RESUMO
Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels1 are essential for pacemaking activity and neural signalling2,3. Drugs inhibiting HCN1 are promising candidates for management of neuropathic pain4 and epileptic seizures5. The general anaesthetic propofol (2,6-di-iso-propylphenol) is a known HCN1 allosteric inhibitor6 with unknown structural basis. Here, using single-particle cryo-electron microscopy and electrophysiology, we show that propofol inhibits HCN1 by binding to a mechanistic hotspot in a groove between the S5 and S6 transmembrane helices. We found that propofol restored voltage-dependent closing in two HCN1 epilepsy-associated polymorphisms that act by destabilizing the channel closed state: M305L, located in the propofol-binding site in S5, and D401H in S6 (refs. 7,8). To understand the mechanism of propofol inhibition and restoration of voltage-gating, we tracked voltage-sensor movement in spHCN channels and found that propofol inhibition is independent of voltage-sensor conformational changes. Mutations at the homologous methionine in spHCN and an adjacent conserved phenylalanine in S6 similarly destabilize closing without disrupting voltage-sensor movements, indicating that voltage-dependent closure requires this interface intact. We propose a model for voltage-dependent gating in which propofol stabilizes coupling between the voltage sensor and pore at this conserved methionine-phenylalanine interface in HCN channels. These findings unlock potential exploitation of this site to design specific drugs targeting HCN channelopathies.
Assuntos
Epilepsia , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização , Ativação do Canal Iônico , Mutação , Canais de Potássio , Propofol , Humanos , Sítios de Ligação , Microscopia Crioeletrônica , Eletrofisiologia , Epilepsia/tratamento farmacológico , Epilepsia/genética , Epilepsia/metabolismo , Células HEK293 , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/antagonistas & inibidores , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/química , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/genética , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/ultraestrutura , Ativação do Canal Iônico/efeitos dos fármacos , Ativação do Canal Iônico/genética , Metionina/genética , Metionina/metabolismo , Modelos Moleculares , Movimento/efeitos dos fármacos , Fenilalanina/genética , Fenilalanina/metabolismo , Polimorfismo Genético , Canais de Potássio/química , Canais de Potássio/genética , Canais de Potássio/metabolismo , Canais de Potássio/ultraestrutura , Propofol/farmacologia , Propofol/químicaRESUMO
The pace of human brain development is highly protracted compared with most other species1-7. The maturation of cortical neurons is particularly slow, taking months to years to develop adult functions3-5. Remarkably, such protracted timing is retained in cortical neurons derived from human pluripotent stem cells (hPSCs) during in vitro differentiation or upon transplantation into the mouse brain4,8,9. Those findings suggest the presence of a cell-intrinsic clock setting the pace of neuronal maturation, although the molecular nature of this clock remains unknown. Here we identify an epigenetic developmental programme that sets the timing of human neuronal maturation. First, we developed a hPSC-based approach to synchronize the birth of cortical neurons in vitro which enabled us to define an atlas of morphological, functional and molecular maturation. We observed a slow unfolding of maturation programmes, limited by the retention of specific epigenetic factors. Loss of function of several of those factors in cortical neurons enables precocious maturation. Transient inhibition of EZH2, EHMT1 and EHMT2 or DOT1L, at progenitor stage primes newly born neurons to rapidly acquire mature properties upon differentiation. Thus our findings reveal that the rate at which human neurons mature is set well before neurogenesis through the establishment of an epigenetic barrier in progenitor cells. Mechanistically, this barrier holds transcriptional maturation programmes in a poised state that is gradually released to ensure the prolonged timeline of human cortical neuron maturation.
Assuntos
Epigênese Genética , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Embrionárias Humanas , Células-Tronco Neurais , Neurogênese , Neurônios , Adulto , Animais , Humanos , Camundongos , Antígenos de Histocompatibilidade/metabolismo , Histona-Lisina N-Metiltransferase/antagonistas & inibidores , Histona-Lisina N-Metiltransferase/metabolismo , Células-Tronco Embrionárias Humanas/citologia , Células-Tronco Embrionárias Humanas/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Neurogênese/genética , Neurônios/citologia , Neurônios/metabolismo , Fatores de Tempo , Transcrição GênicaRESUMO
2,6-di-tert-butylphenol (2,6-DTBP) ameliorates mechanical allodynia and thermal hyperalgesia produced by partial sciatic nerve ligation in mice, and selectively inhibits HCN1 channel gating. We hypothesized that the clinically utilized non-anesthetic dimerized congener of 2,6-DTBP, probucol (2,6-di-tert-butyl-4-[2-(3,5-di-tert-butyl-4-hydroxyphenyl)sulfanylpropan-2-ylsulfanyl]phenol), would relieve the neuropathic phenotype that results from peripheral nerve damage, and that the anti-hyperalgesic efficacy in vivo would correlate with HCN1 channel inhibition in vitro. A single oral dose of probucol (800 mg/kg) relieved mechanical allodynia and thermal hyperalgesia in a mouse spared-nerve injury neuropathic pain model. While the low aqueous solubility of probucol precluded assessment of its possible interaction with HCN1 channels, our results, in conjunction with recent data demonstrating that probucol reduces lipopolysaccharide-induced mechanical allodynia and thermal hyperalgesia, support the testing/development of probucol as a non-opioid, oral antihyperalgesic albeit one of unknown mechanistic action.
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Current operational models for nuclear cloud rise over land were developed and validated using observations from shallow-buried or surface detonations, where lofted soil quickly mixed with fission products from the detonation. These models poorly predict fallout from elevated detonations near the fallout-free height of burst (FFHOB), where interactions with the ground are limited and the mixing of fission products and lofted soil is incomplete. Fallout-free is a misnomer at this HOB, as fallout was observed in these cases, but was below the levels of concern, especially off-grounds of the nuclear test site. To correctly characterize and model fallout from detonations near the FFHOB, models must be developed which can capture the stratified nature of the particle and activity-size distributions within the cloud. Previously, it was shown that the Weather Research and Forecasting (WRF) model can accurately simulate nuclear cloud rise for airbursts with little to no ground interactions (Arthur et al., 2021). That work is expanded here by (1) using a radiation-hydrodynamics code to improve the fireball initialization in WRF, (2) further developing an aerosol package from WRF-Chem to simulate lofted soil, and (3) combining the WRF cloud rise simulations with the operational models used at the National Atmospheric Release Advisory Center (NARAC) for fallout modeling. Using this combination of codes, the Upshot-Knothole Grable detonation, which was just below the FFHOB, is simulated from seconds after detonation through cloud rise and fallout, and results are compared to historical test data. The results show improved prediction of dose rate and highlight the need to correctly characterize the entrainment of material into the cloud and the subsequent mixing of fission products with entrained material.
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Monitoramento de Radiação , Cinza Radioativa , Cinza Radioativa/análise , Monitoramento de Radiação/métodos , Modelos Teóricos , Tempo (Meteorologia) , Aerossóis/análiseRESUMO
BACKGROUND: Neuropathic pain impairs quality of life, is widely prevalent, and incurs significant costs. Current pharmacological therapies have poor/no efficacy and significant adverse effects; safe and effective alternatives are needed. Hyperpolarisation-activated cyclic nucleotide-regulated (HCN) channels are causally implicated in some forms of peripherally mediated neuropathic pain. Whilst 2,6-substituted phenols, such as 2,6-di-tert-butylphenol (26DTB-P), selectively inhibit HCN1 gating and are antihyperalgesic, the development of therapeutically tolerable, HCN-selective antihyperalgesics based on their inverse agonist activity requires that such drugs spare the cardiac isoforms and do not cross the blood-brain barrier. METHODS: In silico molecular dynamics simulation, in vitro electrophysiology, and in vivo rat spared nerve injury methods were used to test whether 'hindered' variants of 26DTB-P (wherein a hydrophilic 'anchor' is attached in the para-position of 26DTB-P via an acyl chain 'tether') had the desired properties. RESULTS: Molecular dynamics simulation showed that membrane penetration of hindered 26DTB-Ps is controlled by a tethered diol anchor without elimination of head group rotational freedom. In vitro and in vivo analysis showed that BP4L-18:1:1, a variant wherein a diol anchor is attached to 26DTB-P via an 18-carbon tether, is an HCN1 inverse agonist and an orally available antihyperalgesic. With a CNS multiparameter optimisation score of 2.25, a >100-fold lower drug load in the brain vs blood, and an absence of adverse cardiovascular or CNS effects, BP4L-18:1:1 was shown to be poorly CNS penetrant and cardiac sparing. CONCLUSIONS: These findings provide a proof-of-concept demonstration that anchor-tethered drugs are a new chemotype for treatment of disorders involving membrane targets.
Assuntos
Agonismo Inverso de Drogas , Neuralgia , Ratos , Animais , Qualidade de Vida , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/uso terapêutico , Neuralgia/tratamento farmacológico , Fenômenos EletrofisiológicosRESUMO
BACKGROUND: Bordetella pertussis epidemics persist as transmission remains unabated despite high acellular pertussis vaccination rates. BPZE1, a live attenuated intranasal pertussis vaccine, was designed to prevent B pertussis infection and disease. We aimed to assess the immunogenicity and safety of BPZE1 compared with the tetanus-diphtheria-acellular pertussis vaccine (Tdap). METHODS: In this double-blind, phase 2b trial at three research centres in the USA, healthy adults aged 18-50 years were randomly assigned (2:2:1:1) via a permuted block randomisation schedule to receive BPZE1 vaccination followed by BPZE1 attenuated challenge, BPZE1 vaccination followed by placebo challenge, Tdap followed by BPZE1 attenuated challenge, or Tdap followed by placebo challenge. On day 1, lyophilised BPZE1 was reconstituted with sterile water and given intranasally (0·4 mL delivered to each nostril), whereas Tdap was given intramuscularly. To maintain masking, participants in the BPZE1 groups received an intramuscular saline injection, and those in the Tdap groups received intranasal lyophilised placebo buffer. The attenuated challenge took place on day 85. The primary immunogenicity endpoint was the proportion of participants achieving nasal secretory IgA seroconversion against at least one B pertussis antigen on day 29 or day 113. Reactogenicity was assessed up to 7 days after vaccination and challenge, and adverse events were recorded for 28 days after vaccination and challenge. Serious adverse events were monitored throughout the study. This trial is registered with ClinicalTrials.gov, NCT03942406. FINDINGS: Between June 17 and Oct 3, 2019, 458 participants were screened and 280 were randomly assigned to the main cohort: 92 to the BPZE1-BPZE1 group, 92 to the BPZE1-placebo group, 46 to the Tdap-BPZE1 group, and 50 to the Tdap-placebo group. Seroconversion of at least one B pertussis-specific nasal secretory IgA was recorded in 79 (94% [95% CI 87-98]) of 84 participants in the BPZE1-BPZE1 group, 89 (95% [88-98]) of 94 in the BPZE1-placebo group, 38 (90% [77-97]) of 42 in the Tdap-BPZE1 group, and 42 (93% [82-99]) of 45 in the Tdap-placebo group. BPZE1 induced broad and consistent B pertussis-specific mucosal secretory IgA responses, whereas Tdap did not induce consistent mucosal secretory IgA responses. Both vaccines were well tolerated, with mild reactogenicity and no serious adverse events related to study vaccination. INTERPRETATION: BPZE1 induced nasal mucosal immunity and produced functional serum responses. BPZE1 has the potential to avert B pertussis infections, which ultimately could lead to reduced transmission and diminished epidemic cycles. These results should be confirmed in large phase 3 trials. FUNDING: ILiAD Biotechnologies.
Assuntos
Vacinas contra Difteria, Tétano e Coqueluche Acelular , Difteria , Tétano , Coqueluche , Adulto , Humanos , Difteria/prevenção & controle , Vacinas contra Difteria, Tétano e Coqueluche Acelular/imunologia , Método Duplo-Cego , Imunoglobulina A Secretora , Tétano/prevenção & controle , Vacinas Atenuadas/imunologia , Coqueluche/prevenção & controle , Adulto Jovem , Pessoa de Meia-Idade , AdolescenteRESUMO
Transplant surgery is an area that gives rise to a number of ethical considerations. As medicine continues to expand the boundaries of what is technically possible, we must consider the ethical implications of our interventions, not solely on patients and society, but also on those asked to provide that care. Here, we consider physician participation in procedures required to provide patient care in the context of the ethical convictions held by the physician, with an emphasis on organ donation after circulatory determination of death. Strategies that can be used to mitigate any potential negative impact on the psychological well-being of members of the patient care team are considered.
Assuntos
Transplante de Órgãos , Obtenção de Tecidos e Órgãos , Humanos , Morte , Equipe de Assistência ao Paciente , Doadores de TecidosRESUMO
William Morton introduced the world to ether anesthesia for use during surgery in the Bullfinch Building of the Massachusetts General Hospital on October 16, 1846. For nearly two centuries, the prevailing wisdom had been that the effects of general anesthetics were rapidly and fully reversible, with no apparent long-term adverse sequelae. Despite occasional concerns of a possible association between surgery and anesthesia with dementia since 1887 (Savage, 1887), our initial belief was robustly punctured following the publication in 1998 of the International Study of Post-Operative Cognitive Dysfunction [ISPOCD 1] study by Moller et al. (1998) in The Lancet, in which they demonstrated in a prospective fashion that there were in fact persistent adverse effects on neurocognitive function up to 3 months following surgery and that these effects were common. Since the publication of that landmark study, significant strides have been made in redefining the terminology describing cognitive dysfunction, identifying those patients most at risk, and establishing the underlying etiology of the condition, particularly with respect to the relative contributions of anesthesia and surgery. In 2018, the International Nomenclature Consensus Working Group proposed new nomenclature to standardize identification of and classify perioperative cognitive changes under the umbrella of perioperative neurocognitive disorders (PND) (Evered et al., 2018a). Since then, the new nomenclature has tried to describe post-surgical cognitive derangements within a unifying framework and has brought to light the need to standardize methodology in clinical studies and motivate such studies with hypotheses of PND pathogenesis. In this narrative review, we highlight the relevant literature regarding recent key developments in PND identification and management throughout the perioperative period. We provide an overview of the new nomenclature and its implications for interpreting risk factors identified by clinical association studies. We then describe current hypotheses for PND development, using data from clinical association studies and neurophysiologic data where appropriate. Finally, we offer broad clinical guidelines for mitigating PND in the perioperative period, highlighting the role of Brain Enhanced Recovery After Surgery (Brain-ERAS) protocols.
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BACKGROUND: Anesthesiologists are increasingly encountering sicker patients that require potentially life-saving surgical interventions, and assess risk using the American Society of Anesthesiology Physical Status (ASA PS) classification system. Here, we examined long-term mortality along with hospital length of stay (LoS) and discharge disposition for survivors in ASA PS 5 and 5E patients. METHODS: Adult surgeries were extracted from New York-Presbyterian Hospital/Weill Cornell Medical Center's Electronic Medical Record (EMR) for cases between January 1, 2013 and December 31, 2017; outcomes were collected from EMRs and the Social Security Death Index Master File. RESULTS: 194,947 cases were identified. Mortality correlated with increasing ASA PS; the same trend was observed within both emergent and non-emergent sub-populations. Two hundred seventy-six cases were identified as 5/5E. This patient population had a higher rate of mortality at 30 days than at 48 hours (25.9% vs. 13.4%, respectively, p < 0.01); there was no difference between survivor functions at 30 or 90 days (p = 0.63, p = 0.09, respectively). Survivors within the 5 or 5E subpopulations did not have significantly different LoSs. Further, survivors after 90 days typically had a disposition of hospice, long-term facilities, inpatient rehabilitation, or self-discharged. CONCLUSIONS: Mortality increases with increases in ASA PS classifications. There is no difference in outcomes for 5 vs 5E at 30- or 90-day postoperatively. Similarly, emergency status did not play a role in LoS. Most 5 or 5E patients are not discharged home but to another facility. These outcomes should be considered during the informed consent process in this high-risk surgical population.
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The phencyclidine-derivative ketamine [2-(2-chlorophenyl)-2-(methylamino)cyclohexan-1-one] was added to the World Health Organization's Model List of Essential Medicines in 1985 and is also on the Model List of Essential Medicines for Children due to its efficacy and safety as an intravenous anesthetic. In sub-anesthetic doses, ketamine is an effective analgesic for the treatment of acute pain (such as may occur in the perioperative setting). Additionally, ketamine may have efficacy in relieving some forms of chronic pain. In 2019, Janssen Pharmaceuticals received regulatory-approval in both the United States and Europe for use of the S-enantiomer of ketamine in adults living with treatment-resistant major depressive disorder. Pre-existing anxiety/depression and the severity of postoperative pain are risk factors for development of chronic postsurgical pain. An important question is whether short-term administration of ketamine can prevent the conversion of acute postsurgical pain to chronic postsurgical pain. Here, we have reviewed ketamine's effects on the biopsychological processes underlying pain perception and affective mood disorders, focusing on non-NMDA receptor-mediated effects, with an emphasis on results from human trials where available.
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BACKGROUND: The coronavirus-2019 (COVID-19) pandemic highlighted the unfortunate reality that many hospitals have insufficient intensive care unit (ICU) capacity to meet massive, unanticipated increases in demand. To drastically increase ICU capacity, NewYork-Presbyterian/Weill Cornell Medical Center modified its existing operating rooms and post-anaesthesia care units during the initial expansion phase to accommodate the surge of critically ill patients. METHODS: This retrospective chart review examined patient care in non-standard Expansion ICUs as compared to standard ICUs. We compared clinical data between the two settings to determine whether the expeditious development and deployment of critical care resources during an evolving medical crisis could provide appropriate care. RESULTS: Sixty-six patients were admitted to Expansion ICUs from March 1st to April 30th, 2020 and 343 were admitted to standard ICUs. Most patients were male (70%), White (30%), 45-64 years old (35%), non-smokers (73%), had hypertension (58%), and were hospitalized for a median of 40 days. For patients that died, there was no difference in treatment management, but the Expansion cohort had a higher median ICU length of stay (q = 0.037) and ventilatory length (q = 0.015). The cohorts had similar rates of discharge to home, but the Expansion ICU cohort had higher rates of discharge to a rehabilitation facility and overall lower mortality. CONCLUSIONS: We found no significantly worse outcomes for the Expansion ICU cohort compared to the standard ICU cohort at our institution during the COVID-19 pandemic, which demonstrates the feasibility of providing safe and effective care for patients in an Expansion ICU.
Assuntos
COVID-19 , Pandemias , Cuidados Críticos , Feminino , Humanos , Unidades de Terapia Intensiva , Masculino , Pessoa de Meia-Idade , Estudos RetrospectivosRESUMO
Clinical research involving human subjects and quality improvement (QI) projects share a common goal of seeking to improve human health, whether by directly changing the standard of care (research) or by improving the process(es) by which that care is delivered (QI). Whether a QI project requires informed consent (written or oral) is a function of the risk-benefit analysis of the study; such a determination should not be at the sole discretion of the investigators, but should come from an appropriately constituted review board with expertise in the ethics of biomedical research.
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Pesquisa Biomédica , Melhoria de Qualidade , Comitês de Ética em Pesquisa , Humanos , Consentimento Livre e Esclarecido , Sujeitos da PesquisaRESUMO
General anesthesia has been administered for over 150 years, and in that time, has become progressively safer. Improvements in outcomes have been driven by multiple advances, including the use of non-invasive monitors to assess cardiovascular and respiratory status. More recent advances have included the development and use of monitors to measure neurologic status by means of "processed" electroencephalography (pEEG), wherein the frontal EEG signal is analyzed by proprietary algorithms to produce a dimensionless number (scaled from 0 to 100), wherein low values are associated with deepening levels of sedation that progresses to loss of consciousness. Such monitors have been shown to enable anesthetic titration so as to expedite emergence and early recovery, and their use is advocated for the prevention of intraoperative awareness in the setting of administration of total intravenous anesthesia and neuromuscular blockade. Whether their use can minimize, or prevent, longer term adverse events is a matter of debate. In this narrative review of the most recent literature, we provide an assessment on the use of pEEG monitors in the prevention of a notable, and important, postoperative adverse outcome - delirium - in elderly patients. As we will discuss, the existing data do not support its routine use for the prevention of postoperative delirium in this, or any other, patient population.
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Pain is a prevalent biopsychosocial condition that poses a significant challenge to healthcare providers, contributes substantially to a disability, and is a major economic burden worldwide. An overreliance on opioid analgesics, which primarily target the µ-opioid receptor, has caused devastating morbidity and mortality in the form of misuse and overdose-related death. Thus, novel analgesic medications are needed that can effectively treat pain and provide an alternative to opioids. A variety of cellular ion channels contribute to nociception, the response of the sensory nervous system to a noxious stimulus that commonly leads to pain. Ion channels involved in nociception may provide a suitable target for pharmacologic modulation to achieve pain relief. This narrative review summarizes the evidence for two ion channels that merit consideration as targets for non-opioid pain medications: ryanodine receptors (RyRs), which are intracellular calcium channels, and hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which belong to the superfamily of voltage-gated K+ channels. The role of these channels in nociception and neuropathic pain is discussed and suitability as targets for novel analgesics and antihyperalgesics is considered.
Assuntos
Cálcio , Neuralgia , Analgésicos/farmacologia , Analgésicos/uso terapêutico , Humanos , Canais Disparados por Nucleotídeos Cíclicos Ativados por HiperpolarizaçãoAssuntos
Lesão Pulmonar Aguda/etiologia , Infecções por Coronavirus/complicações , Obesidade/complicações , Pneumonia Viral/complicações , Apneia Obstrutiva do Sono/complicações , Lesão Pulmonar Aguda/epidemiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , COVID-19 , Infecções por Coronavirus/epidemiologia , Cuidados Críticos/estatística & dados numéricos , Estado Terminal , Feminino , Humanos , Masculino , Síndrome Metabólica/complicações , Pessoa de Meia-Idade , Obesidade/epidemiologia , Pandemias , Pneumonia Viral/epidemiologia , Respiração Artificial , Testes de Função Respiratória , Fatores de Risco , Apneia Obstrutiva do Sono/diagnóstico , Apneia Obstrutiva do Sono/epidemiologia , Resultado do TratamentoRESUMO
Significant progress has been made in the 21st century towards a comprehensive understanding of the mechanisms of action of general anesthetics, coincident with progress in structural biology and molecular, cellular, and systems neuroscience. This review summarizes important new findings that include target identification through structural determination of anesthetic binding sites, details of receptors and ion channels involved in neurotransmission, and the critical roles of neuronal networks in anesthetic effects on memory and consciousness. These recent developments provide a comprehensive basis for conceptualizing pharmacological control of amnesia, unconsciousness, and immobility.
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Anestésicos Gerais/farmacologia , Anestesia Geral/métodos , Anestésicos Gerais/administração & dosagem , Anestésicos Gerais/efeitos adversos , Anestésicos Gerais/química , Animais , Estado de Consciência/efeitos dos fármacos , Humanos , Modelos Moleculares , Neurônios/efeitos dos fármacos , Receptores de GABA-A/química , Receptores de GABA-A/metabolismo , Sono/efeitos dos fármacos , Relação Estrutura-Atividade , Transmissão Sináptica/efeitos dos fármacosRESUMO
BACKGROUND AND PURPOSE: In models of neuropathic pain, inhibition of HCN1 is anti-hyperalgesic. 2,6-di-iso-propyl phenol (propofol) and its non-anesthetic congener, 2,6-di-tert-butyl phenol, inhibit HCN1 channels by stabilizing closed state(s). EXPERIMENTAL APPROACH: Using in vitro electrophysiology and kinetic modeling, we systematically explore the contribution of ligand architecture to alkylphenol-channel coupling. KEY RESULTS: When corrected for changes in hydrophobicity (and propensity for intra-membrane partitioning), the decrease in potency upon 1-position substitution (NCOâ¼OHâ¯>>â¯SHâ¯>>>â¯F) mirrors the ligands' H-bond acceptor (NCOâ¯>â¯OHâ¯>â¯SHâ¯>>>â¯F) but not donor profile (OHâ¯>â¯SHâ¯>>>â¯NCOâ¼F). H-bond elimination (OH to F) corresponds to a ΔΔG of â¼4.5 kCalâ¯mol-1 loss of potency with little or no disruption of efficacy. Substitution of compact alkyl groups (iso-propyl, tert-butyl) with shorter (ethyl, methyl) or more extended (sec-butyl) adducts disrupts both potency and efficacy. Ring saturation (with the obligate loss of both planarity and π electrons) primarily disrupts efficacy. CONCLUSIONS AND IMPLICATIONS: A hydrophobicity-independent decrement in potency at higher volumes suggests the alkylbenzene site has a volume of ≥800â¯Å3. Within this, a relatively static (with respect to ligand) H-bond donor contributes to initial binding with little involvement in generation of coupling energy. The influence of π electrons/ring planarity and alkyl adducts on efficacy reveals these aspects of the ligand present towards a face of the channel that undergoes structural changes during opening. The site's characteristics suggest it is "druggable"; introduction of other adducts on the ring may generate higher potency inverse agonists.
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Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Ativação do Canal Iônico/efeitos dos fármacos , Oócitos/metabolismo , Fenóis/farmacologia , Canais de Potássio/metabolismo , Sequência de Aminoácidos , Animais , Microscopia Crioeletrônica , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/química , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/genética , Camundongos , Modelos Moleculares , Oócitos/efeitos dos fármacos , Fenóis/química , Canais de Potássio/química , Canais de Potássio/genética , Conformação Proteica , Isoformas de Proteínas , Relação Estrutura-Atividade , Xenopus laevisRESUMO
The mechanistic basis of gliogenesis, which occurs late in human development, is poorly understood. Here we identify nuclear factor IA (NFIA) as a molecular switch inducing human glial competency. Transient expression of NFIA is sufficient to trigger glial competency of human pluripotent stem cell-derived neural stem cells within 5 days and to convert these cells into astrocytes in the presence of glial-promoting factors, as compared to 3-6 months using current protocols. NFIA-induced astrocytes promote synaptogenesis, exhibit neuroprotective properties, display calcium transients in response to appropriate stimuli and engraft in the adult mouse brain. Differentiation involves rapid but reversible chromatin remodeling, glial fibrillary acidic protein (GFAP) promoter demethylation and a striking lengthening of the G1 cell cycle phase. Genetic or pharmacological manipulation of G1 length partially mimics NFIA function. We used the approach to generate astrocytes with region-specific or reactive features. Our study defines key mechanisms of the gliogenic switch and enables the rapid production of human astrocytes for disease modeling and regenerative medicine.
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Diferenciação Celular/genética , Fatores de Transcrição NFI/genética , Neurogênese/genética , Células-Tronco Pluripotentes/metabolismo , Animais , Astrócitos/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Camundongos , Fatores de Transcrição NFI/metabolismo , Células-Tronco Neurais/metabolismo , Neuroglia/citologia , Neuroglia/metabolismo , Neurônios/metabolismo , Regiões Promotoras GenéticasRESUMO
BACKGROUND: As minimally invasive spine surgery evolves, spine surgeons increasingly rely on advanced intraoperative computed tomography (iCT). iCT provides rapid acquisition of high-resolution images, reduces radiation exposure, improves surgical accuracy, and decreases operative time. However, all iCT systems currently available pose a patient safety risk as their physical space requirements limit patient access in the event of an emergency, particularly when patients are in the prone position. After a near-cardiac arrest at our institution during posterior cervical spine surgery, it was apparent that the presence of the iCT complicated the ability to rapidly reposition the patient in order to provide appropriate resuscitation. METHODS: To ensure our ability to provide timely care during an emergency, we determined that a process which included all members of the operating room (OR) team was required. We held an initial planning meeting where a detailed plan-of-action was created, reviewed, and revised in response to feedback from all stakeholders. We then simulated a cardiac arrest to test our resuscitation plan with all members of the neurosurgery team. A mannequin was positioned prone on an OR table within the iCT, and a resuscitation plan was created. RESULTS: The team orchestrated the mock resuscitation, and the time of cardiac arrest in the prone position to supine repositioning required 110 seconds. The simulation was recorded for post-"code" performance review. Application of the protocol during an actual cardiac arrest was associated with successful restoration of spontaneous circulation and full recovery. CONCLUSIONS: The development and rehearsal of an emergency plan of action greatly facilitated the timely responsiveness of the neurosurgical OR team during a simulated cardiac arrest and was an effective way to identify and address key logistical issues regarding the use of an iCT system.