Synergetic Effect of Tumor Treating Fields and Zinc Oxide Nanoparticles on Cell Apoptosis and Genotoxicity of Three Different Human Cancer Cell Lines.

Cancer remains a leading cause of death worldwide, despite extraordinary progress. So, new cancer treatment modalities are needed. Tumor-treating fields (TTFs) use low-intensity, intermediate-frequency alternating electric fields with reported cancer anti-mitotic properties. Moreover, nanomedicine is a promising therapy option for cancer. Numerous cancer types have been treated with nanoparticles, but zinc oxide nanoparticles (ZnO NPs) exhibit biocompatibility. Here, we investigate the activity of TTFs, a sub-lethal dose of ZnO NPs, and their combination on hepatocellular carcinoma (HepG2), the colorectal cancer cell line (HT-29), and breast cancer cell lines (MCF-7). The lethal effect of different ZnO NPs concentrations was assessed by sulforhodamine B sodium salt assay (SRB). The cell death percent was determined by flow cytometer, the genotoxicity was evaluated by comet assay, and the total antioxidant capacity was chemically measured. Our results show that TTFs alone cause cell death of 14, 8, and 17% of HepG2, HT-29, and MCF-7, respectively; 10 µg/mL ZnO NPs was the sub-lethal dose according to SRB results. The combination between TTFs and sub-lethal ZnO NPs increased the cell death to 29, 20, and 33% for HepG2, HT-29, and MCF-7, respectively, without reactive oxygen species increase. Increasing NPs potency using TTFs can be a novel technique in many biomedical applications.

Expression screening using a Medaka cDNA library identifies evolutionarily conserved regulators of the p53/Mdm2 pathway.

BACKGROUND: The p53 tumor suppressor protein is mainly regulated by alterations in the half-life of the protein, resulting in significant differences in p53 protein levels in cells. The major regulator of this process is Mdm2, which ubiquitinates p53 and targets it for proteasomal degradation. This process can be enhanced or reduced by proteins that associate with p53 or Mdm2 and several proteins have been identified with such an activity. Furthermore, additional ubiquitin ligases for p53 have been identified in recent years. Nevertheless, our understanding of how p53 abundance and Mdm2 activity are regulated remains incomplete. Here we describe a cell culture based overexpression screen to identify evolutionarily conserved regulators of the p53/Mdm2 circuit. The results from this large-scale screening method will contribute to a better understanding of the regulation of these important proteins. METHODS: Expression screening was based on co-transfection of H1299 cells with pools of cDNA's from a Medaka library together with p53, Mdm2 and, as internal control, Ror2. After cell lysis, SDS-PAGE/WB analysis was used to detect alterations in these proteins. RESULTS: More than one hundred hits that altered the abundance of either p53, Mdm2, or both were identified in the primary screen. Subscreening of the library pools that were identified in the primary screen identified several potential novel regulators of p53 and/or Mdm2. We also tested whether the human orthologues of the Medaka genes regulate p53 and/or Mdm2 abundance. All human orthologues regulated p53 and/or Mdm2 abundance in the same manner as the proteins from Medaka, which underscores the suitability of this screening methodology for the identification of new modifiers of p53 and Mdm2. CONCLUSIONS: Despite enormous efforts in the last two decades, many unknown regulators for p53 and Mdm2 abundance are predicted to exist. This cross-species approach to identify evolutionarily conserved regulators demonstrates that our Medaka unigene cDNA library represents a powerful tool to screen for these novel regulators of the p53/Mdm2 pathway.

Citalopram, an antipsychotic agent, induces G1/G0 phase cell cycle arrest and promotes apoptosis in human laryngeal carcinoma HEP-2 cells.

Human laryngeal squamous carcinoma (LSCC) is a common malignant tumor in the head and neck. Despite the recently developed therapies for the treatment of LSCC, patients' overall survival rate still did not enhance remarkably; this highlights the need to formulate alternative strategies to develop novel treatments. The antitumor effects of antidepressant drugs such as citalopram have been reported on several cancer cells; however, they have yet to be investigated against LSCC. The current study was directed to explore the possible antitumor effects of citalopram on human laryngeal carcinoma cell lines (HEP-2). HEP-2 cells were cultured and treated with different doses of citalopram (50-400 µM) for 24, 48, and 72 h. The effects of citalopram on the viability of cancer cells were determined by the MTT assay. In addition, apoptosis and cell cycle analysis were performed by flow cytometry. Moreover, evaluation of the expression of proapoptotic and apoptotic proteins, such as cytochrome c, cleaved caspases 3 and 9, Bcl-2, and BAX, was performed by western blotting analysis. Our results revealed that citalopram significantly suppressed the proliferation of HEP-2 cells through the upregulation of p21 expression, resulting in the subsequent arrest of the cell cycle at the G0/G1 phase. Furthermore, citalopram treatment-induced HEP-2 cell apoptosis; this was indicated by the significant increase of cytochrome c, cleaved caspases 3 and 9, and BAX protein expression. On the contrary, Bcl-2 protein expression was significantly downregulated following treatment with citalopram. The ultrastructure studies were in accordance with the protein expression findings and showed clear signs of apoptosis with ring chromatin condensation upon treatment with citalopram. These findings suggest that citalopram's anti-tumor activities on HEP-2 cells entailed stimulation of the intrinsic apoptotic pathway, which was mediated via Bcl-2 suppression.

TRIM25 targets p300 for degradation.

p300 is an important transcriptional co-factor. By stimulating the transfer of acetyl residues onto histones and several key transcription factors, p300 enhances transcriptional initiation and impacts cellular processes including cell proliferation and cell division. Despite its importance for cellular homeostasis, its regulation is poorly understood. We show that TRIM25, a member of the TRIM protein family, targets p300 for proteasomal degradation. However, despite TRIM25's RING domain and E3 activity, degradation of p300 by TRIM25 is independent of TRIM25-mediated p300 ubiquitination. Instead, TRIM25 promotes the interaction of p300 with dynein, which ensures a microtubule-dependent transport of p300 to cellular proteasomes. Through mediating p300 degradation, TRIM25 affects p300-dependent gene expression.

Delay in development and behavioural abnormalities in the absence of p53 in zebrafish.

p53 is well-known for its tumour-suppressive activity. However, in the past decade it became clear that p53 is also involved in other processes including stem cell proliferation, differentiation and animal development. To investigate the role of p53 in early embryonic development, we targeted p53 by CRISPR/Cas9 to make a p53 knock-out zebrafish (Danio rerio). Our data show developmental and behavioural effects in p53-deficient zebrafish embryos and larvae. Specifically, we found that early development of zebrafish was clearly delayed in the absence of p53. However, after 1 day (1 dpf), the p53-deficient embryos appeared to recover, as evidenced by a similar level of pigmentation at 26 hpf, similar size of the eye at 4 dpf and only a minor difference in body size at 4 dpf compared to p53 wild-type siblings. The recovery of development after 1 dpf in p53-deficient embryos could be due to a compensatory mechanism involving other p53 family members. p63 and p73 were found over-expressed with respect to wild-type siblings. However, despite this adaptation, the hatching time remained delayed in p53-/- zebrafish. In addition to differences in development, p53-null zebrafish embryos also showed differences in behaviour. We observed an overall reduced activity and a reduced travel distance under non-stressed conditions and after exposing the larvae to vibration. We also observed a longer latency until the larvae started to move after touching with a needle. Overall, these data indicate that p53 is involved in early development and locomotion activities.

Fam83F induces p53 stabilisation and promotes its activity.

p53 is one of the most important tumour suppressor proteins currently known. It is activated in response to DNA damage and this activation leads to proliferation arrest and cell death. The abundance and activity of p53 are tightly controlled and reductions in p53's activity can contribute to the development of cancer. Here, we show that Fam83F increases p53 protein levels by protein stabilisation. Fam83F interacts with p53 and decreases its ubiquitination and degradation. Fam83F is induced in response to DNA damage and its overexpression also increases p53 activity in cell culture experiments and in zebrafish embryos. Downregulation of Fam83F decreases transcription of p53 target genes in response to DNA damage and increases cell proliferation, identifying Fam83F as an important regulator of the DNA damage response. Overexpression of Fam83F also enhances migration of cells harbouring mutant p53 demonstrating that it can also activate mutant forms of p53.

p53 Is Active in Human Amniotic Fluid Stem Cells.

Despite increasing interest in human amniotic fluid cells, very little is known about the regulation and function of p53 in this cell type. In this study, we show that undifferentiated human amniotic fluid cells express p53, yet at lower levels than in cancer cells. The p53 protein in amniotic fluid cells is mainly localized in the nuclei, however, its antiproliferative activity is compromised in these cells. Igf2, a maternal imprinted gene, and c-jun, a proto-oncogene, are regulated by p53 in these cells. DNA damage leads to an increase in p53 abundance in human amniotic fluid cells and to transcriptional activation of its target genes. Interestingly, cell differentiation toward the neural lineage leads to p53 induction as differentiation progresses.

Two Birds with One Stone: Possible Dual-Role of Oxytocin in the Treatment of Diabetes and Osteoporosis.

Oxytocin (OT), a hormone most commonly associated with labor and lactation, may have a wide variety of physiological and pathological functions, which makes OT and its receptor potential targets for drug therapy. In this review, we highlight the newly discovered metabolic role of OT in diabetes and its complication, such as diabetic osteopathy. OT may have positive metabolic effects; this is based on the change in glucose metabolism, lipid profile, and insulin sensitivity. It may modify glucose uptake and insulin sensitivity both through direct and indirect effects. It may also cause regenerative changes in diabetic pancreatic islet cells. Moreover, it has an anabolic effect on the bone biology. So, the activation of the OT receptor pathway by infusion of OT, OT analogs, or OT agonists may represent a promising approach for the treatment of diabetes and some of its complications, including diabetic osteopathy.