Search

Colletotrichum fructicola is a globally significant phytopathogenic fungus. Mycovirus-induced hypovirulence has great potential for biological control and study of fungal pathogenic mechanisms. We previously reported that the mycovirus Colletotrichum alienum partitivirus 1 (CaPV1) is associated with the hypovirulence of C. fructicola , and the present study aimed to further investigate a host factor and its roles in mycovirus-induced hypovirulence. A gene named CfKOB1 , which encodes putative protein homologous to the β-subunit of voltage-gated potassium channels and aldo–keto reductase, is downregulated upon CaPV1 infection and significantly upregulated during the early infection phase of Nicotiana benthamiana by C. fructicola. Deleting the CfKOB1 gene resulted in diminished vegetative growth, decreased production of asexual spores, hindered appressorium formation, reduced virulence, and altered tolerance to abiotic stresses. Transcriptome analysis revealed that CfKOB1 regulates many metabolic pathways as well as the cell cycle and apoptosis. Furthermore, enhanced apoptosis was observed in the Δ CfKOB1 mutants. Viral RNA accumulation was significantly increased in the CfKOB1 deletion mutant. Additionally, our findings demonstrated that CaPV1 infection in the WT strain also induced cell apoptosis. Collectively, these results highlight the diverse biological roles of the CfKOB1 gene in the fungus C. fructicola , while it also participates in mycovirus-induced hypovirulence by regulating apoptosis. • A gene named CfKOB1 was confirmed to regulate development and pathogenicity of C. fructicola. • The CfKOB1 gene is downregulated upon mycovirus CaPV1 infection. • Deletion of CfKOB1 enhances cell apoptosis of C. fructicola. • CfKOB1 plays a negative role in viral RNA accumulation in the CaPV1 infected host fungus. • CfKOB1 associated cell apoptosis participates in mycovirus-induced hypovirulence in C. fructicola. [ABSTRACT FROM AUTHOR]

Increased levels of interleukin‐6 (IL‐6) contribute to the development of polycystic ovary syndrome (PCOS); in renal cell carcinoma, the long non‐coding RNA (lncRNA) SRLR upregulates IL‐6. In this study, we demonstrated that the levels of the lncRNA SRLR were upregulated in PCOS patients with high expression of plasma IL‐6 compared with heathy females. The levels of the lncRNA SRLR in the plasma had a positive correlation with expression of IL‐6 in patients with PCOS but not in healthy females. Upregulation of the lncRNA SRLR in plasma could distinguish PCOS patients from healthy females. Overexpression of the lncRNA SRLR led to upregulation of IL‐6 and promoted apoptosis of human granulosa‐like tumour cells (KGN). Therefore, the lncRNA SRLR participated in PCOS by regulating cell apoptosis and IL‐6 expression. Significance of the Study: The lncRNA SRLR mediates its effects on apoptosis and IL‐6 expression in PCOS and could be used to distinguish PCOS patients from healthy controls. Plasma circulating levels of the lncRNA SRLR may be a potential target for the treatment of PCOS. [ABSTRACT FROM AUTHOR]

Probiotics are known to be a potential agent for colorectal cancer (CRC) inhibition, but the precise mechanisms by which probiotic exert anti-tumorigenic effects remain to be explored. Lactobacillus salivarius (LS) Ren was isolated from centenarians living in Bama of China, which showed an anticancer potent in animal model of oral cancer. Here, we investigated the effect of LS on colorectal carcinogenesis and its putative mechanism. Oral administration of LS effectively suppressed the formation of dimethylhydrazine (DMH)-induced CRC in both initial and post-initial stages. Significant antiproliferation and proapoptotic effects were observed with inhibition of tumor formation by dietary intake of LS. Besides, LS metabolites inhibited growth, arrested cell cycle, and induced apoptosis of HT-29 cells. Furthermore, upon the treatment of LS, protein kinase B (AKT) phosphorylation and the downstream proteins of cyclinD1 and cyclooxygenase-2 (COX-2) were significantly downregulated in both in vivo and in vitro tests. These results showed that LS inhibited the colorectal carcinogenesis through suppressing AKT signaling pathway, resulting in suppressing cell proliferation and inducing cell apoptosis. Our findings suggest that this probiotic may act as a prophylactic agent for CRC prevention. Key points • LS effectively prevented rat colorectal carcinogenesis induced by DMH. • LS modulated the proliferation and apoptosis in both in vivo and in vitro. • LS inhibited AKT phosphorylation and expressions of downstream cyclinD1 and COX-2. [ABSTRACT FROM AUTHOR]

The aim of this article is to study the effect of miR-486-5p in acute lung injury (ALI). MiR-486-5p expression in peripheral blood was determined in ALI patients and healthy volunteers by qRT-PCR. ALI mouse model were reproduced by LPS treatment, and miR-486-5p NC and miRNA-486 inhibitors were injected through trachea. ALI patients' peripheral blood and LPS-induced acute lung injury in mice had significantly higher miR-486-5p levels than control subjects. Inhibition of miR-486-5p by injection with antagomiR-486-5p markedly reduced LPS-induced lung inflammation. Moreover, knockdown of miR-486-5p can reduce protects A549 cell against LPS-induced injury and its corresponding inflammatory response. In addition, Mechanistic analysis indicated that miR-486-5p on the occurrence of ALI is related to the inhibition of OTUD7B activity, which induces the downregulation of inflammatory in ALI. Our results identified miR-486-5p independently associated with ALI. miR-486-5p can mediate the formation of ALI by promoting inflammation. [ABSTRACT FROM AUTHOR]

Background Ursolic acid (UA), a plant extract used in traditional Chinese medicine, exhibits potential anticancer effects in various human cancer cell lines in vitro. In the present study, we evaluated the anti-tumoral properties of UA against gallbladder carcinoma and investigated the potential mechanisms responsible for its effects on proliferation, cell cycle arrest and apoptosis in vitro. Methods The anti-tumor activity of UA against GBC-SD and SGC-996 cells was assessed using MTT and colony formation assays. An annex in V/PI double-staining assay was used to detect cell apoptosis. Cell cycle changes were detected using flow cytometry. Rhodamine 123 staining was used to assess the mitochondrial membrane potential (ΔΨm) and validate UA's ability to induce apoptosis in both cell lines. The effectiveness of UA in gallbladder cancer was further verified in vivo by establishing a xenograft GBC model in nude mice. Finally, the expression levels of cell cycle- and apoptosis-related proteins were analyzed by western blotting. Results Our results suggest that UA can significantly inhibit the growth of gallbladder cancer cells. MTT and colony formation assays indicated dose-dependent decreases in cell proliferation. S-phase arrest was observed in both cell lines after treatment with UA. Annex in V/PI staining suggested that UA induced both early and late phases of apoptosis. UA also decreased ΔΨm and altered the expression of molecules regulating the cell cycle and apoptosis. In vivo study showed intraperitoneally injection of UA can significantly inhibited the growth of xenograft tumor in nude mice and the inhibition efficiency is dose related. Activation of caspase-3,-9 and PARP indicated that mitochondrial pathways may be involved in UA-induced apoptosis. Conclusions Taken together, these results suggest that UA exhibits significant anti-tumor effects by suppressing cell proliferation, promoting apoptosis and inducing 7cell cycle arrest both in vitro and in vivo. It may be a potential agent for treating gallbladder cancer. [ABSTRACT FROM AUTHOR]

Changes of a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor expression could impacts the viability of neurons and brain levels of brain-derived neurotrophic factor (BDNF) expression in the key brain structures in the pathophysiology of depressive disorder. In the present study, chronic unpredictable mild stress (CUMS) degraded performance decreased AMPA receptor expression and increased neuron apoptosis. Treatment with baicalin (20, 40 mg/kg) significantly reversed these changes. This study demonstrates that baicalin has potent antidepressant effect, likely through up-regulated the expression of AMPA receptor and suppression neuron apoptosis in CUMS-treated rats. [ABSTRACT FROM AUTHOR]

Gallbladder carcinoma is the most common malignancy of the biliary tract, with a very low 5-year survival rate and extremely poor prognosis. Thus, new effective treatments and drugs are urgently needed for the treatment of this malignancy. In this study, for the first time we investigated the effects of triptolide on gallbladder cancer cells and identified the mechanisms underlying its potential anticancer effects. The MTT assay showed that triptolide decreased cell viability in a dose- and time-dependent manner. The results of the colony formation assay indicated that triptolide strongly suppressed colony formation ability in GBC-SD and SGC-996 cells. Flow cytometric analysis revealed that triptolide induced S phase arrest in gallbladder cancer cells. In addition, triptolide induced apoptosis, as shown by the results of annexin V/propidium iodide double-staining and Hoechst 33342 staining. Furthermore, triptolide decreased mitochondrial membrane potential (Δψm) in a dose-dependent manner. Finally, western blot analysis of triptolide-treated cells revealed the activation of caspase-3, caspase-9, PARP, and Bcl-2; this result demonstrated that triptolide induced apoptosis in gallbladder cancer cells by regulating apoptosis-related protein expression, and suggests that triptolide may be a promising drug to treat gallbladder carcinoma. [ABSTRACT FROM AUTHOR]

Background: Gallbladder carcinoma is a malignant tumor with a very low 5-year survival rate because of the difficulty with its early diagnosis and the very poor prognosis of the advanced cancer state. The aims of this study were to determine whether curcumin could induce the apoptosis of a gallbladder carcinoma cell line, GBC-SD, and to clarify its related mechanism. Methods: First, the anti-proliferative activities of curcumin-treated and untreated GBC-SD cells were determined using the MTT and colony formation assays. Then, the early apoptosis of cells was detected by the annexin V/ propidium iodide double-staining assay and Hoechst 33342 staining assay. Detection of mitochondrial membrane potential was used to validate the ability of curcumin on inducing apoptosis in GBC-SD cells. Cell cycle changes were detected by flow cytometric analysis. Finally, the expressions of the apoptosis-related proteins or genes caspase-3, PARP, Bcl-2, and Bax were analyzed by western blot and quantitative real time PCR assay. Statistical analyses were performed using the Student’s t-test for comparison of the results obtained from cells with or without curcumin treatment. Results: The MTT assay revealed that curcumin had induced a dose- and a time-dependent decrease in cell viability. Colony counting indicated that curcumin had induced a dose-dependent decrease in the colony formation ability in GBC-SD cells. Cells treated with curcumin were arrested at the S phase, according to the flow cytometric analysis. A significant induction of both the early and late phases of apoptosis was shown by the annexin V-FITC and PI staining. Morphological changes in apoptotic cells were also found by the Hoechst 33342 staining. After treatment with curcumin fluorescence shifted from red to green as ΔΨm decreased. Furthermore, western blot and quantitative real time PCR assays demonstrated that the curcumin induced apoptosis in GBC-SD cells by regulating the ratio of Bcl-2/Bax and activating the expression of cleaved caspase-3. Conclusions: Taken together, the results indicate that curcumin may be a potential agent for the treatment of gallbladder cancer. [ABSTRACT FROM AUTHOR]

Aim: The present study aimed to explore the antitumor effect and action mechanism of cucurbitacin B (CuB) on human T-cell leukemia Jurkat cells. Methods: Cell proliferation was measured by the MTS assay. Cell cycle distribution, mitochondrial membrane potential and annexin V staining were analyzed using flow cytometry. Western blotting was used to determine the levels of apoptosis- and autophagy-related proteins. Results: CuB inhibited the proliferation of Jurkat cells in a dose-dependent manner and induced G2/M phase arrest as well as formation of tetraploid cells. Accompanied with these effects, the actin dynamics was disrupted, and cofilin, a key regulator of actin dynamics, was persistently activated (dephosphorylated). Although CuB induced around 10% cells undergoing apoptosis, most of the cells were alive after CuB treatment for 24 h. Induction of autophagy was also evident by accumulation of LC3-II. CuB-induced autophagy seemed to be a prosurvival response, since suppression of CuB-induced autophagy significantly increased the activation of caspase-3. Conclusion: Our results demonstrated that CuB exhibited antitumor activity in Jurkat cells through induction of cell cycle arrest and apoptosis which was at least partly due to the disruption of actin dynamics. Copyright © 2012 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

In anther development, tapetal cells take part in complex processes, including endomitosis and apoptosis (programmed cell death). The tapetum provides many of the proteins, lipids, polysaccharides and other molecules necessary for pollen development. Several transcription factors, including DYT1, TDF1, AMS, MS188 and MS1, have been reported to be essential for tapetum development and function in Arabidopsis thaliana. Here, we present a detailed cytological analysis of knockout mutants for these genes, along with an in situ RNA hybridization experiment and double mutant analysis showing that these transcription factors form a genetic pathway in tapetum development. DYT1, TDF1 and AMS function in early tapetum development, while MS188 and MS1 are important for late tapetum development. The genetic pathway revealed in this work facilitates further investigation of the function and molecular mechanisms of tapetum development in Arabidopsis. [ABSTRACT FROM AUTHOR]

Abstract: Integrin-linked kinase (ILK), an intracellular serine/threonine kinase, is implicated in cell growth and survival, cell-cycle progression, tumor angiogenesis, and cell apoptosis. Recent studies showed that the expression and activity of ILK increased significantly in many types of solid tumors. However, the exact molecular mechanism of ILK underlie tumor has not been fully ascertained. The purpose of our study was to determine whether knockdown of ILK would inhibit cell growth and induce apoptosis in bladder cancer cells using a plasmid vector based small interfering RNA (siRNA). The experiments showed that knockdown of ILK could remarkably inhibit cell proliferation and growth, regulate cell cycle and induce apoptosis of bladder cancer BIU-87 and EJ cells. We demonstrated that knockdown of ILK inhibited phosphorylation of downstream signaling targets protein kinase B/Akt, glycogen synthase kinase 3-beta (GSK-3β), and reduced expression of β-catenin in BIU-87 as well as EJ cells by Western blot and Immunofluorescence analysis. In addition, down-regulation of ILK also could increase expression of Ribonuclease inhibitor (RI), an important acidic cytoplasmic protein with many functions. BALB/C nude mice injected with the BIU-87 cells transfected ILK siRNA showed a significant inhibition of the tumor growth with lighter tumor weight, lower microvessels density and higher apoptosis rate than those in the other two control groups. In conclusion, these results suggest that ILK might be involved in the development of bladder cancer, and could be served as a novel potential therapy target for human bladder cancer. Our study may be of biological and clinical importance. [Copyright &y& Elsevier]

Diabetes mellitus (DM) is a widespread metabolic disease presenting with various complications, including spermatogenic dysfunction. However, the underlying mechanisms are still unclear. Ferroptosis, a novel type of programmed cell death, is associated with much metabolic diseases. Here, we investigated the role of ferroptosis in spermatogenic dysfunction of streptozotocin (STZ)-induced type 1 diabetic mice (diabetic mice), high glucose (HG)-treated GC-2 cells (HG cells) as well as testicular tissues of diabetic patients. We found an accumulation of iron, elevated malondialdehyde level and reduced glutathione level in the testis tissues of diabetic mice and HG cells. Histological examination showed a decrease in spermatogenic cells and spermatids within the seminiferous tubules as well as mitochondrial shrinkage in the testis tissues of diabetic mice. Ferrostatin-1 (Fer-1), the inhibitor of ferroptosis, mitigated ferroptosis-associated iron overload, lipid peroxidation accumulation and spermatogenic dysfunction of diabetic mice. Furthermore, we observed a downregulation of GPX4, FTL and SLC7A11 in diabetic mice and HG cells. Fer-1 treatment and GPX4 overexpression counteracted the effects of HG on cell viability, reactive oxygen species, lipid peroxidation and glutathione via inhibition of ferroptosis. Moreover, we found an elevation of ferroptosis in testicular tissues of diabetic patients. Taken together, our results identify the crucial role of ferroptosis in diabetic spermatogenic dysfunction and ferroptosis may be a promising therapeutic target to improve spermatogenesis in diabetic patients. [Display omitted] • Increased ferroptosis caused spermatogenic dysfunction in type 1 diabetic mice and patients. • Ferrostatin-1 treatment alleviated spermatogenic dysfunction in diabetic mice. • Ferrostatin-1 treatment and GPX4 overexpression attenuated ferroptosis in high glucose treated GC-2 cells. • Inhibition of ferroptosis may be a potential therapy to alleviate diabetic male infertility. [ABSTRACT FROM AUTHOR]

Methylmercury (MeHg) is a ubiquitous environmental toxicant with neurotoxic effects. Although its neurotoxicity had been more studied, the role of gut microbiota remains unclear. In this study, adult zebrafish and larvae were exposed to MeHgCl at the dose of 0, 1 and 10 ng/mL. MeHgCl exposure impaired the locomotor activity via upregulation of apoptosis and autophagy related genes in the brain. Intestinal and cerebral metabolome indicated that phosphatidylinositol signaling system and inositol phosphate metabolism pathways were significantly impacted in adult zebrafish upon MeHgCl exposure. The levels of myo-inositol (MI) in the intestine and brain were decreased and positively correlated. 16 S rRNA sequencing data from adult zebrafish showed that MeHgCl exposure also shifted the structure of gut microbiota and reduced the relative abundance of Bacteroidetes and Proteobacteria , which were further identified at genus level as Aeromonas and Cetobacterium. Further functional analysis indicated that MeHgCl disrupted inositol phosphate metabolism of gut microbiota. Notably, MI supplementation restored the impairment of locomotor activity and inhibited the upregulation of apoptosis and autophagy related genes, such as bcl-2 and atg5. Thus, this study not only revealed the key role of gut microbiota in MeHgCl-mediated neurotoxicity but also gave new insights into antagonizing its toxicity. • MeHgCl exposure impaired the locomotor activity in adult zebrafish and larvae. • MeHgCl exposure upregulated apoptosis and autophagy related genes in the brain. • MeHgCl exposure decreased the levels of myo-inositol in the intestine and brain. • MeHgCl shifted the structure of gut microbiota and inositol phosphate metabolism. • Myo-inositol supplementation restored MeHgCl-induced neurotoxicity. [ABSTRACT FROM AUTHOR]

Endometrial cancer (EC) is a common gynecological malignancy worldwide whose therapy mainly depends on chemotherapy. In past years, an increasing number of studies indicate that hollow MnO2 could serve as a nanoplatform in the drug delivery system. The Brucea javanica oil emulsion (BJOE) has been illustrated to play a vital role in cancers. However, knowledge about the combined effect of H-MnO2-PEG/BJOE in endometrial cancer remains ambiguous up to now. In the present work, we prepared a drug-delivery vector H-MnO2-PEG by chemical synthesis and found that H-MnO2-PEG significantly inhibited cell proliferation in endometrial cancer cells. Moreover, the combination of H-MnO2-PEG/BJOE could repress cell proliferation more efficiently and promote cell apoptosis. Mechanistically, we found that BJOE exerted its role as a promoter of endometrial apoptosis by regulating relative protein expressions. In general, the present study demonstrates that H-MnO2-PEG functions as a critical vector in the tumor microenvironment of endometrial cancer and the significant effect of H-MnO2-PEG/BJOE on cancer cells, suggesting a new paradigm for the treatment of endometrial cancer. [ABSTRACT FROM AUTHOR]

Acute myeloid leukemia (AML) is an aggressive blood cancer with poor clinical outcomes. Emerging data suggest that mitochondrial oxidative phosphorylation (mtOXPHOS) plays a significant role in AML tumorigenesis, progression, and resistance to chemotherapies. However, how the mtOXPHOS is regulated in AML cells is not well understood. In this study, we investigated the oncogenic functions of ERRα in AML by combining in silico, in vitro, and in vivo analyses and showed ERRα is a key regulator of mtOXPHOS in AML cells. The increased ERRα level was associated with worse clinical outcomes of AML patients. Single cell RNA-Seq analysis of human primary AML cells indicated that ERRα-expressing cancer cells had significantly higher mtOXPHOS enrichment scores. Blockade of ERRα by pharmacologic inhibitor (XCT-790) or gene silencing suppressed mtOXPHOS and increased anti-leukemic effects in vitro and in xenograft mouse models. [ABSTRACT FROM AUTHOR]

Antiproliferative effects of 15 sulfides were investigated in human leukemia Jurkat cells. Treatment with 5–50 μM of nine monosulfides and two linear disulfides did not induce DNA fragmentation. Whereas, furan-containing sulfur flavors including methyl 2-methyl-3-furyl disulfide (MMFDS), bis (2-methyl-3-furyl) disulfide (BMFDS), methyl furfuryl disulfide (MFDS) and difurfuryl disulfide (DFDS) induced DNA fragmentation to a varying extent in Jurkat cells. The cell viability-reduction effect of these sulfur flavors was in the following order: DFDS > BMFDS > MMFDS>>MFDS based on the IC 50 values. MMFDS and BMFDS, but not DFDS, significantly increased the intracellular ROS level by 1.90- and 3.02-fold, respectively. Addition of N -acetylcysteine (NAC) or glutathione (GSH) partially suppressed induction of DNA fragmentation, apoptosis and caspase-3 activation by MMFDS and BMFDS. These results suggest that the furan-containing disulfides have a strong antiproliferative effect, and the oxidative stress and subsequent caspase-3 activation are involved in antiproliferative effect induced by MMFDS and BMFDS in Jurkat cells. [ABSTRACT FROM AUTHOR]

Summary: Pollen protects male sperm and allows flowering plants to adapt to diverse terrestrial environments, thereby leading to the rapid expansion of plants into new regions. The process of anther/pollen development is coordinately regulated by internal and external factors including hormones. Currently, the molecular mechanisms underlying gibberellin (GA)‐mediated male reproductive development in plants remain unknown.We show here that rice DELLA/SLR1, which encodes the central negative regulator of GA signaling, is essential for rice anther development. The slr1‐5 mutant exhibits premature programmed cell death of the tapetum, lacks Ubisch bodies, and has no exine and no mature pollen. SLR1 is mainly expressed in tapetal cells and tetrads, and is required for the appropriate expression of genes encoding key factors of pollen development, which are suggested to be OsMS188‐targeted genes. OsMS188 is the main component in the essential genetic program of tapetum and pollen development.Further, we demonstrate that SLR1 interacts with OsMS188 to cooperatively activate the expression of the sporopollenin biosynthesis and transport‐related genes CYP703A3, DPW, ABCG15 and PKS1 for rapid formation of pollen walls.Overall, the results of this study suggest that the GA hormonal signal is integrated into the anther genetic program and regulates rice anther development through the GA–DELLA–OsMS188 module. [ABSTRACT FROM AUTHOR]

Abstract: Suberoylanilide hydroxamic acid (SAHA), a histone deacetylase inhibitor, has been proven to be an anti-cancer agent. Its anti-inflammatory activities have recently been observed both in in vitro and in vivo models. Yet its action on lymphocytes and the underlying mechanism are still not well known. In this study, in order to evaluate the anti-inflammatory function of SAHA, we analyzed the effects of SAHA on the proliferation, activation, cytokines secretion, cell cycle distribution and apoptosis of murine lymphocytes activated with concanavalin A (Con A). Our results demonstrated that SAHA inhibited the proliferation of Con A-activated lymphocytes in a dose-dependent manner. The expression of CD69 on CD3+ T lymphocytes was significantly inhibited by SAHA. Intracellular cytokine staining analysis showed that SAHA could downregulate the expression of pro-inflammatory cytokines TNF-α, IL-6 and IFN-γ in T lymphocytes. Furthermore, analysis of sub-G0/G1 peaks and annexin V binding populations revealed that SAHA induced apoptotic cell death in Con A-activated lymphocytes. Consistent with these results, SAHA treatment also induced a decrease of mitochondrial membrane potential and cleavage of caspase-3 and PARP in these cells. Moreover, SAHA caused an accumulation of phosphorylated histone H2A.X, indicating increased double strand DNA breaks. These findings suggest that induction of apoptosis through the mitochondrial pathway may contribute to the anti-inflammatory activities of SAHA on activated lymphocytes. [Copyright &y& Elsevier]

Abstract: In this study, cytotoxic effects of structurally related flavones and flavonols on a human esophageal squamous cell carcinoma cell line (KYSE-510) were determined, and the molecular mechanisms responsible for their cytotoxic effects were studied. The results of MTT assay showed that flavones (luteolin, apigenin, chrysin) and flavonols (quercetin, kaempferol, myricetin) were able to induce cytotoxicity in KYSE-510 cells in a dose- and time-dependent manner, and the cytotoxic potency of these compounds was in the order of: luteolin>quercetin>chrysin>kaempferol>apigenin>myricetin. Flow cytometry and DNA fragmentation analysis indicated that the cytotoxicity induced by flavones and flavonols was mediated by G2/M cell cycle arrest and apoptosis. Furthermore, the expression of genes related to cell cycle arrest and apoptosis was assessed by oligonucleotide microarray, real-time RT-PCR and Western blot. It was shown that the treatment of KYSE-510 cells with these compounds caused G2/M arrest through up-regulation of p21waf1 and down-regulation of cyclin B1 at the mRNA and protein levels, and induced p53-independent mitochondrial-mediated apoptosis through up-regulation of PIG3 and cleavage of caspase-9 and caspase-3. The results of western blot analysis further showed that increases of p63 and p73 protein translation or stability might be contributed to the regulation of p21waf1, cyclin B1 and PIG3. [Copyright &y& Elsevier]

Abstract: Dietary flavonoids have been shown to exert specific cytotoxicity towards some cancer cells, but the precise molecular mechanisms are still not completely understood. In our study, cytotoxic effects of structurally related flavones and flavonols on a human oesophageal adenocarcinoma cell line (OE33) were compared, and the molecular mechanisms responsible for their cytotoxic effects were explored. The results of MTT assay showed that flavones (luteolin, apigenin, chrysin) and flavonols (quercetin, kaempferol, myricetin) were all able to induce cytotoxicity in OE33 cells in a dose- and time-dependent manner, and the cytotoxic potency of these compounds was in the order of quercetin>luteolin>chrysin>kaempferol>apigenin>myricetin. Flow cytometry and DNA fragmentation analysis indicated that the cytotoxicity induced by flavones and flavonols was mediated by G2/M cell cycle arrest and apoptosis. Furthermore, the expression of genes related to cell cycle arrest and apoptosis was assessed by oligonucleotide microarray, real-time RT-PCR and Western blot. It was found that the treatment of OE33 cells with flavones and flavonols caused G2/M arrest through up-regulation of GADD45β and 14-3-3σ and down-regulation of cyclin B1 at the mRNA and protein levels, and induced p53-independent mitochondrial-mediated apoptosis through up-regulation of PIG3 and cleavage of caspase-9 and caspase-3. The results of western blot analysis further showed that increases of p63 and p73 protein translation or stability might be contribute to the regulation of GADD45β, 14-3-3σ, cyclin B1 and PIG3. [Copyright &y& Elsevier]

Promyelocytic leukaemia nuclear bodies (PML NBs) are structured protein complexes associated with the nuclear matrix. PML constitutes the scaffold component of NBs and recruits onto these domains a striking variety of proteins, many of which are involved in apoptosis control. Several reports have directly implicated PML in apoptosis and senescence, but the mechanisms by which these are conveyed are still largely unsettled. Recruitment of partner proteins onto NBs is regulated by PML sumolation, a specific post-translational modification also found in many NB-associated proteins. Among these, several are implicated in transcription repression or activation, like the transcriptional repressor Daxx or the transcriptional activator P53. Whether NBs constitute platforms where active sites of enzymatic modifications are carried out, as suggested for P53, sites of intranuclear protein sequestration, as proposed for Daxx or organelles specialized in catabolism, is still debated. A variety of stress-related signalling pathways dramatically modulate the formation of PML NBs, which may provide a clue as to their physiological function.Oncogene (2004) 23, 2819-2824. doi:10.1038/sj.onc.1207533 [ABSTRACT FROM AUTHOR]

miR-92a miRNAs are immune molecules that regulate apoptosis (programmed cell death) during the immune response. Apoptosis helps to maintain the dynamic balance in tissues of fish under hypoxia stress. The aim of this study was to explore the role and potential mechanisms of miR-92a in the liver of tilapia under hypoxia stress. We first confirmed that CaSR (encoding a calcium-sensing receptor) is a target gene of miR-92a in genetically improved farmed tilapia (GIFT) using luciferase reporter gene assays. In GIFT under hypoxia stress, miR-92a was up-regulated and CaSR was down-regulated in a time-dependent manner. Knocked-down CaSR expression led to inhibited expression of p53, TP53INP1, and caspase-3/8, reduced the proportion of apoptotic hepatocytes, and decreased the activity of calcium ions induced by hypoxia in hepatocytes. GIFT injected in the tail vein with an miR-92a agomir showed up-regulation of miR-92a and down-regulation of CaSR, p53, TP53INP1, and caspase-3/8 genes in the liver, resulting in lower serum aspartate aminotransferase and alanine aminotransferase activities under hypoxia stress. These findings suggest that stimulation of miR-92a interferes with hypoxia-induced apoptosis in hepatocytes of GIFT by targeting CaSR, thereby alleviating liver damage. These results provide new insights into the adaptation mechanisms of GIFT to hypoxia stress. [ABSTRACT FROM AUTHOR]

p53 and mouse double minute 2 homolog (MDM2) serve key regulatory roles in the apoptosis of synovial cells. The present study aimed to investigate the effects of electroacupuncture (EA) at the 'Zusanli' (ST36) and 'Xuanzhong' (GB39) acupoints on apoptosis in an adjuvant arthritis (AA) rat model. A total of 40 male Sprague-Dawley rats were randomly divided into Control, AA, AA + EA and AA + sham EA groups (n=10 rats in each group). Rats in all the groups, with the exception of the control group, were injected with Complete™ Freund's adjuvant into the bilateral hindlimb footpad to establish the AA model. Rats in the AA + EA group were treated with EA at the ST36 and GB39 acupoints. Rats in the AA + sham EA group were treated with percutaneous electrical stimulation at a position of 5 mm away from the ST36 and GB39 acupoints. The arthritis index scores and hindlimb paw volumes of the rats in each group were recorded. Subsequently, pathological changes in the synovial tissue were evaluated by hematoxylin and eosin (H&E) staining, and the apoptotic rate of the synovial cells was detected by TUNEL staining. In addition, the expression levels of the apoptosis-associated proteins, Bax, phorbol-12-myristate-13-acetate-induced protein 1 (Noxa) and p53 upregulated modulator of apoptosis (PUMA), were determined by western blot analysis. The expression of both the gene and protein of p53 and MDM2 in synovial tissue was detected by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis, respectively. The results indicated that the arthritis index scores and hindlimb paw volumes upon EA stimulation were significantly decreased compared with those of the AA group (P<0.05). H&E staining revealed that the synovial inflammation of EA stimulation was significantly decreased compared with the AA group (P<0.05). The TUNEL assay results indicated that the apoptotic rate of synovial cells in the AA + EA group was significantly increased compared with that in the AA group (P<0.05). Furthermore, an increased expression of proapoptotic proteins was confirmed by the increased expression levels of Bax, Noxa and PUMA in the AA + EA group. The results of RT-qPCR and western blot analysis demonstrated that, compared with the AA group, EA stimulation led to a marked increase in p53 (P<0.05) and a significant decrease in MDM2 (P<0.05) gene and protein expression. Taken together, these results demonstrated that EA performed on the ST36 and GB39 acupoints led to a significant amelioration in AA injury of model rats, by regulating the p53 signaling pathway and inducing apoptosis. [ABSTRACT FROM AUTHOR]

Artemisinin (Qinghaosu), a sesquiterpene lactone isolated from the Chinese medicinal herb Artemisia annua, is famous as the first‐line antimalarial drug, and reported to also possess anticancer properties. Though many studies have proceeded focusing on artemisinin and bioactive derivatives in clinic or laboratory, their cellular targets and mechanism of actions have still remained obscure. Here, two new 11‐aza‐artemisinin derivatives AAD1 and AAD2, which displayed cytotoxity against multiple cancer cell lines, were synthesized and applied as the probes for target identification and anticancer mechanism research. Using T7 phage display screen method, fatty acid binding protein 6 (FABP6) was identified and verified as one of the potential targets. Further studies showed FABP6 is a direct binding protein of AAD1 and AAD2, and mediated the corresponding anticancer activities of these artemisinin derivatives. Two new anticancer 11‐aza‐artemisinin derivatives AAD1 and AAD2 with terminal amino groups were successfully synthesized, evaluated and applied as the probes for the study of target identification and mechanism of action. FABP6 potein was identified and further confirmed as one target of these azaartemisinin derivatives. [ABSTRACT FROM AUTHOR]

The accurate therapeutic evaluation for chronic myeloid leukemia (CML) drug is of great importance to minimize side effects and enhance efficacy. Herein, a facile and precise surface-enhanced scattering (SERS) approach based on coupled plasmonic field has been introduced to evaluate the therapeutic outcomes of antileukemia drug through ultrasensitive assay of caspase-3 activity in apoptotic cells. Caspase-3 as an apoptosis indicator could specifically cleave the N-terminus of biotinylated DEVD-peptide (biotin-Gly-Asp-Gly-Asp-Glu-Val-Asp-Gly-Cys) immobilized on the Au nanoparticle-decorated TiO 2 nanotube arrays (TiO 2 /Au NTAs) substrate. After the enzyme cleavage with caspase-3, Raman-labelled Au nanostar (AuNS) probes captured the residual DEVD-peptides via the recognition between streptavidin and biotin, thus resulting in an enhanced Raman response on the SERS platform. The variation of Raman intensity revealed caspase-3 activity that reflected the chemotherapeutic effect. On this platform, AuNS nanoprobes offered a large number of binding sites and intrinsic “hot spots” for Raman reporters, while TiO 2 /Au NTAs rendered a homogenously coupled electromagnetic field between the adjacent repeated units over the large area. In particular, a spatially expanding plasmonic field formed by coupling AuNSs with TiO 2 /Au NTAs would further heighten Raman enhancement. Taking these advantages, the strong and uniform Raman signals were achieved. Furthermore, the practicability investigation witnessed that the proposed SERS strategy was available to evaluate the therapeutic effect of dasatinib on CML K562 cells. The developed method possesses fascinating advantages of cost-effectiveness, excellent reproducibility and high sensitivity, which endows it with promising potential in apoptosis monitoring and anticancer drug development. [ABSTRACT FROM AUTHOR]

Receptor for activated protein kinase C 1 (RACK1) is a multifaceted scaffolding protein known to be involved in the regulation of signaling events required for neuronal protection. In the present study, we investigated the role of RACK1 in secondary brain injury in a rat traumatic brain injury (TBI) model. A weight-drop TBI model was established in Sprague Dawley rats, and RACK1 in vivo knockdown and overexpression were performed 24 h before TBI insult. The IRE1 inhibitor 3,5-dibromosalicylaldehyde (DBSA) was administered by intracerebroventricular injection 1 h after TBI insult. Real-time PCR, Western blotting, immunofluorescence, neuronal apoptosis, brain water content, and neurological scores were evaluated. Our results revealed that TBI induced increased expression of endogenous RACK1, phosphorylated inositol-requiring enzyme 1 (p-IRE1), X-box binding protein-1 (XBP1) and glucose-regulated protein 78 (GRP78) in neurons. RACK1 overexpression significantly ameliorated neuronal apoptosis, blood-brain barrier disruption, brain edema and neurological deficits at 48 h after TBI, which was concomitant with upregulation of p-IRE1, XBP1 and GRP78 expression, while its knockdown induced the opposite effects. Furthermore, DBSA administration reversed the protective effects of RACK1 overexpression against brain injury and decreased the expression of p-IRE1, XBP1 and GRP78. In summary, the upregulation of RACK1 following brain contusion exerted neuroprotective effects against secondary brain injury, which were probably mediated by activation of the IRE1-XBP1 pathway. [ABSTRACT FROM AUTHOR]

Gastric cancer most commonly occurs in East Asia, and China accounts for more than half the of world's gastric cancer burden. Despite the efficacy of chemotherapy for patients, this treatment leads to significant patient inconvenience, toxicity and cost. The present study aimed to assess a non-toxic agent, glabridin, as a future chemotherapeutic approach for treating gastric cancer. Using cell proliferation, apoptosis, invasion, and colony formation assays, it was determined that glabridin alone, or in combination with 5-fluorouracil (5-FU), inhibited MKN-45 cell proliferation and invasion, and increased apoptosis. These effects were accompanied by downregulation of p16, E-cadherin and apoptosis regulator Bcl-2 protein, and upregulation of N-cadherin, apoptosis regulator BAX and caspases 3, 8 and 9. The results demonstrated that glabridin may inhibit the malignant proliferation of the human gastric cancer MKN-45 cell line and enhance the efficiency of 5-FU. The data indicate that the p16, and potentially the p16/cyclin-dependent kinase 4/cyclin D1 pathway, may be a novel target for gastric cancer therapy. [ABSTRACT FROM AUTHOR]

Anthocyanins are the polyphenolic phytochemicals which have been shown to scavenge free radicals. In this study, we investigated the effects of anthocyanins extracted from red-fleshed apples (Malus sieversii) on reducing oxidative damage by Rosup in porcine granulosa cells (GCs) by measuring intracellular reactive oxygen species (ROS), content of glutathione (GSH), activities of superoxide dismutase (SOD1), catalase (CAT) and glutathione peroxidase (GPX1) and the gene expression of SOD1, CAT, GPX1. Apoptosis was determined with TdT-mediated dUTP-biotin nick end labeling (TUNEL) and apoptosis-related proteins were quantified with Western blotting. The results indicate that Rosup increases oxidative stress by inducing reactive oxygen species production in porcine GCs and the oxidative stress could be reduced by anthocyanins. The gene expression of SOD1, CAT, GPX1 and the activities of these enzymes were increased when GCs were treated with anthocyanins and Rosup for 6 hours. Anthocyanins inhibit Rosup-induced apoptosis by increasing expression of antiapoptotic protein Bcl-2 and suppressing the expression of pro-apoptotic protein Bax. Collectively, anthocyanins from red-fleshed apples reduce oxidative stress and inhibit apoptosis in porcine GCs in vitro. This approach indicates that antioxidants might be developed from red-fleshed apples. [ABSTRACT FROM AUTHOR]

Kukoamine A (KuA) is a bioactive compound, which is known for a hypotensive effect. Recent studies have shown that KuA has anti-oxidative effect and anti-apoptosis stress in vitro . However, its neuroprotective effect in rats with cerebral ischemia is still unclear. In the study, we investigated whether KuA could attenuate cerebral ischemia induced by permanent middle cerebral artery occlusion (pMCAO) in rats. Results revealed that KuA could significantly reduce infarct volume both pre-treatment and post-treatment, and increase corresponding Garcia neurological scores. Acute KuA postconditioning not only significantly reduced cerebral infarct volume, brain water content and improved neurological deficit scores, but also decreased the number of TUNEL-positive cells. Moreover, it markedly increased the activities of Cu/Zn-SOD and Mn-SOD, reduced levels of MDA and H 2 O 2. Increased expressions of caspase-3, cytochrome c and the ratio of Bax/Bcl-2 were significantly alleviated with KuA treatment. These findings demonstrated that KuA was able to protect the brain against injury induced by pMCAO via mitochondria mediated apoptosis signaling pathway. [ABSTRACT FROM AUTHOR]

Glyphosate-based herbicides (GBHs) are the most widely used agrochemicals worldwide, increasing the risk of their occurrence in the environment. This study aimed to explore effects and mechanisms of GBH exposure on placental development in vivo during pregnancy in mice. Pregnant mice received GBH by gavage at 0, 5, and 50 mg⋅kg−1⋅day−1 doses from gestational day (GD) 1 to GD 13 and were sacrificed on GD 13 or GD19. Our data indicated that GBH administration significantly increased the number of resorbed fetuses, reduced the weight of fetuses and placentas, and inhibited placental growth, as evident from decreased placental total area and spongiotrophoblast area on GD 19. GBH treatment also inhibited proliferation and induced apoptosis of placenta via upregulation of Bax, cleaved caspase-3 and -12 expression, and downregulation of B cell lymphoma (Bcl)-2 expression. Further study showed that GBH exposure significantly increased expression levels of glucose-regulated protein 78 (GRP78), protein kinase RNA-like endoplasmic reticulum kinase (PERK), and C/EBP homologous protein (CHOP) mRNAs and proteins and triggered oxidative stress in placenta on GD 13 and GD 19. In conclusion, our findings suggest that maternal exposure to GBH can impair placental development through the endoplasmic reticulum stress-mediated activation of GRP78/PERK/CHOP signaling pathway in mice. [Display omitted] • Maternal exposure to GBH increased the numbers of resorbed fetus. • GBH inhibited the placental development. • GBH induced placental apoptosis by endoplasmic reticulum stress. • GRP78/PERK/CHOP pathway was involved in GBH-induced placental impairment. [ABSTRACT FROM AUTHOR]

Twenty-one novel derivatives of millepachinewere synthesized andevaluated for their in vitro antiproliferative activity. Among them, 8exhibited the most potent activity, with IC50values of 8–27 nM against panel of cancer cell lines andretained full activity in multidrug resistant cancer cells. Treatedcells were arrested in G2/M phase and resulted in cellular apoptosis.Microtubule dynamics confirmed 8was a novel tubulinpolymerization inhibitor by binding at the colchicine site. 8also exhibited antivascular activity because it concentrationdependently reduced the cell migration and disrupted capillary liketube formation in HUVEC cells. Furthermore, the hydrochloride saltof 8(8·HCl) significantlyimproved the bioavailability up to 47% while retaining the antiproliferativeactivity. Importantly, 8·HClsignificantlyinhibited tumor growths in four xenograft models including resistancetumor-cell-bearing mice models without causing significant loss ofbody weight, suggesting that 8is a promising new orallyanticancer agent to be developed. [ABSTRACT FROM AUTHOR]

Gallbladder cancer, with high aggressivity and extremely poor prognosis, is the most common malignancy of the bile duct. The main objective of the paper was to investigate the effects of schisandrin B (Sch B) on gallbladder cancer cells and identify the mechanisms underlying its potential anticancer effects. We showed that Sch B inhibited the viability and proliferation of human gallbladder cancer cells in a dose-, time -dependent manner through MTT and colony formation assays, and decrease mitochondrial membrane potential (ΔΨm) at a dose-dependent manner through flow cytometry. Flow cytometry assays also revealed G0/G1 phase arrest and apoptosis in GBC-SD and NOZ cells. Western blot analysis of Sch B-treated cells revealed the upregulation of Bax, cleaved caspase-9, cleaved caspase-3, cleaved PARP and downregulation of Bcl-2, NF-κB, cyclin D1 and CDK-4. Moreover, this drug also inhibited the tumor growth in nude mice carrying subcutaneous NOZ tumor xenografts. These data demonstrated that Sch B induced apoptosis in gallbladder cancer cells by regulating apoptosis-related protein expression, and suggests that Sch B may be a promising drug for the treatment of gallbladder cancer. [ABSTRACT FROM AUTHOR]

Gallbladder cancer is the most common malignant tumor of the biliary tract, and this condition has a rather dismal prognosis, with an extremely low five-year survival rate. To improve the outcome of unresectable and recurrent gallbladder cancer, it is necessary to develop new effective treatments and drugs. The purpose of the present study was to evaluate the effects of cordycepin on human gallbladder cells and uncover the molecular mechanisms responsible for these effects. The Cell Counting Kit-8 (CCK-8) and colony formation assays revealed that cordycepin affected the viability and proliferation of human gallbladder cancer cells in a dose- and time-dependent manner. Flow cytometric analysis showed that cordycepin induced S phase arrest in human gallbladder cancer cell lines(NOZ and GBC-SD cells). Cordycepin-induced apoptosis was observed using an Annexin V/propidium iodide (PI) double-staining assay, and the mitochondrial membrane potential (ΔΨm) decreased in a dose-dependent manner. Additionally, western blot analysis revealed the upregulation of cleaved-caspase-3, cleaved-caspase-9, cleaved-PARP and Bax and the downregulation of Bcl-2, cyclin A and Cdk-2 in cordycepin-treated cells. Moreover, cordycepin inhibited tumor growth in nude mice bearing NOZ tumors. Our results indicate that this drug may represent an effective treatment for gallbladder carcinoma. [ABSTRACT FROM AUTHOR]

The use of nanomedicines for cancer treatment has been widespread. Fullerenes have significant effects in the treatment of solid tumors. Here, we are going to study the effects of hydroxylated fullerene C60(OH)n(n = 18–22) treatment on chronic myeloid leukemia cell proliferation and investigate its toxicity. The results showed that hydroxylated fullerene C60(OH)n (n = 18–22) at low concentrations (less than 120 μM) not only had apparent toxic side effects, but also promoted the growth of K562 cells, while a high concentration of C60(OH)n had different degrees of inhibition on K562 cells. When the concentration is higher than 160 μM, the K562 cells showed morphological changes, the mitochondrial membrane potential decreased, the cell cycle was blocked in the stage of G2-phase, and cell apoptosis occurred, which may cause apoptosis, autophagy, and a variety of other damage leading to cell death. Meanwhile, it also indicated that its inhibition of solid tumors might be related to the tumor microenvironment; we verified the safety of fullerene without apparent cellular toxicity at a specific concentration. [ABSTRACT FROM AUTHOR]

Abstract: A novel microfluidic platform integrated with a flexible PDMS-based electrochemical cytosensor was developed for real-time monitoring of the proliferation and apoptosis of HeLa cells. The PDMS-gold film, which had a conductive smooth surface and was semi-transparent, facilitated electrochemical measurements and optical microscope observations. We observed distinct increases and decreases in peak current intensity, corresponding to cell proliferation in culture medium and apoptosis in the presence of an anticancer drug, respectively. This electrochemical analysis method permitted real-time, label-free monitoring of cell behavior, and the electrochemical results were confirmed with optical microscopy. The flexible microfluidic electrochemical platform presented here is suitable for on-site monitoring of cell behavior in microenvironments. [Copyright &y& Elsevier]

Highlight: [•] Construction of shRNA segments expression vectors is valid by the investigation of RT-PCR for IGF1R, EGFR and Bcl-xl mRNA and protein expression. [•] Studies have suggested that the vectors in blocking these genes of the growth factor receptors and anti- apoptosis is capable of breaking the balance of tumor growth so that tumor trend apoptosis and senescence. [•] Simultaneously blocking multiple genes that are abnormally expressed may be more effective in treating cancer cells than silencing a single gene. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the effects of intravitreal injection of soluble erythropoietin (EPO) receptor (sEPOR) on photoreceptor cell apoptosis in an animal model of retinal detachment (RD). Methods: Various dosages of sEPOR (2, 20, or 200 ng) were injected into the vitreous cavities of normal rats. Three days after injection, retinal function was measured by flash electroretinography (ERG). On day 7, histopathology and retinal morphology were examined by light and transmission electron microscopy (TEM), respectively. Rat models of RD were successfully established by injection of 1.4% sodium hyaluronate into the subretinal space, followed by immediate injection of phosphate-buffered saline (PBS) or sEPOR into the vitreous cavity. On day 3, photoreceptor cell apoptosis was evaluated using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL), and caspase-3 activity assayed by Western blotting and immunofluorescence. Light microscopic examination of retinal histopathology was used to determine the thickness of the outer nuclear layer (ONL) 14 days after establishment of RD. Results: There were no significant differences in the latency and amplitude of maximal a, b and oscillatory potential (OP) wave responses by flash ERG before or 3 days after sEPOR injection ( p > 0.05). Retinal tissues showed no obvious pathological changes by either light or transmission electron microscopy. Both Western blotting and immunofluorescence indicated consistent sEPOR enhanced caspase-3 activation aggravated apoptosis of photoreceptor cells in RD rat retinas. On day 14, RD ONLs were thinner, according to increasing dosages of sEPOR. Conclusion: Intravitreal injection of sEPOR exacerbates photoreceptor cell apoptosis in RD models via activation of caspase-3. [ABSTRACT FROM AUTHOR]

Abstract: Tissue factor pathway inhibitor-2 (TFPI-2) is a Kunitz-type serine proteinase inhibitor with inhibitory activity toward activated factor XI, plasma kallikrein, plasmin, certain matrix metalloproteinases, and the tissue factor-activated factor VII complex. In addition, TFPI-2 has other functions such as promoting cell migration and inducing apoptosis. In the present study, we investigated if TFPI-2 induced apoptosis in cultured U937-derived macrophages and the possible signal pathways that involved in the apoptotic process. Apoptotic DNA fragment detection and caspase-3,9 activity measurements indicated that rTFPI-2 promoted U937-derived macrophage apoptosis. Hoechst 33342 assay and flow cytometry further showed that rTFPI-2 induced apoptosis in cultured macrophages in a dose-dependent manner. Because death receptors of the TNF family such as Fas are the best-understood death pathways that recruit Fas-associated death domain (FADD) and procaspase-8 to the receptor in macrophages, we investigated the expression of Fas and its ligand (FasL) and downstream signal caspase-8 by Western blot analysis. The results indicated that the process of apoptosis triggered by rTFPI-2 was, at least in part, actively conducted by U937-derived macrophages possibly through Fas/FasL signal pathway. In brief, rTFPI-2 may have the potential usefulness in inducing macrophages apoptosis, which suggest TFPI-2 might have antiatherogenic effects. [Copyright &y& Elsevier]

Simple Summary: The EZH2-targeted drugs have demonstrated notable therapeutic effects in EZH2 mutant B-cell lymphoma patients. In this study, we demonstrated that the combination of EZH2 inhibitor SHR2554 and HDAC inhibitor HBI8000 exert synergistic anti-proliferative activity in both EZH2 wide-type and mutation B-cell lymphoma. More importantly, gene expression profile analysis revealed simultaneous treatment with these agents led to dramatic inhibition of DNA replication initiator protein ORC1, which might contribute to great efficacy of combination strategy. The combination of EZH2 inhibitor and HDAC inhibitor could provide a potential therapeutic treatment for both EZH2 wide-type and mutation B-cell lymphoma patients. Background: Upregulation of H3K27me3 induced by EZH2 overexpression or somatic heterozygous mutations were implicated in lymphomagenesis. It has been demonstrated that several EZH2-target agents have notable therapeutic effects in EZH2-mutant B-cell lymphoma patients. Here we present a novel highly selective EZH2 inhibitor SHR2554 and possible combination strategy in diffuse large B-cell lymphoma (DLBCL). Methods: Cell proliferation, cell cycle and apoptosis were analyzed by CellTiter-Glo Luminescent Cell Viability Assay and flow cytometry. Western Blot was used to detect the expression of related proteins. The gene expression profiling post combination treatment was analyzed by RNA-Seq. Finally, CDX and PDX models were used to evaluate the synergistic anti-tumor effects of the combination treatment in vivo. Results: The novel EZH2 inhibitor SHR2554 inhibited proliferation and induced G1 phase arrest in EZH2-mutant DLBCL cell lines. The combination of EZH2 inhibitor SHR2554 with histone deacetylase (HDAC) inhibitor chidamide (hereafter referred to as HBI8000) exerted synergistic anti-proliferative activity in vitro and in vivo. Gene expression profile analysis revealed dramatic inhibition of the DNA replication process in combined treatment. Conclusions: SHR2554, a potent, highly selective small molecule inhibitor of EZH2, inhibited EZH2-mutant DLBCL more significantly in vitro and in vivo. The combination of HDAC inhibitor HBI8000 with EZH2 inhibitor SHR2554 exhibited dramatic anti-tumor activity in both mutant and wild-type DLBCL, which may become a potential therapeutic modality for the treatment of DLBCL patients. [ABSTRACT FROM AUTHOR]

Abstract: The aim of the study was to investigate the sensitivity of AHH-1 human lymphoblastoid cells to radiation and its relevance to intracellular events, specifically alteration in cellular energy-producing systems. AHH-1 human lymphoblastoid cells were irradiated with 6Gy of γ radiation, and then were collected at the indicated time points. Parallel studies were conducted to assess the effects of radiation on the cell proliferation and apoptotic index. Mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) production were monitored. A marked decrease of cell viability was observed as early as 12h postirradiation and fraction of apoptotic cells was highest at 24h. Intracellular ROS generation measured with 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) appeared to be highest as early as 30min postirradiation and resumed to normal level at 6h. Unexpectedly, the fluorescence intensity of Rhodamine 123 for measuring MMP did not change during the first 3h after radiation and exhibited an aberrant increase at 6h. The results suggest that AHH-1 cells are sensitive to radiation-induced apoptosis and ROS generation is an early phase in the apoptosis process. Moreover, the results might cast doubts on those studies using Rhodamine 123 which hypothesized that the fall in MMP is one of the early events of apoptosis. [Copyright &y& Elsevier]