Reduced symmetric dimethylation stabilizes vimentin and promotes metastasis in MTAP-deficient lung cancer.

The aggressive nature and poor prognosis of lung cancer led us to explore the mechanisms driving disease progression. Utilizing our invasive cell-based model, we identified methylthioadenosine phosphorylase (MTAP) and confirmed its suppressive effects on tumorigenesis and metastasis. Patients with low MTAP expression display worse overall and progression-free survival. Mechanistically, accumulation of methylthioadenosine substrate in MTAP-deficient cells reduce the level of protein arginine methyltransferase 5 (PRMT5)-mediated symmetric dimethylarginine (sDMA) modification on proteins. We identify vimentin as a dimethyl-protein whose dimethylation levels drop in response to MTAP deficiency. The sDMA modification on vimentin reduces its protein abundance but trivially affects its filamentous structure. In MTAP-deficient cells, lower sDMA modification prevents ubiquitination-mediated vimentin degradation, thereby stabilizing vimentin and contributing to cell invasion. MTAP and PRMT5 negatively correlate with vimentin in lung cancer samples. Taken together, we propose a mechanism for metastasis involving vimentin post-translational regulation.

Altered expression of the L-arginine/nitric oxide pathway in ovarian cancer: metabolic biomarkers and biological implications.

MOTIVATION: Ovarian cancer (OC) is a highly lethal gynecological malignancy. Extensive research has shown that OC cells undergo significant metabolic alterations during tumorigenesis. In this study, we aim to leverage these metabolic changes as potential biomarkers for assessing ovarian cancer. METHODS: A functional module-based approach was utilized to identify key gene expression pathways that distinguish different stages of ovarian cancer (OC) within a tissue biopsy cohort. This cohort consisted of control samples (n = 79), stage I/II samples (n = 280), and stage III/IV samples (n = 1016). To further explore these altered molecular pathways, minimal spanning tree (MST) analysis was applied, leading to the formulation of metabolic biomarker hypotheses for OC liquid biopsy. To validate, a multiple reaction monitoring (MRM) based quantitative LCMS/MS method was developed. This method allowed for the precise quantification of targeted metabolite biomarkers using an OC blood cohort comprising control samples (n = 464), benign samples (n = 3), and OC samples (n = 13). RESULTS: Eleven functional modules were identified as significant differentiators (false discovery rate, FDR < 0.05) between normal and early-stage, or early-stage and late-stage ovarian cancer (OC) tumor tissues. MST analysis revealed that the metabolic L-arginine/nitric oxide (L-ARG/NO) pathway was reprogrammed, and the modules related to "DNA replication" and "DNA repair and recombination" served as anchor modules connecting the other nine modules. Based on this analysis, symmetric dimethylarginine (SDMA) and arginine were proposed as potential liquid biopsy biomarkers for OC assessment. Our quantitative LCMS/MS analysis on our OC blood cohort provided direct evidence supporting the use of the SDMA-to-arginine ratio as a liquid biopsy panel to distinguish between normal and OC samples, with an area under the ROC curve (AUC) of 98.3%. CONCLUSION: Our comprehensive analysis of tissue genomics and blood quantitative LC/MSMS metabolic data shed light on the metabolic reprogramming underlying OC pathophysiology. These findings offer new insights into the potential diagnostic utility of the SDMA-to-arginine ratio for OC assessment. Further validation studies using adequately powered OC cohorts are warranted to fully establish the clinical effectiveness of this diagnostic test.

Urinary excretion of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine is positively related to nitric oxide level in tissues of normotensive and hypertensive rats.

Nitric oxide (NO) is one of the gaseous transmitters which play a very important role in the regulation of the circulatory system. Decreased NO availability is associated with hypertension, cardiovascular and kidney diseases. Endogenous NO is generated enzymatically by nitric oxide synthase (NOS) depending on the availability of the substrate, cofactors, or presence/absence of inhibitors, such as asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA). The objective of this study was to evaluate the potential relationship between NO level in rat tissues (heart and kidneys) and the concentrations of endogenous metabolites related to NO in plasma and urine. The experiment was carried out with 16- and 60-week-old male Wistar Kyoto (WKY) and age-matched male Spontaneously Hypertensive Rats (SHR). NO level in tissue homogenates was determined by the colorimetric method. RT-qPCR was used to verify the expression of the eNOS (endothelial NOS) gene. Plasma and urine concentrations of arginine, ornithine, citrulline, and dimethylarginines were examined by the UPLC-MS/MS method. 16-week-old WKY rats had the highest tissue NO and plasma citrulline levels. Furthermore, 16-week-old WKY rats showed higher urinary excretion of ADMA/SDMA compared to other experimental groups, however, plasma concentrations of arginine, ADMA, and SDMA were comparable between the groups. In conclusion, our research shows that hypertension and aging decrease tissue NO levels and are associated with reduced urinary excretion of NOS inhibitors, i.e., ADMA and SDMA.

Association of Uremic Solutes With Cardiovascular Death in Diabetic Kidney Disease.

RATIONALE & OBJECTIVE: Cardiovascular disease (CVD) is a major cause of mortality among people with diabetic kidney disease (DKD). The pathophysiology is inadequately explained by traditional CVD risk factors. The uremic solutes trimethylamine-N-oxide (TMAO) and asymmetric and symmetric dimethylarginine (ADMA, SDMA) have been linked to CVD in kidney failure with replacement therapy (KFRT), but data are limited in populations with diabetes and less severe kidney disease. STUDY DESIGN: Observational cohort. SETTINGS & PARTICIPANTS: Random subcohort of 555 REGARDS (Reasons for Geographic and Racial Differences in Stroke) study participants with diabetes and estimated glomerular filtration rate (eGFR) <60 mL/min/1.73 m2 at study entry. EXPOSURE: ADMA, SDMA, and TMAO assayed by liquid chromatography-mass spectrometry in plasma and urine. OUTCOME: Cardiovascular mortality (primary outcome); all-cause mortality and incident KFRT (secondary outcomes). ANALYTICAL APPROACH: Plasma concentrations and ratios of urine to plasma concentrations of ADMA, SDMA, and TMAO were tested for association with outcomes. Adjusted Cox regression models were fitted and hazard ratios of outcomes calculated per standard deviation and per doubling, and as interquartile comparisons. RESULTS: The mean baseline eGFR was 44 mL/min/1.73 m2. Cardiovascular death, overall mortality, and KFRT occurred in 120, 285, and 89 participants, respectively, during a mean 6.2 years of follow-up. Higher plasma ADMA and SDMA (HRs of 1.20 and 1.28 per 1-SD greater concentration), and lower ratios of urine to plasma concentrations of ADMA, SDMA, and TMAO (HRs per halving of 1.53, 1.69, and 1.38) were associated with cardiovascular mortality. Higher plasma concentrations of ADMA, SDMA, and TMAO (HRs of 1.31, 1.42, and 1.13 per 1-SD greater concentration) and lower urine to plasma ratios of ADMA, SDMA, and TMAO (HRs per halving of 1.34, 1.37, and 1.26) were associated with all-cause mortality. Higher plasma ADMA and SDMA were associated with incident KFRT by categorical comparisons (HRs of 2.75 and 2.96, comparing quartile 4 to quartile 1), but not in continuous analyses. LIMITATIONS: Single cohort, restricted to patients with diabetes and eGFR < 60 mL/min/1.73 m2, potential residual confounding by GFR, no dietary information. CONCLUSIONS: Higher plasma concentrations and lower ratios of urine to plasma concentrations of uremic solutes were independently associated with cardiovascular and all-cause mortality in DKD. Associations of ratios of urine to plasma concentrations with mortality suggest a connection between renal uremic solute clearance and CVD pathogenesis.

Assessment of potential cardiovascular risk in trichloroethylene exposure by serum methylated arginine levels.

Trichloroethylene (TCE) is a widely used solvent in industrial applications and has toxic effects on various systems. Methylated arginine amino acids (eg asymmetric dimethyl arginine (ADMA), symmetric dimethyl arginine (SDMA)) cause the development of cardiovascular disease by inhibiting NO synthesis, which is considered to be heart-protector. The aim of this study is to determine the risk of cardiovascular diseases in TCE exposure by methylated arginine biomarkers. About 98 controls and 100 TCE-exposed male subjects were included in the study. Trichloroacetic acid (urinary metabolite of TCE), arginine, homoarginine, citrulline ADMA, SDMA, and N-monomethyl L-arginine (L-NMMA) levels were found significantly higher than control group (p < 0.001). The strongest correlation was found between ADMA and Trichloroacetic acid (TCA) level (r = 0.453, p < 0.01). Long-term TCE exposure, may be an important risk factor for cardiovascular diseases by increasing methylated arginine levels.

Esophageal Squamous Cell Carcinoma Is Accompanied by Local and Systemic Changes in L-arginine/NO Pathway.

The L-arginine/NO pathway holds promise as a source of potential therapy target and biomarker; yet, its status and utility in esophageal squamous cell carcinoma (ESCC) is unclear. We aimed at quantifying pathway metabolites in sera from patients with ESCC (n = 61) and benign conditions (n = 62) using LC-QTOF-MS and enzyme expression in esophageal tumors and matched noncancerous samples (n = 40) using real-time PCR with reference to ESCC pathology and circulating immune/inflammatory mediators, quantified using Luminex xMAP technology. ESCC was associated with elevated systemic arginine and asymmetric dimethylarginine. Citrulline decreased and arginine bioavailability increased along with increasing ESCC advancement. Compared to adjacent tissue, tumors overexpressed ODC1, NOS2, PRMT1, and PRMT5 but had downregulated ARG1, ARG2, and DDAH1. Except for markedly higher NOS2 and lower ODC1 in tumors from M1 patients, the pathology-associated changes in enzyme expression were subtle and present also in noncancerous tissue. Both the local enzyme expression level and systemic metabolite concentration were related to circulating inflammatory and immune mediators, particularly those associated with eosinophils and those promoting viability and self-renewal of cancer stem cells. Metabolic reprogramming in ESCC manifests itself by the altered L-arginine/NO pathway. Upregulation of PRMTs in addition to NOS2 and ODC1 and the pathway link with stemness-promoting cytokines warrants further investigation.

Transcriptional and Metabolomic Analysis of L-Arginine/Nitric Oxide Pathway in Inflammatory Bowel Disease and Its Association with Local Inflammatory and Angiogenic Response: Preliminary Findings.

L-arginine/nitric oxide pathway in Crohn's disease (CD) and ulcerative colitis (UC) is poorly investigated. The aim of current study is to quantify pathway serum metabolites in 52 CD (40 active), 48 UC (33 active), and 18 irritable bowel syndrome patients and 40 controls using mass spectrometry and at determining mRNA expression of pathway-associated enzymes in 91 bowel samples. Arginine and symmetric dimethylarginine decreased (p < 0.05) in active-CD (129 and 0.437 µM) compared to controls (157 and 0.494 µM) and active-UC (164 and 0.52 µM). Citrulline and dimethylamine increased (p < 0.05) in active-CD (68.7 and 70.9 µM) and active-UC (65.9 and 73.9 µM) compared to controls (42.7 and 50.4 µM). Compared to normal, CD-inflamed small bowel had downregulated (p < 0.05) arginase-2 by 2.4-fold and upregulated dimethylarginine dimethylaminohydrolase (DDAH)-2 (1.5-fold) and arginine N-methyltransferase (PRMT)-2 (1.6-fold). Quiescent-CD small bowel had upregulated (p < 0.05) arginase-2 (1.8-fold), DDAH1 (2.9-fold), DDAH2 (1.5-fold), PRMT1 (1.5-fold), PRMT2 (1.7-fold), and PRMT5 (1.4-fold). Pathway enzymes were upregulated in CD-inflamed/quiescent and UC-inflamed colon as compared to normal. Compared to inflamed, quiescent CD-colon had upregulated DDAH1 (5.7-fold) and ornithine decarboxylase (1.6-fold). Concluding, the pathway is deregulated in CD and UC, also in quiescent bowel, reflecting inflammation severity and angiogenic potential. Functional analysis of PRMTs and DDAHs as potential targets for therapy is warranted.

The Second Life of Methylarginines as Cardiovascular Targets.

Endogenous methylarginines were proposed as cardiovascular risk factors more than two decades ago, however, so far, this knowledge has not led to the development of novel therapeutic approaches. The initial studies were primarily focused on the endogenous inhibitors of nitric oxide synthases asymmetric dimethylarginine (ADMA) and monomethylarginine (MMA) and the main enzyme regulating their clearance dimethylarginine dimethylaminohydrolase 1 (DDAH1). To date, all the screens for DDAH1 activators performed with the purified recombinant DDAH1 enzyme have not yielded any promising hits, which is probably the main reason why interest towards this research field has started to fade. The relative contribution of the second DDAH isoenzyme DDAH2 towards ADMA and MMA clearance is still a matter of controversy. ADMA, MMA and symmetric dimethylarginine (SDMA) are also metabolized by alanine: glyoxylate aminotransferase 2 (AGXT2), however, in addition to methylarginines, this enzyme also has several cardiovascular protective substrates, so the net effect of possible therapeutic targeting of AGXT2 is currently unclear. Recent studies on regulation and functions of the enzymes metabolizing methylarginines have given a second life to this research direction. Our review discusses the latest discoveries and controversies in the field and proposes novel directions for targeting methylarginines in clinical settings.

RETROSPECTIVE ANALYSIS AND VALIDATION OF SERUM SYMMETRIC DIMETHYLARGININE (SDMA) CONCENTRATIONS IN CHEETAHS ( ACINONYX JUBATUS).

Kidney disease is common among captive cheetahs ( Acinonyx jubatus). Serum creatinine is the most common measurement to estimate glomerular filtration rate (GFR) because of the ease of its clinical use, but it is a crude estimate that only increases after significant disease is already present and is affected by extrarenal factors. Symmetric dimethylarginine (SDMA) is a renal biomarker in humans, dogs, and cats that correlates with serum creatinine and GFR and appears to be an earlier and more specific biomarker for kidney disease. Ninety-two banked serum samples from 11 cheetahs housed at the Oklahoma City Zoo from 1992 to 2012 were retrospectively analyzed. Histopathology results were available for 10/11 cheetahs, and all 10 had histologic renal lesions. General categories of renal lesions included glomerulosclerosis (7/10; 70%), amyloidosis (7/10; 70%), inflammatory (9/10; 90%), and oxalate nephrosis (2/10; 20%). SDMA immunoassay and mass spectrometry were measured for validation and compared with creatinine to assess for correlation. Serum creatinine concentrations were determined by enzymatic colorimetric methods. SDMA immunoassay was validated in cheetahs and correlated well with serum creatinine ( R2=0.687; P < 0.0001). SDMA and serum creatinine measured from freeze-thawed stored samples show high correlation in individual cheetahs ( R2 = 0.972; P < 0.0001). These data support that SDMA could be a promising renal biomarker in cheetahs. Further research is warranted to investigate whether SDMA might be an earlier indicator of kidney disease in cheetahs and whether this assay can be extended to other nondomestic carnivores.

Serum Asymmetric and Symmetric Dimethylarginine and Morbidity and Mortality in Hemodialysis Patients.

BACKGROUND: Asymmetric (ADMA) and symmetric dimethylarginine (SDMA) are putative uremic toxins that may exert toxicity by a number of mechanisms, including impaired nitric oxide synthesis and generation of reactive oxygen species. The study goal was to determine the association between these metabolites and cardiovascular outcomes in hemodialysis patients. STUDY DESIGN: Post hoc analysis of the Hemodialysis (HEMO) Study. SETTING & PARTICIPANTS: 1,276 prevalent hemodialysis patients with available samples 3 to 6 months after randomization. PREDICTOR: ADMA and SDMA measured in stored specimens. OUTCOMES: Cardiac death, sudden cardiac death, first cardiovascular event, and any-cause death. Association with predictors analyzed using Cox regression adjusted for potential confounders (including demographics, clinical characteristics, comorbid conditions, albumin level, and residual kidney function). RESULTS: Mean age of patients was 57±14 (SD) years, 63% were black, and 57% were women. Mean ADMA (0.9±0.2µmol/L) and SDMA levels (4.3±1.4µmol/L) were moderately correlated (r=0.418). Higher dialysis dose or longer session length were not associated with lower predialysis ADMA or SDMA concentrations. In fully adjusted models, each doubling of ADMA level was associated with higher risk (HR per 2-fold higher concentration; 95% CI) of cardiac death (1.83; 1.29-2.58), sudden cardiac death (1.79; 1.19-2.69), first cardiovascular event (1.50; 1.20-1.87), and any-cause death (1.44; 1.13-1.83). Compared to the lowest ADMA quintile (<0.745 µmol/L), the highest ADMA quintile (≥1.07µmol/L) was associated with higher risk (HR; 95% CI) of cardiac death (2.10; 1.44-3.05), sudden cardiac death (2.06; 1.46-2.90), first cardiovascular event (1.75; 1.35-2.27), and any-cause death (1.56; 1.21-2.00). SDMA level was associated with higher risk for cardiac death (HR, 1.40; 95% CI, 1.03-1.92), but this was no longer statistically significant after adjusting for ADMA level (HR, 1.20; 95% CI, 0.86-1.68). LIMITATIONS: Single time-point measurement of ADMA and SDMA. CONCLUSIONS: ADMA and, to a lesser extent, SDMA levels are associated with cardiovascular outcomes in hemodialysis patients.

Admission levels of asymmetric and symmetric dimethylarginine predict long-term outcome in patients with community-acquired pneumonia.

BACKGROUND: During infection, there is an activation of the L-arginine-nitric-oxide pathway, with a shift from nitric oxide synthesis to a degradation of L-arginine to its metabolites, asymmetric and symmetric dimethylarginine (ADMA and SDMA). However, the prognostic implications for short-term or long-term survival remains unclear. We investigated the association of L-arginine, ADMA, and SDMA with adverse clinical outcomes in a well-defined cohort of patients with community-acquired pneumonia (CAP). METHODS: We measured L-arginine, ADMA, and SDMA in 268 CAP patients from a Swiss multicenter trial by mass spectrometry and used Cox regression models to investigate associations between blood marker levels and disease severity as well as mortality over a period of 6 years. RESULTS: Six-year mortality was 44.8%. Admission levels of ADMA and SDMA (µmol/L) were correlated with CAP severity as assessed by the pneumonia severity index (r = 0.32, p < 0.001 and r = 0.56, p < 0.001 for ADMA and SDMA, respectively) and higher in 6-year non-survivors versus survivors (median 0.62 vs. 0.48; p < 0.001 and 1.01 vs. 0.85; p < 0.001 for ADMA and SDMA, respectively). Both ADMA and SDMA were significantly associated with long-term mortality (hazard ratios [HR] 4.44 [95% confidence intervals (CI) 1.84 to 10.74] and 2.81 [95% CI 1.45 to 5.48], respectively). The effects were no longer significant after multivariate adjustment for age and comorbidities. No association of L-arginine with severity and outcome was found. CONCLUSIONS: Both ADMA and SDMA show a severity-dependent increase in patients with CAP and are strongly associated with mortality. This association is mainly explained by age and comorbidities. TRIAL REGISTRATION: ISRCTN95122877 . Registered 31 July 2006.

Evaluation of various biomarkers for kidney monitoring during canine leishmaniosis treatment.

BACKGROUND: The objective of this study was to evaluate and compare the evolution of the profile currently recommended by the International Renal Interest Society (IRIS) (sCr, UPC and sSDMA) with a panel of other different kidney biomarkers during treatment for canine leishmaniosis. This panel included three urinary glomerular biomarkers (uIgG, uCRP and uferritin) and three urinary tubular biomarkers (uGGT, uNAG and uRBP). These biomarkers were measured in two groups of dogs with canine leishmaniosis at IRIS stage I. Group 1: dogs showing proteinuria (UPC > 0.5) before treatment which did not decrease after treatment; Group 2: dogs showing proteinuria before treatment which decreased after treatment. RESULTS: Group 1 showed no significant changes in any biomarker after treatment. In group 2, among the biomarkers recommended by the IRIS, only UPC showed a significant decrease after treatment. However all biomarkers of glomerular damage showed a significant decrease after treatment, with uIgG/Cr and uCRP/Cr showing the greater decreases. In addition uRBP/Cr and uNAG/Cr showed significant decreases after treatment. CONCLUSIONS: In dogs with leishmaniosis at IRIS stage I that reduced UPC after treatment, there were no significant changes in serum creatinine and sSDMA. However, all the urine biomarkers evaluated with exception of uGGT showed a significant decrease. These decreases were more evident in those markers related with glomerular function, being uIgG/Cr the biomarker more associated with UPC. Further studies involving a larger number of animals and histological analysis of the kidney would be recommended to confirm these findings and evaluate the routine practical use of these urine biomarkers in canine leishmaniosis.

Asymmetric Dimethylarginine Predicts Long-Term Outcome in Patients with Acute Exacerbation of Chronic Obstructive Pulmonary Disease.

INTRODUCTION: In chronic obstructive pulmonary disease (COPD), there is an activation of the L-arginine nitric oxide pathway. Pulmonary obstruction causes to elevated nitric oxide (NO) levels, which lead to higher production of the NO-inhibiting metabolites asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA). METHODS: We investigated the association of L-arginine, ADMA, and SDMA with clinical outcomes in a well-defined observational cohort of 150 patients with acute exacerbation of COPD. We measured L-arginine, ADMA, and SDMA by mass spectrometry in patients with pneumonic or non-pneumonic exacerbation of COPD included in a Swiss multicenter trial. We used Cox regression models to investigate the associations between blood marker levels and disease severity as well as all-cause mortality over a follow-up of 6.1 years. RESULTS: Six-year all-cause mortality was 54%. Admission levels of ADMA and SDMA (µmol L-1) were increased in 6-year non-survivors compared to survivors' median (0.60 vs. 0.46, p = 0.004; and 1.05 vs. 0.85, p = 0.012). In a multivariate Cox regression analysis, ADMA was associated with long-term mortality resulting in an age- and comorbidity-adjusted hazard ratio (HR) of 4.55 (95% confidence interval 1.02-20.43, p = 0.048). SDMA was only associated in univariate models and no association of L-arginine with outcome was found. CONCLUSION: ADMA was found to be an independent risk factor for long-term all-cause mortality in patients with acute exacerbation of COPD. Whether therapeutic modification of the L-arginine-nitric oxide pathway has the potential to improve outcome should be evaluated in future interventional trials.

Ammonia Reduces Intracellular Asymmetric Dimethylarginine in Cultured Astrocytes Stimulating Its y⁺LAT2 Carrier-Mediated Loss.

Previously we had shown that ammonia stimulates nitric oxide (NO) synthesis in astrocytes by increasing the uptake of the precursor amino acid, arginine via the heteromeric arginine/glutamine transporter y⁺LAT2. Ammonia also increases the concentration in the brain of the endogenous inhibitor of nitric oxide synthases (NOS), asymmetric dimethylarginine (ADMA), but distribution of ADMA surplus between the intraastrocytic and extracellular compartments of the brain has not been studied. Here we tested the hypothesis that ammonia modulates the distribution of ADMA and its analog symmetric dimethylarginine (SDMA) between the two compartments of the brain by competition with arginine for the y⁺LAT2 transporter. In extension of the hypothesis we analyzed the ADMA/Arg interaction in endothelial cells forming the blood-brain barrier. We measured by high-performance liquid chromatography (HPLC) and mass spectrometry (MS) technique the concentration of arginine, ADMA and SDMA in cultured cortical astrocytes and in a rat brain endothelial cell line (RBE-4) treated with ammonia and the effect of silencing the expression of a gene coding y⁺LAT2. We also tested the expression of ADMA metabolism enzymes: protein arginine methyltransferase (PRMT) and dimethylarginine dimethyl aminohydrolase (DDAH) and arginine uptake to astrocytes. Treatment for 48 h with 5 mM ammonia led to an almost 50% reduction of ADMA and SDMA concentration in both cell types, and the effect in astrocytes was substantially attenuated by silencing of the Slc7a6 gene. Moreover, the y⁺LAT2-dependent component of ammonia-evoked arginine uptake in astrocytes was reduced in the presence of ADMA in the medium. Our results suggest that increased ADMA efflux mediated by upregulated y⁺LAT2 may be a mechanism by which ammonia interferes with intra-astrocytic (and possibly intra-endothelial cell) ADMA content and subsequently, NO synthesis in both cell types.

Computational Study of Symmetric Methylation on Histone Arginine Catalyzed by Protein Arginine Methyltransferase PRMT5 through QM/MM MD and Free Energy Simulations.

Protein arginine methyltransferases (PRMTs) catalyze the transfer of the methyl group from S-adenosyl-l-methionine (AdoMet) to arginine residues. There are three types of PRMTs (I, II and III) that produce different methylation products, including asymmetric dimethylarginine (ADMA), symmetric dimethylarginine (SDMA) and monomethylarginine (MMA). Since these different methylations can lead to different biological consequences, understanding the origin of product specificity of PRMTs is of considerable interest. In this article, the quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) and free energy simulations are performed to study SDMA catalyzed by the Type II PRMT5 on the basis of experimental observation that the dimethylated product is generated through a distributive fashion. The simulations have identified some important interactions and proton transfers during the catalysis. Similar to the cases involving Type I PRMTs, a conserved Glu residue (Glu435) in PRMT5 is suggested to function as general base catalyst based on the result of the simulations. Moreover, our results show that PRMT5 has an energetic preference for the first methylation on Nη1 followed by the second methylation on a different ω-guanidino nitrogen of arginine (Nη2).The first and second methyl transfers are estimated to have free energy barriers of 19-20 and 18-19 kcal/mol respectively. The computer simulations suggest a distinctive catalytic mechanism of symmetric dimethylation that seems to be different from asymmetric dimethylation.

Assessment of cell cycle arrest biomarkers and neutrophil gelatinase-associated lipocalin to distinguish acute kidney injury from other diseases in dogs.

BACKGROUND: Cell cycle arrest biomarkers (tissue inhibitor of metalloproteinase-2 [uTIMP-2] and insulin-like growth factor binding protein 7 [uIGFBP7]), and neutrophil gelatinase-associated lipocalin (NGAL) variables are valuable biomarkers for early diagnosis of acute kidney injury (AKI) in people. OBJECTIVES: To evaluate uTIMP-2, uIGFBP7, fractional excretion of NGAL (FeNGAL), and urinary to serum NGAL ratio (u/sNGAL) in healthy dogs, dogs with AKI, dogs with chronic kidney disease (CKD), and critically ill (CI) dogs. ANIMALS: Forty-two client-owned dogs (healthy, n = 10; AKI, n = 11; CKD, n = 11; CI, n = 10). METHODS: Prospective, observational study. After assessment of routine renal biomarkers, stress (uTIMP-2, uIGFBP7) and damage (NGAL) biomarkers were measured, using ELISA kits, and normalized to urinary creatinine (uCr). RESULTS: Normalized uTIMP-2 and [uTIMP-2] × [uIGFBP7]/uCr were significantly higher in the AKI group (median 151.9 [range, 2.2-534.2] and 62.9 [1.1-266.8] pg/mL respectively), compared to healthy dogs (0.3 [0.2-74.7]; P < .001 and 0.16 [0.1-58.1] pg/mL; P < .001), dogs with CKD (0.7 [0.3-742.5]; P = .04 and 0.37 [0.2-180.1] pg/mL; P = .03) and CI dogs (1.9 [0.2-37.0]; P = .03 and 0.8 [0.1-16.1] pg/mL; P = .02). Fractional excretion of NGAL was significantly higher in dogs with AKI (54.17 [7.93-155.32] %), than in healthy (0.03 [0.01-0.21] %; P < .001) and CI dogs (3.05 [0.05-28.86] %; P = .02). CONCLUSIONS AND CLINICAL IMPORTANCE: Normalized uTIMP-2, [uTIMP-2] × [uIGFBP7]/uCr, and FeNGAL can be valuable renal biomarkers for early diagnosis of AKI in dogs.

Dimethylarginines in acute myocardial infarction: Association with lifestyle, sociodemographic, and somatometric factors.

BACKGROUND: Recent findings associate asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) with the prognosis of acute myocardial infarction (AMI). The purpose of the current study was to associate patients' lifestyle, sociodemographic, and somatometric characteristics with the time course of ADMA and SDMA concentrations in the serum of AMI patients. PATIENTS AND METHODS: In the serum of 66 AMI patients, ADMA, SDMA, troponin T, and C-reactive protein (CRP) were measured upon hospital admission (<24 h) and on the 3rd day following. Lifestyle, sociodemographic, and somatometric characteristics were obtained through a questionnaire, filled on patient discharge. RESULTS: ADMA concentrations on the 1st day positively correlated with daily reported hours of sleep (+0.497, P < 0.001) and delivery or eating out frequency (+0.285, P = 0.02), whereas it negatively correlated with reported physical condition (-0.304, P = 0.013). A personal history of hypertension indicated higher 1st-day ADMA concentration (1.818 vs 1.568, P = 0.042). Age positively correlated with 1st-day SDMA (+0.320, P = 0.009). All of the biomarker concentrations were reduced on the 3rd day measurements (P < 0.001). Self-reported lifetime minimum BMI positively correlated with either absolute (r = +0.366, P = 0.009) or percentage (r = +0.262, P = 0.045) ADMA reduction. A daily sleep in 5-8-h range was inversely correlated with percentage (-0.410, P = 0.001) or absolute (r = -0.369, P = 0.002) SDMA reduction. CONCLUSIONS: Modifiable factors such as BMI, eating habits, physical condition, and sleep seem to affect the baseline levels or time course of ADMA and SDMA in AMI patients. Changes in these factors may affect AMI prognosis by altering dimethylarginine levels.

Renal ultrasonographic strain elastography and symmetric dimethylarginine (SDMA) in canine and feline chronic kidney disease.

Chronic kidney disease (CKD) is a common renal disease in dogs and cats. Renal fibrosis is a main pathologic process leading of CKD progression. Renal biopsy is the gold standard for renal fibrosis assessment. However, it is not routinely performed in clinic due to its invasiveness. Therefore, the aim of this study was to evaluate the use of ultrasonographic strain elastography (SE), which is a non-invasive method for renal tissue stiffness determination and its association with renal function. Renal strain ratios and renal function were evaluated in 13 CKD dogs (CKDD), 38 healthy dogs (HD), 17 CKD cats (CKDC) and 26 healthy cats (HC). There were significantly lower renal cortical strain ratios than medullary strain ratios in all groups (HD; P<0.01, HC; P<0.01, CKDD and CKDC; P<0.05) and significantly lower cortical and medullary strain ratios in both CKDD and CKDC than in healthy control animals of both species (P<0.0001). In dogs, the renal cortical and medullary strain ratios significantly negatively correlated with plasma creatinine (P<0.05), blood urea nitrogen (BUN; P<0.05; P<0.01, respectively), and symmetric dimethylarginine (SDMA; P<0.01). In cats, similar correlations were found for plasma creatinine (P<0.001), BUN (P<0.05; P<0.001, respectively) and SDMA (P<0.05). SE might be a promising imaging diagnostic tool for renal-elasticity evaluation, also correlating with renal functional impairment in canine and feline CKD.

Arginine and Arginine/ADMA Ratio Predict 90-Day Mortality in Patients with Out-of-Hospital Cardiac Arrest-Results from the Prospective, Observational COMMUNICATE Trial.

(1) Background: In patients with shock, the L-arginine nitric oxide pathway is activated, causing an elevation of nitric oxide, asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) levels. Whether these metabolites provide prognostic information in patients after out-of-hospital cardiac arrest (OHCA) remains unclear. (2) Methods: We prospectively included OHCA patients, recorded clinical parameters and measured plasma ADMA, SDMA and Arginine levels by liquid chromatography tandem mass spectrometry (LC-MS). The primary endpoint was 90-day mortality. (3) Results: Of 263 patients, 130 (49.4%) died within 90 days after OHCA. Compared to survivors, non-survivors had significantly higher levels of ADMA and lower Arginine and Arginine/ADMA ratios in univariable regression analyses. Arginine levels and Arginine/ADMA ratio were significantly associated with 90-day mortality (OR 0.51 (95%CI 0.34 to 0.76), p < 0.01 and OR 0.40 (95%CI 0.26 to 0.61), p < 0.001, respectively). These associations remained significant in several multivariable models. Arginine/ADMA ratio had the highest predictive value with an area under the curve (AUC) of 0.67 for 90-day mortality. Results for secondary outcomes were similar with significant associations with in-hospital mortality and neurological outcome. (4) Conclusion: Arginine and Arginine/ADMA ratio were independently associated with 90-day mortality and other adverse outcomes in patients after OHCA. Whether therapeutic modification of the L-arginine-nitric oxide pathway has the potential to improve outcome should be evaluated.

L-Arginine/Nitric Oxide Pathway Is Altered in Colorectal Cancer and Can Be Modulated by Novel Derivatives from Oxicam Class of Non-Steroidal Anti-Inflammatory Drugs.

L-arginine/nitric oxide pathway metabolites are altered in colorectal cancer (CRC). We evaluated underlying changes in pathway enzymes in 55 paired tumor/tumor-adjacent samples and 20 normal mucosa using quantitative-PCR and assessed the impact of classic and novel oxicam analogues on enzyme expression and intracellular metabolite concentration (LC-MS/MS) in Caco-2, HCT116, and HT-29 cells. Compared to normal mucosa, ARG1, PRMT1, and PRMT5 were overexpressed in both tumor and tumor-adjacent tissue and DDAH2 solely in tumor-adjacent tissue. Tumor-adjacent tissue had higher expression of ARG1, DDAH1, and DDAH2 and lower NOS2 than patients-matched tumors. The ARG1 expression in tumors increased along with tumor grade and reflected lymph node involvement. Novel oxicam analogues with arylpiperazine moiety at the thiazine ring were more effective in downregulating DDAHs and PRMTs and upregulating ARG2 than piroxicam and meloxicam. An analogue distinguished by propylene linker between thiazine's and piperazine's nitrogen atoms and containing two fluorine substituents was the strongest inhibitor of DDAHs and PRMTs expression, while an analogue containing propylene linker but no fluorine substituents was the strongest inhibitor of ARG2 expression. Metabolic reprogramming in CRC includes overexpression of DDAHs and PRMTs in addition to ARG1 and NOS2 and is not restricted to tumor tissue but can be modulated by novel oxicam analogues.