Ornithine Decarboxylase Expression Research Articles

Increased hepatic putrescine levels as a new potential factor related to the progression of metabolic dysfunction-associated steatotic liver disease.

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.

Mechanism of electro-acupuncture in alleviating intestinal injury in septic mice via polyamine-related M2-macrophage polarization.

The objective of this study was to investigate the impact of electro-acupuncture (EA) on sepsis-related intestinal injury and its relationship with macrophage polarization. A sepsis model was established using cecal ligation and puncture (CLP) to assess the effectiveness of EA. The extent of pathological injury was evaluated using Chiu's score, the expression of ZO-1 and Ocludin, and the impact on macrophage polarization was examined through flow cytometry and immunofluorescence staining. The expression of spermidine, one type of polyamine, and ornithine decarboxylase (ODC) was measured using ELISA and PCR. Once the efficacy was determined, a polyamine depletion model was created, and the role of polyamines was reassessed by evaluating efficacy and observing macrophage polarization. EA treatment reduced the Chiu's score and increased the expression of ZO-1 and Ocludin in the intestinal tissue of septic mice. It inhibited the secretion of IL-1β and TNF-α, promoted the polarization of M2-type macrophages, increased the secretion of IL-10, and upregulated the expression of Arg-1, spermidine, and ODC. However, after depleting polyamines, the beneficial effects of EA on alleviating intestinal tissue damage and modulating macrophage polarization disappeared. The mechanism underlying the alleviation of intestinal injury associated with CLP-induced sepsis by EA involves with the promotion of M2-type macrophage polarization mediated by spermidine expression.

Abstract 18: The polyamine-hypusine axis is a promising avenue to improve T-cell metabolism and persistence for adoptive cell therapy

Abstract There are significant barriers to the application of Adoptive Cell Therapy (ACT) to many cancer types. One of these barriers is a result of poor metabolic fitness of T cells, leading to poor proliferation, persistence, and dampened anti-tumor function. Leveraging T-cell metabolism to improve their function is becoming increasingly important to make significant strides in immunotherapy. Stat5a has been shown to be a master regulator of energy and metabolism in T cells, highlighting it as a promising avenue to improve T-cell metabolism for ACT. We have established that the expression of Constitutively Active Signal Transducer and Activator of Transcription 5A (CASTAT5) on tumor-specific CD4+ T cells improves their persistence, polyfunctionality, and anti-tumor effects in a mouse model. This study further examines Stat5a and its downstream metabolic targets in the context of ACT. We carried out scRNAseq analysis and found that Stat5a directly interacts with genes involved in polyamine metabolism. More specifically, Stat5a activation increases the expression of ornithine decarboxylase (Odc1), spermidine synthase (Srm), deoxyhypusine synthase (Dhps), and deoxyhypusine hydroxylase (Dohh) in a subset of CD4+ T cells expressing CASTAT5. Cleavage Under Targets and Tagmentation (Cut&Tag) analysis of histone H3K27ac also demonstrated increased H3K27ac at promoter and enhancer regions of polyamine pathway genes in CASTAT5 compared to control CD4+ T cells. To further examine the function of Stat5a, the murine pro-B cell line, Ba/F3, is being used as a model to study Stat5a-mediated signaling. We have found that upon withdrawal of IL-3 from Ba/F3 cells, Stat5a activation is significantly downregulated. Our scRNA-seq-based metabolic flux estimation showed a significant reduction in metabolic fluxes and related genes, including polyamines, following IL-3 withdrawal. Correspondingly, Stat5a ChIP-Seq and Cut&Tag analyses of H3K27ac revealed Stat5a binding to active enhancer regions in the presence of IL-3 that are lost following its withdrawal. Untargeted metabolomics confirmed the marked Stat5a-mediated reduction in metabolite abundance after IL-3 deprivation, in a time-dependent manner. The polyamine-hypusine axis revealed in our studies is a promising avenue to improve T-cell function. This axis has been shown to regulate T-cell function, and potentially modulate mitochondrial respiration and T-cell activation. Hence, our studies have identified polyamine supplementation and gene modulation as promising strategies to improve the metabolism of Adoptive T cells in a Stat5a-mediated manner. Further studies will evaluate the effectiveness of this avenue in improving T-cell function, improving their efficacy for ACT, and improving patient outcomes. Citation Format: Mercy Kehinde-Ige, Ogacheko Okoko, Gang Zhou, Huidong Shi. The polyamine-hypusine axis is a promising avenue to improve T-cell metabolism and persistence for adoptive cell therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 18.

Polyamine-mediated ferroptosis amplification acts as a targetable vulnerability in cancer

Targeting ferroptosis, an iron-dependent form of regulated cell death triggered by the lethal overload of lipid peroxides, in cancer therapy is impeded by our limited understanding of the intersection of tumour’s metabolic feature and ferroptosis vulnerability. In the present study, arginine is identified as a ferroptotic promoter using a metabolites library. This effect is mainly achieved through arginine’s conversion to polyamines, which exerts their potent ferroptosis-promoting property in an H2O2-dependent manner. Notably, the expression of ornithine decarboxylase 1 (ODC1), the critical enzyme catalysing polyamine synthesis, is significantly activated by the ferroptosis signal——iron overload——through WNT/MYC signalling, as well as the subsequent elevated polyamine synthesis, thus forming a ferroptosis-iron overload-WNT/MYC-ODC1-polyamine-H2O2 positive feedback loop that amplifies ferroptosis. Meanwhile, we notice that ferroptotic cells release enhanced polyamine-containing extracellular vesicles into the microenvironment, thereby further sensitizing neighbouring cells to ferroptosis and accelerating the “spread” of ferroptosis in the tumour region. Besides, polyamine supplementation also sensitizes cancer cells or xenograft tumours to radiotherapy or chemotherapy through inducing ferroptosis. Considering that cancer cells are often characterized by elevated intracellular polyamine pools, our results indicate that polyamine metabolism exposes a targetable vulnerability to ferroptosis and represents an exciting opportunity for therapeutic strategies for cancer.

Polyamine Catabolism and Its Role in Renal Injury and Fibrosis in Mice Subjected to Repeated Low-Dose Cisplatin Treatment.

Cisplatin, a chemotherapeutic agent, can cause nephrotoxic and ototoxic injuries. Using a mouse model of repeated low dose cisplatin (RLDC), we compared the kidneys of cisplatin- and vehicle-treated mice on days 3 (early injury phase) and 35 (late injury/recovery phase) after the final treatment. RNA-seq analyses revealed increases in the expression of markers of kidney injury (e.g., lipocalin 2 and kidney injury molecule 1) and fibrosis (e.g., collagen 1, fibronectin, and vimentin 1) in RLDC mice. In addition, we observed increased expression of polyamine catabolic enzymes (spermidine/spermine N1-acetyltransferase, Sat1, and spermine oxidase, Smox) and decreased expression of ornithine decarboxylase (Odc1), a rate-limiting enzyme in polyamine synthesis in mice subjected to RLDC. Upon confirmation of the RNA-seq results, we tested the hypothesis that enhanced polyamine catabolism contributes to the onset of renal injury and development of fibrosis. To test our hypothesis, we compared the severity of RLDC-induced renal injury and fibrosis in wildtype (WT), Sat1-KO, and Smox-KO mice. Our results suggest that the ablation of polyamine catabolic enzymes reduces the severity of renal injury and that modulation of the activity of these enzymes may protect against kidney damage and fibrosis caused by cisplatin treatment.

Elevated expression of HIGD1A drives hepatocellular carcinoma progression by regulating polyamine metabolism through c-Myc–ODC1 nexus

BackgroundHypoxia contributes to cancer progression through various molecular mechanisms and hepatocellular carcinoma (HCC) is one of the most hypoxic malignancies. Hypoxia-inducible gene domain protein-1a (HIGD1A) is typically induced via epigenetic regulation and promotes tumor cell survival during hypoxia. However, the role of HIGD1A in HCC remains unknown.MethodsHIGD1A expression was determined in 24 pairs of human HCC samples and para-tumorous tissues. Loss-of-function experiments were conducted both in vivo and in vitro to explore the role of HIGD1A in HCC proliferation and metastasis.ResultsIncreased HIGD1A expression was found in HCC tissues and cell lines, which was induced by hypoxia or low-glucose condition. Moreover, HIGD1A knockdown in HCC cells arrested the cell cycle at the G2/M phase and promoted hypoxia-induced cell apoptosis, resulting in great inhibition of cell proliferation, migration, and invasion, as well as tumor xenograft formation. Interestingly, these anti-tumor effects were not observed in normal hepatocyte cell line L02. Further, HIGD1A knockdown suppressed the expression of ornithine decarboxylase 1 (ODC1), a rate-limiting enzyme of polyamine metabolism under c-Myc regulation. HIGD1A was found to bind with the c-Myc promoter region, and its knockdown decreased the levels of polyamine metabolites. Consistently, the inhibitory effect on HCC phenotype by HIGD1A silencing could be reversed by overexpression of c-Myc or supplementation of polyamines.ConclusionsOur results demonstrated that HIGD1A activated c-Myc–ODC1 nexus to regulate polyamine synthesis and to promote HCC survival and malignant phenotype, implying that HIGD1A might represent a novel therapeutic target for HCC.

Expression of ornithine decarboxylase in peripheral blood mononuclear cells from patients with pancreatic adenocarcinoma: A preliminary report.

Ductal adenocarcinoma represents 90-95% of pancreatic cancer (PC) cases and it is an aggressive disease with asymptomatic evolution at early stages, non-specific symptoms and a typical late diagnosis with a 5-year survival rate estimated to be 8%. A window of opportunity lies in early diagnosis as there are currently no reliable biomarkers. CA 19-9 is one of the most frequently used biomarkers of PC, with 75 and 77.6% sensitivity (Se) and specificity (Sp), respectively, and the carcinoembryonic antigen (CEA) shows 39.5 and 81.3% of Se and Sp, respectively. A case-control study was conducted including adult patients with a histological diagnosis of PC (n=11) without previous treatment at the Oncology Service of the CMNO-IMSS between 2019 and 2020, and a control group of adult volunteers (n=11) who were clinically healthy or with controlled disease including hypertension, hypothyroidism and diabetes. Clinical, laboratory and sociodemographic data as well as blood, urine and saliva samples were collected following patient consent. Polyamines were quantified using high-performance liquid chromatography with fluorescence detection, CA 19-9 and CEA were evaluated using enzyme-linked immunosorbent assay, and the protein expression of ornithine decarboxylase (ODC) was evaluated using western blotting. Polyamine metabolism and modulation by means of ODC were increased in the serum and saliva of patients with PC, and the expression of ODC alone was increased in peripheral blood mononuclear cells (PBMCs). The present study focused on the evaluation of putrescine, spermine, spermidine and ODC in PBMCs associated with CA 19-9 and CEA as an auxiliary tool in PC diagnosis.

The roles of polyamines in Malaysian herbs as a chemoprevention strategy: A scoping review

Maintaining normal polyamine levels is crucial for the physiological cellular process and differentiation because an imbalance of polyamine concentration affects the occurrence of neoplastic activities in many types of cancer diseases. Malaysian herbs have been purported to have anticancer properties that influence the polyamine’s bioavailability, leading to cancer cell death. Presently, the roles of polyamines in Malaysian herbs as chemoprevention have yet to be documented. Thus, this review aims to determine which Malaysian herbs have chemoprevention properties affecting polyamines and appraise the relationship between polyamine levels and Malaysian herbs and their cytotoxicity on cancer cells. The search, employing keywords like “Malaysian herbs”, “polyamines”, and “anticancer” or “chemoprevention”, yielded 256 articles. The articles were screened for their eligibility, befitting the objectives mentioned. The PRISMA-ScR checklist was employed to report the published findings from the databases. Thirteen articles meeting inclusion criteria were selected, highlighting 13 Malaysian herbs influencing the intracellular polyamines pathway. All herbs showed a significant reduction in ornithine decarboxylase (ODC) expression. In addition, Alpinia galanga and Aloe barbadensis showed an increase in catabolic polyamines expression, spermidine/spermine-N-acetyl transferase (SSAT) and spermine oxidase (SMO), respectively, leading to growth arrest and induction of apoptosis in cancer cells. The in vitro and in vivo studies demonstrated these herbs’ anti-proliferative effects on diverse cancer types, involving polyamine level modulation, intracellular dynamics, and chemopreventive mechanisms through apoptosis and anti-inflammatory responses. The findings underscore the potential of Malaysian herbs in cancer prevention by modulating the polyamine pathway.

Carboxylic acid-functionalized multiwalled carbon nanotubes (COOH-MWCNTs) improved production of atropine in callus of Datura inoxia by influencing metabolism, gene regulation, and DNA cytosine methylation; an in vitro biological assessment

Atropine is a well-known tropane alkaloid commonly employed in medicine class called anticholinergics. This study intends to address biochemical and molecular responses of Datura inoxia calluses to fortifying culture medium with carboxylic acid-functionalized multi-walled carbon nanotubes (COOH-MWCNTs). The application of MWCNTs influenced callogenesis performance and biomass in a dose-dependent manner. The MWCNT at 5 mgL−1 resulted in the highest biomass of calluses by 57%. While, MWCNTs at high concentrations were accompanied by cytotoxicity. On the other hand, MWCNTs at concentrations above 100 mgL−1 exhibited cytotoxicity, decreased callogenesis performance, and reduced Atropine biosynthesis. The MWCNTs increased the activity of phenylalanine ammonia-lyase (PAL) and catalase enzymes. The concentrations of proline and soluble phenols displayed upward trends in response to using MWCNTs. According to the HPLC assessment, enriching culture medium with MWCNTs at 5 mgL−1 elicited Atropine production in calluses by 64%. The quantitative PCR assessment referred to the upregulation in the transcription of the PAL gene. The expression of ornithine decarboxylase (ODC) and putrescine N-methyltransferase 1 (PMT) genes were also upregulated in calluses cultured in a medium supplemented with MWCNTs. Methylation Sensitive Amplification Polymorphism (MSAP) technique indicated that employing MWCNTs altered the DNA methylation profile, reflecting epigenetic modification. Overall, engineering plant cells with MWCNTs as a nano-elicitor can be suggested for large-scale synthesis of industrially-valuable secondary metabolites.

Abstract 5905: Enterotoxigenic Bacteroides fragilis, a colon microbe, dysregulates polyamine catabolism to promote breast cancer progression

Abstract Background: With more than 2.3 million cases worldwide, breast cancer persists as a major health burden and warrants a greater understanding of the molecular triggers underlying this disease. Focusing on microbial dysbiosis, our lab discovered the pivotal role of the enterotoxigenic Bacteroides fragilis (ETBF) in promoting breast tumorigenesis through activation of key oncogenic pathways. A gap in knowledge exists regarding how microbes/microbial toxins modulate the oncogenic pathways in cancer cells. Polyamines are imperative for various physiological processes, including cell growth, survival, differentiation and apoptosis and are strongly associated with breast cancer. We questioned the involvement of polyamine pathway in mediating the oncogenic effects of microbial dysbiosis on breast cancer. Methods: Human Her2 expressing spontaneous breast cancer model, MMTV.f.HuHer2 mice, were infected with ETBF and tumor incidence and progression were monitored for 18 months. We used qRT-PCR, western blotting, immunocytochemistry and immunohistochemistry to examine expression of key nodes of polyamine pathway. Various growth, migration and invasion assays were performed to assess carcinogenic properties in breast cancer cells. We employed a colorimetric-based assay to analyze enzymatic activities of key polyamine metabolic members. ROS generation was examined via DCFA staining. Results: Our in vivo data showed that ETBF colonization in mice led to a considerable increase in the level of spermine oxidase (SMOX) in breast tumors, hinting at a correlation between ETBF and polyamine metabolism. Moreover, a marked rise in expression of γH2AX foci was observed in these tumors and in breast cancer cells, suggesting a critical role of ETBF in SMOX-related DNA damage. In agreement, we found SMOX upregulation in breast cancer cells. Our findings displayed a substantial increase in SMOX enzymatic activity in the presence of ETBF toxin, BFT, in breast tumor cells. BFT caused a marked rise in ROS generation of breast tumor cells, suggesting a plausible association of ETBF in SMOX-mediated oxidative stress. Interestingly, we noted a significant induction of ornithine decarboxylase (ODC) expression and activity in response to ETBF or BFT in breast cancer cells. Treatment of BFT-exposed breast cancer cells with SMOX inhibitors led to a sharp inhibition of neoplastic transformations, like cellular migration and γH2AX induction. Conclusion: Our collated evidences suggest the existence of an important cross-talk between key polyamine metabolic enzymes and ETBF that aids in exacerbating breast tumorigenesis. Of importance, therapeutic inhibition of SMOX dramatically impeded BFT-related breast tumorigenesis. Collectively, our results demonstrated the attractive potential of intervening the polyamine catabolic pathway for the management of breast cancer. Citation Format: Deeptashree Nandi, Sheetal Parida, Sowjanya Thatikonda, Jackson Foley, Tracy Stewart, Cynthia L. Sears, Robert A. Casero, Dipali Sharma. Enterotoxigenic Bacteroides fragilis, a colon microbe, dysregulates polyamine catabolism to promote breast cancer progression. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5905.

Ornithine metabolism and the osmotic stress response in mytilid congeners

ABSTRACT Previous transcriptomic studies have suggested that ornithine decarboxylase (ODC) expression in Mytilids (Mytilus spp.) may be a critical component of the osmotic stress response. Ornithine is catabolized into polyamines by ODC or into glutamate or proline by ornithine aminotransferase (OAT). To better understand how ornithine is metabolized under low salinity conditions, we examined species-specific variation in OAT and ODC expression for three species that vary in their salinity tolerance (M. trossulus, M. edulis, and M. galloprovincialis). We found a consistent decrease in ODC expression during hypoosmotic exposure in all three species but pronounced species-specific increases in OAT expression. During hyperosmotic stress, the patterns of expression of these genes reversed, suggesting that proline or glutamate synthesis is important during low salinity exposure, while polyamine synthesis may be more important during hyperosmotic exposure. These responses were most pronounced in M. galloprovincialis, with an 18-fold increase in OAT during hypoosmotic exposure.

A new 68Ga-labeled ornithine derivative for PET imaging of ornithine metabolism in tumors.

Ornithine metabolism plays a vital role in tumorigenesis. For cancer cells, ornithine is mainly used as a substrate for ornithine decarboxylase (ODC) for the synthesis of polyamines. The ODC as a key enzyme of polyamine metabolism has become an important target for cancer diagnosis and treatment. To non-invasively detect the levels of ODC expression in malignant tumors, we have synthesized a novel 68Ga-labeled ornithine derivative ([68Ga]Ga-NOTA-Orn). The synthesis time of [68Ga]Ga-NOTA-Orn was about 30min with a radiochemical yield of 45-50% (uncorrected), and the radiochemical purity was > 98%. [68Ga]Ga-NOTA-Orn was stable in saline and rat serum. Cellular uptake and competitive inhibition assays using DU145 and AR42J cells demonstrated that the transport pathway of [68Ga]Ga-NOTA-Orn was similar to that of L-ornithine, and it could interact with the ODC after transporting into the cell. Biodistribution and micro-positron emission tomography (Micro-PET) imaging studies showed that [68Ga]Ga-NOTA-Orn exhibited rapid tumor uptake and was rapidly excreted through the urinary system. All above results suggested that [68Ga]Ga-NOTA-Orn is a novel amino acid metabolic imaging agent with great potential of tumor diagnosis.

P53 inhibits the Urea cycle and represses polyamine biosynthesis in glioma cell lines.

Glioma is the most common malignant tumor of the central nervous system. The urea cycle (UC) is an essential pathway to convert excess nitrogen and ammonia into the less toxic urea in humans. However, less is known about the functional significance of the urea cycle in glioma. p53 functions as a tumor suppressor and modulates several cellular functions and disease processes. In the present study, we aimed to explore whether p53 influences glioma progression by regulating the urea cycle. Here, we demonstrated the inhibitory impact of p53 on the expression of urea cycle enzymes and urea genesis in glioma cells. The level of polyamine, a urea cycle metabolite, was also regulated by p53 in glioma cells. Carbamoyl phosphate synthetase-1 (CPS1) is the first key enzyme involved in the urea cycle. Functionally, we demonstrated that CPS1 knockdown suppressed glioma cell proliferation, migration and invasion. Mechanistically, we demonstrated that the expression of ornithine decarboxylase (ODC), which determines the generation of polyamine, was regulated by CPS1. In addition, the impacts of p53 knockdown on ODC expression, glioma cell growth and aggressive phenotypes were significantly reversed by CPS1 inhibition. In conclusion, these results demonstrated that p53 inhibits polyamine metabolism by suppressing the urea cycle, which inhibits glioma progression.

Gasdermin D Deficiency in Vascular Smooth Muscle Cells Ameliorates Abdominal Aortic Aneurysm Through Reducing Putrescine Synthesis.

Abdominal aortic aneurysm (AAA) is a common vascular disease associated with significant phenotypic alterations in vascular smooth muscle cells (VSMCs). Gasdermin D (GSDMD) is a pore-forming effector of pyroptosis. In this study, the role of VSMC-specific GSDMD in the phenotypic alteration of VSMCs and AAA formation is determined. Single-cell transcriptome analyses reveal Gsdmd upregulation in aortic VSMCs in angiotensin (Ang) II-induced AAA. VSMC-specific Gsdmd deletion ameliorates Ang II-induced AAA in apolipoprotein E (ApoE)-/- mice. Using untargeted metabolomic analysis, it is found that putrescine is significantly reduced in the plasma and aortic tissues of VSMC-specific GSDMD deficient mice. High putrescine levels trigger a pro-inflammatory phenotype in VSMCs and increase susceptibility to Ang II-induced AAA formation in mice. In a population-based study, a high level of putrescine in plasma is associated with the risk of AAA (p < 2.2 × 10-16 ), consistent with the animal data. Mechanistically, GSDMD enhances endoplasmic reticulum stress-C/EBP homologous protein (CHOP) signaling, which in turn promotes the expression of ornithine decarboxylase 1 (ODC1), the enzyme responsible for increased putrescine levels. Treatment with the ODC1 inhibitor, difluoromethylornithine, reduces AAA formation in Ang II-infused ApoE-/- mice. The findings suggest that putrescine is a potential biomarker and target for AAA treatment.

Heterologous expression and activity verification of ornithine decarboxylase from a wild strain of Shewanella xiamenensis.

Shewanella xiamenensis is widely found in spoilage fish, shrimp and other seafoods. Under suitable conditions, ornithine can be synthesized into putrescine, which may spoil food or endanger health. Our research used a wild strain of Shewanella xiamenensis isolated from "Yi Lu Xian" salted fish (a salting method for sea bass) as a research object. According to the database of National Center of Biotechnology Information (NCBI), the target ornithine decarboxylase (ODC) gene SpeF was successfully amplified using the wild strain of Shewanella xiamenensis as the template. Sequencing alignment showed that the SpeF of the wild strain had more than 98% homology compared with the standard strain. The amino acid substitution occurred in the residues of 343, 618, 705, and 708 in the wild strain. After optimizing the expression conditions, a heterologous expression system of ODC was constructed to achieve a high yield of expression. The amount of 253.38 mg of ODC per liter of LB broth was finally expressed. High performance liquid chromatography (HPLC) and subsequent ODC activity verification experiments showed that hetero-expressed ODC showed a certain enzyme activity for about 11.91 ± 0.38 U/mg. Our study gives a new way to the development of a low-cost and high-yield strategy to produce ODC, providing experimental materials for further research and elimination of putrescine in food hazards.

Uric Acid Deteriorates Load-Free Cell Shortening of Cultured Adult Rat Ventricular Cardiomyocytes via Stimulation of Arginine Turnover.

Hyperuricemia is a risk factor for heart disease. Cardiomyocytes produce uric acid via xanthine oxidase. The enzymatic reaction leads to oxidative stress in uric-acid-producing cells. However, extracellular uric acid is the largest scavenger of reactive oxygen species, specifically to nitrosative stress, which can directly affect cells. Here, the effect of plasma-relevant concentrations of uric acid on adult rat ventricular cardiomyocytes is analyzed. A concentration- and time-dependent reduction of load-free cell shortening is found. This is accompanied by an increased protein expression of ornithine decarboxylase, the rate-limiting enzyme of the polyamine metabolism, suggesting a higher arginine turnover. Subsequently, the effect of uric acid was attenuated if other arginine consumers, such as nitric oxide synthase, are blocked or arginine is added. In the presence of uric acid, calcium transients are increased in cardiomyocytes irrespective of the reduced cell shortening, indicating calcium desensitization. Supplementation of extracellular calcium or stimulation of intracellular calcium release by β-adrenergic receptor stimulation attenuates the uric-acid-dependent effect. The effects of uric acid are attenuated in the presence of a protein kinase C inhibitor, suggesting that the PKC-dependent phosphorylation of troponin triggers the desensitizing effect. In conclusion, high levels of uric acid stress cardiomyocytes by accelerating the arginine metabolism via the upregulation of ornithine decarboxylase.

Compromised glycolysis contributes to foot process fusion of podocytes in diabetic kidney disease: Role of ornithine catabolism

IntroductionCompromised glycolysis in podocytes contributes to the initiation of diabetic kidney disease (DKD). Podocyte injury is characterized by cytoskeletal remodeling and foot process fusion. Compromised glycolysis in diabetes likely leads to switch of energy supply in podocyte. However, the underlying mechanism by which disturbed energy supply in podocytes affects the cytoskeletal structure of podocytes remains unclear. MethodsMetabolomic and transcriptomic analyses were performed on the glomeruli of db/db mice to examine the catabolism of glucose, fatty, and amino acids. Ornithine catabolism was targeted in db/db and podocyte-specific pyruvate kinase M2 knockout (PKM2-podoKO) mice. In vitro, expression of ornithine decarboxylase (ODC1) was modulated to investigate the effect of ornithine catabolism on mammalian target of rapamycin (mTOR) signaling and cytoskeletal remodeling in cultured podocytes. ResultsMulti-omic analyses of the glomeruli revealed that ornithine metabolism was enhanced in db/db mice compared with that in db/m mice under compromised glycolytic conditions. Additionally, ornithine catabolism was exaggerated in podocytes of diabetic PKM2-podoKO mice compared with that in diabetic PKM2flox/flox mice. In vivo, difluoromethylornithine (DFMO, inhibitor of ODC1) administration reduced urinary albumin excretion and alleviated podocyte foot process fusion in db/db mice. In vitro, 2-deoxy-d-glucose (2-DG) exposure induced mTOR signaling activation and cytoskeletal remodeling in podocytes, which was alleviated by ODC1-knockdown. Mechanistically, a small GTPase Ras homolog enriched in the brain (Rheb), a sensor of mTOR signaling, was activated by exposure to putrescine, a metabolic product of ornithine catabolism. ConclusionThese findings demonstrate that compromised glycolysis in podocytes under diabetic conditions enhances ornithine catabolism. The metabolites of ornithine catabolism contribute to mTOR signaling activation via Rheb and cytoskeletal remodeling in podocytes in DKD.

The sexual dimorphism of kidney growth in mice and humans

Kidney mass and function are sexually determined, but the cellular events and the molecular mechanisms involved in this dimorphism are poorly characterized. By combining female and male mice with castration/replacement experiments, we showed that male mice exhibited kidney overgrowth from five weeks of age. This effect was organ specific, since liver and heart weight were comparable between males and females, regardless of age. Consistently, the androgen receptor was found to be expressed in the kidneys of males, but not in the liver. In growing mice, androgens led to kidney overgrowth by first inducing a burst of cell proliferation and then an increase of cell size. Remarkably, androgens were also required to maintain cell size in adults. In fact, orchiectomy resulted in smaller kidneys in a matter of few weeks. These changes paralleled the changes of the expression of ornithine decarboxylase and cyclin D1, two known mediators of kidney growth, whereas, unexpectedly, mTORC1 and Hippo pathways did not seem to be involved. Androgens also enhanced kidney autophagy, very likely by increasing transcription factor EB nuclear translocation. Functionally, the increase of tubular mass resulted in increased sodium/phosphate transport. These findings were relevant to humans. Remarkably, by studying living gender-paired kidney donors-recipients, we showed that tubular cell size increased three months after transplantation in men as compared to women, regardless of the donor gender. Thus, our results identify novel signaling pathways that may be involved in androgen-induced kidney growth and homeostasis and suggest that androgens determine kidney size after transplantation.

MiR27a, a fine-tuning molecule, interacts with growth hormone (GH) signaling and ornithine decarboxylase (ODC) via targeting STAT5.

Autocrine growth hormone (GH) expression triggers cell proliferation, invasion-metastasis in vitro and in vivo models, but GH gene mutations inhibit postnatal growth. Natural polyamines (PA); putrescine, spermidine, spermine trigger cell growth and differentiation. The importance of miR27a has shown to exert a suppressive effect on ornithine decarboxylase (ODC) expression in dwarf mice models. We aimed to modulate the role of A13S, F166Δ, T24 GH gene mutations' impact on PA metabolism and epithelial-mesencyhmal transition (EMT) pathway through miR27a. Biologically active GH signaling triggered cell viability, growth, and colony formation, but T24A alteration significantly decreases aggressive profiles due to inactive GH signaling through a decline in STAT5 activity and expressions of STAT5, c-myc and ODC. Although statistically significant increase in intracellular PA levels in wt GH signaling HEK293 cells compared to HEK293 cells with a lack of GH signaling, a sharp decline in PA levels measured in each mutant GH expressing HEK293 cells. When we inhibited miR27a, proliferation and colony formation accelerated through a significant increase in putrescine levels and upregulation of ODC, STAT5 expression. In contrast, a substantial decline in GH-mediated colony enlargement observed via ODC, STAT5 downregulation, and PA depletion in both wt and mutant GH expressing HEK293 cell lines by miR27a mimic transfection. In conclusion, T24A mutant GH expression declines the GH signaling through STAT5 activity, and mutant GH signaling decreased cell proliferation, division, and colony formation via EMT inhibition. The autocrine GH-mediated proliferative profiles were under the control of miR27a that depletes intracellular putrescine levels via targeting ODC.

Genome wide survey, and expression analysis of Ornithine decarboxylase gene associated with alkaloid biosynthesis in plants

Plant ODC (ornithine decarboxylase) plays a vital role in normalizing cell division in actively growing tissues. The ODC is a key precursor enzyme for nicotine and nornicotine biosynthesis in plants. ODCs are widely present in many plant families but have not been functionally validated and characterized at the molecular level. In the present study, 58 plant ODCs were identified and were found to contain two putative regulatory motifs, specifically PLP (Pyridoxal 5′-phosphate) and Orn/DAP/Arg decarboxylase family 2 pyridoxal-phosphate, that are highly conserved among diverse plant species. Further, the cis-regulatory elements and interacting partners of the gene revealed the importance of ODC in various metabolic pathways. The qRT-PCR revealed highest relative expression of ODC in floral meristem and roots. Our results suggest that ODC can be effectively used as an ideal candidate for engineering polyamine biosynthesis and would be crucial for developing ultra-low nicotine content tobacco lines via genome editing.