Early epithelial phenotypic changes predict graft fibrosis

Abstract

Chronic allograft nephropathy accounts for the loss of approximately 40% of allografts at 10 yr. Currently, no biomarker is available to detect interstitial fibrosis and tubular atrophy in the renal graft at an early stage, when intervention may be beneficial. Because tubular epithelial cells have been shown to exhibit phenotypic changes suggestive of epithelial-to-mesenchymal transition, we studied whether these changes predict the progression of fibrosis in the allograft. Eighty-three kidney transplant recipients who had undergone a protocol graft biopsy at both 3 and 12 mo after transplantation were enrolled. De novo vimentin expression and translocation of beta-catenin into the cytoplasm of tubular cells were detected on the first biopsy by immunohistochemistry. Patients with expression of these markers in >or=10% of tubules at 3 mo had a higher interstitial fibrosis score at 1 yr and a greater progression of this score between 3 and 12 mo. The intensity of these phenotypic changes positively and significantly correlated with the progression of fibrosis, and multivariate analysis showed that their presence was an independent risk factor for this progression. In addition, the presence of early phenotypic changes was associated with poorer graft function 18 mo after transplantation. In conclusion, early phenotypic changes indicative of epithelial-to-mesenchymal transition predict the progression toward interstitial fibrosis in human renal allografts.

Figure 1.

Immunohistochemical staining of β-catenin and vimentin in 3-mo biopsy. (A) A regular expression of β-catenin was found at the basal surface of normal proximal tubular epithelial cells (★, tubules in which β-catenin staining is interpreted as normal, i.e., not suggestive of EPC). An upregulation and intracellular translocation of β-catenin was observed in the epithelial cells of some proximal and distal tubules, especially in flat epithelial cells: The expression near the basal membrane was reduced or absent, whereas an intense, granular and irregular staining was found in the cytoplasm (arrows indicate tubules in which this translocation is suggestive of EPC). (B) Vimentin staining was negative in the epithelial cells of normal tubules (★) but was observed in flat epithelial cells or atrophic tubules (arrow) surrounded by interstitial fibrosis. The serial sections revealed that in the epithelial cells expressing vimentin, an upregulation and intracellular translocation of β-catenin expression was present. (C) Vimentin staining was also observed in nonatrophic tubules without excessive surrounding matrix.

Figure 2.

Double staining of β-catenin and vimentin in tubular epithelial cells. *Normal tubules with basal localization of β-catenin (A) and no expression of vimentin (B); the arrow indicates a tubule with phenotypic changes: Translocation of β-catenin and de novo expression of vimentin. (C) Merged image.

Figure 3.

Score of IF/TA at 12 mo and progression of IF/TA score (ΔIF/TA) between 3 and 12 mo among patients with or without EPC at 3 mo. (A and B) Grafts with EPC at 3 mo (n = 21) had a higher IF/TA score at 12 mo (A) and had significantly more progressed toward IF/TA (B) than grafts without EPC (n = 62). *P values are based from the Mann Whitney rank test. (C and D) A linear relation is shown between the score of EPC at 3 mo and the score of IF/TA at 12 mo (C) and the progression of the IF/TA score between 3 and 12 mo (D). •, Mean values with SEM.

Figure 4.

Long-term follow-up of serum creatinine in the two groups (EMT+, EMT−). P values are indicated at each time point. The presence of EPC is associated with a slow decline of graft function with time.