Introduction: Among the complex cellular interplay engaged in renal rejection, recent transcriptomic analysis of renal biopsies has detected NK cell transcripts in the setting of antibody-mediated rejection (ABMR). The precise in situ detection of NK cells in renal biopsies supporting this molecular signature remains to be done.

Methods: Formalin fixed paraffin embedded sections from 30 renal biopsies [5 without rejection; 12 with T-cell-mediated rejection (TCMR); 13 with ABMR] was used for immunoperoxidase staining of NK cells. Labeled NK cells in renal cortex were counted and their intravascular (intraglomerular/ intraperitubular capillaries) and interstitial location were determined. The surface area of renal cortex was measured allowing the calculation of NK cell density. To determine the precise intravascular/interstitial location of NK cells and their relationship with T lymphocytes and macrophages, multiplex immunofluorescence labeling (kit Opal,Perkin Elmer[copy]) was used with NK antibody, CD3, CD163 and CD34 for NK cells, T lymphocytes, macrophages and endothelial cells respectively. The sections were observed with Vectra imaging system and analyzed with inForm software[copy].

Results: The density of total NK cells was very low in non-rejecting renal biopsies (mean 0.51/mm²) compared to ABMR and TCMR renal biopsies (mean 13.40/ mm² and 22.51/mm² respectively; p<0.01). ). The density of interstitial NK cells was increased in TCMR vs. ABMR but was not statistically significant (mean 19.41/mm² vs. 7.26/ mm² respectively). The ratio of intravascular to interstitial NK cells was increased in ABMR vs. TCMR (mean 2.06 vs. 0.27 respectively; p<0.001) whereas the ratio of interstitial to total NK cells was increased in TCMR vs. ABMR (mean 0.82 vs. 0.44 respectively; p<0.001). The preliminary results of multiplex immunofluorescence demonstrated clearly both intravascular and interstitial NK cells and their close relationship with T lymphocytes and macrophages.

Conclusion: NK cells are recruited in both ABMR and TCMR. Their compartmentalization differs between TCMR and ABMR. NK cells colocalize with T lymphocytes and appear as a cell type engaged in renal rejection.

CITATION INFORMATION: Terada M., Duong Van Huyen J., Drieux F., Legendre C., Bruneval P., Rabant M. NK Cells in Allograft Kidney Biopsies during Rejection: Detection, Quantification, and Precise Localization Using Multiplex Immunofluorescence Am J Transplant. 2017;17 (suppl 3).

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