RESUMO
The ventrolateral medulla (VLM) is a crucial region in the brain for visceral and somatic control, serving as a significant source of synaptic input to the spinal cord. Experimental studies have shown that gene expression in individual VLM neurons is predictive of their function. However, the molecular and cellular organization of the VLM has remained uncertain. This study aimed to create a comprehensive dataset of VLM cells using single-cell RNA sequencing in male and female mice. The dataset was enriched with targeted sequencing of spinally-projecting and adrenergic/noradrenergic VLM neurons. Based on differentially expressed genes, the resulting dataset of 114,805 VLM cells identifies 23 subtypes of neurons, excluding those in the inferior olive, and five subtypes of astrocytes. Spinally-projecting neurons were found to be abundant in seven subtypes of neurons, which were validated through in situ hybridization. These subtypes included adrenergic/noradrenergic neurons, serotonergic neurons, and neurons expressing gene markers associated with premotor neurons in the ventromedial medulla. Further analysis of adrenergic/noradrenergic neurons and serotonergic neurons identified nine and six subtypes, respectively, within each class of monoaminergic neurons. Marker genes that identify the neural network responsible for breathing were concentrated in two subtypes of neurons, delineated from each other by markers for excitatory and inhibitory neurons. These datasets are available for public download and for analysis with a user-friendly interface. Collectively, this study provides a fine-scale molecular identification of cells in the VLM, forming the foundation for a better understanding of the VLM's role in vital functions and motor control.
Assuntos
Bulbo , Neurônios , Medula Espinal , Animais , Bulbo/citologia , Bulbo/fisiologia , Camundongos , Masculino , Feminino , Neurônios/fisiologia , Medula Espinal/citologia , Medula Espinal/fisiologia , Camundongos Endogâmicos C57BL , Sistema Nervoso Autônomo/fisiologia , Sistema Nervoso Autônomo/citologiaRESUMO
Cardiovagal neurons (CVNs) innervate cardiac ganglia through the vagus nerve to control cardiac function. Although the cardioinhibitory role of CVNs in nucleus ambiguus (CVNNA) is well established, the nature and functionality of CVNs in dorsal motor nucleus of the vagus (CVNDMV) is less clear. We therefore aimed to characterize CVNDMV anatomically, physiologically, and functionally. Optogenetically activating cholinergic DMV neurons resulted in robust bradycardia through peripheral muscarinic (parasympathetic) and nicotinic (ganglionic) acetylcholine receptors, but not beta-1-adrenergic (sympathetic) receptors. Retrograde tracing from the cardiac fat pad labeled CVNNA and CVNDMV through the vagus nerve. Using whole-cell patch-clamp, CVNDMV demonstrated greater hyperexcitability and spontaneous action potential firing ex vivo despite similar resting membrane potentials, compared to CVNNA. Chemogenetically activating DMV also caused significant bradycardia with a correlated reduction in anxiety-like behavior. Thus, DMV contains uniquely hyperexcitable CVNs and is capable of cardioinhibition and robust anxiolysis.
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Central respiratory chemoreceptors are cells in the brain that regulate breathing in relation to arterial pH and PCO2. Neurons located at the retrotrapezoid nucleus (RTN) have been hypothesized to be central chemoreceptors and/or to be part of the neural network that drives the central respiratory chemoreflex. The inhibition or ablation of RTN chemoreceptor neurons has offered important insights into the role of these cells on central respiratory chemoreception and the neural control of breathing over almost 60 years since the original identification of acid-sensitive properties of this ventral medullary site. Here, we discuss the current definition of chemoreceptor neurons in the RTN and describe how this definition has evolved over time. We then summarize the results of studies that use loss-of-function approaches to evaluate the effects of disrupting the function of RTN neurons on respiration. These studies offer evidence that RTN neurons are indispensable for the central respiratory chemoreflex in mammals and exert a tonic drive to breathe at rest. Moreover, RTN has an interdependent relationship with oxygen sensing mechanisms for the maintenance of the neural drive to breathe and blood gas homeostasis. Collectively, RTN neurons are a genetically-defined group of putative central respiratory chemoreceptors that generate CO2-dependent drive that supports eupneic breathing and stimulates the hypercapnic ventilatory reflex.
Assuntos
Células Quimiorreceptoras , Bulbo , Animais , Células Quimiorreceptoras/fisiologia , Bulbo/fisiologia , Hipercapnia , Respiração , Neurônios/fisiologia , Dióxido de Carbono , MamíferosRESUMO
The vagus nerve vitally connects the brain and body to coordinate digestive, cardiorespiratory, and immune functions. Its efferent neurons, which project their axons from the brainstem to the viscera, are thought to comprise "functional units" - neuron populations dedicated to the control of specific vagal reflexes or organ functions. Previous research indicates that these functional units differ from one another anatomically, neurochemically, and physiologically but have yet to define their identity in an experimentally tractable way. However, recent work with genetic technology and single-cell genomics suggests that genetically distinct subtypes of neurons may be the functional units of the efferent vagus. Here we review how these approaches are revealing the organizational principles of the efferent vagus in unprecedented detail.
Assuntos
Neurônios Eferentes , Nervo Vago , Nervo Vago/metabolismo , Neurônios/fisiologiaRESUMO
Cardiovagal neurons (CVNs) innervate cardiac ganglia through the vagus nerve to control cardiac function. Although the cardioinhibitory role of CVNs in nucleus ambiguus (CVNNA) is well established, the nature and functionality of CVNs in dorsal motor nucleus of the vagus (CVNDMV) is less clear. We therefore aimed to characterize CVNDMV anatomically, physiologically, and functionally. Optogenetically activating cholinergic DMV neurons resulted in robust bradycardia through peripheral muscarinic (parasympathetic) and nicotinic (ganglionic) acetylcholine receptors, but not beta-1-adrenergic (sympathetic) receptors. Retrograde tracing from the cardiac fat pad labeled CVNNA and CVNDMV through the vagus nerve. Using whole cell patch clamp, CVNDMV demonstrated greater hyperexcitability and spontaneous action potential firing ex vivo despite similar resting membrane potentials, compared to CVNNA. Chemogenetically activating DMV also caused significant bradycardia with a correlated reduction in anxiety-like behavior. Thus, DMV contains uniquely hyperexcitable CVNs capable of cardioinhibition and robust anxiolysis.
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The interoceptive homeostatic mechanism that controls breathing, blood gases and acid-base balance in response to changes in CO2 /H+ is exquisitely sensitive, with convergent roles proposed for chemosensory brainstem neurons in the retrotrapezoid nucleus (RTN) and their supporting glial cells. For astrocytes, a central role for NBCe1, a Na+ -HCO3 - cotransporter encoded by Slc4a4, has been envisaged in multiple mechanistic models (i.e. underlying enhanced CO2 -induced local extracellular acidification or purinergic signalling). We tested these NBCe1-centric models by using conditional knockout mice in which Slc4a4 was deleted from astrocytes. In GFAP-Cre;Slc4a4fl/fl mice we found diminished expression of Slc4a4 in RTN astrocytes by comparison to control littermates, and a concomitant reduction in NBCe1-mediated current. Despite disrupted NBCe1 function in RTN-adjacent astrocytes from these conditional knockout mice, CO2 -induced activation of RTN neurons or astrocytes in vitro and in vivo, and CO2 -stimulated breathing, were indistinguishable from NBCe1-intact littermates; hypoxia-stimulated breathing and sighs were likewise unaffected. We obtained a more widespread deletion of NBCe1 in brainstem astrocytes by using tamoxifen-treated Aldh1l1-Cre/ERT2;Slc4a4fl/fl mice. Again, there was no difference in effects of CO2 or hypoxia on breathing or on neuron/astrocyte activation in NBCe1-deleted mice. These data indicate that astrocytic NBCe1 is not required for the respiratory responses to these chemoreceptor stimuli in mice, and that any physiologically relevant astrocytic contributions must involve NBCe1-independent mechanisms. KEY POINTS: The electrogenic NBCe1 transporter is proposed to mediate local astrocytic CO2 /H+ sensing that enables excitatory modulation of nearby retrotrapezoid nucleus (RTN) neurons to support chemosensory control of breathing. We used two different Cre mouse lines for cell-specific and/or temporally regulated deletion of the NBCe1 gene (Slc4a4) in astrocytes to test this hypothesis. In both mouse lines, Slc4a4 was depleted from RTN-associated astrocytes but CO2 -induced Fos expression (i.e. cell activation) in RTN neurons and local astrocytes was intact. Likewise, respiratory chemoreflexes evoked by changes in CO2 or O2 were unaffected by loss of astrocytic Slc4a4. These data do not support the previously proposed role for NBCe1 in respiratory chemosensitivity mediated by astrocytes.
Assuntos
Astrócitos , Simportadores , Animais , Camundongos , Astrócitos/fisiologia , Dióxido de Carbono/metabolismo , Células Quimiorreceptoras/metabolismo , Íons/metabolismo , Camundongos Knockout , Simportadores de Sódio-Bicarbonato/genética , Simportadores de Sódio-Bicarbonato/metabolismo , Simportadores/metabolismoRESUMO
Respiratory chemoreceptor activity encoding arterial Pco2 and Po2 is a critical determinant of ventilation. Currently, the relative importance of several putative chemoreceptor mechanisms for maintaining eupneic breathing and respiratory homeostasis is debated. Transcriptomic and anatomic evidence suggests that bombesin-related peptide Neuromedin-B (Nmb) expression identifies chemoreceptor neurons in the retrotrapezoid nucleus (RTN) that mediate the hypercapnic ventilatory response, but functional support is missing. In this study, we generated a transgenic Nmb-Cre mouse and used Cre-dependent cell ablation and optogenetics to test the hypothesis that RTN Nmb neurons are necessary for the CO2-dependent drive to breathe in adult male and female mice. Selective ablation of â¼95% of RTN Nmb neurons causes compensated respiratory acidosis because of alveolar hypoventilation, as well as profound breathing instability and respiratory-related sleep disruption. Following RTN Nmb lesion, mice were hypoxemic at rest and were prone to severe apneas during hyperoxia, suggesting that oxygen-sensitive mechanisms, presumably the peripheral chemoreceptors, compensate for the loss of RTN Nmb neurons. Interestingly, ventilation following RTN Nmb -lesion was unresponsive to hypercapnia, but behavioral responses to CO2 (freezing and avoidance) and the hypoxia ventilatory response were preserved. Neuroanatomical mapping shows that RTN Nmb neurons are highly collateralized and innervate the respiratory-related centers in the pons and medulla with a strong ipsilateral preference. Together, this evidence suggests that RTN Nmb neurons are dedicated to the respiratory effects of arterial Pco2/pH and maintain respiratory homeostasis in intact conditions and suggest that malfunction of these neurons could underlie the etiology of certain forms of sleep-disordered breathing in humans.SIGNIFICANCE STATEMENT Respiratory chemoreceptors stimulate neural respiratory motor output to regulate arterial Pco2 and Po2, thereby maintaining optimal gas exchange. Neurons in the retrotrapezoid nucleus (RTN) that express the bombesin-related peptide Neuromedin-B are proposed to be important in this process, but functional evidence has not been established. Here, we developed a transgenic mouse model and demonstrated that RTN neurons are fundamental for respiratory homeostasis and mediate the stimulatory effects of CO2 on breathing. Our functional and anatomic data indicate that Nmb-expressing RTN neurons are an integral component of the neural mechanisms that mediate CO2-dependent drive to breathe and maintain alveolar ventilation. This work highlights the importance of the interdependent and dynamic integration of CO2- and O2-sensing mechanisms in respiratory homeostasis of mammals.
Assuntos
Bombesina , Dióxido de Carbono , Humanos , Camundongos , Masculino , Feminino , Animais , Bombesina/metabolismo , Respiração , Células Quimiorreceptoras/fisiologia , Hipercapnia , Homeostase , Camundongos Transgênicos , Oxigênio/metabolismo , Neurônios/fisiologia , Centro Respiratório , MamíferosRESUMO
OBJECTIVES: The aim of the study is to determine whether a positive OncoE6 Anal Test result has statistically significant higher odds of being associated with high-grade squamous intraepithelial lesion (HSIL) and to calculate sensitivity and specificity of this test for predicting HSIL in adult men who have sex with men and are living with HIV (MSMLWH). MATERIALS AND METHODS: Men living with HIV 18 years or older having ≥atypical squamous cells of undetermined significance-grade anal cytology results were eligible to enroll in this cross-sectional study. Anal samples were collected just before the high-resolution anoscopy procedure. OncoE6 Anal Test results were compared with histology, the reference standard. Sensitivity, specificity, and odds ratio were calculated using HSIL as the threshold. RESULTS: Two hundred seventy-seven consented MSMLWH were enrolled between June 2017 and January 2022. Of these, 219 (79.1%) had biopsies obtained and histology performed; 81 of 219 participants (37%) had 1 or more biopsies with HSIL results while the remaining 138 of 219 (63%) had only low-grade squamous intraepithelial lesion or were negative for dysplasia. Anal samples from 7 participants (8.6%, 7/81) with HSIL and 3 (2.2%, 3/138) with low-grade squamous intraepithelial lesion had positive OncoE6 Anal Test results. Odds of having HSIL were 4.26 times higher among participants testing positive for HPV16/HPV18 E6 oncoprotein(s) (OR = 4.26, 95% CI = 1.07-16.95, p = .04). The OncoE6 Anal Test demonstrated excellent specificity, 97.83% (93.78-99.55), but poor sensitivity, 8.64% (3.55-17.0). CONCLUSIONS: In this highest-risk population for anal cancer, one could combine the OncoE6 Anal Test, having excellent specificity, with the anal Pap test, having higher sensitivity. Patients found having both an abnormal anal Pap and positive OncoE6 Anal Test result could be triaged for rapid scheduling of their high-resolution anoscopy.
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Neoplasias do Ânus , Carcinoma in Situ , Carcinoma de Células Escamosas , Infecções por HIV , Infecções por Papillomavirus , Minorias Sexuais e de Gênero , Lesões Intraepiteliais Escamosas , Adulto , Masculino , Humanos , Homossexualidade Masculina , Estudos Transversais , Canal Anal/patologia , Carcinoma in Situ/patologia , Carcinoma de Células Escamosas/patologia , Lesões Intraepiteliais Escamosas/patologia , Neoplasias do Ânus/patologia , Infecções por HIV/complicações , Infecções por HIV/epidemiologia , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/epidemiologia , Infecções por Papillomavirus/patologia , PapillomaviridaeRESUMO
The nucleus ambiguus (nAmb) provides parasympathetic control of cardiorespiratory functions as well as motor control of the upper airways and striated esophagus. A subset of nAmb neurons innervates the heart through the vagus nerve to control cardiac function at rest and during key autonomic reflexes such as the mammalian diving reflex. These cardiovagal nAmb neurons may be molecularly and anatomically distinct, but how they differ from other nAmb neurons in the adult brain remains unclear. We therefore classified adult mouse nAmb neurons based on their genome-wide expression profiles, innervation of cardiac ganglia, and ability to control HR. Our integrated analysis of single-nucleus RNA-sequencing data predicted multiple molecular subtypes of nAmb neurons. Mapping the axon projections of one nAmb neuron subtype, Npy2r-expressing nAmb neurons, showed that they innervate cardiac ganglia. Optogenetically stimulating all nAmb vagal efferent neurons dramatically slowed HR to a similar extent as selectively stimulating Npy2r+ nAmb neurons, but not other subtypes of nAmb neurons. Finally, we trained mice to perform voluntary underwater diving, which we use to show Npy2r+ nAmb neurons are activated by the diving response, consistent with a cardiovagal function for this nAmb subtype. These results together reveal the molecular organization of nAmb neurons and its control of heart rate.
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In mammals, the pontine noradrenergic system influences nearly every aspect of central nervous system function. A subpopulation of pontine noradrenergic neurons, called A5, are thought to be important in the cardiovascular response to physical stressors, yet their function is poorly defined. We hypothesized that activation of A5 neurons drives a sympathetically mediated increase in blood pressure (BP). To test this hypothesis, we conducted a comprehensive assessment of the cardiovascular effects of chemogenetic stimulation of A5 neurons in male and female adult rats using intersectional genetic and anatomical targeting approaches. Chemogenetic stimulation of A5 neurons in freely behaving rats elevated BP by 15 mmHg and increased cardiac baroreflex sensitivity with a negligible effect on resting HR. Importantly, A5 stimulation had no detectable effect on locomotor activity, metabolic rate, or respiration. Under anesthesia, stimulation of A5 neurons produced a marked elevation in visceral sympathetic nerve activity (SNA) and no change in skeletal muscle SNA, showing that A5 neurons preferentially stimulate visceral SNA. Interestingly, projection mapping indicates that A5 neurons target sympathetic preganglionic neurons throughout the spinal cord and parasympathetic preganglionic neurons throughout in the brainstem, as well as the nucleus of the solitary tract, and ventrolateral medulla. Moreover, in situ hybridization and immunohistochemistry indicate that a subpopulation of A5 neurons coreleases glutamate and monoamines. Collectively, this study suggests A5 neurons are a central modulator of autonomic function with a potentially important role in sympathetically driven redistribution of blood flow from the visceral circulation to critical organs and skeletal muscle.
Assuntos
Neurônios Adrenérgicos , Neurônios Adrenérgicos/fisiologia , Animais , Pressão Sanguínea/fisiologia , Feminino , Glutamatos/farmacologia , Masculino , Mamíferos , Ponte/fisiologia , Ratos , Sistema Nervoso Simpático/fisiologiaRESUMO
The action of leptin in brain to increase sympathetic nerve activity (SNA) and blood pressure depends upon functional Angiotensin II (AngII) type 1a receptors (AT1aR); however, the sites and mechanism of interaction are unknown. Here we identify one site, the hypothalamic arcuate nucleus (ArcN), since prior local blockade of AT1aR in the ArcN with losartan or candesartan in anesthetized male rats essentially eliminated the sympathoexcitatory and pressor responses to ArcN leptin nanoinjections. Unlike mice, in male and female rats, AT1aR and LepR rarely co-localized, suggesting that this interdependence occurs indirectly, via a local interneuron or network of neurons. ArcN leptin increases SNA by activating pro-opiomelanocortin (POMC) inputs to the PVN, but this activation requires simultaneous suppression of tonic PVN Neuropeptide Y (NPY) sympathoinhibition. Because AngII-AT1aR inhibits ArcN NPY neurons, we propose that loss of AT1aR suppression of NPY blocks leptin-induced increases in SNA; in other words, ArcN-AngII-AT1aR is a gatekeeper for leptin-induced sympathoexcitation. With obesity, both leptin and AngII increase; therefore, the increased AT1aR activation could open the gate, allowing leptin (and insulin) to drive sympathoexcitation unabated, leading to hypertension.
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Núcleo Arqueado do Hipotálamo , Leptina , Angiotensina II/farmacologia , Animais , Núcleo Arqueado do Hipotálamo/metabolismo , Pressão Sanguínea , Feminino , Leptina/metabolismo , Leptina/farmacologia , Masculino , Camundongos , Neuropeptídeo Y/metabolismo , Neuropeptídeo Y/farmacologia , Núcleo Hipotalâmico Paraventricular/metabolismo , Ratos , Sistema Nervoso Simpático/metabolismoRESUMO
Motor control of the striated esophagus originates in the nucleus ambiguus (nAmb), a vagal motor nucleus that also contains upper airway motor neurons and parasympathetic preganglionic neurons for the heart and lungs. We disambiguate nAmb neurons based on their genome-wide expression profiles, efferent circuitry, and ability to control esophageal muscles. Our single-cell RNA sequencing analysis predicts three molecularly distinct nAmb neuron subtypes and annotates them by subtype-specific marker genes: Crhr2, Vipr2, and Adcyap1. Mapping the axon projections of the nAmb neuron subtypes reveals that Crhr2nAmb neurons innervate the esophagus, raising the possibility that they control esophageal muscle function. Accordingly, focal optogenetic stimulation of cholinergic Crhr2+ fibers in the esophagus results in contractions. Activating Crhr2nAmb neurons has no effect on heart rate, a key parasympathetic function of the nAmb, whereas activating all of the nAmb neurons robustly suppresses heart rate. Together, these results reveal a genetically defined circuit for motor control of the esophagus.
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Bulbo , Nervo Vago , Frequência Cardíaca/fisiologia , Bulbo/fisiologia , Neurônios Motores , Músculo Liso , Nervo Vago/fisiologiaRESUMO
Hemorrhage initially triggers a rise in sympathetic nerve activity (SNA) that maintains blood pressure (BP); however, SNA is suppressed following severe blood loss causing hypotension. We hypothesized that adrenergic C1 neurons in the rostral ventrolateral medulla (C1RVLM) drive the increase in SNA during compensated hemorrhage, and a reduction in C1RVLM contributes to hypotension during decompensated hemorrhage. Using fiber photometry, we demonstrate that C1RVLM activity increases during compensated hemorrhage and falls at the onset of decompensated hemorrhage. Using optogenetics combined with direct recordings of SNA, we show that C1RVLM activation mediates the rise in SNA and contributes to BP stability during compensated hemorrhage, whereas a suppression of C1RVLM activity is associated with cardiovascular collapse during decompensated hemorrhage. Notably, re-activating C1RVLM during decompensated hemorrhage restores BP to normal levels. In conclusion, C1 neurons are a nodal point for the sympathetic response to blood loss.
Assuntos
Neurônios Adrenérgicos , Hipotensão , Adrenérgicos , Animais , Pressão Arterial , Pressão Sanguínea/fisiologia , Hemorragia , Bulbo/fisiologia , Ratos , Ratos Sprague-Dawley , Sistema Nervoso Simpático/fisiologiaRESUMO
Nursing service development or innovation projects, even small-scale ones, can be difficult to deliver and evaluate, due to a lack of resources and support. Results can also be difficult to disseminate, limiting transfer of learning. This paper presents findings from a realist evaluation of 10 small projects supported by the Queen's Nursing Institute Homeless and Inclusion Health Programme to deliver innovation in health care for people experiencing homelessness and other marginalised groups. These nurse-led projects were funded by the Queen's Nursing Institute and the Oak Foundation, and were largely successful in achieving outcomes to support the improved health of people experiencing homelessness and other marginalised groups. This realist evaluation explores the factors that contributed to the delivery of positive outcomes. All were impacted by the context and the response (mechanisms) of people experiencing homelessness and staff within these settings. It is hoped that the lessons learned will enable better support for nurse innovation projects in the future.
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Pessoas Mal Alojadas , Papel do Profissional de Enfermagem , HumanosRESUMO
Hyperventilation reliably provokes seizures in patients diagnosed with absence epilepsy. Despite this predictable patient response, the mechanisms that enable hyperventilation to powerfully activate absence seizure-generating circuits remain entirely unknown. By utilizing gas exchange manipulations and optogenetics in the WAG/Rij rat, an established rodent model of absence epilepsy, we demonstrate that absence seizures are highly sensitive to arterial carbon dioxide, suggesting that seizure-generating circuits are sensitive to pH. Moreover, hyperventilation consistently activated neurons within the intralaminar nuclei of the thalamus, a structure implicated in seizure generation. We show that intralaminar thalamus also contains pH-sensitive neurons. Collectively, these observations suggest that hyperventilation activates pH-sensitive neurons of the intralaminar nuclei to provoke absence seizures.
Assuntos
Alcalose Respiratória/patologia , Convulsões , Animais , Dióxido de Carbono , Concentração de Íons de Hidrogênio , Hipóxia , Núcleos Intralaminares do Tálamo/citologia , Masculino , Neurônios/fisiologia , RatosRESUMO
Arousal from sleep in response to CO2 is a life-preserving reflex that enhances ventilatory drive and facilitates behavioural adaptations to restore eupnoeic breathing. Recurrent activation of the CO2 -arousal reflex is associated with sleep disruption in obstructive sleep apnoea. In this review we examine the role of chemoreceptors in the carotid bodies, the retrotrapezoid nucleus and serotonergic neurons in the dorsal raphe in the CO2 -arousal reflex. We also provide an overview of the supra-medullary structures that mediate CO2 -induced arousal. We propose a framework for the CO2 -arousal reflex in which the activity of the chemoreceptors converges in the parabrachial nucleus to trigger cortical arousal.
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Dióxido de Carbono , Células Quimiorreceptoras , Nível de Alerta , Respiração , SonoRESUMO
Although anal cancers represent just 0.5 % of all new cancer cases in U.S., rates have increased markedly, with highest rates in HIV-infected MSM. American Cancer Society estimates there will be â¼9090 new cases and â¼1420 deaths in 2021. We compared Roche Linear Array HPV Genotyping (Roche) and AmpFire HPV Genotyping (AmpFire) assays for concordance and agreement to detect 15â¯hr-HPV types from 151 anal specimens. Within run precision of AmpFire was assessed on 50 anal specimens. Specimens with Roche Combo-positive and HPV33, HPV35 and/or HPV58-positive results were further tested using HPV52-specific TaqMan assay. AmpFire generated valid results on 149/151 (98.7 %) specimens; 135/149 (90.6 %) and 134/149 (89.9 %) had detectable HR-HPV DNA by AmpFire or Roche, respectively. Overall concordance was 89.8 % (2007/2235, κâ¯=â¯0.65). HPV16 showed highest overall concordance at 93.3 % (139/149, κâ¯=â¯0.84). HPV68 had lowest overall concordance at 77.2 % (115/149, κâ¯=â¯0.28). Kappa values were interpreted as being moderate or good for all other HR-HPV types. Within run precision generated 744/750 concordant results; R2 valueâ¯=â¯0.97 (pâ¯<â¯0.0001) (Mantel Test). In conclusion, AmpFire and Roche demonstrated good inter-assay agreement for detecting most HR-HPV types from anal samples, with AmpFire detecting a broader range of HPV68 subtypes and detecting HPV52 without the need for confirmatory testing.