Identification and Characterization of Cancer Stem Cells from Head and Neck Squamous Cell Carcinoma Cell Lines

Background/Aims: Head and neck squamous cell carcinoma (HNSCC) ranks sixth worldwide for tumor-related mortality. A subpopulation of tumor cells, termed cancer stem cells (CSCs), has the ability to support cancer growth. Therefore, profiling CSC-enriched populations could be a reliable tool to study cancer biology. Methods: We performed phenotypic characterization of 7 HNSCC cell lines and evaluated the presence of CSCs. CSCs from Hep-2 cell line and HNSCC primary cultures were enriched through sphere formation and sphere-forming cells have been characterized both in vitro and in vivo. In addition, we investigated the expression levels of Nicotinamide N-methyltransferase (NNMT), an enzyme overexpressed in several malignancies. Results: CSC markers were markedly expressed in Hep-2 cell line, which was found to be highly tumorigenic. CSC-enriched populations displayed increased expression of CSC markers and a strong capability to form tumors in vivo. We also found an overexpression of CSC markers in tumor formed by CSC-enriched populations. Interestingly, NNMT levels were significantly higher in CSC-enriched populations compared with parental cells. Conclusion: Our study provides an useful procedure for CSC identification and enrichment in HNSCC. Moreover, results obtained seem to suggest that CSCs may represent a promising target for an anticancer therapy

Presynaptic c-Jun N-terminal Kinase 2 regulates NMDA receptor-dependent glutamate release

Activation of c-Jun N-terminal kinase (JNK) signaling pathway is a critical step for neuronal death occurring in several neurological conditions. JNKs can be activated via receptor tyrosine kinases, cytokine receptors, G-protein coupled receptors and ligand-gated ion channels, including the NMDA glutamate receptors. While JNK has been generally associated with postsynaptic NMDA receptors, its presynaptic role remains largely unexplored. Here, by means of biochemical, morphological and functional approaches, we demonstrate that JNK and its scaffold protein JIP1 are also expressed at the presynaptic level and that the NMDA-evoked glutamate release is controlled by presynaptic JNK-JIP1 interaction. Moreover, using knockout mice for single JNK isoforms, we proved that JNK2 is the essential isoform in mediating this presynaptic event. Overall the present findings unveil a novel JNK2 localization and function, which is likely to play a role in different physiological and pathological conditions

The synergic action of amyloid-β peptide and LPS in amyloid plaque formation

Introduction: The current model which assume amyloid-β (Aβ) deposition to be an accidental process resulting from the abnormal behavior of an incidental product of catabolism, is now in contrast with the fact that Aβ, a key player in AD, may have a normal function because belongs to the group of AMPs also called host defense peptide. AMPs such as Aβ, are small molecule peptides which are not only intended to kill pathogens through their antimicrobial activity but they also have a high affinity for bacterial lipopolysaccharide (LPS) or membrane receptors. Once that Aβ is deposited can interact with the LPS which acts as a fibrillogenesis promoter; the positive charge of antibacterial peptides can increase the ability of binding to LPS. The interaction between LPS and Aβ occurs at molecular level, on the basis of their common surfactant properties and considering that detergents and fatty acids are able to form micelles. Material and methods: SH-SY5Y (neuroblastoma cells) were treated with Aβ1–42 in the form of monomers at the starting concentration of 10 µg/ml, with LPS dissolved in order to obtain a starting concentration of1 µg/ml and their combination. Each experiment was performed for 24, 48, and 72 h at 37°C. Western blot analysis for Aβ1-42 monomers, Beclin-1 and Lamp-1; Cytokines (IL-1β) quantification with Elisa; Transmission electron microscopy (TEM) and Statistical analysis were performed. Results: LPS leads to increased production and impaired degradation of Aβ mediated by upregulation of proinflammatory cytokines and inhibition of Aβ degrading enzyme, respectively intracellular accumulation at the endoplasmic reticulum and in lysosomes; the overexpression of IL-1 occurs at the beginning of the inflammatory process. Over-production of Aβ peptides directed against pathogenic neuroinvasion can cause accumulation of Aβ in plaques; Aβ deposits seem to trigger autophagy. Conclusions: In this study authors observed the binding of the Aβ spherical oligomers to LPS micellar particles inside neuroblastoma cells, the incorporation of LPS to micellar particles occurs at an early stage of Aβ aggregation acting as a nucleation factor, these results are in agreement with the possibility that microorganisms could play a role in the formation of senile plaques in AD