RESUMEN
Metabolic dysfunction-associated steatotic liver disease (MASLD) is a chronic liver condition that often progresses to more advanced stages, such as metabolic dysfunction-associated steatohepatitis (MASH). MASH is characterized by inflammation and hepatocellular ballooning, in addition to hepatic steatosis. Despite the relatively high incidence of MASH in the population and its potential detrimental effects on human health, this liver disease is still not fully understood from a pathophysiological perspective. Deregulation of polyamine levels has been detected in various pathological conditions, including neurodegenerative diseases, inflammation, and cancer. However, the role of the polyamine pathway in chronic liver disorders such as MASLD has not been explored. In this study, we measured the expression of liver ornithine decarboxylase (ODC1), the rate-limiting enzyme responsible for the production of putrescine, and the hepatic levels of putrescine, in a preclinical model of MASH as well as in liver biopsies of patients with obesity undergoing bariatric surgery. Our findings reveal that expression of ODC1 and the levels of putrescine, but not spermidine nor spermine, are elevated in hepatic tissue of both diet-induced MASH mice and patients with biopsy-proven MASH compared with control mice and patients without MASH, respectively. Furthermore, we found that the levels of putrescine were positively associated with higher aspartate aminotransferase concentrations in serum and an increased SAF score (steatosis, activity, fibrosis). Additionally, in in vitro assays using human HepG2 cells, we demonstrate that elevated levels of putrescine exacerbate the cellular response to palmitic acid, leading to decreased cell viability and increased release of CK-18. Our results support an association between the expression of ODC1 and the progression of MASLD, which could have translational relevance in understanding the onset of this disease. © 2024 The Pathological Society of Great Britain and Ireland.
Asunto(s)
Progresión de la Enfermedad , Hígado , Ornitina Descarboxilasa , Putrescina , Animales , Humanos , Putrescina/metabolismo , Ornitina Descarboxilasa/metabolismo , Hígado/metabolismo , Hígado/patología , Masculino , Ratones Endogámicos C57BL , Hígado Graso/metabolismo , Hígado Graso/patología , Ratones , Modelos Animales de Enfermedad , Femenino , Persona de Mediana Edad , Obesidad/metabolismo , Obesidad/complicaciones , Células Hep G2 , AdultoRESUMEN
RNA interference (RNAi) has not been tested in the pandemic amphibian pathogen, Batrachochytrium dendrobatidis, but developing this technology could be useful to elucidate virulence mechanisms, identify therapeutic targets, and may present a novel antifungal treatment option for chytridiomycosis. To manipulate and decipher gene function, rationally designed small interfering RNA (siRNA) can initiate the destruction of homologous messenger RNA (mRNA), resulting in the "knockdown" of target gene expression. Here, we investigate whether siRNA can be used to manipulate gene expression in B. dendrobatidis via RNAi using differing siRNA strategies to target genes involved in glutathione and ornithine synthesis. To determine the extent and duration of mRNA knockdown, target mRNA levels were monitored for 24-48 h after delivery of siRNA targeting glutamate-cysteine ligase, with a maximum of ~56% reduction in target transcripts occurring at 36 h. A second siRNA design targeting glutamate-cysteine ligase also resulted in ~53% knockdown at this time point. siRNA directed toward a different gene target, ornithine decarboxylase, achieved 17% reduction in target transcripts. Although no phenotypic effects were observed, these results suggest that RNAi is possible in B. dendrobatidis, and that gene expression can be manipulated in this pathogen. We outline ideas for further optimization steps to increase knockdown efficiency to better harness RNAi techniques for control of B. dendrobatidis.
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Quitridiomicetos , Técnicas de Silenciamiento del Gen , Glutamato-Cisteína Ligasa , Interferencia de ARN , ARN Mensajero , ARN Interferente Pequeño , ARN Interferente Pequeño/genética , ARN Mensajero/genética , Animales , Quitridiomicetos/genética , Quitridiomicetos/patogenicidad , Glutamato-Cisteína Ligasa/genética , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Anfibios/microbiología , Glutatión/metabolismoRESUMEN
Bee venom serves as an essential defensive weapon for bees and also finds application as a medicinal drug. MicroRNAs (miRNAs) serve as critical regulators and have been demonstrated to perform a variety of biological functions. However, the presence of miRNAs in bee venom needs to be confirmed. Therefore, we conducted small RNA sequencing and identified 158 known miRNAs, 15 conserved miRNAs and 4 novel miRNAs. It is noteworthy that ame-miR-1-3p, the most abundant among them, accounted for over a quarter of all miRNA reads. To validate the function of ame-miR-1-3p, we screened 28 candidate target genes using transcriptome sequencing and three target gene prediction software (miRanda, PITA and TargetScan) for ame-miR-1-3p. Subsequently, we employed real-time quantitative reverse transcription PCR (qRT-PCR), Western blot and other technologies to confirm that ame-miR-1-3p inhibits the relative expression of antizyme inhibitor 1 (AZIN1) by targeting the 3' untranslated region (UTR) of AZIN1. This, in turn, caused ODC antizyme 1 (OAZ1) to bind to ornithine decarboxylase 1 (ODC1) and mark ODC1 for proteolytic destruction. The reduction in functional ODC1 ultimately resulted in a decrease in polyamine biosynthesis. Furthermore, we determined that ame-miR-1-3p accelerates cell death through the AZIN1/OAZ1-ODC1-polyamines pathway. Our studies demonstrate that ame-miR-1-3p diminishes cell viability and it may collaborate with sPLA2 to enhance the defence capabilities of honeybees (Apis mellifera L.). Collectively, these data further elucidate the defence mechanism of bee venom and expand the potential applications of bee venom in medical treatment.
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Venenos de Abeja , Proteínas de Insectos , MicroARNs , Animales , Abejas/genética , Abejas/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Venenos de Abeja/farmacología , Proteínas de Insectos/metabolismo , Proteínas de Insectos/genética , Supervivencia Celular , Poliaminas/metabolismo , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/genéticaRESUMEN
As demonstrated in previous research, hsa_circ_0052602 (circODC1) is dynamically expressed in HPV-positive cervical cancer (CC). CircODC1 expression was quantified using qRT-PCR, and its role in CC cell growth was assessed via loss-of-function assays. Interactions between miR-607 and circODC1 or ODC1 were confirmed using bioinformatics and mechanistic assays. The association of FOXA1 with the circODC1 promoter was validated through ChIP and luciferase reporter assays. CircODC1 was highly expressed in HPV-positive CC cell lines, and its depletion significantly impeded malignant processes such as proliferation, migration, and invasion. We found that ODC1 also played an oncogenic role in HPV-positive CC cells. CircODC1 was shown to positively regulate ODC1 as a ceRNA, competitively binding to miR-607 to counteract its suppression of ODC1. HPV-associated FOXA1 was identified as a potential transcription factor of circODC1. Restoration experiments showed that overexpression of circODC1 could counterbalance the inhibitory effect of FOXA1 knockdown. These findings offer new insights into therapeutic strategies for HPV-positive CC patients.
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Proliferación Celular , Factor Nuclear 3-alfa del Hepatocito , Ornitina Descarboxilasa , Neoplasias del Cuello Uterino , Femenino , Humanos , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Factor Nuclear 3-alfa del Hepatocito/metabolismo , Factor Nuclear 3-alfa del Hepatocito/genética , MicroARNs/genética , MicroARNs/metabolismo , Infecciones por Papillomavirus/virología , Infecciones por Papillomavirus/genética , ARN Circular/genética , ARN Circular/metabolismo , Neoplasias del Cuello Uterino/virología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología , Neoplasias del Cuello Uterino/metabolismo , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismoRESUMEN
Non-small-cell lung cancer (NSCLC), a common malignant tumor, requires deeper pathogenesis investigation. Autophagy is an evolutionarily conserved lysosomal degradation process that is frequently blocked during cancer progression. It is an urgent need to determine the novel autophagy-associated regulators in NSCLC. Here, we found that pirin was upregulated in NSCLC, and its expression was positively correlated with poor prognosis. Overexpression of pirin inhibited autophagy and promoted NSCLC proliferation. We then performed data-independent acquisition-based quantitative proteomics to identify the differentially expressed proteins (DEPs) in pirin-overexpression (OE) or pirin-knockdown (KD) cells. Among the pirin-regulated DEPs, ornithine decarboxylase 1 (ODC1) was downregulated in pirin-KD cells while upregulated along with pirin overexpression. ODC1 depletion reversed the pirin-induced autophagy inhibition and pro-proliferation effect in A549 and H460 cells. Immunohistochemistry showed that ODC1 was highly expressed in NSCLC cancer tissues and positively related with pirin. Notably, NSCLC patients with pirinhigh/ODC1high had a higher risk in terms of overall survival. In summary, we identified pirin and ODC1 as a novel cluster of prognostic biomarkers for NSCLC and highlighted the potential oncogenic role of the pirin/ODC1/autophagy axis in this cancer type. Targeting this pathway represents a possible therapeutic approach to treat NSCLC.
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Autofagia , Carcinoma de Pulmón de Células no Pequeñas , Proliferación Celular , Progresión de la Enfermedad , Neoplasias Pulmonares , Ornitina Descarboxilasa , Femenino , Humanos , Masculino , Células A549 , Autofagia/genética , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Línea Celular Tumoral , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/metabolismo , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/genética , Pronóstico , Regulación hacia ArribaRESUMEN
Despite the well-known relevance of polyamines to many forms of life, little is known about how polyamines regulate osteogenesis and skeletal homeostasis. Here, we report a series of in vitro studies conducted with human-bone-marrow-derived pluripotent stromal cells (MSCs). First, we show that during osteogenic differentiation, mRNA levels of most polyamine-associated enzymes are relatively constant, except for the catabolic enzyme spermidine/spermine N1-acetyltransferase 1 (SAT1), which is strongly increased at both mRNA and protein levels. As a result, the intracellular spermidine to spermine ratio is significantly reduced during the early stages of osteoblastogenesis. Supplementation of cells with exogenous spermidine or spermine decreases matrix mineralization in a dose-dependent manner. Employing N-cyclohexyl-1,3-propanediamine (CDAP) to chemically inhibit spermine synthase (SMS), the enzyme catalyzing conversion of spermidine into spermine, also suppresses mineralization. Intriguingly, this reduced mineralization is rescued with DFMO, an inhibitor of the upstream polyamine enzyme ornithine decarboxylase (ODC1). Similarly, high concentrations of CDAP cause cytoplasmic vacuolization and alter mitochondrial function, which are also reversible with the addition of DFMO. Altogether, these studies suggest that excess polyamines, especially spermidine, negatively affect hydroxyapatite synthesis of primary MSCs, whereas inhibition of polyamine synthesis with DFMO rescues most, but not all of these defects. These findings are relevant for patients with Snyder-Robinson syndrome (SRS), as the presenting skeletal defects-associated with SMS deficiency-could potentially be ameliorated by treatment with DFMO.
Asunto(s)
Células Madre Mesenquimatosas , Espermidina , Humanos , Espermidina/metabolismo , Espermina/metabolismo , Espermina Sintasa/genética , Ornitina Descarboxilasa/metabolismo , Osteogénesis , Poliaminas/metabolismo , Células Madre Mesenquimatosas/metabolismo , ARN MensajeroRESUMEN
Citrus Huanglongbing, one of the most devastating citrus diseases, is caused by 'Candidatus Liberibacter asiaticus' (CLas). Polyamines are aliphatic nitrogen-containing compounds that play important roles in disease resistance and are synthesized primarily by two pathways: an arginine decarboxylation pathway and an ornithine decarboxylation pathway. However, it is unclear whether polyamines play a role in the tolerance of citrus to infection by CLas and, if so, whether one or both of the core polyamine metabolic pathways are important. We used high-performance liquid chromatography and ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry to detect the contents of nine polyamine metabolism-related compounds in six citrus cultivars with varying levels of tolerance to CLas. We also systematically detected the changes in polyamine metabolism-related compounds and H2O2 contents and compared the gene expression levels and the activities of enzymes involved in the polyamine metabolic pathway among healthy, asymptomatic, and symptomatic leaves of Newhall navel oranges infected with CLas. The tolerant and moderately tolerant varieties showed higher polyamine metabolism-related compound levels than those of susceptible varieties. Compared with the healthy group, the symptomatic group showed significantly increased contents of arginine, ornithine, γ-aminobutyric acid, and putrescine by approximately 180, 19, 1.5, and 0.2 times, respectively, and upregulated expression of biosynthetic genes. Arginase and ornithine decarboxylase enzyme activities were the highest in the symptomatic group, whereas arginine decarboxylase and agmatine deiminase enzyme activities were the highest in the asymptomatic group. The two polyamine biosynthetic pathways showed different trends with the increase of the CLas titer, indicating that polyamines were mainly synthesized through the arginine decarboxylase pathway in the asymptomatic leaves and were synthesized via the ornithine decarboxylase pathway in symptomatic leaves. These findings provide new insight into the changes in polyamine metabolism in citrus infected with CLas.
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Citrus , Enfermedades de las Plantas , Poliaminas , Rhizobiaceae , Poliaminas/metabolismo , Enfermedades de las Plantas/microbiología , Citrus/microbiología , Rhizobiaceae/fisiología , Hojas de la Planta/microbiología , Hojas de la Planta/metabolismo , Peróxido de Hidrógeno/metabolismo , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/genética , Liberibacter/fisiología , Regulación de la Expresión Génica de las Plantas , Redes y Vías MetabólicasRESUMEN
This perspective delves into the investigation of synthetic and naturally occurring inhibitors, their patterns of inhibition, and the effectiveness of newly utilized natural compounds as inhibitors targeting the Ornithine decarboxylase enzyme. This enzyme is known to target the MYC oncogene, thereby establishing a connection between polyamine metabolism and oncogenesis in both normal and cancerous cells. ODC activation and heightened polyamine activity are associated with tumor development in numerous cancers and fluctuations in ODC protein levels exert a profound influence on cellular activity for inhibition or suppressing tumor cells. This perspective outlines efforts to develop novel drugs, evaluate natural compounds, and identify promising inhibitors to address gaps in cancer prevention, highlighting the potential of newly designed synthetic moieties and natural flavonoids as alternatives. It also discusses natural compounds with potential as enhanced inhibitors.
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Inhibidores de la Ornitina Descarboxilasa , Ornitina Descarboxilasa , Humanos , Inhibidores de la Ornitina Descarboxilasa/farmacología , Poliaminas/farmacología , Poliaminas/metabolismo , Flavonoides , Transformación Celular NeoplásicaRESUMEN
Clostridium aceticum DSM1496 is an acid-resistant strain in which ornithine decarboxylase (ODC) plays a crucial role in acid resistance. In this study, we expressed ODC derived from C. aceticum DSM1496 in Escherichia coli BL21 (DE3) and thoroughly examined its enzymatic properties. The enzyme has a molecular weight of 55.27 kDa and uses pyridoxal-5'-phosphate (PLP) as a coenzyme with a Km = 0.31 mM. ODC exhibits optimal activity at pH 7.5, and it maintains high stability even at pH 4.5. The peak reaction temperature for ODC is 30°C. Besides, it can be influenced by certain metal ions such as Mn2+. Although l-ornithine serves as the preferred substrate for ODC, the enzyme also decarboxylates l-arginine and l-lysine simultaneously. The results indicate that ODC derived from C. aceticum DSM1496 exhibits the ability to produce putrescine, cadaverine, and agmatine through decarboxylation. These polyamines have the potential to neutralize acid in an acidic environment, facilitating the growth of microorganisms. These significant findings provide a strong basis for further investigation into the acid-resistant mechanisms contributed by ODC.
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Ornitina Descarboxilasa , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/química , Concentración de Iones de Hidrógeno , Escherichia coli/metabolismo , Escherichia coli/enzimologíaRESUMEN
Alzheimer's disease (AD) is characterized by the loss of memory due to aggregation of misphosphorylated tau and amyloid beta (Aß) plaques in the brain, elevated release of inhibitory neurotransmitter gamma-aminobutyric acid (GABA) and reactive oxygen species from astrocytes, and subsequent neurodegeneration. Recently, it was found that enzyme Ornithine Decarboxylase 1 (ODC1) acts as a bridge between the astrocytic urea cycle and the putrescine-to-GABA conversion pathway in the brain of AD mouse models as well as human patients. In this study, we show that the long-term knockdown of astrocytic Odc1 in APP/PS1 animals was sufficient to completely clear Aß plaques in the hippocampus while simultaneously switching the astrocytes from a detrimental reactive state to a regenerative active state, characterized by proBDNF expression. Our experiments also reveal an effect of astrocytic ODC1 inhibition on the expression of genes involved in synapse pruning and organization, histone modification, apoptotic signaling and protein processing. These genes are previously known to be associated with astrocytic activation and together create a neuroregeneration-supportive environment in the brain. By inhibiting ODC1 for a long period of 3 months in AD mice, we demonstrate that the beneficial amyloid-clearing process of astrocytes can be completely segregated from the systemically harmful astrocytic response to insult. Our study reports an almost complete clearance of Aß plaques by controlling an endogenous degradation process, which also modifies the astrocytic state to create a regeneration-supportive environment in the brain. These findings present the potential of modulating astrocytic clearance of Aß as a powerful therapeutic strategy against AD.
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Enfermedad de Alzheimer , Péptidos beta-Amiloides , Animales , Humanos , Ratones , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Astrocitos/metabolismo , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Ácido gamma-Aminobutírico/metabolismo , Ratones Transgénicos , Placa Amiloide/complicaciones , Ornitina DescarboxilasaRESUMEN
Metabolism of polyamines is of critical importance to physiological processes. Ornithine decarboxylase (ODC) antizyme inhibitors (AZINs) are capable of interacting with antizymes (AZs), thereby releasing ODC from ODC-AZs complex, and promote polyamine biosynthesis. AZINs regulate reproduction, embryonic development, fibrogenesis and tumorigenesis through polyamine and other signaling pathways. Dysregulation of AZINs has involved in multiple human diseases, especially malignant tumors. Adenosine-to-inosine (A-to-I) RNA editing is the most common type of post-transcriptional nucleotide modification in humans. Additionally, the high frequencies of RNA-edited AZIN1 in human cancers correlates with increase of cancer cell proliferation, enhancement of cancer cell stemness, and promotion of tumor angiogenesis. In this review, we summarize the current knowledge on the various contribution of AZINs related with potential cancer promotion, cancer stemness, microenvironment and RNA modification, especially underlying molecular mechanisms, and furthermore explored its promising implication for cancer diagnosis and treatment.
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Ornitina Descarboxilasa , Investigación Biomédica Traslacional , Humanos , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Poliaminas/metabolismo , Transformación Celular Neoplásica , ARN , Microambiente TumoralRESUMEN
The brown planthopper (BPH, Nilaparvata lugens), a major insect pest of rice, can make a shift in wing dimorphism to adapt to complex external environments. Our previous study showed that NlODC (Ornithine decarboxylase in N. lugens) was involved in wing dimorphism of the brown planthopper. Here, further experiments were conducted to reveal possible molecular mechanism of NlODC in manipulating the wing dimorphism. We found that the long-winged rate (LWR) of BPH was significantly reduced after RNAi of NlODC or injection of DFMO (D, L-α-Difluoromethylornithine), and LWR of males and females significantly decreased by 21.7% and 34.6%, respectively. Meanwhile, we also examined the contents of three polyamines under DFMO treatment and found that the contents of putrescine and spermidine were significantly lower compared to the control. After 3rd instar nymphs were injected with putrescine and spermidine, LWR was increased significantly in both cases, and putrescine was a little bit more effective, with 5.6% increase in males and 11.4% in females. Three days after injection of dsNlODC, injection of putrescine and spermidine rescued LWR to the normal levels. In the regulation of wing differentiation in BPH, NlODC mutually antagonistic to NlAkt may act through other signaling pathways rather than the classical insulin signaling pathway. This study illuminated a physiological function of an ODC gene involved in wing differentiation in insects, which could be a potential target for pest control.
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Hemípteros , Ornitina Descarboxilasa , Femenino , Masculino , Animales , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Hemípteros/fisiología , Caracteres Sexuales , Putrescina/metabolismo , Espermidina/metabolismoRESUMEN
PURPOSE: Skin exposure to noxious agents leads to cutaneous lesion marked by an increase in inflammation, cellular proliferation, and hyperplasiogenic reactions. Studies have demonstrated that these damages breach the skin integrity resulting in the aetiology of various cutaneous disorders like atopic dermatitis, eczema, psoriasis, and development of non-melanoma skin cancer. Celecoxib, a cyclooxygenase-2 (COX-2) inhibitor, is an effective treatment for a variety of inflammatory diseases. Its importance in the therapy of skin problems, however, remains under appreciated. METHODS: We tested efficacy of topically applied celecoxib in mitigating skin inflammation, cellular proliferation, and hyperplasia induced by the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) in Swiss albino mice. RESULTS: Celecoxib (5 and 10 µmol) markedly reduced TPA (10 nmol) induced prostaglandin E2 (PGE2) production, oedema formation, myeloperoxidase (MPO) activity, and levels of pro-inflammatory cytokines such as tumour necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), and interleukin-6 (IL-6). It also resulted in a considerable decrease in ornithine decarboxylase (ODC) activity and the incorporation of [3H]-thymidine into DNA. In addition, there was a significant reduction in histoarchitectural abnormalities such as epidermal thickness, number of epidermal cell layers, neutrophil infiltration, intercellular oedema, and vasodilation. CONCLUSION: Our results demonstrate that topical celecoxib can reduce the inflammation, hyperproliferation, and hyperplasiogenic events of skin insults suggesting that it may prove to be a valuable management option for cutaneous lesion and associated illnesses such as atopic dermatitis, eczema, and psoriasis, as well as the emergence of non-melanoma cancer.
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Dermatitis Atópica , Eccema , Psoriasis , Enfermedades de la Piel , Neoplasias Cutáneas , Ratones , Animales , Celecoxib/efectos adversos , Dermatitis Atópica/metabolismo , Dermatitis Atópica/patología , Ornitina Descarboxilasa/metabolismo , Ornitina Descarboxilasa/farmacología , Piel , Acetato de Tetradecanoilforbol/toxicidad , Acetato de Tetradecanoilforbol/metabolismo , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Enfermedades de la Piel/patología , Psoriasis/patología , Edema/metabolismo , Acetatos/efectos adversos , Acetatos/metabolismo , Eccema/metabolismo , Eccema/patología , Neoplasias Cutáneas/patologíaRESUMEN
In preclinical models, α-difluoromethylornithine (DFMO), an ornithine decarboxylase (ODC) inhibitor, delays the onset of type 1 diabetes (T1D) by reducing ß cell stress. However, the mechanism of DFMO action and its human tolerability remain unclear. In this study, we show that mice with ß cell ODC deletion are protected against toxin-induced diabetes, suggesting a cell-autonomous role of ODC during ß cell stress. In a randomized controlled trial (ClinicalTrials.gov: NCT02384889) involving 41 recent-onset T1D subjects (3:1 drug:placebo) over a 3-month treatment period with a 3-month follow-up, DFMO (125-1,000 mg/m2) is shown to meet its primary outcome of safety and tolerability. DFMO dose-dependently reduces urinary putrescine levels and, at higher doses, preserves C-peptide area under the curve without apparent immunomodulation. Transcriptomics and proteomics of DFMO-treated human islets exposed to cytokine stress reveal alterations in mRNA translation, nascent protein transport, and protein secretion. These findings suggest that DFMO may preserve ß cell function in T1D through islet cell-autonomous effects.
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Diabetes Mellitus Tipo 1 , Humanos , Ratones , Animales , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/metabolismo , Inhibidores de la Ornitina Descarboxilasa/farmacología , Eflornitina/farmacología , Eflornitina/uso terapéutico , Putrescina/metabolismoRESUMEN
The enzyme arginase 1 (A1) hydrolyzes the amino acid arginine to form L-ornithine and urea. Ornithine is further converted to polyamines by the ornithine decarboxylase (ODC) enzyme. We previously reported that deletion of myeloid A1 in mice exacerbates retinal damage after ischemia/reperfusion (IR) injury. Furthermore, treatment with A1 protects against retinal IR injury in wild-type mice. PEG-A1 also mitigates the exaggerated inflammatory response of A1 knockout (KO) macrophages in vitro. Here, we sought to identify the anti-inflammatory pathway that confers macrophage A1-mediated protection against retinal IR injury. Acute elevation of intraocular pressure was used to induce retinal IR injury in mice. A multiplex cytokine assay revealed a marked increase in the inflammatory cytokines interleukin 1ß (IL-1ß) and tumor necrosis factor α (TNF-α) in the retina at day 5 after IR injury. In vitro, blocking the A1/ODC pathway augmented IL-1ß and TNF-α production in stimulated macrophages. Furthermore, A1 treatment attenuated the stimulated macrophage metabolic switch to a pro-inflammatory glycolytic phenotype, whereas A1 deletion had the opposite effect. Screening for histone deacetylases (HDACs) which play a role in macrophage inflammatory response showed that A1 deletion or ODC inhibition increased the expression of HDAC3. We further showed the involvement of HDAC3 in the upregulation of TNF-α but not IL-1ß in stimulated macrophages deficient in the A1/ODC pathway. Investigating HDAC3 KO macrophages showed a reduced inflammatory response and a less glycolytic phenotype upon stimulation. In vivo, HDAC3 co-localized with microglia/macrophages at day 2 after IR in WT retinas and was further increased in A1-deficient retinas. Collectively, our data provide initial evidence that A1 exerts its anti-inflammatory effect in macrophages via ODC-mediated suppression of HDAC3 and IL-1ß. Collectively we propose that interventions that augment the A1/ODC pathway and inhibit HDAC3 may confer therapeutic benefits for the treatment of retinal ischemic diseases.
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Daño por Reperfusión , Enfermedades de la Retina , Animales , Ratones , Arginasa/genética , Citocinas , Isquemia , Células Mieloides , Ornitina , Ornitina Descarboxilasa , Factor de Necrosis Tumoral alfaRESUMEN
Rhizosphere soil of aromatic rice inhabits different fungal species that produce many bioactive metabolites including 2-acetyl-1-pyrroline (2AP). The mechanism for the biosynthesis of 2AP in the fungal system is still elusive. Hence, the present study investigates the role of possible nitrogen (N) precursors such as some amino acids and polyamines as well as the enzymes involved in 2AP synthesis in the fungal species isolated from the rhizosphere of aromatic rice varieties. Three fungal isolates were found to synthesize 2AP (0.32-1.07 ppm) and maximum 2AP was synthesized by Aspergillus niger (1.07 ppm) isolated from rhizosphere of Dehradun Basmati (DB). To determine the N source for 2AP synthesis, various N sources such as proline, glutamate, ornithine putrescine, spermine, and spermidine were used in place of putrescine in the synthetic medium (Syn18). The results showed that maximum 2AP synthesis was found with putrescine (1.07 ppm) followed by spermidine (0.89 ppm) and spermine (0.84 ppm). Further, LC-QTOF-MS analysis revealed the mobilization of spermine and spermidine into the putrescine, indicating that putrescine is the key N source for 2AP synthesis. Moreover, higher enzyme activity of DAO, PAO, and ODC as well as higher content of methylglyoxal metabolite in the A. niger NFCCI 5060 as compared to A. niger NFCCI 4064 (control) suggests the prominent role of these enzymes in the synthesis of 2AP. In conclusion, this study showed evidence of the polyamines mediated 2AP biosynthesis in A. niger NFCCI 5060.
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Oryza , Poliaminas , Poliaminas/metabolismo , Espermidina/metabolismo , Putrescina/metabolismo , Espermina/metabolismo , Aspergillus niger/genética , Aspergillus niger/metabolismo , Oryza/metabolismo , Ornitina Descarboxilasa/metabolismoRESUMEN
The pyridoxal 5'-phosphate (PLP) acts as a coenzyme for a large number of biochemical reactions. It exists in mainly two bound forms at the active site of the concerned enzyme: the internal aldimine, in which the PLP is bound with the ϵ-amino group of lysine at the active site, and the external aldimine, where the PLP is bound to the substrate amino acid. Both the internal and external aldimines have Schiff base linkage (N-H-O) and can exist in two tautomeric structures of ketoenamine and enolimine forms. In this work, we have investigated the free energy landscape for the tautomeric proton transfer in the internal and external aldimines at the active site of the ornithine decarboxylase enzyme in an aqueous medium. We performed hybrid quantum-classical metadynamics and force field-based molecular dynamics simulations, which revealed that the ketoenamine tautomer is more stable than the enolimine form. The QM/MM metadynamics calculations show that the free energy difference between the ketoenamine and enolimine forms for the internal aldimine is 3.9 kcal/mol, and it is found to be 5.8 kcal/mol for the external aldimine, with the ketoenamine form being more stable in both cases. The results are further supported by calculations of the binding free energies from classical simulations and static quantum chemical calculations in different environments. We have also analyzed the configurational structure of the microenvironment at the active site in order to have better insights into the interactions of the active site residues with the PLP in its two tautomeric forms.
Asunto(s)
Ornitina Descarboxilasa , Bases de Schiff , Dominio Catalítico , Ornitina Descarboxilasa/metabolismo , Bases de Schiff/química , Protones , Fosfato de Piridoxal/química , FosfatosRESUMEN
S-adenosylmethionine decarboxylase (AdoMetDC/SpeD) is a key polyamine biosynthetic enzyme required for conversion of putrescine to spermidine. Autocatalytic self-processing of the AdoMetDC/SpeD proenzyme generates a pyruvoyl cofactor from an internal serine. Recently, we discovered that diverse bacteriophages encode AdoMetDC/SpeD homologs that lack AdoMetDC activity and instead decarboxylate L-ornithine or L-arginine. We reasoned that neofunctionalized AdoMetDC/SpeD homologs were unlikely to have emerged in bacteriophages and were probably acquired from ancestral bacterial hosts. To test this hypothesis, we sought to identify candidate AdoMetDC/SpeD homologs encoding L-ornithine and L-arginine decarboxylases in bacteria and archaea. We searched for the anomalous presence of AdoMetDC/SpeD homologs in the absence of its obligatory partner enzyme spermidine synthase, or the presence of two AdoMetDC/SpeD homologs encoded in the same genome. Biochemical characterization of candidate neofunctionalized genes confirmed lack of AdoMetDC activity, and functional presence of L-ornithine or L-arginine decarboxylase activity in proteins from phyla Actinomycetota, Armatimonadota, Planctomycetota, Melainabacteria, Perigrinibacteria, Atribacteria, Chloroflexota, Sumerlaeota, Omnitrophota, Lentisphaerota, and Euryarchaeota, the bacterial candidate phyla radiation and DPANN archaea, and the δ-Proteobacteria class. Phylogenetic analysis indicated that L-arginine decarboxylases emerged at least three times from AdoMetDC/SpeD, whereas L-ornithine decarboxylases arose only once, potentially from the AdoMetDC/SpeD-derived L-arginine decarboxylases, revealing unsuspected polyamine metabolic plasticity. Horizontal transfer of the neofunctionalized genes appears to be the more prevalent mode of dissemination. We identified fusion proteins of bona fide AdoMetDC/SpeD with homologous L-ornithine decarboxylases that possess two, unprecedented internal protein-derived pyruvoyl cofactors. These fusion proteins suggest a plausible model for the evolution of the eukaryotic AdoMetDC.
Asunto(s)
Adenosilmetionina Descarboxilasa , Carboxiliasas , Adenosilmetionina Descarboxilasa/genética , Adenosilmetionina Descarboxilasa/metabolismo , Archaea/genética , Archaea/metabolismo , Ornitina , Filogenia , Carboxiliasas/genética , Carboxiliasas/metabolismo , Poliaminas/metabolismo , Bacterias/metabolismo , Ornitina Descarboxilasa/metabolismo , Arginina/genéticaRESUMEN
Polyamines promote cellular proliferation. Their levels are controlled by ornithine decarboxylase antizyme 1 (Az1, encoded by OAZ1), through the proteasome-mediated, ubiquitin-independent degradation of ornithine decarboxylase (ODC), the rate-limiting enzyme of polyamine biosynthesis. Az1-mediated degradation of other substrates such as cyclin D1 (CCND1), DNp73 (TP73) or Mps1 regulates cell growth and centrosome amplification, and the currently known six Az1 substrates are all linked with tumorigenesis. To understand whether Az1-mediated protein degradation might play a role in regulating other cellular processes associated with tumorigenesis, we employed quantitative proteomics to identify novel Az1 substrates. Here, we describe the identification of LIM domain and actin-binding protein 1 (LIMA1), also known as epithelial protein lost in neoplasm (EPLIN), as a new Az1 target. Interestingly, between the two EPLIN isoforms (α and ß), only EPLIN-ß is a substrate of Az1. The interaction between EPLIN-ß and Az1 appears to be indirect, and EPLIN-ß is degraded by Az1 in a ubiquitination-independent manner. Az1 absence leads to elevated EPLIN-ß levels, causing enhanced cellular migration. Consistently, higher LIMA1 levels correlate with poorer overall survival of colorectal cancer patients. Overall, this study identifies EPLIN-ß as a novel Az1 substrate regulating cellular migration.
Asunto(s)
Ornitina Descarboxilasa , Ubiquitina , Humanos , Ornitina Descarboxilasa/genética , Ornitina Descarboxilasa/química , Ornitina Descarboxilasa/metabolismo , Ubiquitina/metabolismo , Isoformas de Proteínas , Carcinogénesis , Proteínas del CitoesqueletoRESUMEN
Visceral leishmaniasis (VL) is caused by Leishmania donovani (Ld), and most cases occur in Brazil, East Africa, and India. The treatment for VL is limited and has many adverse effects. The development of safer and more efficacious drugs is urgently needed. Drug repurposing is one of the best processes to repurpose existing drugs. Ornithine decarboxylase (ODC) is an important target against L. donovani in the polyamine biosynthesis pathway. In this study, we have modeled the 3D structure of ODC and performed high-throughput virtual screening of 8630 ZINC database ligands against Leishmania donovani ornithine decarboxylase (Ld ODC), selecting 45 ligands based on their high binding score. It is further validated through molecular docking simulation and the selection of the top two lead molecules (ceftaroline fosamil and rimegepant) for Molecular Dynamics (MD) simulation, Density functional theory (DFT), and molecular mechanics generalized born surface area (MMGBSA) analysis. The results showed that the binding affinities of ceftaroline fosamil, and rimegepant are, respectively, -10.719 and 10.159 kcal/mol. The docking complexes of the two lead compounds, ceftaroline fosamil, and rimegepant, with the target ODC, were found stable during molecular dynamics simulations. Furthermore, the analysis of MMGBSA revealed that these compounds had a high binding free energy. The DFT analysis showed that the top lead molecules were more reactive than the standard drug (pentamidine). In-silico findings demonstrated that ceftaroline fosamil, and rimegepant might be recognized as potent antagonists against ODC for the treatment of VL.