Mir-192 suppresses apoptosis and promotes proliferation in esophageal aquamous cell caicinoma by targeting Bim

MicroRNAs (miRNAs) are small, non-coding RNAs of endogenous origin. Accumulating studies have shown aberrant miRNA expression plays an important role in many tumor types. However, the mechanisms by which miRNAs regulate esophageal squamous cell carcinoma (ESCC) development remain poorly understood. In the present study, we assayed expression level of miR-192 in ESCC tissues and cell lines by real-time PCR, and defined the target gene and biological function by luciferase reporter assay, Western blot and apoptosis assay. We first verified that the expression level of miR-192 was significantly increased in ESCC tissues and cancer cells. Moreover, miR-192 over-expression inhibited cells apoptosis and promoted ESCC cells proliferation. We further demonstrated that miR-192 directly targeted 3’-UTR of Bim gene, and inhibited its protein expression. Importantly, Bim could reduce ESCC cells apoptosis ability induced by miR-192. These data suggest an important role of miR-192 in the molecular etiology of ESCC and implicate the potential application of miR-192 in ESCC therapy.     

Glycyrrhizic acid prevents ultraviolet‐B‐induced photodamage: a role for mitogen‐activated protein kinases,nuclear factor kappa B and mitochondrial apoptotic pathway

Glycyrrhizic acid (GA), a natural triterpene, has received attention as an agent that has protective effects against chronic diseases including ultraviolet UV‐B‐induced skin photodamage. However, the mechanism of its protective effect remains elusive. Here, we used an immortalized human keratinocyte cell line (HaCaT) and a small animal model (BALB/c mice), to investigate the protective effects of GA against UV‐B‐induced oxidative damage, and additionally, delineated the molecular mechanisms involved in the UV‐B‐mediated inflammatory and apoptotic response. In the HaCaT cells, GA inhibited the UV‐B‐mediated increase in intracellular reactive oxygen species (ROS) and down‐regulated the release of pro‐inflammatory cytokines interleukin (IL)‐1α, ‐1β and ‐6, tumor necrosis factor (TNF)‐α and prostaglandin E2 (PGE2). GA inhibited UV‐B‐mediated activation of p38 and JNK MAP kinases, COX‐2 expression and nuclear translocation of NF‐κB. Furthermore, GA inhibited UV‐B‐mediated apoptosis by attenuating translocation of Bax from the cytosol to mitochondria, thus preserving mitochondrial integrity. GA‐treated HaCaT cells also exhibited elevated antiapoptotic Bcl‐2 protein, concomitant with reduced caspase‐3 cleavage and decreased PARP‐1 protein. In BALB/c mice, topical application of GA on dorsal skin exposed to UV‐B irradiation protected against epidermal hyperplasia, lymphocyte infiltration and expression of several inflammatory proteins, p38, JNK, COX‐2, NF‐κB and ICAM‐1. Based on the above findings, we conclude that GA protects against UV‐B‐mediated photodamage by inhibiting the signalling cascades triggered by oxidative stress, including MAPK/NF‐κB activation, as well as apoptosis. Thus, GA has strong potential to be used as a therapeutic/cosmeceutical agent against photodamage.     

Apoptosis pathway signature for prediction of treatment response and clinical outcome in childhood high risk B-Precursor acute lymphoblastic leukemia

The most common cancer in children is acute lymphoblastic leukemia (ALL) and it had high cure rate, especially for B-precursor ALL. However, relapse due to drug resistance and overdose treatment reach the limitations in patient managements. In this study, integration of gene expression microarray data, logistic regression, analysis of microarray (SAM) method, and gene set analysis were performed to discover treatment response associated pathway-based signatures in the original cohort. Results showed that 3772 probes were significantly associated with treatment response. After pathway analysis, only apoptosis pathway had significant association with treatment response. Apoptosis pathway signature (APS) derived from 15 significantly expressed genes had 88% accuracy for treatment response prediction. The APS was further validated in two independent cohorts. Results also showed that APS was significantly associated with induction failure time (adjusted hazard ratio [HR] = 1.60, 95% confidence interval [CI] = [1.13, 2.27]) in the first cohort and significantly associated with event-free survival (adjusted HR = 1.56, 95% CI = [1.13, 2.16]) or overall survival in the second cohort (adjusted HR = 1.74, 95% CI = [1.24, 2.45]). APS not only can predict clinical outcome, but also provide molecular guidance of patient management.     

Cytoprotective effect of rhamnetin on miconazole-induced H9c2 cell damage

BACKGROUND/OBJECTIVESReactive oxygen species (ROS) formation is closely related to miconazole-induced heart dysfunction. Although rhamnetin has antioxidant effects, it remained unknown whether it can protect against miconazole-induced cardiomyocyte apoptosis. Thus, we investigated the effects of rhamnetin on miconazole-stimulated H9c2 cell apoptosis.MATERIALS/METHODSCell morphology was observed by inverted microscope and cell viability was determined using a WelCount™ cell proliferation assay kit. Miconazole-induced ROS production was evaluated by fluorescence-activated cell sorting with 6-carboxy-2'',7''-dichlorofluoroscein diacetate (H₂DCF-DA) stain. Immunoblot analysis was used to determine apurinic/apyrimidinic endonuclease 1 (APE/Ref-1) and cleaved cysteine-aspartic protease (caspase) 3 expression. NADPH oxidase levels were measured using real-time polymerase chain reaction.RESULTSMiconazole (3 and 10 µM) induced abnormal morphological changes and cell death in H9c2 cells. Rhamnetin enhanced the viability of miconazole (3 µM)-treated cells in a dose-dependent manner. Rhamnetin (1 and 3 µM) treatment downregulated cleaved caspase 3 and upregulated APE/Ref-1 expression in miconazole-stimulated cells. Additionally, rhamnetin significantly reduced ROS generation.CONCLUSIONSOur data suggest that rhamnetin may have cytoprotective effects in miconazole-stimulated H9c2 cardiomyocytes via ROS inhibition. This effect most likely occurs through the upregulation of APE/Ref-1 and attenuation of hydrogen peroxide levels.     

Enhanced Human Neutrophil Vitamin C Status,Chemotaxis and Oxidant Generation Following Dietary Supplementation with Vitamin C-Rich SunGold Kiwifruit

Neutrophils are the body’s primary defenders against invading pathogens. These cells migrate to loci of infection where they engulf micro-organisms and subject them to an array of reactive oxygen species and antimicrobial proteins to effect killing. Spent neutrophils subsequently undergo apoptosis and are cleared by macrophages, thereby resolving the inflammatory episode. Neutrophils contain high concentrations of vitamin C (ascorbate) and this is thought to be essential for their function. This may be one mechanism whereby vitamin C enhances immune function. The aim of our study was to assess the effect of dietary supplementation with vitamin C-rich SunGold kiwifruit on four important functions of neutrophils: chemotaxis, oxidant generation, extracellular trap formation, and apoptosis. Fourteen young men (aged 18–30 years) with suboptimal plasma vitamin C status (<50 μmol/L) were supplemented for four weeks with two SunGold kiwifruit/day. Plasma vitamin C status was monitored weekly and neutrophil vitamin C levels were assessed at baseline and post-intervention. Neutrophil function assays were carried out on cells isolated at baseline and post-intervention. Plasma vitamin C levels increased to >70 μmol/L (p < 0.001) within one week of supplementation and there was a significant increase in neutrophil vitamin C status following four weeks’ intervention (p = 0.016). We observed a significant 20% increase in neutrophil chemotaxis post-intervention (p = 0.041) and also a comparable increase in oxidant generation (p = 0.031). Supplementation did not affect neutrophil extracellular trap formation or spontaneous apoptosis. Our data indicate that supplementation with vitamin C-rich kiwifruit is associated with improvement of important neutrophil functions, which would be expected to translate into enhanced immunity.     

Antagonistic effects of chloroquine on autophagy occurrence potentiate the anticancer effects of everolimus on renal cancer cells

Renal cell carcinoma is an aggressive disease often asymptomatic and weakly chemo-radiosensitive. Currently, new biologic drugs are used among which everolimus, an mTOR inhibitor, that has been approved for second-line therapy. Since mTOR is involved in the control of autophagy, its antitumor capacity is often limited. In this view, chloroquine, a 4-alkylamino substituted quinoline family member, is an autophagy inhibitor that blocks the fusion of autophagosomes and lysosomes. In the present study, we evaluated the effects of everolimus alone or in combination with chloroquine on renal cancer cell viability and verified possible synergism. Our results demonstrate that renal cancer cells are differently sensitive to everolimus and chloroquine and the pharmacological combination everolimus/chloroquine was strongly synergistic inducing cell viability inhibition. In details, the pharmacological synergism occurs when chloroquine is administered before everolimus. In addition, we found a flow autophagic block and shift of death mechanisms to apoptosis. This event was associated with decrease of Beclin-1/Bcl^-2 complex and parallel reduction of anti-apoptotic protein Bcl^-2 in combined treatment. At last, we found that the enhancement of apoptosis induced by drug combination occurs through the intrinsic mitochondrial apoptotic pathway activation, while the extrinsic pathway is involved only partly following its activation by chloroquine. These results provide the basis for new therapeutic strategies for the treatment of renal cell carcinoma after appropriate clinical trial.     

Human interleukin-10 gene inhibits acute rejection by triggering apoptosis in allograft vascular transplantation

The aim this study is to explore effect of IL-10 on apoptosis of VSMCs in allograft arterial transplantation rats, and to investigate mechanism. SD rats were divied into three groups, including control group (CN, with physiological saline), blank vector group (BV, with blank adenovirus) and combined gene group (CG, with adenovirus carried IL-10 gene). The isolated donor vascular was transfected with the adenovirus carried hIL-10 gene for 30 minutes by immersing method. Forty-five days post transplantation, the grafts were harvested. The allografts pathologioc changes were observed and the size of vascular intima and middle layer of allografts were measured. The expression of hIL-10 was detected by RT-PCR, ELISA and immunohistochemistry, respectively. The repression of Fas/Fasl in artery allografts was also examined by immunohistochemistry method. The results indicated that 45 days after transplantation, the intimal and middle hyperplasia ratio in CG group was significantly lower than that in CN and BV group (P < 0.05). The transgene expression of human interleukin-10 was significantly enhanced in CG group compared to CN and BV group by ELISA, RT-PCR and immunohistochemistry (P < 0.05). The expression of Fas/FasL was higher in CG group compared with the other groups (P < 0.05). The level of apoptotic smooth muscle cells were significantly increased in CG group compared to CN and BV group (P < 0.05). In conclusion, adenovirus mediated IL-10 expression could up-regulate Fas/FasL expression, induce smooth muscle cell apoptosis and alleviate angiosclerosis process. The IL-10 gene transfer to allograft artery could inhibit acute rejection reaction of allograft vascular transplantation.     

Differential role of RIP1 in Smac mimetic-mediated chemosensitization of neuroblastoma cells

We explored the potential of Smac mimetics, which antagonize Inhibitor of Apoptosis (IAP) proteins, for chemosensitization of neuroblastoma (NB). Here, we report that Smac mimetics, e.g. BV6, prime NB cells for chemotherapeutics including the topoisomerase II inhibitor doxorubicin (DOX) and vinca alkaloids such as Vincristine (VCR), Vinblastine (VBL) and Vinorelbine (VNR). Additionally, BV6 acts in concert with DOX or VCR to suppress long-term clonogenic growth. While BV6 causes rapid downregulation of cellular IAP (cIAP)1 protein and nuclear factor-kappaB (NF-κB) activation, DOX/BV6- or VCR/BV6-induced apoptosis occurs independently of NF-κB or TNFα signaling, since overexpression of dominant-negative IκBα superrepressor or the Tumor Necrosis Factor (TNF)α-blocking antibody Enbrel fail to block cell death. Mechanistic studies reveal that Receptor-interacting protein (RIP)1 is required for DOX/BV6-, but not for VCR/BV6-induced apoptosis, since transient or stable knockdown of RIP1 or the pharmacological RIP1 inhibitor necrostatin-1 significantly reduce apoptosis. By comparison, VCR/BV6-mediated apoptosis critically depends on the mitochondrial pathway. VCR/BV6 cotreatment causes phosphorylation of BCL-2 during mitotic arrest, enhanced activation of BAX and BAK and loss of mitochondrial membrane potential (MMP). Additionally, overexpression of BCL-2 profoundly suppresses VCR/BV6-induced apoptosis. Thus, BV6 sensitizes NB cells to chemotherapy-induced apoptosis via distinct initial signaling mechanisms depending on the chemotherapeutic drug. These findings provide novel mechanistic insights into Smac mimetic-mediated chemosensitization of NB.     

Melatonin protects testes against lithium‐pilocarpine‐induced temporal lobe epilepsy in rats: a time course study

Male dysfunction is common in patients with temporal lobe epilepsy (TLE). We evaluated whether melatonin, as a supplement, can play a positive role in reducing the epileptogenesis imposing abnormalities of spermatozoa and testes in epileptic rats. Status epilepticus was induced based on the TLE lithium‐pilocarpine model. Two patterns of melatonin were administered to the epileptic animals along the mean durations of latent (14 days) and chronic (60 days) phases. Sperm parameters, different antioxidant enzyme levels, germ cell apoptosis, body and relative sex organ weights were evaluated in all groups 60 days following SE induction. Chronic TLE caused a significant reduction in sperm parameters. In the testis, the reduced level of antioxidant enzymes was accompanied by a significant increase in malondialdehyde concentration. The presence of oxidant condition in the testes of epileptic animals caused expanded apoptosis in the germ cell layer. Moreover, the amount of weight gain in epileptic animals was more prominent. Melatonin administration was able to improve sperm motility by increasing the total antioxidant level. There was also a significant reduction in the spermatogenic cell line apoptosis and the extra weight gain of melatonin‐treated animals. Melatonin supplementation might be considered as an acceptable cotreatment in epileptic patients.