Mertk-expressing microglia influence oligodendrogenesis and myelin modelling in the CNS.

BACKGROUND: Microglia, an immune cell found exclusively within the CNS, initially develop from haematopoietic stem cell precursors in the yolk sac and colonise all regions of the CNS early in development. Microglia have been demonstrated to play an important role in the development of oligodendrocytes, the myelin producing cells in the CNS, as well as in myelination. Mertk is a receptor expressed on microglia that mediates immunoregulatory functions, including myelin efferocytosis. FINDINGS: Here we demonstrate an unexpected role for Mertk-expressing microglia in both oligodendrogenesis and myelination. The selective depletion of Mertk from microglia resulted in reduced oligodendrocyte production in early development and the generation of pathological myelin. During demyelination, mice deficient in microglial Mertk had thinner myelin and showed signs of impaired OPC differentiation. We established that Mertk signalling inhibition impairs oligodendrocyte repopulation in Xenopus tadpoles following demyelination. CONCLUSION: These data highlight the importance of microglia in myelination and are the first to identify Mertk as a regulator of oligodendrogenesis and myelin ultrastructure.

Lung transcriptional unresponsiveness and loss of early influenza virus control in infected neonates is prevented by intranasal Lactobacillus rhamnosus GG.

Respiratory viral infections contribute substantially to global infant losses and disproportionately affect preterm neonates. Using our previously established neonatal murine model of influenza infection, we demonstrate that three-day old mice are exceptionally sensitive to influenza virus infection and exhibit high mortality and viral load. Intranasal pre- and post-treatment of neonatal mice with Lactobacillus rhamnosus GG (LGG), an immune modulator in respiratory viral infection of adult mice and human preterm neonates, considerably improves neonatal mice survival after influenza virus infection. We determine that both live and heat-killed intranasal LGG are equally efficacious in protection of neonates. Early in influenza infection, neonatal transcriptional responses in the lung are delayed compared to adults. These responses increase by 24 hours post-infection, demonstrating a delay in the kinetics of the neonatal anti-viral response. LGG pretreatment improves immune gene transcriptional responses during early infection and specifically upregulates type I IFN pathways. This is critical for protection, as neonatal mice intranasally pre-treated with IFNß before influenza virus infection are also protected. Using transgenic mice, we demonstrate that the protective effect of LGG is mediated through a MyD88-dependent mechanism, specifically via TLR4. LGG can improve both early control of virus and transcriptional responsiveness and could serve as a simple and safe intervention to protect neonates.

Characterization of CD31 expression on murine and human neonatal T lymphocytes during development and activation.

BackgroundCD31, expressed by the majority of the neonatal T-cell pool, is involved in modulation of T-cell receptor signaling by increasing the threshold for T-cell activation. Therefore, CD31 could modulate neonatal tolerance and adaptive immune responses.MethodsLymphocytes were harvested from murine neonates at different ages, human late preterm and term cord blood, and adult peripheral blood. Human samples were activated over a 5-day period to simulate acute inflammation. Mice were infected with influenza; lungs and spleens were harvested at days 6 and 9 post infection and analyzed by flow cytometry.ResultsCD31-expressing neonatal murine CD4+ and CD8a+ T cells increase over the first week of life. Upon in vitro stimulation, human infants' CD4+ and CD8a+ T cells shed CD31 faster in comparison with adults. In the context of acute infection, mice infected at 3 days of age have an increased number of naive and activated CD31+ T lymphocytes at the site of infection at days 6 and 9 post infection, as compared with those infected at 7 days of age; however, the opposite is true in the periphery.ConclusionDifferences in trafficking of CD31+ cytotoxic T lymphocytes (CTLs) during acute influenza infection could modulate tolerance and contribute to a dampened adaptive immune response in neonates.

miRNAs in Microglia: Important Players in Multiple Sclerosis Pathology.

Microglia are the resident immune cells of the central nervous system and important regulators of brain homeostasis. Central to this role is a dynamic phenotypic plasticity that enables microglia to respond to environmental and pathological stimuli. Importantly, different microglial phenotypes can be both beneficial and detrimental to central nervous system health. Chronically activated inflammatory microglia are a hallmark of neurodegeneration, including the autoimmune disease multiple sclerosis (MS). By contrast, microglial phagocytosis of myelin debris is essential for resolving inflammation and promoting remyelination. As such, microglia are being explored as a potential therapeutic target for MS. MicroRNAs (miRNAs) are short non-coding ribonucleic acids that regulate gene expression and act as master regulators of cellular phenotype and function. Dysregulation of certain miRNAs can aberrantly activate and promote specific polarisation states in microglia to modulate their activity in inflammation and neurodegeneration. In addition, miRNA dysregulation is implicated in MS pathogenesis, with circulating biomarkers and lesion specific miRNAs identified as regulators of inflammation and myelination. However, the role of miRNAs in microglia that specifically contribute to MS progression are still largely unknown. miRNAs are being explored as therapeutic agents, providing an opportunity to modulate microglial function in neurodegenerative diseases such as MS. This review will focus firstly on elucidating the complex role of microglia in MS pathogenesis. Secondly, we explore the essential roles of miRNAs in microglial function. Finally, we focus on miRNAs that are implicated in microglial processes that contribute directly to MS pathology, prioritising targets that could inform novel therapeutic approaches to MS.

Determination of respiratory motion for distal esophagus cancer using four-dimensional computed tomography.

PURPOSE: To investigate the motion characteristics of distal esophagus cancer primary tumors using four-dimensional computed tomography (4D CT). METHODS AND MATERIALS: Thirty-one consecutive patients treated for esophagus cancer who received respiratory-gated 4D CT imaging for treatment planning were selected. Deformable image registration was used to map the full expiratory motion gross tumor volume (GTV) to the full-inspiratory CT image, allowing quantitative assessment of each voxel's displacement. These displacements were correlated with patient tumor and respiratory characteristics. RESULTS: The mean (SE) tidal volume was 608 (73) mL. The mean GTV volume was 64.3 (10.7) mL on expiration and 64.1 (10.7) mL on inspiration (no significant difference). The mean tumor motion in the x-direction was 0.13 (0.006) cm (average of absolute values), in the y-direction 0.23 (0.01) cm (anteriorly), and in the z-direction 0.71 (0.02) cm (inferiorly). Tumor motion correlated with tidal volume. Comparison of tumor motion above vs. below the diaphragm was significant for the average net displacement (p = 0.014), motion below the diaphragm was greater than above. From the cumulative distribution 95% of the tumors moved less than 0.80 cm radially and 1.75 cm inferiorly. CONCLUSIONS: Primary esophagus tumor motion was evaluated with 4D CT. According to the results of this study, when 4D CT is not available, a radial margin of 0.8 cm and axial margin of +/-1.8 cm would provide tumor motion coverage for 95% of the cases in our study population.

Introduction of triggers for palliative care consultation improves utilization and satisfaction within a level four NICU.

OBJECTIVE: We sought to explore the beliefs regarding palliative care team utilization, as well as increase consultation and awareness of the palliative care team's role in the NICU. STUDY DESIGN: The study design in this Level 4 NICU included observational time series with multiple planned sequential interventions. Medical chart review was conducted to determine eligibility, and statistical process control charts were used to show performance over time. RESULTS: Prior to implementation of the triggers, 26% received consultation, which increased to 46% after implementation. There was an increase in level of understanding, knowledge of team's role, and improved utilization. The time until initial consultation decreased from ~1.5 months to 1 week. CONCLUSIONS: We observed a 20% increase in consultations. Key interventions included continual education, reminders, and clear postage of the trigger list. Written guidelines increase awareness of a palliative care team's role within a NICU, and provider satisfaction.

Reduction of normal lung irradiation in locally advanced non-small-cell lung cancer patients, using ventilation images for functional avoidance.

PURPOSE: To investigate the ability of four-dimensional computed tomography (4D-CT)-derived ventilation images to identify regions of highly functional lung for avoidance in intensity-modulated radiotherapy (IMRT) planning in locally advanced non-small-cell lung cancer (NSCLC). METHODS AND MATERIALS: The treatment-planning records from 21 patients with Stage III NSCLC were selected. Ventilation images were generated from the 4D-CT sets, and each was imported into the treatment-planning system. Ninetieth percentile functional volumes (PFV90), constituting the 10% of the lung volume where the highest ventilation occurs, were generated. Baseline IMRT plans were generated using the lung volume constraint on V20 (<35%), and two additional plans were generated using constraints on the PFV90 without a volume constraint. Dose-volume and dose-function histograms (DVH, DFH) were generated and used to evaluate the planning target volume coverage, lung volume, and functional parameters for comparison of the plans. RESULTS: The mean dose to the PFV90 was reduced by 2.9 Gy, and the DFH at 5 Gy (F5) was reduced by 9.6% (SE = 2.03%). The F5, F10, V5, and V10 were all significantly reduced from the baseline values. We identified a favorable subset of patients for whom there was a further significant improvement in the mean lung dose. CONCLUSIONS: Four-dimensional computed tomography-derived ventilation regions were successfully used as avoidance structures to reduce the DVH and DFH at 5 Gy in all cases. In a subset, there was also a reduction in the F10 and V10 without a change in the V20, suggesting that this technique could be safely used.

22q11.2 microduplication syndrome with associated esophageal atresia/tracheo-esophageal fistula and vascular ring.

This case report describes a patient with a 22q11.2 duplication. His features, which include VACTERL association with an esophageal atresia/tracheo-esophageal fistula and a vascular ring, expand the previously described phenotype for this duplication.

Improved facial nerve identification during parotidectomy with fluorescently labeled peptide.

OBJECTIVES/HYPOTHESIS: Additional intraoperative guidance could reduce the risk of iatrogenic injury during parotid gland cancer surgery. We evaluated the intraoperative use of fluorescently labeled nerve binding peptide NP41 to aid facial nerve identification and preservation during parotidectomy in an orthotopic model of murine parotid gland cancer. We also quantified the accuracy of intraoperative nerve detection for surface and buried nerves in the head and neck with NP41 versus white light (WL) alone. STUDY DESIGN: Twenty-eight mice underwent parotid gland cancer surgeries with additional fluorescence (FL) guidance versus WL reflectance (WLR) alone. Eight mice were used for additional nerve-imaging experiments. METHODS: Twenty-eight parotid tumor-bearing mice underwent parotidectomy. Eight mice underwent imaging of both sides of the face after skin removal. Postoperative assessment of facial nerve function measured by automated whisker tracking were compared between FL guidance (n = 13) versus WL alone (n=15). In eight mice, nerve to surrounding tissue contrast was measured under FL versus WLR for all nerve branches detectable in the field of view. RESULTS: Postoperative facial nerve function after parotid gland cancer surgery tended to be better with additional FL guidance. Fluorescent labeling significantly improved nerve to surrounding tissue contrast for both large and smaller buried nerve branches compared to WLR visualization and improved detection sensitivity and specificity. CONCLUSIONS: NP41 FL imaging significantly aids the intraoperative identification of nerve braches otherwise nearly invisible to the naked eye. Its application in a murine model of parotid gland cancer surgery tended to improve functional preservation of the facial nerve. LEVEL OF EVIDENCE: NA Laryngoscope, 126:2711-2717, 2016.

Combined TP53 mutation/3p loss correlates with decreased radiosensitivity and increased matrix-metalloproteinase activity in head and neck carcinoma.

OBJECTIVE: Patients with head and neck squamous cell carcinoma (HNSCC) containing TP53 mutation and 3p deletion ("double-hit") have poorer prognosis compared to patients with either event alone ("single-hit"). The etiology for worse clinical outcomes in patients with "double-hit" cancers is unclear. We compared radiosensitivity of cell lines containing both TP53 mutations and deletion of Fragile Histidine Triad (FHIT, the gene most commonly associated with 3p deletion) to "single-hit" lines with only TP53 mutation. We compared radiosensitivity in a "single-hit" cell line with TP53 mutation converted to "double-hit" using RNA interference targeting FHIT. Finally, we compared matrixmetalloproteinase-2/9 (MMP-2/9) activity, a previously-established biomarker for tumor aggressiveness, in xenograft tumors derived from these cell lines. MATERIALS/METHODS: TP53 mutation and FHIT deletion profiles of HNSCC lines were established using Cancer Cell Line Encyclopedia (CCLE). We used RNA-interference to convert a "single-hit" cell line (SCC4) to "double-hit". Cultured cells were examined for radiosensitivity and cisplatin sensitivity. MMP-2/9 activity was evaluated in "double-hit" versus "single-hit" tumors using ratiometric activatable cell-penetrating peptide (RACPP) in tongue (n=17) and flank xenografts (n=4). RESULTS: Radiotherapy caused greater double-stranded DNA breaks in "single-hit" vs naturally occurring and engineered "double-hit" cells. In-vivo, "double-hit" xenografts demonstrated higher MMP-2/9 activity compared to "single-hit" xenografts (p<0.01). There was no difference in cisplatin sensitivity between the cell lines. CONCLUSIONS: TP53 mutation combined with FHIT deletion correlates with decreased radiosensitivity in HNC cell lines. Xenograft from "double-hit" cells exhibit increased MMP-2/9 activity. These findings may in part account for the worse clinical outcome seen in patients with HNSCC "double-hit" tumors.

Fluorescent peptides highlight peripheral nerves during surgery in mice.

Nerve preservation is an important goal during surgery because accidental transection or injury leads to significant morbidity, including numbness, pain, weakness or paralysis. Nerves are usually identified by their appearance and relationship to nearby structures or detected by local electrical stimulation (electromyography), but thin or buried nerves are sometimes overlooked. Here, we use phage display to select a peptide that binds preferentially to nerves. After systemic injection of a fluorescently labeled version of the peptide in mice, all peripheral nerves are clearly delineated within 2 h. Contrast between nerve and adjacent tissue is up to tenfold, and useful contrast lasts up to 8 h. No changes in behavior or activity are observed after treatment, indicating a lack of obvious toxicity. The fluorescent probe also labels nerves in human tissue samples. Fluorescence highlighting is independent of axonal integrity, suggesting that the probe could facilitate surgical repair of injured nerves and help prevent accidental transection.

Clinical utility of electrocochleography in the diagnosis and management of Ménière's disease: AOS and ANS membership survey data.

OBJECTIVE: Evaluate the clinical use of electrocochleography (ECoG) for diagnosis/treatment of Ménière's disease among members of the American Otological Society (AOS) and American Neurotology Society (ANS). SUBJECTS: Clinically active members of the AOS/ANS. MAIN OUTCOME MEASURE: Survey responses. RESULTS: A total of 143 responses were received from 344 possible respondents (41.6%). In suspected cases of Ménière's disease, 45.5% of respondents did not use ECoG at all, 17.5 % used ECoG routinely, and 37.1% used it only in questionable cases. Electrocochleography users differed widely in electrode approach and stimulus modality used, with extratympanic approach and click stimuli used most frequently. Most respondents (73.2%) thought that ECoG is a test of indeterminate value. Only 3.6% required an abnormal ECoG to diagnose endolymphatic hydrops. An abnormal test was a requirement to proceed with ablative therapy for just 8.6% of respondents. Still, 77.9% think that ECoG findings do fluctuate with activity of the disorder, but only 18.0% agree that when the ECoG reverts to normal, one can predict remission of symptoms. Almost half of respondents (46.7%) reported that they have now stopped ordering ECoG due to variability in results and lack of correlation with their patients' symptoms. CONCLUSION: Among AOS/ANS members, there is low clinical use of ECoG in diagnosis/management of Ménière's disease. For approximately half of respondents, ECoG has no role in their clinical practice. Electrocochleography was used routinely by only 1 in 6 respondents. Those who used ECoG differed widely in electrode placement and type of stimuli paradigm used.

Erosive external otitis: a novel distinct clinical entity of the external auditory canal in nonimmunosuppressed individuals.

OBJECTIVE: We introduce erosive external otitis (EEO) as a novel erosive process of the external auditory canal in the absence of diabetes or immune suppression. STUDY DESIGN: Case series and literature review. SETTING: Tertiary referral center. PATIENTS: Primary eligibility criteria included patients who had an erosive process of their external auditory canal in the absence of diabetes or immune suppression. INTERVENTION: Surgical debridement and split-thickness skin grafting. MAIN OUTCOME MEASURES: Uneventful wound healing and disease-free long-term follow-up after surgical debridement and skin grafting. RESULTS: Three cases of EEO in the absence of immune suppression or diabetes were diagnosed and treated. All patients required surgical debridement and skin grafting as part of their management. All recovered uneventfully from surgery and had no evidence of recurrence on long-term follow-up. CONCLUSION: We propose that EEO, which occurs in the absence of immune suppression or diabetes, is a clinical entity that is distinct from the more commonly diagnosed malignant external otitis and that the management of EEO is primarily surgical debridement with skin grafting.

Post-training intrabasolateral amygdala infusions of dopamine modulate consolidation of inhibitory avoidance memory: involvement of noradrenergic and cholinergic systems.

There is extensive evidence that several neurotransmitter systems within the basolateral amygdala (BLA) influence memory consolidation. The present study investigated the influence of dopamine (DA) in the BLA on the consolidation of memory for inhibitory avoidance (IA) training. Male Sprague-Dawley rats (approximately 300 g) were trained on a step-through IA task and, 48 h later, tested for retention as indexed by their latencies to enter the shock compartment on the test day. Drugs were infused into the BLA or central amygdala nucleus (CEA) immediately or 3 h after training via bilateral cannulae. DA infused into the BLA immediately after training enhanced retention, whereas DA infused into the BLA 3 h after training or into the CEA did not affect retention. Infusions of the dopaminergic antagonist cis-Flupenthixol together with DA blocked the DA-induced memory enhancement. Immediate post-training intra-BLA infusions of the D1 receptor antagonist SCH 23390 or the D2 receptor antagonist sulpiride impaired retention. beta-adrenergic or muscarinic cholinergic receptor antagonists coinfused into the BLA with DA blocked the memory enhancing effects of DA. These findings indicate that dopaminergic activation within the BLA modulates memory consolidation and that the modulation involves activation of both D1 and D2 receptors and concurrent activation of beta-adrenergic and cholinergic influences within the BLA.