Invariant NKT cells increase drug-induced osteosarcoma cell death.

BACKGROUND AND PURPOSE: In osteosarcoma (OS) patients, only a limited number of drugs are active and the regimens currently in use include a combination of at least two of these drugs: doxorubicin, cisplatin, methotrexate and ifosfamide. Today, 30-40% of patients still die of OS highlighting the urgent need for new treatments. Invariant NKT (iNKT) cells are a lymphocyte lineage with features of both T and NK cells, playing important roles in tumour suppression. Our aim was to test whether the cytoxicity induced by cisplatin, doxorubicin and methotrexate against OS cells can be enhanced by iNKT cell treatment. EXPERIMENTAL APPROACH: iNKT cells were purified from human peripheral blood mononuclear cells by cell sorting (Vα24Vß11(+) cells) and used as effector cells against OS cells (U2-OS, HOS, MG-63). Cell death (calcein-AM method), perforin/granzyme B and Fas/FasL expressions were determined by flow cytometry. CD1d expression was analysed at both the gene and protein level. KEY RESULTS: iNKT cells were cytotoxic against OS cells through a CD1d-dependent mechanism. This activity was specific for tumour cells, because human CD1d(+) mesenchymal stem cells and CD1d(-) osteoblasts were not affected. iNKT cell treatment enhanced drug-induced OS cell death in a concentration-dependent manner and this effect was reduced in CD1d-silenced OS cells. CONCLUSION AND IMPLICATIONS: iNKT cells kill malignant, but not non-malignant, cells. iNKT cell treatment enhances the cytotoxicity of anti-neoplastic drugs against OS cells in a CD1d-dependent manner. The present data encourage further studies on the use of iNKT cells in OS therapy.

Clovamide and rosmarinic acid induce neuroprotective effects in in vitro models of neuronal death.

BACKGROUND AND PURPOSE: Phenolic compounds exert cytoprotective effects; our purpose was to investigate whether the isosteric polyphenolic compounds clovamide and rosmarinic acid are neuroprotective. EXPERIMENTAL APPROACH: Three in vitro models of neuronal death were selected: (i) differentiated SH-SY5Y human neuroblastoma cells exposed to tert-butylhydroperoxide (t-BOOH), for oxidative stress; (ii) differentiated SK-N-BE(2) human neuroblastoma cells treated with L-glutamate, for excitotoxicity; and (iii) differentiated SH-SY5Y human neuroblastoma cells exposed to oxygen-glucose deprivation/reoxygenation, for ischaemia-reperfusion. Cell death was evaluated by lactate dehydrogenase measurements in the cell media, while the mechanisms underlying the effects by measuring: (i) t-BOOH-induced glutathione depletion and increase in lipoperoxidation; and (ii) L-glutamate-induced intracellular Ca(2+) overload (fura-2 method) and inducible gene expression (c-fos, c-jun), by reverse transcriptase-PCR. The ability of compounds to modulate nuclear factor-kappaB and peroxisome proliferator-activated receptor-gamma activation was evaluated by Western blot in SH-SY5Y cells not exposed to harmful stimuli. KEY RESULTS: Both clovamide and rosmarinic acid (10-100 micromol x L(-1)) significantly protected neurons against insults with similar potencies and efficacies. The EC(50) values were in the low micromolar range (0.9-3.7 micromol x L(-1)), while the maximal effects ranged from 40% to -60% protection from cell death over untreated control at 100 micromol x L(-1). These effects are mediated by the prevention of oxidative stress, intracellular Ca(2+) overload and c-fos expression. In addition, rosmarinic acids inhibited nuclear factor-kappaB translocation and increased peroxisome proliferator-activated receptor-gamma expression in SH-SY5Y cells not exposed to harmful stimuli. CONCLUSION AND IMPLICATIONS: Clovamide and rosmarinic acid are neuroprotective compounds of potential use at the nutritional/pharmaceutical interface.