THE DONOR LUNG: A TROJAN HORSE?

Donor “passenger” leukocytes: unwelcome guests

Mounting evidence suggests that “passenger” leukocytes from the donor lung play a key role in the development of transplant-related ischemia-reperfusion injury and its sequelae including Primary Graft Dysfunction (PGD) and even acute rejection.

Several recent studies indicate that inflammatory cytokines and cascades triggered by the activation and adhesion of circulating platelets and leukocytes after re-establishment of oxygen-rich blood flow to the ischemic microcirculation are key players in the etiology of PGD. The role of these hyperactivated adhesive platelets and leukocytes in the development of ischemia-reperfusion injury under such conditions was first demonstrated in real-time (intravital) studies on the microcirculation some 30 years ago, primarily by pioneers in the field of intravital microscopy lead by the German München group headed by Prof Konrad Messmer (Menger 1993, 1995; Nolte 1991 Steinbauer 1996; Werner 1996; Shasby 1983; Modig 1988).

In poorly perfused tissue, secondary to hypovolemia, arterial stenosis or ligation, (as in the isolated organ), they showed that hypoxia triggers cascades which increase the adhesiveness and rigidity of circulating leukocytes. These hyperadhesive leukocytes impair flow in the microcirculation and play a far greater role in further reducing microvascular flow than do concomitant changes in plasma or perfusate viscosity or red cell flexibility (Chien 1987). They virtually double the resistance to microcirculatory flow despite the fact that leukocytes normally comprise far less than 1% of all blood cells.

In particular, they showed in numerous studies over the succeeding 15 years how the deleterious effects of ischemia-reperfusion could be mitigated by various interventions, not least by the presence of dextran (a key component of PERFADEX and STEEN solution) in the reperfusing medium. (Arfors, Buckley 1997). Since then successive groups have further elaborated on the complex details of the cascades involved.

Recent studies and remedial measures

Several recent related studies have shed further new light on the role of donor “passenger” leukocytes in the development of primary graft dysfunction (PGD), a condition virtually synonymous with the “ischemia-reperfusion injury” described above, which arises when the isolated ischemic organ is finally transplanted and reperfused by the recipient´s own systemic circulation.

Last year we highlighted the important findings of a joint study from Manchester, Oslo and Lund (Stone et al, 2016) in which the removal of donor “passenger” leukocytes by filtration during ex-vivo lung perfusion (EVLP) of porcine lungs was shown to “reduce direct allorecognition and T cell priming, thereby diminishing recipient T cell infiltration, the hallmark of acute rejection”.

Several related studies have been published this year. Tatham et al (2) for example, from the London-based group, subjected isolated mouse lungs to two hours of warm ischaemia followed by two hours of reperfusion under normoxic conditions. As expected, reperfusion triggered the adhesion of substantial numbers of activated leukocytes (monocytes) to the endothelial walls of the microcirculation, accompanied by reduced L-selectin and increased CD86 indicating their activation. Conversely, monocyte depletion resulted in reductions of lung edema, BAL fluid protein and perfusate levels of inflammatory biomarkers RAGE, MIP-2 and KC.

In human lungs, the same group observed correlations between pre-implantation donor monocyte numbers, their CD86 and TREM-1 expression and post-implantation lung dysfunction at 48 and 72 hours, clearly linking donor monocyte activity to poorer outcomes.

And in a related study, the Zurich group, (Iskender 2017) investigated the effects of removing cytokines during EVLP of porcine lungs using a dedicated adsorbent bead device (Cytosorb). The authors found that cytokine removal in this way during EVLP “decreased the development of pulmonary edema and electrolyte imbalance through the suppression of anaerobic glycolysis and neutrophil activation in this setting.”

Yet another related study, in rats, from Pittsburgh (Noda 2017) demonstrated that circulating leukocytes derived from donor lungs, (and not circulating proinflammatory cytokines) substantially impaired the quality of lung grafts through Caspase-1 induced pyroptotic cell death during EVLP. The authors suggested that removing these cells with a leukocyte filter (as, of course, is standard practise with XPS and similar EVLP circuits) and/or inhibiting pyroptosis of the cells “can be a new therapeutic approach leading to long-term success following lung transplantation.”

Conclusion

Taken together, the above evidence clearly indicates that “passenger” leukocytes in the donor lung are NOT particularly welcome guests. Moreover, the donor lung itself, which has functioned during the last few hours of life as the body´s principle filter for all manner of circulating thromboembolic debris, microaggregates of activated adhesive leukocytes, platelets, fibrin, etc, functions in principle as a veritable Trojan horse. And, of course, most lungs are generally retrieved after the “cytokine storm” associated with brain-death or the traumatic stress of DCD donation.

Fortunately, the presence of dextran in PERFADEX® and STEEN® solution suppresses further activation and adhesion of leukocytes to the vessel walls, whilst early retrograde perfusion with PERFADEX will generally flush out most thromboemboli.

And, of course, all lungs subjected to EVLP, (whether sub-optimal or “standard” lungs), will be subjected to continuous “passenger” leukocyte removal thanks to the integrated leukocyte filters nowadays incorporated into the XPS and similar EVLP systems.

Xvivo Insights PB-2017-06-24

References

Menger 1993
Menger M, Thierjung C, Hammersen, F, Messmer, K.
Dextran vs hydoxyethylstarch in inhibition of postischemic leukocyte adherence in striated muscle. Circ Shock. 1993 Dec;41(4):248-55.

Menger 1995
Menger, M D (1995.) Microcirculatory disturbances secondary to ischemia-reperfusion. Transplantation proceedings 27(5):2863 – 2865

Nolte 1991
Nolte D, Lehr H-A, Messmer K (1991). Dextran and adenosine-coupled dextran reduce post-ischemic leukocyte adherence in postcapillary venules of the hamster Progress in Applied Microcirculation 18:103

Steinbauer 1996
Steinbauer M, Harris A, Hoffman T, Messmer K (1996). Pharmacological effects of dextran on the postischemic leukocyte – endothelial interaction. In Messmer K (ed) Compromised Perfusion, Progress in Applied Microcirculation. 22: 114 -125.

Werner 1996
Werner J, Scmidt J, Gebhard MM et al, (1996). Superiority of dextran compared to other colloids and crystalloids in inhibiting the leukocyte-endothelium interaction in experimental necrotizing pancreatitis. Langenbecks Archives of Surgery (supplement 1): 467-470.

Shasby 1983
Shasby DM, Shasby SS, Peach MJ (1983). Granulocytes and PMA increase permeability to albumin of cultured endothelial monolayers and isolated perfused lungs. American Review of Respiratory Disease 127 : 72-76.

Modig 1988
Modig J, (1988). Comparison of dextran 70 and Ringers on pulmonary function, hemodynamics and survival in experimental septic shock. Critical Care Medicine 16: 266-271.

Arfors, Buckley 1997
Arfors, KE, Buckley PB. Pharmacological Characteristics of Artificial Colloids. Bailliere`s Clinical Anaesthesiology, H Haljamäe (ed).Vol 11, no 1, March 1997.

Recent (2016-17) Refs – with abstracts

1. Stone J P et al, 2016. Altered Immunogenicity of Donor Lungs via Removal of Passenger Leukocytes Using Ex Vivo Lung Perfusion. (Link to abstract)
2. Tatham et al, 2017. Intravascular donor monocytes play a central role in lung transplant ischaemia-reperfusion injury. (Link to abstract)
3. Iskender, 2017. Cytokine filtration modulates pulmonary metabolism and edema formation during ex vivo lung perfusion. (Link to abstract)
4. Noda, 2017. Targeting Circulating Leukocytes and Pyroptosis during Ex Vivo Lung Perfusion Improves Lung Preservation. (Link to abstract)