Purinergic signalling links mechanical breath profile and alveolar mechanics with the pro-inflammatory innate immune response causing ventilation-induced lung injury

Abstract

Severe pulmonary infection or vigorous cyclic deformation of the alveolar epithelial type I (AT I) cells by mechanical ventilation leads to massive extracellular ATP release. High levels of extracellular ATP saturate the ATP hydrolysis enzymes CD39 and CD73 resulting in persistent high ATP levels despite the conversion to adenosine. Above a certain level, extracellular ATP molecules act as danger-associated molecular patterns (DAMPs) and activate the pro-inflammatory response of the innate immunity through purinergic receptors on the surface of the immune cells. This results in lung tissue inflammation, capillary leakage, interstitial and alveolar oedema and lung injury reducing the production of surfactant by the damaged AT II cells and deactivating the surfactant function by the concomitant extravasated serum proteins through capillary leakage followed by a substantial increase in alveolar surface tension and alveolar collapse. The resulting inhomogeneous ventilation of the lungs is an important mechanism in the development of ventilation-induced lung injury. The high levels of extracellular ATP and the upregulation of ecto-enzymes and soluble enzymes that hydrolyse ATP to adenosine (CD39 and CD73) increase the extracellular adenosine levels that inhibit the innate and adaptive immune responses rendering the host susceptible to infection by invading microorganisms. Moreover, high levels of extracellular adenosine increase the expression, the production and the activation of pro-fibrotic proteins (such as TGF-β, α-SMA, etc.) followed by the establishment of lung fibrosis.

Keywords: CD39; CD73; Diffuse alveolar damage; Extracellular ATP; Purinergic signalling; Ventilation-induced lung injury.

Fig. 1

Photomicrographs of rat lungs. After en bloc excision of the lungs, one of the lungs was clamped and fixed in formalin at peak inspiration pressure and the other at end expiration pressure for histologic analysis. The respiratory bronchiole, alveolar ducts and alveolar sacs are green; the alveoli are lilac; and the alveolar walls are magenta. Healthy lungs ventilated with controlled continuous mandatory ventilation (CMV) with low tidal volume (≤6 ml/kg ideal body weight) and 5 cm H₂O PEEP (control). APRV: surfactant-deactivated lungs with intratracheal instillation of a detergent ventilated with T _low being interrupted when the end-expiratory flow (EEF) reached 75% (APRV 75%) or 10% (APRV 10%) of the peak expiratory flow (PEF). (PEEP 5) Surfactant-deactivated lungs ventilated with the same settings as ‘control’ or with PEEP 16 cm H₂O (PEEP 16) (see Table 1 for the mechanical breath profile data). Note that the highest alveolar duct surface area is reached during inspiration in APRV 10% followed by PEEP 5; the lowest alveolar duct surface area is observed in the control group. The highest alveolar stability (the smallest difference in alveolar duct surface area between inspiration and expiration) with the lowest microstrain is reached in healthy lungs (control) followed by APRV 75%. The lowest alveolar stability and the highest microstrain are seen in APRV 10% and PEEP 5. Microstrain is calculated as the change in length of the alveolar ducts between inspiration and expiration normalised by their original length. The difference between the control group and PEEP 5 is exclusively attributed to the surfactant function. APRV can either significantly increase (APRV 10%) or decrease (APRV 75%) the microstrain and the redistribution of air towards the alveolar ducts. Figure from Kollisch-Singule et al. [8] with permission

Fig. 2

Schematic presentation of the fusion-activated Ca²⁺ entry (FACE). AT I cells release ATP to the extracellular space through the P2X7R (ATP receptor that can function as an ATP channel) provoked by mechanical deformation (compression or stretching) [, –56]. The extracellular ATP activates the P2Y2Rs on the surface of the AT II cells in a paracrine manner. The G protein-coupled P2Y2 ATP receptor releases DAG and IP₃ into the cytoplasm. DAG release leads to the activation of PKC-dependent pathway of fusion of LBs with the cell membrane. IP₃ release results in the release of intracellular Ca²⁺ by stores that are sensitive to IP₃ and Ca²⁺ entry from the extracellular space through several pathways (TRPV2 and STIM1/Orai1). Increased cytoplasmic Ca²⁺ level also promotes the fusion of the LBs with the cell membrane of the AT II cells. These two processes create a fusion pore causing the P2X4 ATP receptors in the membrane of the LBs to be exposed to extracellular ATP. Activation of these P2X4Rs by extracellular ATP strongly increases the local Ca²⁺ concentration to a much higher level around the membrane of the fused vesicles (FACE). FACE promotes a significant expansion of the fusion pore resulting in surfactant release by the AT II cells (LBs unpacking) and is accompanied by the FACE-induced cations and water migration from the alveolar space to the subepithelial interstitium. Clearance of the ATP molecules from the extracellular space occurs through the stepwise conversion by ecto-enzymes or by soluble extracellular enzymes (CD39 and CD73) to adenosine (ADO). ADO is returned to the cytoplasm through ENTs or CNTs and converted by ADA to inosine in the cytoplasm or converted by soluble ADA in the extracellular space. AT I alveolar epithelial type I cell, AT II alveolar epithelial type II cell, ER endoplasmic reticulum, LB lamellar body, DAG diacylglycerol, PKC protein kinase C, IP ₃ inositol triphosphate, IP ₃ R inositol triphosphate receptor, a membrane bound glycoprotein complex functioning as a Ca²⁺ channel sensitive to activation by inositol triphosphate, TRPV2 transient receptor potential cation channel subfamily V member 2, a non-selective cation channel, STIM1 stromal interaction molecule 1, a calcium sensor, Orai1 calcium release-activated calcium channel protein 1, a calcium selective ion channel, CD39 nucleoside triphosphate diphosphohydrolase 1 (NTPD1), NPP nucleotide pyrophosphatase/phosphodiesterase, CD73 5′-nucleotidase (5′-NT), ADA adenosine deaminase, ENTs equilibrative nucleoside transporters 1 and 2, CNTs concentrative nucleoside transporters 1 and 2

Fig. 3

Putative model of local tissue purinergic signalling, pathogen barriers, adaptive immunity and pro-fibrotic proteins during ARDS and/or VILI [70]. In the very early phase of ARDS and VILI, infection [43, 52, 53] and vigorous mechanical deformation of the alveoli by mechanical ventilation [212, 213] lead to the substantial increase of extracellular ATP. The ATP levels exceed the threshold for the activation of P2X7R and induce pro-inflammatory immune response [70, 211]. This causes capillary congestion and capillary leakage causing interstitial and alveolar oedema. CD39 expression is upregulated in severe sepsis [214] and after several hours of mechanical ventilation [212]. Consequently, extracellular levels of ATP gradually decrease to a certain extent and extracellular adenosine increases. In general, adenosine has potent anti-inflammatory properties. This may lead to immune paralysis against secondary specific infections. Moreover, lung tissue damage due to DAD is accompanied by the disruption of the physical barrier as a component of the innate immunity for the defence against invading pathogens and by decreased Sp-A and Sp-D levels that function as soluble pattern recognition receptors (PRRs) of the innate immune system. This renders the host susceptible to invading pathogens [42]. TGF-β expression is increased by the activation of P2X7Rs [175, 176] and activation of the adenosine receptor AdoRA2B [132]. AdoRA2B activation also increases the expression of TGF-β and other fibrotic factors such as alpha smooth muscle actin (α-SMA), connective tissue growth factor (CTGF or CCN2), IL-6, fibronectin, VEGF, CD206, arginase-1, hyaluronan, basic fibroblast growth factor (bFGF), insulin-like factor-1, etc. (Table 3, rows 40, 44, 47 and 49) [128, 130, 132, 239]

Fig. 4

The summary of the physiological, pathophysiological and immunological consequences of controlled CMV with a V _T ≤ 6 ml/kg ideal body weight (left) and of controlled CMV with extremely high V _T (right) in healthy lungs (see text for explanation). The common cell signalling pathway for the release of surfactant by alveolar epithelial type 2 (AT II) cells and for the activation of the innate immunity (red arrows). Sequential processes related to mechanical ventilation (grey coloured text boxes). CMV continuous mandatory ventilation, V _T tidal volume, APRV 10% airway pressure release ventilation with the expiration termination set at 10% of the peak-expiratory flow rate (PEFR), DAMPs danger-associated molecular patterns, DAD diffuse alveolar damage

Fig. 5

The summary of the physiological, pathophysiological and immunological consequences of controlled CMV with a V _T ≤ 6 ml/kg ideal body weight or APRV 10% (left) and of APRV 75% (right) in infected lungs (see text for explanation). The common cell signalling pathway for the release of surfactant by alveolar epithelial type 2 (AT II) cells and for the activation of the innate immunity (red arrows). Sequential processes related to mechanical ventilation (grey coloured text boxes). CMV continuous mandatory ventilation, V _T tidal volume, APRV 75% airway pressure release ventilation with the expiration termination set at 75% of the peak-expiratory flow rate (PEFR), APRV 10% APRV with the expiration termination set at 10% of the PEFR, DAMPs danger-associated molecular patterns, DAD diffuse alveolar damage