Glutamine prevents acute kidney injury by modulating oxidative stress and apoptosis in tubular epithelial cells
Katharina Thomas, Lisa Zondler, Nadine Ludwig, Marina Kardell, Corinna Lüneburg, Katharina Henke, Sina Mersmann, Andreas Margraf, Tilmann Spieker, Tobias Tekath, Ana Velic, Richard Holtmeier, Juliane Hermann, Vera Jankowski, Melanie Meersch, Dietmar Vestweber, Martin Westphal, Johannes Roth, Michael A. Schäfers, John A. Kellum, Clifford A. Lowell, Jan Rossaint, Alexander Zarbock
Katharina Thomas, Lisa Zondler, Nadine Ludwig, Marina Kardell, Corinna Lüneburg, Katharina Henke, Sina Mersmann, Andreas Margraf, Tilmann Spieker, Tobias Tekath, Ana Velic, Richard Holtmeier, Juliane Hermann, Vera Jankowski, Melanie Meersch, Dietmar Vestweber, Martin Westphal, Johannes Roth, Michael A. Schäfers, John A. Kellum, Clifford A. Lowell, Jan Rossaint, Alexander Zarbock
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Research Article Immunology Nephrology

Glutamine prevents acute kidney injury by modulating oxidative stress and apoptosis in tubular epithelial cells

Abstract

Acute kidney injury (AKI) represents a common complication in critically ill patients that is associated with increased morbidity and mortality. In a murine AKI model induced by ischemia/reperfusion injury (IRI), we show that glutamine significantly decreases kidney damage and improves kidney function. We demonstrate that glutamine causes transcriptomic and proteomic reprogramming in murine renal tubular epithelial cells (TECs), resulting in decreased epithelial apoptosis, decreased neutrophil recruitment, and improved mitochondrial functionality and respiration provoked by an ameliorated oxidative phosphorylation. We identify the proteins glutamine gamma glutamyltransferase 2 (Tgm2) and apoptosis signal-regulating kinase (Ask1) as the major targets of glutamine in apoptotic signaling. Furthermore, the direct modulation of the Tgm2-HSP70 signalosome and reduced Ask1 activation resulted in decreased JNK activation, leading to diminished mitochondrial intrinsic apoptosis in TECs. Glutamine administration attenuated kidney damage in vivo during AKI and TEC viability in vitro under inflammatory or hypoxic conditions.

Authors

Katharina Thomas, Lisa Zondler, Nadine Ludwig, Marina Kardell, Corinna Lüneburg, Katharina Henke, Sina Mersmann, Andreas Margraf, Tilmann Spieker, Tobias Tekath, Ana Velic, Richard Holtmeier, Juliane Hermann, Vera Jankowski, Melanie Meersch, Dietmar Vestweber, Martin Westphal, Johannes Roth, Michael A. Schäfers, John A. Kellum, Clifford A. Lowell, Jan Rossaint, Alexander Zarbock

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Figure 6

Glutamine affects the expression of apoptosis- and oxidative stress–related proteins in renal TECs on the transcriptional and proteomic level.

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Glutamine affects the expression of apoptosis- and oxidative stress–rela...
TECs were identified in kidney homogenates by FACS based on the cell-specific marker prominin-1. RNA was isolated and sequenced. Hierarchical clustering heatmaps of RNA-Seq indicate differentially expressed genes (rows) between TECs of glutamine-treated mice after IRI and glutamine treatment compared with vehicle control (A, n = 3). Blue color indicates downregulation and red color indicates upregulation. Volcano plots generated to compare glutamine versus saline treatment after IRI induction identify 481 differently expressed genes in renal TECs (B). GO pathway enrichment analyses specify the fold enrichment of the transcriptome sequencing relative to the Mus musculus genome in TECs (C). GO terms representing molecular function are presented in red, cellular component in green, and biological processes in blue (Fisher’s exact test, P value < 0.05). TECs were isolated from murine kidneys and exposed to TNF-α for 18 hours or hypoxia for 24 hours. Additionally, TECs were treated with glutamine or saline as well as Tgm2 inhibitor ERW1041E or vehicle control, respectively. Caspase-3 activity was detected after TNF-α (n = 5) or hypoxia stimulation (n = 3, D), and Tgm2 levels were determined by Western blotting (E and F). Tgm2 activity in kidneys was assessed (G, n = 4). Immuno- and coimmunoprecipitated proteins were separated by SDS-PAGE and blotted using the indicated antibodies. Whole-cell lysate (INPUT) was used as protein control (H, n = 3). Mean ± SEM; 1-way ANOVA *P < 0.05; **P < 0.005; ***P < 0.001.