student research day - Case Western Reserve University School of ...

Stephanie Polites
CD55 and CD59 Expression in Normal and Diseased Cardiac Tissue
Stephanie Polites and Nikolai Sopko, David Xu, G.V. Gonzalez-Stawinski, MD and G.V. Gonzalez-Stawinski, MD
Department of Thoracic and Cardiovascular Sugery
The Cleveland Clinic Foundation
Introduction: Antibody-mediated rejection (AMR) following cardiac transplantation is associated with graft failure and
decreased lifespan due to complement fixation and subsequent vasculopathy. Circulating graft tissue alloantibodies cause
fixation of complement, however recent studies suggest that decay accelerating factor (CD55) and membrane inhibitor of
reactive lysis (CD59) may act as innate immunoprotective factors countering the damage of AMR. Little is known about the
expression profile of CD55 and CD59 within cardiac tissue or whether significant quantitative differences of their expression
can exist between individuals to allow for their theoretical role in influencing susceptibility to AMR damage. The aim of this
study is to describe CD55 and CD59 expression in cardiac tissue under physiologically normal and pathologic conditions so
that future studies can effectively analyze the natural history of their expression in transplanted hearts.
Methods: Endocardial biopsy specimens were obtained peri-mortem and flash frozen in liquid nitrogen. Cardiac tissue was
collected from both healthy donor hearts (not transplanted due to reasons unrelated to cardiac function) and diseased hearts
showing histological evidence of myocarditis. Following CD55 and CD59 mRNA isolation and reverse transcription to cDNA,
mRNA expression was measured using quantitative polymerase chain reaction (RT-PCR). Protein levels of CD55 and CD59
were measured by Western Blot. To evaluate the significance of donor origin and cardiac tissue type on the levels of CD55
and CD59, the non-parametric Kruskal-Wallis H-Test was performed. All analyses were performed in SPSS 16.0.
Results: Cardiac specimens from the left atrium, left ventricle, and interventricular septum of ten healthy donor hearts were
used in addition to specimens from the interventricular septum of five diseased hearts. Statistical testing showed no
significant difference between chambers of individual donor hearts. CD55 and CD59 expression are, respectively, consistent
across heart chambers within individual donor hearts (P>.05 for CD55 and CD59 mRNA and protein). Expression between
individuals, however, is inconsistent (Figure 2) and statistically significant differences in mRNA and protein levels of CD55
and CD59 were found from one heart to another with the exception of CD55 protein levels, which varied but did not reach
significance (P=.006 for CD55 mRNA, P=.002 for CD59 mRNA, P=.288 for CD55 protein, P=.046 for CD59 protein). We also
found that it is possible to measure CD55 and CD59 mRNA expression and protein levels in diseased hearts.
Conclusions: There is significant variability in both CD55 and CD59 levels between individual hearts. This has implications
for future studies related to protective effects of CD55 and CD59 in AMR, as quantification of relative increases or decreases
in expression will depend on individual baseline expression levels. There is not significant variability of either CD55 or CD59
across chambers of individual hearts, allowing mRNA expression and protein levels of a single endocardial biopsy to be
applied to the entire heart. This study also demonstrates that methods used to quantify CD55 and CD59 levels in healthy
hearts can be used in diseased hearts.
Support
K.R. McCurry, D.L. Kooyman and C.G. Alvarado et al., Human complement regulatory proteins protect swine-to-primate cardiac
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G.V. Gonzalez-Stawinski, C.D. Tan, N.G. Smedira, R.C. Starling, E.R. Rodriguez, Decay-accelerating Factor Expression May Provide
Immunoprotection Against Antibody-mediated Cardiac Allograft Rejection, J. Heart Lung Transplant 27 (2008) 357-361.
E.H. Hammond, R.L. Yowell and G.D. Price et al., Vascular rejection and its relationship to allograft coronary artery disease, J Heart
Lung Transplant 11 (suppl) (1992), pp. S111–S119.
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