Charged multivesicular body protein 2B ameliorates biliary injury in the liver from donation after cardiac death rats via autophagy with air-oxygenated normothermic machine perfusion.

Normothermic machine perfusion (NMP) could provide a curative treatment to reduce biliary injury in donation after cardiac death (DCD) donor livers; however, the underlying mechanisms remain poorly understood. In a rat model, our study compared air-oxygenated NMP to hyperoxygenated NMP and found that air-oxygenated NMP improved DCD functional recovery. Here, we found that the charged multivesicular body protein 2B (CHMP2B) expression was substantially elevated in the intrahepatic biliary duct endothelium of the cold-preserved rat DCD liver after air-oxygenated NMP or in biliary endothelial cells under hypoxia/physoxia. CHMP2B knockout (CHMP2B-/-) rat livers showed increased biliary injury after air-oxygenated NMP, indicated by decreased bile production and bilirubin level, elevated biliary levels of lactate dehydrogenase and gamma-glutamyl transferase. Mechanically, we demonstrated that CHMP2B was transcriptionally regulated by Kruppel-like transcription factor 6 (KLF6) and alleviated biliary injury through decreasing autophagy. Collectively, our results suggested that air-oxygenated NMP regulates CHMP2B expression through the KLF6, which reduces biliary injury by inhibiting autophagy. Targeting the KLF6-CHMP2B autophagy axis may provide a solution to reducing biliary injury in DCD livers undergoing NMP.

Cytochrome P450 2E1 predicts liver functional recovery from donation after circulatory death using air-ventilated normothermic machine perfusion.

The optimal oxygen concentration is unclear for normothermic machine perfusion (NMP) of livers from donation after circulatory death (DCD). Our purposes were to investigate the effect of air-ventilated NMP on the DCD liver, analyze the underlying mechanism and select the targets to predict liver functional recovery with NMP. NMP was performed using the NMP system with either air ventilation or oxygen ventilation for 2 h in the rat liver following warm ischemia and cold-storage preservation. Proteomics and metabolomics were used to reveal the significant molecular networks. The bioinformation analysis was validated by administering peroxisome proliferator activator receptor-γ (PPARγ) antagonist and agonist via perfusion circuit in the air-ventilated NMP. Results showed that air-ventilated NMP conferred a better functional recovery and a less inflammatory response in the rat DCD liver; integrated proteomics and metabolomics analysis indicated that intrahepatic docosapentaenoic acid downregulation and upregulation of cytochrome P450 2E1 (CYP2E1) expression and activity were associated with DCD liver functional recovery with air-ventilated NMP; PPARγ antagonist worsened liver function under air-oxygenated NMP whereas PPARγ agonist played the opposite role. In conclusion, air-ventilated NMP confers a better liver function from DCD rats through the DAP-PPARγ-CYP2E1 axis; CYP2E1 activity provides a biomarker of liver functional recovery from DCD.

Mode of action and efficacy of quinolinic acid for the control of Ceratocystis fimbriata on sweet potato.

BACKGROUND: Ceratocystis fimbriata is a hazardous fungal pathogen able to cause black rot disease on sweet potato. The management of C. fimbriata strongly relies on the use of toxic fungicides, and there is a lack of efficient alternative strategies. RESULTS: The antifungal properties of quinolinic acid (QA) were studied for the first time, indicating that QA shows selective antifungal activity against C. fimbriata. QA inhibited completely the mycelial growth of C. fimbriata at less than 0.8 mg mL-1 concentration (pH 4), and was able to produce alterations in the fungal cell wall, and to impede spore agglutination and mycelium formation. QA significantly reduced the concentration of ergosterol, and was able to associate to iron (II), suggesting that QA may be a lanosterol 14-α demethylase inhibitor. In preventive applications, QA reduced the disease incidence of C. fimbriata on sweet potato by 75%, achieving higher control efficacy in comparison with commercial fungicides prochloraz and carbendazim. CONCLUSIONS: The first selective antifungal agent against C. fimbriata was discovered in this work, and showed suitable antifungal properties for the management of black rot disease. © 2021 Society of Chemical Industry.

Effects of chitinase-3-like protein 1 on brain death-induced hepatocyte apoptosis via PAR2-JNK-caspase-3.

Hepatocyte apoptosis is a crucial factor affecting liver quality in brain-dead donors. The identification of key molecular proteins involved in brain-death (BD)-induced hepatocyte apoptosis may help determine an effective method for improving the quality of livers from brain-dead donors. In this study, we used in vivo and in vitro models to investigate the role of chitinase-3-like protein 1 (CHI3L1) in promoting liver cell apoptosis after BD. Chitin was used to inhibit CHI3L1 in a rat model of BD. Macrophage polarization of THP-1 cells and hypoxia/reoxygenation (H/R) of LO-2 cells were used to mimic BD-induced cell stress in liver. We found that CHI3L1 played a vital role in promoting liver cell apoptosis. Six hours after BD, CHI3L1 expression was significantly upregulated in liver macrophages and was associated with BD-induced M1 polarization of these cells. In liver cells cultured under H/R conditions, recombinant CHI3L1 activated the protease-activated receptor 2 (PAR2)/c-June N-terminal kinase (JNK) apoptotic pathway and aggravated apoptosis. Compared with the control group, chitin particles inhibited the expression of CHI3L1 in the liver of brain dead rats, thereby reducing activation of the hepatocyte surface receptor, PAR2, and its downstream JNK/caspase-3 signaling pathway, ultimately reducing hepatocyte apoptosis. In conclusion, our results indicate that CHI3L1 relies on a PAR2/JNK-mediated mechanism to promote BD-induced hepatocyte apoptosis.

Hepatic connective tissue growth factor expression and regulation differ between non-steatotic and non-alcoholic steatotic livers from brain-dead donor.

Accurate evaluation of liver steatosis is required from brain-dead donors (BDDs) with nonalcoholic fatty liver disease (NAFLD). Our purposes were to investigate expression and regulation of connective tissue growth factor (CTGF) expression in livers from human and rat after brain death, and further evaluate its potential application. NAFLD and brain death models were established in rats. LX2 cells were cultured under hypoxia/reoxygenation. CTGF protein and mRNA levels were measured in liver samples from BDDs of human and rat by immunohistochemistry and reverse transcription-quantitative polymerase chain reaction. YAP-regulated CTGF expression was investigated in LX2 cells via YAP small interfering RNA and Verteporfin treatment. Blood CTGF level from BDDs was measured by enzyme-linked immunosorbent assay. After brain death, CTGF, transforming growth factor-ß and YAP were overexpressed in non-alcoholic steatotic liver, whereas CTGF was downregulated in non-steatotic liver. Time-series analysis revealed that CTGF and YAP expression was comparable, as confirmed by inhibited YAP expression in LX2 cells. CTGF level and NAFLD activity were linearly correlated. CTGF expression and regulation differ between non-steatosis and nonalcoholic steatosis livers from BDDs. CTGF may be an important factor to evaluate graft quality from BDDs with NAFLD.

Microbiologic assessment of flexible gastrointestinal endoscope reprocessing using a pump-assisted sampling technique: an investigation involving all endoscopy units in Tianjin, China.

BACKGROUND: Microbiologic surveillance of flexible gastrointestinal endoscopes is recommended in several guidelines as the primary means of identifying reprocessing failures. This study aimed to evaluate the contamination level and prevalence of bacteria of post-reprocessing endoscopes and to access whether using a pump-assisted sampling method (PASM) improves the sensitivity of culture. METHODS: All 59 endoscopy units in Tianjin, China, were investigated. The PASM and the conventional flushing sampling method (CFSM) were used to compare the results of the microbial culture. Logistic regression analysis was used to identify the influencing factors. RESULTS: One hundred four (56.52%) flushing channel samples of gastrointestinal endoscopes were positive for culture, and the maximum bacterial concentration was 14,100 colony-forming units (CFU)/channel. One hundred fifty-one (82.07%) flushing samples were qualified according to the national standard of China (≤ 20 CFU/channel). The qualified rate of the samples collected by PASM was significantly lower than the qualified rate by CFSM (65.52% vs 89.68%). Using PASM (odds ratio [OR]: 4.257; 95% confidence interval [CI]: 1.870-9.690) would increase the sensitivity of culture. The use of purified water (OR: 0.288; 95% CI: 0.102-0.814) could reduce the risk of endoscope reprocessing failure. CONCLUSION: Many endoscopes fail to meet the national standard for microbial culture after reprocessing. Our results suggest that using a pump-assisted method could increase the sensitivity of the test.

[Seroepidemiology investigation of neutralizing antibody against enterovirus 71 among healthy people in Tianjin].

OBJECTIVE: To investigate the latent infection caused by enterovirus 71 (EV71) among healthy people in Tianjin and to provide evidence on prevention and control hand-food and mouth diseases (HFMD). METHODS: 1611 sera specimens were collected from healthy people in Tianjin while EV71 antibody was detected by neutralization test, and then the results were analyzed statistically. RESULTS: For determining positivity, the cut-point was set at 1:4. The positive rate was 66.79% (1076/1611) for EV71 neutralizing antibody. The lowest positive rate was 32.71% in the 0 - 5 age group while the highest rate was 76.67% in the 16 - 25 age group. Significant difference was seen in the positive rates among different age groups. The lowest positive rate (59.05%) was seen in the city areas while the highest rate (72.35%) was seen in the surrounding counties. 5.71% of the people being tested showed their neutralizing antibody as ≥ 1:256. The difference was statistically significant on positive rates among different areas. We constructed logistic regression models with the EV71 neutralizing antibody positive rate as the dependent variable and age, sex, floating population, area etc. as independent variables. There appeared statistical significances in all the independent variables. CONCLUSION: Age seemed a risk factor for recessive infection of EV71, and the neutralizing antibody against EV71 might not be kept permanently. In order to prevent and control the HFMD, more attention should be paid to the areas where more floating population were resided.