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Glyoxalases prevent the formation of advanced glycation end products by converting glycolysis-derived methylglyoxal to d-lactate with the help of glutathione. Vander Jagt and colleagues previously showed that erythrocytes release about thirty times more d-lactate after infection with the human malaria parasite Plasmodium falciparum. Functional glyoxalases in the host-parasite unit might therefore be crucial for parasite survival. Here, we determined the antimalarial and hemolytic activity of two tight-binding glyoxalase inhibitors using infected and uninfected erythrocytes. In addition, we synthesized and analyzed a set of diester derivates of both tight-binding inhibitors resulting in up to threefold lower IC50 values and an altered methemoglobin formation and hemolytic activity depending on the type of ester. Inhibitor treatments of uninfected erythrocytes revealed an extremely slow inactivation of the host cell glyoxalase, irrespective of inhibitor modifications, and a potential dispensability of the host cell enzyme for parasite survival. Our study highlights the benefits and drawbacks of different esterifications of glutathione-derived inhibitors and demonstrates the suitability of glyoxalase inhibitors as a tool for deciphering the relevance and mode of action of different glyoxalase systems in a host-parasite unit., Graphical abstract fx1

Objective Diabetic retinopathy (DR) is induced by an accumulation of reactive metabolites such as ROS, RNS, and RCS species, which were reported to modulate the activity of cation channels of the TRPC family. In this study, we use Trpc1/4/5/6−/− compound knockout mice to analyze the contribution of these TRPC proteins to diabetic retinopathy. Methods We used Nanostring- and qPCR-based analysis to determine mRNA levels of TRPC channels in control and diabetic retinae and retinal cell types. Chronic hyperglycemia was induced by Streptozotocin (STZ) treatment. To assess the development of diabetic retinopathy, vasoregression, pericyte loss, and thickness of individual retinal layers were analyzed. Plasma and cellular methylglyoxal (MG) levels, as well as Glyoxalase 1 (GLO1) enzyme activity and protein expression, were measured in WT and Trpc1/4/5/6−/− cells or tissues. MG-evoked toxicity in cells of both genotypes was compared by MTT assay. Results We find that Trpc1/4/5/6−/− mice are protected from hyperglycemia-evoked vasoregression determined by the formation of acellular capillaries and pericyte drop-out. In addition, Trpc1/4/5/6−/− mice are resistant to the STZ-induced reduction in retinal layer thickness. The RCS metabolite methylglyoxal, which represents a key mediator for the development of diabetic retinopathy, was significantly reduced in plasma and red blood cells (RBCs) of STZ-treated Trpc1/4/5/6−/− mice compared to controls. GLO1 is the major MG detoxifying enzyme, and its activity and protein expression were significantly elevated in Trpc1/4/5/6-deficient cells, which led to significantly increased resistance to MG toxicity. GLO1 activity was also increased in retinal extracts from Trpc1/4/5/6−/− mice. The TRPCs investigated here are expressed at different levels in endothelial and glial cells of the retina. Conclusion The protective phenotype in diabetic retinopathy observed in Trpc1/4/5/6−/− mice is suggestive of a predominant action of TRPCs in Müller cells and microglia because of their central position in the retention of a proper homoeostasis of the neurovascular unit., Highlights • Trpc1/4/5/6−/− mice are protected from hyperglycemia-evoked vasoregression and damage to the neurovascular unit. • The level of methylglyoxal (MG), which plays a crucial role in the development of DR is reduced in Trpc1/4/5/6−/− mice. • The protection against DR in Trpc1/4/5/6−/− mice may be mediated by an increase in GLO1 activity and protein expression.