Clinical improvement of Long-COVID is associated with reduction in autoantibodies, lipids, and inflammation following therapeutic apheresis.

In the aftermath of the COVID-19 pandemic, we are witnessing an unprecedented wave of post-infectious complications. Most prominently, millions of patients with Long-Covid complain about chronic fatigue and severe post-exertional malaise. Therapeutic apheresis has been suggested as an efficient treatment option for alleviating and mitigating symptoms in this desperate group of patients. However, little is known about the mechanisms and biomarkers correlating with treatment outcomes. Here, we have analyzed in different cohorts of Long-Covid patients specific biomarkers before and after therapeutic apheresis. In patients that reported a significant improvement following two cycles of therapeutic apheresis, there was a significant reduction in neurotransmitter autoantibodies, lipids, and inflammatory markers. Furthermore, we observed a 70% reduction in fibrinogen, and following apheresis, erythrocyte rouleaux formation and fibrin fibers largely disappeared as demonstrated by dark field microscopy. This is the first study demonstrating a pattern of specific biomarkers with clinical symptoms in this patient group. It may therefore form the basis for a more objective monitoring and a clinical score for the treatment of Long-Covid and other postinfectious syndromes.

Differential functionality of fluoropyrimidine nucleosides for safe cancer therapy.

Chemotherapies are standard care for most cancer types. Pyrimidine analogs including 5-fluorouracil, cytosine arabinoside, 5-azacytidine, and gemcitabine are effective drugs that are utilized as part of a number of anticancer regimens. However, their lack of cell-specificity results in severe side effects. Therefore, there is a capacity to improve the efficacy of such therapies, while decreasing unwanted side effects. Here, we report that while 5-fluorocytosine is not chemotherapeutic in itself, incorporated into a ribonucleoside and more importantly into an RNA oligonucleotide, it induces cytotoxic effects on cancer cells in vitro . Interestingly, these effects are rescued by both uridine and thymidine. Similarly, in-vitro 2'-deoxy-5-fluorocytidine inhibits the growth of tumor cells but has the advantage of being less toxic to human primary cells compared with 5-fluorocytidine, suggesting that the deoxyribonucleoside could exhibit less side-effects in vivo . Thus, this work indicates that the potency of 5-fluorocytidine and 2'-deoxy-5-fluorocytidine should be further explored. In particular, oligonucleotides incorporating 5-fluorocytosine could be novel chemotherapeutic drugs that could be formulated in cancer-specific particles for safe and efficacious cancer treatments.

Knock-out of the critical nitric oxide synthase regulator DDAH1 in mice impacts amphetamine sensitivity and dopamine metabolism.

The enzyme dimethylarginine dimethylaminohydrolase 1 (DDAH1) plays a pivotal role in the regulation of nitric oxide levels by degrading the main endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine (ADMA). Growing evidence highlight the potential implication of DDAH/ADMA axis in the etiopathogenesis of several neuropsychiatric and neurological disorders, yet the underlying molecular mechanisms remain elusive. In this study, we sought to investigate the role of DDAH1 in behavioral endophenotypes with neuropsychiatric relevance. To achieve this, a global DDAH1 knock-out (DDAH1-ko) mouse strain was employed. Behavioral testing and brain region-specific neurotransmitter profiling have been conducted to assess the effect of both genotype and sex. DDAH1-ko mice exhibited increased exploratory behavior toward novel objects, altered amphetamine response kinetics and decreased dopamine metabolite 3,4-dihydroxyphenylacetic acid (DOPAC) level in the piriform cortex and striatum. Females of both genotypes showed the most robust amphetamine response. These results support the potential implication of the DDAH/ADMA pathway in central nervous system processes shaping the behavioral outcome. Yet, further experiments are required to complement the picture and define the specific brain-regions and mechanisms involved.

Divergent Dimethylarginine Dimethylaminohydrolase Isoenzyme Expression in the Central Nervous System.

The endogenous methylated derivative of ʟ-arginine, Nω,Nω'-dimethyl-ʟ-arginine (asymmetric dimethylarginine, ADMA), an independent risk factor in many diseases, inhibits the activity of nitric oxide synthases and, consequently, modulates the availability of nitric oxide. While most studies on the biological role of ADMA have focused on endothelial and inducible nitric oxide synthases modulation and its contribution to cardiovascular, metabolic, and renal diseases, a role in regulating neuronal nitric oxide synthases and pathologies of the central nervous system is less understood. The two isoforms of dimethylarginine dimethylaminohydrolase (DDAH), DDAH1 and DDAH2, are thought to be the main enzymes responsible for ADMA catabolism. A current impediment is limited knowledge on specific tissue and cellular distribution of DDAH enzymes within the brain. In this study, we provide a detailed characterization of the regional and cellular distribution of DDAH1 and DDAH2 proteins in the adult murine and human brain. Immunohistochemical analysis showed a wide distribution of DDAH1, mapping to multiple cell types, while DDAH2 was detected in a limited number of brain regions and exclusively in neurons. Our results provide key information for the investigation of the pathophysiological roles of the ADMA/DDAH system in neuropsychiatric diseases and pave the way for the development of novel selective therapeutic approaches.

COVID-19 and Therapeutic Apheresis.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus, is an unprecedented challenge for the global community. The pathogenesis of COVID-19, its complications and long term sequelae (so called Long/Post-COVID) include, in addition to the direct virus-induced tissues injury, multiple secondary processes, such as autoimmune response, impairment of microcirculation, and hyperinflammation. Similar pathological processes, but in the settings of neurological, cardiovascular, rheumatological, nephrological, and dermatological diseases can be successfully treated by powerful methods of Therapeutic Apheresis (TA). We describe here the rationale and the initial attempts of TA treatment in severe cases of acute COVID-19. We next review the evidence for the role of autoimmunity, microcirculatory changes and inflammation in pathogenesis of Long/Post COVID and the rationale for targeting those pathogenic processes by different methods of TA. Finally, we discuss the impact of COVID-19 pandemic on patients, who undergo regular TA treatments due to their underlying chronic conditions, with the specific focus on the patients with inherited lipid diseases being treated at the Dresden University Apheresis Center.

Post COVID and Apheresis - Where are we Standing?

A continual increase in cases of Long/Post COVID constitutes a medical and socioeconomic challenge to health systems around the globe. While the true extent of this problem cannot yet be fully evaluated, recent data suggest that up to 20% of people with confirmed SARS-CoV-2 suffer from clinically relevant symptoms of Long/Post COVID several weeks to months after the acute phase. The clinical presentation is highly variable with the main symptoms being chronic fatigue, dyspnea, and cognitive symptoms. Extracorporeal apheresis has been suggested to alleviate symptoms of Post/COVID. Thus, numerous patients are currently treated with apheresis. However, at present there is no data from randomized controlled trials available to confirm the efficacy. Therefore, physicians rely on the experience of practitioners and centers performing this treatment. Here, we summarize clinical experience on extracorporeal apheresis in patients with Post/COVID from centers across Germany.

SGLT2 Inhibitors and Peripheral Vascular Events: A Review of the Literature.

Fifty articles comprising 18 randomized controlled trials (RCTs), 16 observational studies, and 16 meta-analyses on the safety and effectiveness of sodium-glucose cotransporter 2 inhibitors were evaluated in the current review. Only one-fourth of the cohorts of recent trials had peripheral arterial disease (PAD), whereas this subgroup was at high risk for amputations. Despite a remarkable heterogeneity of RCTs, only 2 trials on canagliflozin suggested excess amputation rates, whereas several observational studies generated conflicting conclusions and remained short on possible explanations. Preliminary evidence from observational research suggested that patients with PAD may even benefit from SGLT-2 inhibitor treatment due to lower observed heart failure hospitalization rates.

Overexpression of dimethylarginine dimethylaminohydrolase 1 protects from angiotensin II-induced cardiac hypertrophy and vascular remodeling.

Cardiovascular complications are the leading cause of death, and elevated levels of asymmetric dimethyarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, are implicated in their pathophysiology. We investigated the role of dimethylarginine dimethylaminohydrolase 1 (DDAH1), an enzyme hydrolyzing ADMA, in prevention of cardiovascular remodeling during hypertension. We hypothesized that the animals overexpressing DDAH1 will be protected from angiotensin II (ANG II)-induced end organ damage. Angiotensin II (ANG II) was infused in two doses: 0.75 and 1.5 mg/kg/day in DDAH1 transgenic mice (DDAH1 TG) and wild-type (WT) littermates for 2 or 4 wk. Echocardiography was performed in the first and fourth weeks of the infusion, systolic blood pressure (SBP) was measured weekly, and cardiac hypertrophy and vascular remodeling was assessed by histology. Increase in SBP after 1 wk of ANG II infusion was not different between the groups, whereas TG mice had lower SBP at later time points. TG mice were protected from cardiovascular remodeling after 2 wk of ANG II infusion in the high dose and after 4 wk in the moderate dose. TG mice had higher left ventricular lumen-to-wall ratio, lower cardiomyocyte cross-sectional area, and less interstitial fibrosis compared with WT controls. In aorta, TG mice had less adventitial fibrosis, lower medial thickness with preserved elastin content, lower counts of inflammatory cells, lower levels of active matrix metalloproteinase-2, and showed better endothelium-dependent relaxation. We demonstrated that overexpression of DDAH1 protects from ANG II-induced cardiovascular remodeling and progression of hypertension by preserving endothelial function and limiting inflammation.NEW & NOTEWORTHY We showed that overexpression of dimethylarginine dimethylaminohydrolase 1 (DDAH1) protects from angiotensin II-induced cardiovascular damage, progression of hypertension, and adverse vascular remodeling in vivo. This protective effect is associated with decreased levels of asymmetric dimethylarginine, preservation of endothelial function, inhibition of cardiovascular inflammation, and lower activity of matrix metalloproteinase-2. Our findings are highly clinically relevant, because they suggest that upregulation of DDAH1 might be a promising therapeutic approach against angiotensin II-induced end organ damage.

Initiation of SGLT2 Inhibitors and the Risk of Lower Extremity Minor and Major Amputation in Patients with Type 2 Diabetes and Peripheral Arterial Disease: A Health Claims Data Analysis.

OBJECTIVE: To assess the association between long term risk of hospitalisation for heart failure (HHF) and lower extremity minor and major amputation (LEA) in patients initiating sodium glucose cotransporter 2 inhibitors (SGLT2i) suffering from type 2 diabetes and peripheral arterial disease (PAD). Outcomes were compared with patients without PAD and evaluated separately for the time periods before and after the official warning of the European Medicines Agency (EMA) in early 2017. METHODS: This study used BARMER German health claims data including all patients suffering from type 2 diabetes initiating SGLT2i therapy between 1 January 2013 and 31 December 2019 with follow up until the end of 2020. New users of glucagon like peptide 1 receptor agonists (GLP1-RAs) were used as active comparators. Inverse probability weighting with truncated stabilised weights was used to adjust for confounding, and five year risks of HHF and LEA were estimated using Cox regression. Periods before and after the EMA warning were analysed separately and stratified by presence of concomitant PAD. RESULTS: In total, 44 284 (13.6% PAD) and 56 878 (16.3% PAD) patients initiated SGLT2i or GLP1-RA, respectively. Before the EMA warning, initiation of SGLT2i was associated with a lower risk of HHF in patients with PAD (hazard ratio, HR, 0.85, 95% confidence interval, CI, 0.73 - 0.99) and a higher risk of LEA in patients without PAD (HR 1.79, 95% CI 1.04 - 2.92). After the EMA warning, the efficacy and safety endpoints were no longer statistically different between groups. CONCLUSION: The results from this large nationwide real world study highlight that PAD patients exhibit generally high amputation risks. This study refutes the idea that the presence of PAD explains the excess LEA risk associated with initiation of SGLT2i. The fact that differentials among study groups diminished after the EMA warning in early 2017 emphasises that regulatory surveillance measures worked in everyday clinical practice.

Is Homoarginine a Protective Cardiovascular Risk Factor?

A series of recent epidemiological studies have implicated the endogenous nonproteinogenic amino acid l-homoarginine as a novel candidate cardiovascular risk factor. The association between homoarginine levels and the risk of adverse cardiovascular outcomes is inverse (ie, high cardiovascular risk is predicted by low rather than high homoarginine levels), which makes it plausible to normalize systemic homoarginine levels via oral supplementation. The emergence of homoarginine as a potentially treatable protective cardiovascular risk factor has generated a wave of hope in the field of cardiovascular prevention. Herein, we review the biochemistry, physiology, and metabolism of homoarginine, summarize the strengths and weaknesses of the epidemiological evidence linking homoarginine to cardiovascular disease and its potential protective cardiovascular effects, and identify priorities for future research needed to define the clinical utility of homoarginine as a prognostic factor and therapeutic target in cardiovascular disease.

The role of alanine glyoxylate transaminase-2 (agxt2) in ß-alanine and carnosine metabolism of healthy mice and humans.

PURPOSE: Chronic ß-alanine supplementation leads to increased levels of muscle histidine-containing dipeptides. However, the majority of ingested ß-alanine is, most likely, degraded by two transaminases: GABA-T and AGXT2. In contrast to GABA-T, the in vivo role of AGXT2 with respect to ß-alanine metabolism is unknown. The purpose of the present work is to investigate if AGXT2 is functionally involved in ß-alanine homeostasis. METHODS: Muscle histidine-containing dipeptides levels were determined in AGXT2 overexpressing or knock-out mice and in human subjects with different rs37369 genotypes which is known to affect AGXT2 activity. Further, plasma ß-alanine kinetic was measured and urine was obtained from subjects with different rs37369 genotypes following ingestion of 1400 mg ß-alanine. RESULT: Overexpression of AGXT2 decreased circulating and muscle histidine-containing dipeptides (> 70% decrease; p < 0.05), while AGXT2 KO did not result in altered histidine-containing dipeptides levels. In both models, ß-alanine remained unaffected in the circulation and in muscle (p > 0.05). In humans, the results support the evidence that decreased AGXT2 activity is not associated with altered histidine-containing dipeptides levels (p > 0.05). Additionally, following an acute dose of ß-alanine, no differences in pharmacokinetic response were measured between subjects with different rs37369 genotypes (p > 0.05). Interestingly, urinary ß-alanine excretion was 103% higher in subjects associated with lower AGXT2 activity, compared to subjects associated with normal AGXT2 activity (p < 0.05). CONCLUSION: The data suggest that in vivo, ß-alanine is a substrate of AGXT2; however, its importance in the metabolism of ß-alanine and histidine-containing dipeptides seems small.

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.

Acetylation of asymmetric and symmetric dimethylarginine: an undercharacterized pathway of metabolism of endogenous methylarginines.

BACKGROUND: Increased levels of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) are associated with cardiovascular and renal diseases. We and others have shown that both ADMA and SDMA can be Nα-acetylated to form asymmetric and symmetric Nα-acetyldimethylarginine (Ac-ADMA and Ac-SDMA). The current study further investigated this undercharacterized metabolic pathway. METHODS: ADMA and SDMA were infused in C57/BL6 mice for 3 days using osmotic minipumps. Half of the mice underwent bilateral nephrectomy 24 h before completion of the infusion. Plasma and tissue levels of Ac-ADMA and Ac-SDMA were detected by liquid chromatography-tandem mass spectrometry. RESULTS: ADMA and SDMA infusion resulted in a 3.6-fold increase in plasma Ac-ADMA and a 21-fold increase in plasma Ac-SDMA levels, respectively. Plasma Ac-ADMA and Ac-SDMA levels were dramatically increased after bilateral nephrectomy. The highest baseline tissue concentrations of Ac-ADMA and Ac-SDMA in wild-type mice were detected in the liver, kidney, small intestine, pancreas and spleen. Incubation of the tissue lysates with ADMA and SDMA resulted in increased levels of the corresponding Nα-acetylated products only in the liver, kidney and small intestine. CONCLUSIONS: Our results show that overload of ADMA or SDMA leads to an increase in plasma Ac-ADMA and Ac-SDMA levels. This observation is consistent with the hypothesis that Ac-ADMA and Ac-SDMA are formed directly from ADMA and SDMA in vivo. The increase in plasma Ac-ADMA and Ac-SDMA concentrations after bilateral nephrectomy suggests that both compounds are predominantly eliminated via the kidneys. We demonstrated that acetylation of ADMA and SDMA occurs primarily in the liver, kidney and small intestine.

The SGLT2 inhibitor empagliflozin ameliorates early features of diabetic nephropathy in BTBR ob/ob type 2 diabetic mice with and without hypertension.

Diabetic nephropathy is the leading cause of end-stage renal disease in humans in the Western world. The recent development of Na+-glucose cotransporter 2 (SGLT2) inhibitors offers a new antidiabetic therapy via enhanced glucose excretion. Whether this strategy exerts beneficial effects on the development of type 2 diabetic nephropathy is still largely unclear. We investigated the effects of the specific SGLT2 inhibitor empagliflozin in BTBR.Cg-Lep/WiscJ (BTBR ob/ob) mice, which spontaneously develop type 2 diabetic nephropathy. In the first experiment, BTBR ob/ob mice received either a diet containing 300 ppm empagliflozin or equicaloric placebo chow for 12 wk. In the second experiment, BTBR ob/ob mice received 1 µg·kg body wt(-1)·day(-1) ANG II to induce arterial hypertension and were separated into the same two diet groups for 6 wk. In both experiments, empagliflozin treatment enhanced glucosuria, thereby lowering blood glucose. Independently of hypertension, empagliflozin reduced albuminuria in diabetic mice. However, empagliflozin treatment affected diabetes-related glomerular hypertrophy, markers of renal inflammation, and mesangial matrix expansion only in BTBR ob/ob mice without hypertension. In summary, empagliflozin demonstrated significant antihyperglycemic effects, differentially ameliorating early features of diabetic nephropathy in BTBR ob/ob mice with and without hypertension.

Role of alanine:glyoxylate aminotransferase 2 in metabolism of asymmetric dimethylarginine in the settings of asymmetric dimethylarginine overload and bilateral nephrectomy.

BACKGROUND: Asymmetric and symmetric dimethylarginines (ADMA and SDMA) predict complications and mortality in cardiovascular and renal diseases. Alanine:glyoxylate aminotransferase 2 (AGXT2) can metabolize both ADMA and SDMA; however, this metabolic pathway is still poorly understood. The goal of our study was to test the hypothesis that AGXT2 is compensatory upregulated in the settings of ADMA overload and bilateral nephrectomy. METHODS: ADMA was infused for 3 days using osmotic minipumps in mice. Half of the mice underwent bilateral nephrectomy 24 h before the end of the infusion. RESULTS: Infusion of ADMA caused a 3- to 4-fold increase in plasma and urine ADMA levels and a 2- to 3-fold increase in plasma and urine levels of the ADMA-specific metabolite of AGXT2 α-keto-δ-(N,N-dimethylguanidino)valeric acid (DMGV). Bilateral nephrectomy led to an ∼4-fold increase of plasma SDMA levels, but did not change plasma ADMA levels. Interestingly, plasma levels of DMGV were elevated 32-fold in the mice, which underwent bilateral nephrectomy. Neither bilateral nephrectomy nor ADMA infusion caused upregulation of AGXT2 expression or activity. CONCLUSIONS: Our data demonstrate that short-term elevation of systemic levels of ADMA leads to a dramatic increase of DMGV formation without upregulation of AGXT2 expression or activity, which suggests that AGXT2-mediated pathway of ADMA metabolism is not saturated under normal conditions and may play a major role in the maintenance of ADMA homeostasis in the setting of local or systemic elevation of ADMA levels.

Design and Synthesis of Circular RNA Expression Vectors.

The recent success of the synthetic mRNA-based anti-COVID-19 vaccines has demonstrated the broad potential of the mRNA platform for applications in medicine, thanks to the combined efforts of a small community that has vastly improved key determinants such as design and formulation of synthetic mRNA during the past three decades. However, the cost of production and sensitivity to enzymatic degradation are still limiting the broader application of synthetic mRNA for therapeutic applications. The increased interest in mRNA-based technologies has spurred a renaissance for circular RNA (circRNA), as the lack of free 5' and 3' ends substantially increases resistance against enzymatic degradation in biological systems and does not require expensive cap analogs, as translation is controlled by an Internal Ribosome Entry Site (IRES) sequence. Thus, it can be expected that circRNA will play an important role for future mRNA therapeutics. Here we provide a detailed guide to the production of synthetic circRNA.

A Basic Method for Formulating mRNA-Lipid Nanoparticle Vaccines in the Lab.

During recent years, RNA therapeutics have begun to make a substantial impact in the clinic, with the approval of the siRNA-based therapeutic Patisiran in 2018, and of the two mRNA SARS-CoV-2 vaccines, BNT162b2 and mRNA-1273 in 2021. A key to the success of these therapeutics lies in the lipid-based delivery system. The therapeutic RNAs are encapsulated in lipid nanoparticles (LNPs), which protect against enzymatic degradation and efficiently deliver the RNA across the cell membrane into the cytosol. Thereby, the method used for LNP synthesis and its lipid composition are crucial aspects that decide the efficacy of the LNP-RNA hetero system. Here we provide a detailed guide for the simple preparation of LNP-encapsulated mRNA vaccines.

Absence of Type I Interferon Autoantibodies or Significant Interferon Signature Alterations in Adults With Post-COVID-19 Syndrome.

Genetic defects in the interferon (IFN) system or neutralizing autoantibodies against type I IFNs contribute to severe COVID-19. Such autoantibodies were proposed to affect post-COVID-19 syndrome (PCS), possibly causing persistent fatigue for >12 weeks after confirmed SARS-CoV-2 infection. In the current study, we investigated 128 patients with PCS, 21 survivors of severe COVID-19, and 38 individuals who were asymptomatic. We checked for autoantibodies against IFN-α, IFN-ß, and IFN-ω. Few patients with PCS had autoantibodies against IFNs but with no neutralizing activity, indicating a limited role of type I IFNs in PCS pathogenesis. In a subset consisting of 28 patients with PCS, we evaluated IFN-stimulated gene activity and showed that it did not correlate with fatigue. In conclusion, impairment of the type I IFN system is unlikely responsible for adult PCS.

In vivo evidence that Agxt2 can regulate plasma levels of dimethylarginines in mice.

Elevated plasma concentrations of the asymmetric (ADMA) and symmetric (SDMA) dimethylarginine have repeatedly been linked to adverse cardiovascular clinical outcomes. Both dimethylarginines can be degraded by alanine-glyoxylate aminotransferase 2 (Agxt2), which is also the key enzyme responsible for the degradation of endogenously formed ß-aminoisobutyrate (BAIB). In the present study we wanted to investigate the effect of BAIB on Agxt2 expression and Agxt2-mediated metabolism of dimethylarginines. We infused BAIB or saline intraperitoneally for 7days in C57/BL6 mice via minipumps. Expression of Agxt2 was determined in liver and kidney. The concentrations of BAIB, dimethylarginines and the Agxt2-specific ADMA metabolite α-keto-δ-(N(G),N(G)-dimethylguanidino)valeric acid (DMGV) was determined by LC-MS/MS in plasma and urine. As compared to controls systemic administration of BAIB increased plasma and urine BAIB levels by a factor of 26.5 (p<0.001) and 25.8 (p<0.01), respectively. BAIB infusion resulted in an increase of the plasma ADMA and SDMA concentrations of 27% and 31%, respectively, (both p<0.05) and a 24% decrease of plasma DMGV levels (p<0.05), while expression of Agxt2 was not different. Our data demonstrate that BAIB can inhibit Agxt2-mediated metabolism of dimethylarginines and show for the first time that endogenous Agxt2 is involved in the regulation of systemic ADMA, SDMA and DMGV levels. The effect of BAIB excess on endogenous dimethylarginine levels may have direct clinical implications for humans with the relatively common genetic trait of hyper-ß-aminoisobutyric aciduria.