Li, Xiaogang

Abstract Mixed lineage leukemia 1 (MLL1), a histone H3 lysine 4 (H3K4) methyltransferase, exerts its enzymatic activity by interacting with menin and other proteins. It is unclear whether inhibition of the MLL1‐menin interaction influences epithelial‐mesenchymal transition (EMT), renal fibroblast activation, and renal fibrosis. In this study, we investigated the effect of disrupting MLL1‐menin interaction on those events and mechanisms involved in a murine model of renal fibrosis induced by unilateral ureteral obstruction (UUO), in cultured mouse proximal tubular cells and renal interstitial fibroblasts. Injury to the kidney increased the expression of MLL1 and menin and H3K4 monomethylation (H3K4me1); MLL1 and menin were expressed in renal epithelial cells and renal interstitial fibroblasts. Inhibition of the MLL1‐menin interaction by MI‐503 administration or siRNA‐mediated silencing of MLL1 attenuated UUO‐induced renal fibrosis, and reduced expression of α‐smooth muscle actin (α‐SMA) and fibronectin. These treatments also inhibited UUO‐induced expression of transcription factors Snail and Twist and transforming growth factor β1 (TGF‐β1) while expression of E‐cadherin was preserved. Moreover, treatment with MI‐503 and transfection with either MLL siRNA or menin siRNA inhibited TGF‐β1‐induced upregulation of α‐SMA, fibronectin and Snail, phosphorylation of Smad3 and AKT, and downregulation of E‐cadherin in cultured renal epithelial cells. Finally, MI‐503 was effective in abrogating serum or TGFβ1‐induced transformation of renal interstitial fibroblasts to myofibroblasts in vitro. Taken together, these results suggest that targeting disruption of the MLL1‐menin interaction attenuates renal fibrosis through inhibition of partial EMT and renal fibroblast activation.

TP53 is the most common mutated gene in human cancer. Mutant p53 protein loses its tumor-suppressor properties and gains oncogenic activity. Mutant p53 is a therapeutic target in a broad range of cancer types. However, how mutant p53 is epigenetically regulated during tumor progression remains elusive. In this study, we found that the upregulation of mutant p53 is mediated by bromodomain protein BRD4 in triple-negative breast cancer (TNBC) cells. Inhibition of BRD4 with its inhibitor JQ1 or knockdown of BRD4 suppressed the transcription of mutant p53, which led to the re-expression of p21, the inhibition of S-phase entry, and colony formation in TNBC cells. BRD4 also positively regulated the transcription of wild-type p53, whereas JQ1 treatment and knockdown of BRD4 decreased the expression of p21 in MCF-7 cells. Knockdown of BRD4 resulted in attenuation of TNBC tumor growth in vivo. Taken together, our results uncover a novel regulatory mechanism of mutant p53 via BRD4, and suggest that the bromodomain inhibitor suppresses tumorigenesis through targeting mutant p53 in TNBC.

AbstractAutosomal dominant polycystic kidney disease (ADPKD) is driven by mutations in the PKD1 and PKD2 genes, and it is characterized by renal cyst formation, inflammation and fibrosis. Forkhead box protein M1 (FoxM1), a transcription factor of the Forkhead box (Fox) protein super family, has been reported to promote tumor formation, inflammation and fibrosis in many organs. However, the role and mechanism of FoxM1 in regulation of ADPKD progression is still poorly understood. Here, we show that FoxM1 is an important regulator of cyst growth in ADPKD. FoxM1 is upregulated in cyst-lining epithelial cells in Pkd1 mutant mouse kidneys and human ADPKD kidneys. FoxM1 promotes cystic renal epithelial cell proliferation by increasing the expression of Akt and Stat3 and the activation of ERK and Rb. FoxM1 also regulates cystic renal epithelial cell apoptosis through NF-κB signaling pathways. In addition, FoxM1 regulates the recruitment and retention of macrophages in Pkd1 mutant mouse kidneys, a process that is associated with FoxM1-mediated upregulation of monocyte chemotactic protein 1. Targeting FoxM1 with its specific inhibitor, FDI-6, delays cyst growth in rapidly progressing and slowly progressing Pkd1 mutant mouse kidneys. This study suggests that FoxM1 is a central and upstream regulator of ADPKD pathogenesis and provides a rationale for targeting FoxM1 as a therapeutic strategy for ADPKD treatment.

Abstract Macrophage migration inhibitory factor (MIF1) is a pleiotropic cytokine involved in inflammation and cancer. Genetic knockout of Mif1 in the validated N ‐butyl‐ N ‐(4‐hydroxybutyl) nitrosamine (BBN) model of bladder cancer (BCa) resulted in stage arrest at non‐muscle‐invasive disease in prior studies. Small‐molecule inhibition of MIF1 reduced cancer‐associated outcomes, but it did not fully recapitulate genetic models. D‐dopachrome tautomerase (gene symbol DDT ), commonly referred to as MIF2, is a functional homolog of MIF1, and both MIF1 and MIF2 can bind the cell surface receptor CD74 on multiple cell types to initiate a signaling cascade. It has been proposed that this interaction mediates part of the protumorigenic effects of MIF1 and MIF2 and may explain the discordance in prior studies. We hypothesized that MIF2 functions redundantly with MIF1 in BCa development and progression. The Cancer Genome Atlas (TCGA) analysis indicated MIF and DDT expression were increased in BCa patients compared to control. 4‐Iodopyridine (4‐IPP), a combined MIF1/MIF2 inhibitor, was more efficacious than ISO‐1, a MIF1‐only inhibitor, in preventing cellular proliferation in BCa cell lines. To evaluate these findings in vivo , wild‐type (WT) and Mif1 −/− animals were exposed to 0.05% BBN in drinking water for 16 weeks to initiate tumorigenesis and then evaluated over the subsequent 4 weeks for tumor formation and progression in the presence or absence of 4‐IPP. 4‐IPP reduced bladder weights in WT animals and bladder weights/tumor stage in Mif1 −/− animals. To determine whether MIF1/MIF2 functioned through CD74 in BCa, WT or Cd74 −/− animals were used in the same BBN model. Although these animals were partially protected against BBN‐induced BCa, 4‐IPP did not enhance this effect. In conclusion, our data suggest that MIF2 mechanistically functions in a similar protumorigenic manner to MIF1, and this is at least partially through CD74. Dual inhibition of MIF homologs is more efficacious at reducing tumor burden in this model of BCa. © 2022 The Pathological Society of Great Britain and Ireland.

Abstract DNA damage response (DDR) is an important signaling-transduction network that promotes the repair of DNA lesions which can induce and/or support diseases. However, the mechanisms involved in its regulation are not fully understood. Recent studies suggest that the peroxiredoxin 5 (Prdx5) enzyme, which detoxifies reactive oxygen species, is associated to genomic instability and signal transduction. Its role in the regulation of DDR, however, is not well characterized. In this study, we demonstrate a role of Prdx5 in the regulation of the DDR signaling pathway. Knockdown of Prdx5 resulted in DNA damage manifested by the induction of phosphorylated histone H2AX (γ-H2AX) and p53-binding protein 1 (53BP1). We show that Prdx5 regulates DDR through (1) polo-like kinase 1 (Plk1) mediated phosphorylation of ataxia telangiectasia mutated (ATM) kinase to further trigger downstream mediators Chek1 and Chek2; (2) the increase of the acetylation of p53 at lysine 382, stabilizing p53 in the nucleus and enhancing transcription and (3) the induction of autophagy, which regulates the recycling of molecules involved in DDR. We identified Sirt2 as a novel deacetylase of p53 at lysine 382, and Sirt2 regulated the acetylation status of p53 at lysine 382 in a Prdx5-dependent manner. Furthermore, we found that exogenous expression of Prdx5 decreased DNA damage and the activation of ATM in Pkd1 mutant renal epithelial cells, suggesting that Prdx5 may play a protective role from DNA damage in cystic renal epithelial cells. This study identified a novel mechanism of Prdx5 in the regulation of DDR through the ATM/p53/Sirt2 signaling cascade.

Autosomal dominant polycystic kidney disease (ADPKD) is a renal neoplastic disorder characterized by the formation of large fluid-filled cysts that develop from kidney tubules through abnormal cell proliferation and cyst-filling fluid secretion driven by cAMP-dependent Cl − secretion. This study shows that the lonidamine derivative H2-GMZ inhibits Cl − secretion, cell proliferation, and cyst growth, suggesting that it might have therapeutic value for the treatment of ADPKD.

Abstract Lysyl oxidase (LOX) is a copper‐dependent monoamine oxidase whose primary function is the covalent cross‐linking of collagen in the extracellular matrix (ECM). Evidence has shown that LOX is associated with cancer and some fibrotic conditions. We recently found that serum LOX is a potential diagnostic biomarker for renal fibrosis, but the mechanism by which LOX is regulated and contributes to renal fibrosis remains unknown. The current study demonstrates the following: (1) LOX expression was increased in fibrotic kidneys including ischemia‐reperfusion injury‐(IRI‐), unilateral ureteral obstruction‐(UUO‐), and folic acid‐ (FA‐) induced fibrotic kidneys as well as in the paraffin‐embedded sections of human kidneys from the patients with renal fibrosis. (2) The increasing deposition and cross‐linking of collagen induced by LOX was observed in IRI‐, UUO‐ and FA‐kidneys. (3) LOX was regulated by the β‐arrestin‐ERK‐STAT3 pathway in renal fibrosis. STAT3 was the downstream of AT1R‐β‐arrestin‐ERK, ERK entered the nucleus and activated STAT3‐pY705 but not STAT3‐pS727. (4) STAT3 nuclear subtranslocation and binding to the LOX promoter may be responsible for the upregulation of LOX expression. (5) Pharmacologic inhibition of LOX with BAPN in vivo inhibited the upregulation of LOX, decreased collagen over cross‐linking and ameliorated renal fibrosis after ischemic injury. Collectively, these observations suggest that LOX plays an essential role in the development of renal fibrosis by catalyzing collagen over cross‐linking. Thus, strategies targeting LOX could be a new avenue in developing therapeutics against renal fibrosis.

Here, we identified a cross talk between SET and MYND domain-containing lysine methyltransferase 2 (Smyd2) and transforming growth factor (TGF)-β-Smad3 signaling and a synergistic feedback loop between them, in which TGF-β stimulates expression of Smyd2 in a Smad3-dependent manner, and upregulation of Smyd2 regulates the transcription of TGF-β and other fibrotic marker genes through direct binding on their promoters or methylating histone H3 indirectly to regulate the transcription of those genes in fibroblasts. Thus, the Smyd2-TGF-β-Smad3-Smyd2 signaling axis plays an important role in promoting renal fibrosis, and targeting Smyd2 with its specific inhibitor should not only delay cyst growth but also ameliorate renal fibrosis in ADPKD.

Ferroptosis is a newly identified form of regulated cell death driven by iron-dependent phospholipid peroxidation and oxidative stress. Ferroptosis has distinct biological and morphology characteristics, such as shrunken mitochondria when compared to other known regulated cell deaths. The regulation of ferroptosis includes different molecular mechanisms and multiple cellular metabolic pathways, including glutathione/glutathione peroxidase 4(GPX4) signaling pathways, which are involved in the amino acid metabolism and the activation of GPX4; iron metabolic signaling pathways, which are involved in the regulation of iron import/export and the storage/release of intracellular iron through iron-regulatory proteins (IRPs), and lipid metabolic signaling pathways, which are involved in the metabolism of unsaturated fatty acids in cell membranes. Ferroptosis plays an essential role in the pathology of various kidneys diseases, including acute kidney injury (AKI), chronic kidney disease (CKD), autosomal dominant polycystic kidney disease (ADPKD), and renal cell carcinoma (RCC). Targeting ferroptosis with its inducers/initiators and inhibitors can modulate the progression of kidney diseases in animal models. In this review, we discuss the characteristics of ferroptosis and the ferroptosis-based mechanisms, highlighting the potential role of the main ferroptosis-associated metabolic pathways in the treatment and prevention of various kidney diseases.

Abstract Lysine‐specific histone demethylase 1 (LSD1) as the first identified histone/lysine demethylase regulates gene expression and protein functions in diverse diseases. In this study, we show that the expression of LSD1 is increased in mouse kidneys with unilateral ureteral obstruction (UUO) and in cultured NRK‐52E cells undergoing TGF‐β1‐induced epithelial–mesenchymal transition (EMT). Inhibition of LSD1 with its specific inhibitor ORY1001 attenuated renal EMT and fibrosis, which was associated with decreased the deposition of extracellular matrix proteins and the expression of fibrotic markers, including α‐smooth muscle actin (α‐SMA) and fibronectin, and the recovery of E‐cadherin expression and decrease of N‐cadherin expression in UUO kidneys and in NRK‐52E cells induced with TGF‐β1. Targeting LSD1 also decreased the expression of Snail family transcriptional repressor 1 (Snail‐1) and its interaction with LSD1 in UUO kidneys and in NRK‐52E cells treated with TGF‐β1. In addition, we identified a novel LSD1‐14‐3‐3ζ‐PKCα axis in the regulation of the activation of AKT and Stat3 and then the activation of fibroblasts. This study suggests that LSD1 plays a critical role in regulation of renal EMT and fibrosis through activation of diverse signaling pathways and places an emphasis that LSD1 has potential as a therapeutic target for the treatment of renal fibrosis.