estingly, matrix metalloprotease genes, were involved in all three of the top identified pathways at week 1, indicating that the MMP9 and MMP8 are likely important in kidney transplants without rejection. 4 / 14 Differentially Expressed Genes after Kidney Transplant 5 / 14 Differentially Expressed Genes after Kidney Transplant RNA sequencing reads were aligned to human genome via Tophat2. doi:10.1371/journal.pone.0125045.t002 Three months post-transplant. Compared to 1 week post-transplant, similar genes were altered at month 3. However, the number of DEGs at month 3 was substantially lower than at 1 week with 94 genes with increased levels compared to baseline and 174 genes with decreased levels. T-cell receptor signaling remained the top pathway altered with 16 involved genes. However, the fold changes in gene expression were much lower than at week 1. For instance, the CD3D gene was -14.39 fold change at week 1 compared to -4.65 fold change at month 3. This suggests that the expression of some of the genes in the early affected pathways may be gradually returning to pre-transplant expression levels. The second pathway with lower levels than baseline was iCOS-iCOSL signaling in T-helper cells with 16 genes involved. The upregulated pathways at month 3 were slightly different than compared to week 1. The expression pattern indicates that the complement system was activated as well as IL-8 signaling. The genes MMP8 and MMP9 remained at higher levels at month 3 compared to baseline. Interestingly, pathway analysis also identified both the granulocyte LY-411575 adhesion and diapedesis and the agranulocyte adhesion and diapedesis pathways at month 3 among genes with higher levels compared to baseline. This suggests that perhaps by month 3, leukocytes PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19776696 were leaving the blood vessels and potentially extravasating into kidney allograft as part of the immune response. Six months post-transplant. By 6 months post-transplant, the numbers of DEGs compared to baseline were severely reduced compared to week 1 or month 3 post-transplant. At month 6, only 87 genes were differentially expressed compared to baseline with 50 at higher levels and 37 at lower levels. Remarkably, at month 6, the top pathways have changed and now granulocyte adhesion and diapedesis and the agranulocyte adhesion and diapedesis pathways were among genes with lower 
levels compared to baseline. This pathway PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19778700 was primarily composed of chemokines genes. Gene expression changes over time in representative genes are shown in Fig 1. Differentially Expressed Genes after Kidney Transplant Fig 1. Expression of representative genes over time in kidney transplant recipients. Representative time series of fold expression changes relative to baseline of some top genes with higher and lower level compared to baseline. Each line on each graph represents the expression of the particular gene in a separate kidney allograft recipient. Note that all patients do not have data all the time points. CD3E = CD3 Epsilon TCR complex; CD3D = CD3 Delta TCR complex; MMP8 = Matrix Metallopeptidase 8; IL7R = Interleukin 7 Receptor; OLFM4 = Olfactomedin 4; KLRC3 = Killer Cell Lectin-like Receptor subfamily C, member 3. For example the granulocyte and agranulocyte adhesion, and diapedesis pathways were at higher levels at month 3 and then at lower levels at month 6. This was unusual in our study where a pathway was at higher levels after transplant compared to baseline and then at lower levels at the next time point. Th