Exosomes are secreted vesicles formed in late endocytic compartments. Immature dendritic cells (DCs) secrete exosomes, which transfer functional major histocompatibility complex (MHC)–peptide complexes to other DCs. Since immature and mature DCs induce different functional T-cell responses (ie, tolerance versus priming), we asked whether DC maturation also influenced the priming abilities of their exosomes. We show that exosomes secreted by lipopolysaccharide (LPS)–treated mature DCs are 50- to 100-fold more potent to induce antigen-specific T-cell activation in vitro than exosomes from immature DCs. In vitro, exosomes from mature DCs transfer to B lymphocytes the ability to prime naive T cells. In vivo, only mature exosomes trigger effector T-cell responses, leading to fast skin graft rejection. Proteomic and biochemical analyses revealed that mature exosomes are enriched in MHC class II, B7.2, intercellular adhesion molecule 1 (ICAM-1), and bear little milk-fat globule–epidermal growth factor–factor VIII (MFG-E8) as compared with immature exosomes. Functional analysis using DC-derived exosomes from knock-out mice showed that MHC class II and ICAM-1 are required for mature exosomes to prime naive T cells, whereas B7.2 and MFG-E8 are dispensable. Therefore, changes in protein composition and priming abilities of exosomes reflect the maturation signals received by DCs.

Paradoxically, reactive oxygen species (ROS) can promote normal cellular proliferation and carcinogenesis, and can also induce apoptosis of tumor cells. In this report, we study the contribution of ROS to various cellular signals depending on the nature and the level of ROS produced. In nontransformed NIH 3T3 cells, ROS are at low levels and originate from NADPH oxidase. Hydrogen peroxide (H2O2), controlled by the glutathione system, is pivotal for the modulation of normal cell proliferation. In CT26 (colon) and Hepa 1-6 (liver) tumor cells, high levels of ROS, close to the threshold of cytotoxicity, are produced by mitochondria and H2O2 is controlled by catalase. N-acetylcysteine, which decreases H2O2 levels, inhibits mitogen-activated protein kinase and normal cell proliferation but increases tumor cell proliferation as H2O2 concentration drops from the toxicity threshold. In contrast, antioxidant molecules, such as mimics of superoxide dismutase (SOD), increase H2O2 levels through superoxide anion dismutation, as well as in vitro proliferation of normal cells, but kill tumor cells. CT26 tumors were implanted in mice and treated by oxaliplatin in association with one of the three SOD mimics manganese(III)tetrakis(4-benzoic acid) porphyrin, copper(II)(3,5-diisopropylsalicylate)2, or manganese dipyridoxyl diphosphate. After 1 month, the volumes of tumors were respectively 35%, 31%, and 63% smaller than with oxaliplatin alone (P < 0.001). Similar data were gained with Hepa 1-6 tumors. In conclusion, antioxidant molecules may have opposite effects on tumor growth. SOD mimics can act in synergy with cytotoxic drugs to treat colon and liver cancers.

Intracellular events following paclitaxel binding to microtubules that lead to cell death remain poorly understood. Because reactive oxygen species (ROS) are involved in the cytotoxicity of anticancer agents acting through independent molecular targets, we explored the role of ROS in paclitaxel cytotoxicity. Within 15 min after in vitro exposure of A549 human lung cancer cells to paclitaxel, a concentration‐dependent intracellular increase in O°₂⁻ and H₂O₂ levels was detected by spectrofluorometry. Addition of N‐acetylcysteine (NAC) or glutathione, two H₂O₂ scavenger, induced a 4‐fold increase in paclitaxel IC₅₀. Delaying NAC co‐incubation by 4 hr, resulted in a 3‐fold reduction in cell protection. The glutathione synthesis inhibitor, buthionine sulfoximine significantly increased paclitaxel cytotoxicity and H₂O₂ accumulation, but did not modify O°₂⁻ levels. Co‐incubation with diphenylene iodonium suggested that paclitaxel induced‐O°₂⁻ production was in part associated with increased activity of cytoplasmic NADPH oxidase. Concomitant treatment with inhibitors of caspases 3 and 8 increased cell survival but did not prevent the early accumulation of H₂O_2. To evaluate the role of ROS in paclitaxel antitumoral activity, mice were injected with LLC1 lung cancer cells and treated with paclitaxel i.p. and/or NAC. The antitumoral activity of paclitaxel in mice was abolished by NAC. In conclusion, the accumulation of H₂O₂ is an early and crucial step for paclitaxel‐induced cancer cell death before the commitment of the cells into apoptosis. These results suggest that ROS participate in vitro and in vivo to paclitaxel cytotoxicity.

Collagen-induced arthritis (CIA), an animal model for rheumatoid arthritis, is induced in DBA/1 (H-2q) mice following immunization with type II collagen (CII) in CFA. Since we have previously shown that IFN-gamma exerts a biphasic effect during the evolution of CIA in DBA/1 mice, we analyzed the development of this disease in mice with a disruption of the IFN-gamma receptor gene (IFN-gammaR(0/0)). Mutant mice were interbred with the DBA/1 strain to yield IFN-gammaR(0/0) mice expressing the H-2q haplotype. In three consecutive experiments, IFN-gammaR(0/0) male mice were found to exhibit severe clinical and histologic arthritis with an average incidence of 88.5 vs 94.1% for the wild DBA/1 strain. Notably, onset of clinical symptoms occurred significantly earlier than in DBA/1 mice. Although of a lower magnitude than in males, CIA also developed early in IFN-gammaR(0/0) female mice and with higher clinical severity than in control DBA/1 females. Immunization of knockout mice with CII resulted in the generation of CII-specific T cells belonging to the Th1 phenotype that recognize the same immunodominant peptides as do DBA/1 mice. CIA in IFN-gammaR(0/0) mice was associated with a down-regulation of the CII-specific IgG response, and this impairment was essentially due to a strong reduction of Abs of the IgG2a isotype. Taken together, our findings provide evidence that IFN-gammaR deficiency in DBA/1 mice leads to the occurrence of severe CIA with an accelerated onset compared with that in wild-type mice, indicating that the proinflammatory action of IFN-gamma has been bypassed in the IFN-gammaR(0/0) mice.