JNK Contributes to Hif-1α Regulation in Hypoxic Neurons

Hypoxia is an established factor of neurodegeneration. Nowadays, attention is directed at understanding how alterations in the expression of stress-related signaling proteins contribute to age dependent neuronal vulnerability to injury. The purpose of this study was to investigate how Hif-1alpha, a major neuroprotective factor, and JNK signaling, a key pathway in neurodegeneration, relate to hypoxic injury in young (6DIV) and adult (12DIV) neurons. We could show that in young neurons as compared to mature ones, the protective factor Hif-1alpha is more induced while the stress protein phospho-JNK displays lower basal levels. Indeed, changes in the expression levels of these proteins correlated with increased vulnerability of adult neurons to hypoxic injury. Furthermore, we describe for the first time that treatment with the D-JNKI1, a JNK-inhibiting peptide, rescues adult hypoxic neurons from death and contributes to Hif-1alpha upregulation, probably via a direct interaction with the Hif-1alpha protei

JNK plays a key role in tau hyperphosphorylation in Alzheimer's disease models

Alzheimer's disease (AD) is a major clinical concern, and the search for new molecules to combat disease progression remains important. One of the major hallmarks in AD pathogenesis is the hyperphosphorylation of tau and subsequent formation of neurofibrillary tangles. Several kinases are involved in this process. Amongst them, c-Jun N-terminal kinases (JNKs) are activated in AD brains and are also associated with the development of amyloid plaques. This study was designed to investigate the contribution of JNK in tau hyperphosphorylation and whether it may represent a potential therapeutic target for the fight against AD. The specific inhibition of JNK by the cell permeable peptide D-JNKI-1 led to a reduction of p-tau at S202/T205 and S422, two established target sites of JNK, in rat neuronal cultures and in human fibroblasts cultures. Similarly, D-JNKI-1 reduced p-tau at S202/T205 in an in vivo model of AD (TgCRND8 mice). Our findings support the fundamental role of JNK in the regulation of tau hyperphosphorylation and subsequently in AD pathogenesis

A targeted use of DNA microarrays in the pharmacogenetics of antidepressants: pros and cons

Microarray or DNA chip technology is a revolutionary approach, which is reshaping the molecular biology and the way to make research and diagnosis. It is a recently developed technique, which allows analyzing thousand of genes, in a very short time. Microarray technology has a high number of fields of application, starting from rapid sequencing of DNA, to identify rare or more frequent gene variants (polymorphisms) related to a particular disease, until gene and protein expression analysis, to study the way, the time and the physiological/pathological conditions, in which a specific gene is translated or not into the final protein. Due to the powerful nature of this genetic approach, the number of researches using microarray technology boosted in the last years (less than ten papers from 1995 to 1997, up to approximately one thousand in the following 5 years -PubMed). Nevertheless, in spite of the promising fields of application, many drawbacks have been described, in the use of DNA microarrays and need to be carefully considered. For example, many errors of incorporation, during the manufacturing of the chips, loss of alternative variants of the studied genes (alternative splicing variants), variable reliability of differential expression data, low specificity of cDNA microarray probes, discrepancy in fold change calculation for a given gene and so on have been reported. In spite of all those troubles, there is no doubt that this new promising technology could give an overall idea of gene organization and expression and might contribute to understanding the molecular mechanisms involved in processes, as disease diagnosis or drug discovery (Pharmacogenetics and Pharmacogenomics). The aim of the present review is to suggest a targeted use of DNA chip technology, in the field of pharmacogenetics (with the example of antidepressants), suggesting the way to select, among thousands of genes, possibly involved in the pharmacological action of antidepressants, only those ones more probably candidate, trough a step by step analysis of intracellular pathways

Complete analysis of the mwsenchymal stem cells secretome using A label-free approach

Mesenchymal stem cell (MSCs) have recently been shown to have a pivotal role in tissue repair and in local control of inflammation. Therefore, in the last few years there has been a growing interest in using them to treat human inflammatory diseases, including severe (grade III-IV) steroid resistant acute graft versus host disease (aGVHD). In vitro studies indicated that MSCs display immunomodulatory activities: they inhibit T and NK cell activation, B-cell terminal differentiation and dendritic cell maturation and functions. Although there is evidence from in vivo studies that MSCs are able to reduce inflammatory damage, it is not clear whether their immunomodulatory effects rely on soluble factors or cell-cell contacts. In a recent study, by comparing the immunomodulatory potential of mouse MSC either injected intravenously, or subcutaneously entrapped through the use of microcapsules, it has been reported that the encapsulated MSCs (E-MSCs) are extremely efficient in down-modulating in vivo immune responses, such as antigen-specific T cell responses and acute GVHD, and that this activity is based on soluble factors released by MSC into the blood stream [1]. Based on these evidences, in collaboration with Humanitas Clinical and Research Center, we compared the secretome of MSC, isolated from murine bone marrow cells, before & after treatment with pro-inflammatory cytokines (TNFs, IL1b, and IL6). To achieve this goal, we employed a label-free quantitative approach that allows us to examine differences in global protein expression between different samples. Samples have been derivatized, digested and analyzed by LC-nanoESI-LTQ Orbitrap Velos equipped with a RPC18 column for peptides separation prior to MS/MS analysis. For protein identification and quantification, raw data files were processed and analyzed using MaxQuant 1.3.0.5

JNK regulates APP cleavage and degradation in a model of Alzheimer's disease

Secretion of Amyloid-beta peptide (A\uce\ub2) circulating oligomers and their aggregate forms derived by processing of beta-amyloid precursor protein (APP) are a key event in Alzheimer's disease (AD). We show that phosphorylation of APP on threonine 668 may play a role in APP metabolism in H4-APPswcell line, a degenerative AD model. We proved that JNK plays a fundamental role in this phosphorylation since its specific inhibition, with the JNK inhibitor peptide (D-JNKI1), induced APP degradation and prevented APP phosphorylation at T668. This results in a significant drop of \uce\ub2APPs, A\uce\ub2 fragments and A\uce\ub2 circulating oligomers. Moreover the D-JNKI1 treatment produced a switch in the APP metabolism, since the peptide reduced the rate of the amyloidogenic processing in favour of the non-amyloidogenic one. All together our results suggest an important link between APP metabolism and the JNK pathway and contribute to shed light on the molecular signalling pathway of this disease indicating JNK as an innovative target for AD therapy