Utility of guided FNAC and cell block preparation from liver and gall bladder masses: Learning experience from a tertiary care center.

INTRODUCTION: Ultrasound- and CT-guided fine needle aspiration cytology (FNAC) increases the accessibility of intra-abdominal masses to the liver and gall bladder with the advantages of low cost and high diagnostic yield. Cell block technique has been known for further increasing the diagnostic accuracy. AIMS AND OBJECTIVES: We aimed to study the effectiveness of FNAC and the cell block method in cytological diagnosis of liver and gall bladder masses. We also followed a step-wise approach to increase the success rate. MATERIALS AND METHODS: A 2-year observational study was done from July 2020 to June 2022. Total 80 guided (CT and ultrasound) aspirations were done from space occupying/mass lesions in the liver [74 (92.5%)] and gall bladder [6 (7.5%)], out of which cell blocks by the plasma thrombin method were prepared in 12 cases (15%). The on-site radiological details were noted, and rapid on-site evaluation was done in 65 cases (81.25%). The prepared cytology slides were stained with Papanicolaou, H and E and May-Grunwald Giemsa (MGG) stain. The cytological diagnosis was noted, and the uses and limitations (if any) were observed in each case. A step-wise structured questionnaire format was developed to assist the reporting pathologist so as not to miss out on important diagnostic observations, if present. RESULTS: FNAC in 71 cases (88.7%) gave a conclusive diagnosis. The maximum number of cases were of adenocarcinoma [38 (51.3%)] from the liver followed by hepatocellular carcinoma in 10 cases (13.5%). In gall bladder masses, all 6 cases (100%) were positive for malignancy, out of which 4 cases (66.7%) could be characterized as adenocarcinoma. The cell block preparation was helpful in reaching the diagnosis as well as typing the malignancy in 10 cases (83.3%). The chief limitation observed on conventional cytology smears was inadequate cellularity, which caused inconclusive diagnosis in 9 cases (11.25%). The reporting questionnaire was helpful chiefly in terms of time-efficient reporting in 34 cases (42.5%), increasing the ease and confidence in 69 cases (86.25%) and the advantage of reproducibility of data in all cases (100%) according to the case-by-case evaluation by the reporting pathologists. CONCLUSION: Guided FNAC in conjunction with the cell block technique is extremely helpful in the evaluation of mass lesions of the liver and gall bladder for cytological diagnosis. A proper step-wise approach may be useful to reach a quick and effective diagnosis.

Excited States of apo-Guanidine-III Riboswitch Contribute to Guanidinium Binding through Both Conformational and Induced-Fit Mechanisms.

Riboswitches are mRNA segments that regulate gene expression through conformational changes driven by their cognate ligand binding. The ykkC motif forms a riboswitch class that selectively senses a guanidinium ion (Gdm+) and regulates the downstream expression of proteins which aid in the efflux of excess Gdm+ from the cells. The aptamer domain (AD) of the guanidine-III riboswitch forms an H-type pseudoknot with a triple helical domain that binds a Gdm+. We studied the binding of Gdm+ to the AD of the guanidine (ykkC)-III riboswitch using computer simulations to probe the specificity of the riboswitch to Gdm+ binding. We show that Gdm+ binding is a fast process occurring on the nanosecond time scale, with minimal conformational changes to the AD. Using machine learning and Markov-state models, we identified the excited conformational states of the AD, which have a high Gdm+ binding propensity, making the Gdm+ binding landscape complex exhibiting both conformational selection and induced-fit mechanisms. The proposed apo-AD excited states and their role in the ligand-sensing mechanism are amenable to experimental verification. Further, targeting these excited-state conformations in discovering new antibiotics can be explored.

Positive margin rates for colorectal cancer vary significantly by hospital in Michigan: Can we achieve a 0 % positive margin rate?

Background: High quality surgical care for colorectal cancer (CRC) includes obtaining a negative surgical margin. The Michigan Surgical Quality Collaborative (MSQC) is a statewide consortium of hospitals dedicated to quality improvement; a subset of MSQC hospitals abstract quality of care measures for CRC surgery, including positive margin rate. The purpose of this study was to determine whether positive margin rates vary significantly by hospital, and whether positive margin rates should be a target for quality improvement. Methods: We performed a retrospective cohort study of patients who underwent CRC resection from 2016 to 2020. The primary outcome was the presence of a positive margin. Univariate and multivariable analyses were performed to test the association of positive margins with patient, hospital, and tumor characteristics. Results: The cohort consisted of 4211 patients from 42 hospitals (85 % colon cancer and 15 % rectal cancer). The crude positive margin rate was 6.15 % (95 % CI 4.6-7.4 %); this ranged from 0 % to 22 % at individual hospitals. In multivariable analysis, factors independently associated with positive margins included male sex, underweight BMI, metastatic cancer, rectal cancer (vs. colon), T4 T-stage, N1c/N2 N-stage, and open surgical approach. After adjusting for these factors, there remained significant variation by hospital, with 8 hospitals being statistically-significant outliers. Conclusions: Positive margins rates for CRC vary by hospital in Michigan, even after rigorous adjustment for case-mix. Furthermore, several hospitals achieved near-zero positive margin rates, suggesting opportunities for quality improvement through the identification of best practices among CRC surgery centers.

Allosteric Determinants in High Temperature Requirement A Enzymes Are Conserved and Regulate the Population of Active Conformations.

High temperature requirement A (HtrA) are allosterically regulated enzymes wherein effector binding to the PDZ domain triggers proteolytic activity. Yet, it remains unclear if the inter-residue network governing allostery is conserved across HtrA enzymes. Here, we investigated and identified the inter-residue interaction networks by molecular dynamics simulations on representative HtrA proteases, Escherichia coli DegS and Mycobacterium tuberculosis PepD, in effector-bound and free forms. This information was used to engineer mutations that could potentially perturb allostery and conformational sampling in a different homologue, M. tuberculosis HtrA. Mutations in HtrA perturbed allosteric regulation─a finding consistent with the hypothesis that the inter-residue interaction network is conserved across HtrA enzymes. Electron density from data collected on cryo-protected HtrA crystals revealed that mutations altered the topology of the active site. Ensemble models fitted into electron density calculated from room-temperature diffraction data showed that only a fraction of these models had a catalytically competent active site conformation alongside a functional oxyanion hole thus providing experimental evidence that these mutations influenced conformational sampling. Mutations at analogous positions in the catalytic domain of DegS perturbed the coupling between effector binding and proteolytic activity, thus confirming the role of these residues in the allosteric response. The finding that a perturbation in the conserved inter-residue network alters conformational sampling and the allosteric response suggests that an ensemble allosteric model best describes regulated proteolysis in HtrA enzymes.

Adverse Surgical Outcomes Linked to Co-occurring Smoking and Risky Alcohol Use Among General Surgery Patients.

OBJECTIVE: To assess associations between co-occurring preoperative smoking and risky alcohol use on the likelihood of adverse surgical outcomes. BACKGROUND: Risky alcohol use and smoking are the known surgical risk factors with a high co-occurrence and additive adverse effects on multiple organ systems that impact surgical health, yet no research has evaluated the impact of co-occurrence on surgical outcomes. METHODS: This investigation analyzed 200,816 patients from the Michigan Surgical Quality Collaborative database between July 1, 2012, to December 31, 2018. Patients were classified based on past year risky alcohol use (>2 drink/day) and cigarette smoking into 4 groups: (1) risky alcohol and smoking, (2) risky alcohol only, (3) smoking only, and (4) no risky alcohol/smoking. We fitted logistic regression models, applying propensity score weights incorporating demographic, clinical, and surgical factors to assess associations between alcohol and smoking and 30-day postoperative outcomes; surgical complications, readmission, reoperation, and emergency department (ED) visits. RESULTS: Risky alcohol and smoking, risky alcohol only, and smoking only were reported by 2852 (1.4%), 2840 (1.4%), and 44,042 (22%) patients, respectively. Relative to all other groups, the alcohol and smoking group had greater odds of surgical complications, readmission, and reoperation. Relative to the no alcohol and smoking group, the alcohol only group higher odds of reoperation and smoking only group had higher odds of emergency department visits. CONCLUSIONS: The combination of smoking and risky drinking conferred the highest likelihood of complications, readmission, and reoperation before surgery. Co-occurring alcohol and smoking at the time of surgery warrants special attention as a patient risk factor and deserves additional research.

pH Regulates Ligand Binding to an Enzyme Active Site by Modulating Intermediate Populations.

Understanding the mechanism of ligands binding to their protein targets and the influence of various factors governing the binding thermodynamics is essential for rational drug design. The solution pH is one of the critical factors that can influence ligand binding to a protein cavity, especially in enzymes whose function is sensitive to the pH. Using computer simulations, we studied the pH effect on the binding of a guanidinium ion (Gdm+) to the active site of hen egg-white lysozyme (HEWL). HEWL serves as a model system for enzymes with two acidic residues in the active site and ligands with Gdm+ moieties, which can bind to the active sites of such enzymes and are present in several approved drugs treating various disorders. The computed free energy surface (FES) shows that Gdm+ binds to the HEWL active site using two dominant binding pathways populating multiple intermediates. We show that the residues close to the active site that can anchor the ligand could play a critical role in ligand binding. Using a Markov state model, we quantified the lifetimes and kinetic pathways connecting the different states in the FES. The protonation and deprotonation of the acidic residues in the active site in response to the pH change strongly influence the Gdm+ binding. There is a sharp jump in the ligand-binding rate constant when the pH approaches the largest pKa of the acidic residue present in the active site. The simulations reveal that, at most, three Gdm+ can bind at the active site, with the Gdm+ bound in the cavity of the active site acting as a scaffold for the other two Gdm+ ions binding. These results can aid in providing greater insights into designing novel molecules containing Gdm+ moieties that can have high binding affinities to inhibit the function of enzymes with acidic residues in their active site.

Prevalence and Trends in Smoking Among Surgical Patients in Michigan, 2012-2019.

Importance: Surgery is a teachable moment, and smoking cessation interventions that coincide with an episode of surgical care are especially effective. Implementing these interventions at a large scale requires understanding the prevalence and characteristics of smoking among surgical patients. Objectives: To describe the prevalence of smoking in a population of patients undergoing common surgical procedures and to identify any clinical or demographic characteristics associated with smoking. Design, Setting, and Participants: This cross-sectional study included all adult patients (aged ≥18 years) in a statewide registry who underwent general and vascular surgical procedures from 2012 to 2019 at 70 hospitals in Michigan. Data analysis was conducted from August to October 2020. Exposures: Undergoing a surgical procedure in any of the following categories: appendectomy, cholecystectomy, colon procedures, gastric or esophageal procedures, hepatopancreatobiliary procedures, hernia repair, small-bowel procedures, hysterectomy, vascular procedures, thyroidectomy, and other unspecific abdominal procedures. Main Outcomes and Measures: The prevalence of smoking prior to surgery, defined as cigarette use in the year prior to surgery, obtained from medical record review. Multivariable logistic regression was performed to analyze smoking prevalence based on insurance type and year of surgery while adjusting for demographic and clinical factors, including age, sex, race/ethnicity (determined from the medical record), insurance type, geographic region, comorbidities (ie, hypertension, diabetes, congestive heart failure, chronic obstructive pulmonary disease, chronic steroid use, and obstructive sleep apnea), American Society of Anesthesiologists classification, admission status, surgical priority, procedure type, and year of surgery. Results: From 2012 to 2019, 328 578 patients underwent surgery and were included in analysis. Mean (SD) age was 54.0 (17.0) years, and 197 501 patients (60.1%) were women. The overall prevalence of smoking was 24.1% (79 152 patients). Prevalence varied regionally from 21.5% (95% CI, 21.0%-21.9%; 6686 of 31 172 patients) in southeast Michigan to 28.0% (95% CI, 27.1%-28.9%; 2696 of 9614 patients) in northeast Michigan. When adjusting for clinical and demographic factors, there were greater odds of smoking among patients with Medicaid (odds ratio [OR], 2.75; 95% CI, 2.69-2.82) and patients without insurance (OR, 2.21; 95% CI, 2.10-2.33) compared with patients with private insurance. Among procedure categories, patients undergoing vascular surgery had greater odds of smoking (OR, 3.24; 95% CI, 3.11-3.38) than those undergoing cholecystectomy. Compared with 2012, the adjusted odds of smoking decreased significantly each year (eg, 2019: OR, 0.78; 95% CI, 0.74-0.81). In 2019, the adjusted prevalence of smoking was 22.3% (95% CI, 22.0%-22.7%) among all patients, 43.0% (95% CI, 42.4%-43.6%) among patients with Medicaid, and 36.3% (95% CI, 35.2%-37.4%) among patients without insurance. Conclusions and Relevance: In a statewide population of surgical patients, nearly one-quarter of patients smoked cigarettes, which is higher than the national average. The prevalence of smoking was especially high among patients without insurance and among those receiving Medicaid. Given the established association between undergoing a major surgical procedure and health behavior change, targeted smoking cessation interventions at the time of surgery may be an effective strategy to improve population health, especially among at-risk patient groups.

Synergistic effects of ultrasonication and ethanol washing in controlling the stoichiometry, phase-purity and morphology of rare-earth doped ceria nanoparticles.

Over a period of last thirty years, use of ethanol has been historically reported for obtaining nanopowders with low agglomeration for various oxide systems. In addition to these benefits, we show for the first time that treatments in ethanol medium coupled with an ultrasonication step can impart crucial additional advantages in controlling the phase purity and stoichiometry/composition for such systems. This is an important issue especially for any complex multicationic oxide nanoparticles system and hence we selected one of the most popular catalyst systems of doped-ceria (CeO2) nanoparticles with very high (50%) level of rare-earth (lanthanum) doping for this case study. The effect of an ultrasonication combined ethanol treatment was compared with the other solvent media (pure water and ethanol) without ultrasonication. The underlying mechanism for this process involves lowering the deprotonation rate in ethanol medium which eventually reduces the condensation of the individual metal oxides while the ultrasonication ensures the reproducibility of the synthesis by providing a homogeneous colloidal solution for each washing stages. This novel modification in synthesis of nanoparticles aims to provide meaningful solutions in optimising the phase, composition and morphology of multicationic complex system of nanocrystals.

Improved prediction of missing protein interactome links via anomaly detection.

Interactomes such as Protein interaction networks have many undiscovered links between entities. Experimental verification of every link in these networks is prohibitively expensive, and therefore computational methods to direct the search for possible links are of great value. The problem of finding undiscovered links in a network is also referred to as the link prediction problem. A popular approach for link prediction has been to formulate it as a binary classification problem in which class labels indicate the existence or absence of a link (we refer to these as positive links or negative links respectively) between a pair of nodes in the network. Researchers have successfully applied such supervised classification techniques to determine the presence of links in protein interaction networks. However, it is quite common for protein-protein interaction (PPI) networks to have a large proportion of undiscovered links. Thus, a link prediction approach could incorrectly treat undiscovered positive links as negative links, thereby introducing a bias in the learning. In this paper, we propose to denoise the class of negative links in the training data via a Gaussian process anomaly detector. We show that this significantly reduces the noise due to mislabelled negative links and improves the resulting link prediction accuracy. We evaluate the approach by introducing synthetic noise into the PPI networks and measuring how accurately we can reconstruct the original PPI networks using classifiers trained on both noisy and denoised data. Experiments were performed with five different PPI network datasets and the results indicate a significant reduction in bias due to label noise, and more importantly, a significant improvement in the accuracy of detecting missing links via classification.

Atg23 and Atg27 act at the early stages of Atg9 trafficking in S. cerevisiae.

Atg9 is a conserved multipass transmembrane protein with an essential role in autophagy. In Saccharomyces cerevisiae, it travels through the secretory pathway to a unique compartment, the Atg9 peripheral structures. These structures are then targeted to the phagophore assembly site (PAS), where they are proposed to help deliver membrane to the forming autophagosome. We used 'in vivo reconstitution' of this process in a multiple-knockout strain to define four proteins, Atg11, Atg19, Atg23 and Atg27, as the core minimal machinery necessary and sufficient for the trafficking of Atg9 to the PAS. Atg23 and Atg27 function in the formation of the Atg9 peripheral structures. Overexpression of Atg9 can bypass the need for Atg23, suggesting that the amount of Atg9 in each peripheral structure is a critical factor in their targeting to the PAS. In contrast, overexpression of Atg23 or Atg27 interferes with Atg9 trafficking, suggesting that these proteins must be present in the appropriate stoichiometry in order to function properly. These data allow us to resolve existing controversies regarding the role of Atg23 and Atg27, and propose a model that ties together previous observations regarding the role of Atg9 in autophagosome formation.

Thermosensitive periodontal sol of ciprofloxacin hydrochloride and serratiopeptidase: Pharmaceutical and mechanical analysis.

AIM: The aim of the present work was to explore the development of a dual-controlled release periodontal system of a potent broad spectrum first-generation fluoroquinolone, ciprofloxacin, and the anti-inflammatory enzyme serratiopeptidase (STP). MATERIALS AND METHODS: Based on 3(2) full factorial design, thermoreversible periodontal sols capable of controlled dual delivery of ciprofloxacin hydrochloride and STP were designed using pluronic F127 and carbopol 934P as thermosensitive gelling polymers. Sol gel transition characteristics, %cumulative drug release at 48(th) h and exvivo mucoadhesive strength were designated as dependent responses. The sols were mucoadhesive, syringeable, and inverted into gels at simulated periodontal cavity temperature. RESULTS: F9 with optimal drug release was identified as the best formulation. Extra design check point generated using Design Expert software 8.02 (Stat-Ease, USA) validated the experimental design. Textural analysis revealed that the developed sols were syringeable and spreadable enough for periodontal treatment so it can be expected that hardness and compressibility of sols would pose no problem during clinical application. The in vitro release behavior exhibited controlled release of both cipro HCl and STP (>90% release). CONCLUSION: A dual-controlled release thermoreversible periodontal sol of ciproflaxin and STP was successfully developed. Incorporation of STP as anti-inflammatory agent has the potential of developing a therapeutically efficacious system of cipro HCl for treatment of periodontal inflammatory anaerobic infections.