Biochemical and Structural Evidence for Pig Myocardium Adherens Junction Disruption by Cardiopulmonary Bypass

Abstract

Background-Given that cardiopulmonary bypass (CPB) is associated with edema and heart dysfunction and that adherens junctions may regulate vascular permeability barrier integrity and cardiomyocyte function, we investigated adherens junction protein steady-state levels in a pig model of CPB. Methods and Results-Pigs were subjected to normothermic CPB for 90 minutes, followed by post-CPB perfusion for 90 minutes. Atrial and ventricular myocardium tissue samples were harvested before institution of bypass (basal levels) and at the end of post-CPB perfusion. Adherens junctions were analyzed by either total lysate or cadherin immunoprecipitates that were immunoblotted for pan-cadherin, VE-cadherin, ␤-catenin, and ␥-catenin. Adherens junction solubility was addressed with Triton X-100 extraction. Frozen tissue sections were labeled with the same antibodies, and adherens junctions were visualized by confocal microscopy. Immunoblotting of total lysates revealed an increase in smallermolecular-weight fragments of VE-cadherin, ␤-catenin, and ␥ -catenin after post-CPB perfusion, indicating partial protein degradation. Smaller-molecular-weight fragments recognized by VE-cadherin and ␤-catenin antibodies were also obtained from VE-cadherin immunoprecipitation, indicating degradation of endothelial cell adherens junctions. A prominent increase in adherens junction complex solubility was observed in post-CPB perfusion samples. Confocal microscopy of hearts obtained before CPB showed a continuous, homogeneous pattern of cell-cell labeling that contrasted with an irregular, discontinuous, punctuate, or zigzag pattern observed in post-CPB perfusion samples, corroborating biochemical data. Conclusions-These results indicate that CPB is associated with signs of degradation of endothelial and cardiomyocytes adherens junctions, pointing to a molecular mechanism leading to increased vascular permeability and cardiomyocyte dysfunction. (Circulation. 2001;104[suppl I]:I-319-I-324.)

Injury of the renal microvascular endothelium alters barrier function after ischemia

“…2). Whereas VE-cadherin immunostaining was not limited to intracellular endothelial contacts, it was similar to in vivo VE-cadherin immunostaining observed in other studies (4,8). After 32 min of ischemia, VE-cadherin staining remained in the renal microvasculature.…”

“…Increased endothelial permeability in these models has been associated with internalization of VE-cadherin from endothelial adherens junctions (2,21). Although we did not observe internalization of VE-cadherin after ischemia in vivo, other mechanisms including cleavage of VE-cadherin as described in a previous in vivo study (4) may account for the diminished VE-cadherin staining observed in our study.…”

Cyclin A2 Mediates Cardiomyocyte Mitosis in the Postmitotic Myocardium

“…Only cells with clearly delineated borders were measured. Cell lengths were measured utilizing the same program after immunostaining for pancadherin (Sigma), a structural protein found in the intercalated discs (22), again at ϫ40 magnification. Only those myocytes (Ͼ200 cells/ heart) that could be visualized end-to-end were assessed.…”

Actin cytoskeleton in ischemic acute renal failure

“…Recent studies indicate a central role for microvascular injury with attendant disruption of the endothelial cell actin cytoskeleton in the pathophysiology of ischemic ARF [51][52][53]. In particular, endothelial cell injury, including cell swelling, altered cell-cell attachment, and reduced endothelial cell-basement membrane attachment are some of the alterations that have been observed following ischemic injury [52,54].…”