Fighting the obesity pandemic during the COVID-19 pandemic.

BACKGROUND: The COVID-19 pandemic created delays in surgical care. The population with obesity has a high risk of death from COVID-19. Prior literature shows the most effective way to combat obesity is by weight loss surgery. At different times throughout the COVID-19 pandemic, elective inpatient surgeries have been halted due to bed availability. Recognizing that major complications following bariatric surgery are extremely low (bleeding 0-4%, anastomotic leaks 0.8%), we felt outpatient bariatric surgery would be safe for low-risk patients. Complications such as DVT, PE, infection, and anastomotic leaks typically present after 7 days postoperatively, well outside the usual length of stay. Bleeding events, severe postoperative nausea, and dehydration typically occur in the first few days postoperatively. We designed a pathway focused on detecting and preventing these early post-op complications to allow safe outpatient bariatric surgery. METHODS: We used a preoperative evaluation tool to risk stratify bariatric patients. During a 16-month period, 89 patients were identified as low risk for outpatient surgery. We designed a postoperative protocol that included IV hydration and PO intake goals to meet a safe discharge. We sent patients home with a pulse oximeter and had them self-monitor their pulse and oxygen saturation. We called all patients at 10 pm for a postoperative assessment and report of their vitals. Patients returned to clinic the following day and were seen by a provider, received IV hydration, and labs were drawn. RESULTS: 80 of 89 patients (89.8%) were successfully discharged on POD 0. 3 patients were readmitted within 30 days. We had zero deaths in our study cohort and no morbidity that would have been prevented with postoperative admission. CONCLUSION: We demonstrate that by identifying low-risk patients for outpatient bariatric surgery and by implementing remote monitoring of vitals early outpatient follow-up, we were able to safely perform outpatient bariatric surgery.

Are you better off than you were 4 years ago? Measuring the impact of the ABS flexible endoscopy curriculum.

INTRODUCTION: In 2014, the ABS introduced the Flexible Endoscopy Curriculum (FEC). The FEC did not alter the minimum defined category case volumes for endoscopy; however, it did introduce specific cognitive and technical milestones for endoscopy training. It also mandated that residents pass the Fundamentals of Endoscopic Skills (FES) exam to qualify for board certification. Although significant research has been published regarding residents' success on the FES exam, very little is known regarding how the FEC has changed the way general surgery programs train their residents in surgical endoscopy. The aim of this study was to quantify changes in flexible endoscopy education at a large academic program in the 4 years since the FEC was published. METHODS: We classified the impact of FEC into four categories: (a) case volume or distribution, (b) clinical rotations, (c) required didactics or simulation exercises, and (d) FES pass rates. For category (a), we reviewed current and historical case logs for all categorical residents from 2013 to 2018. Mann-Whitney U tests were used to compare endoscopy volumes for each PGY level in 2013-2014 to the respective PGY level in 2017-2018 with p < 0.05 considered significant. For categories (b)-(d), we gathered historical records from the residency coordinator and endoscopy rotation director. RESULTS: Complete data were available for 57 residents in the 2013-2014 academic year and 56 residents in the 2017-2018 academic year. Median total endoscopies performed by PGY2, PGY3, and PGY5 residents all significantly increased during the FEC rollout. Our program's focus on endoscopy also expanded with absolute increases in endoscopy rotations, didactics, and simulation exercises. These changes translated into significantly increased pass rates on the FES exam from 40 to 100%. CONCLUSIONS: Implementation of the FEC at a large academic program led to measurable improvements in clinical experience, program structure, educational programing, and performance on high-stakes assessments.

Dynamic regulation of ß1 subunit trafficking controls vascular contractility.

Ion channels composed of pore-forming and auxiliary subunits control physiological functions in virtually all cell types. A conventional view is that channels assemble with their auxiliary subunits before anterograde plasma membrane trafficking of the protein complex. Whether the multisubunit composition of surface channels is fixed following protein synthesis or flexible and open to acute and, potentially, rapid modulation to control activity and cellular excitability is unclear. Arterial smooth muscle cells (myocytes) express large-conductance Ca(2+)-activated potassium (BK) channel α and auxiliary ß1 subunits that are functionally significant modulators of arterial contractility. Here, we show that native BKα subunits are primarily (∼95%) plasma membrane-localized in human and rat arterial myocytes. In contrast, only a small fraction (∼10%) of total ß1 subunits are located at the cell surface. Immunofluorescence resonance energy transfer microscopy demonstrated that intracellular ß1 subunits are stored within Rab11A-postive recycling endosomes. Nitric oxide (NO), acting via cGMP-dependent protein kinase, and cAMP-dependent pathways stimulated rapid (≤1 min) anterograde trafficking of ß1 subunit-containing recycling endosomes, which increased surface ß1 almost threefold. These ß1 subunits associated with surface-resident BKα proteins, elevating channel Ca(2+) sensitivity and activity. Our data also show that rapid ß1 subunit anterograde trafficking is the primary mechanism by which NO activates myocyte BK channels and induces vasodilation. In summary, we show that rapid ß1 subunit surface trafficking controls functional BK channel activity in arterial myocytes and vascular contractility. Conceivably, regulated auxiliary subunit trafficking may control ion channel activity in a wide variety of cell types.

Weak interactions between folate and osmolytes in solution.

Previous osmotic stress studies on the role of solvent in two structurally unrelated dihydrofolate reductases (DHFRs) found weaker binding of dihydrofolate (DHF) to either enzyme in the presence of osmolytes. To explain these unusual results, weak interactions between DHF and osmolytes were proposed, with a competition between osmolyte and DHFR for DHF. High osmolyte concentrations will inhibit binding of the cognate pair. To evaluate this hypothesis, we devised a small molecule approach. Dimerization of folate, monitored by nuclear magnetic resonance, was weakened 2-3-fold upon addition of betaine or dimethyl sulfoxide (DMSO), supporting preferential interaction of either osmolyte with the monomer (as it possesses a larger surface area). Nuclear Overhauser effect (NOE) spectroscopy experiments found a positive NOE for the interaction of the C3'/C5' benzoyl ring protons with the C9 proton in buffer; however, a negative NOE was observed upon addition of betaine or DMSO. This change indicated a decreased tumbling rate, consistent with osmolyte interaction. Osmotic stress experiments also showed that betaine, DMSO, and sucrose preferentially interact with folate. Further, studies with the folate fragments, p-aminobenzoic acid and pterin 6-carboxylate, revealed interactions for both model compounds with betaine and sucrose. In contrast, DMSO was strongly excluded from the pterin ring but preferentially interacted with the p-aminobenzoyl moiety. These interactions are likely to be important in vivo because of the crowded conditions of the cell where weak contacts can more readily compete with specific binding interactions.

Thermodynamics and solvent effects on substrate and cofactor binding in Escherichia coli chromosomal dihydrofolate reductase.

Chromosomal dihydrofolate reductase from Escherichia coli catalyzes the reduction of dihydrofolate to tetrahydrofolate using NADPH as a cofactor. The thermodynamics of ligand binding were examined using an isothermal titration calorimetry approach. Using buffers with different heats of ionization, zero to a small, fractional proton release was observed for dihydrofolate binding, while a proton was released upon NADP(+) binding. The role of water in binding was additionally monitored using a number of different osmolytes. Binding of NADP(+) is accompanied by the net release of ∼5-24 water molecules, with a dependence on the identity of the osmolyte. In contrast, binding of dihydrofolate is weakened in the presence of osmolytes, consistent with "water uptake". Different effects are observed depending on the identity of the osmolyte. The net uptake of water upon dihydrofolate binding was previously observed in the nonhomologous R67-encoded dihydrofolate reductase (dfrB or type II enzyme) [Chopra, S., et al. (2008) J. Biol. Chem. 283, 4690-4698]. As R67 dihydrofolate reductase possesses a nonhomologous sequence and forms a tetrameric structure with a single active site pore, the observation of weaker DHF binding in the presence of osmolytes in both enzymes implicates cosolvent effects on free dihydrofolate. Consistent with this analysis, stopped flow experiments find betaine mostly affects DHF binding via changes in k(on), while betaine mostly affects NADPH binding via changes in k(off). Finally, nonadditive enthalpy terms when binary and ternary cofactor binding events are compared suggest the presence of long-lived conformational transitions that are not included in a simple thermodynamic cycle.

Radical redesign of a tandem array of four R67 dihydrofolate reductase genes yields a functional, folded protein possessing 45 substitutions.

R67 dihydrofolate reductase (DHFR) is a plasmid-encoded, type II enzyme. Four monomers (78 amino acids long) assemble into a homotetramer possessing 222 symmetry. In previous studies, a tandem array of four R67 DHFR gene copies was fused in frame to generate a functional monomer named Quad1. This protein possessed the essential tertiary structure of the R67 "parent". To facilitate mutagenesis reactions, restriction enzyme sites were introduced in the tandem gene array. S59A and H362L mutations were also added to minimize possible folding topologies; this protein product, named Quad3, possesses 10 substitutions and is functional. Since R67 DHFR possesses a stable scaffold, a large jump in sequence space was performed by the further addition of 45 amino acid substitutions. The mutational design utilized alternate sequences from other type II DHFRs. In addition, most of the mutations were positioned on the surface of the protein as well as in the disordered N-terminal sequence, which serves as the linker between the fused domains. The resulting Quad4 protein is quite functional; however, it is less stable than Quad1, suffering a DeltaDeltaG loss of 5 kcal/mol at pH 5. One unexpected result was formation of Quad4 dimers and higher order oligomers at pH 8. R67 DHFR, and its derivative Quad proteins, possesses a robust scaffold, capable of withstanding introduction of >or=55 substitutions.

Perforated jejunal diverticulitis in a nonagenarian veteran: A case report.

INTRODUCTION: Jejunal diverticular (JD) disease is an uncommon occurrence that frequently present as a diagnostic dilemma. The correct diagnosis from complications of JD is typically made at laparotomy. Most JD are asymptomatic. Of clinically significant small bowel diverticula, only 15% present with surgical problems including obstruction, gastrointestinal bleed, and perforation. PRESENTATION OF CASE: A 90-year-old man presented to the hospital with abdominal pain. He was clinically stable with local tenderness in the left lower quadrant. Computed tomography demonstrated a dot of free air near the sigmoid and sigmoid diverticula. He continued to have pain and clinically deteriorated following a short period of observation. He underwent an exploratory laparotomy that revealed perforated JD. CONCLUSION: Perforation from JD is exceedingly rare. Due to their infrequent clinical significance, complications from JD are difficult to diagnose and therapeutic options are typically made intraoperatively. Any deviation from the expected positive pathway in the management of a suspected entity should prompt an immediate reassessment as well as definitive therapeutic options.

Isothermal titration calorimetry for measuring macromolecule-ligand affinity.

Isothermal titration calorimetry (ITC) is a useful tool for understanding the complete thermodynamic picture of a binding reaction. In biological sciences, macromolecular interactions are essential in understanding the machinery of the cell. Experimental conditions, such as buffer and temperature, can be tailored to the particular binding system being studied. However, careful planning is needed since certain ligand and macromolecule concentration ranges are necessary to obtain useful data. Concentrations of the macromolecule and ligand need to be accurately determined for reliable results. Care also needs to be taken when preparing the samples as impurities can significantly affect the experiment. When ITC experiments, along with controls, are performed properly, useful binding information, such as the stoichiometry, affinity and enthalpy, are obtained. By running additional experiments under different buffer or temperature conditions, more detailed information can be obtained about the system. A protocol for the basic setup of an ITC experiment is given.