Novel nitroxoline derivative combating resistant bacterial infections through outer membrane disruption and competitive NDM-1 inhibition.

ABSTRACTNew Delhi metallo-ß-lactamase-1 (NDM-1) has rapidly disseminated worldwide, leading to multidrug resistance and worse clinical prognosis. Designing and developing effective NDM-1 inhibitors is a critical and urgent challenge. In this study, we constructed a library of long-lasting nitroxoline derivatives and identified ASN-1733 as a promising dual-functional antibiotic. ASN-1733 can effectively compete for Ca2+ on the bacterial surface, causing the detachment of lipopolysaccharides (LPS), thereby compromising the outer membrane integrity and permeability and exhibiting broad-spectrum bactericidal activity. Moreover, ASN-1733 demonstrated wider therapeutic applications than nitroxoline in mouse sepsis, thigh and mild abdominal infections. Furthermore, ASN-1733 can effectively inhibit the hydrolytic capability of NDM-1 and exhibits synergistic killing effects in combination with meropenem against NDM-1 positive bacteria. Mechanistic studies using enzymatic experiments and computer simulations revealed that ASN-1733 can bind to key residues on Loop10 of NDM-1, hindering substrate entry into the enzyme's active site and achieving potent inhibitory activity (Ki = 0.22 µM), even in the presence of excessive Zn2+. These findings elucidate the antibacterial mechanism of nitroxoline and its derivatives, expand their potential application in the field of antibacterial agents and provide new insights into the development of novel NDM-1 inhibitors.

Leukocyte telomere length independently predicts hyperuricemia risk in a longitudinal study of the Chinese population.

BACKGROUND AND AIMS: Leukocyte telomere length (LTL) has been correlated with uric acid levels, although results are inconsistent, and prospective studies are lacking. In this longitudinal, prospective cohort study, we aimed to assess whether a shorter LTL predicts the risk of hyperuricemia. METHODS AND RESULTS: We conducted a longitudinal study in a Chinese cohort of 599 participants. Of these, 266 participants completed a 5.9-year follow-up from June 2014 to December 2021. LTL was assessed at baseline using real-time polymerase chain reaction. Hyperuricemia was defined as serum uric acid ≥420 mmol/L according to Chinese guidelines for diagnosis and treatment of hyperuricemia and gout. Participants who had developed hyperuricemia during follow-up (n = 17) had shorter LTL at baseline. Baseline LTL was independently associated with the development of hyperuricemia at follow-up after adjusting for conventional hyperuricemia risk factors (odds ratio [OR] 2.347 [95% confidence interval [CI] 1.123, 4.906]; P = 0.023). After grouping according to LTL tertiles, the incidence of hyperuricemia was 18.334-fold higher for the first than for the third tertile (OR 18.334 [95%CI 1.786, 191.272]; P = 0.014, P for trend = 0.050). CONCLUSION: Our findings in a prospective cohort suggest that LTL could predict hyperuricemia risk, which might inform the timely prevention and treatment of hyperuricemia.

Tumor necrosis factor-alpha mediates the negative association between telomere length and kidney dysfunction.

Aim/hypothesis: The relationship between peripheral blood leukocyte telomere length (LTL) and kidney dysfunction, especially in people with hypertension, remains unclear. No clinical study has explored the role of inflammation and oxidative stress in the relationship between LTL and kidney dysfunction. Therefore, we examined the relationship between baseline LTL and albuminuria progression and/or rapid renal function decline in Chinese patients with or without hypertension and investigated whether inflammation and oxidative stress played a mediating role in this relationship. Methods: We conducted a prospective study including 262 patients in a 7-year follow-up period from 2014 to 2021. Data on LTL, inflammation, oxidative markers, renal function, and urine protein levels were assessed. Kidney dysfunction was defined as either albuminuria progression, rapid decline in renal function, or the composite endpoint (albuminuria progression and rapid decline in renal function). Logistic regression and simple mediation models were used for the analysis. Results: In this cohort (mean age, 54.3±9.7 years; follow-up period, 5.9±1.1 years), 42(16.0%), 21(8.0%), and 59(22.5%) patients developed albuminuria progression, rapid eGFR decline, and the composite endpoint of kidney dysfunction, respectively. Logistic regression analysis showed that each standard deviation decrease of baseline LTL and the lower quartile (Q) of baseline LTL were significantly correlated with an increased risk of rapid decline in renal function (OR=1.83 [95% CI 1.07, 3.27] per 1SD, P=0.03; OR=7.57 [95% CI 1.25, 145.88] for Q1 vs. Q4, P for trend=0.031); and the composite endpoint of kidney dysfunction (OR=1.37 [95% CI 0.97, 1.96] per 1SD, borderline positive P=0.072; OR=2.96[95% CI 1.15, 8.2] for Q1 vs. Q4, P for trend=0.036). The mediating analysis showed that tumor necrosis factor (TNF)-a partly mediated the relationship between LTL and rapid decline in renal function (direct effect: ß=0.046 95%CI [0.006, 0.090],P=0.02; indirect effect: ß=0.013 95%CI [0.003, 0.020]), and the mediating proportion was 22.4%.In subgroup analyses, LTL was inversely associated with rapid decline in renal function or the composite endpoint of kidney dysfunction only in patients with hypertension (OR=49.07[3.72,211.12] vs.1.32[0.69,2.58] per 1SD, P for interaction=0.045;OR=3.10 [1.48, 7.52] vs.1.08[0.92,1.63] per 1SD, P for interaction=0.036). Conclusion: Baseline LTL could independently predict kidney dysfunction at follow-up, especially in participants with hypertension. TNF-a partially mediated the negative association between LTL and kidney dysfunction.

Pharmacokinetics and Pharmacodynamics of a Novel Vancomycin Derivative LYSC98 in a Murine Thigh Infection Model Against Staphylococcus aureus.

Introduction: LYSC98 is a novel vancomycin derivative used for gram-positive bacterial infections. Here we compared the antibacterial activity of LYSC98 with vancomycin and linezolid in vitro and in vivo. Besides, we also reported the pharmacokinetic/pharmacodynamic (PK/PD) index and efficacy-target values of LYSC98. Methods: The MIC values of LYSC98 were identified through broth microdilution method. A mice sepsis model was established to investigate the protective effect of LYSC98 in vivo. Single-dose pharmacokinetics of LYSC98 was studied in thigh-infected mice and liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was used to determine LYSC98 concentration in plasma. Dose fractionation studies were performed to evaluate different PK/PD indices. Two methicillin-resistant Staphylococcus aureus (MRSA) clinical strains were used in the dose ranging studies to determine the efficacy-target values. Results: LYSC98 showed a universal antibacterial effect in Staphylococcus aureus with a MIC range of 2-4 µg/mL. In vivo, LYSC98 demonstrated distinctive mortality protection in mice sepsis model with an ED50 value of 0.41-1.86 mg/kg. The pharmacokinetics results displayed maximum plasma concentration (Cmax) 11,466.67-48,866.67 ng/mL, area under the concentration-time curve from 0 to 24 h (AUC0-24) 14,788.42-91,885.93 ng/mL·h, and elimination half-life (T1/2) 1.70-2.64 h, respectively. Cmax/MIC (R 2 0.8941) was proved to be the most suitable PK/PD index for LYSC98 to predict its antibacterial efficacy. The magnitude of LYSC98 Cmax/MIC associated with net stasis, 1, 2, 3 and 4 - log 10 kill were 5.78, 8.17, 11.14, 15.85 and 30.58, respectively. Conclusion: Our study demonstrates that LYSC98 is more effective than vancomycin either in killing vancomycin-resistant Staphylococcus aureus (VRSA) in vitro or treating S. aureus infections in vivo, making it a novel and promising antibiotic. The PK/PD analysis will also contribute to the LYSC98 Phase I dose design.

Engineered Artificial Membraneless Organelles in Saccharomyces cerevisiae To Enhance Chemical Production.

Microbial cell factories provide a green and sustainable opportunity to produce value-added products from renewable feedstock. However, the leakage of toxic or volatile intermediates decreases the efficiency of microbial cell factories. In this study, membraneless organelles (MLOs) were reconstructed in Saccharomyces cerevisiae by the disordered protein sequence A-IDPs. A regulation system was designed to spatiotemporally regulate the size and rigidity of MLOs. Manipulating the MLO size of strain ZP03-FM, the amounts of assimilated methanol and malate were increased by 162 % and 61 %, respectively. Furthermore, manipulating the MLO rigidity in strain ZP04-RB made acetyl-coA synthesis from oxidative glycolysis change to non-oxidative glycolysis; consequently, CO2 release decreased by 35 % and the n-butanol yield increased by 20 %. This artificial MLO provides a strategy for the co-localization of enzymes to channel C1 starting materials into value-added chemicals.

Hypoglycemia Caused by Exogenous Insulin Antibody Syndrome: A Large Single-Center Case Series From China.

CONTEXT: Exogenous insulin antibody syndrome (EIAS) can lead to unexpected and potentially life-threatening recurrent hypoglycemia. OBJECTIVE: We aimed to better define autoimmune hypoglycemia caused by EIAS in patients with diabetes and shed light on the improvements in the identification and intervention for this rare but possibly life-threatening condition. METHODS: We summarized the clinical characteristics of autoimmune hypoglycemia caused by EIAS in 23 patients with diabetes. Furthermore, we performed human leukocyte antigen (HLA) genotyping of 10 patients. RESULTS: We identified a high frequency of autoimmune comorbidities (21.7%), food or drug allergy (48%), insulin allergy (30%), lipodystrophy at the insulin injection sites (22%), and antinuclear antibodies (25%) in the patients. Alternation between hyperglycemia and hypoglycemia was observed in more than 90% of the patients. Most patients showed a high insulin autoantibody titer (>90%) and inappropriately increased insulin concentration (insulin/C-peptide molar ratio >7, >85%). We detected similar frequencies of DRB1*0405-DQB1*0401 and DRB1*0901-DQB1*0303 compared with previously reported frequencies in type 1 diabetes, and a lower frequency of DRB1*0406 compared with insulin autoimmune syndrome. The spontaneous remission rate exceeded 70%. CONCLUSION: Predisposing factors for autoimmune hypoglycemia caused by EIAS include a strong autoimmune background. Susceptible HLA genotypes for type 1 diabetes or insulin autoimmune syndrome might not explain susceptibility to this condition. Additionally, insulin autoantibodies and the insulin/C-peptide molar ratio are reliable screening options. The prognosis for this condition is favorable. Monitoring of insulin and insulin autoantibodies may contribute to treatment effectiveness.

Dietary patterns, oxidative Stress, inflammation and biological variation in hemoglobin A1c: Association and Mediation analysis in a rural community in north China.

OBJECTIVE: The aim is to assess the relationship between the hemoglobin glycation index(HGI) and dietary patterns, and investigates whether inflammation and oxidative stress mediate the relationship. METHODS: Cross-sectional data were collected from 453 dwellers in a Chinese rural community. Diet was assessed using 24 h food recalls. Based on the energy intake ratio from three macronutrients, dietary patterns were identified by cluster analysis. The HGI was defined as the observed HbA1c minus predicted HbA1c. Indicators of inflammation and oxidative stress were assessed. RESULT: 3 dietary patterns were clustered, namely "fat(n = 100)", "balance(n = 186)" and "carbohydrate(n = 167)". The fat dietary patterns had lower HGI than the other two dietary patterns. TNFα was higher in the carbohydrate dietary pattern. Linear regression analysis suggested that the carbohydrate dietary pattern was correlated with higher HGI levels(ß = 0.204,95 %CI(0.071,0.338)), compared with the fat dietary pattern. The relationship disappeared after accounting for biomarkers of inflammation and oxidative stress. Mediation analyses indicated that TNFα might explain for 19.15 % effects of the carbohydrate dietary pattern on HGI, compared with the fat dietary pattern. CONCLUSION: The carbohydrate dietary pattern had positive associations with HGI and TNFα. TNFα partly mediated the relationship between dietary patterns and HGI.

The optimal diagnostic criteria of endogenous hyperinsulinemic hypoglycemia based on a large cohort of Chinese patients.

Purpose: An end-of-fast insulin level ≥ 3 µIU/ml, C-peptide level ≥ 0.6 ng/ml, and proinsulin level ≥ 5 pmol/l with end-of-fast glucose level ≤ 3.0 mmol/l have been established as the criteria for endogenous hyperinsulinemic hypoglycemia. However, all these criteria have been proposed based on patients in Western populations. This study aimed to determine the optimal criteria using a large series of Chinese patients. Methods: This retrospective study comprised 144 patients with surgically proven insulinoma and 40 controls who underwent a 72-h fasting test at the Peking Union Medical College Hospital(PUMCH) from 2000 to 2020. Receiver operating characteristic curves were used for analysis. Results: In this series of patients, the optimal diagnostic criteria for endogenous hyperinsulinemic hypoglycemia were insulin ≥ 5.5 µIU/ml, C-peptide ≥ 0.7 ng/ml, and proinsulin ≥ 12 pmol/l with end-of-fast glucose ≤ 2.8 mmol/l; the sensitivity and specificity were 99% and 100% for insulin, 100% and 100% for C-peptide, and 93% and 100% for proinsulin, respectively. The diagnostic efficacy of the criteria based on Western populations was then tested. The sensitivity and specificity of end-of-fast insulin ≥ 3 µIU/ml, C-peptide ≥ 0.6 ng/ml, and proinsulin ≥ 5 pmol/l with end-of-fast glucose ≤ 3.0 mmol/l were 100% and 83%, 100% and 80%, and 97% and 78%, respectively. Conclusions: New and optimized diagnostic criteria for endogenous hyperinsulinemic hypoglycemia in Chinese populations have been proposed, and these criteria yield satisfactory accuracy.

Engineering microbial cell viability for enhancing chemical production by second codon engineering.

Microbial cell factories offer a promising strategy for the sustainable production of industrial chemicals from renewable biomass feedstock. However, their performance is often limited by poor microbial cell viability (MCV). Here, MCV was engineered to enhance chemical production by optimizing the regulation of lifespan-specific genes to reduce the accumulation of reactive oxygen species (ROS). In Escherichia coli, MCV was improved by reducing ROS accumulation using second codon engineering to regulate hypoxia-inducible transcription factor (arcA), resulting in lysine production up to 213 g L-1 with its productivity 5.90 g L-1·h-1. In Saccharomyces cerevisiae, MCV was increased by decreasing ROS accumulation using second codon engineering to fine-tune ceramide synthase (lag1), leading to glucaric acid production up to 9.50 g L-1 with its productivity 0.057 g L-1·h-1. These results demonstrate that engineering MCV is a potential strategy to boost the performance of microbial cell factories in industrial processes.

Enhancing biofuels production by engineering the actin cytoskeleton in Saccharomyces cerevisiae.

Saccharomyces cerevisiae is widely employed as a cell factory for the production of biofuels. However, product toxicity has hindered improvements in biofuel production. Here, we engineer the actin cytoskeleton in S. cerevisiae to increase both the cell growth and production of n-butanol and medium-chain fatty acids. Actin cable tortuosity is regulated using an n-butanol responsive promoter-based autonomous bidirectional signal conditioner in S. cerevisiae. The budding index is increased by 14.0%, resulting in the highest n-butanol titer of 1674.3 mg L-1. Moreover, actin patch density is fine-tuned using a medium-chain fatty acid responsive promoter-based autonomous bidirectional signal conditioner. The intracellular pH is stabilized at 6.4, yielding the highest medium-chain fatty acids titer of 692.3 mg L-1 in yeast extract peptone dextrose medium. Engineering the actin cytoskeleton in S. cerevisiae can efficiently alleviate biofuels toxicity and enhance biofuels production.

The Role of Lactate Exercise Test and Fasting Plasma C-Peptide Levels in the Diagnosis of Mitochondrial Diabetes: Analysis of Clinical Characteristics of 12 Patients With Mitochondrial Diabetes in a Single Center With Long-Term Follow-Up.

Objective: The aim of this study was to analyze the clinical characteristics and the pattern of long-term changes of fasting plasma C-peptide in patients with mitochondrial diabetes (MD), and to provide guidance for the diagnosis and treatment of MD. Methods: We retrieved MD patients with long-term follow-up at Peking Union Medical College Hospital from January 2005 to July 2021 through the medical record retrieval system and retrospectively analyzed their clinical data, biochemical parameters, fasting plasma C-peptide, glycosylated hemoglobin and treatment regimens. Non-parametric receiver operating characteristic (ROC) curves were used to assess the relationship between exercise test-related plasma lactate levels and suffering from MD. Results: A total of 12 MD patients were included, with clinical characteristics of early-onset, normal or low body weight, hearing loss, maternal inheritance, and negative islet-related autoantibodies. In addition, patients with MD exhibited significantly higher mean plasma lactate levels immediately after exercise compared to patients with type 1 diabetes mellitus (T1DM) (8.39 ± 2.75 vs. 3.53 ± 1.47 mmol/L, p=0.003) and delayed recovery time after exercise (6.02 ± 2.65 vs. 2.17 ± 1.27 mmol/L, p=0.011). The optimal cut-off points identified were 5.5 and 3.4 mmol/L for plasma lactate levels immediately after and 30 minutes after exercise, respectively. The fasting plasma C-peptide levels decreased as a negative exponential function with disease progression (Y= 1.343*e-0.07776X, R2 = 0.4154). Treatment regimens in MD patients were varied, with no metformin users and a weak correlation between insulin dosage and body weight. Conclusions: The increased level of plasma lactate during exercise or its delayed recovery after exercise would contribute to the diagnosis of MD. Changes of fasting plasma C-peptide in MD patients over the course of the disease indicated a rapid decline in the early stages, followed by a gradual slowing rate of decline.

High Hemoglobin Glycation Index Is Associated With Telomere Attrition Independent of HbA1c, Mediated by TNFα.

CONTEXT: The hemoglobin glycation index (HGI) is correlated with metabolic diseases and inflammation. Whether the HGI is associated with the aging process and how inflammation and oxidative stress affect the relationship remain unclear. OBJECTIVE: We aimed to analyze links between the HGI and aging biomarkers, and to explore a potential role of inflammation and oxidative stress in the correlations. METHODS: A cross-sectional study of 434 subjects with different glucose intolerances in a rural community was enrolled. The HGI was calculated as the difference between the measured and predicted hemoglobin A1c (HbA1c). The population was categorized into tertiles of the HGI. Telomere length (LTL) and mitochondrial DNA copy number (mtDNAcn) determined by polymerase chain reaction assay. Tumor necrosis factor (TNF) α and interleukin (IL) 6, 8-oxo-2'-deoxyguanosine (8-oxo-dG), superoxide dismutase (SOD) activities, and glutathione reductase (GR) were measured. RESULTS: Participants in the high HGI group were older and reported a shorter LTL, higher levels of TNFα, SOD activities, and HbA1c. Correlation analyses demonstrated that HGI was correlated with LTL (r = -0.25, P < .001) and TNFα (r = 0.19, P < .001) regardless of HbA1c levels. No relationship was found between HGI and mtDNAcn. HGI (ß = -0.238, 95% CI -0.430, -0.046, P = .015) and TNFα (ß = -0.02, 95% CI -0.030, -0.014, P < .001) were proved to be correlated with LTL independently, using multiple linear regression analysis. Ordinal logistic regression models showed that compared with subjects the high HGI group, the possibilities of a higher-level LTL was 5.29-fold in the low HGI group (OR 5.29, 95% CI (2.45, 11.41), P < .001), 2.41-fold in the moderate HGI group (OR 2.41, 95% CI 1.35, 4.30, P = .003) after controlling for confounding variables. Mediation analyses indicated that TNFα accounted for 30.39% of the effects of the HGI on LTL. CONCLUSION: HGI was negatively related to telomere attrition, independent of HbA1c. TNFα acted as a mediator of the relationship between HGI and LTL.