Inhibitory roles of signal transducer and activator of transcription 3 in antitumor immunity during carcinogen-induced lung tumorigenesis.

Stat3 mediates a complex spectrum of cellular responses, including inflammation, cell proliferation, and apoptosis. Although evidence exists in support of a positive role for Stat3 in cancer, its role has remained somewhat controversial because of insufficient study of how its genetic deletion may affect carcinogenesis in various tissues. In this study, we show using epithelium-specific knockout mice (Stat3(Δ/Δ)) that Stat3 blunts rather than supports antitumor immunity in carcinogen-induced lung tumorigenesis. Although Stat3(Δ/Δ) mice did not show any lung defects in terms of proliferation, apoptosis, or angiogenesis, they exhibited reduced urethane-induced tumorigenesis and increased antitumor inflammation and natural killer (NK) cell immunity. Comparative microarray analysis revealed an increase in Stat3(Δ/Δ) tumors in proinflammatory chemokine production and a decrease in MHC class I antigen expression associated with NK cell recognition. Consistent with these findings, human non-small cell lung cancer (NSCLC) cells in which Stat3 was silenced displayed an enhancement of proinflammatory chemokine production, reduced expression of MHC class I antigen, and increased susceptibility to NK cell-mediated cytotoxicity. In addition, supernatants from Stat3-silenced NSCLC cells promoted monocyte migration. Collectively, our findings argue that Stat3 exerts an inhibitory effect on antitumor NK cell immunity in the setting of carcinogen-induced tumorigenesis.

Antioxidant effects of 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran and alpha-tocopherol in hyperlipidemic mice as evaluated by hydroxyoctadecadienoic acid and 7-hydroxycholesterol.

It has been hypothesized that oxidative modification of low density lipoprotein plays a key role in the pathogenesis of atherosclerosis. In order to elucidate the role of lipid oxidation and its inhibition in vivo, apolipoprotein E and alpha-tocopherol (alphaT) transfer protein double knockout (ApoE(-/-)alpha-TTP(-/-)) and apolipoprotein E knockout (ApoE(-/-)) mice fed with a vitamin E-depleted diet and a diet containing 0.002 wt.% alphaT, respectively, were used with or without the treatment of a synthetic antioxidant 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran (BO-653, 0.2 wt.%). The lipid oxidation markers of total hydroxylinoleic acid (tHODE), 8-iso-prostaglandin F(2alpha), and 7-hydroxycholesterol (t7-OHCh) in the blood, liver, and brain were inclusively measured with or without an excessive cholesterol-feeding (Ch-diet). The tHODE levels were elevated by Ch-diet in the plasma and brain of ApoE(-/-)alpha-TTP(-/-) mice and in the liver of ApoE(-/-) mice without BO-653. The levels of t7-OHCh in the liver were also increased due to the Ch-diet, and the ratio of t7-OHCh to the parent compound cholesterol was reduced to the control levels by BO-653. In summary, it was demonstrated by biomarkers, tHODE and t7-OHCh, that the added BO-653 in their diets exerted antioxidative effects in vivo under the condition of reduced vitamin E.

Development of vaccines and passive immunotherapy against SARS corona virus using SCID-PBL/hu mouse models.

We have investigated novel vaccine strategies against severe acute respiratory syndrome (SARS) CoV using cDNA constructs encoding the structural antigens: (S), (M), (E), or (N) protein, derived from SARS CoV. PBL from healthy human volunteers were administered i.p. into IL-2 receptor gamma-chain disrupted SCID mice, and SCID-PBL/hu mice were constructed. These mice can be used to analyze the human immune response in vivo. SARS M DNA vaccine and N DNA vaccine induced human CTL specific for SARS CoV antigens. Alternatively, SARS M DNA vaccines inducing human neutralizing antibodies and human monoclonal antibodies against SARS CoV are now being developed. These results show that these vaccines can induce virus-specific immune responses and should provide a useful tool for development of protective and therapeutic vaccines.

Evaluation of a novel vaccine (HVJ-liposome/HSP65 DNA+IL-12 DNA) against tuberculosis using the cynomolgus monkey model of TB.

We have developed a novel tuberculosis (TB) vaccine; a combination of the DNA vaccines expressing mycobacterial heat shock protein 65 (HSP65) and interleukin 12 (IL-12) delivered by the hemagglutinating virus of Japan (HVJ)-liposome (HSP65+IL-12/HVJ). This vaccine provided remarkable protective efficacy in mouse and guinea pig models compared to the BCG vaccine, on the basis of an induction of the CTL activity and improvement of the histopathological tuberculosis lesions, respectively. Furthermore, we extended our studies to a cynomolgus monkey model, which is currently the best animal model of human tuberculosis. This novel vaccine provided a higher level of the protective efficacy than BCG based upon the assessment of mortality, the ESR, body weight, chest X-ray findings and immune responses. Furthermore, the combination of HSP65+IL-12/HVJ and BCG by the priming-booster method showed a synergistic effect in the TB-infected cynomolgus monkey (100% survival). These data indicate that our novel DNA vaccine might be useful against Mycobacterium tuberculosis for human clinical trials.

Lipid peroxidation in mice fed a choline-deficient diet as evaluated by total hydroxyoctadecadienoic acid.

OBJECTIVE: The relevance of oxidative stress in mice fed a choline-deficient diet (CDD) was investigated in relation to the oxidative stress marker, hydroxyoctadecadienoic acid (HODE) in comparison with F2-isoprostanes. Further, the protective effects of antioxidants against oxidative damage were assessed by using HODE. METHODS: We recently proposed total HODE as a biomarker for oxidative stress in vivo. Biological samples such as plasma, urine, and tissues were first reduced and then saponified to convert various oxidation products of linoleates to HODE. In the present study, this method was applied to measure oxidative damage in mice induced by CDD for 1 mo. RESULTS: CDD, when compared with choline-controlled diet (CCD), increased liver weight and fatty acid accumulation but the increase in body weight was less significant. Remarkable increases in HODE and 8-iso-prostaglandin F(2alpha) in liver and plasma were observed when mice were fed with the CDD for 1 mo compared with the CCD. The HODE level was about two to three orders higher than the F2-isoprostane level. This increase was decreased to the level of the CCD when alpha-tocopherol or 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran, a potent synthetic antioxidant, was mixed with the CDD. The stereoisomer ratio of HODE (9-and-13 (Z,E)-HODE/9-and-13 (E,E)-HODE) was decreased by CDD compared with CCD, which was spared by the addition of alpha-tocopherol and 2,3-dihydro-5-hydroxy-4,6-di-tert-butyl-2,2-dipentylbenzofuran. However, the increase in plasma glutamic-pyruvic transaminase and fatty acids in liver induced by the CDD was not recovered by any antioxidant. CONCLUSIONS: This study clearly demonstrated that oxidative stress was involved in fatty liver formation induced by the CDD and that HODE was a good biomarker for an oxidative stress in vivo.