Primary Pancreatic Neuroblastoma in an Infant.

A 2-month-old girl with conjugated hyperbilirubinemia was found at the surgery and by computed tomography to have a large mass originating in the pancreas. Histopathology, molecular testing, and staging evaluations showed this to be a stage 3, MYCN unamplified, intermediate-risk neuroblastoma. The patient had a partial response to risk-stratified chemotherapy. The mass remained unresectable, but the response was sustained after 18 months. Although fewer than a dozen cases of primary pancreatic neuroblastoma have been reported, our experience and a literature review suggest that these tumors can be managed in the same way as similar-risk neuroblastoma of other sites.

Altering the Y137-K164-K166 triad of mandelate racemase and its effect on the observed pKa of the Brønsted base catalysts.

Mandelate racemase (MR) catalyzes the interconversion of the enantiomers of mandelate using a two-base mechanism with Lys 166 acting as the Brønsted base to abstract the α-proton from (S)-mandelate. The resulting intermediate is subsequently re-protonated by the conjugate acid of His 297 to yield (R)-mandelate. The roles of these amino acids are reversed when (R)-mandelate is the substrate. The side chains of Tyr 137, Lys 164, and Lys 166 form a H-bonding network and the proximity of the two ε-NH3+ groups is believed to lower the pKa of Lys 166. We used site-directed mutagenesis, kinetics, and pH-rate studies to explore the roles of Lys 164 (K164 C/M) and Tyr 137 (Y137  L/F/S/T) in catalysis. The efficiency (kcat/Km) was reduced ∼3.5 × 105-fold for K164C MR, relative to wild-type MR, indicating a major role for this residue in catalysis. The efficiency of Y137F MR, however, was reduced only 25-30-fold. pH-Rate profiles (log kcat vs. pH) revealed that substitution of Tyr 137 by Phe increased the kinetic pKa of Lys 166 from 5.88 ±â€¯0.02 to 7.3 ±â€¯0.2. Hence, Tyr 137 plays an important role in facilitating the reduction of the pKa of the Brønsted base Lys 166 by ∼1.4 units. Interestingly, the Phe substitution also increased the kinetic pKa of His 297 from 5.97 ±â€¯0.04 to 7.1 ±â€¯0.1. Thus, the Tyr 137-Lys 164-Lys 166 H-bonding network plays a broader role in modulating the pKa of catalytic residues by influencing the electrostatic character of the entire active site, not only by decreasing the observed pKa value of Lys 166, but also by decreasing the pKa of His 297 by 1.1 units.

Monosaccharide inhibitors targeting carbohydrate esterase family 4 de-N-acetylases.

The Carbohydrate Esterase family 4 contains virulence factors which modify peptidoglycan and biofilm-related exopolysaccharides. Despite the importance of this family of enzymes, a potent mechanism-based inhibition strategy has yet to emerge. Based on the postulated tridentate binding mode of the tetrahedral de-N-acetylation intermediate, GlcNAc derivatives bearing metal chelating groups at the 2 and 3 positions were synthesized. These scaffolds include 2-C phosphonate, 2-C sulfonamide, 2-thionoacetamide warheads as well as derivatives bearing thiol, amine and azide substitutions at the 3-position. The inhibitors were assayed against a representative peptidoglycan deacetylase, Pgda from Streptococcus pneumonia, and a representative biofilm-related exopolysaccharide deacetylase, PgaB from Escherichia coli. Of the inhibitors evaluated, the 3-thio derivatives showed weak to moderate inhibition of Pgda. The strongest inhibitor was benzyl 2,3-dideoxy-2-thionoacetamide-3-thio-ß-d-glucoside, whose inhibitory potency showed an unexpected dependence on metal concentration and was found to have a partial mixed inhibition mode (Ki = 2.9 ±â€¯0.6 µM).

Metabolic Usage and Glycan Destinations of GlcNAz in E. coli.

Bacteria use a diverse range of carbohydrates to generate a profusion of glycans, with amino sugars, such as N-acetylglucosamine (GlcNAc), being prevalent in the cell wall and in many exopolysaccharides. The primary substrate for GlcNAc-containing glycans, UDP-GlcNAc, is the product of the bacterial hexosamine pathway and a key target for bacterial metabolic glycan engineering. Using the strategy of expressing NahK, to circumvent the hexosamine pathway, it is possible to directly feed the analogue of GlcNAc, N-azidoacetylglucosamine (GlcNAz), for metabolic labeling in Escherichia coli. The cytosolic production of UDP-GlcNAz was confirmed by using fluorescence-assisted polyacrylamide gel electrophoresis. The key question of where GlcNAz is incorporated was interrogated by analyzing potential sites including peptidoglycan (PGN), the biofilm-related exopolysaccharide poly-ß-1,6-N-acetylglucosamine (PNAG), lipopolysaccharide (LPS), and the enterobacterial common antigen (ECA). The highest levels of incorporation were observed in PGN with lower levels in PNAG and no observable incorporation in LPS or ECA. The promiscuity of the PNAG synthase (PgaCD) toward UDP-GlcNAz in vitro and the lack of undecaprenyl-pyrophosphoryl-GlcNAz intermediates generated in vivo confirmed the incorporation preferences. The results of this work will guide the future development of carbohydrate-based probes and metabolic engineering strategies.

The Metabolic Usage and Glycan Destinations of GlcNAz in E. coli.

Bacteria use a diverse range of carbohydrates to generate a profusion of glycans, with amino sugars such as N-acetylglucosamine (GlcNAc) being prevalent in the cell wall and in many exopolysaccharides. The primary substrate for GlcNAc-containing glycans, UDP-GlcNAc, is the product of the bacterial hexosamine pathway, and a key target for bacterial metabolic glycan engineering. Using the strategy of expressing NahK, to circumvent the hexosamine pathway, it is possible to directly feed the analogue of GlcNAc, N-azidoacetylglucosamine (GlcNAz), for metabolic labelling in E. coli. The cytosolic production of UDP-GlcNAz was confirmed using fluorescence assisted polyacrylamide gel electrophoresis. The key question of where GlcNAz is incorporated, was interrogated by analyzing potential sites including: peptidoglycan (PGN), the biofilm-related exopolysaccharide poly-ß-1,6-N-acetylglucosamine (PNAG), lipopolysaccharide (LPS) and the enterobacterial common antigen (ECA). The highest levels of incorporation were observed in PGN with lower levels in PNAG and no observable incorporation in LPS or ECA. The promiscuity of the PNAG synthase (PgaCD) towards UDP-GlcNAz in vitro and lack of undecaprenyl-pyrophosphoryl-GlcNAz intermediates generated in vivo confirmed the incorporation preferences. The results of this work will guide the future development of carbohydrate-based probes and metabolic engineering strategies.

Performance of ACR TI-RADS and the Bethesda System in Predicting Risk of Malignancy in Thyroid Nodules at a Large Children's Hospital and a Comprehensive Review of the Pediatric Literature.

While thyroid nodules are less common in children than in adults, they are more frequently malignant. However, pediatric data are scarce regarding the performance characteristics of imaging and cytopathology classification systems validated to predict the risk of malignancy (ROM) in adults and select those patients who require fine-needle aspiration (FNA) and possibly surgical resection. We retrospectively reviewed the electronic medical records of all patients 18 years of age or younger who underwent thyroid FNA at our institution from 1 July 2015 to 31 May 2022. Based on surgical follow-up from 74 of the 208 FNA cases, we determined the ROM for the American College of Radiology Thyroid Imaging Reporting and Data System (ACR TI-RADS) ultrasound risk stratification system and The Bethesda System for Reporting Thyroid Cytopathology and added our results to those of pediatric cohorts from other institutions already published in the literature. We found the following ROMs for 1458 cases using ACR TI-RADS (TR): TR1. Benign: 2.2%, TR2. Not Suspicious: 9.3%, TR3. Mildly Suspicious: 16.6%, TR4. Moderately Suspicious: 27.0%, and TR5. Highly Suspicious 76.5%; and for 5911 cases using the Bethesda system: Bethesda I. Unsatisfactory: 16.8%, Bethesda II. Benign: 7.2%, Bethesda III: Atypia of Undetermined Significance: 29.6%, Bethesda IV. Follicular Neoplasm: 42.3%, Bethesda V. Suspicious for Malignancy: 90.8%, and Bethesda VI. Malignant: 98.8%. We conclude that ACR TI-RADS levels imply higher ROMs for the pediatric population than the corresponding suggested ROMs for adults, and, in order to avoid missing malignancies, we should consider modifying or altogether abandoning size cutoffs for recommending FNA in children and adolescents whose thyroid glands are smaller than those of adults. The Bethesda categories also imply higher ROMs for pediatric patients compared to adults.

Small Molecule Inhibition of an Exopolysaccharide Modification Enzyme is a Viable Strategy To Block Pseudomonas aeruginosa Pel Biofilm Formation.

Biosynthesis of the Pel exopolysaccharide in Pseudomonas aeruginosa requires all seven genes of the pelABCDEFG operon. The periplasmic modification enzyme PelA contains a C-terminal deacetylase domain that is necessary for Pel-dependent biofilm formation. Herein, we show that extracellular Pel is not produced by a P. aeruginosa PelA deacetylase mutant. This positions PelA deacetylase activity as an attractive target to prevent Pel-dependent biofilm formation. Using a high-throughput screen (n = 69,360), we identified 56 compounds that potentially inhibit PelA esterase activity, the first enzymatic step in the deacetylase reaction. A secondary biofilm inhibition assay identified methyl 2-(2-pyridinylmethylene) hydrazinecarbodithioate (SK-017154-O) as a specific Pel-dependent biofilm inhibitor. Structure-activity relationship studies identified the thiocarbazate as a necessary functional group and that the pyridyl ring could be replaced with a phenyl substituent (compound 1). Both SK-017154-O and compound 1 inhibit Pel-dependent biofilm formation in Bacillus cereus ATCC 10987, which has a predicted extracellular PelA deacetylase in its pel operon. Michaelis-Menten kinetics determined SK-017154-O to be a noncompetitive inhibitor of PelA, while compound 1 did not directly inhibit PelA esterase activity. Cytotoxicity assays using human lung fibroblast cells showed that compound 1 is less cytotoxic than SK-017154-O. This work provides proof of concept that biofilm exopolysaccharide modification enzymes are important for biofilm formation and can serve as useful antibiofilm targets. IMPORTANCE Present in more than 500 diverse Gram-negative and 900 Gram-positive organisms, the Pel polysaccharide is one of the most phylogenetically widespread biofilm matrix determinants found to date. Partial de-N-acetylation of this α-1,4 linked N-acetylgalactosamine polymer by the carbohydrate modification enzyme PelA is required for Pel-dependent biofilm formation in Pseudomonas aeruginosa and Bacillus cereus. Given this and our observation that extracellular Pel is not produced by a P. aeruginosa PelA deactylase mutant, we developed an enzyme-based high-throughput screen and identified methyl 2-(2-pyridinylmethylene) hydrazinecarbodithioate (SK-017154-O) and its phenyl derivative as specific Pel-dependent biofilm inhibitors. Michaelis-Menten kinetics revealed SK-017154-O is a noncompetitive inhibitor and that its noncytotoxic, phenyl derivative does not directly inhibit P. aeruginosa PelA esterase activity. We provide proof of concept that exopolysaccharide modification enzymes can be targeted with small molecule inhibitors to block Pel-dependent biofilm development in both Gram-negative and Gram-positive bacteria.

Termination of Poly-N-acetylglucosamine (PNAG) Polymerization with N-Acetylglucosamine Analogues.

Bacteria require polysaccharides for structure, survival, and virulence. Despite their central role in microbiology, few tools are available to manipulate their production. In E. coli, the glycosyltransferase complex PgaCD produces poly-N-acetylglucosamine (PNAG), an extracellular matrix polysaccharide required for biofilm formation. We report that C6-substituted (H, F, N3, SH, NH2) UDP-GlcNAc substrate analogues are inhibitors of PgaCD. In vitro, the inhibitors cause PNAG chain termination, consistent with the mechanism of PNAG polymerization from the nonreducing terminus. In vivo, expression of the GlcNAc-1-kinase NahK in E. coli provided a non-native GlcNAc salvage pathway that produced the UDP-GlcNAc analogue inhibitors in situ. The 6-fluoro and 6-deoxy derivatives were potent inhibitors of biofilm formation in the transformed strain, providing a tool to manipulate this key exopolysaccharide. Characterization of the UDP-GlcNAc pool and quantification of PNAG generation support PNAG termination as the primary in vivo mechanism of biofilm inhibition by 6-fluoro UDP-GlcNAc.

Allosteric modulation of the adenosine A2A receptor by cholesterol.

Cholesterol is a major component of the cell membrane and commonly regulates membrane protein function. Here, we investigate how cholesterol modulates the conformational equilibria and signaling of the adenosine A2A receptor (A2AR) in reconstituted phospholipid nanodiscs. This model system conveniently excludes possible effects arising from cholesterol-induced phase separation or receptor oligomerization and focuses on the question of allostery. GTP hydrolysis assays show that cholesterol weakly enhances the basal signaling of A2AR while decreasing the agonist EC50. Fluorine nuclear magnetic resonance (19F NMR) spectroscopy shows that this enhancement arises from an increase in the receptor's active state population and a G-protein-bound precoupled state. 19F NMR of fluorinated cholesterol analogs reveals transient interactions with A2AR, indicating a lack of high-affinity binding or direct allosteric modulation. The combined results suggest that the observed allosteric effects are largely indirect and originate from cholesterol-mediated changes in membrane properties, as shown by membrane fluidity measurements and high-pressure NMR.

Bias and discrimination in surgery: Where are we and what can we do about it?

Bias is an inclination or preconceived outlook that favors toward or against an idea, person, or group. It manifests in implicit and explicit ways throughout all aspects and institutions of society. These cognitive shortcuts are often based on stereotypes and can lead to prejudice and discrimination in medicine as they mediate interactions with patients, between providers, and at the institutional level. It is important to understand the drivers and consequences of bias in order to overcome barriers to representation, equity, and inclusion. This paper provides definitions of bias; discusses its manifestations across academic medicine at the institutional and individual levels; and concludes by examining techniques to reduce bias and measure progress. Equity for patients, families, and members of the broader surgical community cannot be achieved without reducing bias and discrimination. We call for action to increase intentional efforts that reduce the influences of bias in healthcare, research, and education, particularly in the field of pediatric surgery.

Pancreaticoduodenectomy is safe in appropriately resourced rural hospitals.

BACKGROUND: Research shows improved safety and treatment outcomes for patients undergoing pancreaticoduodenectomy at high-volume centers. Regionalization of pancreaticoduodenectomy to high-volume urban centers can result in unintended negative consequences for rural patients and communities. This report examines outcomes after pancreaticoduodenectomy performed at a rural hospital and compares them with national standards. METHODS: A prospectively maintained database of pancreatic operations performed at a rural tertiary hospital was queried. Demographic and clinical information for patients undergoing pancreaticoduodenectomy (2007-2019) was analyzed. Primary outcomes were the rates of patient mortality and morbidity. Secondary outcomes were readmission rates, indications, and associations with clinical variables. RESULTS: We included 118 patients in our study. There were 41 postoperative complications (34.7%), including 1 death (0.9%). The 90-day readmission rate was 24.6%. The most common indication for readmission was deep space infection (n = 7, 24.1%). Patients requiring an intraoperative transfusion were more likely to need hospital readmission (41.4% vs 9.0% of patients without transfusion, P = .016). Patients with postoperative complications required readmission more frequently (51.7% vs 29.2%, P = .093). These findings are similar to data from urban hospitals. CONCLUSION: Patient safety and surgical outcomes after pancreaticoduodenectomy performed in appropriately resourced rural hospitals can be comparable with national standards. Safely treating rural patients near their home benefits patients and their communities.

Synthesis of C6-substituted UDP-GlcNAc derivatives.

UDP-sugar analogs are useful for the study of glycosyltransferases and the production of unnatural glycans. The preparation of five UDP-GlcNAc derivatives is reported with 6-deoxy, 6-azido, 6-amino, 6-mercapto, or 6-fluoro substitutions. A concise chemoenzymatic synthesis was developed using the kinase NahK (B. longum JCM1217) and the uridyl transferase GlmU (E. coli K12).

TBAF Effects 3,6-Anhydro Formation from 6-O-Tosyl Pyranosides.

3,6-Anhydro sugars are common structures in algal polysaccharides and occur in the furanodictine and sauropunol natural products. We have found that treatment of 6-O-tosylpyranosides with tetrabutylammonium fluoride provides a mild, high-yielding synthesis of 3,6-anhydro sugars. Using O-glycoside substrates, 3,6-anhydropyranosides are isolated and the use of N,O-dimethyl hydroxylamine glycosides yields 3,6-anhydrofuranosides. Applying this approach, concise synthetic routes to several 3,6-anhydro sugar natural products are reported, including furanodictine A and sauropunols A-D.

Trends in Orthopedic Surgery Reimbursement From 2000 to 2015.

Understanding trends in reimbursement for orthopedic surgery is important, especially considering the changing landscape of health care delivery and payment models. Although other studies have examined these trends using a sampling of common orthopedic procedures compared with non-orthopedic specialties, robust examination across all orthopedic specialties is not available in the current literature. This study aimed to critically analyze the trends in reimbursement in the field of orthopedic surgery. Inflation-adjusted Medicare reimbursement and work relative value units (RVUs) between 2000 and 2016 for more than 200 individual Current Procedural Terminology codes across all major orthopedic subspecialties were analyzed, and inherent value of work RVUs was assessed by dividing reimbursement dollar values by work RVUs annually and tracking the changes. Between 2000 and 2016, reimbursement decreased across all orthopedic subspecialties by an average of 29%, except oncology, which showed a 6% increase. Work RVUs increased by an average of 10%, but the inherent value of work RVUs decreased across all orthopedic subspecialties by an average of 39%. Increased active involvement of orthopedic attending physicians and residents in coding documentation and fee-schedule representation is needed. [Orthopedics. 2020;43(3):187-190.].

Resident Participation as Co-Surgeon Does Not Adversely Impact Patient Outcomes in Pancreatic Surgery.

OBJECTIVE: In academic settings, surgical residents often serve as co-surgeon in complex operations such as pancreatic resections. These operations are typically performed by fellowship-trained primary surgeons with extensive experience in the field. Our study aimed to evaluate how the participation of general surgery residents in these complex operations affected patient outcomes. Our hypothesis was that resident involvement as co-surgeon would not adversely impact key patient outcomes including complications, readmission, and mortality. DESIGN: A REDCap database of perioperative variables for patients undergoing pancreatic resection was established at a single independent academic medical center. The database was populated via retrospective chart review. Patient demographics, surgical indications, operative time, estimated blood loss, postoperative hospital length of stay, intensive care unit length of stay, postoperative complications, and 30- and 90-day survival for patients with and without cancer were reviewed. We further categorized the data based on the designation of a general surgery resident or a second staff surgeon as co-surgeon in each operation. SETTING: The study was performed at the Marshfield Clinic Health System-Marshfield Medical Center, an independent academic medical center located in central Wisconsin. PARTICIPANTS: Data were abstracted from the medical records of all adult patients (18 years of age and older) who underwent pancreatic resection from 2007 to 2018 (n = 173). RESULTS: 173 pancreatic resections were performed by 8 different primary staff surgeons over 10.5 years. All co-surgeons were either another staff surgeon or a senior-level (postgraduate year 4 or 5) general surgery resident. Perioperative and postoperative patient outcomes were statistically similar in both groups. CONCLUSIONS: Resident involvement as co-surgeon in complex pancreatic resections constituted no increased risk for patients at our institution. Senior residents should continue to operate on these important learning cases under appropriate staff supervision.

A systematic review of management options in pediatric rectal prolapse.

PURPOSE: Rectal prolapse is a relatively common condition in infants and young children with a multifactorial etiology. Despite its prevalence, there remains clinical equipoise with respect to secondary treatment in pediatric surgery literature. We conducted a systematic review to evaluate methods of secondary treatment currently used to treat rectal prolapse in children. METHODS: We searched Pubmed, Medline, and Scopus with the terms "rectal prolapse" and "children" for papers published from 1990 to April 2017. Papers satisfying strict criteria were analyzed for patient demographics, intervention, efficacy, and complications. Procedures were grouped by like type. Pooled success rates were calculated. RESULTS: Twenty-seven studies documenting 907 patients were included. Injection sclerotherapy had an overall initial success rate of 79.5%. Ethyl alcohol seemed the best sclerosing agent due to a high first-injection success rate, low complication rate, and ready accessibility. Several perineal repairs were found, with operative success rates ranging from 60.8%-100%. Laparoscopic rectopexy with mesh was the most commonly reported transabdominal procedure and had an overall success rate of 96.1%. Postoperative complications from all procedures were comparable. CONCLUSION: Though many secondary treatment options have been reported for rectal prolapse, sclerotherapy and laparoscopic rectopexy predominate in contemporary literature and appear to have high success and low complication rates. LEVEL OF EVIDENCE: IV.

A dedicated anticoagulation clinic does not improve postoperative management of warfarin after total joint arthroplasty.

BACKGROUND: Periprosthetic joint infections (PJIs) are devastating complications. Excessive anticoagulation with warfarin is an independent risk factor for PJIs. The use of a dedicated anticoagulation clinic to improve warfarin management has not been proven. METHODS: Between 2006 and 2014, we identified 92 patients who were placed on postoperative warfarin, and later developed PJI. These patients were compared to 313 patients who underwent total joint arthroplasty placed on warfarin without developing PJI. Patients were included if they had no history of a venous thromboembolic event, were warfarin naive, and enrolled in the anticoagulation clinic. A univariate analysis compared independent variables, and statistical analysis was performed using Student's t-test and Pearson chi-square test for continuous and categorical variables. RESULTS: Thirty-six PJI patients and 297 control patients met the inclusion criteria. The venous thromboembolism rate was 2.1%. At discharge, 82% of all patients were subtherapeutic. Patients were within their target international normalized ratio (INR) range 26.7% of the time. The mean INR in the initial postoperative period for the PJI group was 1.46 and 1.29 in the control group (P < .001). In the acute postoperative period, 13.3% of the knee PJI group were therapeutic or supratherapeutic compared with 3.5% in the knee control group (P = .002). CONCLUSIONS: Despite utilization of a dedicated anticoagulation clinic, patients were only within their target INR range 27% of the time. Total knee arthroplasty patients who developed a PJI were more likely to be therapeutic or supratherapeutic in the initial postoperative period. Consequently, the risks associated with warfarin as a venous thromboembolism prophylaxis may outweigh the potential benefits.

Teaching corner: an undergraduate medical education program comprehensively integrating global health and global health ethics as core curricula : student experiences of the medical school for international health in Israel.

The Medical School for International Health (MSIH) was created in 1996 by the Faculty of Health Sciences at Ben-Gurion University of the Negev in affiliation with Columbia University's Health Sciences division. It is accredited by the New York State Board of Education. Students complete the first three years of the program on the Ben-Gurion University campus in Be'er-Sheva, Israel, while fourth-year electives are completed mainly in the United States (at Columbia University Medical Center and affiliates as well as other institutions) along with a two-month global health elective at one of numerous sites located around the world (including Canada, Ethiopia, India, Israel, Kenya, Nepal, Peru, the Philippines, Sri Lanka, Uganda, the United States, and Vietnam). The unique four-year, American-style curriculum is designed not only to prepare physicians who will be able to work at both an individual and community level but also at both of these levels anywhere in the world. In this way, it combines elements of medical and public health curricula not limited to an American perspective.