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IRE1 is an important central regulator of unfolded protein response (UPR) in the endoplasmic reticulum (ER) because of its ability to regulate cell fate as a function of stress sensing. When misfolded proteins accumulated in chondrocytes ER, IRE1 disintegrates with BIP/GRP78 and undergoes dimer/oligomerization and transautophosphorylation. These two processes are mediated through an enzyme activity of IRE1 to activate endoribonuclease and generates XBP1 by unconventional splicing of XBP1 messenger RNA. Thereby promoting the transcription of UPR target genes and apoptosis. The deficiency of inositol‐requiring enzyme 1α (IRE1α) in chondrocytes downregulates prosurvival factors XBP1S and Bcl‐2, which enhances the apoptosis of chondrocytes through increasing proapoptotic factors caspase‐3, p‐JNK, and CHOP. Meanwhile, the activation of IRE1α increases chondrocyte viability and reduces cell apoptosis. However, the understanding of IRE1 responses and cell death fate remains controversial. This review provides updated data about the role IRE1 plays in chondrocytes and new insights about the potential efficacy of IRE1 regulation in cartilage repair and osteoarthritis treatment. [ABSTRACT FROM AUTHOR]

Neonatal respiratory distress syndrome (NRDS) is a common disease that occurs in premature infants. However, the mechanisms underlying the disease remain unclear. microRNAs (miRNAs) have been indicated to play a crucial role in the development of NRDS. In this study, we aimed to explore the regulatory mechanisms of miR-296-5p in NRDS. The expression levels of miR-296-5p in preterm infants with NRDS were determined using quantitative reverse-transcription polymerase chain reaction (RTqPCR). A549 cells were transfected with lentiviral vectors encoding miR-296-5p, and the transfection efficiency was determined using RT-qPCR. Flow cytometry and CCK8 assay were performed to measure apoptosis and proliferation of A549 cells, respectively. The protein levels of pulmonary surfactant SP-A (SFTPA1), SP-B, Wnt7b, and ß-catenin were measured using western blotting. We demonstrated an upregulation of miR-296-5p in NRDS. The miR-296-5p was successfully overexpressed in A549 cells via lentivirus transfection, and the upregulation of miR-296-5p inhibited cell proliferation and secretion of SP-A and SP-B and also induced downregulation of the Wnt7b/ß-catenin in vitro. Therefore, miR-296-5p inhibits cell proliferation and secretion of pulmonary surfactants in A549 cells via downregulation of Wnt7b/ß-catenin signaling. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the effect of Camellia nitidissima C. W. Chi extract (CNCE) on apoptosis and proliferation in A549 human lung cancer cells. Methods: Inverted microscope was used to examine morphological changes in A549 cells after exposure to CNCE. Trypan blue staining of living cells was applied to construct the cell growth curve after treatment with varying concentrations of CNCE. The influence of CNCE on cell proliferation, apoptosis and cell cycle were determined by MTT assay. The protein expressions of key apoptosisrelated enzymes were evaluated by immuno-cytochemical method. Results: CNCE inhibited the growth of A549 lung cancer cells at the concentration range of 20 - 160 μg/mL. Flow cytometry showed that CNCE induced apoptosis in the A549 cells. The proportion of cells in G0/G1-phase increased significantly (p < 0.01), while the proportion of cells in S-phase and G2/Mphase decreased correspondingly, indicating that the cells were in G0/G1-phase arrest. Cell cycle arrest and apoptosis-inducing effect gradually increased with increase in CNCE concentration. With increasing concentrations of CNCE, there were significant increases in the expressions of caspase-3 (p < 0.05), caspase-8 (p < 0.01) and caspase-9 (p < 0.05), and significant decreases in Ki-67 (p < 0.01) and p21 ras protein (p < 0.01). Conclusion: CNCE exerts significant inhibitory effect on the proliferation of A549 lung cancer cells, and therefore can potentially be developed for the treatment of lung cancer. [ABSTRACT FROM AUTHOR]

Background: Prolonged isolated thrombocytopenia (PT) is a frequent complication in patients who undergo allogeneic hematopoietic stem cell transplantation (allo-HSCT), and it is associated with an adverse prognosis. In this study, we hypothesized that desialylation on platelet surfaces was associated with PT after allo-HSCT. The mechanisms participating in this process may include NEU1 translocation, platelet apoptosis, and phagocytosis by macrophages. Methods: PT was defined as a peripheral platelet count less than 100 × 109/L without sustained anemia or leukopenia for more than 3 months after allo-HSCT. 34 patients were identified consecutively from a cohort of 255 patients who underwent allo-HSCT for hematologic malignancies between May and October 2014 at Peking University Institute of Hematology. Desialylation, enzyme expression, and phagocytosis were detected using flow cytometry, immunofluorescence, RT-PCR, Western blot, and so on. Results: Platelets from the PT patients had significantly fewer sialic acids (P = .001) and increased β-galactose exposure indicative of desialylation on the surface (P = .042), and serum from the PT patients showed a higher sialic acid concentration (8.400 ± 0.2209 μmol/L, P < .001). The sialidase NEU1 was over-expressed from mRNA to protein levels, and its catalytic activity was increased in platelets from the PT patients. Desialylation of GPIbα in the PT patients was correlated with changes in 14-3-3ζ distribution, which, relative to Bad activation, modulated the expression of Bcl-2 family proteins, depolarized the inner membrane of the mitochondria, and initiated the intrinsic mitochondria-dependent pathway of apoptosis. Macrophages derived from the THP-1 cell line preferred to phagocytize desialylated platelets from the PT patients in vitro. We also revealed that oseltamivir (400 μmol/L) could inhibit 50 % of the sialidase activity on platelets and could protect 20 % of platelets from phagocytosis in vitro. Conclusions: Desialylation of platelets was associated with platelet apoptosis and phagocytosis, whereas oseltamivir could reduce platelet destruction in the periphery, indicating a potential novel treatment for PT after allo-HSCT. [ABSTRACT FROM AUTHOR]

Background: The prognosis for pancreatic cancer (PC) is very poor. The SnoN gene may have a role in cell proliferation and apoptosis in human cancer. However, the influence of SnoN on cell proliferation and apoptosis in human PC cells remains unknown. Methods: SnoN expression was assessed in SW1990 PC cell lines using real-time polymerase chain reaction (PCR). A luciferase reporter assay was used to confirm the target associations. The effect of SnoN on cell proliferation in vitro was confirmed using Cell Counting Kit-8. Apoptosis was confirmed using flow cytometry. Gene and protein expression were examined using real time PCR and Western blotting, respectively. Results: SnoN siRNA significantly inhibited the growth of SW1990 cells by decreasing cell proliferation (P < 0.05) and increasing cell apoptosis (P < 0.05), compared with the blank group and the negative control group. The highest inhibition of cell proliferation appeared at 3 days post-transfection. Cell apoptosis more obvious at 48 h after transfection. Conclusions: In summary, our results reveal that the RNAi-mediated downregulation of SnoN effectively inhibited the proliferation of PC cells. SnoN-siRNA also enhanced SW1990 PC cell apoptosis. These findings indicate that SnoN gene plays an important role in pancreatic cancer development, and might serve as a potential therapeutic target for pancreatic cancer. However, further in vivo studies are needed to clarify the influence of SnoN gene silencing by siRNA on pancreatic cancer therapy. [ABSTRACT FROM AUTHOR]

Rapid growth and formation of new gas exchange units (alveogenesis) are hallmarks of the perinatal lung. Bronchopulmonary dysplasia (BPD), common in very premature infants, is characterized by premature arrest of alveogenesis. Mesenchymal cells (fibroblasts) regulate both lung branching and alveogenesis through mesenchymal-epithelial interactions. Temporal or spatial deficiency of late-gestation lung 1/cysteine-rich secretory protein LD2 (LGL1/CRISPLD2), expressed in and secreted by lung fibroblasts, can impair both lung branching and alveogenesis (LGL1 denotes late gestation lung 1 protein; LGL1 denotes the human gene; Lgl1 denotes the mouse/rat gene). Absence of Lgl1 is embryonic lethal. Lgl1 levels are dramatically reduced in oxygen toxicity rat models of BPD, and heterozygous Lgl1+/- mice exhibit features resembling human BPD. To explore the role of LGL1 in mesenchymal-epithelial interactions in developing lung, we developed a doxycycline (DOX)-inducible RNA-mediated LGL1 knockdown cellular model in human fetal lung fibroblasts (MRC5LGL1KD). We assessed the impact of LGL1 on cell proliferation, cell migration, apoptosis, and wound healing. DOX-induced MRC5LGL1KD suppressed cell growth and increased apoptosis of annexin V+) staining cells and caspase 3/7 activity. LGL1-conditioned medium increased migration of fetal rat primary lung epithelial cells and human airway epithelial cells. Impaired healing by MRC5LGL1KD cells of a wound model was attenuated by addition of LGL1-conditioned medium. Suppression of LGL1 was associated with dysregulation of extracellular matrix genes (downregulated MMP1, ColXVα1, and ELASTIN) and proapoptosis genes (upregulated BAD, BAK, CASP2, and TNFRSF1B) and inhibition of 44/42MAPK phosphorylation. Our findings define a role for LGL1 in fibroblast expansion and migration, epithelial cell migration, and mesenchymal-epithelial signaling, key processes in fetal lung development. [ABSTRACT FROM AUTHOR]

Liver dysfunction has been known to occur frequently in cases of sepsis. Excessive inflammation and apoptosis are pathological features of acute liver failure. Recent studies suggest that activation of glycogen synthase kinase- (GSK-) 3β is involved in inflammation and apoptosis.We aimed to investigate the protective effects of GSK-3β inhibition on polymicrobial sepsis-induced liver injury and to explore the possible mechanisms. Polymicrobial sepsis was induced by cecal ligation and puncture (CLP), and SB216763 was used to inhibit GSK-3β in C57BL/6 mice. GSK-3β was activated followingCLP.Administration of SB216763 decreased mortality, ameliorated liver injury, and reduced hepatic apoptosis. The inhibition of GSK-3β also reduced leukocyte infiltration and hepatic inflammatory cytokine expression and release.Moreover, GSK-3β inhibition suppressed the transcriptional activity of nuclear factor-kappa B (NF-κB) but enhanced the transcriptional activity of cAMP response element binding protein (CREB) in the liver. In in vitro studies, GSK-3β inhibition reduced inflammatory cytokine production via modulation of NF-κB and CREB signaling pathways in lipopolysaccharide-stimulated macrophages. In conclusion, these findings suggest that GSK-3β blockade protects against CLP-induced liver via inhibition of inflammation by modulating NF-κB and CREB activity and suppression of hepatic apoptosis. [ABSTRACT FROM AUTHOR]

β1-integrin induction enhances breast cancer cell survival after exposure to ionizing radiation (IR), but the mechanisms of this effect remain unclear. Although NF-κB initiates prosurvival signaling pathways post-IR, the molecular function of NF-κB with other key elements in radioresistance, particularly with respect to extracellular matrix-induced signaling, is not known. We discovered a typical NF-κB-binding site in the β1-integrin promoter region, indicating a possible regulatory role for NF-κB. Using three-dimensional laminin-rich extracellular matrix (3D lrECM) culture, we show that NF-kB is required for β1-integrin transactivation in T4-2 breast cancer cells post-IR. Inhibition of NF-kB reduced clonogenic survival and induced apoptosis and cytostasis in formed tumor colonies. In addition, T4-2 tumors with inhibition of NF-κB activity exhibit decreased growth in athymic mice, which was further reduced by IR with downregulated b1-integrin expression. Direct interactions between b1-integrin and NF-κB p65 were induced in nonmalignant breast epithelial cells, but not in malignant cells, indicating context-specific regulation. As b1-integrin also activates NF-κB, our findings reveal a novel forward feedback pathway that could be targeted to enhance therapy. [ABSTRACT FROM AUTHOR]

Chalcones, which have characteristic 1,3-diaryl-2-propen-1-one skeleton, are mainly produced in roots, rhizomes, heartwood, leaves, and seeds of genera Angelica, Sophora, Glycyrrhiza, Humulus, Scutellaria, Parartocarpus, Ficus, Dorstenia, Morus, Artocarpus, and so forth. They have become of interest in the research and development of natural antitumor agents over the past decades due to their broad range of mechanisms including anti-initiation, induction of apoptosis, antiproliferation, antimetastasis, antiangiogenesis, and so forth. This review summarizes the studies on the antitumor activity of naturally occurring chalcones and their underlying mechanisms in detail during the past decades. [ABSTRACT FROM AUTHOR]

Acetaminophen (APAP) overdose is the most frequent cause of acute liver failure and is primarily caused by cytochrome P450 (CYP) 2E1-driven conversion of APAP into hepatotoxic metabolites. Several reports showed that melatonin attenuated APAP-induced acute liver failure. Nevertheless, the exact mechanism remains obscure. In the present study, we investigated the effects of melatonin on apoptosis-inducing factor (AIF)-dependent cell death in APAP-induced acute liver failure. Mice were intraperitoneally (i.p.) injected with different doses of melatonin (1.25, 5, 20 mg/kg) 30 min before APAP (300 mg/kg, i.p.). As expected, melatonin significantly alleviated APAP-induced cell death, as determined by TdT-mediated dUTP-biotin nick end labeling (TUNEL) assay. Further analysis showed that melatonin significantly attenuated APAP-induced activation of the serine/threonine kinase receptor interacting protein 1 (RIP1). In addition, melatonin inhibited APAP-induced hepatic c- Jun N-terminal kinase (JNK) phosphorylation and mitochondrial Bax translocation. Correspondingly, melatonin inhibited APAP-induced translocation of AIF from mitochondria to nuclei. Interestingly, no changes were induced by melatonin on hepatic CYP2E1 expression. In addition, melatonin had little effect on APAP-induced hepatic glutathione (GSH) depletion. In conclusion, melatonin protects against AIF-dependent cell death during APAP-induced acute liver failure through its direct inhibition of hepatic RIP1 and subsequent JNK phosphorylation and mitochondrial Bax translocation. [ABSTRACT FROM AUTHOR]

Streptococcus equi subsp. zooepidemicus (S. zooepidemicus), which belongs to Lancefield group C streptococci, is an important pathogen of domesticated species, causing septicemia, meningitis and mammitis. M-like protein (SzP) is an important virulence factor of S. zooepidemicus and contributes to bacterial infection and antiphagocytosis. To increase our knowledge of the mechanism of SzP in infection, we profiled the response of porcine pulmonary alveolar macrophage (PAM) to infection with S. zooepidemicus ATCC35246 wild strain (WD) and SzP-knockout strain (KO) using the Roche NimbleGen Porcine Genome Expression Array. We found SzP contributed to differential expression of 446 genes, with upregulation of 134 genes and downregulation of 312 genes. Gene Ontology category and KEGG pathway were analyzed for relationships among differentially expressed genes. These genes were represented in a variety of functional categories, including genes involved in immune response, regulation of chemokine production, signal transduction and regulation of apoptosis. The reliability of the data obtained from the microarray was verified by performing quantitative real-time PCR on 12 representative genes. The data will contribute to understanding of SzP mediated mechanisms of S. zooepidemicus pathogenesis. [ABSTRACT FROM AUTHOR]

Various mechanical stresses in vivo induce disc cell apoptosis and intervertebral disc (IVD) degeneration, but the underlying molecular mechanism is not fully known. The aim of this study was to investigate the role of endoplasmic reticulum stress in cyclic stretch-induced apoptosis of rat annulus fibrosus (AF) cells. Flexercell Tension Plus system was used to apply cyclic stretch to rat annulus fibrosus cells at a frequency of 0.5 Hz with 20% elongation for 12, 24, 36, or 48 h. Apoptosis was detected by flow cytometry, and nuclei morphologic changes were visualized by Hoechst 33258 staining and caspase-8, 9 activity assays. The expression of the markers of endoplasmic reticulum stress including CHOP, GRP78, and caspase-12 were determined by RT-PCR and Western blot. Mitochondrial membrane potential change was observed by JC-1 staining in situ. In addition, the levels of the nitric oxide (NO) were determined with the Griess reaction and fluorescence staining. The results indicated that cyclic stretch at a frequency of 0.5 Hz with 20% elongation-induced apoptosis in rat AF cells. Prolonged exposure of the unphysiologically cyclic stretch to AF cells caused NO overproduction, up-regulation of endoplasmic reticulum stress markers including CHOP, GRP78, and caspase-12, depolarization of mitochondria and activation of caspase-9. However, cyclic stretch at this level had no effect on caspase-8 activity. In addition, specific inhibitor of caspase-12 (Z-ATAD-FMK) and caspase-9 (Z-LEHD-FMK) partly suppressed cyclic stretch-induced AF cell apoptosis and the anti-apoptotic effects of the caspase inhibitors were additive. Our data suggest that endoplasmic reticulum stress, likely mediated by NO, contributes to the AF cell apoptosis induced by cyclic stretch in addition to the mitochondrial pathway. These findings could be helpful to understand the mechanism of disc cell apoptosis, the root cause of IVD degeneration. [ABSTRACT FROM AUTHOR]

Background: Drug resistance remains a great challenge in the treatment of gastric cancer. The goal of this study was to explore the anti-tumor effects and mechanism of cytokine-induced killer (CIK) cell combined with oxaliplatin (L-OHP) in human oxaliplatin-resistant gastric cancer cells. Methods: After producing oxaliplatin-resistant gastric cancer cells, cell morphology, growth and doubling time were observed, followed by detection of cell cycle distribution and apoptosis, drug sensitivity (e.g., L-OHP) and expression of P-gp and livin. MTT assay, in vivo pharmacodynamics and pathomorphology experiments were used to detect killing activities of CIK combined with L-OHP. Results: Compared with parental gastric cancer cells, oxaliplatin-resistant gastric cancer cells in S phase were reduced and cell apoptosis rate was increased (P < 0.05), the inhibition rate of 10 chemotherapeutics on oxaliplatin-resistant gastric cancer cells was significantly lower and the expression of P-gp was significantly higher (P < 0.05). However, there was no significant difference in livin expression between parental gastric cancer cells and oxaliplatin-resistant gastric cancer cells (P > 0.05). The in vitro killing activity of CIK combined with L-OHP on parental cells and oxaliplatin-resistant cells were significantly enhanced compared with L-OHP or CIK alone. And it showed greater synergetic effects against oxaliplatin-resistant cells compared with parental cells (P < 0.05). In addition, survival rate, abdominal circumference and pathomorphology results revealed stronger in vivo anti-tumor effects when the two therapies were combined. Conclusions: The mechanism of oxaliplatin-resistant cell secondary multidrug resistance was correlated with the variation of cell cycle distribution, extension of doubling time and upregulation of P-gp expression. The synergistic effect of CIK in combination with L-OHP on killing activity against oxaliplatin-resistant cells was shown in vivo and in vitro. [ABSTRACT FROM AUTHOR]

Background: Chronic alcohol consumption reduces the percentage and number of peripheral natural killer (NK) cells in mice and in humans. The underlying mechanism for these changes is only partly known. We recently found that chronic alcohol consumption inhibits NK cell release from the bone marrow (BM) and that this is associated with a decrease in splenic NK cells. The number of peripheral NK cells is tightly controlled by homeostatic proliferation. It is not known whether this mechanism is initiated in response to the reduction in splenic NK cells, or if so, why the steady state levels of NK cells are not restored. Methods: To examine this mechanism, female C57BL/6 mice were given 20% w/v alcohol in the drinking water for 3 months. NK cell proliferation and apoptosis were determined before and after treatment with IL-15 alone or combined with its alpha receptor. Results: Chronic alcohol consumption invoked homeostatic proliferation of splenic NK cells in an attempt to return NK cells to normal levels; however, this did not happen due to enhanced apoptosis of NK cells relative to proliferation. Chronic alcohol consumption decreased IL-15 producing cells in the spleen but not in the BM. The numbers of NK cells in the alcohol-consuming mice returned to normal levels in the spleen and were higher than normal in the BM after 2 daily injections of IL-15; however, the enhanced rate of apoptosis due to alcohol consumption was not decreased in the spleen or BM. Combined IL-15 and IL-15Rα treatment decreased apoptosis of NK cells from alcohol-consuming mice to levels similar to untreated water-drinking mice and greatly increased the percentage and number of NK cells in both the spleen and BM. Conclusion: Chronic alcohol consumption causes a self-unrecoverable loss of NK cells in the spleen by compromising NK cell release from the BM and enhancing splenic NK cell apoptosis that can be reversed with IL-15/IL-15Rα treatment. [ABSTRACT FROM AUTHOR]

Danshen is commonly used in China for the treatment of atherosclerosis-related disorders such as cardiovascular and cerebrovascular diseases. Research shows that it also has immunostimulation properties. The present study evaluates the protective effect of danshensu, an active water-extractable component isolated from danshen, on an endothelial cell line (CRL-1730) treated with hydrogen peroxide (H2O2). Danshensu significantly inhibited endothelial cell viability induced by H2O2. The treatment of endothelial cells with danshensu resulted in most cells being arrested in the S and G2/M phases of the cell cycle. The fraction of cells in G0/G1 phase was markedly decreased by danshensu treatment compared to the control groups. The apoptosis was also markedly decreased after danshensu treatment. Additionally, danshensu restrains decreased nitric oxide level, increased the release of lactate dehydrogenase and expression of cluster of differentiation 40 (CD40) significantly. These results suggest that danshensu protects endothelial cells from the damage induced by H2O2 through its CD40 anti-inflammatory approach and cell apoptosis inhibition. [ABSTRACT FROM AUTHOR]

We consider a procedure for cancer therapy which consists of injecting replication-competent viruses into the tumor. The viruses infect tumor cells, replicate inside them, and eventually cause their death. As infected cells die, the viruses inside them are released and then proceed to infect adjacent tumor cells. However, a major factor influencing the efficacy of virus agents is the immune response that may limit the replication and spread of the replication-competent virus. The competition between tumor cells, a replication-competent virus and an immune response is modelled as a free boundary problem for a nonlinear system of partial differential equations, where the free boundary is the surface of the tumor. In this model, the immune response equation is a semilinear parabolic equation, including a chemotaxis term which is used to describe the movement of the immune response induced by gradients of the infected cell density. Under the assumption that the chemotactic sensitivity coefficient is small compared with the diffusion coefficient of the immune response, we prove the global existence and uniqueness of the solution of this free boundary problem. For large chemotactic coefficient, the global existence is still open. [ABSTRACT FROM AUTHOR]

Relative specific amino acid dependency is one of the metabolic abnormalities of cancer cells, and restriction of specific amino acids induces apoptosis of prostate cancer cells. This study shows that restriction of tyrosine and phenylalanine (Tyr/Phe), glutamine (Gln), or methionine (Met), modulates Raf and Akt survival pathways and affects the function of mitochondria in DU145 and PC3, in vitro. These three restrictions inhibit energy production (ATP synthesis) and induce generation of reactive oxygen species (ROS). Restriction of Tyr/Phe or Met in DU145 and Met in PC3 reduces mitochondrial membrane potential (ΔΨm) and induces caspase-dependent and -independent apoptosis. In DU145, Tyr/Phe or Met restriction reduces activity of Akt, mitochondrial distribution of phosphorylated Raf and apoptosis inducing factor (AIF), and increases mitochondrial distribution of Bak. Mitochondrial Bcl-XL is increased in Tyr/Phe-restricted but decreased in Met-restricted cells. Under Tyr/Phe or Met restriction, reduced mitochondrial Raf does not inactivate the pro-apoptotic function of Bak. Tyr/Phe restriction also inhibits Bcl-2 and Met restriction inhibits Bcl-XL in mitochondria. These comprehensive actions damage the integrity of the mitochondria and induce apoptosis of DU145. In PC3, apoptosis induced by Met restriction was not associated with alterations in intracellular distribution of Raf, Bcl-2 family proteins, or AIF. All of the amino acid restrictions inhibited Akt activity in this cell line. We conclude that specific amino acid restriction differentially interferes with homeostasis/balance between the Raf and Akt survival pathways and with the interaction of Raf and Bcl-2 family proteins in mitochondria to induce apoptosis of DU145 and PC3 cells. J. Cell. Physiol. 209: 522–534, 2006. © 2006 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Caspase-3 is a programmed cell death protease involved in neuronal apoptosis during physiological development and under pathological conditions. It is a promising therapeutic target for treatment of neurodegenerative diseases. We reported previously that isoquinoline-1,3,4-trione and its derivatives inhibit caspase-3. In this report, we validate isoquinoline-1,3,4-trione and its derivatives as potent, selective, irreversible, slow-binding and pan-caspase inhibitors. Furthermore, we show that these inhibitors attenuated apoptosis induced by β-amyloid(25–35) in PC12 cells and primary neuronal cells. [ABSTRACT FROM AUTHOR]

1. The present work was designed to observe the early structural changes in the aortic wall in Sprague-Dawley rats 1, 2 and 4 weeks after sinoaortic denervation (SAD). 2. Rats were examined 1, 2 and 4 weeks after SAD. Blood pressure (BP) was recorded in the conscious state. The thoracic aortas were taken for investigations, including: light microscopy, electron microscopy, immunohistochemistry and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labelling (TUNEL). 3. Blood pressure variability (BPV) was significantly increased in the SAD groups 1, 2 and 4 after the operation when compared with the sham-operated ones. 4. Two weeks after SAD the percentage proportion of smooth muscle cell density (SMC%) was obviously increased. 5. Four weeks after SAD: the SMC%, percentage proportion of collagen density (CD%) and aortic wall thickness (WT) were obviously increased with vascular smooth muscle cells blebbing concomitantly. Endothelial cells showed degenerative changes and swelling with blebbing of the cell membrane and increased condensation of peripheral nuclear chromatin and cytoplasmic vacuolization. It was also found that the number of apoptotic endothelial cells was increased and expression of eNOS was reduced. 6. This is the first study that shows the time-course of aortic wall and endothelial cell changes induced by SAD. Increased BPV might be the priming factor in the development of organ damage induced by SAD. [ABSTRACT FROM AUTHOR]

Apoptin, a viral death protein derived from chicken anemia virus, displays a number of tumor-specific behaviors. In particular, apoptin is phosphorylated, translocates to the nucleus, and induces apoptosis specifically in tumor or transformed cells, whereas it is nonphosphorylated and remains primarily inactive in the cytoplasm of nontransformed normal cells. Here, we show that in normal cells apoptin can also be activated by the transient transforming signals conferred by ectopically expressed simian virus 40 (SV40) large T antigen (LT), which rapidly induces apoptin's phosphorylation, nuclear accumulation, and the ability to induce apoptosis. Further analyses with mutants of LT showed that the minimum domain capable of inducing all three of apoptin's tumor-specific properties resided in the N-terminal J domain, a sequence which is largely shared by SV40 small t antigen (st). Interestingly, the J domain in st, which lacks its own nuclear localization signal (NLS), required nuclear localization to activate apoptin. These results reveal the existence of a cellular pathway shared by conditions of transient transformation and the stable cancerous or precancerous state, and they support a model whereby a transient transforming signal confers on apoptin both the upstream activity of phosphorylation and the downstream activity of nuclear accumulation and apoptosis induction. Such a pathway may reflect a general lesion contributing to human cancers. [ABSTRACT FROM AUTHOR]

The chicken anemia virus-derived protein Apoptin induces apoptosis specifically in human tumor and transformed cells and not in normal, untransformed cells. The cell killing activity correlates with a predominantly nuclear localization of Apoptin in tumor cells, whereas in normal cells, it is detected mainly in cytoplasmic structures. To explore the role of nuclear localization for Apoptin-induced cell death in tumor cells, we employed a mutagenesis strategy. First, we demonstrated that the C terminus of Apoptin contains a bipartite-type nuclear localization signal. Strikingly, further investigation showed that Apoptin contains two different domains that induce apoptosis independently, and for both domains, we found a strong correlation between localization and killing activity. Using inhibitors, we ruled out the involvement of de novo gene transcription and translation and further showed that Apoptin itself does not have any significant transcriptional repression activity, suggesting that Apoptin exerts its effects in the nucleus by some other method. To determine whether nuclear localization is sufficient to enable Apoptin to kill normal, untransformed cells, we expressed fulllength Apoptin fused to a heterologous nuclear localization signal in these cells. However, despite its nuclear localization, no apoptosis was induced, which suggests that nuclear localization per se is not sufficient for Apoptin to become active. These studies increase our understanding of the molecular pathway of Apoptin and may also shed light on the mechanism of cellular transformation. [ABSTRACT FROM AUTHOR]

Cites the inducement of tumor-specific apoptosis as a globular multimer through apoptin. Formation of noncovalent globular aggregates by recombinant apoptin proteins; Irreversibility of the multimerization; Mapping of the structural motifs responsible for multimerization in vitro.

Tibial dyschondroplasia (TD) is a metabolic bone disease that occurs in fastgrowing chickens and can lead to substantial economic damages and compromise the poultry welfare. Triptolide is a traditional Chinese medicine (TCM), which has been broadly used as an anti-inflammatory, and anti-cancerous drug. However, its therapeutic role against tibial dyschondroplasia is under-reported yet. Therefore, the study aimed for investigating the protective effect of triptolide against autophagy and apoptosis in TD affected chickens. Random classification of chickens (n=30) was done into three groups: CON, TD and triptolide treatment group (TP). All chickens were given standard diet until the end of the experiment for 18 days. In this study, tibial bone was collected for analyzing several aspects, serum was collected for biochemical analysis, and microscopic assessment was done by H&E staining. Immunohistochemistry, immunofluorescence, western blotting, and RT-qPCR were used to detect autophagy and apoptosis-related proteins and genes. Observations illustrated that after triptolide treatment, the symptoms of TD group were improved, and tibial parameters were changed. At the same time, the expression of m-TOR and P62 mRNA in TP group showed significant decrease comparing to TD group. Precisely, thiram induced TD was found to be involved in hypertrophic chondrocytes apoptosis and autophagy, and triptolide showed protective effects against TD. [ABSTRACT FROM AUTHOR]

Hepatocellular carcinoma (HCC) is one of the most malignant types of human primary tumor and has a poor prognosis, therefore, the development of novel therapeutic modalities is necessary. Fatsioside A is a novel baccharane-type triterpenoid glycoside, which is extracted from the fruits of Fatsia japonica. Previous data has revealed that fatsioside A can exert growth inhibition, cell cycle arrest and induce apoptosis in human glioma cells. However, no detailed investigations have been performed to determine its action on human hepatocellular cells, and the exact mechanisms underlying the induction of apoptosis remain to be elucidated. The aim of the present study was to investigate the anticancer effect of fatsioside A in the HepG2 human HCC cell line, and to investigate the underlying mechanisms by focusing on the AMP-activated protein kinase (AMPK) signaling cascade. The results of the present study demonstrated that fatsioside A induced apoptotic death of the human HepG2 HCC cells, which was associated with a marked activation of AMPK and increased expression of the downstream acetyl-CoA carboxylase carboxylase. Inhibition of AMPK by RNA interference or by its inhibitor, compound C, suppressed fatsioside A-induced caspase-3 cleavage and apoptosis in the HepG2 cells, while AICAR, the AMPK activator, elicited marked cytotoxic effects. Together, these results suggested that fatsioside A-induced apoptotic death requires AMPK activation in HepG2 cells. [ABSTRACT FROM AUTHOR]

Objectives. To investigate the effect of decreased alpha-enolase (ENO1) expression on rheumatoid arthritis fibroblasts-like synoviocytes (RA-FLSs) proliferation in response to hypoxia, and elucidate the possible mechanisms involved. Methods. RA-FLSs and osteoarthritis fibroblasts-like synoviocytes (OA-FLSs) were cultured in trigas incubators with different oxygen concentrations (3% O2, 7% O2, and 21% O2). 3% O2 (hypoxia) and 7% O2 conditions simulated intra-articular oxygen concentrations as observed in RA and healthy individual, respectively. 21% O2 represented oxygen condition for normal cell culture. ENO1-knockdown FLSs were established using ENO1-siRNA. The expression level of ENO1 was detected using reverse transcription polymerase chain reaction or RT-PCR and Western blot. Proliferation and apoptosis of RA-FLSs and OA-FLSs were assessed using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H-tetrazolium or MTS assay and flow cytometry, respectively. Western blot analysis was used to detect key proteins involved in apoptosis. Results. ENO1 gene expression was remarkably upregulated, as well as its translation into protein, in RA-FLSs and OA-FLSs that were cultured in 3% O2 concentration. RA-FLSs and OA-FLSs that were cultured under hypoxic conditions hyperproliferated compared with similar cells under normaxic conditions. Neither 7% O2 nor 21% O2 condition had any significant effect on ENO1 expression. ENO1-siRNA-transfected FLSs, but not control-siRNA FLSs, showed markedly decreased proliferation. Additionally, ENO1 expression was found to promote significantly higher expression levels of the anti-apoptotic proteins Bcl-2, surviving, and cyclinB1, but inhibited the expression of cleaved caspase3. Conclusion. ENO1 may be crucial in the regulation of the proliferation and survival of synovial fibroblasts. [ABSTRACT FROM AUTHOR]

Background Berberine, an isoquinoline alkaloid, has shown inhibitory effects on growth of several tumor cell lines in vitro. The aim of this study was to investigate chemopreventive effects of berberine on intestinal tumor development in Apcmin/+ mice. Methods Four-week old Apcmin/+ mice were treated with 0.05% or 0.1% berberine in drinking water for twelve weeks. The number and the size of tumors were measured to evaluate intestinal tumor development. Tissue sections were prepared for PCNA and Ki-67 immunostaining to detect cell proliferation, and TUNEL assay and cleaved caspase-3 immunostaining for apoptosis. Western blot analysis and immunostaining were performed to detect the activation of Wnt and epidermal growth factor receptor (EGFR) signaling pathways and COX-2 expression in the intestinal tumor cells. The prostaglandin E2 level in the small intestine was detected using ELISA. Results Compared with untreated Apcmin/+ mice, the total numbers of tumors in the small intestine and the colon were reduced by 39.6% and 62.5% in 0.05% and 0.1% berberine-treated mice, respectively. The numbers of tumors in proximal, middle, and distal segments of the small intestine in 0.1% berberine-treated mice were significantly reduced by 53.7%, 55.3%, and 76.5% respectively. Berberine treatment also decreased the numbers of all sizes of tumors (>2 mm, 1-2 mm, and <1 mm) in the small intestine. Berberine suppressed tumor cell proliferation and increased apoptosis. Furthermore, berberine decreased the activation levels of Wnt and EGFR signaling pathways, and down-regulated COX-2 expression in intestinal tumor cells and prostaglandin E2 production in the small intestine. Conclusions Berberine inhibits intestinal tumor development, which is correlated with its activity to suppress tumor cell proliferation and increase apoptosis in Apcmin/+ mice. Down-regulation of Wnt and EGFR signaling pathways and COX-2 expression by berberine may be involved in its anti-tumorigenic effects. [ABSTRACT FROM AUTHOR]

TBWSP31 is a novel antitumor protein that was isolated from tartary buckwheat water-soluble extracts. The objective of this paper was to investigate the anti-proliferative effects of TBWSP31 on breast cancer Bcap37cells and to explore its possible mechanism. After treatment of Bcap37 cells with TBWSP31, typical apoptotic morphological changes were observed by inverted microscopy and scanning electron microscopy (SEM), such as detachment from the culture plate, change to a round shape, cell shrinkage, the absence of obvious microvilli, plasma membrane blebbing, and formation of apoptotic bodies. Cell-cycle analysis revealed that treatment with TBWSP31 resulted in a G0/G1 arrest and prevented the cells from growing from G0/G1 phase to S phase, which was most prominent at 48 h. The expression of bcl-2 and Fas were detected quantitatively by FCM, which showed that TBWSP31 induced-apoptosis may be involved with the participation of Fas and bcl-2. These results suggest that TBWSP31 is a potential antitumor compound and that apoptosis induced by TBWSP31 is a key antitumor mechanism. [ABSTRACT FROM AUTHOR]

The effects of sophorolipid on the growth and apoptosis of H7402 human liver cancer cells were investigated. By treatment with sophorolipid, a dose- and time-dependent inhibition of cell proliferation was observed. The cells developed many features of apoptosis, including condensation of chromatin, nuclear fragmentation, and appearance of apoptotic bodies, and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling positive cells were stained dark brown. Sophorolipid treatment induced apoptosis in H7402 cells by blocking cell cycle at G1 phase and partly at S phase, activating caspase-3, and increasing Ca2+ concentration in cytoplasm. These findings may suggest a potential use of sophorolipid for liver cancer treatment. [ABSTRACT FROM AUTHOR]

The metabolic regulation of cell death is sophisticated. A growing body of evidence suggests the existence of multiple metabolic checkpoints that dictate cell fate in response to metabolic fluctuations. However, whether microRNAs (miRNAs) are able to respond to metabolic stress, reset the threshold of cell death, and attempt to reestablish homeostasis is largely unknown. Here, we show that miR-378/378* KO mice cannot maintain normal muscle weight and have poor running performance, which is accompanied by impaired autophagy, accumulation of abnormal mitochondria, and excessive apoptosis in skeletal muscle, whereas miR-378 overexpression is able to enhance autophagy and repress apoptosis in skeletal muscle of mice. Our in vitro data show that metabolic stress-responsive miR-378 promotes autophagy and inhibits apoptosis in a cell-autonomous manner. Mechanistically, miR-378 promotes autophagy initiation through the mammalian target of rapamycin (mTOR)/unc-51-like autophagy activating kinase 1 (ULK1) pathway and sustains autophagy via Forkhead box class O (FoxO)-mediated transcriptional reinforcement by targeting phosphoinositide-dependent protein kinase 1 (PDK1). Meanwhile, miR-378 suppresses intrinsic apoptosis initiation directly through targeting an initiator caspase- Caspase 9. Thus, we propose that miR-378 is a critical component of metabolic checkpoints, which integrates metabolic information into an adaptive response to reduce the propensity of myocytes to undergo apoptosis by enhancing the autophagic process and blocking apoptotic initiation. Lastly, our data suggest that inflammationinduced down-regulation of miR-378 might contribute to the pathogenesis of muscle dystrophy. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • AF suppressed glioma cells. • AF induced apoptosis and inhibited glycolysis. • AF upregulated miR-124-3p by repressing DNMT1 through Sp1, which caused by activation of ROS/AMPK. Abstract Malignant glioma is the most common type of brain tumor with poor clinical outcome and survival. Therefore, it is imperative to develop novel therapeutic agents for managing glioma. The aim of this study was to investigate the role of amentoflavone (AF), an active flavonoid component in Selaginella tamariscina Spring, in glioma cells and the underlying mechanism of its action. Our results showed that miR-124-3p expression was significantly down-regulated in glioma tissues relative to normal brain tissues. AF decreased cell viability and triggered apoptosis in both glioma cell lines in a dose-dependent manner. AF induced apoptosis and inhibited glycolysis in the glioma cells by upregulating miR-124-3p. Furthermore, AF upregulated miR-124-3p by repressing DNMT1 through Sp1, which in turn was caused by the activation of ROS/AMPK signaling pathway by AF. In conclusion, AF could induce apoptosis and inhibited glycolysis in glioma cells via miR-124-3p. Our findings provide preliminary experimental data that support further investigation on the therapeutic efficacy of AF in glioma. [ABSTRACT FROM AUTHOR]

Background/Aims: Recently, we observed an increase in O-GlcNAc (O-linked-ß-Nacetylglucosamine) modification, and signal transducer and activator of transcription proteins 3 (STAT3) expression in primary retinal vascular endothelial cells (RVECs) under high glucose conditions and tissues altered by diabetic retinopathy (DR). In this study, we focused on the correlations between O-GlcNAcylation and STAT3 phosphorylation, and their potential effects with regards to DR. Methods: Expression of O-GlcNAcylation and STAT3 were detected in DR-affected tissues and primary RVECs. The relationship between O-GlcNAcylation and STAT3 was further delineated by immunoprecipitation and Western blot analysis. Effects of O-GlcNAcylation on human RVEC apoptosis and involved protein expression were assayed with flow cytometry and Western blot. Results: Global O-GlcNAcylation and pSTAT3 levels were significantly elevated in diabetic rat retina and primary RVECs under high glucose conditions. In vitro assays demonstrated that the Tyr705 site was sensitive to high glucose. While O-GlcNAcylation inhibited p727STAT3 expression, augmented O-GlcNAcylation could balance p705STAT3 expression within relatively high levels corresponding to vascular endothelial growth factor (VEGF) changes. Immunoprecipitation revealed that STAT3 was modified by O-GlcNAcylation and phosphorylation simultaneously. Next, we observed that overexpression of O-GlcNAcylation could relieve human RVEC apoptosis related to the JAK2- Tyr705STAT3-VEGF pathway. Conclusion: O-GlcNAcylation could relieve RVECs apoptosis through the STAT3 pathway in DR, and O-GlcNAcylation combined with STAT3 phosphorylation might open up new insights into the mechanisms of DR and other diabetic complications. [ABSTRACT FROM AUTHOR]

Introduction: The risk of oral lichen planus (OLP), a chronic inflammatory oral mucosal disease, becoming malignant increases by 21-fold in patients with fungal infection. This study examined the impact of Candida albicans exposure on Toll-like receptor (TLR) signaling in primary keratinocyte cultures obtained from OLP patients. Methodology: Following co-culture of primary OLP keratinocyte cultures with C. albicans for 24 hours, inflammatory cytokine concentrations were determined by ELISA. TLR2, MyD88, and NF-κBp65 mRNA and protein expression were assessed using quantitative RT-PCR and Western blot analyses, respectively. Keratinocyte apoptosis was also determined by flow cytometry. Results: IL-10, IL-8, IL-2, and TNF-α levels were significantly higher following co-culture with C. albicans (all p ≤ 0.034). MyD88, NF-κB p65, and TLR2 mRNA (all p < 0.001) and protein (all p ≤ 0.004) expression levels were significantly higher in OLP keratinocytes following C. albicans exposure. Finally, the apoptosis rates of OLP keratinocytes were 21.2%, 29.4%, and 25.4% for the control cells and 3.9%, 5.6%, and 4.4% for those exposed to C. albicans, suggesting that co-culture with C. albicans inhibits the apoptosis of OLP keratinocytes. Conclusions: C. albicans activates the TLR2/MyD88/NF-κB signaling pathway in OLP keratinocytes, resulting in increased cytokine expression and decreased keratinocyte apoptosis. Two key events in the pathogenesis of OLP and its progression to malignancy, namely increased inflammation and decreased apoptosis, were induced by exposure to C. albicans. Thus, targeting this signaling pathway may represent a novel therapeutic strategy to prevent OLP malignant transformation. [ABSTRACT FROM AUTHOR]

p21WAF1/CIP1 is a broad-acting cyclin-dependent kinase inhibitor. Its stability is essential for proper cell-cycle progression and cell fate decision. Ubiquitylation by the multiple E3 ubiquitin ligase complexes is the major regulatory mechanism of p21, which induces p21 degradation. However, it is unclear whether ubiquitylated p21 can be recycled. In this study, we report USP11 as a deubiquitylase of p21. In the nucleus, USP11 binds to p21, catalyzes the removal of polyubiquitin chains conjugated onto p21, and stabilizes p21 protein. As a result, USP11 reverses p21 polyubiquitylation and degradation mediated by SCFSKP2, CRL4CDT2, and APC/CCDC20 in a cell-cycle-independent manner. Loss of USP11 causes the destabilization of p21 and induces the G1/S transition in unperturbed cells. Furthermore, p21 accumulation mediated by DNA damage is completely abolished in cells depleted of USP11, which results in abrogation of the G2 checkpoint and induction of apoptosis. Functionally, USP11-mediated stabilization of p21 inhibits cell proliferation and tumorigenesis in vivo. These findings reveal an important mechanism by which p21 can be stabilized by direct deubiquitylation, and they pinpoint a crucial role of the USP11-p21 axis in regulating cell-cycle progression and DNA damage responses. [ABSTRACT FROM AUTHOR]

Background: Curcumin, a natural compound derived from the turmeric rhizome Curcuma longa Linn, has anticancer and chemoresistance reduction biological activities. We evaluated the efficacy of curcumin in sensitizing chemotherapy drugs through regulation of Bcl-2-mediated apoptosis in breast cancer stem-like cells (BCSCs). Methods: Cell survival was measured using MTT assay. Apoptosis-related proteins were observed using western blot analysis. Apoptosis was detected with flow cytometric analysis and by Hoechst 33258 staining. The mitochondrial membrane potential was observed with flow cytometric analysis. Results: The ability of BCSCs to propagate decreased gradually along the passages and was completely lost at the fifth passage [0.1 μmol/L mitomycin C (MMC) with 5 μmol/L curcumin in MCF-7 and 0.5 μmol/L MMC with 5 μmol/L curcumin in MDA-MB-231 cells]. Curcumin combined with MMC treatment significantly decreased the levels of antiapoptotic Bcl-2 and Bcl-w expression, increased the levels of proapoptotic Bax, Bak, Bad, Bik, and Bim expression, and activated caspase-3 and caspase-9 in MCF-7 BCSCs. In the presence of Bcl-2 siRNA, the apoptosis rate increased by 15% in cells treated with curcumin and MMC. The mitochondrial membrane potential decreased by approximately 20% in MCF-7 BCSCs undergoing the combination treatment of curcumin and MMC. The combination-induced decrease in Bcl-2 was regulated by the presence of the Wnt-specific inhibitor PFK115-584 and PI3k inhibitor LY294002. Conclusions: Our study indicates that curcumin might represent a novel therapeutic agent for treating breast cancer chemoresistance induced by MMC. [ABSTRACT FROM AUTHOR]

Objective: The purpose of this study was to evaluate the frequency of RASSF1A hypermethylation in patients with acute myeloid leukemia (AML), in an attempt to modify the current molecular model for disease prognosis. Materials and methods: Aberrant RASSF1A promoter methylation levels were assessed in 226 newly diagnosed non-M3 AML patients and 30 apparently healthy controls, by quantitative methylation-specific polymerase chain reaction. Meanwhile, RASSF1A mRNA levels were detected by real-time quantitative polymerase chain reaction. Furthermore, hematological characteristics, cytogenetic abnormalities, and genetic aberrations were assessed. Finally, associations of RASSF1A hypermethylation with clinical outcomes were evaluated. Results: RASSF1A hypermethylation was observed in 23.0% of patients with non-M3 AML (52/226), but not in controls. Meanwhile, hypermethylation of the RASSF1A promoter was significantly associated with ASXL1 mutation. Furthermore, the log-rank test revealed that RASSF1A hypermethylation indicated decreased relapse-free survival (RFS) and overall survival (OS) in patients with non-M3 AML (P=0.012 and P=0.014, respectively). In multivariate analysis, RASSF1A hypermethylation was an independent prognostic factor for RFS (P=0.040), but not for OS (P=0.060). Conclusion: Hypermethylation of the RASSF1A promoter is associated with ASXL1 mutation in non-M3 AML patients, likely indicating poor outcome. These findings provide a molecular basis for stratified diagnosis and prognostic evaluation. [ABSTRACT FROM AUTHOR]

B-cell translocation gene 2 (BTG2) proteins have been reported to be putative tumor suppressors in various cancer types. The present study first assessed BTG2 expression in 44 human liver cancer tissue specimens, then investigated BTG2 expression in the regulation of hepatocellular carcinoma (HCC) cell apoptosis with or without radiotherapy in vitro and in vivo. The results revealed that BTG2 protein expression was significantly reduced in HCC tissues, and associated with better survival for HCC patients (P=0.05). BTG2 overexpression also sensitized Huh7 cells to radiation-induced apoptosis in vitro and in a nude mouse model, although restoration of BTG2 expression per se did not affect the viability and apoptosis of HCC cells. Future studies would confirm the role of BTG2 in hepatoma, and further develop BTG2 as a therapeutic strategy for controlling HCC. [ABSTRACT FROM AUTHOR]

In the HIV-1 replication cycle, the endosomal sorting complex required for transport (ESCRT) machinery promotes viral budding and release in the late stages. In this process, the ESCRT proteins, ALIX and TSG101, are recruited through interactions with HIV-1 Gag p6. ALG-2, also known as PDCD6, interacts with both ALIX and TSG101 and bridges ESCRT-III and ESCRT-I. In this study, we show that ALG-2 affects HIV-1 production negatively at both the exogenous and endogenous levels. Through a yeast two-hybrid screen, we identified HEBP2 as the binding partner of ALG-2, and we solved the crystal structure of the ALG-2·HEBP2 complex. The function of ALG- 2·HEBP2 complex in HIV-1 replication was further explored. ALG-2 inhibits HIV-1 production by affecting Gag expression and distribution, and HEBP2 might aid this process by tethering ALG-2 in the cytoplasm. [ABSTRACT FROM AUTHOR]

microRNA (miRNA) is abnormally expressed in numerous diseases, and it was intimately associated with cell proliferation and apoptosis. However, the mechanism by which miRNAs control cataractogenesis remains unclear. In the current study, it was demonstrated that miR-34a was highly expressed in the cataractous lens by stem-loop reverse transcription-quantitative polymerase chain reaction. Trying to investigate the role of miR-34a in human lens epithelial cells, miR-34a mimics were transfected into SRA01/04 cells, and this suppressed proliferation and induced apoptosis. Subsequently, E2F3 was confirmed as a direct target of miR-34a. Downregulation of E2F3 by small interfering (si) RNA siE2F3 resulted in proliferation inhibition and apoptosis of SRA01/04 cells. Furthermore, it was demonstrated that miR-34a and siE2F3 downregulated E2F3 expression at a protein level. In summary, the current study demonstrated that miR-34a suppressed the proliferation and induced apoptosis of SRA01/04 cells by downregulating E2F3. These observations provide novel insights with potential therapeutic applications for the treatment of cataracts. [ABSTRACT FROM AUTHOR]

Brucellosis is a globally distributed zoonotic disease that causes animal and human diseases. Brucella pathogenesis depends on their ability of inhibiting apoptosis and establishes a replicative niche inside host cells. SUMOylation is a major regulator of protein function that is essential for some pathogenic bacteria during infection. However, the relationship between Brucella and SUMOylation remains largely unknown. In our report, we demonstrated that the Brucella melitensis 16M (16M) infection leads to a decrease in the expression of SUMO1 proteins with time-delay, and 16M intracellular replication was inhibited by SUMO-1 overexpression and promoted by SUMO-1 depletion, and 16M-dependent apoptosis and the secretion of gamma interferon and interleukin-6 were induced by SUMO-1 overexpression and restricted by SUMO-1 depletion in the RAW264.7 macrophages. Together, this study shows that SUMO-1 plays an essential role in regulating 16M infection and may help to unravel pathogenic mechanisms of 16M. [ABSTRACT FROM AUTHOR]

Background: Hirsutanol A is a novel sesquiterpene compound purified from fungus Chondrostereum sp. in Sarcophyton tortuosum. Our previous studies had demonstrated that hirsutanol A exhibited potent cytotoxic effect on many kinds of cancer cell lines. In the current study, the antitumor activity of hirsutanol A and its molecular mechanisms were investigated. Methods: Hirsutanol A induced growth inhibition and apoptotic cell death of human colon cancer SW620 cells and human breast cancer MDA-MB-231 cells were determined using MTT assay and flow cytometry assay, respectively. The effect of hirsutanol A on intrinsic ROS level and change in mitochondrial membrane potential (ΔΨm) of different cell lines were also measured by flow cytometry assay. The function of JNK was compromised by JNK siRNA or JNK inhibitor SP600125. The expression of cytochrome c, p-JNK, p-c-Jun after treatment with hirsutanol A were detected by Western blot analysis. Finally, the in vivo anti-tumor effect of hirsutanol A was examined in human cancer cell SW620 xenograft model. Results: The results showed that hirsutanol A significantly induced apoptosis, mitochondrial-independent increase of Reactive Oxygen Species (ROS) level, change of mitochondrial membrane potential, release of cytochrome c in human cancer cells. Preventing increase of ROS level using the potent antioxidant N-acetyl-L-cysteine (NAC) markedly decreased hirsutanol A-induced apoptosis. In addition, JNK signaling pathway was activated by hirsutanol A through elevating ROS level. Blockade of JNK signaling pathway by JNK specific inhibitor SP600125 enhanced apoptosis and hirsutanol A-induced ROS accumulation. Also, hirsutanol A exhibited antitumor activity in human cancer cell SW620 xenograft model. Conclusion: These data suggested that hirsutanol A inhibited tumor growth through triggering ROS production and apoptosis. [ABSTRACT FROM AUTHOR]

Summary: The objective of this study was to explore the influence of isoflurane on the cell proliferation, apoptosis, and invasion of Tca8113 and HSC2 cell lines in vitro. MTT test was used to detect the cell proliferation. It was performed 72h after exposure to isoflurane to make sure that a time for normal cell cycle progression was allowed. The cell apoptosis of Tca8113 and HSC2 cell lines were detected by flow cytometry. We used transwell chamber to detect the cell invasion of Tca8113 and HSC2 cell lines. There was a statistically significant increase of cell proliferation in Tca8113 and HSC2 cell lines after exposure to 2% isoflurane for 3 and 6h. The difference between 3 and 6h group is statistically significant in Tca8113 and HSC2 cell lines. Flow cytometry showed that there was a decrease of cell apoptosis in Tca8113 and HSC2 cell lines after exposure to 2% isoflurane for 3 and 6h. Transwell test showed there was a statistically significant increase of cell invasion in Tca8113 and HSC2 cell lines after exposure to 2% isoflurane for 3 and 6h, and it showed a significant difference between 3h group and 6h group of Tca8113 cell line. Our results demonstrated that isoflurane increased malignancy of head and neck squamous cell carcinoma cell lines in vitro. Isoflurane might enhance tumor development and promote metastasis of tumor cells in HNSCC patients. It is suggested that it might be more suitable to choose total intravenous anesthesia for HNSCC patients. [ABSTRACT FROM AUTHOR]

In this article, we have focused on the structure identification of Euphorbia factor L3 belonging to the lathyrane diterpenoids isolated from Caper Euphorbia Seed. Its anticancer activity in vitro against lung cancer A549 cells was also investigated and the IC50 values were 34.04 ± 3.99 μM. Furthermore, Euphorbia factor L3 could induce apoptosis in A549 cells via the mitochondrial pathway including loss of mitochondrial potential and release of cytochrome c. [ABSTRACT FROM AUTHOR]

Background: Pharmacological intervention of redox balance in cancer cells often results in oxidative stress-mediated apoptosis, attracting much attention for the development of a new generation of targeted therapy in cancer. However, little is known about mechanisms underlying the conversion from oxidative signaling to downstream activities leading cells to death. Methodology/Principal Findings: We here report a systematic detection of transcriptome changes in response to oxidative signals generated in leukemia cells upon fenretinide treatment, implicating the occurrence of numerous stress-responsive events during the fenretinide induced apoptosis, such as redox response, endoplasmic reticulum stress/unfolded protein response, translational repression and proteasome activation. Moreover, the configuration of these relevant events is primarily orchestrated by stress responsive transcription factors, as typically highlighted by NF-E2-related factor-2 (NRF2) and heat shock factor 1 (HSF1). Several lines of evidence suggest that the coordinated regulation of these transcription factors and thus their downstream genes are involved in converting oxidative signaling into downstream stress-responsive events regulating pro-apoptotic and apoptotic activities at the temporal and spatial levels, typifying oxidative stressmediated programmed death rather than survival in cancer cells. Conclusions/Significance: This study provides a roadmap for understanding oxidative stress-mediated apoptosis in cancer cells, which may be further developed into more sophisticated therapeutic protocols, as implicated by synergistic induction of cell apoptosis using proteasome inhibitors with fenretinide. [ABSTRACT FROM AUTHOR]

Caspase-3 is an attractive therapeutic target for treatment of diseases involving disregulated apoptosis. We report here the mechanism of caspase-3 inactivation by isoquinoline- 1,3,4-trione derivatives. Kinetic analysis indicates the compounds can irreversibly inactivate caspase-3 in a 1,4-dithiothreitol (DTT)- and oxygen-dependent manner, implying that a redox cycle might take place in the inactivation process. Reactive oxygen species detection experiments using a chemical indicator, together with electron spin resonance measurement, suggest that ROS can be generated by reaction of isoquinoline-1,3,4-trione derivatives with DTT. Oxygen-free radical scavenger catalase and superoxide dismutase eliciting the inactivation of caspase-3 by the inhibitors confirm that ROS mediates the inactivation process. Crystal structures of caspase-3 in complexes with isoquinoline-1,3,4-trione derivatives show that the catalytic cysteine is oxidized to sulfonic acid (-SO3H) and isoquinoline-1,3,4-trione derivatives are bound at the dimer interface of caspase-3. Further mutagenesis study shows that the binding of the inhibitors with caspase-3 appears to be nonspecific. Isoquinoline-1,3,4-trione derivative-catalyzed caspase-3 inactivation could also be observed when DTT is substituted with dihydrolipoic acid, which exists widely in cells and might play an important role in the in vivo inactivation process in which the inhibitors inactivate caspase-3 in cells and then prevent the cells from apoptosis. These results provide valuable information for further development of small molecular inhibitors against caspase-3 or other oxidation-sensitive proteins. [ABSTRACT FROM AUTHOR]

The avian virus-derived protein apoptin induces p53-independent apoptosis in a tumor-specific way. Apoptin acts as a multimeric complex and forms superstructures upon binding to DNA. In tumor cells, apoptin is phosphorylated and mainly nuclear, whereas in nor-real cells it is unphosphorylated, cytoplasmic, and becomes readily neutralized. Interestingly, apoptin phosphorylation, nuclear translocation, and apoptosis can transiently be induced in normal cells by cotransfecting SV40 large T oncogenic, indicating that apoptin recognizes early stages of oncogenic transformation. In cancer cells, apoptin appears to recognize survival signals, which it is able to redirect into cell death impulses. Apoptin targets include DEDAF, Nur77, Nmi, Hippi, and the potential drug target APC1. Apoptin-transgenic mice and animal tumor models have revealed apoptin as a safe and efficient antitumor agent, resulting in significant tumor regression. Future antitumor therapies could use apoptin either as a therapeutic bullet or as an early sensor of druggable tumor-specific processes. [ABSTRACT FROM AUTHOR]

The use of exogenous cytokines is part of translational immune-antiretroviral approaches to induce immune reconstitution and possibly eliminate the persistence of human immunodeficiency virus type 1 (HIV-1) in virally suppressed infected individuals on highly active antiretroviral therapy (HAART). Recently, our laboratories demonstrated that interleukin-7 (IL-7) has significant efficiency in stimulating HIV-1 replication from proviral latency in CD4+ T lymphocytes of infected patients. The authors now investigated the possible role of IL-7 in HIV-1–associated dementia (HAD). The authors demonstrated that the IL-7 receptor is expressed on both human neurons (i.e., differentiated NT2 cells) and human astrocytes, with relatively higher mRNA levels in neurons. The translational protein levels of IL-7 receptor α were not proportional to those of the mRNA levels in these central nervous system (CNS)-based cell types. Exogenous IL-7 was observed to only slightly down-regulate IL-7 receptor α expression on both neurons and astrocytes, as assayed by Western blotting. Instead of promoting survival, surprisingly, exogenous IL-7 induced neuronal apoptosis, as detected by TUNEL assays. Furthermore, IL-7 augmented neuronal apoptosis induced by HIV-1 gp120. Human apoptosis genomic microarray analyses of IL-7–treated human neurons showed up-regulated expression of proapoptotic genes: protein kinases, caspase-10, FAST kinase, tumor necrosis factor (TNF) receptor, and BCL2-antagonist of cell death. These data suggest that IL-7 leads to neuronal apoptosis by a molecular mechanism(s) that occurs via Fas-mediated activation-induced cell death. These studies may therefore not only be key in evaluating the potential use of IL-7 in vivo as a therapeutic modality, but also suggest that IL-7, which is increased endogenously in HIV-1–infected individuals late in disease, may be involved in the neuronal apoptosis demonstrated during HAD. [ABSTRACT FROM AUTHOR]

Glial cell line-derived neurotrophic factor (GDNF) has been shown to protect and restore dopamine (DA) neurons in injury models and is being evaluated for the treatment of Parkinson's disease. Nevertheless, little is known of its physiological role. We have shown that GDNF suppresses apoptosis in DA neurons of the substantia nigra (SN) postnatally both in vitro and during their first phase of natural cell death in viva. Furthermore, intrastriatal injection of neutralizing antibodies augments cell death, suggesting that endogenous GDNF plays a role as a target-derived factor. Such a role would predict that overexpression of GDNF in striatum would increase the surviving number of SN DA neurons. To test this hypothesis, we used the tetracycline-dependent transcription activator (tTA)/tTA-responsive promoter system to create mice that overexpress GDNF selectively in the striatum, cortex, and hippocampus. These mice demonstrate an increased number of SN DA neurons after the first phase of natural cell death. However, this increase does not persist into adulthood. As adults, these mice also do not have increased dopaminergic innervation of the striatum. They do, however, demonstrate increased numbers of ventral tegmental area (VTA) neurons and increased innervation of the cortex. This morphologic phenotype is associated with an increased locomotor response to amphetamine. We conclude that striatal GDNF is necessary and sufficient to regulate the number of SN DA neurons surviving the first phase of natural cell death, but it is not sufficient to increase their final adult number. GDNF in VTA targets, however, is sufficient to regulate the adult number of DA neurons. [ABSTRACT FROM AUTHOR]