Evaluation of the antioxidant, anti-inflammatory, and anti-tumoral properties of bioactive compounds extracted from murta berries (Ugni molinae T.) dried by different methods.

This study evaluated the effects of different drying methods (freeze drying, vacuum drying, infrared drying, convective drying, and sun drying) on the biological properties of berries from the Chilean murta (Ugni molinae Turcz) shrub. Physical-chemical properties (proximal composition, dietary fiber, sugars) were determined. Total phenolic content through the method of Folin-Ciocalteau, the profile of phenol compounds was determined by HPLC, and antioxidant potential by DPPH and ORAC assays were also evaluated. The topic anti-inflammatory effect was evaluated by mice´s ear edema, and in vitro anti-tumoral activity was tested by MTT assay. The chemical properties of dried berries differed significantly based on the drying method: freeze-dried murta berries showed increased total phenolic content extracted over fresh and dried samples. In addition, this lyophilized extract stood out in its antioxidant potential, in both assays evaluated (DPPH and ORAC), compared to the other drying methods. Notwithstanding, vacuum- and infrared-dried murta also showed a higher ORAC value. Antioxidant potential was significantly associated with phenolic compounds catechin and pyrogallol, which were the most abundant phenolic compounds present in all samples. The anti-inflammatory activity was most effective under freeze-drying and vacuumdrying conditions. Moreover, vacuum drying and infrared drying best preserved the anti-tumoral effect on cancer cells.

Modulation of Synaptic Plasticity Genes Associated to DNA Damage in a Model of Huntington's Disease.

Huntington's disease (HD) is a disease characterized by the progressive degeneration of nerve cells in the brain. DNA damage has been implicated in many neurological disorders; however, the association between this damage and the impaired signaling related to neurodegeneration is still unclear. The transcription factor c-AMP-responsive element binding protein (CREB) has a relevant role in the neuronal plasticity process regulating the expression of several genes, including brain-derived neurotrophic factor (BDNF). Here we analyzed the direct link between DNA damage and the expression of genes involved in neuronal plasticity. The study was performed in model cell lines STHdhQ7 (wild type) and STHdhQ111 (HD model). Treatment with Etoposide (Eto) was used to induce double-strand breaks (DSBs) to evaluate the DNA damage response (DDR) and the expression of synaptic plasticity genes. Eto treatment induced phosphorylation of ATM (p-ATM) and H2AX (γH2AX), markers of DDR, in both cell lines. Interestingly, upon DNA damage, STHdhQ7 cells showed increased expression of activity-regulated cytoskeleton associated protein (Arc) and BDNF when compared to the HD cell line model. Additionally, Eto induced CREB activation with a differential localization of its co-activators in the cell types analyzed. These results suggest that DSBs impact differentially the gene expression patterns of plasticity genes in the normal cell line versus the HD model. This effect is mediated by the impaired localization of CREB-binding protein (CBP) and histone acetylation in the HD model. Our results highlight the role of epigenetics and DNA repair on HD and therefore we suggest that future studies should explore in depth the epigenetic landscape on neuronal pathologies with the goal to further understand molecular mechanisms and pinpoint therapeutic targets.

Metabolic changes induced by DNA damage in Ramos cells: exploring the role of mTORC1 complex.

DNA damage induces the activation of many different signals associated with repair or cell death, but it is also connected with physiological events, such as adult neurogenesis and B-cell differentiation. DNA damage induces different signaling pathways, some of them linked to important metabolic changes. The mTORC1 pathway has a central role in the regulation of growth processes and cell division in response to environmental changes and also controls protein synthesis, lipid biogenesis, nucleotide synthesis, and expression of glycolytic genes. Here, we report that double-strand breaks induced with etoposide affect the expression of genes encoding different enzymes associated with specific metabolic pathways in Ramos cells. We also analyzed the role of mTOR signaling, demonstrating that double-strand breaks induce downregulation of mTOR signaling. Specific inhibition of mTORC1 using rapamycin also induced changes in the expression of metabolic genes. Finally, we demonstrated that DNA damage and rapamycin can regulate glucose uptake. In summary, our findings show that etoposide and rapamycin affect the expression of metabolic genes as well as apoptotic and proliferation markers in Ramos cells, increasing our understanding of cancer metabolism.

The Effect of Resveratrol on Cell Viability in the Burkitt's Lymphoma Cell Line Ramos.

Resveratrol is a polyphenolic natural compound produced by a variety of crops. Currently, resveratrol is considered a multi-target anti-cancer agent with pleiotropic activity, including the ability to prevent the proliferation of malignant cells by inhibiting angiogenesis and curtailing invasive and metastatic factors in many cancer models. However, the molecular mechanisms mediating resveratrol-specific effects on lymphoma cells remain unknown. To begin tackling this question, we treated the Burkitt's lymphoma cell line Ramos with resveratrol and assessed cell survival and gene expression. Our results suggest that resveratrol shows a significant anti-proliferative and pro-apoptotic activity on Ramos cells, inducing the DNA damage response, DNA repairing, and modulating the expression of several genes that regulate the apoptotic process and their proliferative activity.

Herpes Simplex Virus Type 1 Neuronal Infection Perturbs Golgi Apparatus Integrity through Activation of Src Tyrosine Kinase and Dyn-2 GTPase.

Herpes simplex virus type 1 (HSV-1) is a ubiquitous pathogen that establishes a latent persistent neuronal infection in humans. The pathogenic effects of repeated viral reactivation in infected neurons are still unknown. Several studies have reported that during HSV-1 epithelial infection, the virus could modulate diverse cell signaling pathways remodeling the Golgi apparatus (GA) membranes, but the molecular mechanisms implicated, and the functional consequences to neurons is currently unknown. Here we report that infection of primary neuronal cultures with HSV-1 triggers Src tyrosine kinase activation and subsequent phosphorylation of Dynamin 2 GTPase, two players with a role in GA integrity maintenance. Immunofluorescence analyses showed that HSV-1 productive neuronal infection caused a scattered and fragmented distribution of the GA through the cytoplasm, contrasting with the uniform perinuclear distribution pattern observed in control cells. In addition, transmission electron microscopy revealed swollen cisternae and disorganized stacks in HSV-1 infected neurons compared to control cells. Interestingly, PP2, a selective inhibitor for Src-family kinases markedly reduced these morphological alterations of the GA induced by HSV-1 infection strongly supporting the possible involvement of Src tyrosine kinase. Finally, we showed that HSV-1 tegument protein VP11/12 is necessary but not sufficient to induce Dyn2 phosphorylation. Altogether, these results show that HSV-1 neuronal infection triggers activation of Src tyrosine kinase, phosphorylation of Dynamin 2 GTPase, and perturbation of GA integrity. These findings suggest a possible neuropathogenic mechanism triggered by HSV-1 infection, which could involve dysfunction of the secretory system in neurons and central nervous system.

Modulation of the AMPK/Sirt1 axis during neuronal infection by herpes simplex virus type 1.

Currently, it is unclear whether a neuron that undergoes viral reactivation and produces infectious particles survives and resumes latency or is killed, which is intriguing even if still unanswered. Previous reports have shown that herpes simplex virus type 1 (HSV-1) inhibits apoptosis during early infection, but is pro-apoptotic during productive infection. Taking in consideration that the stress sensors AMPK and Sirt1 are involved in neuronal survival and neuroprotection, we hypothesized that HSV-1 could activate the AMPK/Sirt1 axis as a strategy to establish latency through inhibition of apoptosis and restoration of the energy status. These effects could be accomplished through deacetylation of pro-apoptotic protein p53 and regulation of the master regulator of mitochondrial biogenesis and function PGC-1α and its target gene TFAM. Accordingly, we evaluated the AMPK/Sirt1 axis and its targets p53, PGC-1α, and acetyl CoA carboxylase in mice neuronal cultures infected with HSV-1 by western blot, RT-qPCR, and immunofluorescence analyses. Herein, we show that HSV-1 differentially modulates the AMPK/Sirt1 axis during the course of infection. In fact, during early infection (2 hpi) activated AMPK (p-AMPK) was down-regulated, but thereafter recovered gradually. In contrast, the levels of acetylated-p53 increased during the first hours post infection, but afterwards were reduced in parallel with the activation of Sirt1. However, acetylated-p53 peaked again at 18 hpi during productive infection, suggesting an activation of apoptosis. Strikingly, acetylated-p53, Sirt1, and p-AMPK apparently translocate from the nucleus to the cytoplasm after 4 hpi, where they accumulate in discrete foci in the perinuclear region. These results suggest that HSV-1 modulates the AMPK/Sirt1 axis differentially during the course of infection interfering with pro-apoptotic signaling and regulating mitochondrial biogenesis.