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Abstract Primary Hyperparathyroidism (PHPT) is characterized by excessive parathormone (PTH) secretion and disrupted calcium homeostasis. Untargeted metabolomics offers a valuable approach to understanding the complex metabolic alterations associated with different diseases, including PHPT. Plasma untargeted metabolomics was applied to investigate the metabolic profiles of PHPT patients compared to a control group. Two complementary liquid-phase separation techniques were employed to comprehensively explore the metabolic landscape in this retrospective, single-center study. The study comprised 28 female patients diagnosed following the current guidelines of PHPT diagnosis and a group of 30 healthy females as a control group. To evaluate their association with PHPT, we identified changes in plasma metabolic profiles in patients with PHPT compared to the control group. The primary outcome measure included detecting plasma metabolites and discriminating PHPT patients from controls. The study unveiled specific metabolic imbalances that may link l-amino acids with peptic ulcer disease, gamma-glutamyls with oxidative stress, and asymmetric dimethylarginine (ADMA) with cardiovascular complications. Several metabolites, such as gamma-glutamyls, caffeine, sex hormones, carnitine, sphingosine-1-phosphate (S-1-P), and steroids, were connected with reduced bone mineral density (BMD). Metabolic profiling identified distinct metabolic patterns between patients with PHPT and healthy controls. These findings provided valuable insights into the pathophysiology of PHPT.

Rhizobium leguminosarum bv. viciae (Rlv) UPM791 effectively nodulates pea and lentil, but bacteroids contain a number of proteins differentially expressed depending on the host. One of these host-dependent proteins (C189) is similar to a diaminobutyrate-2-oxoglutarate aminotransferase (DABA-AT). DABA-AT activity was demonstrated with cell extracts and with purified protein, so C189 was renamed as Dat. The dat gene was strongly induced in the central, active area of pea nodules, but not in lentil. Mutants defective in dat were impaired in symbiotic performance with pea plants, exhibiting reduced shoot dry weight, smaller nodules, and a lower competitiveness for nodulation. In contrast, there were no significant differences between mutant and wild-type in symbiosis with lentil plants. A comparative metabolomic approach using cell-free extracts from bacteroids induced in pea and lentil showed significant differences among the strains in pea bacteroids whereas no significant differences were found in lentil. Targeted metabolomic analysis revealed that the dat mutation abolished the presence of 2,4-diaminobutyrate (DABA) in pea nodules, indicating that DABA-AT reaction is oriented toward the production of DABA from L-aspartate semialdehyde. This analysis also showed the presence of L-homoserine, a likely source of aspartate semialdehyde, in pea bacteroids but not in those induced in lentil. The dat mutant showed impaired growth when cells were grown with L-homoserine as nitrogen source. Inclusion of DABA or L-homoserine as N source suppressed pantothenate auxotropy in Rlv UPM791, suggesting DABA as source of the pantothenate precursor β-alanine. These data indicate that Rlv UPM791 Dat enzyme is part of an adaptation mechanism of this bacterium to a homoserine-rich environment such as pea nodule and rhizosphere.

Abstract Background Cholangiocarcinoma (CCA) is still a deadly tumour. Histological and molecular aspects of thioacetamide (TAA)-induced intrahepatic CCA (iCCA) in rats mimic those of human iCCA. Carcinogenic changes and therapeutic vulnerabilities in CCA may be captured by molecular investigations in bile, where we performed bile proteomic and metabolomic analyses that help discovery yet unknown pathways relevant to human iCCA. Methods Cholangiocarcinogenesis was induced in rats (TAA) and mice (Jnk Δhepa + CCl4 + DEN model). We performed proteomic and metabolomic analyses in bile from control and CCA-bearing rats. Differential expression was validated in rat and human CCAs. Mechanisms were addressed in human CCA cells, including Huh28-KRASG12D cells. Cell signaling, growth, gene regulation and [U-13C]-D-glucose-serine fluxomics analyses were performed. In vivo studies were performed in the clinically-relevant iCCA mouse model. Results Pathways related to inflammation, oxidative stress and glucose metabolism were identified by proteomic analysis. Oxidative stress and high amounts of the oncogenesis-supporting amino acids serine and glycine were discovered by metabolomic studies. Most relevant hits were confirmed in rat and human CCAs (TCGA). Activation of interleukin-6 (IL6) and epidermal growth factor receptor (EGFR) pathways, and key genes in cancer-related glucose metabolic reprogramming, were validated in TAA-CCAs. In TAA-CCAs, G9a, an epigenetic pro-tumorigenic writer, was also increased. We show that EGFR signaling and mutant KRASG12D can both activate IL6 production in CCA cells. Furthermore, phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine-glycine pathway, was upregulated in human iCCA correlating with G9a expression. In a G9a activity-dependent manner, KRASG12D promoted PHGDH expression, glucose flow towards serine synthesis, and increased CCA cell viability. KRASG12D CAA cells were more sensitive to PHGDH and G9a inhibition than controls. In mouse iCCA, G9a pharmacological targeting reduced PHGDH expression. Conclusions In CCA, we identified new pro-tumorigenic mechanisms: Activation of EGFR signaling or KRAS mutation drives IL6 expression in tumour cells; Glucose metabolism reprogramming in iCCA includes activation of the serine-glycine pathway; Mutant KRAS drives PHGDH expression in a G9a-dependent manner; PHGDH and G9a emerge as therapeutic targets in iCCA.

Leishmaniases are widespread neglected diseases with an incidence of 1.6 million new cases and 40 thousand deaths per year. Leishmania parasites may show distinct, species-specific patterns of virulence that lead to different clinical manifestations. It is well known that successive in vitro passages (SIVP) lead to the attenuation of virulence, but neither the metabolism nor the pathways involved in these processes are well understood. Herein, promastigotes of a virulent L. amazonensis strain recently isolated from mice was compared to SIVP derived and attenuated promastigotes, submitted to 10, 40, and 60 axenic passages and named R10, R40, and R60, respectively. In vitro assays and in vivo tests were performed to characterize and confirmed the attenuation profiles. A metabolomic fingerprint comparison of R0, R10, and R60 was performed by means of capillary electrophoresis, liquid and gas chromatography coupled to mass spectrometry. To validate the metabolomic data, qPCR for selected loci, flow cytometry to measure aPS exposure, sensitivity to antimony tartrate and ROS production assays were conducted. The 65 identified metabolites were clustered in biochemical categories and mapped in eight metabolic pathways: ABC transporters; fatty acid biosynthesis; glycine, serine and threonine metabolism; β-alanine metabolism; glutathione metabolism; oxidative phosphorylation; glycerophospholipid metabolism and lysine degradation. The obtained metabolomic data correlated with previous proteomic findings of the SVIP parasites and the gene expression of 13 selected targets. Late SIVP cultures were more sensitive to SbIII produced more ROS and exposed less phosphatidylserine in their surface. The correspondent pathways were connected to build a biochemical map of the most significant alterations involved with the process of attenuation of L. amazonensis. Overall, the reported data pointed out to a very dynamic and continuous metabolic reprogramming process, accompanied by changes in energetic, lipid and redox metabolisms, membrane remodeling and reshaping of parasite-host cells interactions, causing impacts in chemotaxis, host inflammatory responses and infectivity at the early stages of infection.

There is a rising resistance against antimony drugs, the gold-standard for treatment until some years ago. That is a serious problem due to the paucity of drugs in current clinical use. In a research to reveal how these drugs affect the parasite during treatment and to unravel the underlying basis for their resistance, we have employed metabolomics to study treatment in Leishmania infantum promastigotes. This was accomplished first through the untargeted analysis of metabolic snapshots of treated and untreated parasites both resistant and responders, utilizing a multiplatform approach to give the widest as possible coverage of the metabolome, and additionally through novel monitoring of the origin of the detected alterations through a 13C traceability experiment. Our data stress a multi-target metabolic alteration with treatment, affecting in particular the cell redox system that is essential to cope with detoxification and biosynthetic processes. Additionally, relevant changes were noted in amino acid metabolism. Our results are in agreement with other authors studying other Leishmania species.