Dynamic control of toxic natural product biosynthesis by an artificial regulatory circuit.

To mimic the delicately regulated metabolism in nature for improved efficiency, artificial and customized regulatory components for dynamically controlling metabolic networks in multiple layers are essential in laboratory engineering. For this purpose, a novel regulatory component for controlling vanillin biosynthetic pathway was developed through directed evolution, which was responsive to both the product vanillin and substrate ferulic acid, with different capacities. This regulatory component facilitated pathway expression via dynamic control of the intracellular substrate and product concentrations. As vanillin is an antimicrobial compound, low pathway expression and vanillin formation levels enabled better cell growth at an early stage, and the product feedback-activated pathway expression at later stages significantly improved biosynthesis efficiency. This novel multiple-layer dynamic control was demonstrated effective in managing the trade-off between cell growth and production, leading to improved cell growth and vanillin production compared to the conventional or quorum-sensing promoter-controlled pathway. The multiple-layer dynamic control enabled by designed regulatory components responsive to multiple signals shows potential for wide applications in addition to the dynamic controls based on biosynthetic intermediate sensing and quorum sensing reported to date.

Development of a novel uric-acid-responsive regulatory system in Escherichia coli.

A novel uric-acid-responsive regulatory system was developed in Escherichia coli by adapting the HucR-related regulatory elements from Deinococcus radiodurans into E. coli. The induction performance of this system was compared to the performance of both the pBAD and pET systems. Our novel regulatory system was induced in a dose-dependent manner in the presence of uric acid and exhibited low basal expression in its absence. The system was characterized by a wide dynamic range of induction, being compatible with various E. coli strains and not requiring genomic modifications of the bacterial host. E. coli DH5α and DH10B were the most suitable host strains for optimal performance of this system. In conclusion, we developed a regulatory system with potential for applications in both recombinant protein expression and metabolic optimization.

Indobufen alleviates ischemic stroke injury by regulating transcription factor NRF2 and inhibiting ATG5 expression.

BACKGROUND: Ischemic stroke (IS) is a detrimental neurological disease and IS lacks valuable methods to recover body function. Indobufen (IND) could alleviate IS. However, the possible mechanism remains undefined. METHODS: SH-SY5Y cells were cultured under the oxygen-glucose deprivation/reoxygenation (OGD/R) environment and then were treated with small interfering RNA (siRNA) of NRF2 and ATG5. The influence of various concentrations of IND (50 µM, 100 µM, 200 µM, and 400 µM) was determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide. Levels of superoxide dismutase (SOD) and malonaldehyde (MDA) were examined by ELISA. Reactive oxygen species (ROS) production was determined by DCFH-DA staining. The protein levels of LC3II/LC3I, Beclin1, p62, NRF2, and ATG5 were detected by western blot. RESULTS: IND increased cell viability, while depressed the rate of apoptosis in SH-SY5Y cells of OGD/R environment. IND inhibited autophagy by suppressing the levels of LC3II/LC3I, Beclin1 protein, and increasing p62 protein expression in SH-SY5Y cells of OGD/R environment. IND limited the contents of ROS and MDA, while amplifying the activity of SOD in SH-SY5Y cells with OGD/R exposure. IND also promoted NRF2 expression in OGD/R environment. CONCLUSION: IND could inhibit autophagy, oxidative stress, and apoptosis in SH-SY5Y cells with OGD/R exposure, further alleviating IS injury by regulating transcription factor NRF2 and inhibiting ATG5 expression.

The predictive and prognostic value of risk factors in patients receiving hybrid coronary revascularization with postoperative pulmonary complications.

Background: The mortality rate of coronary artery disease ranks first in developed countries, and coronary revascularization therapy is an important cornerstone of its treatment. The postoperative pulmonary complications (PPCs) in patients receiving one-stop hybrid coronary revascularization (HCR) aggravate the dysfunction of multiple organs such as the heart and lungs, therefore increasing mortality. However, the risk factors are still unclear. The objective of this study was to explore the risk factors of PPCs after HCR surgery. Methods: In this study, the perioperative data of 311 patients undergoing HCR surgery were reviewed. All patients were divided into two groups according to whether the PPCs occurred. The baseline information and surgery-related indicators in preoperative laboratory examination, intraoperative fluid management, and anesthesia management were compared between the two groups. Results: Advanced age [odds ratio (OR): 1.065, 95% confidence interval (CI): 1.030-1.101, P<0.001], high body mass index (BMI; OR: 1.113, 95% CI: 1.011-1.225, P=0.02), history of percutaneous coronary intervention (PCI) surgery (OR: 2.831, 95% CI: 1.388-5.775, P=0.004), one-lung volume ventilation (OR: 3.804, 95% CI: 1.923-7.526, P<0.001), inhalation of high concentration oxygen (OR: 3.666, 95% CI: 1.719-7.815, P=0.001), the application of positive end-expiratory pressure (PEEP; OR: 2.567, 95% CI: 1.338-4.926, P=0.005), and long one-lung ventilation time (OR: 1.015, 95% CI: 1.006-1.023, P=0.001) may be risk factors for postoperative PPCs in patients undergoing one-stop coronary revascularization surgery. Using the above seven factors to jointly predict the risk of PPCs in patients undergoing one-stop coronary revascularization surgery, the receiver operating characteristic (ROC) curve showed an area under the curve (AUC) =0.873, 95% CI: 0.835-0.911, sensitivity: 84.81%, and specificity: 75.82%; the predictive model was shown to be effective. Conclusions: Patients undergoing HCR surgery with advanced age, high BMI, a history of PCI surgery, one-lung volume ventilation, inhalation of high concentration oxygen, use of PEEP, and prolonged single lung ventilation are more prone to PPCs.

Effect of Postoperative Prolonged sedation with Dexmedetomidine after successful reperfusion with Endovascular Thrombectomy on long-term prognosis in patients with acute ischemic stroke (PPDET): study protocol for a randomized controlled trial.

BACKGROUND: Endovascular thrombectomy (EVT) is a standard treatment for acute ischemic stroke (AIS) with large vessel occlusion. Hypertension and increased blood pressure variability within the first 24 h after successful reperfusion are related to a higher risk of symptomatic intracerebral hemorrhage and higher mortality. AIS patients might suffer from ischemia-reperfusion injury following reperfusion, especially within 24 h. Dexmedetomidine (DEX), a sedative commonly used in EVT, can stabilize hemodynamics by inhibiting the sympathetic nervous system and alleviate ischemia-reperfusion injury through anti-inflammatory and antioxidative properties. Postoperative prolonged sedation for 24 h with DEX might be a potential pharmacological approach to improve long-term prognosis after EVT. METHODS: This single-center, open-label, prospective, randomized controlled trial will include 368 patients. The ethics committee has approved the protocol. After successful reperfusion (modified thrombolysis in cerebral infarction scores 2b-3, indicating reperfusion of at least 50% of the affected vascular territory), participants are randomly assigned to the intervention or control group. In the intervention group, participants will receive 0.1~1.0 µg/kg/h DEX for 24 h. In the control group, participants will receive an equal dose of saline for 24 h. The primary outcome is the functional outcome at 90 days, measured with the categorical scale of the modified Rankin Scale, ranging from 0 (no symptoms) to 6 (death). The secondary outcome includes (1) the changes in stroke severity between admission and 24 h and 7 days after EVT, measured by the National Institute of Health Stroke Scale (ranging from 0 to 42, with higher scores indicating greater severity); (2) the changes in ischemic penumbra volume/infarct volume between admission and 7 days after EVT, measured by neuroimaging scan; (3) the length of ICU/hospital stay; and (4) adverse events and the all-cause mortality rate at 90 days. DISCUSSION: This randomized clinical trial is expected to verify the hypothesis that postoperative prolonged sedation with DEX after successful reperfusion may promote the long-term prognosis of patients with AIS and may reduce the related socio-economic burden. TRIAL REGISTRATION: ClinicalTrials.gov NCT04916197. Prospectively registered on 7 June 2021.

Relationship among melatonin, postoperative delirium, and postoperative cognitive dysfunction.

BACKGROUND: Postoperative delirium (POD) and postoperative cognitive dysfunction (POCD) are the most common central nervous system dysfunctions during the perioperative period. Melatonin protects nerve cells and impacts cognitive functioning in patients after surgery. METHODS: A total of 120 patients undergoing elective non-cardiac surgery were evaluated with the confusion assessment method (CAM) for diagnosis of POD on the day before and the 1st, 2nd, 3rd, and 7th day after surgery. Also, a neuropsychological test for the diagnosis of POCD was performed on the day before and 1 week after surgery. Patients' urine was collected to examine the concentration of 6-sulfatoxymelatonin (6-SMT), the metabolite of melatonin, with the enzyme-linked immunosorbent assay method. Meanwhile, urine creatinine values were examined to calculate the 6-SMT/creatinine ratio (M/C). RESULTS: The incidence rates of POD and POCD were 7% and 44%, respectively. There were no statistically differences for the M/C on the 1st, 2nd, 3rd, and 7th day after surgery between the POD and the non-POD groups (P>0.05). However, there were statistically significant differences (P<0.05) in the rates of M/C change [(preoperative value-postoperative value)/(postoperative value) ×100%] on the 1st and 7th day after surgery between both groups. Patients were divided into Group I1 (≥100%) and Group II1 (<100%) based on the M/C rate changes on the 1st day, Group I7 (≥200%) and Group II7 (<200%) based on the M/C rate changes on the 7th day, and Group Iw (≥100%) and Group IIw (<100%) based on the M/C rate changes during the 1st week after surgery. The incidence rates of POD for Group I1 and Group II1 were 21.1% and 3.7%, respectively; for Group I7 and Group II7 were 50% and 1.1%, respectively; for Group Iw and Group IIw were 17.2% and 2.8%, respectively. For 7 patients with POD had POCD, the occurrence of POCD was related to POD (P<0.05). CONCLUSIONS: Increased melatonin after surgery may be a risk factor for POD. There may be no correlation between melatonin and POCD. POD may be a risk factor of POCD.

Development and Verification of an Immune-Related Gene Pairs Prognostic Signature in Hepatocellular Carcinoma.

With the increasing prevalence of Hepatocellular carcinoma (HCC) and the poor prognosis of immunotherapy, reliable immune-related gene pairs (IRGPs) prognostic signature is required for personalized management and treatment of patients. Gene expression profiles and clinical information of HCC patients were obtained from the TCGA and ICGC databases. The IRGPs are constructed using immune-related genes (IRGs) with large variations. The least absolute shrinkage and selection operator (LASSO) regression analysis was used to construct IRGPs signature. The IRGPs signature was verified through the ICGC cohort. 1,309 IRGPs were constructed from 90 IRGs with high variability. We obtained 50 IRGPs that were significantly connected to the prognosis and constructed a signature that included 17 IRGPs. In the TCGA and ICGC cohorts, patients were divided into high and low-risk patients by the IRGPs signature. The overall survival time of low-risk patients is longer than that of high-risk patients. After adjustment for clinical and pathological factors, multivariate analysis showed that the IRGPs signature is an independent prognostic factor. The Receiver operating characteristic (ROC) curve confirmed the accuracy of the signature. Besides, gene set enrichment analysis (GSEA) revealed that the signature is related to immune biological processes, and the immune microenvironment status is distinct in different risk patients. The proposed IRGPs signature can effectively assess the overall survival of HCC, and provide the relationship between the signature and the reactivity of immune checkpoint therapy and the sensitivity of targeted drugs, thereby providing new ideas for the diagnosis and treatment of the disease.

Effects of hydroxyethyl starch and gelatin on the risk of acute kidney injury following orthotopic liver transplantation: A multicenter retrospective comparative clinical study.

OBJECTIVES: This multicenter retrospective study aimed to compare the effects of HES and gelatin (GEL) on the risk of post-OLT AKI. METHOD: A total of 1,672 patients undergoing OLT were enrolled from major transplant centers in China between 2005 and 2013. These patients were divided into three groups: GEL, hydroxyethyl starch (HES), and GEL + HES group. RESULTS: There was no significant difference in the incidence of post-OLT AKI among the GEL, HES, and GEL + HES groups. The GEL + HES group had a lower incidence of stage II post-OLT AKI than the other two groups. Compared with patients receiving GEL, patients receiving HES did not harbor an increased risk of AKI. Our results showed that MELD score (adjusted odds ratio [OR], 1.579; 95% confidence interval [CI], 1.123-2.219; P = 0.009) and preoperative anemia (adjusted OR, 1.533; 95% CI, 1.212-1.939; P < 0.001) were independent risk factors for post-OLT AKI, and normal preoperative Scr level (vs abnormal; adjusted OR, 0.402; 95% CI, 0.222-0.729; P = 0.003) was independent protective factors for post-OLT AKI. CONCLUSION: This large-scale multicenter retrospective study found that the intraoperative use of HES did not increase the overall incidence of post-OLT AKI in patients when compared with GEL, and whether to increase the risk of post-OLT AKI needs to be further explored.

Chronic severe hepatitis and preoperative creatinine are independent risk factors for acute kidney injury after liver transplantation.

BACKGROUND: Orthotopic liver transplantation (OLT) offers the highest chance of cure in comparison with all other treatment for liver tumors and other end stage liver disease. However, the complications caused by liver transplantation significantly affect its therapeutic effect, and acute kidney injury (AKI) is one of the most common of these. It is, therefore, necessary to identify the risk factors of AKI after liver transplantation. METHODS: A single-center, retrospective study of patients receiving liver transplantation at the Beijing Chao-Yang Hospital between January 2015 to January 2019 was conducted.Patients were divided into a normal control group and AKI group based on their previous medical history. Preoperative and intraoperative indicators including preoperative creatinine, uric acid, and the intraoperative input of protein were then recorded. RESULTS: A total of 419 patients were enrolled into the study. The control group consisted of 336 patients while 83 patients formed an AKI group based on the grading criteria of AKI. There were significant differences in chronic severe hepatitis (P=0.001), liver cancer (P=0.044), intraoperative input of sodium bicarbonate (P=0.019), input of red blood cell suspension (P=0.004), the input of blood plasma (P=0.043), intraoperative urine output (P=0.006), and preoperative creatinine (P=0.041) between the control and AKI group. Multivariate analysis indicated that chronic severe hepatitis (OR: 2.872; P=0.003) and preoperative creatinine (OR: 1.083; P=0.011) were independent risk factors for AKI in patients receiving liver transplantation. CONCLUSIONS: Chronic severe hepatitis and preoperative creatinine may be potential risk factors for the occurrence of AKI after liver transplantation.

Effect of Tropisetron on Prevention of Emergence Delirium in Patients After Noncardiac Surgery: A Trial Protocol.

Importance: Postoperative delirium is a frequent disorder for patients undergoing surgery and is associated with poor outcomes. Delirium may occur in the immediate period after anesthesia administration and surgery. Tropisetron, which is frequently administrated for postoperative nausea and vomiting, is also a partial agonist of α7 nicotinic acetylcholine receptors associated with neuroprotective effects. Tropisetron may be the potential pharmacological treatment to decrease delirium after noncardiac surgery. Objective: To perform a randomized clinical trial to determine the efficacy and safety of tropisetron for prevention of emergence delirium in patients undergoing noncardiac surgery. Design, Setting, and Participants: This single-center, 2-arm randomized, double-blind, placebo-controlled trial will include 1508 patients undergoing noncardiac surgery. The intervention group will receive 5 mg of intravenous tropisetron before anesthesia induction, and patients in the control group will receive a placebo. The primary end point is the incidence of emergence delirium within 1 hour after tracheal tube removal, measured by the Confusion Assessment Method for the Intensive Care Unit score. The main secondary outcome is the incidence of postoperative delirium measured at 3 days of follow-up. An intention-to-treat principle will be used for all analyses. Discussion: Delirium remains the most common neuropsychiatric complication for patients after surgery. This will be the first randomized clinical study to evaluate whether tropisetron is effective in preventing emergence delirium. Results from this study will provide evidence for alteration of daily practice. Trial Registration: ClinicalTrials.gov Identifier: NCT04027751.

Design and application of a novel high-throughput screening technique for 1-deoxynojirimycin.

High-throughput screening techniques for small molecules can find intensive applications in the studies of biosynthesis of these molecules. A sensitive, rapid and cost-effective technique that allows high-throughput screening of endogenous production of the natural iminosugar 1-deoxynojirimycin (1-DNJ), an α-glucosidase inhibitor relevant to the pharmaceutical industry, was developed in this study, based on the inhibitory effects of 1-DNJ on the activity of the ß-glycosidase LacS from Sulfolobus solfataricus. This technique has been demonstrated effective in engineering both the key enzyme and the expression levels of enzymes in the 1-DNJ biosynthetic pathway from Bacillus atrophaeus cloned in E. coli. Higher biosynthetic efficiency was achieved using directed evolution strategies.