|Surgical organ perfusion method for somatic gene transfer: An experimental study on gene transfer into the kidney, spleen, lung and mammary gland|
The present experimental work resulted in the development of a novel perfusion-based method for targeted gene transfer into different organs. The other main findings of the study can be summarized as follows:
Adenovirus-mediated gene transfer does not result in remarkable gene expression in the target organ when single in vivo intra-arterial infusion of vectors is used or pharmacological vasodilative adjuvant agents are administered in conjunction with the viral infusion.
By prolonging the incubation time with the adenoviral vectors and the target kidney cells up to 2 hours at body temperature, efficient gene transfer can be accomplished into renal glomerular cells. Organ-specific, efficient and relatively selective in vivo glomerular gene transfer is possible by using the closed-circuit warm organ perfusion method developed in this study.
Apart from gene transfer into the renal glomeruli, the surgical closed-circuit warm organ perfusion method for gene transfer is feasible to be used for gene transfer into the spleen, lungs and mammary gland. In the spleen, adenovirus-mediated gene transfer by the perfusion method yields gene transfer into the cells of the perifollicular area, mostly macrophages and endothelial cells. In the lungs, this method results in the transfection of alveolar epithelial cells (type I and II pneumocytes), bronchiolar epithelial cells, alveolar macrophages and endothelial cells of arterioles. The transfection efficiency obtained by the perfusion method is lower in the spleen and lung than in the kidney. For exogenous protein secretion into milk, retrovirus-mediated gene transfer by perfusion is feasible, but yields very low expression of the transgene.
The closed-circuit organ perfusion method is invasive, as it requires general anesthesia and major surgery. After the operation itself, however, the perfusion procedure is relatively well tolerated and does not give rise to clinical signs of infection or inflammation in the test animals. In the kidney and lungs, signs of a cytotoxic immunological response, evidently against the adenoviral vector, were present after adenoviral gene transfer by the perfusion method. This adenoviral gene transfer method results in slight macroscopic alterations in the kidney and lungs, but none in the spleen. Retroviral gene transfer by perfusion into mammary gland is well tolerated.