Ketogenic Diets Alter the Gut Microbiome Resulting in Decreased Intestinal Th17 Cells.

Very low-carbohydrate, high-fat ketogenic diets (KDs) induce a pronounced shift in metabolic fuel utilization that elevates circulating ketone bodies; however, the consequences of these compounds for host-microbiome interactions remain unknown. Here, we show that KDs alter the human and mouse gut microbiota in a manner distinct from high-fat diets (HFDs). Metagenomic and metabolomic analyses of stool samples from an 8-week inpatient study revealed marked shifts in gut microbial community structure and function during the KD. Gradient diet experiments in mice confirmed the unique impact of KDs relative to HFDs with a reproducible depletion of bifidobacteria. In vitro and in vivo experiments showed that ketone bodies selectively inhibited bifidobacterial growth. Finally, mono-colonizations and human microbiome transplantations into germ-free mice revealed that the KD-associated gut microbiota reduces the levels of intestinal pro-inflammatory Th17 cells. Together, these results highlight the importance of trans-kingdom chemical dialogs for mediating the host response to dietary interventions.

Caloric restriction disrupts the microbiota and colonization resistance.

Diet is a major factor that shapes the gut microbiome1, but the consequences of diet-induced changes in the microbiome for host pathophysiology remain poorly understood. We conducted a randomized human intervention study using a very-low-calorie diet (NCT01105143). Although metabolic health was improved, severe calorie restriction led to a decrease in bacterial abundance and restructuring of the gut microbiome. Transplantation of post-diet microbiota to mice decreased their body weight and adiposity relative to mice that received pre-diet microbiota. Weight loss was associated with impaired nutrient absorption and enrichment in Clostridioides difficile, which was consistent with a decrease in bile acids and was sufficient to replicate metabolic phenotypes in mice in a toxin-dependent manner. These results emphasize the importance of diet-microbiome interactions in modulating host energy balance and the need to understand the role of diet in the interplay between pathogenic and beneficial symbionts.

An Integrated Hepatocyte Stability Assay for Simultaneous Metabolic Stability Assessment and Metabolite Profiling.

The determination of metabolic stability is critical for drug discovery programs, allowing for the optimization of chemical entities and compound prioritization. As such, it is common to perform high-volume in vitro metabolic stability experiments early in the lead optimization process to understand metabolic liabilities. Additional metabolite identification experiments are subsequently performed for a more comprehensive understanding of the metabolic clearance routes to aid medicinal chemists in the structural design of compounds. Collectively, these experiments require extensive sample preparation and a substantial amount of time and resources. To overcome the challenges, a high-throughput integrated assay for simultaneous hepatocyte metabolic stability assessment and metabolite profiling was developed. This assay platform consists of four parts: 1) an automated liquid-handling system for sample preparation and incubation, 2) a liquid chromatography and high-resolution mass spectrometry-based system to simultaneously monitor the parent compound depletion and metabolite formation, 3) an automated data analysis and report system for hepatic clearance assessment; and 4) streamlined autobatch processing for software-based metabolite profiling. The assay platform was evaluated using eight control compounds with various metabolic rates and biotransformation routes in hepatocytes across three species. Multiple sample preparation and data analysis steps were evaluated and validated for accuracy, repeatability, and metabolite coverage. The combined utility of an automated liquid-handling instrument, a high-resolution mass spectrometer, and multiple streamlined data processing software improves the process of these highly demanding screening assays and allows for simultaneous determination of metabolic stability and metabolite profiles for more efficient lead optimization during early drug discovery. SIGNIFICANCE STATEMENT: Metabolic stability assessment and metabolite profiling are pivotal in drug discovery to fully comprehend metabolic liabilities for chemical entity optimization and lead selection. Process of these assays can be repetitive and resource demanding. Here, we developed an integrated hepatocyte stability assay that combines automation, high-resolution mass spectrometers, and batch-processing software to improve and combine the workflow of these assays. The integrated approach allows simultaneous metabolic stability assessment and metabolite profiling, significantly accelerating screening and lead optimization in a resource-effective manner.

Safety and feasibility of laparoscopic liver resection for intrahepatic cholangiocarcinoma: a propensity score-matched study.

BACKGROUND: Laparoscopic liver resection (LLR) is controversial in treating intrahepatic cholangiocarcinoma (ICC). Therefore, this study aimed to evaluate the safety and feasibility of LLR for the treatment of ICC and explored the independent factors affecting the long-term prognosis of ICC. METHODS: We included 170 patients undergoing hepatectomy for ICC from December 2010 to December 2021 and divided them into LLR group and open liver resection (OLR) group. We used propensity score matching (PSM) analysis to reduce the impact of data bias and confounding variables and then compared the short-term and long-term prognosis of LLR and OLR in treating ICC; Cox proportional hazards regression model was adopted to explore the independent factors affecting the long-term prognosis of ICC. RESULTS: A total of 105 patients (70 in the LLR group and 35 in the OLR group) were included after 2:1 PSM analysis. There was no difference in demographic characteristics and preoperative indexes between the two groups. The perioperative results of the OLR group were worse than those of the LLR group, that is, the intraoperative blood transfusion rate (24 (68.6) vs 21 (30.0)), blood loss (500 (200-1500) vs 200 (100-525)), and the morbidity of major postoperative complications (9 (25.7) vs 6 (8.5)) in the OLR group were worse than those in LLR group. LLR could enable patients to obtain an equivalent long-term prognosis compared to OLR. The Cox proportional hazards regression model exhibited that no matter before or after PSM, preoperative serum CA12-5 and postoperative hospital stay were independent factors affecting overall survival, while only lymph node metastasis independently influenced recurrence-free survival. CONCLUSIONS: Compared with ICC treated by OLR, the LLR group obtained superior perioperative period outcomes. In the long run, LLR could enable ICC patients to receive an equivalent long-term prognosis compared to OLR. In addition, ICC patients with preoperative abnormal CA12-5, lymph node metastasis, and more extended postoperative hospital stay might suffer from a worse long-term prognosis. However, these conclusions still need multicenter extensive sample prospective research to demonstrate.

Impact of Dental Institutions' Values on Students' Intent to Practice in Underserved Communities: A Cross-sectional Study.

AIM: To evaluate how an institution's values can impact students' intent to practice in underserved areas. MATERIALS AND METHODS: The values of 71 accredited dental schools in the United States were searched and tabulated. The American Dental Education Association (ADEA) survey results were used to tabulate the desire to serve the underserved population before entering dental school and upon graduation. Additionally, responses on the total amount of educational debt on graduation was compiled for LLUSD and all other dental schools. Fisher's exact test was performed to compare the difference between the two cohorts and Wilcoxon test was used to assess difference within the groups. Tests were conducted at an alpha level of 0.05 with SAS v 9.1.3 (SAS Institute, Cary, NC, USA). RESULTS: The top five values of US dental institutions based on frequency were excellence, diversity/inclusion, integrity, innovation, and respect. There was no statistically significant difference between LLUSD and all other dental schools in their desire to serve the underserved community upon graduation (p > 0.05, in all instances). Overall, there was a trend that upon graduation, the desire to serve the underserved had less impact on students' decision-making on their career choices. This drop in the desire to serve the underserved was statistically significant within both cohorts in the year 2021 (p < 0.001). CONCLUSIONS: Dental institutions should focus on better understanding of how their values impact their students' career choices so that they can develop strategies to better align their values with the mission of addressing the dentist shortage in underserved areas. CLINICAL SIGNIFICANCE: Dental institutions' values play a major role in impacting students' career choices upon graduation and should be assessed using metrics that are measurable.

High-throughput protein binding assay using cross-sample pooling in combination with high-resolution mass spectrometry.

RATIONALE: The fraction of unbound drugs (ƒu ) is a useful pharmacokinetic parameter in understanding drug disposition (Absorption, Distribution, Metabolism, Excretion), pharmacological activity and toxicity. Therefore, protein binding assays are frequently performed in drug development, creating a high demand for biological, experimental and analytical resources. Our work aims to increase binding assay throughput and comprehensiveness, while reducing biological and experimental consumption without compromising data quality by introducing cross-pooling and cassetting procedures, followed by a rapid and informative high-resolution mass spectrometry (HRMS) analysis. METHODS: Individual drugs were spiked into a test matrix and incubated in a rapid equilibrium dialysis device. After incubation, a cross-pooling procedure was performed, in which the samples of one drug were equalized with the complementary matrix provided from a different drug. The same drugs were also assayed with a conventional method, in which samples were equalized with the newly prepared complementary matrix. Cross-pooled samples were further cassetted to increase throughput. The samples were analyzed by high-performance liquid chromatography coupled with an Orbitrap HRMS, and the fu values were calculated and compared between the cross-pooling and conventional sampling procedures. RESULTS: Highly comparable human plasma fu values of 27 drugs representing different chemical classes and wide-ranging fu values were obtained by conventional and cross-pooling procedures, The tight correlation was further validated in other species (rat, mouse) and matrices (microsomes, brain). In addition, the cassetted samples showed highly consistent fu values compared to their noncassetted counterparts. Moreover, HRMS analysis not only showed highly consistent and repeatable quantification results compared to the "gold standard" triple quadrupole (QqQ) analysis, but also demonstrated outstanding advantage over QqQ in enabling a high-throughput, informative and versatile analysis. CONCLUSIONS: This work demonstrates that the cross-pooling procedure with further sample cassetting using HRMS is experimentally and analytically feasible to allow a higher throughput (increased by up to 8-fold), resource-effective (reducing matrix consumption by 50%, minimizing time spent on method development and platemap design), analytically dependable (accurate quantification), and versatile (metabolite elucidation and low recovery troubleshooting) analysis.

Efficacy of laparoscopic repeat hepatectomy compared with open repeat hepatectomy: a single-center, propensity score matching study.

INTRODUCTION: Laparoscopic repeat hepatectomy (LRH) is considered to be a technically challenging procedure which has not been widely applied. This study aimed to assess the accessibility and security of LRH for patients with hepatic tumor recurrence. METHODS: Between January 2010 and October 2020, we performed 48 LRHs and 31 open repeat hepatectomies (ORHs) for recurrent liver cancer. LRHs were matched to ORHs (1:1) using propensity score matching (PSM) created by comparing preoperative factors. The perioperative data of patients were retrospectively analyzed, including baseline data, operative time, intraoperative blood loss, pathology, days of postoperative stay, complication morbidity, and mortality within 30 days. Overall survival and recurrence-free survival rates with appropriate follow-up were obtained to evaluate the long-term outcomes. RESULTS: Compared with the ORH, LRH was related with shorter operative duration (169.9 versus 232.9 ml, p < 0.01), less intraoperative bleeding (100.0 versus 500.0 ml, p < 0.01), lower rate of blood transfusion (8.3% versus 58.1%, p < 0.01), and shorter hospitalization (5.0 versus 11.0 days, p < 0.01). The median follow-up was 31 months. The LRH 1-, 3-, and 5-year overall survival were 77.1%, 61.6%, and 46.2% versus 82.3%, 66.5%, and 29.5% for ORH (p = 0.77). The 1-, 3-, and 5-year disease-free survival rates of the two groups were 73.4%, 62.0%, and 44.3% versus 66.1%, 44.1%, and 14.7%, respectively (p = 0.22). CONCLUSIONS: Laparoscopic repeated hepatectomy is safe and practicable with great short-term results for selected patients.

Safety and feasibility of laparoscopic liver resection for hepatocellular carcinoma with clinically significant portal hypertension: a propensity score-matched study.

BACKGROUND: The presence of clinically significant portal hypertension (CSPH) remains a relative contraindication to liver resection for patients with resectable hepatocellular carcinoma (HCC). The goal of this study was to explore whether a laparoscopic approach could extend the indications for hepatectomy to patients with PH. METHOD: Patients who underwent laparoscopic liver resection (LLR) from February 2016 to September 2019 performed by a single medical team were included in this study. We analyzed the surgical and oncological outcomes between groups with and without CSPH before and after propensity score matching (PSM). RESULT: We enrolled 156 patients divided into two groups according to the presence (CSPH, n = 26) or absence (non-CSPH, n = 130) of CSPH. CSPH group was associated with more clinical signs of liver dysfunction (p < 0.05). After PSM (n = 48 patients), the CSPH group tended to have a longer postoperative hospital stay (p = 0.054); however, there was no difference in operation time (p = 0.329), blood loss volume (p = 0.392), transfusion rates (p = 0.701), rate of conversion to open surgery (p = 0.666), surgical margin (p = 0.306), surgical mortality (n = 0), or comprehensive complication index (p = 0.844) between the two groups. The median follow-up time for the entire cohort was 19.6 months (range 0.2-40.6 months). The 3-year overall survival rate was 62.9% in the CSPH group and 84.3% in the non-CSPH group (p = 0.1090), and results were similar after PSM (p = 0.5734). CONCLUSIONS: LLR is safe and feasible for HCC with PH. The introduction of minimally invasive surgery, represented by LLR, can appropriately expand the indications for hepatectomy.

Comparative assessment for rat strain differences in metabolic profiles of 14 drugs in Wistar Han and Sprague Dawley hepatocytes.

Knowledge of inter-strain and inter-gender differences in drug metabolism studies is important for animal selection in pharmacokinetic and toxicological studies. The effects of rat strain and gender in in vitro metabolism were investigated in Sprague Dawley (SD) and Wister Han (WH) rats based on the hepatocyte metabolic profiles of 14 small molecule drugs. Similarities were found between the hepatocyte metabolic clearances of SD and WH strains, suggesting that only one strain can be confidently used for the evaluation of hepatic clearance. Neither strain of rat was preferable over the other to cover human metabolites. Higher similarities in metabolic pathways were found between the same gender than the same strain. Differences in metabolite identities, metabolite formation rates and potential biotransformation pathways were observed between SD and WH rat strains. Eleven metabolites from six drugs were "disproportionally" formed between SD and WH rats. The use of a specific rat strain model and gender for ADME and toxicity testing should, therefore, be carefully considered as metabolic profiles may differ, even though metabolic clearance was similar between SD and WH rats.

Safety and effectiveness evaluation of a two-handed technique combining harmonic scalpel and laparoscopic Peng's multifunction operative dissector in laparoscopic hemihepatectomy.

OBJECTIVES: This study was designed to evaluate the safety and effectiveness of a two-hand technique combining harmonic scalpel (HS) and laparoscopic Peng's multifunction operative dissector (LPMOD) in patients who underwent laparoscopic hemihepatectomy (LHH). METHODS: We designed and conducted a case-control study nested in a prospectively collected laparoscopic liver surgery database. Patients who underwent LHH for liver parenchyma transection using HS + LPMOD were defined as cases (n = 98) and LPMOD only as controls (n = 47) from January 2016 to May 2018. Propensity score matching (1:1) between the case and control groups was used in the analyses. RESULTS: The case group had significantly less intraoperative blood loss in milliliters (169.4 ± 133.5 vs. 221.5 ± 176.3, P = 0.03) and shorter operative time in minutes (210.5 ± 56.1 vs. 265.7 ± 67.1, P = 0.02) comparing to the control group. The conversion to laparotomy, postoperative hospital stay, resection margin, the mean peak level of postoperative liver function parameters, bile leakage rate, and others were comparable between the two groups. There was no perioperative mortality. CONCLUSIONS: We demonstrated that the two-handed technique combing HS and LPMOD in LHH is safe and effective which is associated with shorter operative time and less intraoperative blood loss compared with LPMOD alone. The technique facilitates laparoscopic liver resection and is recommended for use.

Differentiation and management of hepatobiliary mucinous cystic neoplasms: a single centre experience for 8 years.

BACKGROUND: Hepatobiliary mucinous cystic neoplasms (H-MCNs) are relatively rare cystic neoplasms in the liver. The differential diagnosis of H-MCNs remains big challenging, and the management and prognosis between the hepatic simple cyst (HSC) and H-MCNs are quite different. This study aimed to present our experience in the management of H-MCNs and provide a preoperative H-MCNs risk prediction nomogram to differentiating H-MCNs from liver cystic lesions. METHODS: 29 patients diagnosed with H-MCNs and 75 patients diagnosed with HSC between June 2011 and June 2019 at Zhejiang University School of medicine, Sir Run-Run Shaw Hospital were reviewed in this study. We analyzed the demographic and clinicopathological variables. RESULTS: US, CT, and MRI could accurately diagnose only 3.4%, 46.1%, and 57.1% of H-MCNs, respectively. After univariate analysis and multivariate logistic regression analysis, the variables significantly associated with H-MCNs were enhancement after contrast (p = 0.009), tumour located in the left lobe (p = 0.02) and biliary ductal dilation (p = 0.027). An H-MCNs risk predictive nomogram was constructed, which showed excellent discrimination (areas under the receiver operating characteristic curve were 0.940) and consistent calibration between the predicted probability and actual probability. CONCLUSION: Among patients with H-MCNs, the location of the tumour, enhancement in CT scan, and biliary duct dilation are significantly independent risk factors. The appropriate treatment of H-MCNs is radical resection. Using our Nomogram could facilitate screening and identification of patients with liver cystic lesions.

Laparoscopic anatomical portal territory hepatectomy using Glissonean pedicle approach (Takasaki approach) with indocyanine green fluorescence negative staining: how I do it.

BACKGROUND: Laparoscopic anatomical resection (LAR) is a highly challenging procedure. This study aimed to describe our experience of the LAR with an indocyanine green fluorescence negative staining (ICGNS) by the Glissonean pedicle transection (Takasaki) approach. METHODS: From April 2017 to December 2019, 43 consecutive patients underwent LAR with ICGNS strategy in our medical team. The details of the ICGNS strategy were described. The demographic and clinicopathological data of the included patients were retrospectively analyzed. RESULTS: The extent of resections included right hemihepatectomy (n = 12), left hemihepatectom (n = 4), left lateral sectionectomy (n = 3), Right anterior sectionectomy (n = 3), Right posterior sectionectomy (n = 6), central hepatectomy (n = 2), single anterolateral segmentectomy (n = 5), single posterosuperior segmentectomy (n = 6), and bisegmentectomy (n = 2). The mean operation time was 212 ± 53 min, and the median estimated blood loss was 200 (100-300) ml. The overall complication rate was 30.2% (grade I, 14%; grade II, 14%; grade III, 2.3%). The median duration of postoperative hospital stay was 6 (4-7) days. CONCLUSION: ICGNS is a safe and feasible LAR strategy that greatly facilitates selecting the liver transection plane, although its benefits need to be verified by large-sample comparative studies.

Laparoscopic repeat hepatectomy for treating recurrent liver cancer.

BACKGROUND: Laparoscopic repeat hepatectomy (LRH) is a technically challenging procedure, so LRH for recurrent liver cancer has not been widely accepted. The aim of this study was to perform a systematic review of the current literature to identify and evaluate available data of LRH for recurrent hepatocellular carcinoma (rHCC) and metastases tumour of liver, especially of colorectal liver metastases (CRLM), focusing on the safety and feasibility. METHODS: A comprehensive search of the PubMed database was performed for all studies published in English evaluating LRH for rHCC and recurrent metastases tumour of liver from 1st January, 2005 to 1st June, 2019. RESULTS: A total of 15 studies which comprised 444 patients and reported outcomes for the efficacy and safety of LRH in the treatment of rHCC or CRLM were included in the present review. Moreover, nine studies compared the perioperative outcomes of LRH versus open repeat hepatectomy (ORH). LRH was superior to ORH with reduced blood loss, shorter operative time, shorter hospital stay and lower morbidity rates. CONCLUSIONS: LRH can safely performed in rHCC or CRLM patients with cirrhosis, previous open hepatectomy, multiple recurrent lesions and tumours located in difficult posterosuperior segments.

Ursodeoxycholic Acid (UDCA) Mitigates the Host Inflammatory Response during Clostridioides difficile Infection by Altering Gut Bile Acids.

Clostridioides difficile infection (CDI) is associated with increasing morbidity and mortality posing an urgent threat to public health. Recurrence of CDI after successful treatment with antibiotics is high, thus necessitating discovery of novel therapeutics against this enteric pathogen. Administration of the secondary bile acid ursodeoxycholic acid (UDCA; ursodiol) inhibits the life cycles of various strains of C. difficilein vitro, suggesting that the FDA-approved formulation of UDCA, known as ursodiol, may be able to restore colonization resistance against C. difficilein vivo However, the mechanism(s) by which ursodiol is able to restore colonization resistance against C. difficile remains unknown. Here, we confirmed that ursodiol inhibits C. difficile R20291 spore germination and outgrowth, growth, and toxin activity in a dose-dependent manner in vitro In a murine model of CDI, exogenous administration of ursodiol resulted in significant alterations in the bile acid metabolome with little to no changes in gut microbial community structure. Ursodiol pretreatment resulted in attenuation of CDI pathogenesis early in the course of disease, which coincided with alterations in the cecal and colonic inflammatory transcriptome, bile acid-activated receptors nuclear farnesoid X receptor (FXR) and transmembrane G-protein-coupled membrane receptor 5 (TGR5), which are able to modulate the innate immune response through signaling pathways such as NF-κB. Although ursodiol pretreatment did not result in a consistent decrease in the C. difficile life cycle in vivo, it was able to attenuate an overly robust inflammatory response that is detrimental to the host during CDI. Ursodiol remains a viable nonantibiotic treatment and/or prevention strategy against CDI. Likewise, modulation of the host innate immune response via bile acid-activated receptors FXR and TGR5 represents a new potential treatment strategy for patients with CDI.

Laparoscopic Anatomical Portal Territory Hepatectomy with Cirrhosis by Takasaki's Approach and Indocyanine Green Fluorescence Navigation (with Video).

BACKGROUND: Anatomical liver resection has shown advantages in the treatment of hepatocellular carcinoma (HCC).1 Pure laparoscopic hepatectomy for some deep lesions remains challenging, especially for anatomical resection.2 Because of many kinds of hepatic venous variations, resection along the hepatic vein may not be a "real" anatomical resection. We used a three-dimensional visualization technique to construct a portal territory model which represented the patient-specific anatomy. During the operation, the territory was visualized by indocyanine green (ICG) navigation. PATIENT: A 48-year-old man was admitted to our institution with a single hepatic mass of 4.5 cm in segment 7. The patient suffered hepatitis B related cirrhosis and portal hypertension. METHODS: A resection plan was put forward by 3-D visualization technique in advance (Fig. 1a). The patient was placed in a supine position with pillows underneath the upper right semi-lateral body. The position of the trocar is shown in Fig. 1b. After removal of the gallbladder and overhang of the G6, the G7 was dissected and ligated by Takasaki's Glissonean pedicle approach (Fig. 1c).3 The ischemic line appeared and was consistent with the demarcation line of portal territory (Fig. 1d). A parenchyma transection was performed along the boundary of the unstained side of the ICG fluorescence. Fig. 1 Some important images from the video. a The trocar position of this laparoscopic surgery. Operator-1 or -2: the first and the second trocar for the operator; assistant: the trocar for the assistant; operator/assistant: the trocar can be used by operator and assistant interchangeably; scope: the trocar for the laparoscope. b The transection plan constructed by preoperative 3-D visualization technique. The blue area was the tumor-bearing portal territory and targeted part of the liver that should be removed. c The intrahepatic anterior and posterior Glissonean pedicle of G6 and G7. G6: glissonean pedicle of segment 6; G7: glissonean pedicle of segment 7. d The ischemic line, the demarcation line of portal territory staining by ICG and the target territory constructed by 3-D visualization technique before the operation RESULT: The operation time was 205 min, the estimated blood loss was 150 ml. With no postoperative complications, the patient was discharged on the fourth day. Hepatocellular carcinoma was confirmed in histopathology. The resection margin was free of tumor involvement. CONCLUSION: A preoperative 3-D visualization technique combined with intraoperative ICG fluorescence navigation could facilitate a precise and safe laparoscopic anatomical hepatectomy.

Correction to: Laparoscopic Anatomical Portal Territory Hepatectomy with Cirrhosis by Takasaki's Approach and Indocyanine Green Fluorescence Navigation (with Video).

In the original version of the article all of the authors' first and last names were transposed. The original article has been updated.

Bioactivation of α,ß-Unsaturated Carboxylic Acids Through Acyl Glucuronidation.

After oral administration to monkeys of [14C]GDC-0810, an α,ß-unsaturated carboxylic acid, unchanged parent and its acyl glucuronide metabolite, M6, were the major circulating drug-related components. In addition, greater than 50% of circulating radioactivity in plasma was found to be nonextractable 12 hours post-dose, suggesting possible covalent binding to plasma proteins. In the same study, one of the minor metabolites was a cysteine conjugate of M6 (M11) that was detected in plasma and excreta (urine and bile). The potential mechanism for the covalent binding to proteins was further investigated using in vitro methods. In incubations with glutathione (GSH) or cysteine (5 mM), GSH and cysteine conjugates of M6 were identified, respectively. The cysteine reaction was efficient with a half-life of 58.6 minutes (k react = 0.04 1/M per second). Loss of 176 Da (glucuronic acid) followed by 129 Da (glutamate) in mass fragmentation analysis of the GSH adduct of M6 (M13) suggested the glucuronic acid moiety was not modified. The conjugation of N-glucuronide M4 with cysteine in buffer was >1000-fold slower than with M6. Incubations of GDC-0810, M4, or M6 with monkey or human liver microsomes in the presence of NADPH and GSH did not produce any oxidative GSH adducts, and the respective substrates were qualitatively recovered. In silico analysis quantified the inherent reactivity differences between the glucuronide and its acid precursor. Collectively, these results show that acyl glucuronidation of α,ß-unsaturated carboxylic acids can activate the compound toward reactivity with GSH, cysteine, or other biologically occurring thiols and should be considered during the course of drug discovery. SIGNIFICANCE STATEMENT: Acyl glucuronidation of the α,ß-unsaturated carboxylic acid in GDC-0810 activates the conjugated alkene toward nucleophilic addition by glutathione or other reactive thiols. This is the first example that a bioactivation mechanism could lead to protein covalent binding to α,ß-unsaturated carboxylic acid compounds.

How gut microbiome interactions affect nutritional traits of Drosophila melanogaster.

Most research on the impact of the gut microbiome on animal nutrition is designed to identify the effects of single microbial taxa and single metabolites of microbial origin, without considering the potentially complex network of interactions among co-occurring microorganisms. Here, we investigated how different microbial associations and their fermentation products affect host nutrition, using Drosophila melanogaster colonized with three gut microorganisms (the bacteria Acetobacter fabarum and Lactobacillus brevis, and the yeast Hanseniaspora uvarum) in all seven possible combinations. Some microbial effects on host traits could be attributed to single taxa (e.g. yeast-mediated reduction of insect development time), while other effects were sex specific and driven by among-microbe interactions (e.g. male lipid content determined by interactions between the yeast and both bacteria). Parallel analysis of nutritional indices of microbe-free flies administered different microbial fermentation products (acetic acid, acetoin, ethanol and lactic acid) revealed a single consistent effect: that the lipid content of both male and female flies is reduced by acetic acid. This effect was recapitulated in male flies colonized with both yeast and A. fabarum, but not for any microbial treatment in females or males with other microbial complements. These data suggest that the effect of microbial fermentation products on host nutritional status is strongly context dependent, with respect to both the combination of associated microorganisms and host sex. Taken together, our findings demonstrate that among-microbe interactions can play a critically important role in determining the physiological outcome of host-microbiome interactions in Drosophila and, likely, in other animal hosts.

A novel simple intra-corporeal Pringle maneuver for laparoscopic hemihepatectomy: how we do it.

INTRODUCTION: To prevent and control hemorrhage is the key to successfully perform laparoscopic hemihepatectomy (LHH). Pringle's maneuver (PM) is the standard hepatic inflow occlusion technique. Our study was to describe a novel simple way to perform totally intra-corporeal laparoscopic PM and to explore the feasibility of combining PM and selective hemihepatic vascular occlusion technique in LHH. METHODS: We extracted and analyzed the data of patients who consecutively underwent LHH to validate this new surgery technique. Between January, 2016 and December, 2017, 34 patients were included. Data of pre-operation, operation and post-operation were collected, including some demographic data, operative time, operative blood loss, transfusion rate, hepatic hilum occlusion rate and time, pathologic results, short-term complication, and postoperative hospitalization days. RESULTS: Only one patient (3.0%) in our series required conversion to laparotomy as a result of the severe adhesion. The average operative time was 216.9 ± 60.3 min. The mean hepatic inflow occlusion time was 25.3 ± 14.5 min. The average estimated blood loss was 192.9 ± 152.2 ml. All patients received R0 resection. CONCLUSION: The novel hepatic inflow occlusion device is a safe reliable and convenient technique for LHH that is associated with favorable perioperative outcomes and low risk of conversion.

Microbiota fermentation-NLRP3 axis shapes the impact of dietary fibres on intestinal inflammation.

OBJECTIVE: Diets rich in fermentable fibres provide an array of health benefits; however, many patients with IBD report poor tolerance to fermentable fibre-rich foods. Intervention studies with dietary fibres in murine models of colonic inflammation have yielded conflicting results on whether fibres ameliorate or exacerbate IBD. Herein, we examined how replacing the insoluble fibre, cellulose, with the fermentable fibres, inulin or pectin, impacted murine colitis resulting from immune dysregulation via inhibition of interleukin (IL)-10 signalling and/or innate immune deficiency (Tlr5KO). DESIGN: Mice were fed with diet containing either cellulose, inulin or pectin and subjected to weekly injections of an IL-10 receptor (αIL-10R) neutralising antibody. Colitis development was examined by serological, biochemical, histological and immunological parameters. RESULTS: Inulin potentiated the severity of αIL10R-induced colitis, while pectin ameliorated the disease. Such exacerbation of colitis following inulin feeding was associated with enrichment of butyrate-producing bacteria and elevated levels of caecal butyrate. Blockade of butyrate production by either metronidazole or hops ß-acids ameliorated colitis severity in inulin-fed mice, whereas augmenting caecal butyrate via tributyrin increased colitis severity in cellulose containing diet-fed mice. Elevated butyrate levels were associated with increased IL-1ß activity, while inhibition of the NOD-like receptor protein 3 by genetic, pharmacologic or dietary means markedly reduced colitis. CONCLUSION: These results not only support the notion that fermentable fibres have the potential to ameliorate colitis but also caution that, in some contexts, prebiotic fibres can lead to gut dysbiosis and surfeit colonic butyrate that might exacerbate IBD.