Despite reports suggesting that tissue-resident natural killer (trNK) cells cause ischemic kidney injury, their contribution to the development of tubulointerstitial fibrosis has not been determined. This study hypothesized that depletion of trNK cells may ameliorate renal fibrosis by affecting transglutaminase 2/syndecan-4 interactions. C57BL/6 mice treated with anti-asialo GM1 (ASGM1) or NK1.1 antibody were subjected to aristolochic acid nephropathy (AAN) as an experimental model of kidney fibrosis. Although both ASGM1 and NK1.1 antibody suppressed renal NKp46+DX5+ NK cells, renal NKp46+DX5− cells were resistant to suppression by ASGM1 or NK1.1 antibody during the development of tubulointerstitial fibrosis in the AAN model. Western blot analysis showed that both antibodies increased the expression of fibronectin, transglutaminase 2 and syndecan-4. These findings indicate that trNK cells play an excerbative role in tubulointerstitial fibrosis by activating transglutaminase 2 and syndecan-4 in the AAN model.