The Learning Curve for Robotic Lateral Pelvic Lymph Node Dissection for Rectal Cancer: A View from the West.

BACKGROUND: Lateral pelvic lymph-node dissection is performed for selected patients with rectal cancer with persistent lateral nodal disease after neoadjuvant therapy. This technique has been slow to be adopted in the West due to concerns regarding technical difficulty. This is the first report on the learning curve for lateral pelvic lymph node dissection in the US or Europe. OBJECTIVE: The aim of this study was to analyze the learning curve associated with robotic lateral pelvic lymph node dissection. DESIGN: Retrospective observational cohort. SETTING: Tertiary academic cancer center. PATIENTS: Consecutive patients from 2012 to 2021. INTERVENTION: All patients underwent robotic lateral pelvic lymph node dissection. MAIN OUTCOME MEASURES: The primary endpoints were the learning curves for maximum number of nodes retrieved and urinary retention which was evaluated with simple cumulative-sum and two-sided Bernoulli cumulative-sum charts. RESULTS: Fifty-four procedures were included. A single-surgeon (n = 35) and an institutional learning curve are presented in the analysis. In the single-surgeon learning curve, a turning point marking the end of a learning phase was detected at the 12th procedure for the number of retrieved nodes and at the 20th for urinary retention. In the institutional learning curve analysis, two turning points were identified at the 13th and 26th procedures indicating progressive improvements for the number of retrieved nodes and at the 27th for urinary retention. No sustained alarm signals were detected at any time point. LIMITATIONS: The retrospective nature, small sample size and the referral center nature of the reporting institution that may limit generalizability. CONCLUSIONS: In a setting of institutional experience with robotic colorectal surgery including beyond TME resections, the learning curve for robotic lateral pelvic lymph node dissection is acceptably short. Our results demonstrate feasibility of acquisition of this technique in a controlled setting, with sufficient case volume and proctoring can optimize the learning curve. See Video Abstract.

Perioperative and oncological outcomes following robotic en bloc multivisceral resection for colorectal cancer.

AIM: As multidisciplinary treatment strategies for colorectal cancer have improved, aggressive surgical resection has become commonplace. Multivisceral and extended resections offer curative-intent resection with significant survival benefit. However, limited data exist regarding the feasibility and oncological efficacy of performing extended resection via a minimally invasive approach. The aim of this study was to determine the perioperative and long-term outcomes following robotic extended resection for colorectal cancer. METHOD: We describe the population of patients undergoing robotic multivisceral resection for colorectal cancer at our single institution. We evaluated perioperative details and investigated short- and long-term outcomes, using the Kaplan-Meier method to analyse overall and recurrence-free survival. RESULTS: Among the 86 patients most tumours were T3 (47%) or T4 (47%) lesions in the rectum (78%). Most resections involved the anterior compartment (72%): bladder (n = 13), seminal vesicle/vas deferens (n = 27), ureter (n = 6), prostate (n = 15) and uterus/vagina/adnexa (n = 27). Three cases required conversion to open surgery; 10 patients had grade 3 complications. The median hospital stay was 4 days. Resections were R0 (>1 mm) in 78 and R1 (0 to ≤1 mm) in 8, with none being R2. The average nodal yield was 26 and 48 (55.8%) were pN0. Three-year overall survival was 88% and median progression-free survival was 19.4 months. Local recurrence was 6.1% and distant recurrence was 26.1% at 3 years. CONCLUSION: Performance of multivisceral and extended resection on the robotic platform allows patients the benefit of minimally invasive surgery while achieving oncologically sound resection of colorectal cancer.

Domestic violence towards children in Kenya and Zambia during the COVID-19 pandemic.

OBJECTIVES: Although research has demonstrated that the COVID-19 pandemic has increased the likelihood of children experiencing domestic abuse, few empirical studies have focused on the most vulnerable communities in developing countries. This study aimed to evaluate the impact of the COVID-19 pandemic on domestic violence towards children in Kenya and Zambia. STUDY DESIGN: In collaboration with an international non-governmental organisation (NGO), an original door-to-door survey of 842 children and their parents (or adult guardians) was conducted in Kenya and Zambia. The survey sites were carefully selected to target economically underprivileged communities with the presence of NGOs as well as those without. METHODS: Using multivariate regression analysis of child and parent survey responses, we attempted to identify how parents' experiences of COVID-19 affected children's exposure to domestic violence. Our analysis of domestic violence included both emotional (psychological) and physical violence. RESULTS: Children from households whose guardians perceived a strongly negative effect of COVID-19 were more likely to experience domestic violence. The correlation was statistically significant for both emotional and physical violence. In Kenya, children in communities with NGO presence were less likely to experience domestic violence. CONCLUSIONS: The COVID-19 pandemic has exposed children to an increased risk of experiencing domestic violence. Children from underprivileged communities in developing countries are particularly vulnerable to such risk, as they tend to lack social support from communities and governments. International organisations must pay more attention to these children during the pandemic.

Expression of Immune-Related and Inflammatory Markers and Their Prognostic Impact in Colorectal Cancer Patients.

The tumor microenvironment of colorectal cancer (CRC) is heterogenous; thus, it is likely that multiple immune-related and inflammatory markers are simultaneously expressed in the tumor. The aim of this study was to identify immune-related and inflammatory markers expressed in freshly frozen CRC tissues and to investigate whether they are related to the clinicopathological features and prognosis of CRC. Seventy patients with CRC who underwent curative surgical resection between December 2014 and January 2017 were included in this study. Tissue samples were obtained from tumor and non-tumor areas in the patients' colons. The concentrations of immune-related markers (APRIL/TNFSF13, BAFF, LAG-3, PD-1, PD-L1, and CTLA-4) and inflammatory markers (CHIT, MMP-3, osteocalcin, pentraxin-3, sTNF-R1, and sTNF-R2) in the samples were measured using the Bio-plex Multiplex Immunoassay system. The concentrations of APRIL/TNFSF13, BAFF, and MMP-3 in the samples were significantly high; thus, we conducted analyses based on the cut-off values for these three markers. The high-APRIL/TNFSH13-expression group showed a significantly higher rate of metastatic lesions than the low-expression group, whereas the high-MMP-3-expression group had higher CEA levels, more lymph node metastases, and more advanced disease stages than the low-expression group. The five-year disease-free survival of the high-MMP-3-expression group was significantly shorter than that of the low-expression group (65.1% vs. 90.2%, p = 0.033). This study provides evidence that the APRIL/TNFSF13, BAFF, and MMP-3 pathway is overexpressed in CRC tissues and is associated with unfavorable clinicopathological features and poor prognosis in CRC patients. These markers could serve as diagnostic or prognostic biomarkers for CRC.

Magnetic Resonance Imaging Directed Surgical Decision Making for Lateral Pelvic Lymph Node Dissection in Rectal Cancer After Total Neoadjuvant Therapy (TNT).

OBJECTIVE: Lateral pelvic lymph node (LPLN) metastases are an important cause of preventable local failure in rectal cancer. The aim of this study was to evaluate clinical and oncological outcomes following magnetic resonance imaging (MRI)-directed surgical selection for lateral pelvic lymph node dissection (LPLND) after total neoadjuvant therapy (TNT). METHODS: A retrospective consecutive cohort analysis was performed of rectal cancer patients with enlarged LPLN on pretreatment MRI. Patients were categorized as LPLND or non-LPLND. The main outcomes were lateral local recurrence rate, perioperative and oncological outcomes and factors associated with decision making for LPLND. RESULTS: A total of 158 patients with enlarged pretreatment LPLN and treated with TNT were identified. Median follow-up was 20 months (interquartile range 10-32). After multidisciplinary review, 88 patients (56.0%) underwent LPLND. Mean age was 53 (SD±12) years, and 54 (34.2%) were female. Total operative time (509 vs 429 minutes; P =0.003) was greater in the LPLND group, but median blood loss ( P =0.70) or rates of major morbidity (19.3% vs 17.0%) did not differ. LPLNs were pathologically positive in 34.1%. The 3-year lateral local recurrence rates (3.4% vs 4.6%; P =0.85) did not differ between groups. Patients with LPLNs demonstrating pretreatment heterogeneity and irregular margin (odds ratio, 3.82; 95% confidence interval: 1.65-8.82) or with short-axis ≥5 mm post-TNT (odds ratio 2.69; 95% confidence interval: 1.19-6.08) were more likely to undergo LPLND. CONCLUSIONS: For rectal cancer patients with evidence of LPLN metastasis, the appropriate selection of patients for LPLND can be facilitated by a multidisciplinary MRI-directed approach with no significant difference in perioperative or oncologic outcomes.

High Versus Low Ligation of the Inferior Mesenteric Artery in Colorectal Cancer Surgery: A Systematic Review and Meta-Analysis.

Background and Objectives: This study aimed to compare the effects of high ligation (HL) versus low ligation (LL) in colorectal cancer surgery. Materials and Methods: We performed a comprehensive search using multiple databases (trial registries and ClinicalTrials.gov), other sources of grey literature, and conference proceedings, with no restrictions on the language or publication status, up until 10 March 2021. We included all parallel-group randomized controlled trials (RCTs) and considered cluster RCTs for inclusion. The risk of bias domains were "low risk," "high risk," or "unclear risk." We performed statistical analyses using a random-effects model and interpreted the results according to the Cochrane Handbook for Systematic Reviews of Interventions. We used the GRADE guidelines to rate the certainty of evidence (CoE) of the randomized controlled trials. Results: We found 12 studies (24 articles) from our search. We were very uncertain about the effects of HL on overall mortality, disease recurrence, cancer-specific mortality, postoperative mortality, and anastomotic leakage (very low CoE). There may be little to no difference between HL and LL in postoperative complications (low CoE). For short-term follow-up (within 6 months), HL may reduce defecatory function (constipation; low CoE). While HL and LL may have similar effects on sexual function in men, HL may reduce female sexual function compared with LL (low CoE). For long-term follow-up (beyond 6 months), HL may reduce defecatory function (constipation; low CoE). There were discrepancies in the effects regarding urinary dysfunction according to which questionnaire was used in the studies. HL may reduce male and female sexual function (low CoE). Conclusions: We are very uncertain about the effects of HL on survival outcomes, and there is no difference in the incidence of postoperative complications between HL and LL. More rigorous RCTs are necessary to evaluate the effect of HL and LL on functional outcomes.

Perioperative Probiotics Application for Preventing Postoperative Complications in Patients with Colorectal Cancer: A Systematic Review and Meta-Analysis.

Background and Objectives: Perioperative probiotic administration in patients who undergo gastrointestinal surgery can reduce postoperative infectious complications. This systematic review and meta-analysis aimed to evaluate the effect of probiotics on postoperative outcomes in patients who underwent colorectal cancer surgery. Materials and Methods: For this study, we followed the protocol published by PROSPERO (registration number: CRD42021247277). We included studies on patients undergoing open, laparoscopic, or robotic colorectal cancer surgery for curative intent. We conducted a comprehensive search with online databases (trial registries and ClinicalTrials.gov), other literature sources, and conference proceedings, with no language restriction, up until 12 August 2022. We assessed risk of bias, extracted data, and conducted statistical analyses by using a random-effects model and interpreted the results based on the Cochrane Handbook for Systematic Reviews of Interventions. We rated the certainty of evidence (CoR) according to the GRADE approach. Results: We identified 20 published full-text studies. The use of probiotics probably results in little to no difference in perioperative mortality (risk ratio (RR): 0.17, 95% CI: 0.02 to 1.38; I2 = 0%; moderate CoE) and may result in reducing the overall postoperative infectious complications (RR: 0.45, 95% CI: 0.27 to 0.76; I2 = 38%; low CoE) after colorectal cancer surgery. Probiotics may result in little to no difference in probiotics-related adverse events (RR: 0.73, 95% CI: 0.45 to 1.19; I2 = 0%; low CoE). While probiotics may result in reducing the overall postoperative complications (RR: 0.47, 95% CI: 0.30 to 0.74; I2 = 8%; low CoE), it may result in little to no difference in hospital length of stay (LOS) (MD: -1.06, 95% CI: -1.64 to -0.47; I2 = 8%; low CoE) and postoperative quality of life (QOL) (MD: +5.64, 95% CI: 0.98 to 10.3; low CoE). Conclusions: Perioperative probiotic administration may reduce complications, including overall infectious complications, in patients undergoing colorectal cancer surgery without any additional adverse effects. In addition, probiotics may have similar effects on perioperative mortality; procedure-related complications such as anastomotic leakage, and hospital LOS; or improve the QOL. Thus, probiotics may be considered a beneficial supplement to routine perioperative care for colorectal cancer surgery.

pH-promoted O-α-glucosylation of flavonoids using an engineered α-glucosidase mutant.

Retaining glycosidase mutants lacking its general acid/base catalytic residue are originally termed thioglycoligases which synthesize thio-linked disaccharides using sugar acceptor bearing a nucleophilic thiol group. A few thioglycoligases derived from retaining α-glycosidases have been classified into a new class of catalysts, O-glycoligases which transfer sugar moiety to a hydroxy group of sugar acceptors, resulting in the formation of O-linked glycosides or oligosaccharides. In this study, an efficient O-α-glucosylation of flavonoids was developed using an O-α-glycoligase derived from a thermostable α-glucosidase from Sulfolobus solfataricus (MalA-D416A). The O-glycoligase exhibited efficient transglycosylation activity with a broad substrate spectrum for all kinds of tested flavonoids including flavone, flavonol, flavanone, flavanonol, flavanol and isoflavone classes in yields of higher than 90%. The glucosylation by MalA-D416A preferred alkaline conditions, suggesting that pH-promoted deprotonation of hydroxyl groups of the flavonoids would accelerate turnover of covalent enzyme intermediate via transglucosylation. More importantly, the glucosylation of flavonoids by MalA-D416A was exclusively regioselective, resulting in the synthesis of flavonoid 7-O-α-glucosides as the sole product. Kinetic analysis and molecular dynamics simulations provided insights into the acceptor specificity and the regiospecificity of O-α-glucosylation by MalA-D416A. This pH promoted transglycosylation using O-α-glycoligases may prove to be a general synthesis route to flavonoid O-α-glycosides.

Differences in clinical features and oncologic outcomes between metastatic right and left colon cancer.

Approximately 20% to 25% of patients with colorectal cancer (CRC) have distant organ metastasis at the time of initial diagnosis. The primary tumor location has been suggested as a prognostic factor for patients with metastatic CRC. In recent years, the distinction between right colon cancer (RCC) and left colon cancer (LCC) has been brought into focus due to their different outcomes, prognoses, and clinical responses to chemotherapy. In this article we aimed to review the underlying differences between metastatic RCC and LCC in terms of epidemiology, clinical features, and oncologic outcomes. The outcomes of patients with left-sided tumors were better than those of patients with right-sided tumors in terms of overall survival (OS) and objective response rate (ORR) after treatment with chemotherapy + panitumumab in the PRIME and 20050181 trials. The outcomes of patients with LCC were better than those of patients with RCC in terms of OS, progression-free survival (PFS) and ORR after treatment with FOLFIRI + cetuximab in the CRYSTAL and CALGB 80405 trials. In the FIRE-3 trial, the OS and PFS, but not the ORR, of patients with LCC were superior to those of patients with RCC. LCC and RCC exhibit distinctive clinical features and epidemiology. However, we must further investigate the impact of these distinctive features and how they influence the differential oncologic outcomes.

Color Fine-Tuning of Optical Materials Through Rational Design.

We report on the feasibility for color fine-tuning of optical materials using rational design principles based on chemical reasoning. For this purpose, a modular framework for the construction of symmetrical cap-linker-cap compounds, using triarylamine caps and oligothiophene linkers, is applied. The chosen structural scaffolds are heavily used in recent industrial applications and provide five possibilities for altering their electronic and steric properties: electron donor/acceptor groups, planarization/deplanarization, and modulation of the π-conjugation length. Permutation of the used building blocks leads to a set of 54 different molecules, out of which 32 are synthesized and characterized in solution as well as in example fabricated OLED devices. This setup allows for color fine-tuning in the range of 412 nm to 540 nm with typical steps of 4 nm. In addition, to further benefit from the large experimental data set the spectroscopic results are used to benchmark quantum chemical computations, which show excellent agreement thus highlighting the potential of these calculations to guide future syntheses.

Construction of an antimyoglobin single-chain variable fragment with rapid reaction kinetics.

Antibodies with rapid reaction kinetics (high association and dissociation rates), named reversible antibodies, are used to perform continuous monitoring of sensitive disease biomarkers. In cases of acute myocardial infarction (AMI), continuous monitoring and early diagnosis are important. Human myoglobin (Myo) is a useful biomarker for AMI during the early stage after the onset of symptoms. In this study, a single-chain variable fragment (scFv) specific to Myo was derived from an IgG antibody that has rapid reaction kinetics. Enzyme-linked immunosorbent assay revealed that recombinant scFv exhibited 3.8-fold reduced affinity compared with the parent IgG antibody based on the antibody concentration necessary for 50% of the maximum signal. The scFv retained the rapid reaction kinetic mode with average kon and koff of 2.63 × 10(5) M(-1) Sec(-1) and 3.25 × 10(-3) Sec(-1) , respectively, which were reduced to 10- and 2.3-fold compared with those of the parent antibody. The equilibrium constant for the association of the scFv (KA = 8.09 × 10(7) M(-1) ) was 4.6-fold lower than that of its parent IgG antibody. This scFv may be a starting point for further mutagenesis/kinetic and structural analyses providing valuable insight into the mechanism of reversible antibodies.

The Indian Ocean tsunami and private donations to NGOs.

Non-governmental organisations (NGOs) are widely believed to raise their flag in humanitarian hotspots with a strong media presence in order to attract higher private donations. We assess this hypothesis by comparing the changes in donations between US-based NGOs with and without aid operations in the four countries most affected by the tsunami in the Indian Ocean in 2004. Simple before-after comparisons tend to support the hypothesis that 'flying the flag' helps attract higher private donations. However, performing a difference-in-difference-in-differences (DDD) approach, we find only weak indications that private donors systematically and strongly preferred NGOs with operations in the region. Extended specifications of the baseline regressions reveal that our major findings are robust. NGO heterogeneity matters in some respects, but the DDD results hold when accounting for proxies of the NGOs' reputation and experience.

Structural and biochemical characterization of the broad substrate specificity of Bacteroides thetaiotaomicron commensal sialidase.

Sialidases release the terminal sialic acid residue from a wide range of sialic acid-containing polysaccharides. Bacteroides thetaiotaomicron, a symbiotic commensal microbe, resides in and dominates the human intestinal tract. We characterized the recombinant sialidase from B. thetaiotaomicron (BTSA) and demonstrated that it has broad substrate specificity with a relative activity of 97, 100 and 64 for 2,3-, 2,6- and 2,8-linked sialic substrates, respectively. The hydrolysis activity of BTSA was inhibited by a transition state analogue, 2-deoxy-2,3-dehydro-N-acetyl neuraminic acid, by competitive inhibition with a Ki value of 35µM. The structure of BSTA was determined at a resolution of 2.3Å. This structure exhibited a unique carbohydrate-binding domain (CBM) at its N-terminus (a.a. 23-190) that is adjacent to the catalytic domain (a.a. 191-535). The catalytic domain has a conserved arginine triad with a wide-open entrance for the substrate that exposes the catalytic residue to the surface. Unlike other pathogenic sialidases, the polysaccharide-binding site in the CBM is near the active site and possibly holds and positions the polysaccharide substrate directly at the active site. The structural feature of a wide substrate-binding groove and closer proximity of the polysaccharide-binding site to the active site could be a unique signature of the commensal sialidase BTSA and provide a molecular basis for its pharmaceutical application.

Characterization of a galactosynthase derived from Bacillus circulans ß-galactosidase: facile synthesis of D-lacto- and D-galacto-N-bioside.

Glycosynthases-retaining glycosidases mutated at their catalytic nucleophile-catalyze the formation of glycosidic bonds from glycosyl fluorides as donor sugars and various glycosides as acceptor sugars. Here the first glycosynthase derived from a family 35 ß-galactosidase is described. The Glu→Gly mutant of BgaC from Bacillus circulans (BgaC-E233G) catalyzed regioselective galactosylation at the 3-position of the sugar acceptors with α-galactosyl fluoride as the donor. Transfer to 4-nitophenyl α-D-N-acetyl-glucosaminide and α-D-N-acetylgalactosaminide yielded 4-nitophenyl α-lacto-N-biose and α-galacto-N-biose, respectively, in high yields (up to 98%). Kinetic analysis revealed that the high affinity of the acceptors contributed mostly to the BgaC-E233G-catalyzed transglycosylation. BgaC-E233G showed no activity with ß-(1,3)-linked disaccharides as acceptors, thus suggesting that this enzyme can be used in "one-pot synthesis" of LNB- or GNB-containing glycans.

Refolded scFv antibody fragment against myoglobin shows rapid reaction kinetics.

Myoglobin is one of the early biomarkers for acute myocardial infarction. Recently, we have screened an antibody with unique rapid reaction kinetics toward human myoglobin antigen. Antibodies with rapid reaction kinetics are thought to be an early IgG form produced during early stage of in vivo immunization. We produced a recombinant scFv fragment for the premature antibody from Escherichia coli using refolding technology. The scFv gene was constructed by connection of the V(H)-V(L) sequence with a (Gly4Ser)3 linker. The scFv fragment without the pelB leader sequence was expressed at a high level, but the solubility was extremely low. A high concentration of 8 M urea was used for denaturation. The dilution refolding process in the presence of arginine and the redox reagents GSH and GSSH successfully produced a soluble scFv protein. The resultant refolded scFv protein showed association and dissociation values of 9.32 × 10⁻4 M⁻¹·s⁻¹ and 6.29 × 10⁻³ s⁻¹, respectively, with an affinity value exceeding 107 M⁻¹ (k(on)/k(off)), maintaining the original rapid reaction kinetics of the premature antibody. The refolded scFv could provide a platform for protein engineering for the clinical application for diagnosis of heart disease and the development of a continuous biosensor.

The Expression Patterns of Immune Checkpoint Molecules in Colorectal Cancer: An Analysis Based on Microsatellite Status.

Recently, immunotherapy has arisen as a novel treatment approach for patients with colorectal cancer (CRC), but the effectiveness of immunotherapy varies in these patients. We hypothesized that immune checkpoint molecules (ICMs), which are the targets of immunotherapy, are often exhibited concomitantly. Our objective was to investigate the patterns of ICM expression in patients with CRC and the differences in ICM expression based on microsatellite instability status. The immunohistochemical expression of programmed cell death protein 1 (PD-1), programmed cell death ligand 1 (PD-L1), T cell immunoglobulin and mucin-domain containing-3 (TIM-3), and lymphocyte-activation gene 3 (LAG-3) in the tumor center and periphery was assessed in patients with non-metastatic colorectal cancer. We enrolled 83 patients with CRC: a total of 40 microsatellite-stable (MSS) and 43 microsatellite-instability-high (MSI-H) cancer patients. PD-L1 was more frequently expressed in the tumor center in the MSI-H patients with than that in the MSS patients (18 [41.9%] vs. 3 [7.5%], respectively; p < 0.001), and the same trend was observed for TIM-3 expression (30 [69.8%] vs. 19 [47.5%], respectively; p = 0.047). The concomitant expression of two or more ICMs was more frequently observed than no expression or the expression of a single molecule in both the MSS and MSI-H groups; a total of 34 (79.7%) patients with MSI-H cancer and 23 (57.5%) with MSS cancer showed ICM expression at the tumor center, whereas 34 (79.7%) patients with MSI-H cancer and 22 (55%) with MSS cancer showed expression at the tumor periphery. Patients with the genetic characteristics of MSI-H cancer showed higher expression levels of ICMs than those in patients with MSS cancer, and predominantly, two or more ICMs were concurrently expressed. Our findings highlight the potential efficacy of the dual-blockade approach in immunotherapy, particularly in patients with MSI-H CRC.

Overproduction of a thermostable 4-α-glucanotransferase by codon optimization at N-terminus region.

BACKGROUND: 4-α-Glucanotransferases are useful enzymes to modify starch owing to their transglycosylation activity. In this study, codon optimizations were conducted to overproduce a thermostable 4-α-glucanotransferase from Thermus thermophilus (TTαGT). RESULTS: Two variants, termed TTαGT-P4CCG and TTαGT-mut6, were constructed, which have the optimized codon at the first rare codon and optimized codons at all six chosen rare codons at the N-terminus of TTαGT, respectively. In the Escherichia coli system, the expression of both optimized genes was enhanced by about 100-fold relative to that of the original gene, whereas all six mutated codons contributed to the overall enhancement of TTαGT production in Bacillus subtilis. On the basis of the αGTase activity of the crude cell extracts, relative activities of 1:2.9:5.8 were determined for TTαGT, TTαGT-P4CCG and TTαGT-mut6, respectively, in B. subtilis. In addition, the activity of TTαGT-mut6 from B. subtilis grown without antibiotics was as much as that with the antibiotics. Finally, after heat treatment, the specific activity of TTαGT-mut6 from B. subtilis was 1.5-fold greater than that from E. coli. CONCLUSION: The codon-optimized TTαGT that was produced in a GRAS microorganism, B. subtilis, without the selection antibiotics is potentially useful in the food industry as a food-grade enzyme.

Characterization of amine oxidases from Arthrobacter aurescens and application for determination of biogenic amines.

Biogenic amines (BAs) that are produced through naturally occurring decarboxylation of amino acids have toxicological effects on humans. Bacterial amine oxidases are useful tools for the rapid quantification of BAs in foods. To develop amine oxidases for the rapid detection of BAs, the genes for amine oxidases from Arthobacter aurescens TC-1, designated AMAO1, AMAO2, and AMAO3, respectively, were cloned and expressed in Escherichia coli. AMAO1 was catalytically inactive to BAs, and AMAO3 showed a narrow substrate spectrum. In contrast, AMAO2 exhibited activity with relative k cat/K M values of 100:49.6:7.6 for 2-phenylethylamine, tyramine, and histamine, respectively. AMAO2 also utilized putrescine and spermidine as substrates, with four or five orders of magnitude lower k cat/K M values than that of 2-phenylethylamine. AMAO2 and AMAO3 were seriously affected by substrate inhibition. Using BA mixtures (consisting of 2-phenylethylamine, tyramine, and histamine) as samples, the detection range of the enzymatic analysis of BA using AMAO2 was determined to be 2.5-120 µM, and its detection limit was 2.3 µM. Analysis of five commercial cheese products revealed that the BA contents determined by the enzymatic methods showed a good agreement with the sum of three monoamines and histamine by HPLC. Therefore, the enzymatic assay using AMAO2 can be used in quality control of food products through rapid, sensitive, and preliminary estimation of major BAs including the most important TyrN and HisN in foods.

Reporting of Circumferential Resection Margin in Rectal Cancer Surgery.

Importance: Circumferential resection margin (CRM) in rectal cancer surgery is a major prognostic indicator associated with local recurrence and overall survival. Facility rates of CRM positivity have recently been established as a new quality measure by the Commission on Cancer (CoC); however, the completeness of CRM status reporting is not well characterized. Objective: To describe the changes in CRM reporting and factors associated with low rates of reporting. Design, Setting, and Participants: A retrospective cohort study was conducted using data from the National Cancer Database between January 2010 and December 2019. Data were analyzed between October 1, 2021, and February 1, 2022. Data from the National Cancer Database included patients diagnosed with nonmetastatic rectal adenocarcinoma receiving surgical treatment at CoC-accredited facilities throughout the US. Exposures: Patient, tumor, and facility-level factors. Facilities were divided by surgical volume, safety-net status, and CoC facility type. Main Outcomes and Measures: Circumferential resection margin missingness rates. Results: A total of 110 571 patients (59.3% men) with rectal adenocarcinoma who underwent curative-intent surgery at 1307 CoC-accredited hospitals were included for analysis. Reporting of CRM improved over the study period, with a mean (SE) missing 12.0% (0.32%) decreased from 16.3% (0.36%). Academic facilities had a higher missingness than other facility types (14.3% vs 10.5%-12.7%; P < .001). Mean (SE) rates of missingness were similar between hospitals of varying volume (lowest quartile: 12.2% [0.93%] vs highest quartile: 12.4% [0.53%]; P = .96). Cases in which fewer than 12 lymph nodes were removed had higher rates of missingness (18.1% vs 11.4%; P < .001). Increased odds of CRM missingness were noted with T category (odds ratio [OR], 1.50; 95% CI, 1.35-1.65) and N category (OR, 2.00; 95% CI, 1.82-2.20). Black race was associated with missingness (OR, 1.13; 95% CI, 1.06-1.14). Conclusion and Relevance: Although CRM positivity reporting has improved over the last decade, the findings of this study suggest there is substantial room for improvement as it becomes a quality standard. Missingness appears to be associated with poor performance on other quality metrics and facility type. This measure appears to be ideal for targeted institution-level feedback to improve quality of care nationally.