Zinc deficiency primes the lung for ventilator-induced injury
Francis Boudreault, Miguel Pinilla-Vera, Joshua A. Englert, Alvin T. Kho, Colleen Isabelle, Antonio J. Arciniegas, Diana Barragan-Bradford, Carolina Quintana, Diana Amador-Munoz, Jiazhen Guan, Kyoung Moo Choi, MICU Registry, Lynette Sholl, Shelley Hurwitz, Daniel J. Tschumperlin, Rebecca M. Baron
Francis Boudreault, Miguel Pinilla-Vera, Joshua A. Englert, Alvin T. Kho, Colleen Isabelle, Antonio J. Arciniegas, Diana Barragan-Bradford, Carolina Quintana, Diana Amador-Munoz, Jiazhen Guan, Kyoung Moo Choi, MICU Registry, Lynette Sholl, Shelley Hurwitz, Daniel J. Tschumperlin, Rebecca M. Baron
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Research Article Inflammation Pulmonology

Zinc deficiency primes the lung for ventilator-induced injury

Abstract

Mechanical ventilation is necessary to support patients with acute lung injury, but also exacerbates injury through mechanical stress–activated signaling pathways. We show that stretch applied to cultured human cells, and to mouse lungs in vivo, induces robust expression of metallothionein, a potent antioxidant and cytoprotective molecule critical for cellular zinc homeostasis. Furthermore, genetic deficiency of murine metallothionein genes exacerbated lung injury caused by high tidal volume mechanical ventilation, identifying an adaptive role for these genes in limiting lung injury. Stretch induction of metallothionein required zinc and the zinc-binding transcription factor MTF1. We further show that mouse dietary zinc deficiency potentiates ventilator-induced lung injury, and that plasma zinc levels are significantly reduced in human patients who go on to develop acute respiratory distress syndrome (ARDS) compared with healthy and non-ARDS intensive care unit (ICU) controls, as well as with other ICU patients without ARDS. Taken together, our findings identify a potentially novel adaptive response of the lung to stretch and a critical role for zinc in defining the lung’s tolerance for mechanical ventilation. These results demonstrate that failure of stretch-adaptive responses play an important role in exacerbating mechanical ventilator–induced lung injury, and identify zinc and metallothionein as targets for lung-protective interventions in patients requiring mechanical ventilation.

Authors

Francis Boudreault, Miguel Pinilla-Vera, Joshua A. Englert, Alvin T. Kho, Colleen Isabelle, Antonio J. Arciniegas, Diana Barragan-Bradford, Carolina Quintana, Diana Amador-Munoz, Jiazhen Guan, Kyoung Moo Choi, MICU Registry, Lynette Sholl, Shelley Hurwitz, Daniel J. Tschumperlin, Rebecca M. Baron

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Figure 3

Stretch-induced metallothionein (MT) expression is zinc and MTF1 dependent.

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Stretch-induced metallothionein (MT) expression is zinc and MTF1 depende...
(A) CCL-151 cells were treated with 100 μM extracellular zinc (basal extracellular free Zn2+ level in media is ~1 μM) for 2, 4, 16, and 24 hours and probed for MT transcript levels by real-time quantitative PCR (qPCR) (n = 2 per time point). (B) CCL-151 cells were treated with extracellular zinc for 4 hours and probed for MT1M and MT1G transcript levels by qPCR. Responses were normalized to basal extracellular free Zn2+ level ~1 μM (n = 1 per dose). (C) Intracellular and extracellular zinc were chelated with membrane-permeant TPEN (10 μM) to an estimated level below 1 pM intra- and extracellularly, 15 minutes prior to stretching CCL-151 cells for 4 hours. Stretched and time-matched control cells were then probed for MT transcript levels and responses normalized to TPEN-free stretch induction (n = 4 samples per measurement). *P < 0.05, Student’s 2-tailed t test. (D) A549 cells transfected with MTF1-responsive luciferase reporter were exposed to tonic stretch (30% strain) for 4 hours prior to measuring luminescence (n = 3). *P < 0.05, Student’s 2-tailed t test. (E) Treatment with siRNA (50 nM) targeting MTF1 (+RNAi) for 72 hours decreased transcript levels in CCL-151 and A549 cells relative to control (fold change 1.0) (n = 3), while scrambled siRNA (+RNAi scr) did not affect MTF1 transcript levels (n = 3). *P < 0.05 versus both control and scrambled siRNA, Student’s 2-tailed t test. (F) Knockdown of MTF1 with siRNA (+RNAi) prevented stretch-induced expression of MT1X in A549 and CCL-151 cells, with similar trends observed for MT1G (n = 2). *P < 0.05, Student’s 2-tailed t test.