S FERNANDO 1,2, W MULLEY 1,2, D NIKOLIC-PATERSON D1,2, F MA 1,2
1Department of Nephrology Monash Health , Clayton , Australia, 2Monash University Department of Medicine, Clayton, Australia
Aim: To investigate the contribution of JUN in the inflammatory and fibrotic responses of renal tubular epithelial cells.
Background: JUN is one component of the AP-1 transcription factor that transcribes genes in the inflammatory and fibrotic responses. Although not detected under homeostatic conditions, JUN is prominently expressed in tubules in both acute and chronic kidney injury; however, the precise role of JUN in kidney disease is unknown.
Method:Mouse primary tubular epithelial cells were cultured using standard conditions. Cells were cultured for 4 days with 50nM of a locked nucleic acid anti-sense Jun oligonucleotide (LNA-JUN), or control oligonucleotide (LNA-Ctl). Cells were then stimulated with 10ng/mL Tumor necrosis factor (TNF) for 6hr or with 5ng/mL Transforming growth factor-b1 (TGF-b1) for 24hr and analysed by real-time polymerase chain reaction (RT-PCR). Cell proliferation was induced by 10ng/mL Epidermal growth factor (EGF) for 48hr.
Results:Cells treated with LNA-JUN showed a 3-fold knockdown of JUN protein levels by western blotting. TNF stimulation induced 15-fold and 10-fold increases in CCL-2 and IL-36A mRNA levels (markers of inflammation) in LNA-Ctl treated cells; this was reduced by 67% and 50% respectively, with LNA-JUN treatment (both p<0.0001). TGF-b1 stimulation of LNA-Ctl treated cells induced 2-fold and 2.3-fold increases in TGF-b1 and collagen IV mRNA levels (markers of fibrosis), respectively; this was reduced by 100% and 77% respectively in LNA-JUN treated cells (both p<0.001). By contrast, EGF-stimulation of epithelial cell proliferation was not affected by LNA-JUN treatment.
Conclusion: JUN/AP1 plays a key role in the transcription of both pro-inflammatory and pro-fibrotic molecules in mouse tubular epithelial cells, without affected cell proliferation. These findings identify JUN as a potential therapeutic target in chronic kidney disease.
Sanduni is a Melbourne trained nephrologist (admitted to FRACP Dec 2019) who is currently completing her PhD at Monash University in the area of Chronic kidney disease and fibrosis at the clinical school based at Monash Health. She also works part time as a Fellow at Monash Health and has an interest in chronic kidney disease, renal transplantation and medical education.