Rheumatoid Arthritis (RA) is a chronic autoimmune disease associated with inflammation and joint remodeling. Adenosine deaminase (ADA), a risk factor in RA, degrades adenosine, an anti-inflammatory molecule, resulting in an inflammatory bias. We present an integrative analysis of clinical data, cytokines, serum metabolomics in RA patients and mechanistic studies on ADA-mediated effects on in vitro cell culture models. ADA activity differentiated patients into low and high ADA sets. The levels of the cytokines TNFα, IFNγ, IL-10, TGFβ and sRANKL were elevated in RA and more pronounced in high ADA sets. Serum metabolomic analysis shows altered metabolic pathways in RA which were distinct between low and high ADA sets. Comparative analysis with previous studies shows similar pathways are modulated by DMARDs and biologics. Random forest analysis distinguished RA from control by methyl-histidine and hydroxyisocaproic acid, while hexose-phosphate and fructose-6-phosphate distinguished high ADA from low ADA. The deregulated metabolic pathways of High ADA datasets significantly overlapped with high ADA expressing PBMCs GEO transcriptomics dataset. ADA induced the death of chondrocytes, synoviocyte proliferation, both inflammation in macrophages and their differentiation into osteoclasts and impaired differentiation of mesenchymal stem cells to osteoblasts and mineralization. PBMCs expressing elevated ADA had increased expression of cytokines and P2 receptors compared to synovial macrophages which has low expression of ADA. Our data demonstrates increased cytokine levels and distinct metabolic signatures of RA based on the ADA activity, suggests an important role for ADA in the pathophysiology of RA joints and as a potential marker and therapeutic target in RA patients.
Glaucoma of which primary open angle glaucoma (POAG) constitutes 75%, is the second leading cause of blindness. Elevated intra ocular pressure and Nitric oxide synthase (NOS) dysfunction are hallmarks of POAG. We analyzed clinical data, cytokine profile, ATP level, metabolomics and GEO datasets to identify features unique to POAG. N9 microglial cells are used to gain mechanistic insights. Our POAG cohort showed elevated ATP in aqueous humor and cytokines in plasma. Metabolomic analysis showed changes in 21 metabolites including Dimethylarginine (DMAG) and activation of tryptophan metabolism in POAG. Analysis of GEO data sets and previously published proteomic data sets bins genes into signaling and metabolic pathways. Pathways from reanalyzed metabolomic data from literature significantly overlapped with those from our POAG data. DMAG modulated purinergic signaling, ATP secretion and cytokine expression were inhibited by N-Ethylmaleimide, NO donors, BAPTA and purinergic receptor inhibitors. ATP induced elevated intracellular calcium level and cytokines expression were inhibited by BAPTA. Metabolomics of cell culture supernatant from ATP treated sets showed metabolic deregulation and activation of tryptophan metabolism. DMAG and ATP induced IDO1/2 and TDO2 were inhibited by N-Ethylmaleimide, sodium nitroprusside and BAPTA. Our data obtained from clinical samples and cell culture studies reveal a strong association of elevated DMAG, ATP, cytokines and activation of tryptophan metabolism with POAG. DMAG mediated ATP signaling, inflammation and metabolic remodeling in microglia might have implications in management of POAG.
Huntington's disease (HD) is a neurodegenerative disease associated with polyglutamine expansion in the protein Huntingtin. Though the polyglutamine repeat length correlates with the age of onset and severity, the complication points to disease modifiers. Mitochondrial dysfunction and metabolic deregulation are associated with the disease. Despite multi-omic characterization of patients and model systems, the mechanisms have remained elusive. Systems analysis of multi-omics data and its validation using yeast model could help to elucidate pathways that modulate protein aggregation. Metabolomic analysis of HD patients and yeast model of HD was carried out. Our analysis shows a considerable overlap of deregulated metabolic pathways. Further, our multi-omic analysis shows deregulated pathways that are common to human, mice and yeast model systems and those that are unique to them. The deregulated pathways include metabolism of various amino acids, glutathione metabolism, longevity, autophagy and mitophagy. Addition of selected metabolites and gene knockout from the deregulated pathways in yeast model system shows that they modulate protein aggregation. Taken together our results show modulation of deregulated pathways influences protein aggregation in HD with implications for progression and prognosis.
Amyotrophic lateral sclerosis (ALS) is a multi-systemic, incurable, amyloid disease affecting the motor neurons, resulting in the death of patients. The disease is either sporadic or familial with SOD1, C9orf72, FUS, and TDP-43 constituting the majority of familial ALS. Multi-omics studies on patients and model systems like mice and yeast have helped in understanding the association of various signaling and metabolic pathways with the disease. The yeast model system has played a pivotal role in elucidating the gene amyloid interactions. We carried out an integrated transcriptomic and metabolomic analysis of the TDP-43 expressing yeast model to elucidate deregulated pathways associated with the disease. The analysis shows the deregulation of the TCA cycle, single carbon metabolism, glutathione metabolism, and fatty acid metabolism. Transcriptomic analysis of GEO datasets of TDP-43 expressing motor neurons from mice models of ALS and ALS patients shows considerable overlap with experimental results. Furthermore, a yeast model was used to validate the obtained results using metabolite addition and gene knock-out experiments. Taken together, our result shows a potential role for the TCA cycle, cellular redox pathway, NAD metabolism, and fatty acid metabolism in disease. Supplementation of reduced glutathione, nicotinate, and the keto diet might help to manage the disease.
Neem tree (Azadirachta indica) is one of the richest sources of
secondary metabolites. More than 250 natural products have been
characterized from various parts of the neem tree. These include
diterpenoids, triterpenoids, steroids, flavonoids, coumarins, hydrocarbons,
and fatty acids. Many of these products possess therapeutic properties. Neem
exudate or toddy is a milky white liquid with a strong smell secreted from
the angle between the 2 main branches of old trees. Profiling of neem toddy
for the presence of active metabolites was done by an in-house database
using UHPLC-QTOF-MS. Fifty-seven metabolites were identified from the full
scan of electrospray ionization positive and negative mode from the neem
toddy extract by using UHPLC-QTOF-MS. Further confirmation of 31 of these
metabolites was done by obtaining MS/MS spectrum from UHPLC-QTOF-MS.
Principal component analysis study of metabolites from neem toddy with
leaves, seed, seed coat, and bark revealed that they are closely related to
those contained in neem seeds and seed coats. Azadirachtin, nimbidiol,
22,23-dihydroazadirachtin, nimbonone, nimbonolone, nimosone, and
6-deacetylnimbinene were found to be some of the most abundant metabolites
in neem toddy. The neem toddy extract showed significant anti-inflammatory
activity when tested in N9 murine microglial cells with 25 ng of recombinant
mouse tumor necrosis factor alpha protein (active) using qRT-PCR. The active
metabolites in neem toddy could be further explored for their therapeutic
potentials.
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