Pyroptosis, a form of programmed cell death (PCD), has garnered increasing attention as it relates to innate immunity and diseases. However, the involvement of pyroptosis in the mechanism by which lobaplatin acts against colorectal cancer (CRC) is unclear. Our study revealed that treatment with lobaplatin reduced the viability of HT-29 and HCT116 cells in a dose-dependent manner. Morphologically, HT-29 and HCT116 cells treated with lobaplatin exhibited microscopic features of cell swelling and large bubbles emerging from the plasma membrane, and transmission electron microscopy (TEM) revealed multiple pores in the membrane. GSDME, rather than GSDMD, was cleaved in lobaplatin-induced pyroptosis in HT-29 and HCT116 cells due to caspase-3 activation. Knocking out GSDME switched lobaplatin-induced cell death from pyroptosis to apoptosis but did not affect lobaplatin-mediated inhibition of growth and tumour formation of HT-29 and HCT116 cells in vivo and in vitro. Further investigation indicates that lobaplatin induced reactive oxygen species (ROS) elevation and JNK phosphorylation. NAC, a ROS scavenger, completely reversed the pyroptosis of lobaplatin-treated HT-29 and HCT116 and JNK phosphorylation. Activated JNK recruited Bax to mitochondria, and thereby stimulated cytochrome c release to cytosol, followed by caspase-3/-9 cleavage and pyroptosis induction. Therefore, in colon cancer cells, GSDME mediates lobaplatin-induced pyroptosis downstream of the ROS/JNK/Bax-mitochondrial apoptotic pathway and caspase-3/-9 activation. Our study indicated that GSDME-dependent pyroptosis is an unrecognized mechanism by which lobaplatin eradicates neoplastic cells, which may have important implications for the clinical application of anticancer therapeutics.

The number and morphology of mitochondria within a cell are precisely regulated by the mitochondrial fission and fusion machinery. The human protein, hFis1, participates in mitochondrial fission by recruiting the Drp1 into the mitochondria. Using short hairpin RNA, we reduced the expression levels of hFis1 in mammalian cells. Cells lacking hFis1 showed sustained elongation of mitochondria and underwent significant cellular morphological changes, including enlargement, flattening, and increased cellular granularity. In these cells, staining for acidic senescence-associated ␤-galactosidase activity was elevated, and the rate of cell proliferation was greatly reduced, indicating that cells lacking hFis1 undergo senescence-associated phenotypic changes. Reintroduction of the hFis1 gene into hFis1-depleted cells restored mitochondrial fragmentation and suppressed senescence-associated ␤-galactosidase activity. Moreover, depletion of both hFis1 and OPA1, a critical component of mitochondrial fusion, resulted in extensive mitochondrial fragmentation and markedly rescued cells from senescence-associated phenotypic changes. Intriguingly, sustained elongation of mitochondria was associated with decreased mitochondrial membrane potential, increased reactive oxygen species production, and DNA damage. The data indicate that sustained mitochondrial elongation induces senescence-associated phenotypic changes that can be neutralized by mitochondrial fragmentation. Thus, one of the key functions of mitochondrial fission might be prevention of the sustained extensive mitochondrial elongation that triggers cellular senescence.Mitochondria are dynamic organelles that can change in number and morphology within a cell during development, the cell cycle, or when challenged with various cytotoxic conditions. Size, shape, and interconnectivity of mitochondria are determined by fusion and fission. In mammals, the key molecules for mitochondrial fission are hFis1 and Drp1. The hFis1 protein is anchored to the outer mitochondrial membrane via a C-terminal transmembrane domain, and overexpression of hFis1 was found to induce mitochondrial fragmentation (1, 2). The Drp1 is predominantly distributed in the cytoplasm and partially associates with the mitochondrial outer membrane (3). A portion of cytosolic Drp1 can be recruited to mitochondria through an interaction with hFis1 (4 -6). The opposing process, mitochondrial fusion, is controlled in mammalian cells by Mitofusins (Mfn) 3 and OPA1. Mitofusin1 and -2 (Mfn1 and Mfn2) localize on the outer membrane of mitochondria and may directly mediate mitochondrial fusion (7-9). OPA1 (optic atrophy 1) is a dynamin family GTPase that resides in the intermembrane space of mitochondria and is essential for mitochondrial fusion (10, 11). However, the functional mechanism by which these proteins cooperate to induce mitochondrial fission and fusion remains unidentified.Despite relatively intensive studies on the components of the mitochondrial fission and fusion machineries, a link between mitochondrial...

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Caudal-related homeobox transcription factor 2 (CDX2), an intestine-specific nuclear transcription factor, has been strongly implicated in the tumourigenesis of various human cancers. However, the functional role of CDX2 in the development and progression of colorectal cancer (CRC) is not well known. In this study, CDX2 knockdown in colon cancer cells promoted cell proliferation in vitro, accelerated tumor formation in vivo, and induced a cell cycle transition from G0/G1 to S phase, whereas CDX2 overexpression inhibited cell proliferation. TOP/FOP-Flash reporter assay showed that CDX2 knockdown or CDX2 overexpression significantly increased or decreased Wnt signaling activity. Western blot assay showed that downstream targets of Wnt signaling, including β-catenin, cyclin D1 and c-myc, were up-regulated or down-regulated in CDX2-knockdown or CDX2-overexpressing colon cancer cells. In addition, suppression of Wnt signaling by XAV-939 led to a marked suppression of the cell proliferation enhanced by CDX2 knockdown, whereas activation of this signaling by CHIR-99021 significantly enhanced the cell proliferation inhibited by CDX2 overexpression. Dual-luciferase reporter and quantitative chromatin immunoprecipitation (qChIP) assays further confirmed that CDX2 transcriptionally activates glycogen synthase kinase-3β (GSK-3β) and axis inhibition protein 2 (Axin2) expression by directly binding to the promoter of GSK-3β and the upstream enhancer of Axin2. In conclusion, these results indicated that CDX2 inhibits the proliferation and tumor formation of colon cancer cells by suppressing Wnt/β-catenin signaling.

Bile acids serve a critical role in the induction of gastric intestinal metaplasia (IM) and gastric carcinogenesis. The present study investigated the effects of bile acids on the induction of gastric IM formation. The results demonstrated that the expression levels of caudal-related homeobox transcription factor 2 (CDX2), mucin 2 (MUC2) and farnesoid X receptor (FXR) were increased in vitro and in vivo following treatment with bile acids, and CDX2 transcriptionally activated MUC2 expression. Furthermore, knockdown of FXR attenuated bile acid-enhanced CDX2 promoter activity and protein expression. Conversely, the FXR agonist GW4064 synergistically enhanced bile acid-induced CDX2 promoter activity. Bile acid treatment led to an increase in nuclear factor (NF)-κB activity and protein expression. Treatment with GW4064 or the FXR antagonist Z-guggulsterone enhanced or attenuated bile acid-induced NF-κB activity, respectively. In addition, quantitative chromatin immunoprecipitation confirmed that bile acids led to enhanced binding of p50 to the CDX2 promoter, whereas this effect was not observed for p65. Treatment with GW4064 or Z-guggulsterone enhanced and attenuated the binding activity of p50 to the CDX2 promoter, respectively. These results indicated that bile acids may activate the FXR/NF-κB signalling pathway, thereby upregulating CDX2 and MUC2 expression in normal gastric epithelial cells.