Metabolic Characteristics Research Articles

5045 The Effects Of Non-functioning Adrenal Incidentalomas On Metabolic Comorbidities And Clinical Characteristics Changes: A Systematic Review And Meta-Analysis

Abstract Disclosure: X. Li: None. H. Lan: None. J. Wen: None. G. Chen: None. W. Lin: None. The Effects of Non-functioning Adrenal Incidentalomas on Metabolic Comorbidities and Clinical Characteristics Changes: A Systematic Review and Meta-analysis. Context: Adrenal incidentalomas (AI) are frequently detected during abdominal imaging, with non-functioning adrenal incidentalomas (NFAI) being particularly common, even in elderly individuals. This meta-analysis investigates the influence of NFAI on metabolic comorbidities, comparing non-surgical and surgical patient groups, and assessing alterations in NFAI characteristics. Design: Our comprehensive meta-analysis utilized data from multiple databases, encompassing studies from January 2000 to August 21, 2023.Setting: Data sources included PubMed, Embase, Cochrane Library, and Web of Science. Participants: The analysis comprises 24 articles involving 2,770 NFAI patients, categorized into non-surgical and surgical groups. Intervention: The non-surgical group underwent no surgery, while the surgical group underwent tumor resection. Main Outcome Measure: The study investigated changes in metabolic comorbidities (hypertension, diabetes, obesity, lipid metabolism disorders), tumor characteristics, and hormone production in NFAI patients during follow-up or after surgery. Results: In the non-surgical NFAI patient group (20 studies), metabolic comorbidities exhibited a discernible upward trajectory during follow-up. The initial hypertension prevalence of 58.3% increased by 4.2% over a 4-5 year follow-up period. Moreover, diabetes, obesity, and lipid metabolism disorders experienced significant increases of 5.8%, 2.2%, and 8.8%, respectively. In contrast, the surgical group (6 studies) demonstrated a notable reduction in hypertension, with 43.9% of initially hypertensive patients achieving hypertension-free status after surgery. As for tumor changes in the non-surgical group, they were generally inconsequential, with only 5% of patients witnessing tumor growth exceeding 10mm over approximately 3.3 years, and 8% undergoing functional changes. These findings underscore the divergent impact of non-surgical and surgical interventions on metabolic comorbidities in NFAI patients. Conclusion: The study advocates for regular monitoring of blood glucose and blood pressure in NFAI patients, especially those with metabolic comorbidities. When necessary, adenoma resection should be considered. Further investigation is warranted to elucidate the mechanisms connecting NFAI with metabolic factors. Presentation: 6/1/2024

12500 Objective Evaluation Of Clustering Methods For Resolving The Heterogeneity Of PCOS

Abstract Disclosure: K. Brewer: None. R. Sisk: None. M. Dapas: None. C. Li: None. A. Dunaif: Consulting Fee; Self; AcaciaBio, Inc, Neurocine Biosciences, Inc. Speaker; Self; Quest Diagnostics. Other; Self; Co-Editor Endocrine Today, Healio, Slack Inc. Clustering methods have been used to resolve heterogeneity within complex diseases to elucidate underlying biologic mechanisms. We (Dapas et al. PLoS Med, 2020) and others have applied these methods to PCOS and identified discrete subtypes, including those that capture distinct reproductive or metabolic features. We objectively compared two widely used methods, hierarchical clustering (HC), which recursively merges subjects based on their similarity, and k-means (Km), which iteratively groups individuals to minimize their distance to cluster centroids. The number of clusters (k) was predefined in both approaches. We compared HC vs Km in 874 European ancestry PCOS cases diagnosed by NIH criteria using 8 traits (BMI, testosterone, SHBG, DHEAS, LH, FSH, fasting insulin, fasting glucose) and 9 traits with the addition of AMH. The ConsensusClusterPlus R package was used to evaluate cluster stability of both approaches with k=2, 3, or 4, using 8 or 9 traits. Genomewide association study (GWAS) meta-analysis was performed as reported with a discovery cohort (620 cases, 2951 controls) and a replication cohort (371 cases, 926 controls), except that genotypes were imputed to the newer TOPMED (r2) panel. Clustering using 8 traits and k=3 resulted in the best cluster stability for both HC and Km (pairwise consensus proportion: HC 0.98, Km 0.93) compared to k=2 with 8 traits (HC 0.88, Km 0.88) or 9 traits (HC 0.95, Km 0.92). No stability was observed for k=4 with 8 or 9 traits using either method (range: 0.35-0.66). Km showed slightly better cluster separation than HC (average silhouette width: Km 0.12 vs HC 0.09) with k=3 and 8 traits. The biologic relevance of the three PCOS subtypes, which we have designated as reproductive (cases with higher LH, FSH, SHBG), metabolic (higher BMI, insulin, glucose), and background, was assessed by GWAS, an orthogonal (i.e., uncorrelated) approach for subtype confirmation. The subtypes identified with HC had genomewide significant associations (metabolic subtype, c9orf3/FANCC, rs10761370, minor allele frequency [MAF] 0.47, p=1.21 x 10-8; background subtype, FSHB/ARL14EP, rs10835649, MAF 0.17, p=8.49 x 10-[1]0). There were no genomewide significant signals when the subtypes were identified with Km despite having a better cluster separation. In summary, only HC clusters appeared to capture biologically meaningful differences since two of the three subtypes thus identified were associated with genomewide significant loci. Neither the addition of AMH nor increasing the number of groups improved clustering metrics. Our results emphasize the importance of independent validation of clustering approaches using an orthogonal confirmation strategy such as GWAS. We conclude that HC clustering using 8 traits is superior to Km for resolving the genetic heterogeneity of PCOS. Presentation: 6/2/2024

12500 Objective Evaluation Of Clustering Methods For Resolving The Heterogeneity Of PCOS

Abstract Disclosure: K. Brewer: None. R. Sisk: None. M. Dapas: None. C. Li: None. A. Dunaif: Consulting Fee; Self; AcaciaBio, Inc, Neurocine Biosciences, Inc. Speaker; Self; Quest Diagnostics. Other; Self; Co-Editor Endocrine Today, Healio, Slack Inc. Clustering methods have been used to resolve heterogeneity within complex diseases to elucidate underlying biologic mechanisms. We (Dapas et al. PLoS Med, 2020) and others have applied these methods to PCOS and identified discrete subtypes, including those that capture distinct reproductive or metabolic features. We objectively compared two widely used methods, hierarchical clustering (HC), which recursively merges subjects based on their similarity, and k-means (Km), which iteratively groups individuals to minimize their distance to cluster centroids. The number of clusters (k) was predefined in both approaches. We compared HC vs Km in 874 European ancestry PCOS cases diagnosed by NIH criteria using 8 traits (BMI, testosterone, SHBG, DHEAS, LH, FSH, fasting insulin, fasting glucose) and 9 traits with the addition of AMH. The ConsensusClusterPlus R package was used to evaluate cluster stability of both approaches with k=2, 3, or 4, using 8 or 9 traits. Genomewide association study (GWAS) meta-analysis was performed as reported with a discovery cohort (620 cases, 2951 controls) and a replication cohort (371 cases, 926 controls), except that genotypes were imputed to the newer TOPMED (r2) panel. Clustering using 8 traits and k=3 resulted in the best cluster stability for both HC and Km (pairwise consensus proportion: HC 0.98, Km 0.93) compared to k=2 with 8 traits (HC 0.88, Km 0.88) or 9 traits (HC 0.95, Km 0.92). No stability was observed for k=4 with 8 or 9 traits using either method (range: 0.35-0.66). Km showed slightly better cluster separation than HC (average silhouette width: Km 0.12 vs HC 0.09) with k=3 and 8 traits. The biologic relevance of the three PCOS subtypes, which we have designated as reproductive (cases with higher LH, FSH, SHBG), metabolic (higher BMI, insulin, glucose), and background, was assessed by GWAS, an orthogonal (i.e., uncorrelated) approach for subtype confirmation. The subtypes identified with HC had genomewide significant associations (metabolic subtype, c9orf3/FANCC, rs10761370, minor allele frequency [MAF] 0.47, p=1.21 x 10-8; background subtype, FSHB/ARL14EP, rs10835649, MAF 0.17, p=8.49 x 10-[1]0). There were no genomewide significant signals when the subtypes were identified with Km despite having a better cluster separation. In summary, only HC clusters appeared to capture biologically meaningful differences since two of the three subtypes thus identified were associated with genomewide significant loci. Neither the addition of AMH nor increasing the number of groups improved clustering metrics. Our results emphasize the importance of independent validation of clustering approaches using an orthogonal confirmation strategy such as GWAS. We conclude that HC clustering using 8 traits is superior to Km for resolving the genetic heterogeneity of PCOS. Presentation: 6/2/2024

8415 Rapid Generation Of An In Vivo Loss-Of-Function Genetic Model Of sdhb Zebrafish: Functional And Metabolic Characterization

Abstract Disclosure: S. Parisien-La Salle: None. F. Nobilleau: None. J. Lamontagne: None. S. Éric: None. I. Bourdeau: None. Introduction: Animal research has been limited in pheochromocytomas and paragangliomas (PPGLs). Recently, Dona et al., described a sdhb mutated zebrafish model by introducing a 13 bp frameshift mutation at the sdhb exon1—intron1 boundary into the zebrafish sdhb gene that recapitulated features of PPGLs. Objective: To generate a new model of mutated sdhb zebrafish to further investigate the underlying metabolic and biochemical perturbations in vivo. Methods: We took advantage of the genetic accessibility of zebrafish embryo to generate loss-of-function model for sdhb using CRISPR/Cas9 technology. Three guide RNAs were designed and microinjected in one-cell stage zebrafish embryos to target the coding sequence of zebrafish sdhb gene. In order to control for the stress brought on by injections, we injected a group of zebrafish with Cas9 endonuclease as a control. At five days post fertilization, mutant and control zebrafish were flash frozen and sent for liquid chromatography mass spectrometry measurements of Krebs cycle metabolites and normetanephrines/metanephrines. All results were normalized to the control groups consisting of wild-type or Cas9 injected larvae. Sdhb expression was analyzed by RT-qPCR in sdhb mutants and wild-type zebrafish larvae. We also monitored survival and heart rate in sdhb mutant zebrafish, compared to wild-type and Cas9. Results: Firstly, we confirmed the significant reduction of sdhb expression in CRISPR-injected larvae compared to wild-type larvae at 5 dpf (0.0613 vs 1.000 p<0.0001). Our metabolic panel showed that succinate was up to 26 times more elevated in sdhb mutants than in wild-type fish extracts (sdhb: 26.51 p<0.0001). Lactate (sdhb: 5.27 p<0.0001), leucine (sdhb: 2.88 p<0.0001), arginine (sdhb: 1.50 p=0.0034) and HMG-CoA (sdhb: 1.36 p=0.0033) were also higher in sdhb mutants. Whereas aspartate (sdhb: 0.244 p<0.0001) and oxidized gluthiatone (sdhb: 0.64 p=0.0011) were significantly lower in sdhb mutants than in wild-types. In regards to functionality, normetanephrine and metanephrine levels were at least four folds higher in sdhb mutants than in Cas9 injected larvaes (4.06 p<0.0001 and 3.42 p<0.0001 respectively). Interestingly, this difference was more noticeable in the fish bathing medium than in larvae tissue extracts, suggesting an increased hormonal excretion (normetanephrines: 4.81 p<0.0001 and metanephrines: 9.40 p<0.0001). Finally, sdhb mutated zebrafish presented with a higher heart rate (148 vs 133 p<0.0001) and reduced lifespan (p<0.0001) when compared to the wild-type/Cas9 group. Conclusion: These findings confirm that the zebrafish is an ideal model for studying PPGLs and their associated genetic defects. Our approach allows the in vivo assessment of a large metabolic panel, opening avenues on new potential clinical metabolic biomarkers in a loss-of-function of the SDHB gene. Ref: Dona M, et al. Endocr Relat Cancer. 2021 Presentation: 6/1/2024

6889 Potential Impact of Parental Origin of Inheritance in Presentation of Familial Partial Lipodystrophy

Abstract Disclosure: D. Gilio: None. M.C. Foss de Freitas: Advisory Board Member; Self; PTC Therapeutics. Speaker; Self; Amryt. O. Besci: None. M. Celik Guler: None. A. Neidert: None. I. Yildirim: None. B. Akinci: Advisory Board Member; Self; Amryt. Consulting Fee; Self; Regeneron Pharmaceuticals, Alnylam Pharmaceuticals, Amryt, AstraZeneca, Boehringer Ingelheim, Lilly, MSD, Novartis Pharmaceuticals, Novo Nordisk, Sanofi-Aventis, Servier, Third Rock Ventures. E.A. Oral: Advisory Board Member; Self; Amryt, Regeneron Pharmaceuticals. Consulting Fee; Self; Regeneron Pharmaceuticals, Amryt, Third Rock Ventures. Grant Recipient; Self; Regeneron Pharmaceuticals, Amryt. Research Investigator; Self; Novo Nordisk, Fractyl, Ionis Pharmaceuticals Inc., GI Dynamics, Rhythm Pharmaceuticals. Other; Self; Amryt. Background: Familial partial lipodystrophy Type 2 (FPLD2) caused by autosomal dominant LMNA variants is characterized by partial loss of subcutaneous adipose tissue in the limbs, trunk, and gluteal region with accumulation in other parts of the body, such as the face and neck. It is usually associated with different degrees of insulin resistance and hypertriglyceridemia and body composition patterns. We wanted to know the contributing factors for the different metabolic abnormalities and body composition presentations of FPLD2. We hypothesized that parental inheritance line may be a contributory factor to the expression of the metabolic phenotype and body composition characteristics. Methods: We retrospectively collected clinical data of 77 FPLD2 cases from the Divisions of Metabolism and Endocrinology of 3 different sites: the University of Michigan, the University of São Paulo and The Turkish Lipodystrophy Study Group (TuLip). We further selected 53 published cases curated from 15 studies in the literature. For inclusion of the studies, we used web search tools including PubMed, OMIM and Google Scholar. Only patients where we could assess the genetic inheritance were included in this study. Data collected included demographic information, medical history and laboratory results. Results: 79 patients inherited the LMNA variant from the mother (Group 1; 18 males and 61 females), and 51 patients inherited it from the father (Group 2; 14 males and 37 females). The median age at lipodystrophy diagnosis was lower in Group 1 than in Group 2. Diabetes was reported in more than half of the groups; however, the prevalence rates were not significantly different. The mean age (± SEM) at first diagnosis of diabetes was 30 ± 15 years in Group 1 and 32 ± 11 years in Group 2; the microvascular complications were present in 8 patients with diabetes in Group 1 (36.3%) and in 5 patients in Group 2 (23.8%). Both groups' mean triglycerides levels were >150 mg/dL, with a trend of higher triglycerides in Group 2 than in Group 1. Episodes of acute pancreatitis were identified in 9 patients (19.5%) of Group 1 and 10 patients (31.2%) of Group 2; however, patients in Group 1 showed 1.7 episodes of pancreatitis per person while Group 2 showed 8.1 episodes per person on average. The fatty liver disease was the most common liver abnormality identified and it seemed to be more frequent in Group 2 than in Group 1. There was no difference between the two groups in relation to the body mass index (BMI) and the ratio of percent fat mass of the trunk to percent fat mass of the legs (FMR); however, the DEXA (dual x-ray absorptiometry) showed a higher total fat mass in Group 1 compared to Group 2 (22.2 versus 17.3 Kg, p=0.040) in the presence of comparable total lean mass. Conclusion: Our data suggest that there is a difference in body fat mass according to the genetic inheritance, however the metabolic impact is not completely clear and require more detailed analysis. Presentation: 6/1/2024

Immune-related glycosylation genes based classification predicts prognosis and therapy options of osteosarcoma

Osteosarcoma is the most common primary bone malignancy, with a very poor prognosis. Aberrant glycosylation is close involvement in osteosarcoma. Accordingly, this study aimed at investigating the role of glycosylation genes in the prognosis and therapy options of osteosarcoma. The microenvironment of osteosarcoma was assessed using estimate algorithm. A total of 20 immune-related glycosylation genes (IRGGs) was identified using Pearson correlation analysis. Accordingly, osteosarcoma patients were divided into C1 and C2 type using consensus clustering. Multiple algorithms (Xcell, MCP-counter, ssGSEA, epic, quantiseq), cancer immune cycle analysis, and GSVA were applied to estimate the immune, molecule and metabolism characteristics of osteosarcoma, indicating that C1 type was featured with high immune infiltration, high glycosylation, enriched MEK signaling, and good prognosis, while C2 type was characterized by more metastasis, enriched immunotherapy-positive gene signatures, high tumor mutation burden, and poor prognosis. Results from TIDE algorithm and immunotherapy datasets suggested the C2 type’s preference of immune checkpoint inhibitors (ICIs), while data of GDSC, CMap analysis and cell experiments indicated that C1 type was sensitivity to MEK inhibitor PD0325901. In addition, univariate Cox and Lasso analysis was combined to establish an IRGGs’ risk score containing 6 genes (B3GNT8, FUT7, GAL3ST4, GALNT14, HS3ST2, and MFNG). The data of DCA and ROC indicated its well prediction of prognosis in osteosarcoma. Finally, cellular location analysis showed that the 6 genes not only distributed in tumor cells but also in immune cells. In summary, the classification and risk score based on IRGGs effectively predicted the prognosis and therapy options of osteosarcoma. Further studies on IRGGs may contribute to the understanding of cancer immunity in osteosarcoma.

8532 Circulating Neprilysin in Pregnancy

Abstract Disclosure: A. Kulovic-Sissawo: None. E. Weiss: None. E. Jantscher-Krenn: None. U. Hiden: None. Background: In the complex nature of pregnancy, the female organism undergoes pivotal vascular and metabolic changes orchestrated by a tightly regulated interplay of bioactive regulatory molecules. The neutral endopeptidase, neprilysin, exists both in a membrane-bound and a soluble form (sNEP), of with the latter present in the circulatory system. Neprilysin degrades an exciting range of bioactive peptides involved in glycemic control (insulin B chain, GLP-1) and vascular tone regulation (angiotensin I and II, bradykinin, ANP, BNP), and both of them are altered in pregnancy. Objective: Due to the substrate-specificity of neprilysin and its multifaceted role we hypothesize that neprilysin may be involved in metabolic and vascular adaptions during pregnancy. However, the link between sNEP and maternal metabolism in pregnancy has not yet been investigated. Methodology: Serum sNEP concentration (n=102), longitudinally collected from metabolically healthy women (trial number NTC05496712), was measured at three visits throughout gestation (gestational age: 12.5±0.7; 20±0.8; 33±1.6 weeks) and post-partum (2±1.2 days after delivery) using commercially available sandwich ELISA. In addition, sNEP levels in pregnancy were compared to a cohort of non-pregnant women within reproductive age (n=50). Clinical and anthropometric measurements were determined and the association of sNEP and metabolic characteristics elucidated. Results: Circulating sNEP levels were non-normally distributed (median ± IQR: 2334±232624 pg/ml, n=82, gestational age 12.5±0.7 weeks) and showed high biological variability, with individual levels remaining constant over gestation. After delivery, however, sNEP concentration decreased (median± IQR: 1968±171060 pg/ml n=82, p=0.0001). sNEP showed a positive correlation with systolic blood pressure (Spearman, n=56, r=0.321, p=0.02) at early stages of pregnancy and non-fasting GLP-1 (Spearman, n=98, r=0.309, p=0.002) throughout gestation, but not with blood glucose, insulin resistance or BMI. In the non-pregnant state sNEP concentration (median±IQR: 4322±392152 pg/ml, n=38) positively correlated with body fat percentage (Pearson, n=38, r=0.405, p=0.01), fasting glucose (Pearson, n=43, r=0.301, p=0.05) and body mass index (Spearman, n=38, r=0.369, p=0.02) but did not correlate with systolic blood pressure (Pearson, n=36, r=-0.120, p=0.48). Conclusion: For the first time, the link between sNEP, metabolism and blood pressure during uncomplicated pregnancy was investigated. The decrease in post-partum circulating sNEP as compared to levels throughout pregnancy suggests a particular way of sNEP regulation and release during gestation. Our data indicate that in both, pregnant and non-pregnant state, sNEP may be involved in metabolic and cardiovascular regulation, with circulating sNEP levels showing massive individual differences. Presentation: 6/3/2024

Characteristics of Myocardial Structure and Central Carbon Metabolism during the Early and Compensatory Stages of Cardiac Hypertrophy.

Cardiac hypertrophy is a classical forerunner of heart failure and myocardial structural and metabolic remodeling are closely associated with cardiac hypertrophy. We aim to investigate the characteristics of myocardial structure and central carbon metabolism of cardiac hypertrophy at different stages. Using echocardiography and pathological staining, early and compensatory cardiac hypertrophy were respectively defined as within 7 days and from 7 to 14 days after transverse aortic constriction (TAC) in mice. Among mass-spectrometry-based metabolomics, we identified 45 central carbon metabolites. Differential metabolite analysis showed that six metabolites, including citrate, cis-aconitate and so on, decreased significantly on day 1 after TAC. Ten metabolites, including l-lactate, (S)-2-hydroxyglutarate and so on, were obviously changed on days 10 and 14. Pathway analysis showed that these metabolites were involved in seven metabolic pathways, including carbohydrates, amino acids and so on. Western blot showed the expression of ATP-citrate lyase, malate dehydrogenase 1 and lactate dehydrogenase A in myocardium changed markedly on day 3, while the phosphorylation level of AMP-activated protein kinase did not show significantly difference. We hope our research will promote deeper understanding and early diagnosis of cardiac hypertrophy in clinical practice. All raw data were deposited in MetaboLights (MTBLS10555).

Metabolic energetic adaptation of Atlantic salmon phagocytes to changes in carbon sources and exposure to PAMPs

Phagocytic cells are pivotal for host homeostasis and infection defense, necessitating metabolic adaptations in glycolysis, the tricarboxylic acid (TCA) cycle, and oxidative phosphorylation (OXPHOS). While mammalian phagocytes shift towards glycolysis and glutaminolysis during polarization, research on fish phagocyte metabolic reprogramming is limited. To address this, the Atlantic salmon phagocytic cell line, SHK-1, serves as a valuable model. Using the Seahorse XFe96 Flux Analyzer, this study compares SHK-1 bioenergetics under glucose-restricted (L-15 medium) and glucose-supplemented (PM) conditions, providing insights into metabolic characteristics and responses to Piscirickettsia salmonis bacterium Pathogen-associated molecular patterns (PAMPs). A standardized protocol for the study of real-time changes in the metabolism study of SHK-1 in PM and L-15 media, determining oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) is shown. Exhibiting metabolic adaptations, SHK-1 cells in the PM medium have higher basal and maximal OCR and spare capacity (SRC), while those grown in the L-15 medium favor OXPHOS, showing minimal glycolytic function. Despite metabolic differences, intracellular ATP levels are comparable, highlighting the metabolic plasticity and adaptability of SHK-1 cells to various carbon sources. Exposure to PAMPs from Piscirickettsia salmonis induces a metabolic shift, increasing glycolysis and OXPHOS, influencing ATP, lactate, glutamine, and glutamate levels. These findings highlight the role of mitochondrial bioenergetics and metabolic plasticity in salmon phagocytes, offering novel nutritional strategies for host-pathogen interventions based on energy metabolism.

Metabolic profile characterization of voxelotor in human urine based on in vivo and in vitro models for doping control.

Voxelotor was approved for the treatment of sickle cell anemia as a potent hemoglobin S polymerization inhibitor. Owing to its ability to affect blood components and its potential to enhance athletic performance, voxelotor was included in the prohibited list issued by the World Anti-Doping Agency in 2023, banning its use both in and out of competition. This study aimed to comprehensively investigate the metabolic profile of voxelotor in human urine and identify suitable metabolites for long-term analytical retrospectivity in doping control. A novel strategy for metabolite identification was established by combining in vivo human administration with isotope labeling-based in vitro metabolism analysis. A single microdose of voxelotor was administered orally to five volunteers, and urine samples were collected for up to 28 days post-administration. Concurrently, in vitro incubation of human liver microsomes with voxelotor and D3-voxelotor was conducted, and the microsomal incubates were analyzed via liquid chromatography-high-resolution mass spectrometry. Targeted metabolite searches in human urine samples and automated nontargeted screening of isotope metabolite ion pairs in incubation samples led to the discovery of 9 phase I metabolites and 23 phase II metabolites. Analysis of the urine excretion curves revealed that 4 metabolites, along with voxelotor, were suitable for long-term anti-doping monitoring, with a detection window exceeding 20 days. Using both in vivo and in vitro metabolic models, this study provides comprehensive insight into the metabolic profile of voxelotor in human urine for the first time, enhancing the capacity for doping screening and extending the retrospectivity of voxelotor detection.

Targeted and non-targeted metabolic characteristics of 6,7-dihydroxy-2,4- dimethoxyphenanthrene during simulated gastrointestinal digestion

6,7-dihydroxy-2,4-dimethoxyphenanthrene (PC4), isolated and identified from Chinese yam, was one of the important characteristic active ingredients. The present study established simulated gastrointestinal digestion and caco-2 intestinal epithelial models to investigate the digestive and metabolic characteristics of PC4. The results indicated that PC4 was mainly digested and metabolized in the intestine, with a digestion rate of 94 %, producing active characteristic metabolites including 2,6-dimethoxy-4-phenanthrenol (MC1) and 1-methoxyphenanthrene (MC2). Molecular docking indicated that the COX-2 enzyme inhibitory activity of MC1 might be superior to that of the prototype PC4. During 24 h co-incubation of PC4 with caco-2 monolayer epithelial, the signal pathway related to lipid decomposition was up-regulated within 0–12 h, while the diabetes complications AGE-RAGE was inhibited just in 0–6 h period significantly. The research database provided scientific basis for the digestion and metabolism of PC4, and laid theoretical foundation for the scientific development of Chinese yam functional foods.

Low-intensity transcranial ultrasound stimulation modulates neurovascular coupling in mouse models of Alzheimer's disease.

Neurovascular coupling plays an important role in the progression of Alzheimer's disease. However, it is unclear how ultrasound stimulation modulates neurovascular coupling in Alzheimer's disease. Here, we found that (i) transcranial ultrasound stimulation modulates the time domain and frequency domain characteristics of cerebral blood oxygen metabolism in Alzheimer's disease mice; (ii) transcranial ultrasound stimulation can significantly modulate the relative power of theta and gamma frequency of local field potential in Alzheimer's disease mice; and (iii) transcranial ultrasound stimulation can significantly modulate the neurovascular coupling in time domain and frequency domain induced by forepaw electrical stimulation in Alzheimer's disease mice. It provides a research basis for the clinical application of transcranial ultrasound stimulation in Alzheimer's disease patients.

Transcriptomic data support phylogenetic congruence and reveal genomic changes associated with the repeated evolution of annualism in aplocheiloid killifishes (Cyprinodontiformes)

Repeated evolution of novel life histories that are correlated with ecological variables offers opportunities to study convergence in genetic, developmental, and metabolic features. Nearly half of the 800 species of Aplocheiloid killifishes, a clade of teleost fishes with a circumtropical distribution, are “annual” or seasonal species that survive in ephemeral bodies of water that desiccate and are unfeasible for growth, reproduction, or survival for weeks to months every year. But the repeated evolution of adaptations that are key features of the annual life history among these fishes remains poorly known without a robust phylogenetic framework. We present a large-scale phylogenomic reconstruction of aplocheiloid killifishes evolution using newly sequenced transcriptomes obtained from a diversity of killifish lineages representing putative independent origins of annualism. Ancestral state estimation shows that developmental dormancy (diapause), a key trait of the killifish annual life cycle, may have originated up to seven times independently among African and South American lineages. To further explore the genetic basis of this unique trait, we measure changes in evolutionary rates among orthologous genes across the killifish tree of life by quantifying codon evolution using dN/dS ratios. We show that some genes have higher dN/dS ratios in lineages leading to species with annual life history. Many of them constitute key developmental genes or nuclear-encoded metabolic genes that control oxidative phosphorylation. Lastly, we compare these genes with higher ω to genes previously associated to developmental dormancy and metabolic shifts in killifishes and other vertebrates, and thereby identify molecular evolutionary signatures of repeated transitions to extreme environments.

Generation of a synthetic autochthonous microbiota responsible for the essential flavors and brewing properties of Daqu

Based on the 26 strains of medium–high-temperature Daqu, seed microbial inocula and mature microbial inoculants were prepared and inoculated into crushed wheat to prepare cleaner medium–high-temperature Daqu (XQ). The physicochemical properties, flavor, and microbial profile of XQ were studied using traditional medium–high-temperature Daqu (CQ) or industry standards as controls. The results showed that the liquefaction ability, saccharification activity, esterification activity, and moisture content of XQ were better than those of Daqu industry standards. The results of GC–MS and E-nose analyses showed that the main flavor compounds in the two types of Daqu were esters, alcohols, acids, and aldehydes, which had similar flavor profiles. Microbial sequencing revealed that the most abundant fungi in XQ were Aspergillus subflavus, Wickerhamomyces anomalus, Rhizopus oryzae, and Candida tropicalis, whereas the most abundant bacteria were Lactobacillus curvatus, Weissella cibaria, and Bacillus subtilis. Both types of Daqu contained a high abundance of W. anomalus. In addition, the COG (Clusters of Orthologous Groups of Proteins) analyses displayed that the two types of Daqu had their dominant functions and generally had the pathways and functional characteristics of organism growth and metabolism. This study laid some experimental research bases for the preparation of cleaner Daqu and could provide insights for producing safer microbial fermented foods.

In Silico SwissADME Analysis of Antibacterial NHC–Silver Acetates and Halides Complexes

This study investigates the antibacterial N-heterocyclic carbene (NHC)–silver complexes using the SwissADME platform, a web-based tool developed by the Swiss Institute of Bioinformatics (SIB). NHCs, particularly their silver complexes, have gained significant interest in medicinal chemistry for their potential as antibacterial and anticancer agents. The effectiveness of these complexes is closely linked to their structure, including factors like lipophilicity, which enhance their ability to penetrate bacterial cells and sustain the release of active silver ions. SwissADME provides computational estimates of pharmacokinetic properties, including absorption, distribution, metabolism, and excretion (ADME) characteristics, as well as drug-likeness and toxicity assessments. By evaluating parameters like molecular weight, topological polar surface area, lipophilicity (LogP), and water solubility, SwissADME offers insights into the drug-like potential of compounds. This study is inspired by a comprehensive review of antibacterial NHC–silver complexes published from 2006 to 2023, which identified superior structures with notable biological activity. The primary aim is to determine whether these active complexes exhibit distinct SwissADME parameters compared to others, providing a deeper understanding of the factors that influence their biological efficacy and aiding in the identification of promising drug candidates. Finally, experimental stabilities of exemplary complexes were confronted with absolute LUMO values derived from DFT calculations.

Metabolomic Alterations Associated with Phthalate Exposures among Pregnant Women in Puerto Rico.

Although phthalate exposure has been linked with multiple adverse pregnancy outcomes, their underlying biological mechanisms are not fully understood. We examined associations between biomarkers of phthalate exposures and metabolic alterations using untargeted metabolomics in 99 pregnant women and 86 newborns [mean (SD) gestational age = 39.5 (1.5) weeks] in the PROTECT cohort. Maternal urinary phthalate metabolites were quantified using isotope dilution high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS), while metabolic profiles in maternal and cord blood plasma were characterized via reversed-phase LC-MS. Multivariable linear regression was used in metabolome-wide association studies (MWAS) to identify individual metabolic features associated with elevated phthalate levels, while clustering and correlation network analyses were used to discern the interconnectedness of biologically relevant features. In the MWAS adjusted for maternal age and prepregnancy BMI, we observed significant associations between specific phthalates, namely, di(2-ethylhexyl) phthalate (DEHP) and mono(3-carboxypropyl) phthalate (MCPP), and 34 maternal plasma metabolic features. These associations predominantly included upregulation of fatty acids, amino acids, purines, or their derivatives and downregulation of ceramides and sphingomyelins. In contrast, fewer significant associations were observed with metabolic features in cord blood. Correlation network analysis highlighted the overlap of features associated with phthalates and those identified as differentiating markers for preterm birth in a previous study. Overall, our findings underscore the complex impact of phthalate exposures on maternal and fetal metabolism, highlighting metabolomics as a tool for understanding associated biological processes. Future research should focus on expanding the sample size, exploring the effects of phthalate mixtures, and validating identified metabolic features in larger, more diverse populations.

Regulation of histidine metabolism by Lactobacillus Reuteri mediates the pathogenesis and treatment of ischemic stroke

Increasing evidence has underscored the significance of post-stroke alterations along gut–brain axis, while its role in pathogenesis and treatment of ischemic stroke (IS) remains largely unexplored. This study aimed to elucidate the therapeutic effects and action targets of Panax notoginseng saponins (PNS) on IS and explore a novel pathogenesis and treatment strategy of IS via profiling the microbial community and metabolic characteristics along gut–brain axis. Our findings revealed for the first time that the therapeutic effect of PNS on IS was microbiota-dependent. Ischemia/reperfusion (I/R) modeling significantly down-regulated Lactobacilli in rats, and PNS markedly recovered Lactobacilli, particularly Lactobacillus reuteri (L.Reu). Metabolomics showed a significant reduction in serum histidine (HIS) in clinical obsolete IS patients and rehabilitation period I/R rats. Meanwhile, the L.Reu colonization in I/R rats exhibited significant neuroprotective activity and greatly increased HIS in serum, gut microbiota, and brain. Moreover, exogenous HIS demonstrated indirect neuroprotective effects through metabolizing to histamine. Notably, vagus nerve severance in I/R rats was performed to investigate HIS's neuroprotective mechanism. The results innovatively revealed that PNS could promote HIS synthesis in gut by enhancing L.Reu proportion, thereby increasing intracerebral HIS through peripheral pathway. Consequently, our data provided novel insights into HIS metabolism mediated by L.Reu in the pathogenesis and treatment of IS.

Depression symptoms, wellbeing, health-related quality of life, and diabetes-related distress in novel subtypes of recent-onset diabetes in Germany: a 5-year observational follow-up study.

The subjective experiences of individuals living with diabetes is commonly assessed with patient-reported outcomes (PROs; eg, depression symptoms, wellbeing, health-related quality of life [HRQOL], and diabetes-related distress). Cluster analyses have identified novel diabetes subtypes differing in phenotypic and metabolic characteristics. We aimed to investigate associations between these subtypes and PROs and whether subtype predicted PROs 5 years later. Baseline (<12 months after a diabetes diagnosis) and 5-year follow-up data were collected from German Diabetes Study (GDS) participants. Multiple regressions were applied to analyse associations between diabetes subtypes and depression symptoms (Center for Epidemiologic Studies Depression Scale), wellbeing (WHO-5), HRQOL (SF-36), and diabetes-related distress (Problem Areas in Diabetes Scale). Cluster analyses at baseline (n=1391) identified participants with severe autoimmune diabetes (SAID, 417 [30%]), severe insulin-deficient diabetes (SIDD, 33 [2%]), severe insulin-resistant diabetes (SIRD, 150 [11%]), mild obesity-related diabetes (MOD, 354 [25%]), and mild age-related diabetes (MARD, 437 [31%]). At baseline, multiple regression analyses showed that participants with SIRD had higher depression symptoms than participants with MARD and lower physical HRQOL than all other subtypes. Participants with SAID reported higher depression symptoms and lower mental HRQOL than participants with MARD, higher physical HRQOL than participants with MARD and MOD, and higher diabetes-related distress than most other subtypes. At the 5-year follow-up, clustering predicted no statistically significant changes in PROs after adjustment for multiple testing, whereas descriptive analyses demonstrated that individuals with SIRD were more likely to experience clinically relevant depression symptoms (16% vs 6%) and low wellbeing (31% vs 14%), respectively, than individuals with MARD. Diabetes subtypes already differ in PROs at diabetes diagnosis. Our analyses had limited predictive power during follow-up. However, our findings suggest that clustering could predict future changes in depression symptoms. The GDS was initiated and financed by the German Diabetes Center, which is funded by the German Federal Ministry of Health, the Ministry of Culture and Science of the state of North Rhine-Westphalia, and by the German Federal Ministry of Education and Research to the German Center for Diabetes Research. For the German translation of the abstract see Supplementary Materials section.

In Vitro Treatment with Metformin Significantly Reduces Senescent B Cells Present in the Adipose Tissue of People with Obesity

BackgroundOur previous work has shown that senescent B cells accumulate in the human adipose tissue (AT) from people with obesity, where they express transcripts for markers associated with the senescence-associated secretory phenotype (SASP) and secrete multiple inflammatory mediators. These functions of AT-derived B cells are metabolically supported. ObjectivesTo show that Metformin (MET), a widely used hypoglycemic and antidiabetic drug, is able at least in vitro to decrease frequencies, secretory profile, and metabolic requirements of senescent B cells isolated from the AT from people with obesity. MethodsWe recruited adult females with obesity (n = 8, age 40 ± 2 y, BMI range: 33–42) undergoing breast reduction surgery, who donated their discarded subcutaneous AT. B cells from stromal vascular fractions isolated after collagenase digestion of the AT were evaluated after in vitro incubation with MET (1 mM × 106 B cells) or with a medium for the following measures. The expression of transcripts for SASP-associated markers (p16INK4a and p21CIP1/WAF1) measured by quantitative polymerase chain reaction (qPCR); secretion of inflammatory cytokines (TNF-α, IL-6, IFN-γ and IL-17A) measured by a Cytometric Bead Array); metabolic characteristics as identified by a glycolytic test and Seahorse technology, and by the expression of transcripts for glucose transporters and metabolic enzymes involved in glucose metabolic pathways, measured by qPCR. To examine differences between MET-treated compared with untreated groups, paired Student’s t tests (two-tailed) were employed. ResultsMET in vitro was able to reduce frequencies and numbers of senescent B cells, as identified by staining with β-galactosidase, as well as the secretion of inflammatory cytokines, the expression of transcripts for SASP, and metabolic markers that support intrinsic B cell inflammation. ConclusionsOur results provide evidence to support the beneficial effects of MET in reducing AT-related inflammation through its effects on senescent B cells.

Differential serum metabolites in patients with pregnancy-associated venous thromboembolism analyzed using GC-MS/LC-MS untargeted metabolomics

Untargeted metabolomics can be used for the comprehensive analysis of metabolite profiles in biological samples without preset targets, making them particularly suitable for exploring metabolic characteristics and potential mechanisms in complex diseases. Therefore, in this study, we employed gas chromatography–mass spectrometry (GC-MS) and liquid chromatography–mass spectrometry (LC-MS) techniques to analyze the serum metabolic characteristics of patients with pregnancy-associated venous thromboembolism (PA-VTE). In this study, 11 pregnant women with VTE and 11 healthy pregnant women were included in the experimental and control groups, respectively. Using GC-MS, we identified 325 metabolites, with the highest proportion being organic oxygen compounds. Using LC-MS, we identified 3104 metabolites, with the highest proportion being acylcarnitine. The results revealed significant differences in the levels of lipids, organic compounds, and other metabolites between patients compared to healthy pregnant women. Pathways such as pyrimidine metabolism, linoleic acid metabolism, and mineral absorption differed between patients with PA-VTE and controls. Furthermore, we identified biomarkers associated with metabolic processes, such as fatty acids and amino acids (2-hydroxyhexanedioic acid, hexadecenal, palmitoylethanolamide, glycerol-1-phosphate, and N-acetyl-beta-D-glucosamine). These findings revealed the metabolic characteristics of PA-VTE and provided important clues for further research on its pathophysiological mechanisms. Our findings may contribute to the development of new diagnostic markers and support early diagnosis and treatment of PA-VTE.