The ontogenesis of the circadian clock in the suprachiasmatic nuclei of the hypothalamus (SCN) and its sensitivity to maternal signals are not fully understood. Here, we investigated the development of the clock in the rat SCN from the fetal to the postweaning period and identified rhythmic metabolic signals from the mother to the fetal SCN. We determined daily expression profiles of clock genes (Per2, Nr1d1, Bmal1) and clock- and metabolism-related genes (Dbp, E4bp4) and performed time-resolved analysis of the metabolome and lipidome in the SCN and plasma of 19-day-old embryos (E19) and 2-, 10-, 20-, and 28-day-old pups (P02-28). Our data show that rhythms in the expression of canonical clock genes are absent at E19 and develop gradually until P10, but the Dbp rhythm was still developing between P20 and P28. Expression of the metabolism-sensitive gene E4bp4 and levels of essential amino acids and other metabolites supplied by maternal food are rhythmic in the fetal SCN, which is lost after birth at P02 and reappears later in the postnatal period. Maternal food-derived metabolites were also rhythmic in fetal plasma. The temporal coherence of the fetal SCN metabolome and lipidome declines markedly and its rhythmicity disappears immediately after birth. The results revealed previously unforeseen pathways by which the fetal SCN may receive rhythmic information from the mother before its clock develops.

• MeSH
• Circadian Clocks * physiology   genetics   MeSH
• Period Circadian Proteins genetics   metabolism   MeSH
• Circadian Rhythm physiology   MeSH
• Rats MeSH
• Metabolome MeSH
• Suprachiasmatic Nucleus * metabolism   embryology   physiology   MeSH
• CLOCK Proteins genetics   metabolism   MeSH
• Pregnancy MeSH
• ARNTL Transcription Factors genetics   metabolism   MeSH
• Gene Expression Regulation, Developmental MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Pregnancy MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Period Circadian Proteins MeSH
• CLOCK Proteins MeSH
• ARNTL Transcription Factors MeSH

Circadian rhythms regulate key physiological processes through clock genes in central and peripheral tissues. While circadian gene expression during development has been well studied, the temporal dynamics of metabolism across tissues remain less understood. Here, we present the Circadian Ontogenetic Metabolomics Atlas (COMA), which maps circadian metabolic rhythms across 16 rat anatomical structures. The brain (suprachiasmatic nuclei, medial prefrontal cortex) and periphery (liver, plasma) span developmental stages from embryonic E19 to postnatal P2, P10, P20, and P28. Fecal samples include all four postnatal stages, while additional peripheral tissues were analyzed at P20 and P28. Using a multiplatform liquid chromatography-mass spectrometry approach, we annotated 851 metabolites from 1610 samples. We identified distinct circadian shifts, particularly during the transition from nursing to solid food intake (P10-P20), with an average of 24% of metabolites exhibiting circadian oscillations across sample types, as determined by JTK_CYCLE. Our study also underscores the importance of standardized sampling, as metabolite intensities fluctuate with both circadian rhythms and development. COMA serves as an open-access resource ( https://coma.metabolomics.fgu.cas.cz ) for exploring circadian metabolic regulation and its role in developmental biology.

• Keywords
• Atlas, Circadian rhythm, Lipidomics, Metabolomics, Resource,
• MeSH
• Chromatography, Liquid MeSH
• Circadian Rhythm * physiology   MeSH
• Feces * chemistry   MeSH
• Liver metabolism   MeSH
• Rats MeSH
• Metabolome * MeSH
• Metabolomics * methods   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH

AIM: Exposure to light at night and meal time misaligned with the light/dark (LD) cycle-typical features of daily life in modern 24/7 society-are associated with negative effects on health. To understand the mechanism, we developed a novel protocol of complex chronodisruption (CD) in which we exposed female rats to four weekly cycles consisting of 5-day intervals of constant light and 2-day intervals of food access restricted to the light phase of the 12:12 LD cycle. METHODS: We examined the effects of CD on behavior, estrous cycle, sleep patterns, glucose homeostasis and profiles of clock- and metabolism-related gene expression (using RT qPCR) and liver metabolome and lipidome (using untargeted metabolomic and lipidomic profiling). RESULTS: CD attenuated the rhythmic output of the central clock in the suprachiasmatic nucleus via Prok2 signaling, thereby disrupting locomotor activity, the estrous cycle, sleep patterns, and mutual phase relationship between the central and peripheral clocks. In the periphery, CD abolished Per1,2 expression rhythms in peripheral tissues (liver, pancreas, colon) and worsened glucose homeostasis. In the liver, it impaired the expression of NAD+, lipid, and cholesterol metabolism genes and abolished most of the high-amplitude rhythms of lipids and polar metabolites. Interestingly, CD abolished the circadian rhythm of Cpt1a expression and increased the levels of long-chain acylcarnitines (ACar 18:2, ACar 16:0), indicating enhanced fatty acid oxidation in mitochondria. CONCLUSION: Our data show the widespread effects of CD on metabolism and point to ACars as biomarkers for CD due to misaligned sleep and feeding patterns.

• Keywords
• acylcarnitine, chronodisruption, clock, female, glucose homeostasis, liver, metabolome, pancreas, rat, sleep, suprachiasmatic nucleus,
• MeSH
• Circadian Clocks * physiology   MeSH
• Circadian Rhythm * physiology   MeSH
• Photoperiod MeSH
• Liver * metabolism   MeSH
• Carnitine * analogs & derivatives   metabolism   MeSH
• Rats MeSH
• Metabolome * physiology   MeSH
• Rats, Sprague-Dawley MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• acylcarnitine MeSH Browser
• Carnitine * MeSH

Fasting induces significant shifts in substrate utilization with signs of acute insulin resistance (IR), while obesity is associated with chronic IR. Nonetheless, both states substantially influence adipose tissue (AT) function. Therefore, in this interventional study (NCT04260542), we investigated if excessive adiposity in premenopausal women alters insulin sensitivity and AT metabolic and endocrine activity in response to a 60-h fast and a subsequent 48-h refeeding period. Using physiological methods, lipidomics, and AT explants, we showed that obesity partially modified AT endocrine activity and blunted the dynamics of AT insulin resistance in response to the fasting/refeeding challenge compared to that observed in lean women. AT adapted to its own excess by reducing lipolytic activity/free fatty acids (FFA) flux per mass. This adaptation persisted even after a 60-h fast, resulting in lower ketosis in women with obesity. This could be a protective mechanism that limits the lipotoxic effects of FFA; however, it may ultimately impede desirable weight loss induced by caloric restriction in women with obesity.

• Publication type
• Journal Article MeSH

Metabolic syndrome is a growing concern in developed societies and due to its polygenic nature, the genetic component is only slowly being elucidated. Common mitochondrial DNA sequence variants have been associated with symptoms of metabolic syndrome and may, therefore, be relevant players in the genetics of metabolic syndrome. We investigate the effect of mitochondrial sequence variation on the metabolic phenotype in conplastic rat strains with identical nuclear but unique mitochondrial genomes, challenged by high-fat diet. We find that the variation in mitochondrial rRNA sequence represents risk factor in the insulin resistance development, which is associated with diacylglycerols accumulation, induced by tissue-specific reduction of the oxidative capacity. These metabolic perturbations stem from the 12S rRNA sequence variation affecting mitochondrial ribosome assembly and translation. Our work demonstrates that physiological variation in mitochondrial rRNA might represent a relevant underlying factor in the progression of metabolic syndrome.

• MeSH
• Diet, High-Fat adverse effects   MeSH
• Genetic Predisposition to Disease MeSH
• Haplotypes * MeSH
• Insulin Resistance genetics   MeSH
• Rats MeSH
• Metabolic Syndrome * genetics   metabolism   MeSH
• DNA, Mitochondrial genetics   metabolism   MeSH
• Mitochondria metabolism   genetics   MeSH
• RNA, Mitochondrial genetics   metabolism   MeSH
• RNA, Ribosomal * genetics   metabolism   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA, Mitochondrial MeSH
• RNA, Mitochondrial MeSH
• RNA, Ribosomal * MeSH
• RNA, ribosomal, 12S MeSH Browser

Metabolomics and lipidomics have emerged as tools in understanding the connections of metabolic syndrome (MetS) with cardiovascular diseases (CVD), type 1 and type 2 diabetes (T1D, T2D), and metabolic dysfunction-associated steatotic liver disease (MASLD). This review highlights the applications of these omics approaches in large-scale cohort studies, emphasizing their role in biomarker discovery and disease prediction. Integrating metabolomics and lipidomics has significantly advanced our understanding of MetS pathology by identifying unique metabolic signatures associated with disease progression. However, challenges such as standardizing analytical workflows, data interpretation, and biomarker validation remain critical for translating research findings into clinical practice. Future research should focus on optimizing these methodologies to enhance their clinical utility and address the global burden of MetS-related diseases.

• MeSH
• Biomarkers metabolism   MeSH
• Diabetes Mellitus, Type 1 metabolism   complications   MeSH
• Diabetes Mellitus, Type 2 * metabolism   MeSH
• Cardiovascular Diseases * metabolism   diagnosis   MeSH
• Humans MeSH
• Lipidomics * methods   MeSH
• Metabolic Syndrome * metabolism   MeSH
• Metabolomics * methods   MeSH
• Fatty Liver metabolism   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH
• Names of Substances
• Biomarkers MeSH

Contrast-enhanced computed tomography offers a nondestructive approach to studying adipose tissue in 3D. Several contrast-enhancing staining agents (CESAs) have been explored, whereof osmium tetroxide (OsO4) is the most popular nowadays. However, due to the toxicity and volatility of the conventional OsO4, alternative CESAs with similar staining properties were desired. Hf-WD 1:2 POM and Hexabrix have proven effective for structural analysis of adipocytes using contrast-enhanced computed tomography but fail to provide chemical information. This study introduces isotonic Lugol's iodine (IL) as an alternative CESA for adipose tissue analysis, comparing its staining potential with Hf-WD 1:2 POM and Hexabrix in murine caudal vertebrae and bovine muscle tissue strips. Single and sequential staining protocols were compared to assess the maximization of information extraction from each sample. The study investigated interactions, distribution, and reactivity of iodine species towards biomolecules using simplified model systems and assesses the potential of the CESA to provide chemical information. (Bio)chemical analyses on whole tissues revealed that differences in adipocyte gray values post-IL staining were associated with chemical distinctions between bovine muscle tissue and murine caudal vertebrae. More specific, a difference in the degree of unsaturation of fatty acids was identified as a likely contributor, though not the sole determinant of gray value differences. This research sheds light on the potential of IL as a CESA, offering both structural and chemical insights into adipose tissue composition.

• Keywords
• 3D histology, DICECT, Lugol’s iodine, adipocytes, adipose tissue, bone marrow, lipids/chemistry, muscle,
• MeSH
• Staining and Labeling methods   MeSH
• Contrast Media * chemistry   MeSH
• Mice, Inbred C57BL MeSH
• Mice MeSH
• Tomography, X-Ray Computed * methods   MeSH
• Cattle MeSH
• Adipose Tissue * diagnostic imaging   metabolism   MeSH
• Adipocytes cytology   metabolism   MeSH
• Animals MeSH
• Check Tag
• Mice MeSH
• Cattle MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Contrast Media * MeSH

Liquid chromatography with mass spectrometry (LC-MS)-based metabolomics detects thousands of molecular features (retention time-m/z pairs) in biological samples per analysis, yet the metabolite annotation rate remains low, with 90% of signals classified as unknowns. To enhance the metabolite annotation rates, researchers employ tandem mass spectral libraries and challenging in silico fragmentation software. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) may offer an additional layer of structural information in untargeted metabolomics, especially for identifying specific unidentified metabolites that are revealed to be statistically significant. Here, we investigate the potential of hydrophilic interaction liquid chromatography (HILIC)-HDX-MS in untargeted metabolomics. Specifically, we evaluate the effectiveness of two approaches using hypothetical targets: the post-column addition of deuterium oxide (D2O) and the on-column HILIC-HDX-MS method. To illustrate the practical application of HILIC-HDX-MS, we apply this methodology using the in silico fragmentation software MS-FINDER to an unknown compound detected in various biological samples, including plasma, serum, tissues, and feces during HILIC-MS profiling, subsequently identified as N1-acetylspermidine.

• Keywords
• hydrogen/deuterium exchange, liquid chromatography, mass spectrometry, metabolomics, unknown identification,
• MeSH
• Chromatography, Liquid methods   MeSH
• Deuterium MeSH
• Hydrophobic and Hydrophilic Interactions MeSH
• Metabolomics * methods   MeSH
• Hydrogen Deuterium Exchange-Mass Spectrometry * MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Deuterium MeSH

Cardiac tolerance to ischaemia can be increased by dietary interventions such as fasting, which is associated with significant changes in myocardial gene expression. Among the possible mechanisms of how gene expression may be altered are epigenetic modifications of RNA - epitranscriptomics. N6-methyladenosine (m6A) and N6,2'-O-dimethyladenosine (m6Am) are two of the most prevalent modifications in mRNA. These methylations are reversible and regulated by proteins called writers, erasers, readers, and m6A-repelled proteins. We analysed 33 of these epitranscriptomic regulators in rat hearts after cardioprotective 3-day fasting using RT-qPCR, Western blot, and targeted proteomic analysis. We found that the most of these regulators were changed on mRNA or protein levels in fasting hearts, including up-regulation of both demethylases - FTO and ALKBH5. In accordance, decreased methylation (m6A+m6Am) levels were detected in cardiac total RNA after fasting. We also identified altered methylation levels in Nox4 and Hdac1 transcripts, both of which play a role in the cytoprotective action of ketone bodies produced during fasting. Furthermore, we investigated the impact of inhibiting demethylases ALKBH5 and FTO in adult rat primary cardiomyocytes (AVCMs). Our findings indicate that inhibiting these demethylases reduced the hypoxic tolerance of AVCMs isolated from fasting rats. This study showed that the complex epitranscriptomic machinery around m6A and m6Am modifications is regulated in the fasting hearts and might play an important role in cardiac adaptation to fasting, a well-known cardioprotective intervention.

• Keywords
• ALKBH5, FTO, Fasting, epitranscriptomics, heart, m6A, m6Am,
• MeSH
• Adenosine * genetics   metabolism   MeSH
• Rats MeSH
• RNA, Messenger genetics   MeSH
• Fasting MeSH
• Proteomics * MeSH
• RNA metabolism   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Adenosine * MeSH
• RNA, Messenger MeSH
• RNA MeSH

Liquid chromatography-mass spectrometry (LC-MS) is the key technique for analyzing complex lipids in biological samples. Various LC-MS modes are used for lipid separation, including different stationary phases, mobile-phase solvents, and modifiers. Quality control in lipidomics analysis is crucial to ensuring the generated data's reliability, reproducibility, and accuracy. While several quality control measures are commonly discussed, the impact of organic solvent quality during LC-MS analysis is often overlooked. Additionally, the annotation of complex lipids remains prone to biases, leading to potential misidentifications and incomplete characterization of lipid species. In this study, we investigate how LC-MS-grade isopropanol from different vendors may influence the quality of the mobile phase used in LC-MS-based untargeted lipidomic profiling of biological samples. Furthermore, we report the occurrence of an unusual, yet highly abundant, ethylamine adduct [M+46.0651]+ that may form for specific lipid subclasses during LC-MS analysis in positive electrospray ionization mode when acetonitrile is part of the mobile phase, potentially leading to lipid misidentification. These findings emphasize the importance of considering solvent quality in LC-MS analysis and highlight challenges in lipid annotation.

• Keywords
• MS/MS annotation, adduct formation, lipidomics, lipids, liquid chromatography, mass spectrometry, metabolomics, method development, misidentification, solvent quality,
• Publication type
• Journal Article MeSH