Martin Reichel

Aims: Cardiovascular disease (CVD) is the leading cause of mortality in chronic kidney disease (CKD). While CKD is known to give rise to systemic inflammation, its inciting factors remain poorly defined. Oxalate, long implicated in rare genetic kidney disorders, accumulates with decreased kidney function and has emerged as a driver of inflammation and independent risk factor for CVD. Here, we investigate the immunological mechanisms linking oxalate nephropathy to systemic inflammation, cardiac damage and kidney injury. Methods and results: Oxalate nephropathy was induced in C57BL6/N mice through an oxalate-enriched diet. Oxalate induced systemic immune activation, renal fibrosis, and adverse cardiac remodelling, including pulmonary congestion with systolic and diastolic dysfunction. Flow cytometry analysis identified interleukin (IL)-17A as a dominant inflammatory effector, with expansion of Th17 and Th17-like Treg in the kidney, intestine, and spleen. Bulk mRNA sequencing confirmed these findings in kidney and heart. In line, plasma IL-17A was increased in oxalate-fed mice. Confirming the oxalate-IL-17A relationship, plasma IL-17A was elevated in patients with primary hyperoxaluria. Gut microbiome analysis by 16S amplicon sequencing showed only mild oxalate-induced alterations in mice. However, soluble oxalate directly enhanced Th17 polarization and disrupted mitochondrial respiration in vitro. In vivo, antibody-mediated IL-17A blockade improved kidney function, cardiac fibrosis, reduced neutrophil infiltration, and partially restored cardiac function in oxalate-fed mice. Conclusion: Our study identifies oxalate as a systemic immunometabolic stressor and IL-17A as a central mediator of oxalate-induced cardiorenal injury. These findings establish the oxalate-IL-17A axis as a mechanistic link between CKD and CVD and suggest IL-17A inhibition as a potential therapeutic strategy to reduce cardiovascular damage in CKD.

Aims: Cardiovascular disease (CVD) is the leading cause of mortality in chronic kidney disease (CKD). While CKD is known to give rise to systemic inflammation, its inciting factors remain poorly defined. Oxalate, long implicated in rare genetic kidney disorders, accumulates with decreased kidney function and has emerged as a driver of inflammation and independent risk factor for CVD. Here, we investigate the immunological mechanisms linking oxalate nephropathy to systemic inflammation, cardiac damage and kidney injury. Methods and Results: Oxalate nephropathy was induced in C57Bl6/N mice via an oxalate-enriched diet. Oxalate induced systemic immune activation, renal fibrosis, and cardiac remodeling, including pulmonary congestion with systolic and diastolic dysfunction. Flow cytometry analysis identified interleukin (IL)-17A as a dominant inflammatory effector, with expansion of Th17 and Th17-like Treg in the kidney, intestine, and spleen. Bulk mRNA sequencing confirmed these findings. Confirming the oxalate-IL-17A relationship, plasma IL-17A was elevated in patients with primary hyperoxaluria. Gut microbiome analysis by 16S amplicon sequencing showed only mild oxalate-induced alterations in mice. However, soluble oxalate directly enhanced Th17 polarization and disrupted mitochondrial respiration in vitro . In vivo, antibody-mediated (clone: 17F3) IL-17A blockade decreased cardiac fibrosis, reduced neutrophil infiltration, and partially restored cardiac function in oxalate-fed mice. Conclusions: Our study identifies oxalate as a systemic immunometabolic stressor and IL-17A as a central mediator of oxalate-induced cardiorenal injury. These findings establish the oxalate–IL-17A axis as a mechanistic link between CKD and CVD and suggest IL-17A inhibition as a potential therapeutic strategy to reduce cardiovascular damage in CKD. Translational Perspective: This study advances our understanding of targetable mediators of the cardiovascular risk in chronic kidney disease (CKD). We identify the organic anion oxalate, beyond its traditional role in crystal-induced kidney damage, as a systemic immunometabolic stressor. We demonstrate that oxalate induces IL-17A–mediated inflammation and thereby contributes to maladaptive cardiac remodeling. Therapeutic blockade of IL-17A protects against oxalate-induced cardiorenal injury, highlighting the oxalate–IL-17A axis as a relevant and druggable link between CKD and cardiovascular disease.

Significance Statement: Oxalate is a toxic end product of metabolism that is highly elevated in patients with kidney failure requiring chronic dialysis. Basic research has demonstrated that oxalate is a potential trigger of systemic inflammation, progression of CKD, and cardiovascular complications. The authors conducted a post-hoc analysis of the randomized German Diabetes Dialysis Study of 1255 European patients with diabetes on hemodialysis. In their analysis of 1108 patients, they found those with higher serum oxalate concentrations were more likely than those with lower levels of serum oxalate to experience cardiovascular mortality, particularly sudden cardiac death. These findings suggest oxalate-lowering therapeutic strategies might have potential for decreasing cardiovascular mortality in patients with kidney disease. Background: The clinical significance of accumulating toxic terminal metabolites such as oxalate in patients with kidney failure is not well understood. Methods: To evaluate serum oxalate concentrations and risk of all-cause mortality and cardiovascular events in a cohort of patients with kidney failure requiring chronic dialysis, we performed a post-hoc analysis of the randomized German Diabetes Dialysis (4D) Study; this study included 1255 European patients on hemodialysis with diabetes followed-up for a median of 4 years. The results obtained via Cox proportional hazards models were confirmed by competing risk regression and restricted cubic spline modeling in the 4D Study cohort and validated in a separate cohort of 104 US patients on dialysis after a median follow-up of 2.5 years. Results: A total of 1108 patients had baseline oxalate measurements, with a median oxalate concentration of 42.4 µM. During follow-up, 548 patients died, including 139 (25.4%) from sudden cardiac death. A total of 413 patients reached the primary composite cardiovascular end point (cardiac death, nonfatal myocardial infarction, and fatal or nonfatal stroke). Patients in the highest oxalate quartile (≥59.7 µM) had a 40% increased risk for cardiovascular events (adjusted hazard ratio [aHR], 1.40; 95% confidence interval [95% CI], 1.08 to 1.81) and a 62% increased risk of sudden cardiac death (aHR, 1.62; 95% CI, 1.03 to 2.56), compared with those in the lowest quartile (≤29.6 µM). The associations remained when accounting for competing risks and with oxalate as a continuous variable. Conclusions: Elevated serum oxalate is a novel risk factor for cardiovascular events and sudden cardiac death in patients on dialysis. Further studies are warranted to test whether oxalate-lowering strategies improve cardiovascular mortality in patients on dialysis.

Major depressive disorder (MDD) is a severe psychiatric condition with key symptoms of low mood and lack of motivation, joy, and pleasure. Recently, the acid sphingomyelinase (ASM)/ceramide system has been implicated in the pathogenesis of MDD. ASM is a lysosomal glycoprotein that catalyzes the hydrolysis of sphingomyelin, an abundant component of membranes, into the bioactive sphingolipid ceramide, which impacts signaling pathways. ASM activity is inhibited by several common antidepressant drugs. Human and murine studies have confirmed that increased ASM activity and ceramide levels are correlated with MDD. To define a molecular marker for treatment monitoring, we investigated the mRNA expression of SMPD1, which encodes ASM, in primary cell culture models, a mouse study, and a human study with untreated MDD patients before and after antidepressive treatment. Our cell culture study showed that a common antidepressant inhibited ASM activity at the enzymatic level and also at the transcriptional level. In a genetically modified mouse line with depressive-like behavior, Smpd1 mRNA expression in dorsal hippocampal tissue was significantly decreased after treatment with a common antidepressant. The large human study showed that SMPD1 mRNA expression in untreated MDD patients decreased significantly after antidepressive treatment. This translational study shows that SMPD1 mRNA expression could serve as a molecular marker for treatment and adherence monitoring of MDD.

Significance Statement: Fecal oxalate excretion is critical in oxalate balance with progression of CKD. However, the identity of the transporter(s) responsible for increased intestinal oxalate secretion in CKD is unknown. Intestinal expression of oxalate transporter Slc26a6 is strongly upregulated in two murine models of CKD. Deletion of Slc26a6 completely abrogates enhanced fecal oxalate excretion in CKD, increasing plasma oxalate concentration. This study demonstrates that Slc26a6 mediates intestinal oxalate secretion and mitigates hyperoxalemia in murine CKD models. The findings suggest that pharmacologic approaches enhancing intestinal Slc26a6 activity may stimulate extrarenal clearance of oxalate and prevent hyperoxalemia in CKD. Future studies are needed to address whether similar mechanisms contribute to intestinal oxalate elimination in humans to enhance extrarenal oxalate clearance. Background: A state of oxalate homeostasis is maintained in patients with healthy kidney function. However, as GFR declines, plasma oxalate (P_ox) concentrations start to rise. Several groups of researchers have described augmentation of oxalate secretion in the colon in models of CKD, but the oxalate transporters remain unidentified. The oxalate transporter Slc26a6 is a candidate for contributing to the extrarenal clearance of oxalate via the gut in CKD. Methods: Feeding a diet high in soluble oxalate or weekly injections of aristolochic acid induced CKD in age- and sex-matched wild-type and Slc26a6 ^−/− mice. qPCR, immunohistochemistry, and western blot analysis assessed intestinal Slc26a6 expression. An oxalate oxidase assay measured fecal and P_ox concentrations. Results: Fecal oxalate excretion was enhanced in wild-type mice with CKD. This increase was abrogated in Slc26a6 ^−/− mice associated with a significant elevation in plasma oxalate concentration. Slc26a6 mRNA and protein expression were greatly increased in the intestine of mice with CKD. Raising P_ox without inducing kidney injury did not alter intestinal Slc26a6 expression, suggesting that changes associated with CKD regulate transporter expression rather than elevations in P_ox. Conclusions: Slc26a6-mediated enteric oxalate secretion is critical in decreasing the body burden of oxalate in murine CKD models. Future studies are needed to address whether similar mechanisms contribute to intestinal oxalate elimination in humans to enhance extrarenal oxalate clearance.

Human and murine studies identified the lysosomal enzyme acid sphingomyelinase (ASM) as a target for antidepressant therapy and revealed its role in the pathophysiology of major depression. In this study, we generated a mouse model with overexpression of Asm (Asm-tg^fb) that is restricted to the forebrain to rule out any systemic effects of Asm overexpression on depressive-like symptoms. The increase in Asm activity was higher in male Asm-tg^fb mice than in female Asm-tg^fb mice due to the breeding strategy, which allows for the generation of wild-type littermates as appropriate controls. Asm overexpression in the forebrain of male mice resulted in a depressive-like phenotype, whereas in female mice, Asm overexpression resulted in a social anxiogenic-like phenotype. Ceramides in male Asm-tg^fb mice were elevated specifically in the dorsal hippocampus. mRNA expression analyses indicated that the increase in Asm activity affected other ceramide-generating pathways, which might help to balance ceramide levels in cortical brain regions. This forebrain-specific mouse model offers a novel tool for dissecting the molecular mechanisms that play a role in the pathophysiology of major depression.

Oxalate crystal-induced renal inflammation is associated with progressive kidney failure due to activation of the NLRP3/CASP-1 inflammasome. It has been suggested previously that purinergic P2X7 receptor signaling is critical for crystal-induced inflammasome activation and renal injury. Therefore, we investigated the role of the P2X7 receptor in response to crystal-induced cytokine release, inflammation, and kidney failure using in vitro and in vivo models. Dendritic cells and macrophages derived from murine bone marrow and human peripheral blood mononucleated cells stimulated with calcium-oxalate crystals, monosodium urate crystals, or ATP lead to the robust release of interleukin-1beta (IL-1ß). Treatment with the P2X7 inhibitor A740003 or the depletion of ATP by apyrase selectively abrogated ATP-induced, but not oxalate and urate crystal-induced IL-1ß release. In line with this finding, dendritic cells derived from bone marrow (BMDCs) from P2X7^−/− mice released reduced amounts of IL-1ß following stimulation with ATP, while oxalate and urate crystal-induced IL-1ß release was unaffected. In sharp contrast, BMDCs from Casp1^−/− mice exhibited reduced IL-1ß release following either of the three stimulants. In addition, P2X7^−/− mice demonstrated similar degrees of crystal deposition, tubular damage and inflammation when compared with WT mice. In line with these findings, increases in plasma creatinine were no different between WT and P2X7^−/− mice. In contrast to previous reports, our results indicate that P2X7 receptor is not required for crystal-induced CKD and it is unlikely to be a suitable therapeutic target for crystal-induced progressive kidney disease.

Recent investigations propose the acid sphingomyelinase (ASM)/ceramide system as a novel target for antidepressant action. ASM catalyzes the breakdown of the abundant membrane lipid sphingomyelin to the lipid messenger ceramide. This ASM‐induced lipid modification induces a local shift in membrane properties, which influences receptor clustering and downstream signaling. Canonical transient receptor potential channels 6 (TRPC6) are non‐selective cation channels located in the cell membrane that play an important role in dendritic growth, synaptic plasticity and cognition in the brain. They can be activated by hyperforin, an ingredient of the herbal remedy St. John’s wort for treatment of depression disorders. Because of their role in the context of major depression, we investigated the crosstalk between the ASM/ceramide system and TRPC6 ion channels in a pheochromocytoma cell line 12 neuronal cell model (PC12 rat pheochromocytoma cell line). Ca²⁺ imaging experiments indicated that hyperforin‐induced Ca²⁺ influx through TRPC6 channels is modulated by ASM activity. While antidepressants, known as functional inhibitors of ASM activity, reduced TRPC6‐mediated Ca²⁺ influx, extracellular application of bacterial sphingomyelinase rebalanced TRPC6 activity in a concentration‐related way. This effect was confirmed in whole‐cell patch clamp electrophysiology recordings. Lipidomic analyses revealed a decrease in very long chain ceramide/sphingomyelin molar ratio after ASM inhibition, which was connected with changes in the abundance of TRPC6 channels in flotillin‐1–positive lipid rafts as visualized by western blotting. Our data provide evidence that the ASM/ceramide system regulates TRPC6 channels likely by controlling their recruitment to specific lipid subdomains and thereby fine‐tuning their physical properties. image

Major depression and alcohol use disorder are severe psychiatric diseases affecting the world's population with high comorbidity level. However, the pathogenesis of this comorbidity remains unclear, and no selective treatment for this condition is available. A pathogenic pathway and a possible therapeutic target for the treatment of depression-alcoholism comorbidity based on the hyperfunction of acid sphingomyelinase (Asm) were recently suggested. Here we analyzed the effects of alcohol on the depression/anxiety state of homozygous Asm-knockout mice (Asm − /−), which can be considered as a model of an early stage of Niemann-Pick disease, as well as their drinking pattern under normal and stress conditions. It was observed that forced treatment with alcohol (2 g/kg, i.p.) reduces the anxiety level of Asm−/− mice as measured in the elevated plus maze (EPM) test, but enhances the depression level in the forced swim test (FST). The analysis of drinking pattern of these animals in a free-choice alcohol drinking paradigm revealed higher alcohol intake and preference in Asm−/− mice compared to wild type (wt) littermates. However, this difference was overwritten by the stress exposure. Stronger sedating effects of alcohol were observed in Asm−/− mice compared to wt animals in the loss of righting reflex test after single and repeated alcohol injections (3 g/kg, i.p.). Altogether, the present findings might indicate an Asm involvement in the mechanisms of comorbidity between alcoholism and anxiety/depression.