Magic™ Anti-FADD (phospho S194) polyclonal antibody [Alexa Fluor® 555]

While prior studies have established various interacting mechanisms and neural consequences (i.e. monoaminergic nerve terminal damage) that might contribute to the adverse effects caused by methamphetamine administration, the precise mechanisms that mediate relapse during withdrawal remain unknown. This study evaluated the long-term consequences of binge methamphetamine administration (three pulses/day, every 3 hours, 4 days, i.p.; dose-response: 2.5, 5 and 7.5 mg/kg) in adult Sprague-Dawley rats at two behavioral levels following 25 days of withdrawal: (1) negative affect (behavioral despair-forced-swim test, and anhedonia-1% sucrose consumption, two-bottle choice test) and (2) voluntary methamphetamine consumption (20 mg/l, two-bottle choice test). Striatal and hippocampal brain samples were dissected to quantify monoamines content by high-performance liquid chromatography and to evaluate neurotoxicity (dopaminergic and serotonergic markers) and neuroplasticity markers [i.e. cell fate regulator (Fas-associated protein with death domain) FADD] by Western blot. The results showed that methamphetamine administration induced dose-dependent negative effects during prolonged withdrawal in adult rats. In particular, rats treated repeatedly with methamphetamine (7.5 mg/kg) showed (1) enhanced negative affect-increased anhedonia associated with behavioral despair, (2) increased voluntary methamphetamine consumption, (3) enhanced neurotoxicity-decreased dopamine and metabolites in striatum and decreased serotonin in hippocampus, (4) altered neuroplasticity markers-decreased FADD protein and increased p-FADD/FADD balance selectively in hippocampus and (5) higher consumption rates of methamphetamine that were associated with lower FADD content in hippocampus. These results confirm that methamphetamine withdrawal dose-dependently induced negative affect and decreased monoamines content, while also increased voluntary methamphetamine consumption and suggested a role for hippocampal FADD neuroplasticity in these drug-withdrawal adaptations.

Head and neck squamous cell carcinoma (HNSCC) remains one of the most common malignancies worldwide. Although the treatment outcomes of HNSCC have improved in recent years, the prognosis of patients with advanced-stage disease remains poor. Current treatment strategies for HNSCC include surgery as a primary therapy, while radio-, chemo-, and biotherapeutics can be applied as second-line therapy. Although tumor necrosis factor-alpha (TNF-alpha) is a potent tumor suppressor cytokine, the stimulation of opposing signals impairs its clinical utility as an anticancer agent. The aim of this study was to elucidate the mechanisms regulating TNF-alpha-induced opposing signals and their biological consequences in HNSCC cell lines. We determined the molecular mechanisms of TNF-alpha-induced opposing signals in HNSCC cells. Our in vitro analysis indicated that one of these signals triggers apoptosis, while the other induces both apoptosis and cell survival. The TNF-alpha-induced survival of HNSCC cells is mediated by the TNF receptor-associated factor 2 (TRAF2)/nuclear factor (NF)-kappa B-dependent pathway, while TNF-alpha-induced apoptosis is mediated by mitochondrial and non-mitochondrial-dependent mechanisms through FADD-caspase-8-caspase-3 and ASK-JNK-p53-Noxa pathways. The localization of Noxa protein to both the mitochondria and endoplasmic reticulum (ER) was found to cause mitochondrial dysregulation and ER stress, respectively. Using inhibitory experiments, we demonstrated that the FADD-caspase-8-caspase-3 pathway, together with mitochondrial dysregulation and ER stress-dependent pathways, are essential for the modulation of apoptosis, and the NF-kappa B pathway is essential for the modulation of anti-apoptotic effects/cell survival during the exposure of HNSCC cells to TNF-alpha. Our data provide insight into the mechanisms of TNF-alpha-induced opposing signals in HNSCC cells and may further help in the development of novel therapeutic approaches with which to minimize the systemic toxicity of TNF-alpha.