Oxygen free radical formation in macrophages of bronchoalveolar lavage and monocytes of the pulmonary artery and vein was assessed in terms of the pathogenesis of rejection with or without pneumonia on lung allografts on the condition of the use of cyclosporine and azathioprine. It was clearly observed that oxygen free radicals are activated after transplantation. However, the immunosuppressive drugs of cyclosporine and azathioprione individually revealed a different behavior for the activation of oxygen free radicals. There was not close correlation between the activation of oxygen free radicals and occurrence of rejection with or without pneumonia

Akoskin Santiag

Ayabe Hiroyoshi

Hisano Hiroshi

Honjou Seiji

Ikari Hideki

Kawahara Katsunobu

Takahashi Takao

Tomita Masao

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Acta Med. Nagasaki 35:221-224Graft Rejection in Tems of Oxygen Free RadicalsMasao TOMITA, Hiroyoshi AYABE, Katsunobu KAWAHARA   Takao TAKAHASHI, Seiji HONJOU, Hiroshi HISANO         Hideki IKARI and Santiago AKOSKINFirst Department of Surgery, Nagasaki University School of Medicine,                       Nagasaki, Japan.Received for publication, June 25, 1990ABSTRACT : Oxygen free radical formation in macrophages of bronchoalveolar lavage and monocytes of the pulmonary artery and vein was assessed in terms of the pathogenesis of rejection with or without pneumonia on lung allografts on the condition of the use of cyclosporine and azathioprine.  It was clearly observed that oxygen free radicals are activated after transplantation. However, the immunosuppressive drugs of cyclosporine and azathioprione individually revealed a different behavior for the activation of oxygen free radicals. There was not close correlation between the activation of oxygen free radicals and occurrence of rejection with or without pneumonia.       INTRODUCTION  Recent studies have been elucidated a role of oxygen free radicals which plays a key role in developing reperfusion injury, in particular, in a ischemic state.  Needless to say, it is important to prevent a donor graft from the injury associated with restarting circulation at ischemic state. For this purpose, much research work focuses on reduc-ing oxygen free radical formation.  It is of great concern as to whether the activation of immunosuppressive drugs ralates to oxygen free radical formation or not.  The purpose of this study is to clarify changes in oxygen free radical formation in accordance with the appearance of rejection phenomenon and/or pneumonia on lung allogafts in variety of the use of cyclosporine (CsA) and azathi-oprine (AZA).   MATERIALS AND METHODS  A pair of mongrel dogs with almost the same body weigt each other were used for left lung allotransplantation. Left atrial cuff anastomosis was applied for the anastomosis of the pul-monary vein. The transplantation techniques were standarized and kept uniformity in our institute. The dogs with obvious technical failure such as vascular and bronchial strictures at anastomosis were excluded from this study by postperative angiographic and bronchoscopic evaluations. Cyclosporine (20mg/kg/day) in Group I and azathioprine (5mg/kg/day) in Group II were given as postoperative immunosup-pressive drugs in comparison with the group with reimplantation (autotransplantation).  The function of transplanted donor graft was evaluated by the degrees of aeration on daily chest X-ray film and by the help of perfusion scintigram. After transplantation, macrophages in bronchoalveolar lavage and monocytes in the pulmonary artery and vein were collected every5 days.  Oxygen free radical formation was measured according to Bass method') by using DCFHDA which is converted to DCFH when taken into monocytes and subsequently converted to DCF by oxygen free radicals. DCF was assessed by using Flow cytometric method under stimula-tion by PMA.  At the time when dogs expired, autopsy clarified a presence of rejection and/or pneu-monia on a donor graft. The presence of rejec-tion was determind by the histologic findings of perivascular cuffing.          RESULTS  Fig. 1 showed oxygen free radical formation in BALF in accordance with a presence of rejeciton phenomenon with or without pneu-monia. Oxygen free radical formation after transplantation was facilitated in both groups as compared with that before transplantion. Its tendency had become much more significant at rejection. However, when pneumonia coexisted, oxygen free radical formation was depressed in Group II in contrast with still enhancement of oxygen free radical formation in Group I.  Oxygen free radical formation in monocytesin the pulmonary artery and vein were shown in Fig. 2 and 3. These were kept high after transplantation as compared with those prior to transplantation. Furthermore, when rejection occurred, oxygen free radical formation in the cases with the use of cyclosporine was inhibited, in particular, in macrophages of the pulmonary vein. In the case with pneumonia, there were kept high, however in particular, much less in Group I.         DISCUSSION  Free radical formation now focuses on the pathogenesis of tissue damage ralated to ischemia with subsequent reperfusion. It is well known that free radical plays an important role in reperfusion injury to ischemic organs with subsequent reperfusion.  In this study, changes in oxygen free radical formation in lung allografts were carefully observed in macrophages of the pulmonary artery and vein and bronchioalveolar space in accordance with occurrence of rejection and pneumonia, identifying the difference between the effects of cyclosporine and azathioprine. When prescribing for cyclosporine from azathioprine the responses of free radicalFig. 1. Oxygen free radicals production in neutrophiles of PA in comparison with       rejection and pneumoniaFig. 2. Oxygen free radicals production in neutrophiles of PV in com.       narison with resection and pneumoniaFig. 3. Oxygen free radicals production in neutrophiles of BALF in       comparison with rejection andpneumoniaformation in macrophages to rejection and/or pneumonia were different.  Until recently, many reports indicate that oxygen free radicals play a major role in theinjury associated with ischemia2-5). The ex-planation is that adenosine triphosphate in ischemic tissues breaks down to hypoxanthine and xanthin dehydrogenase in converted toxanthine oxidase6). With the perfusion molecu-lar oxygen produces a burst of superoxide anion, hydrogen peroxide and hydroxyl radical. It is these oxygen radicals that causes tissue dam-age7 .  In this study, it was clarifiend as to whether oxygen free radicals relate to tissue damage association with rejection and/of pneumonia to a donor graft according to the kinds of immuno-suppressive drugs used. This study indicates that the activation of oxygen free radicals vary with rejection and pneuimonia in accordance with the use of cyclosporine and azathioprine. It should be emhasized that cyclosporine some-what inhibits activated oxygen free radicals at rejection. Whereas azathioprine did not suppress the actipon of oxygen free radicals at rehection.  Moreover, it is important to stress that oxygen free radicals in BALF are inhibited by using azathioprine when pneumonia is occurring.  On the contrary, these are not true with the use of cyclosporuine. It is concluded that macrophages in BALF do not act as a direct inducer of pneumonic and rejection in view of activated oxygen. free radicals. The beneficial effects of cyclosporine were observed on lung transplantation.  It is accepted that the lung is most susceptible to oxygen toxicity because of its direct exposure to inspired oxygen8'9). Oxygen therapy has been clearly beneficial in major surgery. However surgeons should be aware of inverse toxic effect. The conclusion drawn from this study is that there is not close relationship in the pathogenisis between activated oxygen free radicals and occurrence of rejection with or without pneumonia.     ACKNOWLEDGEMENT  The authors thank to the experimental dog supply from the Labolatory Animal Center for Biomedical Research of Nagasaki University School of Medicine.          REFERENCES 1) Bassda, Parce WJ, Dechatelet LR et al: Flow    cytometric studies of oxydative product forma-   tion by neutrophils A graded response of mem-   brane stimulation. I Immunology 130: 1910-1917,   1983. 2) Stewart JR, Blackwell WH, Crute et al:    Inhibition of surgically induced ischemia/   reperfusion injury by oxygen free radical scav-   engers. I Thorac. Cardiovasc. Surg. 86:262-272,   1983. 3) Shlefer M, Kane PF, Kirsch MM : Superoxide    dismutase plus catalase enhances the efficacy    of hypothermic cardioplegia to protect the    globally ischemic reperfused heart. J. Thorac.    Carciovas. Surg, 83:830-839, 1982. 4) Perry MA, Wadhwa S, Parks PA, Pickard W et    al: Role of oxygen radicals in ischemia -   induced lesions in the cat stomach. Gastro-   enterolgy 90: 362-367, 1986. 5) IM MJ, Shen WH, Pak CJ, Manson PN, Bulkley    GB : Effect of allopurinol on survival of hyeremic    island skin flaps. Plast Reconstr. Surg. 73: 276-   278, 1984. 6) Hansson R, Gustafasson B, Jonsson 0 et al:    Effect of xanthine oxydase inhibition on renal    circulation after ischemia. Tranepl Proc, 14: 51-   58, 1982. 7) McCord JM : The superoxide free radical : Its    biochemistry and pathophysiology Surgery 94:    412-412, 1988. 8) Gregory EM, Fridovich I: Oxygen toxicity and    the superoxide dismutase J. Bacteriology 114:    1193-1197, 1973. 9) Deneke SM, Fanburg BI: Normobaric oxygen    toxicity of the lung. New Eng. J. Med. 303: 76-86,   1980.

Graft Rejection in Tems of Oxygen Free Radicals

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Graft Rejection in Tems of Oxygen Free Radicals

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