EP1 Receptor Antagonism Mitigates Early and Late-Stage Renal Fibrosis

Background: Lupus nephritis (LN) is caused by autoimmune responses and is a significant driver of end-stage renal disease in systemic lupus erythematosus patients. Complement activation, pro-inflammatory cytokine production, and the influx of macrophages have all been implicated in LN pathogenesis. The anaphylatoxin complement 5a (C5a) receptor 1 (C5aR) is a major driver of the pro-inflammatory functions of complement activation. We examined C5aR’s expression in kidney in lupus nephritis and investigated its role in controlling pro-fibrotic functions of macrophages. Methods: C5aR expression, infiltrating immune cells, and fibrosis were examined by immunohistochemistry in LN patient kidney biopsies. M1 and M2 macrophages derived from human peripheral blood monocytes were used in in vitro assays to examine the effect of C5a stimulation and avacopan, a specific C5aR inhibitor, on the secretion of cytokines and other factors. Results: In LN kidney biopsies, large numbers of macrophages, identified by CD68 staining, were observed in areas with severe fibrosis, and expressed C5aR. In addition, C5aR was detected on distal tubules in biopsies of both normal and lupus nephritis kidneys. C5a increased the production of inflammatory cytokines TNFa and IL-6 from both M1 and M2 macrophages in vitro. Chemokines (MCP-3, MIP-1a, MIP-1b and MIP-3a), matrix metalloproteinases (MMP3 and MMP8), and pro-fibrotic growth factors (fibroblast activation protein, platelet-derived growth factor-AA) were strongly increased in M2 macrophages with C5a stimulation, and these increases were blocked by the C5aR inhibitor avacopan. Conclusions: C5aR activation induced macrophage secretion of factors that are known to drive inflammation, fibroblast activation and tissue fibrosis, and thus may contribute to LN disease progression. Inhibiting C5aR activity with avacopan blocks these pathological changes, and may provide therapeutic benefit to LN patients.

Background: Meta-analyses reveal show a significant association of chronic kidney disease (CKD) with severe COVID-19.The double stranded RNA virus SARS-CoV-2 can evoke a damaging inflammatory response.To understand the mechanism for the greater severity of the disease in patients with CKD, we studied an animal model of CKD exposed to polyinosinic-polycytidylic acid [poly(I:C), a synthetic analog of double-stranded RNA that recapitulates the innate immune response provoked by SARS-CoV-2].
Methods: C57Bl6j mice were injected with 2 doses of cisplatin at 15 mg / kg or vehicle control subcutaneously, 2 weeks apart.After CKD established, control and CKD mice were subsequently injected with poly(I:C) at 30 mg/kg intravenously and monitored for body weight loss and mortality.Bone marrow cells were isolated 2 weeks post poly(I:C) treatment and grown in serum-free medium supplemented with macrophage colony stimulation factor for 7 days to obtain bone marrow derived macrophages (BMDM).These cells were stimulated with 10 ug / ml poly (I:C), followed by measurement of proinflammatory cytokines.Single cell RNA sequencing was used to compare transcriptome between normal and CKD kidneys.
Conclusions: Our results show that CKD mice are more sensitive to foreign double strand RNA insult.BMDM isolated from cisplatin-induced CKD demonstrated a greater innate immune response during CKD.We propose that the inherent hyperinflammatory nature of CKD drives a greater innate immune response in this model of viral injury and may be responsible, at least partially, for the poor outcomes in CKD patients with Covid-19 infection.
Funding: NIDDK Support Background: Epoxyeicosatrienoic acids (EETs) are arachidonic acid metabolites with biological effects, including anti-apoptotic, anti-inflammatory, and anti-fibrotic functions.Soluble epoxide hydrolase (sEH)-mediated hydrolysis of EETs to dihydroxyeicosatrienoic acids (DHET) attenuates these effects.Recent studies have demonstrated inhibition of sEH prevents renal tubulointerstitial fibrosis and inflammation in chronic kidney disease (CKD) model.Here, we demonstrated the role and underlying mechanism of EETs in unilateral ureteral obstruction (UUO)-induced renal fibrogenesis.
Results: EETs administration abolished tubulointerstitial fibrogenesis, as demonstrated by reduced fibroblast activation and collagen deposition after UUO.Furthermore, inflammatory response was prevented as demonstrated by decreased macrophage infiltration and expression of inflammatory cytokines (TGF-β, IL-1β and IL-6) in EETs-administered UUO kidneys.The genetic inhibition of sEH also mitigated UUO-induced renal inflammation and interstitial fibrogenesis.The combination of EET administration and genetic sEH inhibition also attenuated inflammation and renal interstitial fibrogenesis after UUO, but no additive or synergic effect of combined sEH inhibition and EETs administration.
Conclusions: Taken together, our findings provide that the underlying mechanism of EETs in kidney fibrogenesis during obstructive nephropathy, suggesting EETs as a potential therapeutic target of kidney fibrosis progression.
Funding: NIDDK Support
Background: Renal fibrosis is a hallmark of Chronic Kidney Disease (CKD), which affects 10-16% of the world's adult population.Yet current treatment strategies are ineffective in attenuating renal fibrogenesis.Therefore, we are in urgent need for new therapeutic strategies against renal fibrosis.The cyclooxygenase/prostaglandin (COX/PG) system plays a key role in renal fibrosis and holds great promise as a suitable therapeutic target.Here, we used a translational approach to evaluate the role of the PGE 2 -EP 1 receptor in the pathogenesis of renal fibrosis in several models of kidney injury, including human (fibrotic) kidney slices.
Methods: The anti-fibrotic effect of SC-19220 -an EP 1 receptor antagonist -was studied in Madin-Darby Canine Kidney (MDCK) cells, mice subjected to seven days of unilateral ureteral obstruction (UUO), and healthy and fibrotic human precision-cut kidney slices (PCKS).Progression of fibrosis was evaluated on gene and protein level using qPCR, Western blot and immunohistochemistry.
Results: Pharmacological inhibition of the EP 1 receptor using SC-19220 reduced TGF-β-induced fibronectin (FN) expression, ERK1/2 phosphorylation and epithelialto-mesenchymal transition in MDCK cells.Moreover, SC-19220 diminished fibrosis in UUO mice, measured by decreased protein expression of FN and α-smooth muscle actin (αSMA), and a reduction in collagen deposition.In addition, treatment of healthy human PCKS with SC-19220 reduced TGF-β-induced fibrosis as shown by decreased gene levels of collagen 1A1, FN and αSMA as well as reduced collagen deposition.Moreover, similar observations were made using fibrotic human PCKS.
Conclusions: This study highlights that the EP 1 receptor is a promising target for preventing both the onset and late stage of renal fibrosis.Moreover, we provide strong evidence that the effect of SC-19220 may translate to clinical care since its effects were observed in UUO mice and human kidney slices.