Expanded clinical phenotype and untargeted metabolomics analysis in RARS2-related mitochondrial disorder: a case report.

BACKGROUND: RARS2-related mitochondrial disorder is an autosomal recessive mitochondrial encephalopathy caused by biallelic pathogenic variants in the gene encoding the mitochondrial arginyl-transfer RNA synthetase 2 (RARS2, MIM *611524, NM_020320.5). RARS2 catalyzes the transfer of L-arginine to its cognate tRNA during the translation of mitochondrially-encoded proteins. The classical presentation of RARS2-related mitochondrial disorder includes pontocerebellar hypoplasia (PCH), progressive microcephaly, profound developmental delay, feeding difficulties, and hypotonia. Most patients also develop severe epilepsy by three months of age, which consists of focal or generalized seizures that frequently become pharmacoresistant and lead to developmental and epileptic encephalopathy (DEE). CASE PRESENTATION: Here, we describe a six-year-old boy with developmental delay, hypotonia, and failure to thrive who developed an early-onset DEE consistent with Lennox-Gastaut Syndrome (LGS), which has not previously been observed in this disorder. He had dysmorphic features including bilateral macrotia, overriding second toes, a depressed nasal bridge, retrognathia, and downslanting palpebral fissures, and he did not demonstrate progressive microcephaly. Whole genome sequencing identified two variants in RARS2, c.36 + 1G > T, a previously unpublished variant that is predicted to affect splicing and is, therefore, likely pathogenic and c.419 T > G (p.Phe140Cys), a known pathogenic variant. He exhibited significant, progressive generalized brain atrophy and ex vacuo dilation of the supratentorial ventricular system on brain MRI and did not demonstrate PCH. Treatment with a ketogenic diet (KD) reduced seizure frequency and enabled him to make developmental progress. Plasma untargeted metabolomics analysis showed increased levels of lysophospholipid and sphingomyelin-related metabolites. CONCLUSIONS: Our work expands the clinical spectrum of RARS2-related mitochondrial disorder, demonstrating that patients can present with dysmorphic features and an absence of progressive microcephaly, which can help guide the diagnosis of this condition. Our case highlights the importance of appropriate seizure phenotyping in this condition and indicates that patients can develop LGS, for which a KD may be a viable therapeutic option. Our work further suggests that analytes of phospholipid metabolism may serve as biomarkers of mitochondrial dysfunction.

Novel phenotype of aortic root dilatation and late-onset metabolic decompensation in a patient with TMEM70 deficiency.

TMEM70 deficiency causing mitochondrial complex V deficiency, nuclear type 2 (MIM: 614052) is the most common nuclear encoded defect affecting ATP synthase and has been well described in the literature as being characterized by neonatal or infantile onset of poor feeding, hypotonia, lethargy, respiratory compromise, heart failure, lactic acidosis, hyperammonemia, and 3-methylglutaconic aciduria progressing to a phenotype of developmental delay, failure to thrive, short stature, nonprogressive cardiomyopathy, microcephaly, facial dysmorphisms, hypospadias, persistent pulmonary hypertension of the newborn, and Wolff-Parkinson-White syndrome, as well as metabolic crises followed by developmental regression. The patient with TMEM70 deficiency herein reported has the unique presentation of aortic root dilatation, differing facial dysmorphisms, and no history of neonatal metabolic decompensation or developmental delay, as well as a plasma metabolomics signature, including elevated 3-methylglutaconic acid, 3-methylglutarylcarnitine, alanine, and lactate, in addition to the commonly described increased 3-methylglutaconic acid on urine organic acid analysis that helped aid in the diagnostic interpretation of variants of uncertain significance in TMEM70.

rpeA, a global regulator involved in mupirocin biosynthesis in Pseudomonas fluorescens NCIMB 10586.

Mupirocin, a polyketide antibiotic produced by Pseudomonas fluorescens, is used as a topical antimicrobial treatment to cure various skin infections. Quorum sensing system plays an important role in regulation of mupirocin biosynthesis in P. fluorescens NCIMB 10586. In Pseudomonas, the RpeA/RpeB two-component signal transduction (TCST) system regulates quorum sensing system. However, the influences of the RpeA/RpeB TCST system on mupirocin production or other cell activities have not been studied. In this work, the homologous genes of rpeA and rpeB in P. fluorescens NCIMB 10586 were identified and inactivated in the chromosome, respectively. The deletion of rpeA reduced the mupirocin production from 160 in the wild-type to 21.3 mg/L along with slightly decreased cell growth, while no significant effected on mupirocin production in the rpeB mutant. Next, it was found that the RpeA/RpeB TCST system regulated the biosynthesis of mupirocin by modulating the quorum sensing system. Furthermore, untargeted metabolomics analysis was employed to detect the influences of RpeA on other cell activities modulated by quorum sensing system. Combined with quantitative real-time PCR, the results demonstrated that RpeA also regulated other cell activities including central carbon, amino acids, fatty acids, and purine metabolism. Overall, this study expands the current understanding of the RpeA/RpeB TCST system and provides several targets for increasing yields of mupirocin. KEY POINTS: • In P. fluorescens, the RpeA/RpeB TCST system regulates the biosynthesis of mupirocin. • RpeA modulates the cell activities through effecting the central carbon metabolism.

Alteration of plasma metabolites associated with chemoradiosensitivity in esophageal squamous cell carcinoma via untargeted metabolomics approach.

BACKGROUND: To investigate the differences in plasma metabolomic characteristics between pathological complete response (pCR) and non-pCR patients and identify biomarker candidates for predicting the response to neoadjuvant chemoradiotherapy (nCRT) in esophageal squamous cell carcinoma (ESCC). METHODS: A total of 46 ESCC patients were included in this study. Gas chromatography time-of- flight mass spectrometry (GC-TOF/MS) technology was applied to detect the plasma samples collected before nCRT via untargeted metabolomics analysis. RESULTS: Five differentially expressed metabolites (out of 109) was found in plasma between pCR and non-pCR groups. Compared with non-pCR group, isocitric acid (p = 0.0129), linoleic acid (p = 0.0137), citric acid (p = 0.0473) were upregulated, while L-histidine (p = 0.0155), 3'4 dihydroxyhydrocinnamic acid (p = 0.0339) were downregulated in the pCR plasma samples. Pathway analyses unveiled that citrate cycle (TCA cycle), glyoxylate and dicarboxylate metabolic pathway were associated with ESCC chemoradiosensitivity. CONCLUSION: The present study provided supporting evidence that GC-TOF/MS based metabolomics approach allowed identification of metabolite differences between pCR and non-pCR patients in plasma levels, and the systemic metabolic status of patients may reflect the response of ESCC patient to neoadjuvant chemoradiotherapy.

Hematologic presentation and the role of untargeted metabolomics analysis in monitoring treatment for riboflavin transporter deficiency.

Riboflavin transporter deficiency (RTD) (MIM #614707) is a neurogenetic disorder with its most common manifestations including sensorineural hearing loss, peripheral neuropathy, respiratory insufficiency, and bulbar palsy. Here, we present a 2-year-old boy whose initial presentation was severe macrocytic anemia necessitating multiple blood transfusions and intermittent neutropenia; he subsequently developed ataxia and dysarthria. Trio-exome sequencing detected compound heterozygous variants in SLC52A2 that were classified as pathogenic and a variant of uncertain significance. Bone marrow evaluation demonstrated megaloblastic changes. Notably, his anemia and neutropenia resolved after treatment with oral riboflavin, thus expanding the clinical phenotype of this disorder. We reiterate the importance of starting riboflavin supplementation in a young child who presents with macrocytic anemia and neurological features while awaiting biochemical and genetic work up. We detected multiple biochemical abnormalities with the help of untargeted metabolomics analysis associated with abnormal flavin adenine nucleotide function which normalized after treatment, emphasizing the reversible pathomechanisms involved in this disorder. The utility of untargeted metabolomics analysis to monitor the effects of riboflavin supplementation in RTD has not been previously reported.

Physicochemical, microbiological and metabolomics changes in yogurt supplemented with lactosucrose.

Lactosucrose (LS) is a known prebiotic that has gained recognition for its low caloric content and various health benefits. However, its potential in food applications remains largely unexplored. In this study the effects of adding LS to milk at concentrations (0 %, 2 %, 5 % and 8 % w/v) for yogurt production, and the relevant changes in yogurt texture, microbial composition and metabolomics were investigated. Our findings revealed that LS played a role in promoting the formation of a structured gel during fermentation, resulting in increased elasticity and viscosity while reducing fluidity. Additionally incorporating high doses of LS into yogurt led to reduced post-acidification, enhanced survival of starter bacteria, improved water retention capacity and overall texture throughout a refrigerated storage period of 21 days. Notably higher concentrations of LS (8 % w/v) exhibited effects on enhancing yogurt quality. Furthermore, untargeted metabolomics analysis using UPLC Q TOF MS/MS revealed 45 differentially expressed metabolites, including up-regulated L-arginine, L-proline and L-glutamic acid along with the down-regulated glutathione, L-tyrosine, L-phenylalanyl and L-proline. These differential metabolites were primarily associated with amino acid metabolism such as thiamine metabolism, nicotinic acid salt and nicotinamide metabolism, and pyrimidine metabolism. As a result, the inclusion of LS in yogurt had an impact on the production of various beneficial metabolites in yogurt, highlighting the importance of combining prebiotic LS with probiotics to obtain desired physiological benefits of yogurt.

Untargeted metabolomics analyses to identify a new sweet compound released during post-fermentation maceration of wine.

The sensory quality of a wine is mainly based on its aroma and flavor. Sweetness contributes in the gustatory balance of red wines. The investigation of compounds involved in this flavor was based on empirical observations, such as the increase in wine sweetness during yeast autolysis, concomitant to post-fermentation maceration in red winemaking. An untargeted metabolomics approach using UHPLC-HRMS has been developed to discover a new sweet molecule released during this stage. Among several markers highlighted, one compound was selected to be isolated by various separative techniques. It was unambiguously identified by NMR as N6-succinyladenosine and is reported for the first time in wine at an average concentration of 3.16 mg/L in 85 red wines. Furthermore, sensory analysis has highlighted its sweetness. In addition to discovering a new sweet compound in wine, this study proposes new tools for studying taste-active compounds in natural matrices.

Metagenomics and untargeted metabolomics analyses to unravel the formation mechanism of characteristic metabolites in Cantonese soy sauce during different fermentation stages.

Cantonese soy sauce (CSS) is an important Chinese condiment due to its distinctive flavor. Microorganisms play a significant role in the flavor formation of CSS during fermentation. However, the correlation between microbes and flavor compounds as well as the potential fermentation mechanism remained poorly uncovered. Here we revealed the dynamic changes of microbial structure and characteristics metabolites as well as their correlation of CSS during the fermentation process. Metagenomics sequencing analysis showed that Tetragenococcus halophilus, Weissella confusa, Weissella paramesenteroides, Aspergillus oryzae, Lactiplantibacillus plantarum, Weissella cibaria were top six dominant species from day 0 to day 120. Sixty compounds were either positively or tentatively identified through untargeted metabolomics profile and they were 27 peptides, amino acids and derivatives, 8 carbohydrates and conjugates, 14 organic acids and derivatives, 5 amide compounds, 3 flavonoids and 3 nucleosides. Spearman correlation coefficient indicated that Tetragenococcus halophilus, Zygosaccharomyces rouxii, Pediococcus pentosaceus and Aspergillus oryzae were significantly related with the formation of taste amino acids and derivatives, peptides and functional substances. Additionally, the metabolisms of flavor amino acids including 13 main free amino acids were also profiled. These results provided valuable information for the production practice in the soy sauce industry.

Differential in vitro cytotoxic effects and metabolomic insights into raw and powdered Manuka honey through UPLC-Q-TOF-MS.

Manuka honey (MH) has garnered much attention due to its remarkable antimicrobial, anticancer, immunomodulatory and wound-healing properties. This study compared the antiproliferative effects of raw and powdered MH (pMH) on various human and murine cancer cell lines. A detailed metabolomics analysis was also carried out using untargeted ultrahigh-performance liquid chromatography-quadrupole-time-of-flight-mass spectrometry (UPLC-Q-TOF-MS) to compare the constituents in raw MH and pMH. The results of the viability studies showed that both raw MH and pMH caused a dose-dependent inhibition of tumor cell growth at concentrations of > 1% w/v (equivalent to ~ 10 mg/ml). A differential susceptibility to MH was observed among the cell lines with the human MDA-MB-231 and A549 cells and murine B16.F10 cells being relatively resistant to MH while the murine MC38 colorectal adeno-carcinoma cells showing the most sensitivity. The effect of raw MH and pMH on cell viability was validated using 2 indepndent assays. Metabolomics analysis detected 2440 compounds, out of which 833 were successfully identified. Among these, 90 phytochemical compounds, predominantly comprising terpenoids, flavonoids, coumarins and derivatives, and phenylpropanoic acids, and 79 lipids were identifiable. Significant differences in 5 metabolite classes, including flavonoids, phenols, terpenoids, carbohydrates, and organic acids were observed between the raw and pMH. Moreover, several altered metabolic pathways were identified in pMH compared to raw MH, such as energy metabolism, amino acid metabolism, and various other pathways that collectively influence biological functions associated with cellular growth, signaling, and stress response.

Untargeted Metabolomics Reveals Alterations of Rhythmic Pulmonary Metabolism in IPF.

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive condition characterized by the impairment of alveolar epithelial cells. Despite continued research efforts, the effective therapeutic medication is still absent due to an incomplete understanding of the underlying etiology. It has been shown that rhythmic alterations are of significant importance in the pathophysiology of IPF. However, a comprehensive understanding of how metabolite level changes with circadian rhythms in individuals with IPF is lacking. Here, we constructed an extensive metabolite database by utilizing an unbiased reference system culturing with 13C or 15N labeled nutrients. Using LC-MS analysis via ESI and APCI ion sources, 1300 potential water-soluble metabolites were characterized and applied to evaluate the metabolic changes with rhythm in the lung from both wild-type mice and mice with IPF. The metabolites, such as glycerophospholipids and amino acids, in WT mice exhibited notable rhythmic oscillations. The concentrations of phospholipids reached the highest during the fast state, while those of amino acids reached their peak during fed state. Similar diurnal variations in the metabolite rhythm of amino acids and phospholipids were also observed in IPF mice. Although the rhythmic oscillation of metabolites in the urea cycle remained unchanged, there was a significant up-regulation in their levels in the lungs of IPF mice. 15N-ammonia in vivo isotope tracing further showed an increase in urea cycle activity in the lungs of mice with IPF, which may compensate for the reduced efficiency of the hepatic urea cycle. In sum, our metabolomics database and method provide evidence of the periodic changes in lung metabolites, thereby offering valuable insights to advance our understanding of metabolic reprogramming in the context of IPF.

FG-4592 relieves diabetic kidney disease severity by influencing metabolic profiles via gut microbiota reconstruction in both human and mouse models.

Objective: Diabetic kidney disease (DKD) is one of the most prevalent complications of diabetes mellitus (DM) and is highly associated with devastating outcomes. Hypoxia-inducible factor (HIF), the main transcription factor that regulates cellular responses to hypoxia, plays an important role in regulating erythropoietin (EPO) synthesis. FG-4592 is the HIF stabilizer that is widely used in patients with renal anemia. We investigated the effect of FG-4592 on DKD phenotypes and the pharmacologic mechanism from the perspective of gut microbiota and systemic metabolism. Design: We collected the clinical data of 73 participants, including 40 DKD patients with combined renal anemia treated with FG-4592, and 33 clinical index-matched DKD patients without FG-4592 treatment from The First Affiliated Hospital of Zhengzhou University at the beginning and after a 3-6-month follow-up period. We established DKD mouse models treated by FG-4592 and performed fecal microbiota transplantation from FG-4592-treated DKD mice to investigate the effects of FG-4592 on DKD and to understand this mechanism from a microbial perspective. Untargeted metabolome-microbiome combined analysis was implemented to globally delineate the mechanism of FG-4592 from both microbial and metabolomic aspects. Result: DKD phenotypes significantly improved after 3-6 months of FG-4592 treatment in DKD patients combined with renal anemia, including a decreased level of systolic blood pressure, serum creatinine, and increased estimated glomerular infiltration rate. Such effects were also achieved in the DKD mouse model treated with FG-4592 and can be also induced by FG-4592-influenced gut microbiota. Untargeted plasma metabolomics-gut microbiota analysis showed that FG-4592 dramatically altered both the microbial and metabolic profiles of DKD mice and relieved DKD phenotypes via upregulating beneficial gut microbiota-associated metabolites. Conclusion: FG-4592 can globally relieve the symptoms of DKD patients combined with renal anemia. In the animal experiment, FG-4592 can reconstruct the intestinal microbial profiles of DKD to further upregulate the production of gut-associated beneficial metabolites, subsequently improving DKD phenotypes.

Unveiling Disrupted Lipid Metabolism in Benign Prostate Hyperplasia, Prostate Cancer, and Metastatic Patients: Insights from a Colombian Nested Case-Control Study.

Prostate cancer is a significant global health concern, and its prevalence is increasing worldwide. Despite extensive research efforts, the complexity of the disease remains challenging with respect to fully understanding it. Metabolomics has emerged as a powerful approach to understanding prostate cancer by assessing comprehensive metabolite profiles in biological samples. In this study, metabolic profiles of patients with benign prostatic hyperplasia (BPH), prostate cancer (PCa), and metastatic prostate cancer (Met) were characterized using an untargeted approach that included metabolomics and lipidomics via liquid chromatography and gas chromatography coupled with high-resolution mass spectrometry. Comparative analysis among these groups revealed distinct metabolic profiles, primarily associated with lipid biosynthetic pathways, such as biosynthesis of unsaturated fatty acids, fatty acid degradation and elongation, and sphingolipid and linoleic acid metabolism. PCa patients showed lower levels of amino acids, glycerolipids, glycerophospholipids, sphingolipids, and carnitines compared to BPH patients. Compared to Met patients, PCa patients had reduced metabolites in the glycerolipid, glycerophospholipid, and sphingolipid groups, along with increased amino acids and carbohydrates. These altered metabolic profiles provide insights into the underlying pathways of prostate cancer's progression, potentially aiding the development of new diagnostic, and therapeutic strategies.

Association of Plasma Selenium and Its Untargeted Metabolomic Profiling with Cervical Cancer Prognosis.

Selenium is an essential trace element that shows beneficial or adverse health effects depending on the dose. However, its role in the prognosis of cervical cancer (CC) has been less reported. We aimed to explore the association between selenium status and prognosis in CC patients with different prognoses and to elucidate the underlying mechanism of selenium in CC prognosis. This cross-sectional observational study had a case-control design at the Harbin Medical University Cancer Hospital and was conducted using 29 CC cases with poor prognosis and 29 CC cases with good prognosis. Plasma selenium levels were measured using an atomic fluorescence spectrometer. Untargeted metabolomics was used to identify metabolites. Plasma selenium levels of the poor prognosis group (49.90 ± 13.81 µg/L) were lower than that of the good prognosis group (59.38 ± 13.00 µg/L, t = 2.69, P = 0.009). In the logistic regression analysis, plasma selenium levels were associated with lower poor prognosis risk [odds ratio (OR) = 0.952, 95% CI: 0.909-0.998]. Receiver operating characteristic curve analysis revealed an optimal cut-off point of plasma selenium levels ≤ 47.68 µg/L for poor prognosis of CC. Based on the cut-off selenium levels, patients with different prognoses were divided into high and low selenium groups. Metabolomic analysis revealed six differential metabolites among different prognoses with low and high selenium levels, and the glycerophospholipid (GPL) metabolism was enriched. Plasma selenium levels were positively correlated with metabolite levels. Our findings provided evidence that low plasma selenium levels may associate with a poor prognosis of CC. Low plasma selenium levels might suppress GPL metabolism and influence the prognosis of CC. This finding requires confirmation in future prospective cohort studies.

Comprehensive analysis of the microbiome and metabolome in pus from pyogenic liver abscess patients with and without diabetes mellitus.

Introduction: Pyogenic liver abscess (PLA) patients combined with diabetes mellitus (DM) tend to have more severe clinical manifestations than without DM. The mechanism responsible for this phenomenon is not entirely clear. The current study therefore aimed to comprehensively analyze the microbiome composition and metabolome in pus from PLA patients with and without DM, to determine the potential reasons for these differences. Methods: Clinical data from 290 PLA patients were collected retrospectively. We analyzed the pus microbiota using 16S rDNA sequencing in 62 PLA patients. In addition, the pus metabolomes of 38 pus samples were characterized by untargeted metabolomics analysis. Correlation analyses of microbiota, metabolites and laboratory findings were performed to identify significant associations. Results: PLA patients with DM had more severe clinical manifestations than PLA patients without DM. There were 17 discriminating genera between the two groups at the genus level, among which Klebsiella was the most discriminating taxa. The ABC transporters was the most significant differential metabolic pathway predicted by PICRUSt2. Untargeted metabolomics analysis showed that concentrations of various metabolites were significantly different between the two groups and seven metabolites were enriched in the ABC transporters pathway. Phosphoric acid, taurine, and orthophosphate in the ABC transporters pathway were negatively correlated with the relative abundance of Klebsiella and the blood glucose level. Discussion: The results showed that the relative abundance of Klebsiella in the pus cavity of PLA patients with DM was higher than those without DM, accompanied by changes of various metabolites and metabolic pathways, which may be associated with more severe clinical manifestations.

Untargeted metabolomics analysis in drug-naïve patients with severe obsessive-compulsive disorder.

Introduction: Obsessive-compulsive disorder (OCD), characterized by the presence of obsessions and/or compulsions, is often difficult to diagnose and treat in routine clinical practice. The candidate circulating biomarkers and primary metabolic pathway alteration of plasma in OCD remain poorly understood. Methods: We recruited 32 drug-naïve patients with severe OCD and 32 compared healthy controls and applied the untargeted metabolomics approach by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) to assess their circulating metabolic profiles. Both univariate and multivariate analyses were then utilized to filtrate differential metabolites between patients and healthy controls, and weighted Correlation Network Analysis (WGCNA) was utilized to screen out hub metabolites. Results: A total of 929 metabolites were identified, including 34 differential metabolites and 51 hub metabolites, with an overlap of 13 metabolites. Notably, the following enrichment analyses underlined the importance of unsaturated fatty acids and tryptophan metabolism alterations in OCD. Metabolites of these pathways in plasma appeared to be promising biomarkers, such as Docosapentaenoic acid and 5-Hydroxytryptophan, which may be biomarkers for OCD identification and prediction of sertraline treatment outcome, respectively. Conclusion: Our findings revealed alterations in the circulating metabolome and the potential utility of plasma metabolites as promising biomarkers in OCD.

First Application of Fecal Microbiota Transplantation in Adult Asperger Syndrome With Digestive Symptoms-A Case Report.

Asperger syndrome (AS) is a chronic neurodevelopmental disorder. Although all of the clinically diagnosed cases display normal intelligence and speech functions, barriers in social interaction and communication seriously affect mental health and psychological function. In addition to traditional psychological/behavioral training and symptomatic medication, in-depth studies of intestinal microbiota and mental health have indicated that probiotics (e.g., Lactobacillus rhamnosus) can effectively reduce the occurrence of AS. Fecal microbiota transplantation (FMT) is a type of biological therapy that involves the transplant of intestinal microbiota from healthy donors into the patient's gastrointestinal tract to improve the gut microenvironment. In this case report, we describe the first case of adult AS treated with FMT. The patient suffered from diarrhea-predominant irritable bowel syndrome for 6 years with symptoms of diarrhea and abdominal pain. After three rounds of FMT, the diarrhea and abdominal pain were significantly improved. Moreover, the symptoms of AS were also significantly ameliorated. We found that FMT changed the structure of the intestinal microbiota as well as the patient's serum metabolites, and these changes were consistent with the patient's symptoms. The metabolites may affect signaling pathways, as revealed by Kyoto Encyclopedia of Genes and Genomes enrichment analysis. The changes in microbial metabolites following FMT may affect other regions (e.g., the nervous system) via the circulatory system, such that the bacteria-gut-blood-brain axis may be the means through which FMT mitigates AS.

Serum untargeted metabolomics delineates the metabolic status in different subtypes of non-alcoholic fatty liver disease.

AIMS: The aim of this study was to identify novel serum metabolites associated with non-alcoholic fatty liver disease (NAFLD), and to explore the metabolic discrepancies between Lean-NAFLD and Obese-NAFLD. METHODS: Serum samples from patients with NAFLD (n = 161) and healthy participants (n = 149) were collected, and metabolites were analyzed with UPLC-Q-TOF MS/MS. Subgroup analyses were performed to explore the metabolic differences among Lean-NAFLD, Obese-NAFLD and healthy controls RESULTS: A total of 24 differentially present metabolites were found between patients with NAFLD and healthy controls. Marked metabolic pathway differences were observed among the NAFLD subtypes, including in fatty acid and amino acid metabolism. Ultimately, five metabolites (prasterone, indoxylsulfuric acid, sebacic acid, arachidonic acid and pregnenolone sulfate) were used to establish a diagnostic model to distinguish patients with NAFLD regardless of Lean- or Obese-NAFLD type. CONCLUSIONS: This study suggested that significant metabolic differences existed among subtypes of NAFLD, and our model might be useful to distinguish patients with NAFLD. These findings may lay a foundation for the detection and treatment of NAFLD subtypes.

Gut Microbial Dysbiosis and Changes in Fecal Metabolic Phenotype in Precancerous Lesions of Gastric Cancer Induced With N-Methyl-N'-Nitro-N-Nitrosoguanidine, Sodium Salicylate, Ranitidine, and Irregular Diet.

Background and Aims: Precancerous lesions of gastric cancer (PLGC) are the most important pathological phase with increased risk of gastric cancer (GC) and encompass the key stage in which the occurrence of GC can be prevented. In this study, we found that the gut microbiome changed significantly during the process of malignant transformation from chronic gastritis to GC in N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) multiple factors-induced rat model. Accumulating evidence has shown that alterations in gut microbiota and metabolism are potentially linked to chronic inflammation and cancer of the gastrointestinal tract. However, the correlation of gut microbiota and metabolites, inflammatory factors, and the potential mechanism in the formation of PLGC have not yet been revealed. Methods: In this study, multiple factors including MNNG, sodium salicylate drinking, ranitidine feed, and irregular diet were used to establish a PLGC rat model. The pathological state of the gastric mucosa of rats was identified through HE staining and the main inflammatory cytokine levels in the serum were detected by the Luminex liquid suspension chip (Wayen Biotechnologies, Shanghai, China). The microbial composition and metabolites in the stool samples were tested by using 16S ribosomal RNA (rRNA) gene sequencing and non-targeted metabolomics. The correlation analysis of gut microbiota and inflammatory cytokines in the serum and gut microbiota and differential metabolites in feces was performed to clarify their biological function. Results: The results showed that compared to the control group, the gastric mucosa of the model rats had obvious morphological and pathological malignant changes and the serum levels of inflammatory cytokines including interleukin-1ß (IL-1ß), interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-10 (IL-10), interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and macrophage colony-stimulating factor (M-CSF) increased significantly, while the level of chemokine (C-X-C motif) ligand 1 (CXCL1) in serum reduced significantly. There were significant differences in the composition of the gut microbiota and fecal metabolic profiles between the model and control rats. Among them, Lactobacillus and Bifidobacterium increased significantly, while Turicibacter, Romboutsia, Ruminococcaceae_UCG-014, Ruminococcaceae_UCG-005, and Ruminococcus_1 reduced significantly in the model rats compared to the control rats. The metabolites related to the lipid metabolism and peroxisome proliferator-activated receptor (PPAR) signaling pathway have also undergone significant changes. In addition, there was a significant correlation between the changes of the differential inflammatory cytokines in the serum, fecal metabolic phenotypes, and gut microbial dysbiosis in model rats. Conclusion: The activation of the inflammatory response, disturbance of the gut microbiota, and changes in the fecal metabolic phenotype could be closely related to the occurrence of PLGC. This study provides a new idea to reveal the mechanism of risk factors of chronic gastritis and GC from the perspective of inflammation-immune homeostasis, gut microbiota, and metabolic function balance.