Acute Kidney Injury-Induced Circulating TNFR1/2 Elevations Correlate with Persistent Kidney Injury and Progression to Fibrosis.

Elevated levels of circulating tumor necrosis factor receptors 1 and 2 (cTNFR1/2) predict chronic kidney disease (CKD) progression; however, the mechanisms of their release remain unknown. Whether acute kidney injury (AKI) drives cTNFR1/2 elevations and whether they predict disease outcomes after AKI remain unknown. In this study, we used AKI patient serum and urine samples, mouse models of kidney injury (ischemic, obstructive, and toxic), and progression to fibrosis, nephrectomy, and related single-cell RNA-sequencing datasets to experimentally test the role of kidney injury on cTNFR1/2 levels. We show that TNFR1/2 serum and urine levels are highly elevated in all of the mouse models of kidney injury tested, beginning within one hour post injury, and correlate with its severity. Consistent with this, serum and urine TNFR1/2 levels are increased in AKI patients and correlate with the severity of kidney failure. Kidney tissue expression of TNFR1/2 after AKI is only slightly increased and bilateral nephrectomies lead to strong cTNFR1/2 elevations, suggesting the release of these receptors by extrarenal sources. The injection of the uremic toxin indoxyl sulfate in healthy mice induces moderate cTNFR1/2 elevations. Moreover, TNF neutralization does not affect early cTNFR1/2 elevations after AKI. These data suggest that cTNFR1/2 levels in AKI do not reflect injury-induced TNF activity, but rather a rapid response to loss of kidney function and uremia. In contrast to traditional disease biomarkers, such as serum creatinine or BUN, cTNFR1/2 levels remain elevated for weeks after severe kidney injury. At these later timepoints, cTNFR1/2 levels positively correlate with remaining kidney injury. During the AKI-to-CKD transition, elevations of TNFR1/2 kidney expression and of cTNFR2 levels correlate with kidney fibrosis levels. In conclusion, our data demonstrate that kidney injury drives acute increases in cTNFR1/2 serum levels, which negatively correlate with kidney function. Sustained TNFR1/2 elevations after kidney injury during AKI-to-CKD transition reflect persistent tissue injury and progression to kidney fibrosis.

TNF or EGFR inhibition equally block AKI-to-CKD transition: opportunities for etanercept treatment.

BACKGROUND: Inflammation is a key driver of the transition of acute kidney injury to progressive fibrosis and chronic kidney disease (AKI-to-CKD transition). Blocking a-disintegrin-and-metalloprotease-17 (ADAM17)-dependent ectodomain shedding, in particular of epidermal growth factor receptor (EGFR) ligands and of the type 1 inflammatory cytokine tumor necrosis factor (TNF), reduces pro-inflammatory and pro-fibrotic responses after ischemic AKI or unilateral ureteral obstruction (UUO), a classical fibrosis model. Metalloprotease or EGFR inhibition show significant undesirable side effects in humans. In retrospective studies anti-TNF biologics reduce the incidence and progression of CKD in humans. Whether TNF has a role in AKI-to-CKD transition and how TNF inhibition compares to EGFR inhibition is largely unknown. METHODS: Mice were subjected to bilateral renal ischemia-reperfusion injury or unilateral ureteral obstruction. Kidneys were analyzed by histology, immunohistochemistry, qPCR, western blot, mass cytometry, scRNA sequencing, and cytokine profiling. RESULTS: Here we show that TNF or EGFR inhibition reduce AKI-to-CKD transition and fibrosis equally by about 25%, while combination has no additional effect. EGFR inhibition reduced kidney TNF expression by about 50% largely by reducing accumulation of TNF expressing immune cells in the kidney early after AKI, while TNF inhibition did not affect EGFR activation or immune cell accumulation. Using scRNAseq data we show that TNF is predominantly expressed by immune cells in AKI but not in proximal tubule cells (PTC), and PTC-TNF knockout did not affect AKI-to-CKD transition in UUO. Thus, the anti-inflammatory and anti-fibrotic effects of the anti-TNF biologic etanercept in AKI-to-CKD transition rely on blocking TNF that is released from immune cells recruited or accumulating in response to PTC-EGFR signals. CONCLUSION: Short-term anti-TNF biologics during or after AKI could be helpful in the prevention of AKI-to-CKD transition.

FOXM1 drives proximal tubule proliferation during repair from acute ischemic kidney injury.

The proximal tubule has a remarkable capacity for repair after acute injury, but the cellular lineage and molecular mechanisms underlying this repair response are incompletely understood. Here, we developed a Kim1-GFPCreERt2 knockin mouse line (Kim1-GCE) in order to perform genetic lineage tracing of dedifferentiated cells while measuring the cellular transcriptome of proximal tubule during repair. Acutely injured genetically labeled clones coexpressed KIM1, VIMENTIN, SOX9, and KI67, indicating a dedifferentiated and proliferative state. Clonal analysis revealed clonal expansion of Kim1+ cells, indicating that acutely injured, dedifferentiated proximal tubule cells, rather than fixed tubular progenitor cells, account for repair. Translational profiling during injury and repair revealed signatures of both successful and unsuccessful maladaptive repair. The transcription factor Foxm1 was induced early in injury, was required for epithelial proliferation in vitro, and was dependent on epidermal growth factor receptor (EGFR) stimulation. In conclusion, dedifferentiated proximal tubule cells effect proximal tubule repair, and we reveal an EGFR/FOXM1-dependent signaling pathway that drives proliferative repair after injury.

Indole-3- carbinol enhances sorafenib cytotoxicity in hepatocellular carcinoma cells: A mechanistic study.

Sorafenib is the only chemotherapeutic agent currently approved for unresectable hepatocellular carcinoma (HCC). However, poor response rates have been widely reported. Indole-3-carbinol (I3C) is a potential chemopreventive phytochemical. The present study aimed to explore the potential chemomodulatory effects of I3C on sorafenib in HCC cells as well as the possible underlying mechanisms. I3C exhibited a greater cytotoxicity in HepG2 cells compared to Huh-7 cells (p < 0.0001). Moreover, the co-treatment of HepG2 cells with I3C and sorafenib was more effective (p = 0.002). Accordingly, subsequent mechanistic studies were carried on HepG2 cells. The results show that the ability of I3C to enhance sorafenib cytotoxicity in HCC cells could be partially attributed to increasing the apoptotic activity and decreasing the angiogenic potentials. The combination had a negative effect on epithelial-mesenchymal transition (EMT). Increased NOX-1 expression was also observed which may indicate the involvement of NOX-1 in I3C chemomodulatory effects. Additionally, the combination induced cell cycle arrest at the G0/G1 phase. In conclusion, these findings provide evidence that I3C enhances sorafenib anti-cancer activity in HCC cells.