5.4. Tissue and systemic effects of gene transfer by the closed-circuit organ perfusion method

Peroperative mortality in the kidney, spleen and lung perfusion experiments was 24%, 20% and 27%, respectively. All deaths were on the table and they were due to sudden cardiac arrhythmias of unknown reason, usually at the beginning of the operation. As a whole, the animals seemed to tolerate the perfusion experiments well after the operation itself. They were able to drink water on the operation day and eat on the day after the operation. They moved and breathed without help on the next day and urinated normally. No clinical signs of inflammation or infection were observed.

During the kidney perfusion, only a small fraction of the adenoviral vectors (< 0.1%) were detected in the urine flowing out of the ureter. Four weeks after the kidney perfusion experiments, no remarkable titers could be measured for serum anti‐β -galactosidase antibodies. Only one animal showed slight elevation of the IgG‐type anti‐AdV antibodies, the others being negative. The serum creatinine levels were within the normal reference range. Kidney function was followed up on excretory urograms four weeks after the perfusion (Fig. 14). Macroscopically, the viral-perfused kidney was of the same size as the contralateral kidney, but the cut surface was slightly paler than that of the contralateral kidney (Fig. 15). The perfused control kidney was macroscopically normal. In histological analysis, the control perfusion appeared to have no effect on cell morphology, but following the viral perfusion, there were inflammatory changes, such as mononuclear cell clusters mainly around blood vessels, on the fourth day after the perfusion already (Fig. 16). Additionally, after four weeks, the viral-perfused kidneys also showed some signs of tubular necrosis, such as reduction of the epithelial brush border in the tubules in restricted areas.

The macroscopic appearance of the perfused spleen was unaltered four days after the operation. No remarkable cytotoxic or inflammatory alterations in splenic tissue were seen. There were no inflammatory cell clusters of the kind found previously in the kidney after kidney perfusion experiments. The specimens of perfused spleen were also compared with a normal spleen from a non-perfused animal.

The macroscopic appearance of the lung lobe was slightly darkish compared to the other lobes seven days after the operation. The possible toxic and cytopathic effects were examined in paraffin-embedded sections after HE, PAS and some other special stainings (see Material and methods). The special stainings showed only some scant chronic inflammatory infiltrations and intra-alveolar edema in most cases and alveolar hemorrhage in one case, but neither signs of diffuse alveolar damage nor hypertrophy of pulmonary muscular arterioles, excluding pulmonary hypertension during perfusion.

The goats tolerated the perfusion of the mammary gland well. Their milk secretion was normal. One of the treated goats gave birth to a kid more than a year after the perfusion, and she had normal lactation in the previously treated mammary gland.