Properties of the iron-sulfur cluster electron transfer relay in an [FeFe]-hydrogenase that is tuned for H2 oxidation catalysis.

[FeFe]-hydrogenases catalyze the reversible oxidation of H2 from electrons and protons at an organometallic active site cofactor named the H-cluster. In addition to the H-cluster, most [FeFe]-hydrogenases possess accessory FeS cluster (F-cluster) relays that function in mediating electron transfer with catalysis. There is significant variation in the structural properties of F-cluster relays among the [FeFe]-hydrogenases; however, it is unknown how this variation relates to the electronic and thermodynamic properties, and thus the electron transfer properties, of enzymes. Clostridium pasteurianum [FeFe]-hydrogenase II (CpII) exhibits a large catalytic bias for H2 oxidation (compared to H2 production), making it a notable system for examining if F-cluster properties contribute to the overall function and efficiency of the enzyme. By applying a combination of multifrequency and potentiometric electron paramagnetic resonance, we resolved two electron paramagnetic resonance signals with distinct power- and temperature-dependent properties at g = 2.058 1.931 1.891 (F2.058) and g = 2.061 1.920 1.887 (F2.061), with assigned midpoint potentials of -140 ± 18 mV and -406 ± 12 mV versus normal hydrogen electrode, respectively. Spectral analysis revealed features consistent with spin-spin coupling between the two [4Fe-4S] F-clusters, and possible functional models are discussed that account for the contribution of coupling to the electron transfer landscape. The results signify the interplay of electronic coupling and free energy properties and parameters of the FeS clusters to the electron transfer mechanism through the relay and provide new insight as to how relays functionally complement the catalytic directionality of active sites to achieve highly efficient catalysis.

An uncharacteristically low-potential flavin governs the energy landscape of electron bifurcation.

Electron bifurcation, an energy-conserving process utilized extensively throughout all domains of life, represents an elegant means of generating high-energy products from substrates with less reducing potential. The coordinated coupling of exergonic and endergonic reactions has been shown to operate over an electrochemical potential of ∼1.3 V through the activity of a unique flavin cofactor in the enzyme NADH-dependent ferredoxin-NADP+ oxidoreductase I. The inferred energy landscape has features unprecedented in biochemistry and presents novel energetic challenges, the most intriguing being a large thermodynamically uphill step for the first electron transfer of the bifurcation reaction. However, ambiguities in the energy landscape at the bifurcating site deriving from overlapping flavin spectral signatures have impeded a comprehensive understanding of the specific mechanistic contributions afforded by thermodynamic and kinetic factors. Here, we elucidate an uncharacteristically low two-electron potential of the bifurcating flavin, resolving the energetic challenge of the first bifurcation event.

Docetaxel-Based Neoadjuvant Chemotherapy Followed by En Bloc Resection for Esophageal Adenocarcinoma: A 15-Year Retrospective Analysis from a Regional Upper Gastrointestinal Cancer Network.

BACKROUND: Real-world, long-term survival outcomes of neoadjuvant, docetaxel-based therapy for esophageal and junctional adenocarcinoma are lacking. This study describes the long-term survival outcomes of patients with esophageal and junctional adenocarcinoma treated with neoadjuvant docetaxel-based chemotherapy and en bloc transthoracic esophagectomy. METHODS: A retrospective cohort analysis of a prospectively maintained database from a regional upper gastrointestinal cancer network in Quebec, Canada, was performed. From January 2007 to December 2021, all patients with locally advanced (cT3 and/or N1) esophageal/Siewert I/II adenocarcinoma treated with neoadjuvant DCFx3 (Docetaxel/Cisplatin/5FU) or FLOTx4 (5FU/Leucovorin/Oxaliplatin/Docetaxel) and transthoracic en bloc esophagectomy were identified. Postoperative, pathological, and survival outcomes were compared. RESULTS: Overall, 236 of 420 patients met the inclusion criteria. Tumor location was esophageal/Siewert I/Siewert II (118/33/85), most were cT3-4 (93.6%) and cN+ (61.0%). DCF and FLOT were used in 127 of 236 (53.8%) and 109 of 236 (46.2%). All neoadjuvant cycles were completed in 87.3% with no difference between the regimens. Operative procedures included Ivor Lewis (81.8%), left thoraco-abdominal esophagectomy (10.6%) and McKeown (7.6%) with an R0 resection in 95.3% and pathological complete response in 9.7% (DCF 12.6%/FLOT 6.4%, p = 0.111). The median lymph node yield was 32 (range 4-79), and 60.6% were ypN+. Median follow-up was longer for the DCF group (74.8 months 95% confidence interval [CI] 4-173 vs. 37.8 months 95% CI 2-119, p <0.001. Overall survival was similar between the groups (FLOT 97.3 months, 78.6-115.8 vs. DCF 92.9, 9.2-106.5, p = 0.420). CONCLUSIONS: Neoadjuvant DCF and FLOT followed by transthoracic en bloc resection are both highly effective regimens for locally advanced esophageal adenocarcinoma with equivalent survival outcomes despite high disease load.

High Affinity Electrostatic Interactions Support the Formation of CdS Quantum Dot:Nitrogenase MoFe Protein Complexes.

Nitrogenase MoFe protein can be coupled with CdS nanocrystals (NCs) to enable photocatalytic N2 reduction. The nature of interactions that support complex formation is of paramount importance in intermolecular electron transfer that supports catalysis. In this work we have employed microscale thermophoresis to examine binding interactions between 3-mercaptopropionate capped CdS quantum dots (QDs) and MoFe protein over a range of QD diameters (3.4-4.3 nm). The results indicate that the interactions are largely electrostatic, with the strength of interactions similar to that observed for the physiological electron donor. In addition, the strength of interactions is sensitive to the QD diameter, and the binding interactions are significantly stronger for QDs with smaller diameters. The ability to quantitatively assess NC protein interactions in biohybrid systems supports strategies for understanding properties and reaction parameters that are important for obtaining optimal rates of catalysis in biohybrid systems.

Cryo-annealing of Photoreduced CdS Quantum Dot-Nitrogenase MoFe Protein Complexes Reveals the Kinetic Stability of the E4(2N2H) Intermediate.

A critical step in the mechanism of N2 reduction to 2NH3 catalyzed by the enzyme nitrogenase is the reaction of the four-electron/four-proton reduced intermediate state of the active-site FeMo-cofactor (E4(4H)). This state is a junction in the catalytic mechanism, either relaxing by the reaction of a metal bound Fe-hydride with a proton forming H2 or going forward with N2 binding coupled to the reductive elimination (re) of two Fe-hydrides as H2 to form the E4(2N2H) state. E4(2N2H) can relax to E4(4H) by the oxidative addition (oa) of H2 and release of N2 or can be further reduced in a series of catalytic steps to release 2NH3. If the H2 re/oa mechanism is correct, it requires that oa of H2 be associative with E4(2N2H). In this report, we have taken advantage of CdS quantum dots in complex with MoFe protein to achieve photodriven electron delivery in the frozen state, with cryo-annealing in the dark, to reveal details of the E-state species and to test the stability of E4(2N2H). Illumination of frozen CdS:MoFe protein complexes led to formation of a population of reduced intermediates. Electron paramagnetic resonance spectroscopy identified E-state signals including E2 and E4(2N2H), as well as signals suggesting the formation of E6 or E8. It is shown that in the frozen state when pN2 is much greater than pH2, the E4(2N2H) state is kinetically stable, with very limited forward or reverse reaction rates. These results establish that the oa of H2 to the E4(2N2H) state follows an associative reaction mechanism.

Postoperative Outcomes and Quality of Life After Left Thoracoabdominal Esophagogastrectomy: Contrasting Esophagogastrostomy with Esophagojejunostomy.

BACKGROUND: Following left thoracoabdominal (LTA) esophagogastrectomy, gastrointestinal continuity can be re-established via esophagogastrostomy or esophagojejunostomy. We explored how the method of reconstruction impacted postoperative outcomes and quality of life (QoL). METHODS: From January 2007 to January 2022, patients undergoing LTA were identified from a single center's prospectively maintained database. Following esophagogastrectomy or extended total gastrectomy, an esophagogastrostomy (GAS) or Roux-en-Y esophagojejunostomy (R-Y) was fashioned. Postoperative outcomes were compared according to the method of reconstruction. The Functional Assessment of Cancer Therapy-Esophagus (FACT-E) questionnaire compared QoL. RESULTS: Of the 147 LTA patients identified, 135 (92%) were included-97 GAS (72%) and 38 R-Y patients (28%). R-Y patients had more ypT3/4 lesions (97% vs. 61%, p ≤ 0.001) and a similar incidence of ypN+/M+ disease. Anastomotic leaks were more common among GAS patients (17% vs. 3%, p = 0.023), however grade 3/4 complications (26.6% vs. 19.4%, p = 0.498), reoperation, intensive care admission, hospital representation and readmission were similar. FACT-E data were available for 68/97 (70%) GAS patients and 22/38 (58%) R-Y patients, with scores for 80/21/24/18/23/24 patients at baseline/preoperatively/1 month/3-6 months/1-3 years/3+ years postoperatively, respectively. Comparing between the groups, the scores were similar at each timepoint. FACT-E improved between baseline and preoperatively (79, 34-124 vs. 102, 81-123, p = 0.027). Only at 3+ years were postoperative scores equivalent to preoperative values. GAS patients had more reflux and esophagitis >6 months postoperatively (54% vs. 13%, p = 0.048; 62% vs. 0%, p ≤ 0.001). CONCLUSION: While the type of reconstruction did not affect QoL, it did affect the postoperative course.

Low-temperature trapping of N2 reduction reaction intermediates in nitrogenase MoFe protein-CdS quantum dot complexes.

The biological reduction of N2 to ammonia requires the ATP-dependent, sequential delivery of electrons from the Fe protein to the MoFe protein of nitrogenase. It has been demonstrated that CdS nanocrystals can replace the Fe protein to deliver photoexcited electrons to the MoFe protein. Herein, light-activated electron delivery within the CdS:MoFe protein complex was achieved in the frozen state, revealing that all the electron paramagnetic resonance (EPR) active E-state intermediates in the catalytic cycle can be trapped and characterized by EPR spectroscopy. Prior to illumination, the CdS:MoFe protein complex EPR spectrum was composed of a S = 3/2 rhombic signal (g = 4.33, 3.63, and 2.01) consistent with the FeMo-cofactor in the resting state, E0. Illumination for sequential 1-h periods at 233 K under 1 atm of N2 led to a cumulative attenuation of E0 by 75%. This coincided with the appearance of S = 3/2 and S = 1/2 signals assigned to two-electron (E2) and four-electron (E4) reduced states of the FeMo-cofactor, together with additional S = 1/2 signals consistent with the formation of E6 and E8 states. Simulations of EPR spectra allowed quantification of the different E-state populations, along with mapping of these populations onto the Lowe-Thorneley kinetic scheme. The outcome of this work demonstrates that the photochemical delivery of electrons to the MoFe protein can be used to populate all of the EPR active E-state intermediates of the nitrogenase MoFe protein cycle.

Dissecting Electronic-Structural Transitions in the Nitrogenase MoFe Protein P-Cluster during Reduction.

The [8Fe-7S] P-cluster of nitrogenase MoFe protein mediates electron transfer from nitrogenase Fe protein during the catalytic production of ammonia. The P-cluster transitions between three oxidation states, PN, P+, P2+ of which PN↔P+ is critical to electron exchange in the nitrogenase complex during turnover. To dissect the steps in formation of P+ during electron transfer, photochemical reduction of MoFe protein at 231-263 K was used to trap formation of P+ intermediates for analysis by EPR. In complexes with CdS nanocrystals, illumination of MoFe protein led to reduction of the P-cluster P2+ that was coincident with formation of three distinct EPR signals: S = 1/2 axial and rhombic signals, and a high-spin S = 7/2 signal. Under dark annealing the axial and high-spin signal intensities declined, which coincided with an increase in the rhombic signal intensity. A fit of the time-dependent changes of the axial and high-spin signals to a reaction model demonstrates they are intermediates in the formation of the P-cluster P+ resting state and defines how spin-state transitions are coupled to changes in P-cluster oxidation state in MoFe protein during electron transfer.

Changes in Perioperative Platelet Lymphocyte Ratio Predict Survival in Oesophago-Gastric Adenocarcinoma.

BACKGROUND: Platelet to lymphocyte ratio (PLR) is associated with survival in oesophageal cancer. We explored whether PLR changes during different stages of treatment correlate with survival outcomes. METHODS: A retrospective single-centre study was performed. Consecutive patients who received neoadjuvant chemotherapy followed by surgery for oesophageal adenocarcinoma were identified. Changes in PLR were calculated during two time periods: the first spanning the neoadjuvant period (T1); the second the perioperative period (T2). Differences in PLR were calculated for clinicopathological variables during both T1 and T2 and for variables with regards to their association with median overall survival (OS). Variables found to be significant on univariate analysis were included in a multivariate Cox regression model. Using ROC analysis, optimal cut-offs for PLR changes were identified and plotted on a Kaplan-Meir curve. RESULTS: Of the 370 patients identified, 110 (29.7%) were included in the analysis. During T1 a positive correlation was noted between higher positive lymph node ratio and PLR change. During T2, PLR change was positively higher in patients who suffered major postoperative complications. Median survival for the cohort as a whole was 42.3 months and 5-year OS was 57.3%. Survival at 5 years was associated with lower PLR changes during T2. On univaraite analysis, median OS was significantly less for patients with a tumour size > 3 cm, poor differentiation and change in PLR ≥ 43.4 during T2. The latter two variables remained significant on multivariate analysis. Using the same PLR threshold, the survival curve comparing changes in PLR during T2 remained statistically significant. CONCLUSION: Perioperative PLR changes are highly prognostic of survival outcomes in patients treated for oesophageal adenocarcinoma.

The structure and reactivity of the HoxEFU complex from the cyanobacterium Synechocystis sp. PCC 6803.

Cyanobacterial Hox is a [NiFe] hydrogenase that consists of the hydrogen (H2)-activating subunits HoxYH, which form a complex with the HoxEFU assembly to mediate reactions with soluble electron carriers like NAD(P)H and ferredoxin (Fdx), thereby coupling photosynthetic electron transfer to energy-transforming catalytic reactions. Researchers studying the HoxEFUYH complex have observed that HoxEFU can be isolated independently of HoxYH, leading to the hypothesis that HoxEFU is a distinct functional subcomplex rather than an artifact of Hox complex isolation. Moreover, outstanding questions about the reactivity of Hox with natural substrates and the site(s) of substrate interactions and coupling of H2, NAD(P)H, and Fdx remain to be resolved. To address these questions, here we analyzed recombinantly produced HoxEFU by electron paramagnetic resonance spectroscopy and kinetic assays with natural substrates. The purified HoxEFU subcomplex catalyzed electron transfer reactions among NAD(P)H, flavodoxin, and several ferredoxins, thus functioning in vitro as a shuttle among different cyanobacterial pools of reducing equivalents. Both Fdx1-dependent reductions of NAD+ and NADP+ were cooperative. HoxEFU also catalyzed the flavodoxin-dependent reduction of NAD(P)+, Fdx2-dependent oxidation of NADH and Fdx4- and Fdx11-dependent reduction of NAD+ MS-based mapping identified an Fdx1-binding site at the junction of HoxE and HoxF, adjacent to iron-sulfur (FeS) clusters in both subunits. Overall, the reactivity of HoxEFU observed here suggests that it functions in managing peripheral electron flow from photosynthetic electron transfer, findings that reveal detailed insights into how ubiquitous cellular components may be used to allocate energy flow into specific bioenergetic products.

Transition from open to minimally invasive en bloc esophagectomy can be achieved without compromising surgical quality.

BACKGROUND: En bloc esophagectomy results in higher lymph node (LN) retrieval than standard esophagectomy. Minimally invasive esophagectomy (MIE) has gained traction due to improved short-term outcomes, but many large series report LN yields well below the international benchmark of 23. We sought to determine if an established approach to open en bloc resection can be safely transferred to MIE using LN yield as a quality benchmark. METHODS: An open approach to en bloc esophagectomy (OE) was established over 5 years (~ 300 cases) before en bloc MIE was introduced in 2010. Patients undergoing curative-intent en bloc Ivor-Lewis and McKeown esophagectomy for cancer from 2010 to 2019 by a single surgeon with formal minimally invasive surgery training were identified from a prospectively collected database. Mann-Whitney U and χ2 tests and cumulative sum analysis were used for statistical analysis. "Failure" was defined as LN yield less than AJCC's 8th edition guidelines: 10 LNs for pT1 cancers, 20 for pT2 and 30 for pT3-4. RESULTS: A total of 269 esophageal resections met inclusion criteria [193(72%) OE; 76(28%) MIE]. Age, sex, BMI and comorbidities were comparable between groups. Tumors were larger and more often locally advanced in OE. Median LN retrieval was sufficient by international standards in both groups [OE:34(27-46); MIE:28(22-39); p = 0.01]. "Failures" occurred in 33(17%) of OE and 12(16%) MIE cases (p = 0.63). No learning effect was observed for LN yield. R0 resection rate was comparable [OE:191(99%); MIE:73(96%); p = 0.90]. Operative time was longer for MIE [275(246-300)] than OE [240(210-270) minutes], p < 0.0001, while estimated blood loss (OE:350(250-500)mL; MIE:300(200-400)mL; p = 0.02] and length of stay [OE:8(6-13); MIE7(6-9) days; p = 0.02] were higher for OE. Morbidity and mortality were comparable between groups and LN yield did not impact survival. CONCLUSIONS: Under appropriate conditions, an established approach to open en bloc esophagectomy can be safely transferred to MIE without compromising surgical quality.

The catalytic mechanism of electron-bifurcating electron transfer flavoproteins (ETFs) involves an intermediary complex with NAD.

Electron bifurcation plays a key role in anaerobic energy metabolism, but it is a relatively new discovery, and only limited mechanistic information is available on the diverse enzymes that employ it. Herein, we focused on the bifurcating electron transfer flavoprotein (ETF) from the hyperthermophilic archaeon Pyrobaculum aerophilum The EtfABCX enzyme complex couples NADH oxidation to the endergonic reduction of ferredoxin and exergonic reduction of menaquinone. We developed a model for the enzyme structure by using nondenaturing MS, cross-linking, and homology modeling in which EtfA, -B, and -C each contained FAD, whereas EtfX contained two [4Fe-4S] clusters. On the basis of analyses using transient absorption, EPR, and optical titrations with NADH or inorganic reductants with and without NAD+, we propose a catalytic cycle involving formation of an intermediary NAD+-bound complex. A charge transfer signal revealed an intriguing interplay of flavin semiquinones and a protein conformational change that gated electron transfer between the low- and high-potential pathways. We found that despite a common bifurcating flavin site, the proposed EtfABCX catalytic cycle is distinct from that of the genetically unrelated bifurcating NADH-dependent ferredoxin NADP+ oxidoreductase (NfnI). The two enzymes particularly differed in the role of NAD+, the resting and bifurcating-ready states of the enzymes, how electron flow is gated, and the two two-electron cycles constituting the overall four-electron reaction. We conclude that P. aerophilum EtfABCX provides a model catalytic mechanism that builds on and extends previous studies of related bifurcating ETFs and can be applied to the large bifurcating ETF family.

Defining Intermediates of Nitrogenase MoFe Protein during N2 Reduction under Photochemical Electron Delivery from CdS Quantum Dots.

Coupling the nitrogenase MoFe protein to light-harvesting semiconductor nanomaterials replaces the natural electron transfer complex of Fe protein and ATP and provides low-potential photoexcited electrons for photocatalytic N2 reduction. A central question is how direct photochemical electron delivery from nanocrystals to MoFe protein is able to support the multielectron ammonia production reaction. In this study, low photon flux conditions were used to identify the initial reaction intermediates of CdS quantum dot (QD):MoFe protein nitrogenase complexes under photochemical activation using EPR. Illumination of CdS QD:MoFe protein complexes led to redox changes in the MoFe protein active site FeMo-co observed as the gradual decline in the E0 resting state intensity that was accompanied by an increase in the intensity of a new "geff = 4.5" EPR signal. The magnetic properties of the geff = 4.5 signal support assignment as a reduced S = 3/2 state, and reaction modeling was used to define it as a two-electron-reduced "E2" intermediate. Use of a MoFe protein variant, ß-188Cys, which poises the P cluster in the oxidized P+ state, demonstrated that the P cluster can function as a site of photoexcited electron delivery from CdS to MoFe protein. Overall, the results establish the initial steps for how photoexcited CdS delivers electrons into the MoFe protein during reduction of N2 to ammonia and the role of electron flux in the photochemical reaction cycle.

Tuning Catalytic Bias of Hydrogen Gas Producing Hydrogenases.

Hydrogenases display a wide range of catalytic rates and biases in reversible hydrogen gas oxidation catalysis. The interactions of the iron-sulfur-containing catalytic site with the local protein environment are thought to contribute to differences in catalytic reactivity, but this has not been demonstrated. The microbe Clostridium pasteurianum produces three [FeFe]-hydrogenases that differ in "catalytic bias" by exerting a disproportionate rate acceleration in one direction or the other that spans a remarkable 6 orders of magnitude. The combination of high-resolution structural work, biochemical analyses, and computational modeling indicates that protein secondary interactions directly influence the relative stabilization/destabilization of different oxidation states of the active site metal cluster. This selective stabilization or destabilization of oxidation states can preferentially promote hydrogen oxidation or proton reduction and represents a simple yet elegant model by which a protein catalytic site can confer catalytic bias.

Feasibility analysis for the development of a video-assisted thoracoscopic (VATS) lobectomy 23-hour recovery pathway.

Background: Video-assisted thoracoscopic (VATS) lobectomy has been demonstrated to offer several benefits over open surgery. The purpose of this study was to assess the feasibility and safety of an ultra-fast-track 23-hour recovery pathway for VATS lobectomy. Methods: A prospectively maintained institutional database was queried for patients who underwent VATS lobectomy from 2006 to 2016 at the McGill University Health Centre in Montreal, Quebec, and data were supplemented with focused chart review. Patients discharged with a length of stay (LOS) of 23 hours or less were compared with those with an LOS of 2 days or more. Logistic regression was performed to identify predictors of LOS of 23 hours or less. Results: Two hundred and five patients were included in the study. Perioperative 30-day mortality for our cohort was 0% and the major complication rate was 8.3%. The median LOS was 3 days (interquartile range [IQR] 2-4 d). Thirty-four patients were discharged within 23 hours and none of them required readmission; 171 patients were discharged on postoperative day 2 or later and 9 of them (5.3%) required readmission (p = 0.36). The proportion of patients discharged within 23 hours increased in 2016 compared with previous years (25.8% v. 12.0%, p = 0.05). Patients discharged within 23 hours had shorter chest tube duration (odds ratio [OR] 0.20, 95% confidence interval [CI] 0.09-0.46, p < 0.001), lower clinical stage disease (stages II-III v. stage I OR 0.07, 95% CI 0.01-0.52, p = 0.011), lower pathologic stage lesions (stages II-III v. stage I OR 0.26, 95% CI 0.07-0.91, p = 0.035), fewer surgical complications (OR 0.04, 95% CI 0.01-0.30, p = 0.002) and shorter operative time (surgery duration > 120 min OR 0.42, 95% CI 0.18-0.95, p = 0.04). Our exploratory prediction modelling showed that chest tube duration, clinical stage and surgeon were the most influential predictors of discharge within 23 hours. Conclusion: The only preoperative factors that predicted shorter LOS in our cohort were clinical stage and surgeon. A significant proportion of patients can be discharged safely by adopting a VATS lobectomy 23-hour enhanced recovery pathway.

Contexte: Il a été démontré que la lobectomie par chirurgie thoracique vidéoassistée (CTVA) offre plusieurs avantages comparativement à la chirurgie ouverte. La présente étude avait pour but d'évaluer la faisabilité et la sûreté d'un protocole de récupération ultrarapide en 23 heures pour la lobectomie par CTVA. Méthodes: Nous avons extrait d'une base de données d'établissement maintenue de manière prospective des données sur les patients ayant subi une lobectomie par CTVA entre 2006 et 2016 au Centre universitaire de santé McGill à Montréal (Québec), complétées par un examen ciblé des dossiers. Les patients ayant reçu leur congé après une hospitalisation de 23 heures ou moins ont été comparés à ceux dont l'hospitalisation avait duré 2 jours ou plus. Nous avons ensuite mis en évidence les facteurs prédictifs d'une hospitalisation de 23 heures ou moins par une analyse de régression logistique. Résultats: Deux cent cinq patients ont été inclus dans l'étude. La mortalité périopératoire dans les 30 jours suivant l'intervention était de 0 % dans notre cohorte, et le taux de complications majeures était de 8,3 %. La durée d'hospitalisation médiane était de 3 jours (écart interquartile [EI] 2 à 4 jours). Trente-quatre patients ont reçu leur congé dans les 23 heures suivant l'intervention, et aucun n'a dû être réhospitalisé; comparativement, 171 patients ont reçu leur congé au deuxième jour ou après, et 9 d'entre eux (5,3 %) ont dû être réhospitalisés (p = 0,36). Le pourcentage de patients ayant reçu leur congé dans les 23 heures a augmenté en 2016 par rapport aux années précédentes (25,8 % c. 12,0 %, p = 0,05). Les patients au congé dans les 23 heures conservaient leur drain thoracique moins longtemps (rapport de cotes [RC] 0,20, intervalle de confiance [IC] de 95 % 0,09 à 0,46, p < 0,001); leur stade clinique était moins élevé (stades II à III c. stade I ­ RC 0,07, IC de 95 % 0,01 à 0,52, p = 0,011); le stade pathologique de leurs lésions était plus faible (stades II à III c. stade I ­ RC 0,26, IC de 95 % 0,07 à 0,91, p = 0,035); ils avaient moins de complications chirurgicales (RC 0,04, IC de 95 % 0,01 à 0,30, p = 0,002); et la durée de leur intervention était plus courte (durée de la chirurgie > 120 minutes ­ RC 0,42, IC de 95 % 0,18 à 0,95, p = 0,04). Notre modèle prédictif exploratoire a montré que le délai avant le retrait du drain thoracique, le stade clinique et le chirurgien était les facteurs prédictifs les plus importants du congé dans les 23 heures. Conclusion: Les seuls facteurs préopératoires permettant de prédire une hospitalisation plus courte dans notre cohorte étaient le stade clinique et le chirurgien. Un pourcentage important des patients peuvent recevoir leur congé sans danger si on suit un protocole de récupération optimisée en 23 heures après une lobectomie par CTVA.

Mechanistic insights into energy conservation by flavin-based electron bifurcation.

The recently realized biochemical phenomenon of energy conservation through electron bifurcation provides biology with an elegant means to maximize utilization of metabolic energy. The mechanism of coordinated coupling of exergonic and endergonic oxidation-reduction reactions by a single enzyme complex has been elucidated through optical and paramagnetic spectroscopic studies revealing unprecedented features. Pairs of electrons are bifurcated over more than 1 volt of electrochemical potential by generating a low-potential, highly energetic, unstable flavin semiquinone and directing electron flow to an iron-sulfur cluster with a highly negative potential to overcome the barrier of the endergonic half reaction. The unprecedented range of thermodynamic driving force that is generated by flavin-based electron bifurcation accounts for unique chemical reactions that are catalyzed by these enzymes.

CO-Bridged H-Cluster Intermediates in the Catalytic Mechanism of [FeFe]-Hydrogenase CaI.

The [FeFe]-hydrogenases ([FeFe] H2ases) catalyze reversible H2 activation at the H-cluster, which is composed of a [4Fe-4S]H subsite linked by a cysteine thiolate to a bridged, organometallic [2Fe-2S] ([2Fe]H) subsite. Profoundly different geometric models of the H-cluster redox states that orchestrate the electron/proton transfer steps of H2 bond activation have been proposed. We have examined this question in the [FeFe] H2ase I from Clostridium acetobutylicum (CaI) by Fourier-transform infrared (FTIR) spectroscopy with temperature annealing and H/D isotope exchange to identify the relevant redox states and define catalytic transitions. One-electron reduction of Hox led to formation of HredH+ ([4Fe-4S]H2+-FeI-FeI) and Hred' ([4Fe-4S]H1+-FeII-FeI), with both states characterized by low frequency µ-CO IR modes consistent with a fully bridged [2Fe]H. Similar µ-CO IR modes were also identified for HredH+ of the [FeFe] H2ase from Chlamydomonas reinhardtii (CrHydA1). The CaI proton-transfer variant C298S showed enrichment of an H/D isotope-sensitive µ-CO mode, a component of the hydride bound H-cluster IR signal, Hhyd. Equilibrating CaI with increasing amounts of NaDT, and probed at cryogenic temperatures, showed HredH+ was converted to Hhyd. Over an increasing temperature range from 10 to 260 K catalytic turnover led to loss of Hhyd and appearance of Hox, consistent with enzymatic turnover and H2 formation. The results show for CaI that the µ-CO of [2Fe]H remains bridging for all of the "Hred" states and that HredH+ is on pathway to Hhyd and H2 evolution in the catalytic mechanism. These results provide a blueprint for designing small molecule catalytic analogs.

Electron Bifurcation: Thermodynamics and Kinetics of Two-Electron Brokering in Biological Redox Chemistry.

How can proteins drive two electrons from a redox active donor onto two acceptors at very different potentials and distances? And how can this transaction be conducted without dissipating very much energy or violating the laws of thermodynamics? Nature appears to have addressed these challenges by coupling thermodynamically uphill and downhill electron transfer reactions, using two-electron donor cofactors that have very different potentials for the removal of the first and second electron. Although electron bifurcation is carried out with near perfection from the standpoint of energy conservation and electron delivery yields, it is a biological energy transduction paradigm that has only come into focus recently. This Account provides an exegesis of the biophysical principles that underpin electron bifurcation. Remarkably, bifurcating electron transfer (ET) proteins typically send one electron uphill and one electron downhill by similar energies, such that the overall reaction is spontaneous, but not profligate. Electron bifurcation in the NADH-dependent reduced ferredoxin: NADP+ oxidoreductase I (Nfn) is explored in detail here. Recent experimental progress in understanding the structure and function of Nfn allows us to dissect its workings in the framework of modern ET theory. The first electron that leaves the two-electron donor flavin (L-FAD) executes a positive free energy "uphill" reaction, and the departure of this electron switches on a second thermodynamically spontaneous ET reaction from the flavin along a second pathway that moves electrons in the opposite direction and at a very different potential. The singly reduced ET products formed from the bifurcating flavin are more than two nanometers distant from each other. In Nfn, the second electron to leave the flavin is much more reducing than the first: the potentials are said to be "crossed." The eventually reduced cofactors, NADH and ferredoxin in the case of Nfn, perform crucial downstream redox processes of their own. We dissect the thermodynamics and kinetics of electron bifurcation in Nfn and find that the key features of electron bifurcation are (1) spatially separated transfer pathways that diverge from a two-electron donor, (2) one thermodynamically uphill and one downhill redox pathway, with a large negative shift in the donor's reduction potential after departure of the first electron, and (3) electron tunneling and activation factors that enable bifurcation, producing a 1:1 partitioning of electrons onto the two pathways. Electron bifurcation is found in the CO2 reducing pathways of methanogenic archaea, in the hydrogen pathways of hydrogenases, in the nitrogen fixing pathway of Fix, and in the mitochondrial charge transfer chain of complex III, cytochrome bc1. While crossed potentials may offer the biological advantage of producing tightly regulated high energy reactive species, neither kinetic nor thermodynamic considerations mandate crossed potentials to generate successful electron bifurcation. Taken together, the theoretical framework established here, focusing on the underpinning electron tunneling barriers and activation free energies, explains the logic of electron bifurcation that enables energy conversion and conservation in Nfn, points toward bioinspired schemes to execute multielectron redox chemistry, and establishes a roadmap for examining novel electron bifurcation networks in nature.

Same day discharge for benign laparoscopic hiatal surgery: a feasibility analysis.

BACKGROUND: Advances in minimally invasive surgery and the development of enhanced recovery pathways have favored the spread of day-surgery programs. Despite laparoscopic approaches being accepted as the standard of care for benign hiatal diseases, safety and feasibility of same day discharges for laparoscopic hiatal surgeries other than fundoplication has yet to be established. OBJECTIVE: This study aimed to assess the feasibility of same day discharge for primary and revisional laparoscopic hiatal surgeries including paraesophageal hernia repairs (PEHR), fundoplication for reflux, and Heller myotomy (±diverticulectomy). METHODS: A retrospective cohort study including all patients undergoing elective laparoscopic hiatal procedures in the division of Thoracic surgery between 2011 and 2016 at McGill University Health Centre was performed. Planned day-surgery (DAYCASE) was compared to planned inpatient (INPATIENT) cohorts with respect to operative and postoperative outcomes, length of stay, readmission, and emergency room visits. RESULTS: A total of 261 patients were identified, 161 female (62%); median age 62 (20). The case distribution was: PEHR (123; 47.1%), Heller myotomy (94;36%, 7 diverticulectomy), and fundoplication (44; 16.9%). Twenty patients had revisional procedures (7.7%). Same day discharge was planned in 98 cases (38%) and was successful in 80 (81.6%). Proportion of DAYCASE increased form 12% prior to 2013 to 67% in 2016. INPATIENTs were older (median 66 vs. 60 years), and had a higher proportion of PEHR (55 vs. 34%), p < 0.05. Both cohorts were comparable in gender proportion, ASA classification, and length of surgery. Complications, readmission, and emergency visits did not differ between the two cohorts. On multivariate analysis, female gender (OR 37, 95% CI 1.46-936, p = 0.028), surgery beginning after noon (OR 5.4, 95% CI 1.1-26.9, p = 0.038), intraoperative complications (OR 20.4 95% CI 1.5-286, p = 0.025), and postoperative complications (OR 52.1, 95% CI 4.5-602, p = 0.002) were independently associated with unplanned admission. CONCLUSIONS: Day-case surgery for complex laparoscopic hiatal procedures is feasible and can be achieved in a significant number of patients without compromising safety.