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Yu-Qin Yan,1 Yu-Qing Huang,2 Ying-Qing Feng2 1Department of Cardiology, People’s Hospital of Shenzhen Baoan District, Shenzhen, People’s Republic of China; 2Department of Cardiology, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, Guangzhou, People’s Republic of ChinaCorrespondence: Yu-Qin Yan, Department of Cardiology, People’s Hospital of Shenzhen Baoan District, No. 118, Longjing Second Road, Xinan Street, Baoan District, Shenzhen, 518100, People’s Republic of China, Tel/Fax +86-755-27788311, Email graceyuqin@163.comBackground: The Great Chinese Famine, as the famine of 1959– 1961 was often known. Famine exposure during early life was proven to be associated with some kidney diseases but has not been studied with kidney stone. We aimed to investigate the relationship between exposure to the Great Chinese Famine in early life and the incidence of kidney stone in adulthood.Methods: From 1 January 2017 to 31 December 2018, a total of 19,658 eligible adults were recruited in a cross-sectional survey who were born between 1 October 1952 and 30 September 1964 in Guangdong, China. Participants were separated into kidney stone and none-kidney stone groups based on kidney stone status. According to birth data, participants were divided into non-exposed, fetal-exposed, early-, mid-, and late-childhood-exposed groups. Multivariate logistic regression, subgroup analysis and interaction test were used to estimate the odds ratios (ORs) and confidence intervals (CIs) between famine exposure and kidney stone.Results: In total, 19,658 (12,246 female, mean age 59.31 ± 3.68 years) subjects were enrolled, and 3219 (16.38%) participants with kidney stone. The prevalence of kidney in none-, fetal-, early-, mid-, and late-childhood-exposed groups were 645 (14.9%), 437 (15.9%), 676 (16.3%), 743 (17.0%), and 718 (17.6%), respectively (P< 0.001). When compared with the unexposed group, the fully adjusted ORs for kidney stone from fetal-exposed, early-, mid- to late-childhood-exposed groups were 1.37 (95% CI: 1.13, 1.68, P=0.002), 1.98 (95% CI: 1.45, 2.72, P< 0.001), 2.94 (95% CI: 1.96, 4.42, P< 0.001), and 3.48 (95% CI: 2.11, 5.72, P< 0.001), respectively (P for trend< 0.001). Subgroup analyses revealed no interactions between the famine effect on kidney stones and body mass index, gender, smoking status, history of diabetes or hypertension (all P for interaction > 0.05).Conclusion: This study found that exposure to the Great Chinese Famine during early life was independently associated with the increased incidence of kidney stone in adulthood.Keywords: Great Chinese Famine, early life, famine exposure, adulthood, kidney stone

Chao-Lei Chen,1,* Lin Liu,1,* Jia-Yi Huang,1 Yu-Ling Yu,1 Kenneth Lo,1,2 Yu-Qing Huang,1 Ying-Qing Feng1 1Department of Cardiology, Guangdong Cardiovascular Institute, Guangdong Provincial Key Laboratory of Coronary Heart Disease Prevention, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, School of Medicine, South China University of Technology, Guangzhou 510080, People’s Republic of China; 2Centre for Global Cardiometabolic Health, Department of Epidemiology, School of Public Health, Brown University, Providence, RI, USA*These authors contributed equally to this workCorrespondence: Yu-Qing Huang; Ying-Qing FengDepartment of Cardiology,Guangdong Cardiovascular Institute, Guangdong Provincial People’s Hospital, Guangdong Provincial Key Laboratory of Coronary Heart Disease Prevention, Guangdong Academy of Medical Sciences, School of Medicine, South China University of Technology, Guangzhou 510080, People’s Republic of ChinaTel/ Fax +86-20-83827812Email hyq513@126.com; 651792209@qq.comBackground: The optimal blood pressure (BP) level for diabetic patients remains controversial, while studies provided limited evidence on BP management for individuals with normoglycemia or prediabetes. We aimed to investigate the associations between systolic blood pressure (SBP) and all-cause and cardiovascular (CVD) mortality among US adults with different glycemic profiles.Methods: We used data from the 1999– 2014 National Health and Nutrition Examination Survey (NHANES, n=40,046) with comprehensive baseline examination and follow-up assessment. Restricted cubic spline analysis was performed to examine dose–response relationship between SBP and mortality risk. Cox regression models were used to estimate hazard ratios of all-cause mortality and CVD mortality for SBP categories.Results: U-shaped associations between SBP and all-cause mortality were observed regardless of glucose status. The relationship between SBP and CVD mortality was found to be U-shaped only in normoglycemic participants. The lowest mortality risk of optimal SBP (mmHg) by group was 115– 120 (normoglycemia), 120– 130 (prediabetes), and 125– 135 (diabetes). When compared with the reference group, SBP < 100 mmHg was significantly associated with 49% (HR=1.49, 95% CI: 1.13– 1.96), 57% (1.57, 1.07– 2.3), and 59% (1.59, 1.12– 2.25) higher all-cause mortality risk in people with normoglycemia, prediabetes, and diabetes, respectively. The multivariable-adjusted HRs of all-cause mortality for SBP ≥ 150 mmHg were 1.51 (1.25– 1.82), 1.56 (1.27– 1.93), and 1.33 (1.08– 1.64), respectively. As for CVD mortality, the multivariable-adjusted HRs were 2.85 (1.29– 6.33) for SBP < 100 mmHg and 2.71 (1.56– 4.69) for SBP ≥ 150 mmHg in normoglycemia; HR was 1.66 (1.05– 2.63) for SBP ≥ 150 mmHg in diabetic patients.Conclusion: U-shaped relationships between SBP and all-cause mortality were observed regardless of diabetes status. The optimal SBP range was gradually higher with worsening glucose status.Keywords: systolic blood pressure, mortality, normoglycemia, prediabetes, diabetes mellitus

Ying-Qi Feng,1,* Bo-An Li,2,* Fan Feng,2,* Yong-Shou Chen,1 Yi-Xin Ren,1 Heng Zhang,1 Shuang Cao1 1Key Laboratory for Green Chemical Process of Ministry of Education, School of Chemical Engineering and Pharmacy, Wuhan Institute of Technology, Wuhan 430072, People’s Republic of China; 2Center for Clinical Laboratory, The Fifth Medical Center, General Hospital of Chinese PLA, Beijing 100039, People’s Republic of China*These authors contributed equally to this workCorrespondence: Shuang Cao; Heng ZhangKey Laboratory for Green Chemical Process of Ministry of Education, School of Chemical Engineering and Pharmacy, Wuhan Institute of Technology, Wuhan 430072, People’s Republic of ChinaTel/ Fax +86-18701418117; +86-15994288097Email caoshuangknight@163.com; zhzpthm@163.comBackground: Although molecular-targeted agents are still the first choice for advanced hepatocellular carcinoma (HCC) treatment, the therapeutic efficacy of these agents is not satisfactory. Recently, the mammalian target of rapamycin (mTOR) is considered to be a promising molecular target that can enhance the sensitivity of HCC cells to antitumor therapy. However, the reported mTOR inhibitors have some shortcomings, and novel mTOR inhibitors need to be developed to enhance the antitumor effect of molecularly targeted agents on advanced HCC.Methods: In this study, five small-molecular compounds that could serve as potential mTOR-specific inhibitors were identified by virtual screening. The activity of tert-butyl (4-(9-(2-(1,3-dioxolan-2-yl)ethyl)-6-morpholino-9H-purin-2-yl)phenyl)carbamate (compound 4) was measured by enzyme test and Western blot, and its antitumor effect on HCC was examined in nude mice subcutaneous tumor model.Results: The results showed that 4 is the most effective one in inhibiting the activation of mTOR kinase (mTOR IC50 = 17.52± 3.67 nmol/L) among the five lead compounds. Further research in this study indicated that treatment with 4 enhanced the sensitivity of HCC cells to the molecular-targeted agents, such as sorafenib, regorafenib, lenvatinib, anlotinib, and apatinib. In addition, this research indicated that mTOR was correlated with the poor prognosis in patients with advanced HCC who received sorafenib.Conclusion: Our study identified a new type of small-molecular inhibitors of mTOR and confirmed their ability to enhance the antitumor effect of molecular-targeted agents on advanced HCC.Keywords: hepatocellular carcinoma, mTOR, molecular-targeted agents, virtual screening

Ying-Qi Feng,1,* Shuang-Xi Gu,1,* Yong-Shou Chen,1,* Xu-Dong Gao,2 Yi-Xin Ren,1 Jian-Chao Chen,3 Yin-Ying Lu,2 Heng Zhang,1 Shuang Cao1,4 1Key Laboratory for Green Chemical Process of Ministry of Education, School of Chemical Engineering and Pharmacy, Wuhan Institute of Technology, Wuhan 430072, People’s Republic of China; 2Comprehensive Liver Cancer Department, The Fifth Medical Center, Chinese PLA General Hospital, Beijing 100039, People’s Republic of China; 3Key Laboratory of Structure-Based Drug Design and Discovery, Shenyang Pharmaceutical University, Ministry of Education, Shenyang 110016, People’s Republic of China; 4National Engineering Research Center for the Emergency Drug, Beijing Institute of Pharmacology and Toxicology, Beijing 100850, People’s Republic of China*These authors contributed equally to this workCorrespondence: Shuang Cao; Heng ZhangKey Laboratory for Green Chemical Process of Ministry of Education, School of Chemical Engineering and Pharmacy, Wuhan Institute of Technology, Wuhan 430072, People’s Republic of ChinaTel/ Fax +86-18701418117; +86-15994288097Email caoshuang@wit.edu.cn; zhzpthm@163.comBackground: Radiotherapy has an ameliorative effect on a wide variety of tumors, but hepatocellular carcinoma (HCC) is insensitive to this treatment. Overactivated mammalian target of rapamycin (mTOR) plays an important part in the resistance of HCC to radiotherapy; thus, mTOR inhibitors have potential as novel radiosensitizers to enhance the efficacy of radiotherapy for HCC.Methods: A lead compound was found based on pharmacophore modeling and molecular docking, and optimized according to the differences between the ATP-binding pockets of mTOR and PI3K. The radiosensitizing effect of the optimized compound ( 2a) was confirmed by colony formation assays and DNA double-strand break assays in vitro. The discovery and preclinical characteristics of this compound are described.Results: The key amino acid residues in mTOR were identified, and a precise virtual screening model was constructed. Compound 2a, with a 4,7-dihydro-[1,2,4]triazolo[1,5-a]pyrimidine scaffold, exhibited promising potency against mTOR (mTOR IC50=7.1 nmol/L (nM)) with 126-fold selectivity over PI3Kα. Moreover, 2a significantly enhanced the sensitivity of HCC to radiotherapy in vitro in a dose-dependent manner.Conclusion: A new class of selective mTOR inhibitors was developed and their radiosensitization effects were confirmed. This study also provides a basis for developing mTOR-specific inhibitors for use as radiosensitizers for HCC radiotherapy.Keywords: virtual docking, HCC, kinase inhibitor, mTOR, radiosensitizer

Jie Jiang,1 Yu-Jie Zhou,2 Jian-Jun Li,3 Jun-Bo Ge,4 Ying-Qing Feng,5 Yong Huo1 On behalf of ASAP Study Investigators 1Department of Cardiology, Peking University First Hospital, Xicheng District, Beijing 100034, China; 2Department of Cardiology, Beijing Anzhen Hospital, Chaoyang District, Beijing 100029, China; 3Department of Cardiology, Fuwai Hospital, Xicheng District, Beijing 100037, China; 4Department of Cardiology, Fudan University Zhongshan Hospital, Xuhui District, Shanghai 200032, China; 5Department of Cardiology, Guangdong General Hospital, Yuexiu District, Guangzhou, Guangdong Province 510080, China Objective: Despite current standard of care, the overall lipid goal attainment rate for hyperlipidemia patients, especially those who have experienced acute coronary syndrome (ACS), is suboptimal, which predisposes them to a higher residual risk of atherothrombotic events. This study aimed to describe characteristics of Chinese patients who recently experienced an ACS event and were on lipid-lowering treatment, yet failing to reach targeted goal. Methods: A multicenter, cross-sectional study was conducted to recruit 2,034 Chinese patients who experienced an ACS (ST segment elevation myocardial infarction [STEMI], non-STEMI, or unstable angina) event within the past 4–40 weeks and were on statin treatment (>2 weeks) from March 2015 to December 2016. All eligible patients underwent a fasting lipid test after enrollment and data on medical history were collected. Results: The mean age of 1,994 eligible patients was 61.0±9.84 years. Among them, 1,493 (74.9%) patients received intensive statin therapy (defined as atorvastatin 40 or 80 mg, or rosuvastatin 20 mg per protocol) and 499 (25.0%) patients were on maximum tolerated dose statin. Of the 1,994 eligible subjects, 1,273 (63.8%) patients did not achieve the lipid goal at the time of enrollment. Among the not-at-goal patients, 910 (71.5%) received intensive statin therapy; the majority (73.4%) of them were male; the mean age was 61.2±10.1 years old; 699 (54.9%) patients had a history of hypertension; 25.3% had diabetes mellitus; and 29.5% were current smokers. The mean low-density lipoprotein-cholesterol (LDL-C), non-high-density lipoprotein-cholesterol (non-HDL-C), and ApoB levels at enrollment of this group of patients were 2.460±0.7139 mmol/L, 3.094±0.8861 mmol/L, and 0.840±0.3015 g/L, respectively. Conclusion: The study result demonstrates that overall more than half of the patients who recently (4–40 weeks) experienced ACS who were treated did not reach the guideline-recommended LDL-C and non-HDL-C goal. These results highlight the potential necessity for a new drug beyond statins to further reduce disease burden in the future. Keywords: acute coronary syndrome, intensive statin treatment, lipid goal attainment, lipid-lowering treatment

Rationale: Spermatogenesis is a highly organized cell differentiation process in mammals, involving mitosis, meiosis, and spermiogenesis. DIS3L2, which is primarily expressed in the cytoplasm, is an RNA exosome-independent ribonuclease. In female mice, Dis3l2 -deficient oocytes fail to resume meiosis, resulting in arrest at the germinal vesicle stage and complete infertility. However, the role of DIS3L2 in germ cell development in males has remained largely unexplored. Methods: We established a pre-meiotic germ cell conditional knockout mouse model and investigated the biological function of DIS3L2 in spermatogenesis and male fertility through bulk RNA-seq and scRNA-seq analyses. Results: This study unveils that conditional ablation of Dis3l2 in pre-meiotic germ cells with Stra8-Cre mice impairs spermatogonial differentiation and hinders spermatocyte meiotic progression coupled with cell apoptosis. Such conditional ablation leads to defective spermatogenesis and sterility in adults. Bulk RNA-seq analysis revealed that Dis3l2 deficiency significantly disrupted the transcriptional expression pattern of genes related to the cell cycle, spermatogonial differentiation, and meiosis in Dis3l2 conditional knockout testes. Additionally, scRNA-seq analysis indicated that absence of DIS3L2 in pre-meiotic germ cells causes disrupted RNA metabolism, downregulated expression of cell cycle genes, and aberrant expression of spermatogonial differentiation genes, impeding spermatogonial differentiation. In meiotic spermatocytes, loss of DIS3L2 results in disturbed RNA metabolism, abnormal translation, and disrupted meiotic genes that perturb meiotic progression and induce cell apoptosis, leading to subsequent failure of spermatogenesis and male infertility. Conclusions: Collectively, these findings highlight the critical role of DIS3L2 ribonuclease-mediated RNA degradation in safeguarding the correct transcriptome during spermatogonial differentiation and spermatocyte meiotic progression, thus ensuring normal spermatogenesis and male fertility., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

DIS3L, a catalytic exoribonuclease associated with the cytoplasmic exosome complex, degrades cytoplasmic RNAs and is implicated in cancers and certain other diseases in humans. Epididymis plays a pivotal role in the transport, maturation, and storage of sperm required for male fertility. However, it remains unclear whether DIS3L-mediated cytoplasmic RNA degradation plays a role in epididymis biology and functioning. Herein, we fabricated a Dis3l conditional knockout (Dis3l cKO) mouse line in which DIS3L was ablated from the principal cells of the initial segment (IS). Morphological analyses showed that spermatogenesis and IS differentiation occurred normally in Dis3l cKO mice. Additionally, the absence of DIS3L had no dramatic influence on the transcriptome of IS. Moreover, the sperm count, morphology, motility, and acrosome reaction frequency in Dis3l cKO mice were comparable to that of the control, indicating that the Dis3l cKO males had normal fertility. Collectively, our genetic model demonstrates that DIS3L inactivation in the IS is nonessential for sperm maturation and male fertility., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of interest regarding the publication of this research., (Copyright © 2024 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

One new 2-arylbenzo[ b ]furan named iteafuranal F ( 1 ) as well as two known analogues ( 2-3 ) were isolated from the 95% EtOH extract of aerial parts of Itea omeiensis . Their chemical structures were constructed based on extensive analyses of UV, IR, 1D/2D NMR and HRMS spectra. Antioxidant assays revealed significant superoxide anion radical scavenging capacity of 1 with IC 50 value of 0.66 mg/mL, which was comparable to the efficiency of positive control of luteolin. In addition, the preliminary MS fragmentation patterns in negative ion mode were established to distinguish 2-arylbenzo[ b ]furans with C-10 in different oxidation states: the characteristic loss of CO molecule [M-H-28] - was observed for 3-formyl-2-arylbenzo[ b ]furans, and the loss of CH 2 O fragment [M-H-30] - for 3-hydroxymethyl-2-arylbenzo[ b ]furans, and the loss of CO 2 fragment [M-H-44] - for 2-arylbenzo[ b ]furan-3-carboxylic acids.

Translational control plays a crucial role in the regulation of apoptosis, with the EIF4 family serving as one of the mRNA translation factors that modulate the process of mRNA translation based on mRNA characteristics. To address this potential causal role of EIF4 family proteins and breast cancer, Mendelian randomization was employed. The study incorporated four sets of genetics instrumental variables, namely EIF4E, EIF4B, EIF4A, and EIF4EBP2. The outcome variables selected for analysis were the BCAC consortium, which included estrogen receptor positive (ER+) and estrogen receptor negative (ER-) samples. To assess the potential violations of the MR assumption, the primary MR analysis employed inverse variance weighted (IVW), and several sensitivity analyses were conducted. The findings of the two-sample MR analysis indicate that EIF4E has an adverse effect on breast cancer risk (p = 0.028). However, the evidence for the relationship between EIF4E and ER status of breast cancer suggests a weak association with ER+ breast cancer (p = 0.054), but not with ER- breast cancer (p > 0.05). The study findings indicate that EIF4A is not causally linked to the risk of ER+ breast cancer, but is significantly associated with an elevated risk of ER- breast cancer (p = 0.028). However, the evidence is inadequate to support the effects of EIF4B and EIF4EBP2 on breast cancer (p > 0.05). Our results suggest that EIF4 may be a potential factor in the occurrence and development of breast cancer, which may lead to a better understanding of its causes and prevention., (© 2024. The Author(s).)

Acyl-Coenzyme As (acyl-CoAs) are essential intermediates to incorporate carboxylic acids into the bioactive metabolic network across all species, which play important roles in lipid remodeling, fatty acids, and xenobiotic carboxylic metabolism. However, due to the poor liquid chromatographic behavior, the relatively low mass spectrometry (MS) sensitivity, and lack of authentic standards for annotation, the in-depth untargeted profiling of acyl-CoAs is challenging. We developed a chemical derivatization strategy of acyl-CoAs by employing 8-(diazomethyl) quinoline (8-DMQ) as the labeling reagent, which increased the detection sensitivity by 625-fold with good peak shapes. By applying the MS/MS fragmentation rules learned from the MS/MS spectra of 8-DMQ-acyl-CoA authentic standards, an 8-DMQ-acyl-CoA in silico mass spectral library containing 33,344 high-resolution tandem mass spectra of 8,336 acyl-CoA species was created. The in silico library facilitated the high-throughput and automatic annotation of acyl-CoA using multiple metabolomic data processing tools, such as NIST MS Search and MSDIAL. The feasibility of the in silico library in a complex sample was demonstrated by profiling endogenous acyl-CoAs in multiple organs of an aging mouse. 53 acyl-CoA species were annotated, including 12 oxidized fatty acyl-CoAs and 3 novel nonfatty acyl-CoAs. False positive annotations were further screened by developing an eXtreme Gradient Boosting (XGBoost) based retention time prediction model. The organ distribution and the aging dynamics of acyl-CoAs in a mouse model were discussed for the first time, which helped to elucidate the organ-specific function of acyl-CoAs and the role of different acyl-CoA species during aging.

Background: Masked hypertension is associated with target organ damage (TOD) and adverse health outcomes, but whether antihypertensive treatment improves TOD in patients with masked hypertension is unproven., Methods: In this multicentre, randomised, double-blind, placebo-controlled trial at 15 Chinese hospitals, untreated outpatients aged 30-70 years with an office blood pressure (BP) of <140/<90 mm Hg and 24-h, daytime or nighttime ambulatory BP of ≥130/≥80, ≥135/≥85, or ≥120/≥70 mm Hg were enrolled. Patients had ≥1 sign of TOD: electrocardiographic left ventricular hypertrophy (LVH), brachial-ankle pulse wave velocity (baPWV) ≥1400 cm/s, or urinary albumin-to-creatinine ratio (ACR) ≥3.5 mg/mmol in women and ≥2.5 mg/mmol in men. Exclusion criteria included secondary hypertension, diabetic nephropathy, serum creatinine ≥176.8 μmol/L, and cardiovascular disease within 6 months of screening. After stratification for centre, sex and the presence of nighttime hypertension, eligible patients were randomly assigned (1:1) to receive antihypertensive treatment or placebo. Patients and investigators were masked to group assignment. Active treatment consisted of allisartan starting at 80 mg/day, to be increased to 160 mg/day at month 2, and to be combined with amlodipine 2.5 mg/day at month 4, if the ambulatory BP remained uncontrolled. Matching placebos were used likewise in the control group. The primary endpoint was the improvement of TOD, defined as normalisation of baPWV, ACR or LVH or a ≥20% reduction in baPWV or ACR over the 48-week follow-up. The intention-to-treat analysis included all randomised patients, the per-protocol analysis patients who fully adhered to the protocol, and the safety analysis all patients who received at least one dose of the study medication. This study is registered with ClinicalTrials.gov, NCT02893358., Findings: Between February 14, 2017, and October 31, 2020, 320 patients (43.1% women; mean age ± SD 53.7 ± 9.7 years) were enrolled. Baseline office and 24-h BP averaged 130 ± 6.0/81 ± 5.9 mm Hg and 136 ± 8.6/84 ± 6.1 mm Hg, and the prevalence of elevated baPWV, ACR and LVH were 97.5%, 12.5%, and 7.8%, respectively. The 24-h BP decreased on average (±SE) by 10.1 ± 0.9/6.4 ± 0.5 mm Hg in 153 patients on active treatment and by 1.3 ± 0.9/1.0 ± 0.5 mm Hg in 167 patients on placebo. Improvement of TOD occurred in 79 patients randomised to active treatment and in 49 patients on placebo: 51.6% (95% CI 43.7%, 59.5%) versus 29.3% (22.1, 36.5%; p < 0.0001). Per-protocol and subgroup analyses were confirmatory. Adverse events were generally mild and occurred in 38 (25.3%) and 43 (26.4%) patients randomised to active treatment and placebo, respectively (p = 0.83)., Interpretation: Our results suggest that antihypertensive treatment improves TOD in patients with masked hypertension, highlighting the need of treatment. However, the long-term benefit in preventing cardiovascular complications still needs to be established., Funding: Salubris China., Competing Interests: JGW reports having received research grants, lecture and consulting fees from A&D, Bayer, Novartis, Omron, Servier and Viatris. YL reports having received research grants from A&D, Bayer, Omron, Salubris, and Shyndec and lecture fees from A&D, Omron, Servier, Salubris and Shyndec. All other authors declare no competing interests., (© 2024 The Author(s).)

Background: Acute kidney injury (AKI) is a common and serious complication after cardiac surgery that significantly affects patient outcomes. Given the limited treatment options available, identifying modifiable risk factors is critical. Frailty and obesity, two heterogeneous physiological states, have significant implications for identifying and preventing AKI. Our study investigated the interplay among frailty, body composition, and AKI risk after cardiac surgery to inform patient management strategies., Material and Methods: This retrospective cohort study included three international cohorts. Primary analysis was conducted in adult patients who underwent cardiac surgery between 2014 and 2019 at Wuhan XX Hospital, China. We tested the generalizability of our findings with data from two independent international cohorts, the Medical Information Mart for Intensive Care IV (MIMIC-IV) and the eICU Collaborative Research Database. Frailty was assessed using a clinical lab-based frailty index (FI-LAB), while total body fat percentage (BF%) was calculated based on a formula accounting for BMI, sex, and age. Logistic regression models were used to analyze the associations between frailty, body fat, and AKI, adjusting for pertinent covariates., Results: A total of 8785 patients across three international cohorts were included in the study. In the primary analysis of 3,569 patients from Wuhan XX Hospital, moderate and severe frailty were associated with an increased AKI risk after cardiac surgery. Moreover, a nonlinear relationship was observed between body fat percentage and AKI risk. When stratified by the degree of frailty, lower body fat correlated with a decreased incidence of AKI. Extended analyses using the MIMIC-IV and eICU cohorts (n=3,951 and n=1,265, respectively) validated these findings and demonstrated that a lower total BF% was associated with decreased AKI incidence. Moderation analysis revealed that the effect of frailty on AKI risk was moderated by the body fat percentage. Sensitivity analyses demonstrated results consistent with the main analyses., Conclusion: Higher degrees of frailty were associated with an elevated risk of AKI following cardiac surgery, and total BF% moderated this relationship. This research underscores the significance of integrating frailty and body fat assessments into routine cardiovascular care to identify high-risk patients for AKI and implement personalized interventions to improve patient outcomes., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Iridocyclitis and the use of glucocorticoid medication have been widely studied as susceptibility factors for cataracts. However, the causal relationship between them remains unclear. This study aimed to investigate the causal relationship between the development of iridocyclitis and the genetic liability of glucocorticoid medication use on the risk of senile cataracts occurrence by performing Two-sample Mendelian randomization (MR) analyses. Instrumental variables (IVs) significantly associated with exposure factors (P < 5 × 10 -8 ) were identified using published genome-wide association data from the FinnGen database and UK Biobank. Reliability analyses were conducted using five approaches, including inverse-variance weighted (IVW), MR-Egger regression, simple median, weighted median, and weighted mode. A sensitivity analysis using the leave-one-out method was also performed. Genetic susceptibility to glucocorticoid use was associated with an increased risk of developing senile cataracts (OR, 1.10; 95% CI, 1.02-1.17; P < 0.05). Moreover, iridocyclitis was significantly associated with a higher risk of developing senile cataracts (OR, 1.03; 95% CI, 1.01-1.05; P < 0.05). Nonetheless, some heterogeneity in the IVs was observed, but the MR results remained consistent after penalizing for outliers. The estimates were consistent in multivariate analyses by adjusting for body mass index (BMI) and diabetes mellitus type 2 (T2DM). This study provides new insights into the prevention and management of senile cataracts by highlighting the increased risk associated with iridocyclitis and the use of glucocorticoids.

Adenosine-to-inosine (A-to-I) editing and N 6 -methyladenosine (m 6 A) modifications are pivotal RNA modifications with widespread functional significance in physiological and pathological processes. Although significant effort has been dedicated to developing methodologies for identifying and quantifying these modifications, traditional approaches have often focused on each modification independently, neglecting the potential co-occurrence of A-to-I editing and m 6 A modifications at the same adenosine residues. This limitation has constrained our understanding of the intricate regulatory mechanisms governing RNA function and the interplay between different types of RNA modifications. To address this gap, we introduced an innovative technique called deamination-assisted reverse transcription stalling (DARTS), specifically designed for the simultaneous quantification of A-to-I editing and m 6 A at the same RNA sites. DARTS leverages the selective deamination activity of the engineered TadA-TadA8e protein, which converts adenosine residues to inosine, in combination with the unique property of Bst 2.0 DNA polymerase, which stalls when encountering inosine during reverse transcription. This approach enables the accurate quantification of A-to-I editing, m 6 A, and unmodified adenosine at identical RNA sites. The DARTS method is remarkable for its ability to directly quantify two distinct types of RNA modifications simultaneously, a capability that has remained largely unexplored in the field of RNA biology. By facilitating a comprehensive analysis of the co-occurrence and interaction between A-to-I editing and m 6 A modifications, DARTS opens new avenues for exploring the complex regulatory networks modulated by different RNA modifications.

The possibility of cyanoacetohydrazide usage as a novel derivatizing agent is demonstrated in the presented article, and a comparison with hydroxylamine as the most commonly used reagent is provided. Optimal conditions for steroid derivatization with cyanoacetohydrazide are provided. According to the collected data, the maximum yield of derivatives was observed at pH 2.8 within 70 min at 40 °C with 5 ng/mL limit of detection for all investigated analytes. It was shown that cyanoacetohydrazide derivatives produces both syn- and anti-forms as well as hydroxylamine, and their ratios were evaluated and shown in presented work. An efficiency enchantment from two to up to five times was achieved with a novel derivatization reagent. Its applicability for qualitative analysis of steroids in urine was presented at real samples. Additionally, the reproducible fragmentation of the derivatizing agent in collision-induced dissociation offers opportunities for simplified non-targeted steroidomic screening. Furthermore, cyanoacetohydrazide increases ionization efficiency in positive mode, which can eliminate the need for redundant high-resolution instrument runs required for both positive and negative mode analyses.

Cytosine modifications, particularly 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), play crucial roles in numerous biological processes. Current analytical methods are often constrained to the separate detection of either 5mC or 5hmC, or the combination of both modifications. The ability to simultaneously detect C, 5mC, and 5hmC at the same genomic locations with precise stoichiometry is highly desirable. Herein, we introduce a method termed engineered deaminase-assisted sequencing (EDA-seq) for the simultaneous quantification of C, 5mC, and 5hmC at the same genomic sites. EDA-seq utilizes a specially engineered protein, derived from human APOBEC3A (A3A), known as eA3A-M5. eA3A-M5 exhibits distinct deamination capabilities for C, 5mC, and 5hmC. In EDA-seq, C undergoes complete deamination and is sequenced as T. 5mC is partially deaminated resulting in a mixed readout of T and C, and 5hmC remains undeaminated and is read as C. Consequently, the proportion of T readouts ( P T ) reflects the collective occurrences of C and 5mC, regulated by the deamination rate of 5mC ( R 5mC ). By determining R 5mC and P T values, we can deduce the precise levels of C, 5mC, and 5hmC at particular genomic locations. We successfully used EDA-seq to simultaneously measure C, 5mC, and 5hmC at specific loci within human lung cancer tissue and their normal counterpart. The results from EDA-seq demonstrated a strong concordance with those obtained from the combined application of BS-seq and ACE-seq methods. EDA-seq eliminates the need for bisulfite treatment, DNA oxidation or glycosylation and uniquely enables simultaneous quantification of C, 5mC and 5hmC at the same genomic locations., Competing Interests: The authors declare no competing financial interest., (This journal is © The Royal Society of Chemistry.)

Rationale: In recent years, nicotinamide adenine dinucleotide (NAD + ) precursors (Npre) have been widely employed to ameliorate female reproductive problems in both humans and animal models. However, whether and how Npre plays a role in the male reproductive disorder has not been fully clarified. Methods: In the present study, a busulfan-induced non-obstructive azoospermic mouse model was used, and Npre was administered for five weeks following the drug injection, with the objective of reinstating spermatogenesis and fertility. Initially, we assessed the NAD + level, germ cell types, semen parameters and sperm fertilization capability. Subsequently, testis tissues were examined through RNA sequencing analysis, ELISA, H&E, immunofluorescence, quantitative real-time PCR, and Western blotting techniques. Results: The results indicated that Npre restored normal level of NAD + in blood and significantly alleviated the deleterious effects of busulfan (BU) on spermatogenesis, thereby partially reestablishing fertilization capacity. Transcriptome analysis, along with recovery of testicular Fe 2+ , GSH, NADPH, and MDA levels, impaired by BU, and the fact that Fer-1, an inhibitor of ferroptosis, restored spermatogenesis and semen parameters close to CTRL values, supported such possibility. Interestingly, the reduction in SIRT2 protein level by the specific inhibitor AGK2 attenuated the beneficial effects of Npre on spermatogenesis and ferroptosis by affecting PGC-1α and ACLY protein levels, thus suggesting how these compounds might confer spermatogenesis protection. Conclusion: Collectively, these findings indicate that NAD + protects spermatogenesis against ferroptosis, probably through SIRT2 dependent mechanisms. This underscores the considerable potential of Npre supplementation as a feasible strategy for preserving or restoring spermatogenesis in specific conditions of male infertility and as adjuvant therapy to preserve male fertility in cancer patients receiving sterilizing treatments., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Background: Danggui Sini decoction (DSD), a traditional Chinese medicine formula, has the function of nourishing blood, warming meridians, and unblocking collaterals. Our clinical and animal studies had shown that DSD can effectively protect against oxaliplatin (OXA)-induced peripheral neuropathy (OIPN), but the detailed mechanisms remain uncertain. Multiple studies have confirmed that gut microbiota plays a crucial role in the development of OIPN. In this study, the potential mechanism of protective effect of DSD against OIPN by regulating gut microbiota was investigated., Methods: The neuroprotective effects of DSD against OIPN were examined on a rat model of OIPN by determining mechanical allodynia, biological features of dorsal root ganglia (DRG) as well as proinflammatory indicators. Gut microbiota dysbiosis was characterized using 16S rDNA gene sequencing and metabolism disorders were evaluated using untargeted and targeted metabolomics. Moreover the gut microbiota mediated mechanisms were validated by antibiotic intervention and fecal microbiota transplantation., Results: DSD treatment significantly alleviated OIPN symptoms by relieving mechanical allodynia, preserving DRG integrity and reducing proinflammatory indicators lipopolysaccharide (LPS), IL-6 and TNF-α. Besides, DSD restored OXA induced intestinal barrier disruption, gut microbiota dysbiosis as well as systemic metabolic disorders. Correlation analysis revealed that DSD increased bacterial genera such as Faecalibaculum, Allobaculum, Dubosiella and Rhodospirillales&#95;unclassified were closely associated with neuroinflammation related metabolites, including positively with short-chain fatty acids (SCFAs) and sphingomyelin (d18:1/16:0), and negatively with pi-methylimidazoleacetic acid, L-glutamine and homovanillic acid. Meanwhile, antibiotic intervention apparently relieved OIPN symptoms. Furthermore, fecal microbiota transplantation further confirmed the mediated effects of gut microbiota., Conclusion: DSD alleviates OIPN by regulating gut microbiota and potentially relieving neuroinflammation related metabolic disorder., (© 2024. The Author(s).)

Background: Ferroptosis has recently been associated with multiple degenerative diseases. Ferroptosis induction in cancer cells is a feasible method for treating neoplastic diseases. However, the association of iron proliferation-related genes with prognosis in HER2+ breast cancer (BC) patients is unclear., Aim: To identify and evaluate fresh ferroptosis-related biomarkers for HER2+ BC., Methods: First, we obtained the mRNA expression profiles and clinical information of HER2+ BC patients from the TCGA and METABRIC public databases. A four-gene prediction model comprising PROM2 , SLC7A11 , FANCD2 , and FH was subsequently developed in the TCGA cohort and confirmed in the METABRIC cohort. Patients were stratified into high-risk and low-risk groups based on their median risk score, an independent predictor of overall survival (OS). Based on these findings, immune infiltration, mutations, and medication sensitivity were analyzed in various risk groupings. Additionally, we assessed patient prognosis by combining the tumor mutation burden (TMB) with risk score. Finally, we evaluated the expression of critical genes by analyzing single-cell RNA sequencing (scRNA-seq) data from malignant vs normal epithelial cells., Results: We found that the higher the risk score was, the worse the prognosis was ( P < 0.05). We also found that the immune cell infiltration, mutation, and drug sensitivity were different between the different risk groups. The high-risk subgroup was associated with lower immune scores and high TMB. Moreover, we found that the combination of the TMB and risk score could stratify patients into three groups with distinct prognoses. HRisk-HTMB patients had the worst prognosis, whereas LRisk-LTMB patients had the best prognosis ( P < 0.0001). Analysis of the scRNA-seq data showed that PROM2 , SLC7A11 , and FANCD2 were significantly differentially expressed, whereas FH was not, suggesting that these genes are expressed mainly in cancer epithelial cells ( P < 0.01)., Conclusion: Our model helps guide the prognosis of HER2+ breast cancer patients, and its combination with the TMB can aid in more accurate assessment of patient prognosis and provide new ideas for further diagnosis and treatment., Competing Interests: Conflict-of-interest statement: All the authors report no relevant conflicts of interest for this article., (©The Author(s) 2024. Published by Baishideng Publishing Group Inc. All rights reserved.)

DNA cytosine methylation (5-methylcytosine, 5mC) is a predominant epigenetic modification that plays a critical role in a variety of biological and pathological processes in mammals. In active DNA demethylation, the 10-11 translocation (TET) dioxygenases can sequentially oxidize 5mC to generate three modified forms of cytosine, 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). Beyond being a demethylation intermediate, recent studies have shown that 5fC has regulatory functions in gene expression and chromatin organization. While some methods have been developed to detect 5fC, genome-wide mapping of 5fC at base resolution is still highly desirable. Herein, we propose a chemical labeling enrichment and deamination sequencing (CLED-seq) method for detecting 5fC in genomic DNA at single-base resolution. The CLED-seq method utilizes selective labeling and enrichment of 5fC-containing DNA fragments, followed by deamination mediated by apolipoprotein B mRNA-editing catalytic polypeptide-like 3A (APOBEC3A or A3A) and sequencing. In the CLED-seq process, while all C, 5mC, and 5hmC are interpreted as T during sequencing, 5fC is still read as C, enabling the precise detection of 5fC in DNA. Using the proposed CLED-seq method, we accomplished genome-wide mapping of 5fC in mouse embryonic stem cells. The mapping study revealed that promoter regions enriched with 5fC overlapped with H3K4me1, H3K4me3, and H3K27ac marks. These findings suggest a correlation between 5fC marks and active gene expression in mESCs. In conclusion, CLED-seq is a straightforward, bisulfite-free method that offers a valuable tool for detecting 5fC in genomes at a single-base resolution.

Introduction: Salvia L., representing the largest genus within the mint family, is noted for its global distribution of approximately 1000 species, with East Asia, and particularly China, recognized as a critical center of diversity for the genus., Methods: Our research was conducted through extensive fieldwork in Guidong County, Hunan Province, China, where we identified a previously undescribed species of Salvia . The identification process involved detailed morphological observations, phylogenetic analyses, and plastid genomics., Results: The newly discovered species, Salvia guidongensis , exhibits unique characteristics not commonly observed in the East Asian lineage of Salvia, including dual floral colors within natural populations-either pale purple or pale yellow. Morphologically, while it shares similarities with members of sect. Glutinaria , S. guidongensis is distinct in its floral morphology, stature, and specific foliar traits. Phylogenetic analysis places S. guidongensis in a unique clade within the East Asian lineage of Salvia , suggesting it may serve as an important evolutionary link. Additionally, we explored the plastome features of S. guidongensis , comparing them with those of closely related species., Discussion: The discovery of S. guidongensis not only entriches the taxonomic tapestry of Salvia but also provides critical insights into the biogeography and evolutionary pathways of the genus in East Asia. By integrating morphological and molecular data, we validate the novel status of S. guidongensis and highlight its significance in bridging taxonomic and evolutionary gaps within Sect. Glutinaria of Salvia ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Huang, Qi, Feng, Ge, Huang, Feng, Wu, Wei, Ito, Kokubugata, Li and Wei.)

Various separation methods in combination with spectral data analysis, X-ray single crystal diffraction analysis, and litera-ture data comparison were employed to clarify the chemical constituents of Itea yunnanensis. Seven compounds were obtained from I. yunnanensis, which were identified as(S)-3-[1-(4-hydroxyphenyl)propane-2-yl]-4-methoxybenzoate methyl ester(1), iteafuranal B(2), syringaresinol(3), dihydrokaempferol(4), trimethoxybenzene(5), eicosane(6), and nonacosane(7), respectively. Among them, compound 1 was a new nor-neolignan compound named iteanorneoligan A, and the rest of the compounds were identified from I. yunnanensis for the first time. The anti-hepatocellular carcinoma effect of the compound was evaluated based on Sk-hep-1 cells model via MTT assay, and compound 2 showed a significant inhibitory effect on the proliferation of Sk-hep-1 cells with an IC&#95;(50) of 9.4 μmol·L~(-1). The antioxidant capacity was determined via DPPH, ABTS~(·+), and O■ radical scavenging ability, and compound 1 exhibited a significant ABTS~(·+) radical scavenging effect with an IC&#95;(50) of 0.178 mg·mL~(-1).

In vitro fertilization (IVF) is a highly effective treatment for infertility; however, it poses challenges for women with decreased ovarian reserve (DOR). Despite the importance of understanding the impact of DOR on IVF outcomes, limited research has explored this relationship, particularly using omics approaches. Hence, we conducted a study to investigate the association between DOR and IVF outcomes, employing a metabolomic approach. We analyzed serum samples from 207 women undergoing IVF treatment, including 89 with DOR and 118 with normal ovarian reserve (NOR). Our findings revealed that DOR was significantly associated with unfavorable IVF outcomes, characterized by a reduced oocyte count, lower embryo quality, and decreased rates of pregnancy and live births. Furthermore, we identified 82 metabolites that displayed significant alterations in DOR patients, impacting diverse metabolic pathways. Notably, a distinct panel of metabolites, including palmitic acid, stearic acid, LysoPC(9:0(CHO)/0:0), PC(18:0/9:0(CHO)), and PC(16:0/9:0(CHO)), exhibited discriminatory power between the DOR and NOR groups, showcasing a strong correlation with IVF outcomes. These findings emphasize the crucial role of metabolomic disruptions in influencing IVF outcomes among women with DOR.

Background: Wuhu Oral Liquid (WHOL) is a modified preparation derived from the famous Wuhu Powder, which has a long history of use in treating traumatic injuries. This preparation has anti-inflammatory and analgesic properties and accelerates recovery following acute soft tissue injuries. Aims: To evaluate the efficacy and safety of WHOL in treating acute soft tissue injury associated with qi stagnation and blood stasis syndrome and to provide a basis for applying for the protection of varieties of Chinese medicine for WHOL. Methods: This study was a randomized, controlled, double-blind, multicenter clinical trial in which Fufang Shang Tong Capsule (FFSTC) was selected as the control drug. A total of 480 subjects with acute soft tissue injury associated with qi stagnation and blood stasis syndrome were randomly divided into a test and control group in a 3:1 ratio. The duration of drug treatment was 10 days. The primary outcome was Visual Analogue Scale (VAS) score for pain (including pain at rest and pain on activity). Secondary outcomes included the disappearance time of the pain at rest and on activity; the curative effect of TCM syndrome and improvement in the individual symptoms of TCM (swelling, ecchymosis, and dysfunction); and changes in C-reactive protein (CRP) and interleukin-6 (IL-6) levels. Safety was assessed using vital signs, laboratory examinations, electrocardiograms, and physical examinations. Results: Patient compliance was satisfactory in both groups (all between 80% and 120%). After 4 days of treatment, the WHOL group was superior to the FFSTC group in decreasing the VAS scores for pain at rest (-1.88 ± 1.13 vs . -1.60 ± 0.93, p < 0.05) and on activity (-2.16 ± 1.18 vs . -1.80 ± 1.07, p < 0.05). After 7 days of treatment, the WHOL group was superior to the FFSTC group in decreasing the VAS scores for pain on activity (-3.87 ± 1.60 vs . -3.35 ± 1.30, p < 0.01) and improving swelling (cure rate: 60.4% vs . 46.2%, p < 0.05; obvious effective rate: 60.7% vs . 47.0%, p < 0.05). After 10 days of treatment, the WHOL group was superior to the FFSTC group in decreasing the levels of CRP (-0.13 ± 2.85 vs . 0.25 ± 2.09, p < 0.05) and improving the TCM syndrome (cure rate: 44.1% vs . 30.8%, p < 0.05) and swelling (cure rate: 75.6% vs . 67.5%, p < 0.01; obvious effective rate: 75.6% vs . 68.4%, p < 0.05; effective rate: 77.0% vs . 71.8%, p < 0.05). The disappearance time of pain at rest was 8 days in both groups and 9 days on activity in both groups. In addition, there was no statistical difference between the incidence of adverse events (4.5% vs . 2.6%, p > 0.05) and adverse reactions (0.3% vs . 0%, p > 0.05) between the WHOL group and the FFSTC group. No serious adverse events occurred in either group, and no subjects were withdrawn because of adverse events. Conclusion: WHOL relieves the symptoms caused by acute soft tissue injury associated with qi stagnation and blood stasis syndrome more rapidly than FFSTC, and it is effective and safe in the treatment of acute soft tissue injury. Future studies still need a larger sample size to verify its efficacy and safety. Clinical Trial Registration: https:// www.chictr.org.cn/showproj.html?proj=149531, Identifier ChiCTR2200056411., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhu, Shen, Xiao, Shi, Fan, Feng, Wan, Qu, Zhao, Zhang, Xu, Wu and Tang.)

Peak alignment is a crucial data-processing step in untargeted metabolomics analysis that aims to integrate metabolite data from multiple liquid chromatography-mass spectrometry (LC-MS) batches for enhanced comparability and reliability. However, slight variations in the chromatographic separation conditions can result in retention time (RT) shifts between consecutive analyses, adversely affecting peak alignment accuracy. In this study, we present a retention index (RI)-based chromatographic peak-shift correction (CPSC) strategy to address RT shifts and align chromatographic peaks for metabolomics studies. A series of N -acyl glycine homologues (C2-C23) was synthesized as calibrants, and an LC RI system was established. This system effectively corrected RT shifts arising from variations in flow rate, gradient elution, instrument systems, and chromatographic columns. Leveraging the RI system, we successfully adjusted the RT of raw data to mitigate RT shifts and then implemented the Joint Aligner algorithm for peak alignment. We assessed the accuracy of the RI-based CPSC strategy using pooled human fecal samples as a test model. Notably, the application of the RI-based CPSC strategy to a long-term dataset spanning 157 d as an illustration revealed a significant enhancement in peak alignment accuracy from 15.5% to 80.9%, indicating its ability to substantially improve peak-alignment precision in multibatch LC-MS analyses.

Gibberellins (GAs) play a pivotal role in modulating plant growth and development. Glucose-conjugated gibberellins (Glc-GAs), a prevalent conjugated form of GAs, regulate intracellular GA levels by the coupling and decoupling of glucose groups. However, the diversity of Glc-GAs identified within individual species remains limited, hinting at a multitude of yet undiscovered gibberellin metabolites. This lacuna poses considerable impediments to research efforts dedicated to comprehensively delineating the GA metabolic pathway. In this study, we developed a structure-oriented screening and identification method for Glc-GAs in plant species by employing LC-MS/MS coupled with chemical derivatization. Through the application of chemical derivatization technique, carboxyl groups on Glc-GAs were labeled which effectively enhanced the sensitivity and selectivity of mass spectrometry detection for these compounds. Concurrently, the integration of mass spectrometry fragmentation and chromatographic retention behavior facilitated the efficient screening and identification of potential Glc-GAs. With this strategy, we screened and identified 12 potential Glc-GAs from six plant species. These findings expand the Glc-GA diversity in plants and contribute to understanding GA metabolic pathways.

Background: Immunosuppressive treatment in heart transplant (HTx) recipient causes osteoporosis. The urinary proteomic profile (UPP) includes peptide fragments derived from the bone extracellular matrix. Study aims were to develop and validate a multidimensional UPP biomarker for osteoporosis in HTx patients from single sequenced urinary peptides identifying the parent proteins., Methods: A single-center HTx cohort was analyzed. Urine samples were measured by capillary electrophoresis coupled with mass spectrometry. Cases with osteoporosis and matching controls were randomly selected from all available 389 patients. In derivation case-control dataset, 1576 sequenced peptides detectable in ≥30 % of patients. Applying statistical analysis on these, an 18-peptide multidimensional osteoporosis UPP biomarker (OSTEO18) was generated by support vector modeling. The 2 replication datasets included 118 and 94 patients. For further validation, the whole cohort was analyzed. Statistical methods included logistic regression and receiver operating characteristic curve (ROC) analysis., Results: In derivation dataset, the AUC, sensitivity and specificity of OSTEO18 were 0.83 (95 % CI: 0.76-0.90), 74.3 % and 87.1 %, respectively. In replication datasets, results were confirmatory. In the whole cohort (154 osteoporotic patients [39.6 %]), the ORs for osteoporosis increased ( p  < 0.0001) across OSTEO18 quartiles from 0.39 (95 % CI: 0.25-0.61) to 3.14 (2.08-4.75). With full adjustment for known osteoporosis risk factors, OSTEO18 improved AUC from 0.708 to 0.786 ( p  = 0.0003) for OSTEO18 categorized (optimized threshold: 0.095) and to 0.784 ( p  = 0.0004) for OSTEO18 as continuously distributed classifier., Conclusion: OSTEO18 is a clinically meaningful novel biomarker indicative of osteoporosis in HTx recipients and is being certified as in-vitro diagnostic., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: uPROPHET was supported by the European Research Council (Advanced Researcher Grant-2011-294713-EPLORE and Proof-of-Concept Grant-713601-uPROPHET awarded to JAS). OMRON Healthcare, Co., Ltd., Kyoto, Japan provided a non-binding grant to the Non-Profit Research Association Alliance for the Promotion of Preventive Medicine (APPREMED), Mechelen, Belgium. Dries S. Martens holds a postdoctoral grant by the Research Foundations Flanders (FWO grants 12X9623N)., (© 2024 The Author(s).)

RNA molecules undergo various chemical modifications that play critical roles in a wide range of biological processes. N 6 , N 6 -Dimethyladenosine (m 6,6 A) is a conserved RNA modification and is essential for the processing of rRNA. To gain a deeper understanding of the functions of m 6,6 A, site-specific and accurate quantification of this modification in RNA is indispensable. In this study, we developed an AlkB-facilitated demethylation (AD-m 6,6 A) method for the site-specific detection and quantification of m 6,6 A in RNA. The N 6 , N 6 -dimethyl groups in m 6,6 A can cause reverse transcription to stall at the m 6,6 A site, resulting in truncated cDNA. However, we found that Escherichia coli AlkB demethylase can effectively demethylate m 6,6 A in RNA, generating full-length cDNA from AlkB-treated RNA. By quantifying the amount of full-length cDNA produced using quantitative real-time PCR, we were able to achieve site-specific detection and quantification of m 6,6 A in RNA. Using the AD-m 6,6 A method, we successfully detected and quantified m 6,6 A at position 1851 of 18S rRNA and position 937 of mitochondrial 12S rRNA in human cells. Additionally, we found that the level of m 6,6 A at position 1007 of mitochondrial 12S rRNA was significantly reduced in lung tissues from sleep-deprived mice compared with control mice. Overall, the AD-m 6,6 A method provides a valuable tool for easy, accurate, quantitative, and site-specific detection of m 6,6 A in RNA, which can aid in uncovering the functions of m 6,6 A in human diseases.

The classification and phylogenetic relationships within the Phaseoleae tribe (Leguminosae) have consistently posed challenges to botanists. This study addresses these taxonomic intricacies, with a specific focus on the Glycininae subtribe, by conducting a comprehensive analysis of the highly conserved plastome in Amphicarpaea edgeworthii Benth., a critical species within this subtribe. Through meticulous genomic sequencing, we identified a plastome size of 148,650 bp, composed of 128 genes, including 84 protein-coding genes, 36 tRNA genes, and 8 rRNA genes. Comparative genomic analysis across seven Glycininae species illuminated a universally conserved circular and quadripartite structure, with nine genes exhibiting notable nucleotide diversity, signifying a remarkable genomic variability. Phylogenetic reconstruction of 35 Phaseoleae species underscores the affinity of Amphicarpaea with Glycine , placing Apios as a sister lineage to all other Phaseoleae species, excluding Clitorinae and Diocleinae subtribes. Intriguingly, Apios , Butea , Erythrina , and Spatholobus , traditionally clumped together in the Erythrininae subtribe, display paraphyletic divergence, thereby contesting their taxonomic coherence. The pronounced structural differences in the quadripartite boundary genes among taxa with unresolved subtribal affiliations demand a reevaluation of Erythrininae's taxonomic classification, potentially refining the phylogenetic contours of the tribe.

Background: The detection of 25-hydroxyvitamin D (25OHD) from dried blood spots (DBS) has been widely studied. However, the existing pretreatment methods suffer from limitations in terms of throughput (usually exceeding 2 h), complexity (involving liquid-liquid extraction or solid-phase extraction), and contamination (including multiple steps of organic solvent evaporation)., Results: We first released 25OHD from DBS samples by 50% acetonitrile solution through ultrasonication. Subsequently, the cold-induced phase separation technique was introduced for in-situ concentration and purification. Afterward, the PTAD derivatization of 25OHD was performed directly, profiting from the high acetonitrile content in the concentrated solution. In the end, the resulting solution was submitted to LC-MS/MS for quantification. The established LC-MS/MS methodology possessed favorable analytical performance, possessing lower limit of quantification of 1 ng/mL pointing to plasma, accuracy of 86.8-110.1% and imprecision of 5.4-16.8%. Method comparison with plasma samples demonstrated that over 93% of the detections met the acceptance limit for cross-validation of ±20%., Significance and Novelty: The novel sample preparation can be finished within 15 min and eliminated the traditional steps of extraction and organic solvent evaporation. Based on this high-throughput, reliable and applicable LC-MS/MS method, the detection of 25OHD in DBS samples can be better achieved for clinical patients and researchers with relevant demands., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The epigenetic modification 5-hydroxymethylcytosine (5hmC) plays a crucial role in the regulation of gene expression. Although some methods have been developed to detect 5hmC, direct genome-wide mapping of 5hmC at base resolution is still highly desirable. Herein, we proposed a single-step deamination sequencing (SSD-seq) method, designed to precisely map 5hmC across the genome at single-base resolution. SSD-seq takes advantage of a screened engineered human apolipoprotein B mRNA-editing catalytic polypeptide-like 3A (A3A) protein, known as eA3A-v10, to selectively deaminate cytosine (C) and 5-methylcytosine (5mC) but not 5hmC. During sequencing, the deaminated C and 5mC are converted to uracil (U) and thymine (T), read as T in the sequencing data. However, 5hmC remains unaffected by eA3A-v10 and is read as C during sequencing. Consequently, the presence of C in the sequence reads indicates the original 5hmC. We applied SSD-seq to generate a base-resolution map of 5hmC in human lung tissue. Our findings revealed that 5hmC was predominantly localized to CpG dinucleotides. Furthermore, the base-resolution map of 5hmC generated by SSD-seq demonstrated a strong correlation with prior ACE-seq results. The advantages of SSD-seq are its single-step process, absence of bisulfite treatment or DNA glycosylation, cost effectiveness, and ability to detect and quantify 5hmC directly at single-base resolution., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Vitamin D (VD) metabolites are involved in a variety of important metabolic processes and physiological effects in organisms. Profiling of VD metabolites favors a deep understanding of the physiological role of VD. However, VD metabolites are difficult to detect due to their high chemical structural rigidity, structural similarity, and low sensitivities under liquid chromatography-tandem mass spectrometry (LC-MS). Herein, we present a chemical derivatization assisted LC-MS/MS strategy for the detection of VDs, in which 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) is employed to derivatize the conjugated diene of VD metabolites and provides sensitizing reporters for MS detection. After PTAD derivatization, the sensitivities of seven VD metabolites increased by 24-276 folds, with the limits of detection ranging from 3 to 20 pg mL -1 . Using this method, we achieved a sensitive and accurate quantification of 7 VD metabolites (vitamin D 2 , vitamin D 3 , 25-hydroxyvitamin D 2 , 25-hydroxyvitamin D 3 , 1,25-dihydroxyvitamin D 2 , 1,25-dihydroxyvitamin D 3 , and 1,24,25-trihydroxyvitamin D 3 ) of the VD metabolic pathway in different trace biological samples, including human serum, mouse tissues (namely liver, kidney, lung, and spleen), and cells. We believe that the present method can provide a promising tool for an in-depth analysis of VD metabolism.

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are a class of bioactive lipids that show therapeutic potential for diabetes, anti-cancer and inflammation. These FAHFAs can be obtained through dietary intake, potentially improving human health. However, there is currently inadequate knowledge regarding the presence and variety of FAHFAs in different foods. Herein, we profile FAHFAs from 12 typical food samples and 4 medicinal food samples with the aid of our previous established chemical isotope labeling-assisted liquid chromatography-mass spectrometry method and build a comprehensive dataset of FAHFA diversity. The dataset comprised a total of 1207 regioisomers belonging to 298 different families, with over 100 families being newly discovered for the first time. Therefore, our findings contribute valuable insights into the molecular diversity and presence of FAHFA in a range of foods. This dataset serves as a foundation for further exploration of the nutritional and medicinal functions of FAHFAs., (© 2023. The Author(s).)

Background: Apolipoprotein B (apoB) is a crucial component that directly reflects the number of atherogenic lipoprotein particles and is closely related to atherosclerosis. However, there was an inconsistency among previous studies in its relationship with mortality. Using nationally representative data, we aimed to investigate the association of apoB with cardiovascular and all-cause mortality., Methods: We retrospectively included participants from the National Health and Nutrition Examination Survey (2007-2014), and mortality was ascertained through December 31, 2015. Hazard ratios (HRs) with 95% confidence intervals (CIs) of apoB in quartiles (Q1-Q4) for mortality risk were calculated using multivariable-adjusted Cox proportional hazards models, and restricted cubic spline regressions were performed to test dose relationships., Results: We enrolled 10,375 participants with a mean age of 46.3 years, of which 47.88% were men. During a mean follow-up time of 69.2 months, 533 (5.14%) and 91 (0.88%) deaths were due to all causes and cardiovascular disease, respectively. After adjusting for confounders, per SD, increment of apoB was associated with an elevated risk of cardiovascular mortality (HR, 1.13; 95% CI, 1.03-1.24). The risk of all-cause mortality was significantly reduced in the third quartile (Q3) of apoB (HR, 0.71; 95% CI, 0.56-0.91) compared with the reference quartile (Q1). Moreover, spline analyses showed that the relationship of apoB with all-cause mortality was U-shaped, and the threshold value was 108 mg/dL., Conclusions: ApoB was linearly associated with increased risk of cardiovascular mortality and non-linearly associated with all-cause mortality in a U-shaped manner, independently of other cardiovascular risk factors., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests., (Copyright © 2023 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.)

Objective: To explore the prognostic value of circulating tumor DNA (ctDNA) testing in patients with refractory/relapsed diffuse large B-cell lymphoma (R/R DLBCL) undergoing chimeric antigen receptor T-cell (CAR-T) therapy, and to guide the prevention and subsequent treatment of CAR-T-cell therapy failure. Methods: In this study, 48 patients with R/R DLBCL who received CAR-T-cell therapy at the First Affiliated Hospital of Zhejiang University School of Medicine between December 2017 and March 2022 were included. Furthermore, ctDNA testing of 187 lymphoma-related gene sets was performed on peripheral blood samples obtained before treatment. The patients were divided into complete remission and noncomplete remission groups. The chi-square test and t -test were used to compare group differences, and the Log-rank test was used to compare the differences in survival. Results: Among the patients who did not achieve complete remission after CAR-T-cell therapy for R/R DLBCL, the top ten genes with the highest mutation frequencies were TP53 (41%), TTN (36%), BCR (27%), KMT2D (27%), IGLL5 (23%), KMT2C (23%), MYD88 (23%), BTG2 (18%), MUC16 (18%), and SGK1 (18%). Kaplan-Meier survival analysis revealed that patients with ctDNA mutation genes >10 had poorer overall survival (OS) rate (1-year OS rate: 0 vs 73.8%, P <0.001) and progression-free survival (PFS) rate (1-year PFS rate: 0 vs 51.8%, P =0.011) compared with patients with ctDNA mutation genes ≤10. Moreover, patients with MUC16 mutation positivity before treatment had better OS (2-year OS rate: 56.8% vs 26.7%, P =0.046), whereas patients with BTG2 mutation positivity had poorer OS (1-year OS rate: 0 vs 72.5%, P =0.005) . Conclusion: ctDNA detection can serve as a tool for evaluating the efficacy of CAR-T-cell therapy in patients with R/R DLBCL. The pretreatment gene mutation burden, mutations in MUC16 and BTG2 have potential prognostic value.

Peak alignment is a crucial step in liquid chromatography-mass spectrometry (LC-MS)-based large-scale untargeted metabolomics workflows, as it enables the integration of metabolite peaks across multiple samples, which is essential for accurate data interpretation. Slight differences or fluctuations in chromatographic separation conditions, however, can cause the chromatographic retention time (RT) shift between consecutive analyses, ultimately affecting the accuracy of peak alignment between samples. Here, we introduce a novel RT shift correction method based on the retention index (RI) and apply it to peak alignment. We synthesized a series of N -acyl glycine (C2-C23) homologues via the amidation reaction between glycine with normal saturated fatty acids (C2-C23) as calibrants able to respond proficiently in both mass spectrometric positive- and negative-ion modes. Using these calibrants, we established an N -acyl glycine RI system. This RI system is capable of covering a broad chromatographic space and addressing chromatographic RT shift caused by variations in flow rate, gradient elution, instrument systems, and LC separation columns. Moreover, based on the RI system, we developed a peak shift correction model to enhance peak alignment accuracy. Applying the model resulted in a significant improvement in the accuracy of peak alignment from 15.5 to 80.9% across long-term data spanning a period of 157 days. To facilitate practical application, we developed a Python-based program, which is freely available at https://github.com/WHU-Fenglab/RI-based-CPSC.

N 6 -Methyladenine (6mA) is a naturally occurring DNA modification in both prokaryotes and eukaryotes. Herein, we developed a deaminase-mediated sequencing (DM-seq) method for genome-wide mapping of 6mA at single-nucleotide resolution. The method capitalizes on the selective deamination of adenine, but not 6mA, in DNA mediated by an evolved adenine deaminase, ABE8e. By employing this method, we achieved genome-wide mapping of 6mA in Escherichia coli and in mammalian mitochondrial DNA (mtDNA) at single-nucleotide resolution. We found that the 6mA sites are mainly located in the GATC motif in the E. coli genome. We also identified 17 6mA sites in mtDNA of HepG2 cells, where all of the 6mA sites are distributed in the heavy strand of mtDNA. We envision that DM-seq will be a valuable tool for uncovering new functions of 6mA in DNA and for exploring its potential roles in mitochondria-related human diseases., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Central carbon metabolism pathway (CCM) is one of the most important metabolic pathways in all living organisms and play crucial function in aspect of organism life. However, the simultaneous detection of CCM intermediates remains challenging. Here, we developed a chemical isotope labeling combined with LC-MS method for simultaneous determination of CCM intermediates with high coverage and accuracy. By chemical derivatization with 2-(diazo-methyl)-N-methyl-N-phenyl-benzamide (2-DMBA) and d 5 -2-DMBA, all CCM intermediates obtain better separation and accurate quantification at a single LC-MS run. The obtained limits of detection of CCM intermediates ranged from 5 to 36 pg/mL. Using this method, we achieved simultaneous and accurate quantification of 22 CCM intermediates in different biological samples. Take account of the high detection sensitivity of the developed method, this method was further applied to the quantification of CCM intermediates at single-cell level. Finally, 21 CCM intermediates were detected in 1000 HEK-293T cells and 9 CCM intermediates were detected in mouse kidney glomeruli optical slice samples (10∼100 cells)., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Objectives: In this study we aimed to evaluate the nocebo response rate in patients with functional dyspepsia (FD) and to explore its influencing factors., Methods: A literature search of the EMBASE, PubMed, and Cochrane Library databases was conducted for all articles published up to March 2021. Randomized, parallel-designed, placebo-controlled trials on pharmacological interventions for patients with FD were included. A meta-analysis that utilized random effects to analyze the incidence of adverse events (AEs) among participants who were given placebo was conducted, and the correlation between trial characteristics and the magnitude of the nocebo response rate was analyzed., Results: Altogether, 27 studies including 1866 paitents were deemed eligible and included in the analysis. The total nocebo response rate was 26% (95% confidence interval [CI] 18%-33%). The most frequently reported AEs included nasopharyngitis (9%), constipation (6%), headache (5%), and diarrhea (3%). There were significant differences in nocebo response rates among studies conducted in different country or region, treatment duration, types of medication, sponsorship and different versions of the Rome criteria used for FD diagnosis. While number of centers engaged in the study, types of FD diagnosis and dosing frequency were not significantly associated with the nocebo response rate., Conclusions: Patients with FD exhibit notable nocebo response strength in clinical trials. The researchers should adopt a more careful approach when analyzing the relationships between AEs and interventions in such trials., (© 2023 Chinese Medical Association Shanghai Branch, Chinese Society of Gastroenterology, Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine and John Wiley & Sons Australia, Ltd.)

Gut microbiota-host co-metabolites serve as essential mediators of communication between the host and gut microbiota. They provide nutrient sources for host cells and regulate gut microenvironment, which are associated with a variety of diseases. Analysis of gut microbiota-host co-metabolites is of great significance to explore the host-gut microbiota interaction. In this study, we integrated chemical derivatization, liquid chromatography-mass spectrometry, and molecular networking (MN) to establish a novel CD-MN strategy for the analysis of carboxylated metabolites in gut microbial-host co-metabolism. Using this strategy, 261 carboxylated metabolites from mouse feces were detected, which grouped to various classes including fatty acids, bile acids, N -acyl amino acids, benzoheterocyclic acids, aromatic acids, and other unknown small-scale molecular clusters in MN. Based on the interpretation of the bile acid cluster, a novel type of phenylacetylated conjugates of host bile acids was identified, which were mediated by gut microbiota and exhibited a strong binding ability to Farnesoid X receptor and Takeda G protein-coupled receptor 5. Our proposed strategy offers a promising platform for uncovering carboxylated metabolites in gut microbial-host co-metabolism.

N 6 -Methyladenosine (m 6 A) is one of the most abundant and prevalent natural modifications occurring in diverse RNA species. m 6 A plays a wide range of roles in physiological and pathological processes. Revealing the functions of m 6 A relies on the faithful detection of individual m 6 A sites in RNA. However, developing a simple method for the single-base resolution detection of m 6 A is still a challenging task. Herein, we report an adenosine deamination sequencing (AD-seq) technique for the facile detection of m 6 A in RNA at single-base resolution. The AD-seq approach capitalizes on the selective deamination of adenosine, but not m 6 A, by the evolved tRNA adenosine deaminase (TadA) variant of TadA8e or the dimer protein of TadA-TadA8e. In AD-seq, adenosine is deaminated by TadA8e or TadA-TadA8e to form inosine, which pairs with cytidine and is read as guanosine in sequencing. m 6 A resists deamination due to the interference of the methyl group at the N 6 position of adenosine. Thus, the m 6 A base pairs with thymine and is still read as adenosine in sequencing. The differential readouts from A and m 6 A in sequencing can achieve the single-base resolution detection of m 6 A in RNA. Application of the proposed AD-seq successfully identified individual m 6 A sites in Escherichia coli 23S rRNA. Taken together, the proposed AD-seq allows simple and cost-effective detection of m 6 A at single-base resolution in RNA, which provides a valuable tool to decipher the functions of m 6 A in RNA.

Intense ultraviolet (UV) exposure can cause phototoxic reactions, such as skin inflammation, resulting in injury. UV is a direct cause of DNA damage, but the mechanisms underlying transcriptional regulation within cells after DNA damage are unclear. The bioinformatics analysis of transcriptome sequencing data from UV-irradiated and non-UV-irradiated skin showed that transcription-related proteins, such as HSF4 and COIL, mediate cellular response to UV irradiation. HSF4 and COIL can form a complex under UV irradiation, and the preference for binding target genes changed because of the presence of a large number of R-loops in cells under UV irradiation and the ability of COIL to recognize R-loops. The regulation of target genes was altered by the HSF4-COIL complex, and the expression of inflammation and ageing-related genes, such as Atg7, Tfpi, and Lims1, was enhanced. A drug screen was performed for the recognition sites of COIL and R-loop. N6-(2-hydroxyethyl)-adenosine can competitively bind COIL and inhibit the binding of COIL to the R-loop. Thus, the activation of downstream inflammation-related genes and inflammatory skin injury was inhibited., (© 2023 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Purpose: The present study aimed to assess the association of elevated serum uric acid (SUA) and hypouricemia with all-cause mortality and cardiovascular mortality in Chinese hypertensive patients., Methods: In the present prospective cohort, 9325 hypertensive patients from Dongguan, China were enrolled from 2014 to 2018 for analysis. Participants were categorised by quintiles of SUA. The HRs and 95% CIs for the association between SUA, all-cause and cardiovascular mortality were evaluated using the multivariate Cox regression model. After adjusting for multiple confounders, restricted cubic spline analysis was conducted to demonstrate the shape of relationship., Results: After a median follow-up of 4.18 years for 9325 participants, there were 409 (4.4%) and 151 (1.6%) reported cases of all-cause and cardiovascular mortality, respectively. By using the third quintile of SUA (6.68 mg/dL to <7.55 mg/dL for men, 5.63 mg/dL to <6.42 mg/dL for women) as reference, the highest quintiles of SUA were associated with an elevated risk of all cause (HR: 1.34, 95% CI 1.00 to 1.80) in the crude model, but the association was not significant after adjusting for multiple comparisons. The association between low SUA and mortality and the dose-response analysis on the non-linearity of SUA-mortality relationship were not statistically significant., Conclusions: Although the association between SUA levels, all-cause and cardiovascular disease mortality did not appear to be significant among Chinese hypertensive patients, the findings might be confounded by their medical conditions. Further studies are needed to verify the optimal SUA levels for hypertensive patients., (© The Author(s) 2022. Published by Oxford University Press on behalf of Postgraduate Medical Journal. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Di(2-ethylhexyl) phthalate (DEHP), the most widely used plasticizers in the world, has been regarded as an endocrine disrupting chemical with serious adverse health outcomes. Accumulating evidence strongly suggests that the undesirable biological effects of DEHP are meditated by its metabolites rather than itself. However, the metabolic footprints of DEHP in vivo are still unclear. Here we developed a click chemistry-assisted mass spectrometry (CC-MS) strategy for in-depth profiling DEHP metabolites in rats. An alkyne-modified DEHP analogue (alkyne-DEHP) was synthesized as a tracer for in vivo tracing, and a pair of MS probes (4-azido-nphenylbenzamide, 4-ANPA, and its deuterated reagent d 5 -4-ANPA) were prepared to specifically label the alkyne-DEHP metabolites, and prominently improve their detection sensitivity and selectivity. Using the CC-MS strategy, we successfully screened 247 alkyne-DEHP metabolites from rat urine, feces, and serum, including many unrevealed metabolites, such as oxidized phthalate diester metabolites and glucuronides of phthalate monoester metabolites. The discovery of new DEHP metabolites provides additional insights for understanding the metabolism of DEHP, which may be beneficial in exploring the mechanism underlying DEHP induced-toxicity in the future., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reports in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Pine wilt disease (caused by the nematode Bursaphelenchus xylophilus) is extremely harmful to pine forests in East Asia. As a low-resistance pine species, Pinus thunbergii is more vulnerable to pine wood nematode (PWN) than Pinus densiflora and Pinus massoniana. Field inoculation experiments were conducted on PWN-resistant and -susceptible P. thunbergii, and the difference in transcription profiles 24 h after inoculation was analyzed. We identified 2603 differentially expressed genes (DEGs) in PWN-susceptible P. thunbergii, while 2559 DEGs were identified in PWN-resistant P. thunbergii. Before inoculation, DEGs between PWN-resistant and PWN-susceptible P. thunbergii were enriched in the REDOX (Oxidation-Reduction) activity pathway (152 DEGs), followed by the oxidoreductase activity pathway (106 DEGs). After inoculation with PWN, however, the opposite was observed; DEGs were enriched in the oxidoreductase activity pathway (119 DEGs), followed by the REDOX activity pathway (84 DEGs). Before inoculation, according to the metabolic pathway analysis results, we found more genes upregulated in phenylpropanoid metabolic pathways and enriched in lignin synthesis pathways; cinnamoyl-CoA reductase-coding genes related to lignin synthesis were upregulated in PWN-resistant P. thunbergii and downregulated in PWN-susceptible P. thunbergii, and the lignin content was always higher in resistant than in susceptible P. thunbergii. These results reveal distinctive strategies of resistant and susceptible P. thunbergii in dealing with PWN infections., (© The Author(s) 2023. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Chemical modifications in DNA have profound influences on the structures and functions of DNA. Uracil, a naturally occurring DNA modification, can originate from the deamination of cytosine or arise from misincorporation of dUTP into DNA during DNA replication. Uracil in DNA will imperil genomic stability due to their potential in producing detrimental mutations. An in-depth understanding of the functions of uracil modification requires the accurate determination of its site as well as content in genomes. Herein, we characterized that a new member of the uracil-DNA glycosylase (UDG) family enzyme (UdgX-H109S) could selectively cleave both uracil-containing single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA). Based on this unique property of UdgX-H109S, we developed an enzymatic cleavage-mediated extension stalling (ECES) method for the locus-specific detection and quantification of uracil in genomic DNA. In the ECES method, UdgX-H109S specifically recognizes and cleaves the N -glycosidic bond of uracil from dsDNA and generates an apurinic/apyrimidinic (AP) site, which could be broken by APE1 to form a one-nucleotide gap. The specific cleavage by UdgX-H109S is then evaluated and quantified by qPCR. With the developed ECES approach, we demonstrated that the level of uracil at position Chr4:50566961 in genomic DNA of breast cancer tissues was significantly decreased. Collectively, the ECES method has been proved to be accurate and reproducible in the locus-specific quantification of uracil in genomic DNA from biological and clinical samples.

The link between lead and blood pressure was debatable, and whether it was mediated by renal function was unknown. The purpose was to investigate the relationship between blood lead concentrations and blood pressure and hypertension, as well as the mediating role of estimated glomerular filtration rate (eGFR) in this relationship. Participants aged 18 were recruited from the National Health and Nutrition Examination Survey (1999-2014) and provided with lead and blood pressure data. Multivariate linear and logistic regression, stratification, interaction tests, and a restricted cubic spline curve were used to assess the association of blood lead with systolic/diastolic blood pressure (SBP/DBP) and hypertension, and mediation effect analysis was used to investigate the role of eGFR in this relationship. A total of 20,073 subjects were enrolled, and among them, 9837 (49.01%) were male and 7800 (38.86%) were hypertensive patients. Multivariate linear and logistic regression analysis showed that blood lead levels were significantly associated with SBP (β = 3.14, 95%CI: 2.03, 4.25; P < 0.001), DBP (β = 3.50, 95%CI: 2.69, 4.30; P < 0.001), and hypertension (OR = 1.29, 95%CI: 1.09, 1.52; P = 0.0026). In comparison to the lowest blood lead quartile, the highest lead group was significantly associated with SBP (= 2.55, 95%CI: 1.66, 3.44; P = 0.0001), DBP (= 2.60, 95%CI: 1.95, 3.24; P = 0.0001), and hypertension (OR = 1.26, 95%CI: 1.10, 1.45; P = 0.0007). Mediation analysis showed that the proportion of blood lead mediated for SBP, DBP, and hypertension was 3.56% (95%CI: 0.42%, 7.96%; P = 0.0320), 6.21% (95%CI: 4.02%, 9.32%; P < 0.0001), and 17.39% (95%CI: 9.34%, 42.71%; P < 0.0001), respectively. Adjusted restricted cubic spline curves presented a non-linear correlation of blood lead levels with DBP (P-non-linearity < 0.001), linear with SBP (P-non-linearity = 0.203), and hypertension (P-non-linearity = 0.763). Our findings demonstrated that blood lead levels were non-linear with DBP, but linear with SBP and hypertension, and this relationship was mediated by eGFR., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Abnormal tumor vasculature blocks the extravasation of T lymphocytes into the tumor, thereby suppressing anti-tumor immunity. Recently, metformin has been shown to affect tumor vasculature and enhance T lymphocyte anti-tumor immunity. However, whether or how metformin affects T lymphocyte anti-tumor immunity via a vascular mechanism remains poorly understood. Herein, we show that a large number of CD8 + lymphocytes gathered in the peri-tumoral region, while very few infiltrated the tumor. Metformin administration increased the expression of anti-tumor immunity-associated genes and the number of tumor-infiltrating CD8 + lymphocytes. Injection of CD8 but not CD4 neutralization antibody into tumor-bearing mice significantly abrogated the anti-tumor effect of metformin. Critically, CD8 + lymphocytes were found to pass through the wall of perfused vessel. Further results of immunofluorescent staining showed that metformin greatly elevated tumor perfusion, which was accompanied by increased vascular maturity in the intratumoral region (ITR) but not peritumoral region (PTR). These findings provide evidence for the vascular mechanism involved in metformin-induced enhancement of T lymphocyte anti-tumor immunity. By remodeling the abnormal tumor vasculature, also called vessel normalization metformin increases vascular maturity and tumor perfusion, thus allowing more CD8 + lymphocytes to infiltrate the tumor., Competing Interests: Declaration of competing interest No potential conflicts of interest were disclosed by all the contributing authors., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)