The factors involved in the induction of neutrophil gelatinase-associated lipocalin overexpression in renal tubular epithelial cells under endoplasmic reticulum stress.

Abstract

Our previous work found that neutrophil gelatinase-associated lipocalin (NGAL) expression increases when endoplasmic reticulum stress (ERS) occurs in human kidney-2 (HK-2) tubular epithelial cells. However, the reason for this is not yet known. This study investigated the factors involved when inducing NGAL overexpression in HK-2 cells during ERS. The cells were divided into six groups: the control group (normal HK-2 cells), the ERS group (HK-2 cells cultured in complete medium with thapsigargin (TG)), the transfection group (HK-2 cells transfected with activating transcription factor 4 small interfering ribonucleic acid (ATF4 siRNA), the ERS after transfection group (HK-2 cells transfected with ATF4 siRNA, then cultured in complete medium with TG), the negative control group (HK-2 cells transfected with siRNA-negative contrast), and the dimethyl sulfoxide (DMSO) group (HK-2 cells cultured in complete medium with DMSO). Western blot and a real-time polymerase chain reaction were used to measure the expression of protein and messenger ribonucleic acid (mRNA). As a result NGAL, ATF4, C/EBP homologous protein, glucose-regulated protein 78 kDa, ATF4 mRNA, and NGAL mRNA were clearly overexpressed in the ERS group compared with the control group (p < 0.05). The expression of NGAL and ATF4 were similar in the control group, the negative control group, and the DMSO group (p > 0.05). Meanwhile, ATF4, NGAL, ATF4 mRNA, and NGAL mRNA in the ERS after transfection group were significantly lower compared with the ERS group (p < 0.05), which showed that NGAL was affected by ATF4. There was a close correlation between NGAL and ATF4; when the expression of ATF4 was inhibited, NGAL was significantly lower. Therefore, ATF4 may be one of the upstream regulators of NGAL.