SARS-CoV-2 variant of concern fitness and adaptation in primary human airway epithelia.

The severe acute respiratory syndrome coronavirus 2 pandemic is characterized by the emergence of novel variants of concern (VOCs) that replace ancestral strains. Here, we dissect the complex selective pressures by evaluating variant fitness and adaptation in human respiratory tissues. We evaluate viral properties and host responses to reconstruct forces behind D614G through Omicron (BA.1) emergence. We observe differential replication in airway epithelia, differences in cellular tropism, and virus-induced cytotoxicity. D614G accumulates the most mutations after infection, supporting zoonosis and adaptation to the human airway. We perform head-to-head competitions and observe the highest fitness for Gamma and Delta. Under these conditions, RNA recombination favors variants encoding the B.1.617.1 lineage 3' end. Based on viral growth kinetics, Alpha, Gamma, and Delta exhibit increased fitness compared to D614G. In contrast, the global success of Omicron likely derives from increased transmission and antigenic variation. Our data provide molecular evidence to support epidemiological observations of VOC emergence.

Reoperation Following Primary Greater Occipital Nerve Decompression Surgery: Incidence, Risk Factors and Outcomes.

BACKGROUND: Although nerve decompression surgery is an effective treatment for refractory occipital neuralgia (ON), a proportion of patients experience recurrence of pain and undergo reoperation. This study analyzes the incidence, risk factors, and outcomes of reoperation following primary greater occipital nerve (GON) decompression. METHODS: 215 patients who underwent 399 primary GON decompressions were prospectively enrolled. Data included patient demographics, past medical and surgical history, reoperation rates, intraoperative findings, surgical technique, and postoperative outcomes in terms of pain frequency (days/month), duration (hours/day), intensity (scale 0-10), and migraine headache index (MHI). Bivariate analyses, univariable and multivariable logistic regression analysis was performed. RESULTS: 27 (6.8%) GON decompressions required reoperation with neurectomy at a median follow-up time of 15.5 months (9.8-40.5). Cervical spine disorders on imaging that did not warrant surgical intervention (OR, 4.88; 95% 1.61-14.79; p<0.01) and radiofrequency ablation (RFA) (OR, 4.20; 95% CI, 1.45-15.2; p<0.05) were significantly associated with higher rates of reoperation. At 12 months postoperatively, patients who underwent reoperation achieved similar mean reductions in pain frequency, duration, intensity and MHI, as compared to patients who underwent only primary decompression (p>0.05). CONCLUSION: Patients with ON who have a history of cervical spine disorders or RFA should be counseled that primary decompression has a higher risk of reoperation, but outcomes are ultimately comparable.

Viral afterlife: SARS-CoV-2 as a reservoir of immunomimetic peptides that reassemble into proinflammatory supramolecular complexes.

It is unclear how severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection leads to the strong but ineffective inflammatory response that characterizes severe Coronavirus disease 2019 (COVID-19), with amplified immune activation in diverse cell types, including cells without angiotensin-converting enzyme 2 receptors necessary for infection. Proteolytic degradation of SARS-CoV-2 virions is a milestone in host viral clearance, but the impact of remnant viral peptide fragments from high viral loads is not known. Here, we examine the inflammatory capacity of fragmented viral components from the perspective of supramolecular self-organization in the infected host environment. Interestingly, a machine learning analysis to SARS-CoV-2 proteome reveals sequence motifs that mimic host antimicrobial peptides (xenoAMPs), especially highly cationic human cathelicidin LL-37 capable of augmenting inflammation. Such xenoAMPs are strongly enriched in SARS-CoV-2 relative to low-pathogenicity coronaviruses. Moreover, xenoAMPs from SARS-CoV-2 but not low-pathogenicity homologs assemble double-stranded RNA (dsRNA) into nanocrystalline complexes with lattice constants commensurate with the steric size of Toll-like receptor (TLR)-3 and therefore capable of multivalent binding. Such complexes amplify cytokine secretion in diverse uninfected cell types in culture (epithelial cells, endothelial cells, keratinocytes, monocytes, and macrophages), similar to cathelicidin's role in rheumatoid arthritis and lupus. The induced transcriptome matches well with the global gene expression pattern in COVID-19, despite using <0.3% of the viral proteome. Delivery of these complexes to uninfected mice boosts plasma interleukin-6 and CXCL1 levels as observed in COVID-19 patients.

AGC kinase inhibitors regulate STING signaling through SGK-dependent and SGK-independent mechanisms.

Type 1 IFN expression is critical in the innate immune response, but aberrant expression is associated with autoimmunity and cancer. Here, we identify N-[4-(1H46 pyrazolo[3,4-b] pyrazin-6-yl)-phenyl]-sulfonamide (Sanofi-14h), a compound with preference for inhibition of the AGC family kinase SGK3, as an inhibitor of Ifnb1 gene expression in response to STING stimulation of macrophages. Sanofi-14h abrogated SGK activity and also impaired activation of the critical TBK1/IRF3 pathway downstream of STING activation, blocking interaction of STING with TBK1. Deletion of SGK1/3 in a macrophage cell line did not block TBK1/IRF3 activation but decreased expression of transcription factors, such as IRF7 and STAT1, required for the innate immune response. Other AGC kinase inhibitors blocked TBK1 and IRF3 activation suggesting common action on a critical regulatory node in the STING pathway. These studies reveal both SGK-dependent and SGK-independent mechanisms in the innate immune response and indicate an approach to block aberrant Ifnb1 expression.

Tumor MK2 transcript levels are associated with improved response to chemotherapy and patient survival in non-small cell lung cancer.

Non-small cell lung cancers (NSCLCs) demonstrate intrinsic resistance to cell death, even after chemotherapy. Previous work suggested defective nuclear translocation of active caspase-3 in observed resistance to cell death. We have identified mitogen-activated protein kinase-activated protein kinase 2 (MK2; encoded by the gene MAPKAPK2) is required for caspase-3 nuclear translocation in the execution of apoptosis in endothelial cells. The objective was to determine MK2 expression in NSCLCs and the association between MK2 and clinical outcomes in patients with NSCLC. Clinical and MK2 mRNA data were extracted from two demographically distinct NSCLC clinical cohorts, North American (The Cancer Genome Atlas, TCGA) and East Asian (EA). Tumor responses following first round of chemotherapy were dichotomized as clinical response (complete response, partial response, and stable disease) or progression of disease. Multivariable survival analyses were performed using Cox proportional hazard ratios and Kaplan-Meier curves. NSCLC exhibited lower MK2 expression than SCLC cell lines. In patients, lower tumor MK2 transcript levels were observed in those presenting with late-stage NSCLC. Higher MK2 expression was associated with clinical response following initial chemotherapy and independently associated with improved 2-yr survival in two distinct cohorts, 0.52 (0.28-0.98) and 0.1 (0.01-0.81), TCGA and EA, respectively, even after adjusting for common oncogenic driver mutations. Survival benefit of higher MK2 expression was unique to lung adenocarcinoma when comparing across various cancers. This study implicates MK2 in apoptosis resistance in NSCLC and suggests prognostic value of MK2 transcript levels in patients with lung adenocarcinoma.

MK2 nonenzymatically promotes nuclear translocation of caspase-3 and resultant apoptosis.

We have previously identified mitogen-activated protein kinase-activated protein kinase 2 (MK2) is required for caspase-3 nuclear translocation in the execution of apoptosis; however, little is known of the underlying mechanisms. Therefore, we sought to determine the role of kinase and nonkinase functions of MK2 in promoting nuclear translocation of caspase-3. We identified two non-small cell lung cancer cell lines for use in these experiments based on low MK2 expression. Wild-type, enzymatic and cellular localization mutant MK2 constructs were expressed using adenoviral infection. Cell death was evaluated by flow cytometry. In addition, cell lysates were harvested for protein analyses. Phosphorylation of caspase-3 was determined using two-dimensional gel electrophoresis followed by immunoblotting and in vitro kinase assay. Association between MK2 and caspase-3 was evaluated using proximity-based biotin ligation assays and co-immunoprecipitation. Overexpression of MK2 resulted in nuclear translocation of caspase-3 and caspase-3-mediated apoptosis. MK2 directly phosphorylates caspase-3; however, phosphorylation status of caspase-3 or MK2-dependent phosphorylation of caspase-3 did not alter caspase-3 activity. The enzymatic function of MK2 was dispensable in nuclear translocation of caspase-3. MK2 and caspase-3 associated together and a nonenzymatic function of MK2, chaperoned nuclear trafficking, is required for caspase-3-mediated apoptosis. Taken together, our results demonstrate a nonenzymatic role for MK2 in the nuclear translocation of caspase-3. Furthermore, MK2 may function as a molecular switch in regulating the transition between the cytosolic and nuclear functions of caspase-3.

Circadian regulator CLOCK promotes tumor angiogenesis in glioblastoma.

Glioblastoma (GBM) is one of the most aggressive tumors in the adult central nervous system. We previously revealed that circadian regulation of glioma stem cells (GSCs) affects GBM hallmarks of immunosuppression and GSC maintenance in a paracrine and autocrine manner. Here, we expand the mechanism involved in angiogenesis, another critical GBM hallmark, as a potential basis underlying CLOCK's pro-tumor effect in GBM. Mechanistically, CLOCK-directed olfactomedin like 3 (OLFML3) expression results in hypoxia-inducible factor 1-alpha (HIF1α)-mediated transcriptional upregulation of periostin (POSTN). As a result, secreted POSTN promotes tumor angiogenesis via activation of the TANK-binding kinase 1 (TBK1) signaling in endothelial cells. In GBM mouse and patient-derived xenograft models, blockade of the CLOCK-directed POSTN-TBK1 axis inhibits tumor progression and angiogenesis. Thus, the CLOCK-POSTN-TBK1 circuit coordinates a key tumor-endothelial cell interaction and represents an actionable therapeutic target for GBM.

Reset Neurectomy for Cutaneous Nerve Injuries.

Diffuse cutaneous nerve injuries, often caused by a crush mechanism, are challenging for the nerve surgeon. Discrete nerve transections and focal neuromas are easier to identify and have a more distinct treatment algorithm. Following crush injury to a noncritical sensory nerve, a successful local anesthetic block proximal to the injury may help determine the possibility of surgical intervention. In these cases, we describe a technique of "reset neurectomy" whereby a neurectomy is performed proximal to the zone of injury, and immediate repair or reconstruction (with or without a nerve graft) is performed. This technique may be useful in cases of diffuse, nontransection nerve injuries in which neuropathic pain is the primary symptom.

Effects of IFN-γ on immune cell kinetics during the resolution of acute lung injury.

The immunologic responses that occur early in the acute respiratory distress syndrome (ARDS) elicit immune-mediated damage. The mechanisms underlying the resolution of ARDS, particularly the role of signaling molecules in regulating immune cell kinetics, remain important questions. Th1-mediated responses can contribute to the pathogenesis of acute lung injury (ALI). Interferon-gamma (IFN-γ) orchestrates early inflammatory events, enhancing immune-mediated damage. The current study investigated IFN-γ during resolution in several experimental models of ALI. The absence of IFN-γ resulted in altered kinetics of lymphocyte and macrophage responses, suggesting that IFN-γ present in this microenvironment is influential in ALI resolution. Genetic deficiency of IFN-γ or administering neutralizing IFN-γ antibodies accelerated the pace of resolution. Neutralizing IFN-γ decreased the numbers of interstitial and inflammatory macrophages and increased alveolar macrophage numbers during resolution. Our results underline the complexity of lung injury resolution and provide insight into the effects through which altered IFN-γ concentrations affect immune cell kinetics and the rate of resolution. These findings suggest that therapies that spatially or temporally control IFN-γ signaling may promote ALI resolution. Identifying and elucidating the mechanisms critical to ALI resolution will allow the development of therapeutic approaches to minimize collateral tissue damage without adversely altering the response to injury.

The Greater Occipital Nerve and Obliquus Capitis Inferior Muscle: Anatomical Interactions and Implications for Occipital Pain Syndromes.

BACKGROUND: The compression/injury of the greater occipital nerve has been identified as a trigger of occipital headaches. Several compression points have been described, but the morphology of the myofascial unit between the greater occipital nerve and the obliquus capitis inferior muscle has not been studied yet. METHODS: Twenty fresh cadaveric heads were dissected, and the greater occipital nerve was tracked from its emergence to its passage around the obliquus capitis inferior. The intersection point between the greater occipital nerve and the obliquus capitis inferior, and the length and thickness of the obliquus capitis inferior, were measured. In addition, the nature of the interaction and whether the nerve passed through the muscle were also noted. RESULTS: All nerves passed either around the muscle loosely (type I), incorporated in the dense superficial muscle fascia (type II), or directly through a myofascial sleeve within the muscle (type III). The obliquus capitis inferior length was 5.60 ± 0.46 cm. The intersection point between the obliquus capitis inferior and the greater occipital nerve was 6.80 ± 0.68 cm caudal to the occiput and 3.56 ± 0.36 cm lateral to the midline. The thickness of the muscle at its intersection with the greater occipital nerve was 1.20 ± 0.25 cm. Loose, tight, and intramuscular connections were found in seven, 31, and two specimens, respectively. CONCLUSIONS: The obliquus capitis inferior remains relatively immobile during traumatic events, like whiplash injuries, placing strain as a tethering point on the greater occipital nerve. Better understanding of the anatomical relationship between the greater occipital nerve and the obliquus capitis inferior can be clinically useful in cases of posttraumatic occipital headaches for diagnostic and operative planning purposes.

Supraorbital Rim Syndrome: Definition, Surgical Treatment, and Outcomes for Frontal Headache.

BACKGROUND: Supraorbital rim syndrome (SORS) is a novel term attributed to a composite of anatomically defined peripheral nerve entrapment sites of the supraorbital rim region. The SORS term establishes a more consistent nomenclature to describe the constellation of frontal peripheral nerve entrapment sites causing frontal headache pain. In this article, we describe the anatomical features of SORS and evidence to support its successful treatment using the transpalpebral approach that allows direct vision of these sites and the intraconal space. METHODS: A retrospective review of 276 patients who underwent nerve decompression or neurectomy procedures for frontal or occipital headache was performed. Of these, treatment of 96 patients involved frontal surgery, and 45 of these patients were pure SORS patients who underwent this specific frontal trigger site deactivation surgery only. All procedures involved direct surgical approach through the upper eyelid to address the nerves of the supraorbital rim at the bony rim and myofascial sites. RESULTS: Preoperative and postoperative data from the Migraine Disability Assessment Questionnaire were analyzed with paired t test. After surgical intervention, Migraine Disability Assessment Questionnaire scores decreased significantly at 12 months postoperatively (P < 0.0001). CONCLUSIONS: SORS describes the totality of compression sites both at the bony orbital rim and the corrugator myofascial unit for the supraorbital rim nerves. Proper diagnosis, full anatomical site knowledge, and complete decompression allow for consistent treatment. Furthermore, the direct, transpalpebral surgical approach provides significant benefit to allow complete decompression.

A review of current evidence in the surgical treatment of migraine headaches.

SUMMARY: Migraines affect 18% of women and 6% of men and result in an estimated $1 billion in medical costs and $16 billion productivity loss in the United States annually. Migraine headaches persist as a problem of this scale because pharmacologic treatments for migraines are frequently incompletely effective, resulting in a population of patients with significant residual disability. In the last decade, novel approaches to the treatment of migraines have been developed based on the theory that extracranial sensory branches of the trigeminal and cervical spinal nerves can be irritated, entrapped, or compressed at points throughout their anatomic course, ultimately leading to the cascade of physiologic events that results in migraine. Botulinum toxin (Botox) injection and surgical decompression of these trigger points have been shown to reduce or eliminate migraines in patients who are incompletely treated by traditional medical management. Despite the recent advances made with Botox, this treatment strategy most commonly results in only temporary migraine prevention. However, the evidence supporting the efficacy and safety of permanent surgical decompression of peripheral trigger points is accumulating rapidly, and the overall success rate of surgery has approached 90%. In addition, an abundance of literature investigating the precise anatomical dissections associated with trigger points has been published concurrently. This article reviews the most up-to-date clinical and anatomic evidence available and seeks to provide a comprehensive, concise resource for the current state of the art in the surgical treatment of migraine headaches.

Anatomy of the supratrochlear nerve: implications for the surgical treatment of migraine headaches.

BACKGROUND: Migraine headaches have been linked to compression, irritation, or entrapment of peripheral nerves in the head and neck at muscular, fascial, and vascular sites. The frontal region is a trigger for many patients' symptoms, and the possibility for compression of the supratrochlear nerve by the corrugator muscle has been indirectly implied. To further delineate their relationship, a fresh tissue anatomical study was designed. METHODS: Dissection of the brow region was undertaken in 25 fresh cadaveric heads. The corrugator muscle was identified on both sides, and its relationship with the supratrochlear nerve was investigated. RESULTS: The supratrochlear nerve was found in all 50 hemifaces. Three potential points of compression were uncovered in this investigation: the nerve entrance into the brow through the frontal notch or foramen, the entrance of the nerve into the corrugator muscle, and the exit of the nerve from the corrugator muscle. The nerve generally bifurcates within the retro-orbicularis oculi fat pad, and these branches enter into one of four relationships with the corrugator muscle: both branches enter the muscle, one branch enters the muscle and one remains deep, both branches remain deep, and the branches further branch into ever smaller filaments that cannot be identified cranially. CONCLUSIONS: Some patients are nonresponders to migraine decompression techniques that address the supraorbital nerve. The supratrochlear nerve may be compressed in these patients. A standard corrugator resection that comes more medially within 1.8 cm of the midline may be beneficial. The morphology of the frontal notch/foramen must be examined and addressed if necessary.

The anatomical morphology of the supraorbital notch: clinical relevance to the surgical treatment of migraine headaches.

BACKGROUND: Current literature for surgical deactivation of frontal migraine trigger points does not incorporate decompression of the supraorbital foramen or fascial bands at the supraorbital rim (frontal exit) as part of the surgical procedure. To evaluate this primary compression site for the supraorbital nerve, anatomical dissections were performed and a classification system was developed. METHODS: Sixty supraorbital regions from 30 ethylene glycol-preserved cadaveric heads were dissected. Particular attention was focused on the morphology of the supraorbital rim, specifically, the presence of a supraorbital notch or supraorbital foramen. The presence or absence of a fascial band completing the notch and the patterns of fascial band variations were documented. RESULTS: A supraorbital foramen was identified 27 percent of the time and a notch was identified 83 percent of the time. When a notch was encountered, a fascial band forming the floor of the notch that completed the encirclement of the supraorbital nerve was noted in 86 percent of supraorbital regions. A classification system was developed to categorize the four common fascial band variation patterns observed. CONCLUSIONS: This study verifies the presence of a primary compression site for the supraorbital nerve that is proximal to the glabellar myofascial complex. Knowledge of this compression site and its possible anatomical variations will enable surgeons to perform a more complete supraorbital nerve decompression for migraine amelioration.

Chronic daily headaches secondary to greater auricular and lesser occipital neuromas following endolymphatic shunt surgery.

In head and neck surgery, peripheral sensory nerves are at risk for traumatic injury. These injuries are known to be peripheral triggers of chronic headaches if left untreated or unrecognised. We report the case of a patient that presented with a severe headache, nausea and emesis of 2 years duration following endolymphatic shunt placement for Meniere's disease. Nerve blockade suggested a peripheral trigger, and surgical exploration of the incision site revealed traumatic neuromas of the greater auricular and lesser occipital nerves. Subsequent nerve resection yielded complete symptomatic relief. This is the first case report of a peripherally triggered migraine headache due to the development of neuromas of the greater auricular and lesser occipital nerves, also representing a previously unreported complication of endolymphatic shunt placement. It is recommended that in patients presenting with intractable migraines and a history of head and neck surgery, diagnostic nerve blockage be used to assess for neuromas.

Soleal sling syndrome (proximal tibial nerve compression): results of surgical decompression.

BACKGROUND: Although distal tibial nerve compression is well recognized, proximal tibial nerve compression remains a rarely recognized clinical condition. This report defines the presentation, diagnosis, surgical decompression technique, and clinical outcome of neurolysis of the tibial nerve at this soleal sling compression site. METHODS: Forty-nine patients with 69 proximal tibial nerves (20 bilateral) were stratified retrospectively into three groups: neuropathy (n = 10), failed tarsal tunnel syndrome (n = 25), and trauma (n = 14). Pain level, strength of the flexor hallucis longus muscle, neurosensory testing of the hallux, and subjective sensory improvement were evaluated. Each proximal tibial nerve compression was subjected to neurolysis with division of the soleal sling. RESULTS: Results were stratified into poor, fair, good, and excellent based on the amount of pain relief and improvement in motor and sensory function. In all groups combined, there were 13 excellent (26.5 percent), 13 good (26.5 percent), 18 fair (36.7 percent), and five poor (10.2 percent) results. Results in the neuropathy group were excellent in two patients, good in three, fair in four, and poor in one (mean follow-up, 18.7 months). Results in the failed tarsal tunnel syndrome group were excellent in two, good in six, fair in 13, and poor in four patients (mean follow-up, 13.9 months). The trauma subgroup had the best outcomes: excellent in nine patients, good in four, fair in one, and poor in zero (mean follow-up, 13.4 months). CONCLUSION: Regardless of cause, if a proximal tibial nerve compression beneath the soleal sling is identified, neurolysis may improve pain and sensory and motor function. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Phylogenetic conservation of the cell-type-specific Lan3-2 glycoepitope in Caenorhabditis elegans.

The biological function of a cell-type-specific glycosylation of an adhesion molecule belonging to the L1CAM immunoglobulin superfamily was previously determined in the nervous system of the embryonic leech, Hirudo medicinalis. The Lan3-2 glycoepitope is a surface marker of sensory afferent neurons and is required for their appropriate developmental collateral branching and synaptogenesis in the CNS. The chemical structure of the Lan3-2 glycoepitope consists of beta-(1,4)-linked mannopyranose. Here, we show the conservation of the cell-type-specific expression of this mannose polymer in Caenorhabditis elegans. The Lan3-2 glycoepitope is expressed on the cell surface of a subset of dissociated embryonic neurons and, in the adult worm, by the pharyngeal motor neuron, M5, and the chemosensory afferents, the amphids. Additionally, the vulval epithelium expresses the Lan3-2 glycoepitope in late L4 larvae and in adult hermaphrodites. To investigate proteins carrying this restrictively expressed glycoepitope, worm extract was immunoaffinity purified with Lan3-2 monoclonal antibody and Western blotted. A polyclonal antibody reactive with the cytoplasmic tail of LAD-1/SAX-7, a C. elegans member of the L1CAM family, recognizes a 270 kDa protein band while Lan3-2 antibody also recognizes a 190 kDa glycoform, its putative Lan3-2 ectodomain. Thus, in C. elegans, as in leech, the Lan3-2 epitope is located on a L1CAM homologue. The cell-type-specific expression of the Lan3-2 glycoepitope shared by leech and C. elegans will be useful for understanding how cell-type-specific glycoepitopes mediate cell-cell interactions during development.

Mongolia: establishing a national palliative care program.

Mongolia has established the foundation measures for a national palliative care program. Generic cost-effective opioids have been made available. Prescription regulations have been changed to allow opioid use according to good medical practice. Education on palliative care has been incorporated into the undergraduate curricula of doctors, nurses, and social workers, and the training of specialists in palliative care has been initiated concurrent with established standards for palliative care.