Non‐acidotic hypercapnia limits the loss of diaphragm force at single fiber level in mechanically ventilated rats for 5 days

Abstract

The diaphragm loss of function resulting from Mechanical Ventilation (MV) in Intensive Care Unit (ICU) patients has been termed “Ventilator Induced Diaphragmatic Dysfunction” (VIDD) and is characterized by a rapid and progressive loss of diaphragm muscle mass and function. Hypercapnia is a frequent condition in mechanically ventilated ICU patients and accepted as “permissive hypercapnia” in ARDS. There are numerous negative effects of this gas imbalance on different tissues and their functions are well known, whereas the positive effects are still poorly understood. Acute hypercapnic exposure has shown benefit to in vivo diaphragm function in porcine and rat ICU models, however our understanding of the effects of longer‐term ventilation and the downstream signaling are less known. Here, we hypothesize that normoxic non acidotic hypercapnia would augment diaphragm muscle function during development of VIDD. Adult female Sprague‐Dawley rats were deeply sedated, pharmacologically paralyzed, hydrated, nourished and controlled mechanical ventilated (CMV) for 5 days in normoxic‐normocapnic or normoxic, hypercapnic conditions. Hypercapnic conditions maintained EtCO2 between 55 and 70 mmHg for the duration of the experiment ensuring normal arterial blood pH (7.37 – 7.43), thus they were non‐acidotic hypercapnic conditions. This unique experimental rat model is more representative of the prolonged nature and effects of VIDD and thus provides clinically relevant assessment of hypercapnic treatment efficacy. Diaphragm muscles were dissected and prepared for contractile function assessment and downstream signalling assessed at the gene and protein level.Our results show that: 1) cross sectional area (CSA) was decreased by 40% following normocapnic CMV when compared with CTR (p< 0.001), which was unaltered by hypercapnia treatment; 2) Specific force (SF; force normalised to CSA) was decreased to 58% of CTR following normocapnic CMV (p< 0.001). Non‐acidotic hypercapnia increased SF by 15% when compared with normocapnic rats (p< 0.05); 3) Muscle E3 ligases Murf1 and Atrogin‐1 protein expression is unchanged following 5 days of CMV, irrespective of C02 conditions. LC3B is a known marker for autophagy, with LC3B II:I ratio signifying activation. Following 5 days normocapnic CMV LC3B II:I ratio (p< 0.05) was reduced by 70% in diaphragm muscle. Non‐acidotic hypercapnic conditions failed to alter the activation of LC3B or these muscle degradation pathways; 4) Non‐acidotic hypercapnic conditions however increased TNFα and Interleukin 1β transcript expression in the diaphragm when compared with normocapnic CMV and CTR groups; 5) Non‐acidotic hypercapnic conditions increased stability of respiratory peak pressure, peripheral oxygenation and perfusion, hemodynamic conditions, thus promoting the benefits of non‐acidotic hypercapnic conditions.Our results suggest that non‐acidotic hypercapnic conditions have beneficial effects on single fibre diaphragm function irrespective of significant increase in muscle inflammation in non‐septic animals. This, combined with the positive respiratory and hemodynamic effects, encourage further study of the therapeutic potential of CO2 in ICU contexts.