Epigenetic modifications of the genome play important roles in controlling gene transcription thus regulating several molecular and cellular processes. A novel epigenetic modification - 5-hydroxymethylcytosine (5hmC) - has been recently described and attracted a lot of attention due to its possible involvement in the active DNA demethylation mechanism. TET enzymes are dioxygenases capable of oxidizing the methyl group of 5-methylcytosines (5mC) and thus converting 5mC into 5hmC. Although most of the work on TET enzymes and 5hmC has been carried out in embryonic stem (ES) cells, the highest levels of 5hmC occur in the brain and in neurons, pointing to a role for this epigenetic modification in the control of neuronal differentiation, neural plasticity and brain functions. Here we review the most recent advances on the role of TET enzymes and DNA hydroxymethylation in neuronal differentiation and function.

One of the cancer molecular hallmarks is a deviant energetic metabolism, known as the Warburg effect, whereby the rate of glucose uptake is significantly increased and a high rate of glycolysis and lactic acid production occurs even when oxygen is present-"aerobic lactatogenesis". Accordingly, GLUT1 and MCT1, which are the main glucose and lactate transporters in cancer cells, respectively, have been proposed as oncogenes and are currently seen as potential therapeutic targets in cancer treatment. Polyphenols, commonly contained in fruits and vegetables, have long been associated with a protective role against cancer. Generally considered as nontoxic, dietary polyphenols are considered ideal chemopreventive and possibly chemotherapeutic agents. Several mechanisms of action of polyphenols in breast cancer cells have been proposed including modulation of intracellular signaling, induction of apoptosis through redox regulation or modulation of epigenetic alterations. Additionally, in vitro studies have shown that several polyphenols act as specific inhibitors of glucose transport in breast cancer cell lines and an association between their anticarcinogenic effect and inhibition of glucose cellular uptake has been described. Also, some polyphenols were found to inhibit lactate transport. Importantly, some polyphenols behave as inhibitors of both glucose and lactate cellular uptake by breast cancer cells and these compounds are thus very interesting in the context of a chemopreventive effect, because they deplete breast cancer cells of their two most important energy suppliers. So, the antimetabolic effect of polyphenols should be regarded as a mechanism of action contributing to their chemopreventive/chemotherapeutic potential in relation to breast cancer.

Silk fibroin (SF) hydrogels can be obtained via self-assembly, but this process takes several days or weeks, being unfeasible to produce cell carrier hydrogels. In this work, a phospholipid, namely, 1,2-dimyristoyl-sn-glycero-3-phospho-(1′-rac-glycerol) sodium salt (DMPG), was used to induce and accelerate the gelation process of SF solutions. Due to the amphipathic nature and negative charge of DMPG, electrostatic and hydrophobic interactions between the phospholipids and SF chains will occur, inducing the structural transition of SF chains to the beta sheet and consequently a rapid gel formation is observed (less than 50 min). Moreover, the gelation time can be controlled by varying the lipid concentration. To assess the potential of the hydrogels as cell carriers, several mammalian cell lines, including L929, NIH/3T3, SaOS-2, and CaSki, were encapsulated into the hydrogel. The silk-based hydrogels supported the normal growth of fibroblasts, corroborating their cytocompatibility.Interestingly, an inhibition in the growth of cancer-derived cell lines was observed.Therefore, DMPG-induced SF hydrogels can be successfully used as a 3D platform for in situ cell encapsulation, opening promising opportunities in biomedical applications, such as in cell therapies and tissue regeneration.Bombyx mori "Nangnoi Srisaket 1" Thai silk cocoons were obtained from Queen Sirikit Sericulture Center, Srisaket province, Thailand.DMPG and 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) were purchased from Avanti Lipids Polar, USA. Cell culture medium and all other reagents were purchased from Thermo Fisher Scientific.All reagents were of analytical grade. | SF solution preparation and its zeta potential determinationSF solution was prepared using the method adapted from Kim, Park, Joo Kim, Wada, and Kaplan (2005). In brief, silk cocoons were boiled in 0.02 M Na 2 CO 3 for 20 min, followed by their thoroughly washing with distilled water. The extracted silk fibre was then dissolved in 9.3 M LiBr at 60°C for 4 hr. The silk solution was dialysed against distilled water using a dialysis tube (MWCO 12-16 kDa; Sekisui, Japan) for 48 hr. The final concentration was approximately 6%, which was determined from dried solid weight.Analysis of zeta potential of the SF solution was conducted using laser doppler electrophoresis in a Zetasizer Nano ZS equipment (Malvern Instruments) at 25°C.

TET enzymes oxidize 5-methylcytosine (5mC) into 5-hydroxymethylcytosine (5hmC), a process thought to be intermediary in an active DNA demethylation mechanism. Notably, 5hmC is highly abundant in the brain and in neuronal cells. Here, we interrogated the function of Tet3 in neural precursor cells (NPCs), using a stable and inducible knockdown system and an in vitro neural differentiation protocol. We show that Tet3 is upregulated during neural differentiation, whereas Tet1 is downregulated. Surprisingly, Tet3 knockdown led to a de-repression of pluripotency-associated genes such as Oct4, Nanog or Tcl1, with concomitant hypomethylation. Moreover, in Tet3 knockdown NPCs, we observed the appearance of OCT4positive cells forming cellular aggregates, suggesting de-differentiation of the cells. Notably, Tet3 KD led to a genome-scale loss of DNA methylation and hypermethylation of a smaller number of CpGs that are located at neurogenesis-related genes and at imprinting control regions (ICRs) of Peg10, Zrsr1 and Mcts2 imprinted genes. Overall, our results suggest that TET3 is necessary to maintain silencing of pluripotency genes and consequently neural stem cell identity, possibly through regulation of DNA methylation levels in neural precursor cells.

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