Acute Rejection From Infection In A Rat Heart Transplant

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Acute cardiac allograft rejection and infection remain significant sources of morbidity and mortality after heart transplantation, accounting for nearly 50% of reported deaths. It is often difficult to clinically distinguish between rejection and infection because they are both inflammatory processes with similar, nonspecific symptoms. However, this differential is essential for determining therapy. Identifying laboratory methods that will permit safe and concise early differentiation between rejection and infection in the transplant patient will improve outcome substantially. The primary goal of this study is to determine whether gene microarray analysis of peripheral blood mononuclear cells (PBMC) will reliably differentiate acute heart rejection from infection in the transplanted rat. Our protocol will combine two well-established rat models, the first is a heterotopic heart transplantation model and the second is an E. coli pulmonary infection model. The study is divided into 2 stages In the first stage of the study, cardiac transplant techniques in the rat will be refined and the optimal doses and duration of both cyclosporin (CSA) therapy and E. coli bacterial infection will be determined. In our first pilot study, we will establish a dose and duration of CSA therapy in this model that reliably suppresses rejection during its administration, but will permit the emergence of rejection upon its discontinuation . Our second pilot study will establish a dose and duration of intrabronchial E. coli pneumonia that is sufficient to cause a systemic inflammatory response without being immediately lethal in transplanted rats receiving CSA. The second stage of our study will determine whether gene microarray analysis of PBMCs is capable of differentiating cardiac rejection from infection. In this stage all rats will undergo heart transplantation on day 0 in conjunction with daily CSA (dose determined in Stage 1) to suppress rejection. After transplant, animals will be randomized (timing determined in stage 1) to have CSA discontinued, in order to initiate rejection, or continued, in order to further suppress rejection. After discontinuing CSA the animals will again be randomized (timing determined in stage 1) to receive intrabronchial E. coli inoculation or saline inoculation. Consequently, four groups will be studied: No rejection (i.e. receiving CSA) without infection, no rejection (i.e. receiving CSA) with infection, rejection (i.e. not receiving CSA) without infection, and rejection (i.e. not receiving CSA) with infection. At the end of the study, all animals will be sacrificed and the blood and heart removed for gene microarray analysis. Other analytic tools that may be employed include: RT-PCR, western blot, in-situ hybridization, proteomics, and immunohistochemistry, and histopathology. In addition, the animals? lungs, spleen, and liver will also be procured in the primary study and preserved for potential future analysis.