Jorge Cardoso

Hyperacute xenograft rejection in the swine-to-human donor-recipient combination : in vitro analysis of complement activation

Complement activation is central to the rejection of discordant xenografts. In order to assess the respective roles of direct and alternative pathways, an in vitro model of hyperacute rejection in the swine-to-human donor-recipient combination was designed, using a complement-dependent cytotoxicity test with swine endothelial cells in culture as targets, and fresh human serum as the source of xenogeneic antibodies and complement. The cytotoxic activity of the sera was evaluated by a colorimetric assay using (3-[4,5-dimethyldiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT). Pure human serum lysed 58 +/- 5% of swine endothelial cells. Selective inhibition of the direct pathway by adding EGTA to the serum reduced cytolysis to 51 +/- 2% (P < 0.01 versus normal serum). Similarly, when using C1q-deficient human sera, only 37 +/- 7% of swine endothelial cells were killed (P < 0.001 versus normal serum). When the alternative pathway was selectively inhibited by heating for 20 min at 50 degrees C, the lytic activity of human serum dropped to 42 +/- 5% (P < 0.001 versus normal serum). Factor B-deficient human serum could only lyse 42 +/- 10% of porcine endothelial cells (P < 0.001 versus normal serum). Syngeneic normal swine serum and heat-inactivated serum were not cytotoxic. Mixing serum with deficient direct pathway and serum with deficient alternative pathway restored the cytotoxicity to normal levels. Similarly, the cytotoxic activity of deficient serum supplemented with purified C1q or factor B at physiological concentrations reached that of normal human serum. In this model of in vitro hyperacute rejection, both pathways of complement activation are involved, suggesting that regimens designed to inhibit hyperacute rejection of swine xenografts into humans should take into account the dual activation of complement in this donor-recipient combination.

Organ xenografting between rodents: an evolutionary perspective

Rejection times of heart xenografts in several donor-recipient combinations including the guinea pig, rat, hamster, and mouse are examined in light of the paleontological history of rodents and the resulting phylogenetic distances between taxa. This multidisciplinary review at the molecular, chromosomal and morphological levels suggests that xenograft rejection time is inversely proportional to the time divergence or phylogenetic distance, and that the binomial terminology concordant/discordant does not reflect the amplitude of phylogenetic distances.

Target antigens of hyperacute xenogeneic rejection in the rat to guinea pig and guinea pig to rat discordant combinations

The increasing shortage in allografts has led to a renewed interest in xenogeneic transplantation. Discordant combinations are characterized by hyperacute rejection partly due to the presence of natural antixenogeneic antibodies in the recipient. The aim of this work was to characterize the target antigens, using 2 discordant models. In the rat into guinea pig model, analysis of organ homogenates by immunoblotting revealed numerous bands. Some of these bands were organ specific, whereas others, namely in the 55-kDa region, were detected in liver, heart, lung, and kidney. Using membrane extracts of liver cells or of aortic endothelial cells, only bands of 55 kDa were revealed. No band could be seen using extracts of isolated hepatocytes. Two bands of 55 kDa disappeared after preabsorption of guinea pig sera on the various rat tissue homogenates, suggesting that they represent xenoantigens common to these tissues. In order to investigate the in vivo relevance of these 55-kDa antigens, isolated rat livers were perfused with decomplemented guinea pig sera. Eluates revealed one single print of 55 kDa on rat tissue homogenates. Finally, preincubation of rat mononuclear cells with various xenogeneic sera did not inhibit the binding of mAb specific for rat class I or class II MHC antigens, suggesting that the latter are not recognized by natural xenoantibodies. In the guinea pig to rat model, the antigens detected had a molecular mass ranging from 95 to 110 kDa. Absorption and perfusion experiments also showed that these antigens were common to various tissues and involved in the binding of rat natural antibodies ex vivo. In conclusion, our results indicate that rat xenoantigens of about 55 kDa are recognized by guinea pig natural antibodies, while guinea pig xenoantigens of 95–110 kDa are bound by rat natural antibodies. These antigens are common to liver, heart, lung, and kidney, are borne by endothelial cells, and cannot be found on hepatocytes.

Hepatitis E–associated subacute liver failure: a rare indication for liver transplantation

blood pressure (digital photopletysmography [Finapres, Ohmeda, Tewksbury, MA]) and heart rate (surface electrocardiography) were monitored continuously. At 40 cpm on insufflation, the patient became profoundly hypotensive in association with a persistent bradycardia of ,30 bpm requiring atropine. The procedure reproduced syncopal symptoms. Because of the persistent bradycardia during symptom reproduction this woman had a pacemaker inserted. Six months after dual chamber physiological pacemaker implantation (Kappa 700, Medtronic, Minneapolis, MN) she has had no further episodes of syncope. This patient had symptom reproduction with a hypotensive response and a profound bradycardia during colonoscopy; in addition, there has been clear benefit from pacing intervention, suggesting a diagnosis of cardioinhibitory situational vasovagal syncope. We were able to detect these subtle changes because we used a real-time beat by beat method of recording blood pressure, combined with continuous measurement of heart rate. To our knowledge this is the first reported case of defecation syncope where hemodynamic changes have been documented in real time, where an abnormality of autonomic function has been demonstrated, and where treatment (in this case, permanent pacemaker implantation) has proved curative.