw), and 10.0 °C ± 3.4 °C and 17.2 °C ± 3.5 °C (juxta-stainless steel) screw; differences among groups did not reach significance by analysis of variance (P = .87). Mixed-effects linear regression revealed a statistically significant increase in temperature over time in all 3 groups (4.2 °C/min ± 0.4 °C/min, P < .001). Compared with the control, there was no significant difference in the temperature change over time for titanium (-0.3 °C/min ± 0.5 °C/min, P = .53) or steel groups (-0.4 °C/min ± 0.5 °C/min, P = .38). The mean screw temperature at the final time point did not show a statistically significant change compared with baseline in either the titanium group (-1.2 °C ± 2.3 °C, P = .50) or steel group (2.6 °C ± 2.9 °C, P = .11). MR imaging and pathologic analyses revealed homogeneous ablation without sparing of the peri-hardware zones. CONCLUSIONS: Adjacent metallic instrumentation did not affect the rate of or absolute increase in temperature in the ablation zone, did not create peri-metallic ablation inhomogeneities, and did not result in significant nontarget heating of muscle tissue in contact with the metal instrumentation.

Association of cystic fibrosis transmembrane conductance regulator with epithelial sodium channel subunits carrying Liddle's syndrome mutations.

The association of the cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial sodium channel (ENaC) in the pathophysiology of cystic fibrosis (CF) is controversial. Previously, we demonstrated a close physical association between wild-type (WT) CFTR and WT ENaC. We have also shown that the F508del CFTR fails to associate with ENaC unless the mutant protein is rescued pharmacologically or by low temperature. In this study, we present the evidence for a direct physical association between WT CFTR and ENaC subunits carrying Liddle's syndrome mutations. We show that all three ENaC subunits bearing Liddle's syndrome mutations (both point mutations and the complete truncation of the carboxy terminus), could be coimmunoprecipitated with WT CFTR. The biochemical studies were complemented by fluorescence lifetime imaging microscopy (FLIM), a distance-dependent approach that monitors protein-protein interactions between fluorescently labeled molecules. Our measurements revealed significantly increased fluorescence resonance energy transfer between CFTR and all tested ENaC combinations as compared with controls (ECFP and EYFP cotransfected cells). Our findings are consistent with the notion that CFTR and ENaC are within reach of each other even in the setting of Liddle's syndrome mutations, suggestive of a direct intermolecular interaction between these two proteins.

Stellate Ganglion Blockade: an Intervention for the Management of Ventricular Arrhythmias.

PURPOSE OF REVIEW: To highlight the indications, procedural considerations, and data supporting the use of stellate ganglion blockade (SGB) for management of refractory ventricular arrhythmias. RECENT FINDINGS: In patients with refractory ventricular arrhythmias, unilateral or bilateral SGB can reduce arrhythmia burden and defibrillation events for 24-72 h, allowing time for use of other therapies like catheter ablation, surgical sympathectomy, or heart transplantation. The efficacy of SGB appears to be consistent despite the type (monomorphic vs polymorphic) or etiology (ischemic vs non-ischemic cardiomyopathy) of the ventricular arrhythmia. Ultrasound-guided SGB is safe with low risk for complications, even when performed on anticoagulation. SGB is effective and safe and could be considered for patients with refractory ventricular arrhythmias.

Stellate Ganglion Blockade for the Treatment of Refractory Ventricular Arrhythmias.

OBJECTIVES: This study sought to describe our institutional experience with establishing a neurocardiology service in an attempt to provide autonomic modulation as a treatment for ventricular arrhythmias (VAs). BACKGROUND: Treatment-refractory VAs are commonly driven and exacerbated by heightened sympathetic tone. METHODS: Among patients referred to the neurocardiology service (August 2016 to December 2018), we performed ultrasound-based, bilateral, temporary stellate ganglion blockade (SGB) in 20 consecutive patients. We analyzed outcomes of interest including sustained VA or VA requiring defibrillation in the 24 and 48 h before and 24 and 48 h after SGB. RESULTS: The majority of patients were men (n = 19, 95%), with a mean age of 58 ± 14 years. At the time of SGB, 10 (50%) were on inotropic support and 9 (45%) were on mechanical circulatory support. Besides 1 case of hoarseness, there were no apparent procedural complications. SGB was associated with a reduction in the number of VA episodes from the 24 h before (median 5.5 [interquartile range (IQR): 2.0 to 15.8]) to 24 h after SGB (median 0 [IQR: 0 to 3.8]) (p < 0.001). The number of defibrillation events decreased from 2.5 (IQR: 0 to 10.3) to 0 (IQR: 0 to 2.5) (p = 0.002). Similar findings were observed over the 48-h period before and after the SGB. Overall, 9 of 20 (45%) patients had a complete response with no recurrence of ventricular tachycardia (VT) or ventricular fibrillation (VF) for 48 h after SGB. Four (20%) patients had no recurrent VT or VF following SGB through discharge. Similar response rates were observed in those with ischemic (median 6 [IQR: 1.8 to 18.8] to 0.5 [IQR: 0 to 5.3] events; p = 0.031) and nonischemic (median 3.5 [IQR: 1.8 to 6.8] to 0 [IQR: 0 to 1.3] events; p = 0.012) cardiomyopathy. CONCLUSIONS: Minimally invasive, ultrasound-guided bilateral SGB appears safe and provides substantial reduction in VA burden with approximately 1 in 2 patients exhibiting complete suppression of VT or VF for 48 h.

Implications for Neuromodulation Therapy to Control Inflammation and Related Organ Dysfunction in COVID-19.

COVID-19 is a syndrome that includes more than just isolated respiratory disease, as severe acute respiratory syndrome-coronavirus 2 (SARS-CoV2) also interacts with the cardiovascular, nervous, renal, and immune system at multiple levels, increasing morbidity in patients with underlying cardiometabolic conditions and inducing myocardial injury or dysfunction. Emerging evidence suggests that patients with the highest rate of morbidity and mortality following SARS-CoV2 infection have also developed a hyperinflammatory syndrome (also termed cytokine release syndrome). We lay out the potential contribution of a dysfunction in autonomic tone to the cytokine release syndrome and related multiorgan damage in COVID-19. We hypothesize that a cholinergic anti-inflammatory pathway could be targeted as a therapeutic avenue. Graphical Abstract .

Procedural Treatments for Knee Osteoarthritis: A Review of Current Injectable Therapies.

Knee osteoarthritis is a common painful degenerative condition affecting the aging Canadian population. In addition to pain and disability, osteoarthritis is associated with depression, comorbid conditions such as diabetes, and increased caregiver burden. It is predicted to cost the Canadian healthcare system $7.6 billion dollars by 2031. Despite its high cost and prevalence, controversy persists in the medical community regarding optimal therapies to treat knee osteoarthritis. A variety of medications like nonsteroidal anti-inflammatories and opioids can cause severe side effects with limited benefits. Total knee arthroplasty, although a definitive management, comes with risk such as postoperative infections, revisions, and chronic pain. Newer injectable therapies are gaining attention as alternatives to medications because of a safer side effect profile and are much less invasive than a joint replacement. Platelet-rich plasma is beginning to replace the more common injectable therapies of intra-articular corticosteroids and hyaluronic acid, but larger trials are needed to confirm this effect. Small studies have examined prolotherapy and stem cell therapy and demonstrate some benefits. Trials involving genicular nerve block procedures have been successful. As treatments evolve, injectable therapies may offer a safe and effective pathway for patients suffering from knee osteoarthritis.

Erythromyeloid progenitors give rise to a population of osteoclasts that contribute to bone homeostasis and repair.

Osteoclasts are multinucleated cells of the monocyte/macrophage lineage that degrade bone. Here, we used lineage tracing studies-labelling cells expressing Cx3cr1, Csf1r or Flt3-to identify the precursors of osteoclasts in mice. We identified an erythromyeloid progenitor (EMP)-derived osteoclast precursor population. Yolk-sac macrophages of EMP origin produced neonatal osteoclasts that can create a space for postnatal bone marrow haematopoiesis. Furthermore, EMPs gave rise to long-lasting osteoclast precursors that contributed to postnatal bone remodelling in both physiological and pathological settings. Our single-cell RNA-sequencing data showed that EMP-derived osteoclast precursors arose independently of the haematopoietic stem cell (HSC) lineage and the data from fate tracking of EMP and HSC lineages indicated the possibility of cell-cell fusion between these two lineages. Cx3cr1+ yolk-sac macrophage descendants resided in the adult spleen, and parabiosis experiments showed that these cells migrated through the bloodstream to the remodelled bone after injury.

Microglia in Pain: Detrimental and Protective Roles in Pathogenesis and Resolution of Pain.

The previous decade has seen a rapid increase in microglial studies on pain, with a unique focus on microgliosis in the spinal cord after nerve injury and neuropathic pain. Numerous signaling molecules are altered in microglia and contribute to the pathogenesis of pain. Here, we discuss how microglial signaling regulates spinal cord synaptic plasticity in acute and chronic pain conditions with different degrees and variations of microgliosis. We highlight that microglial mediators such as pro- and anti-inflammatory cytokines are powerful neuromodulators that regulate synaptic transmission and pain via neuron-glial interactions. We also reveal an emerging role of microglia in the resolution of pain, in part via specialized pro-resolving mediators including resolvins, protectins, and maresins. We also discuss a possible role of microglia in chronic itch.

Stellate ganglion blockade for the treatment of refractory ventricular arrhythmias: A systematic review and meta-analysis.

INTRODUCTION: Treatment refractory ventricular arrhythmias (VAs) are often driven and exacerbated by heightened sympathetic tone. We aim to conduct a systematic review and meta-analysis of published studies of a temporary percutaneous stellate ganglion block (SGB) on VA burden and defibrillation episodes in patients with treatment refractory VAs. METHODS: Relevant studies from January 1960 through May 2017 were identified in PubMed and Google Scholar. We performed a patient-level analysis using Student's t-test to compare outcomes before and after SGB. RESULTS: We identified 22 unique case series with a total of 35 patients. Patients were 57 ± 17 years old and 69% were males with a high burden of VA. A unilateral (left)-sided SGB was used in 85.7% (30 of 35) of cases and the remaining were bilateral SGB. The use of a unilateral or bilateral SGB resulted in a significant reduction of VA episodes (24-hours pre: mean 16.5 [CI 9.7-23.1] events vs. post: mean 1.4 [CI 0.85-2.01] events; P = 0.0002) and need for defibrillation (24-hours pre: mean 14.2 [CI 6.8-21.6] vs. post: mean 0.6 [CI 0.3-0.9]; P = 0.0026). Furthermore, SGB was significantly associated with a reduction of VA burden regardless of etiology of cardiomyopathy, type of ventricular rhythm, and degree of contractile dysfunction. SGB was followed by surgical sympathectomy in 21% of cases. CONCLUSIONS: Early experience suggests that SGB is associated with an acute reduction in the VA burden and offers potential promise for a broader use in high-risk populations. Randomized controlled studies are needed to confirm the safety and efficacy of this therapy.

5-Hydroxymethylcytosine (5hmC) and Ten-eleven translocation 1-3 (TET1-3) proteins in the dorsal root ganglia of mouse: Expression and dynamic regulation in neuropathic pain.

Epigenetic mechanisms are increasingly implicated in chronic pain pathology. In this study, we demonstrate that the novel epigenetic mark 5-hydroxymethylcytosine (5hmC) is present in dorsal root ganglia (DRG) neurons and glia, and its levels increase following nerve injury. Furthermore, we show that the 5hmC-generating Ten-eleven translocation 1-3 (TET1-3) proteins are expressed in a cell-type specific manner in the DRG, with Tet3 displaying differential upregulation after injury, suggesting a potential role in neuropathic pain.

Genetic Polymorphisms in the Dopamine Receptor 2 Predict Acute Pain Severity After Motor Vehicle Collision.

OBJECTIVES: Dopaminergic signaling is implicated in nociceptive pathways. These effects are mediated largely through dopamine receptors and modulated in part by dopamine transporters. This study tested the hypothesis that genetic variants in the genes encoding dopamine receptor 2 (DRD2) and the dopamine active transporter (SLC6A3) influence acute pain severity after motor vehicle collision. MATERIALS AND METHODS: European Americans presenting to the emergency department after motor vehicle collision were recruited. Overall pain intensity in emergency department was assessed using a 0 to 10 numeric rating scale. DNA was extracted from blood samples and genotyping of single-nucleotide polymorphisms (SNPs) in the DRD2 and SLC6A3 gene was performed. RESULTS: A total of 948 patients completed evaluation. After correction for multiple comparisons, SNP rs6276 at DRD2 showed significant association with pain scores, with individuals with the A/A genotype reporting lower mean pain scores (5.3; 95% confidence interval [CI], 5.1-5.5) than those with A/G (5.9; 95% CI, 5.6-6.1) or G/G (5.7; 95% CI, 5.2-6.2) genotypes (P=0.0027). Secondary analyses revealed an interaction between sex and DRD2 SNPs rs4586205 and rs4648318 on pain scores: females with 2 minor alleles had increased pain intensity, whereas males with 2 minor alleles had less pain than individuals with a major allele (interaction P=0.0019). DISCUSSION: Genetic variants in DRD2 are associated with acute pain after a traumatic stressful event. These results suggest that dopaminergic agents may be useful for the treatment of individuals with acute posttraumatic pain as part of a multimodal opioid-sparing analgesic regimen.

Low temperature and chemical rescue affect molecular proximity of DeltaF508-cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial sodium channel (ENaC).

An imbalance of chloride and sodium ion transport in several epithelia is a feature of cystic fibrosis (CF), an inherited disease that is a consequence of mutations in the cftr gene. The cftr gene codes for a Cl(-) channel, the cystic fibrosis transmembrane conductance regulator (CFTR). Some mutations in this gene cause the balance between Cl(-) secretion and Na(+) absorption to be disturbed in the airways; Cl(-) secretion is impaired, whereas Na(+) absorption is elevated. Enhanced Na(+) absorption through the epithelial sodium channel (ENaC) is attributed to the failure of mutated CFTR to restrict ENaC-mediated Na(+) transport. The mechanism of this regulation is controversial. Recently, we have found evidence for a close association of wild type (WT) CFTR and WT ENaC, further underscoring the role of ENaC along with CFTR in the pathophysiology of CF airway disease. In this study, we have examined the association of ENaC subunits with mutated ΔF508-CFTR, the most common mutation in CF. Deletion of phenylalanine at position 508 (ΔF508) prevents proper processing and targeting of CFTR to the plasma membrane. When ΔF508-CFTR and ENaC subunits were co-expressed in HEK293T cells, we found that individual ENaC subunits could be co-immunoprecipitated with ΔF508-CFTR, much like WT CFTR. However, when we evaluated the ΔF508-CFTR and ENaC association using fluorescence resonance energy transfer (FRET), FRET efficiencies were not significantly different from negative controls, suggesting that ΔF508-CFTR and ENaC are not in close proximity to each other under basal conditions. However, with partial correction of ΔF508-CFTR misprocessing by low temperature and chemical rescue, leading to surface expression as assessed by total internal reflection fluorescence (TIRF) microscopy, we observed a positive FRET signal. Our findings suggest that the ΔF508 mutation alters the close association of CFTR and ENaC.

ENaCs and ASICs as therapeutic targets.

The epithelial Na(+) channel (ENaC) and acid-sensitive ion channel (ASIC) branches of the ENaC/degenerin superfamily of cation channels have drawn increasing attention as potential therapeutic targets in a variety of diseases and conditions. Originally thought to be solely expressed in fluid absorptive epithelia and in neurons, it has become apparent that members of this family exhibit nearly ubiquitous expression. Therapeutic opportunities range from hypertension, due to the role of ENaC in maintaining whole body salt and water homeostasis, to anxiety disorders and pain associated with ASIC activity. As a physiologist intrigued by the fundamental mechanics of salt and water transport, it was natural that Dale Benos, to whom this series of reviews is dedicated, should have been at the forefront of research into the amiloride-sensitive sodium channel. The cloning of ENaC and subsequently the ASIC channels has revealed a far wider role for this channel family than was previously imagined. In this review, we will discuss the known and potential roles of ENaC and ASIC subunits in the wide variety of pathologies in which these channels have been implicated. Some of these, such as the role of ENaC in Liddle's syndrome are well established, others less so; however, all are related in that the fundamental defect is due to inappropriate channel activity.

CFTR regulation of epithelial sodium channel.

Cystic fibrosis (CF) is a lethal genetic disorder, characterized by both clinical and genetic complexities, and arises as a result of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. The gene encodes a Cl(-) channel belonging to the ABC (ATP Binding Cassette) superfamily of transporters. The members of this superfamily use ATP hydrolysis to fulfill their function as active transporters. So far, CFTR is the only member of this family to function as a cAMP-activated Cl(-) channel. Intense research following the cloning of the CFTR gene has extended the role of the CFTR beyond that of a Cl(-) channel. One of the best recognized, yet still controversial, functions of the CFTR is its ability to modulate the functioning of other transporters. The modulation of epithelial Na(+) channel (ENaC) function serves as a prime example of regulatory function of the CFTR. In this chapter, we will briefly describe an integrated protocol consisting of biochemical and electrophysiological approaches to study the regulation of ENaC by CFTR.

Amiloride docking to acid-sensing ion channel-1.

Amiloride is a small molecule diuretic, which has been used to dissect sodium transport pathways in many different systems. This drug is known to interact with the epithelial sodium channel and acid-sensing ion channel proteins, as well as sodium/hydrogen antiporters and sodium/calcium exchangers. The exact structural basis for these interactions has not been elucidated as crystal structures of these proteins have been challenging to obtain, though some involved residues and domains have been mapped. This work examines the interaction of amiloride with acid-sensing ion channel-1, a protein whose structure is available using computational and experimental techniques. Using molecular docking software, amiloride and related molecules were docked to model structures of homomeric human ASIC-1 to generate potential interaction sites and predict which analogs would be more or less potent than amiloride. The predictions made were experimentally tested using whole-cell patch clamp. Drugs previously classified as NCX or NHE inhibitors are shown to also inhibit hASIC-1. Potential docking sites were re-examined against experimental data to remove spurious interaction sites. The voltage sensitivity of inhibitors was also examined. Using the aggregated data from these computational and experimental experiments, putative interaction sites for amiloride and hASIC-1 have been defined. Future work will experimentally verify these interaction sites, but at present this should allow for virtual screening of drug libraries at these putative interaction sites.

Knockdown of ASIC1 and epithelial sodium channel subunits inhibits glioblastoma whole cell current and cell migration.

High grade gliomas such as glioblastoma multiforme express multiple members of the epithelial sodium channel (ENaC)/Degenerin family, characteristically displaying a basally active amiloride-sensitive cation current not seen in normal human astrocytes or lower grade gliomas. Using quantitative real time PCR, we have shown higher expression of ASIC1, alphaENaC, and gammaENaC in D54-MG human glioblastoma multiforme cells compared with primary human astrocytes. We hypothesize that this glioma current is mediated by a hybrid channel composed of a mixture of ENaC and acid-sensing ion channel (ASIC) subunits. To test this hypothesis we made dominant negative cDNAs for ASIC1, alphaENaC, gammaENaC, and deltaENaC. D54-MG cells transfected with the dominant negative constructs for ASIC1, alphaENaC, or gammaENaC showed reduced protein expression and a significant reduction in the amiloride-sensitive whole cell current as compared with untransfected D54-MG cells. Knocking down alphaENaC or gammaENaC also abolished the high P(K)(+)/P(Na)(+) of D54-MG cells. Knocking down deltaENaC in D54-MG cells reduced deltaENaC protein expression but had no effect on either the whole cell current or K(+) permeability. Using co-immunoprecipitation we show interactions between ASIC1, alphaENaC, and gammaENaC, consistent with these subunits interacting with each other to form an ion channel in glioma cells. We also found a significant inhibition of D54-MG cell migration after ASIC1, alphaENaC, or gammaENaC knockdown, consistent with the hypothesis that ENaC/Degenerin subunits play an important role in glioma cell biology.

Psalmotoxin-1 docking to human acid-sensing ion channel-1.

Acid-sensing ion channel-1 (ASIC-1) is a proton-gated ion channel implicated in nociception and neuronal death during ischemia. Recently the first crystal structure of a chicken ASIC was obtained. Expanding upon this work, homology models of the human ASICs were constructed and evaluated. Energy-minimized structures were tested for validity by in silico docking of the models to psalmotoxin-1, which potently inhibits ASIC-1 and not other members of the family. The data are consistent with prior radioligand binding and functional assays while also explaining the selectivity of PcTX-1 for homomeric hASIC-1a. Binding energy calculations suggest that the toxin and channel create a complex that is more stable than the channel alone. The binding is dominated by the coulombic contributions, which account for why the toxin-channel interaction is not observed at low pH. The computational data were experimentally verified with single channel and whole-cell electrophysiological studies. These validated models should allow for the rational design of specific and potent peptidomimetic compounds that may be useful for the treatment of pain or ischemic stroke.

Assessment of the CFTR and ENaC association.

Cystic fibrosis (CF) is one of the most common lethal genetic disorders. It results primarily from mutations in the cystic fibrosis transmembrane conductance regulator (cftr) gene. These mutations cause inadequate functioning of CFTR, which in turn leads to the severe disruption of transport function in several epithelia across various organs. Affected organs include the sweat glands, the intestine, and the reproductive system, with the most devastating consequences due to the effects of the disease on airways. Despite aggressive treatment, gradual lung failure is the major life limiting factor in patients with CF. Understanding of the exact manner by which defects in the CFTR lead to lung failure is thus critical. In the CF airway, decreased chloride secretion and increased salt absorption is observed. The decreased chloride secretion appears to be a direct consequence of defective CFTR; however, the increased salt absorption is believed to result from the failure of CFTR to restrict salt absorption through a sodium channel named the epithelial Na(+) channel, ENaC. The mechanism by which CFTR modulates the function of ENaC proteins is still obscure and somewhat controversial. In this short review we will focus on recent findings of a possible direct CFTR and ENaC association.