Beta-adrenergic receptor blocker propranolol triggers anti-tumor immunity and enhances irinotecan therapy in mice colorectal cancer.

Colorectal cancer (CRC) stands as the second leading cause of cancer-related deaths worldwide with limited available medicines. While drug repurposing comes as a promising strategy for cancer treatment, we discovered that propranolol (Prop), a non-selective ß1 and ß2 adrenergic receptor blocker, significantly inhibited the development of subcutaneous CT26 CRC and AOM/DSS-induced CRC models. The RNA-seq analysis highlighted the activated immune pathways after Prop treatment, with KEGG analysis enriched in T-cell differentiation. Routine analyses of blood revealed a decrease in neutrophil to lymphocyte ratio, a biomarker of systemic inflammation, and a prognostic indicator in the Prop-treated groups in both CRC models. Analysis of the tumor-infiltrating immune cells exhibited that Prop regressed the exhaustion of CD4+ and CD8+ T cells in the CT26-derived graft models, which was further corroborated in the AOM/DSS-induced models. Furthermore, bioinformatic analysis fitted well with the experimental data, showing that ß2 adrenergic receptor (ADRB2) was positively correlated with T-cell exhaustion signature in various tumors. The in vitro experiment showed no direct effect of Prop on CT26 cell viability, while T cells were activated with significantly-upregulated production of IFN-γ and Granzyme B. Consistently, Prop was unable to restrain CT26 tumor growth in nude mice. At last, the combination of Prop and the chemotherapeutic drug Irinotecan acted out the strongest inhibition in CT26 tumor progress. Collectively, we repurpose Prop as a promising and economical therapeutic drug for CRC treatment and highlight T-cell as its target.

TRPC absence induces pro-inflammatory macrophages and gut microbe disorder, sensitizing mice to colitis.

The transient receptor potential canonical (TRPC) channels, encoded in seven non-allelic genes, are important contributors to calcium fluxes, are strongly associated with various diseases. Here we explored the consequences of ablating all seven TRPCs in mice focusing on colitis. We discovered that absence of all seven TRPC proteins in mice (TRPC HeptaKO mice) promotes the development of dextran sulfate sodium (DSS)-induced colitis. RNA-sequence analysis highlighted an extremely pro-inflammatory profile in colons of DSS-treated TRPC HeptaKO mice, with an amount of increased pro-inflammatory cytokines and chemokines. Flow cytometry analysis showed that the infiltration of Ly6Chi monocytes and neutrophils in colonic lamina propria was significantly increased in DSS-treated TRPC HeptaKO mice. Results also revealed that macrophages from TRPC HeptaKO mice exhibited M1 polarization and enhanced secretion of pro-inflammatory factors. In addition, the composition of gut microbiota was markedly disturbed in DSS-treated TRPC HeptaKO mice. However, upon antibiotic cocktail (Abx)-treatment, TRPC HeptaKO mice showed no significant differences with WT mice in disease severity. Collectively, these data suggest that ablation of all TRPCs promotes the development of DSS-induced colitis by inducing pro-inflammatory macrophages and gut microbiota disorder.

ACSL4 deficiency confers protection against ferroptosis-mediated acute kidney injury.

The term ferroptosis coined in 2012 causes acute kidney injury (AKI). However, its pathway mechanism in AKI is poorly understood. In this study, we conducted an RNA-sequence analysis of kidneys in AKI and normal mice to explore the pathway mechanism of ferroptosis. Consequently, differentially expressed genes highlighted Acyl-CoA synthetase long-chain family (ACSL4), a known promotor for ferroptosis. Besides, RT-PCR, Western blot, and immunohistochemical analyses confirmed its upregulation. HIF-1α was downregulated in I/R-AKI mice, and in vitro studies confirmed a negative regulation of HIF-1α on ACSL4. To explore the role of ACSL4 in AKI, we constructed ACSL4 knockout in kidney tubules of mice-as Cdh16Cre-ACSL4F/F mice. Results revealed that ACSL4 knockout significantly reduced ferroptosis and inhibited the functional and pathological injury of AKI mice. Meanwhile, the kidneys of Cdh16Cre-ACSL4F/F mice demonstrated a significantly decreased inflammation and macrophage infiltration. Further, additional explorations were explored to decipher a more thorough understanding of ferroptotic immunogenicity. As a result, neutrophils were not directly recruited by ferroptotic cells, but by ferroptotic cell-induced macrophages. Further, ACSL4 inhibitor rosiglitazone significantly inhibited AKI. Collectively, these data provide novel insights into the AKI pathogenesis, and defined ACSL4 as an effective target in AKI.

IL-6 deficiency promotes colitis by recruiting Ly6Chi monocytes into inflamed colon tissues in a CCL2-CCR2-dependent manner.

Interleukin 6 (IL-6) is a pleiotropic cytokine that is elevated in inflammatory bowel disease. However, the role of IL-6 deficiency in colitis is not well-defined. Some IL-6 and IL-6 receptor antagonists are associated with severe gastrointestinal immune adverse effects, but the mechanisms of the effects are not clear. This study aimed to investigate the effect of IL-6 in ulcerative colitis in Il6-/- mice. Results indicated that physiological deficiency of IL-6 promoted the development of colitis. Moreover, IL-6 deficiency significantly increased the mRNA levels of monocytes chemokine Ccl2 and its receptor Ccr2 in colon tissues. Similarly, the percentage of Ly6Chigh monocytes and neutrophils were increased in the colon of Il6-/- mice. Intestinal crypts more strongly increased the migration of Il6-/- macrophages than wild-type ones. Moreover, Il6-/- macrophages promoted the migration of neutrophils. Most importantly, RS102895, an antagonist of CCR2, diminished chemotaxis of macrophages and inhibited colitis in Il6-/- mice. Collectively, these results indicate that Il6-/- macrophages migrate to inflamed colon tissues and recruit neutrophils, thereby promoting the effect of Il6-/- on colitis. This study expands our understanding on the effect of IL-6 deficiency in colitis and the development of gastrointestinal immune adverse effects.

Quercetin alleviates acute kidney injury by inhibiting ferroptosis.

INTRODUCTION: Ferroptosis is an iron-dependent regulated necrosis and has been proven to contribute to the progress of acute kidney injury (AKI). Quercetin (QCT), a natural flavonoid which is commonly found in numerous fruits and vegetables, has extensive pharmacological effects, such as anti-oxidant, anti-inflammatory and anti-senescence effects. OBJECTIVES: This study aims to explain whether ferroptosis is a therapeutic strategy to AKI, and to explore the effect of QCT on AKI ferroptosis. METHODS: NRK-52E cells and HK-2 cells were used for in vitro ferroptosis studies. Morphology of cells was detected by transmission electron microscopy. Lipid ROS was assayed using flow cytometry. In vivo, AKI was induced by ischemia-reperfusion (I/R) or folic acid (FA). To explore the molecular mechanisms, RNA-sequence analysis was performed. Transwell was used to detect macrophage migration. RESULTS: We discovered that quercetin (QCT), a natural flavonoid, inhibited ferroptosis in renal proximal tubular epithelial cells. QCT blocked the typical morphologic changes of ferroptotic cells by reducing the levels of malondialdehyde (MDA) and lipid ROS and increasing the levels of glutathione (GSH). Moreover, QCT ameliorated AKI induced by I/R or FA. RNA-sequence analysis highlighted activation transcription factor 3 (ATF3), as it was the dominant one among all the 299 down-regulated genes by QCT. Knockdown of ATF3 could significantly increase the levels of SLC7A11, GPX4 and increased the cell viability. In addition, ferroptotic cells were found to be extremely pro-inflammatory by recruiting macrophages through CCL2, while QCT inhibited the chemotaxis of macrophages induced by ferroptosis in AKI. CONCLUSIONS: Collectively, these results identify QCT as a ferroptosis inhibitor and provide new therapeutic strategies for diseases related to ferroptosis.

Ferroptosis involves in renal tubular cell death in diabetic nephropathy.

Ferroptosis is a novel type of programmed cell death characterized by iron-dependent accumulation of lipid hydroperoxides to lethal levels. Accumulative studies have indicated diabetic nephropathy (DN) as an inflammatory disorder, which involved immune modulation both in the occurrence and progression of the disease. In addition, DN is also considered as the major threatening complication of Diabetes mellitus (DM). However, other forms of programmed cell death, such as autophagy, apoptosis and necrosis, have been reported to be associated with DN, while there are no effective drugs to alleviate the damage of DN. In this study, we explored whether ferroptosis was involved in the progression of DN both in vivo and in vitro. We first established DN models using streptozotocin (STZ) and db/db mice. Results showed significant changes of ferroptosis associated markers, like increased expression levels of acyl-CoA synthetase long-chain family member 4 (ACSL4) and decreased expression levels of glutathione peroxidase 4 (GPX4) in DN mice. Also lipid peroxidation products and iron content were increased in DN mice. Next, in vitro, ferroptosis inducer erastin or RSL3 could induce renal tubular cell death, while iron and high ACSL4 levels sensitised ferroptosis. Finally, ACSL4 inhibitor rosiglitazone (Rosi) was used in the development of DN, which improved survival rate and kidney function, reduced lipid peroxidation product MDA and iron content. In summary, we first found ferroptosis was involved in DN and ferroptosis might be as a future direction in the treatment of DN.