ABSTRACT
Aberrant transglutaminase 2 (TG2) expression and protein cross-linking activity have been associated with several chronic neurodegenerative disorders in which inflammatory processes triggered by activated microglia and monocytes play a key role, such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and multiple sclerosis. Interestingly, mild-to-moderate hyperhomocysteinemia (HHcy), corresponding to increased plasma homocysteine (Hcy) concentrations in the range 16-60 µM, have recently been associated with the above-cited diseases. Using THP-1 monocytes, here we investigated the role of TG2 in cell response to mildly elevated Hcy concentrations. A five-day incubation with Hcy (â¼25 µM) increased reactive oxygen species, peroxide lipids, as well as 8-hydroxyguanosine levels by twofold, and decreased the endogenous cell antioxidant defenses, that is reduced glutathione, by 50% in Hcy-exposed cultures compared with controls (p < 0.01). Hcy-induced oxidative stress was associated with increases in TG2 expression and activity, as well as nuclear factor kappa B activation. Notably, the latter was reduced in the presence of the TG-specific inhibitor R283. Hcy exposure also significantly increased the mRNA levels of tumor necrosis factor alpha, interleukin (IL)-6, and IL-1ß, as well as the level of Hcy-inducible endoplasmic reticulum (ER) stress protein, a marker of ER stress, in Hcy-exposed cultures compared with controls. Notably, these effects were dramatically reduced by R283. These preliminary findings indicate that TG2 plays a key role in Hcy-induced activation of THP-1 monocytes, involving oxidative as well as ER stress and inflammation. This underlines the potential of TG2 inhibition in the therapeutic management of inflammatory processes contributing to neurodegenerative disorders associated with mild HHcy.
Subject(s)
GTP-Binding Proteins/metabolism , Homocysteine/pharmacology , Monocytes/enzymology , Transglutaminases/metabolism , Antioxidants/metabolism , Cell Line , Cytokines/genetics , Endoplasmic Reticulum Stress , GTP-Binding Proteins/genetics , Humans , Monocytes/immunology , NF-kappa B/metabolism , Oxidative Stress , Protein Glutamine gamma Glutamyltransferase 2 , Transglutaminases/genetics , Up-RegulationABSTRACT
Neuroblastoma is the most common extracranial pediatric solid tumor with poor prognosis in children with disseminated stage of disease. A number of studies show that molecules largely distributed in commonly consumed fruits and vegetables may have anti-tumor activity. In this study we evaluate the effect of Citrus bergamia (bergamot) juice (BJ) in vitro and in a spontaneous metastatic neuroblastoma SCID mouse model. Qualitative and quantitative characterizations of BJ flavonoid fractions were performed by RP-HPLC/PDA/MS. We show that BJ significantly affects SK-N-SH and LAN-1 cell proliferation in vitro, but fails to reduce primary tumor weight in vivo. Moreover, BJ reduced cell adhesiveness and invasion of LAN-1 and SK-N-SH cells in vitro and the number of pulmonary metastases under consideration of the number of tumor cells in the blood in mice inoculated with LAN-1 cells in vivo. These effects without any apparent sign of systemic toxicity confirm the potential clinical interest of BJ and lay the basis for further investigation in cancer.
Subject(s)
Beverages , Citrus/chemistry , Flavonoids/therapeutic use , Lung Neoplasms/drug therapy , Neuroblastoma/drug therapy , Animals , Cell Adhesion/drug effects , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Cell Survival/drug effects , Disease Models, Animal , Female , Flavonoids/chemistry , Flavonoids/isolation & purification , Fruit/chemistry , Heterografts , Humans , Male , Mice , Mice, SCID , Neoplasm Invasiveness , Neoplasm MetastasisABSTRACT
High levels of homocysteine promote cell damage mainly through induction of oxidative stress, endoplasmic reticulum (ER) stress, and activation of pro-inflammatory factors. The effects of homocysteine were here examined in the continuously dividing neuroblastoma cell line Neuro2a. Cell treatment with homocysteine (100-500 microM) for 4 h increased ROS production while reducing cell viability in a dose-dependent manner. Cell exposure to 250 microM homocysteine was able to induce transglutaminase 2 up-regulation and increased in situ transglutaminase activity. These effects were prevented by the incubation with the transglutaminase activity inhibitor cystamine. Homocysteine also induced NF-kappaB activation that seemed associated with transglutaminase 2 up-regulation since the specific NF-kappaB inhibition by SN50 was able to reduce transglutaminase expression and activity levels. In the light of these observations, it may be postulated that TG2 up-regulation is involved in cell response to homocysteine-induced stress, in which NF-kappaB activation plays also a pivotal role.
