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 3

Glutamine administration functionally attenuates renal tubular cell apoptosis.

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Glutamine administration functionally attenuates renal tubular cell apop...
WT mice were subjected to sham or IRI surgery and received glutamine or saline. Paraffin-embedded tissue sections were prepared and TUNEL staining was performed (A, TUNEL-positive cells [%]; B, representative TUNEL-stained cortex and medulla tissue) (n = 4; scale bar: 50 μm). Caspase-3 activity was detected in kidney lysates (C, n = 4). Western blotting was performed to assess the expression level of HSP70 (D and E), p-JNK (D and F), 14-3-3ζ and p–14‑3‑3ζ (Thr232) (D and G), Bad and p-Bad (Ser136) (D and H), caspase-3 (D and I), as well as apoptosis signal-regulating kinase (Ask1) and p-Ask1 (D and J). TECs were transfected with Ask1 siRNA and control siRNA using the Lipofectamine RNAiMAX Reagent. Knockdown efficiency determined by Western blot analysis was ~10% protein expression. Transfected TECs were treated with glutamine or saline, then subjected to hypoxia. TEC lysates were analyzed by Western blotting to assess the expression levels of HSP70 (K and L, n = 3) and caspase-3 (K and M, n = 3). MALDI imaging mass spectrometry (MALDI-IMS) of kidney sections was performed to analyze protein distribution of HSP70, 14-3-3ζ, and Bad (N). The scale represents the relative intensity of the protein (m/z, mass-to-charge ratio). Mean ± SEM, 1-way ANOVA *P < 0.05; **P < 0.005; ***P < 0.001.