Cytokine and chemokine expression in cigarette smoke-induced lung injury in guinea pigs.

The guinea pig model of cigarette smoke (CS)-induced lung injury is known to exhibit many pathophysiological similarities to chronic obstructive pulmonary disease (COPD), but the expression profiles of inflammatory mediators in the lung are poorly understood. Quantitative real-time RT-PCR was used in this study to investigate the pulmonary expression profiles of cytokine and chemokine mRNA in response to single or repeated CS exposure in guinea pigs. A single CS exposure did not induce obvious inflammatory cell infiltration into the lungs, but it led to significant increases in the mRNA expression of tumour necrosis factor-alpha, interleukin (IL)-1beta, IL-8, and monocyte chemoattractant protein (MCP)-1, and decreases in IL-5 and granulocyte-macrophage colony-stimulating factor. Repeated CS exposure induced many features of COPD, such as marked accumulation of macrophages and neutrophils, augmented protease activities, lung structural alterations and increased airway resistance, accompanied by significant increases in the mRNA expression of IL-1beta and MCP-1 and decreases in IL-2, IL-5, transforming growth factor-beta, and eotaxin. In conclusion, in guinea pigs, inflammatory mediator changes in the lungs following cigarette smoke exposure are largely similar to those reported for smokers and/or chronic obstructive pulmonary disease patients. This model will therefore be useful to further understand the pathogenesis of chronic obstructive pulmonary disease.

Dendritic cells transduced with gp100 gene by RGD fiber-mutant adenovirus vectors are highly efficacious in generating anti-B16BL6 melanoma immunity in mice.

Dendritic cells (DCs) are the most potent professional antigen-presenting cells for the initiation of antigen-specific immune responses, and antigen-loaded DCs have been regarded as promising vaccines in cancer immunotherapy. We previously demonstrated that RGD fiber-mutant adenovirus vector (AdRGD) could attain highly efficient gene transduction into human and murine DCs. The aim of the present study is to demonstrate the predominance of ex vivo genetic DC manipulation using AdRGD in improving the efficacy of DC-based immunotherapy targeting gp100, a melanoma-associated antigen (MAA). Vaccination with murine bone marrow-derived DCs transduced with AdRGD encoding gp100 (AdRGD-gp100/mBM-DCs) dramatically improved resistance to B16BL6 melanoma challenge and pulmonary metastasis as compared with immunization with conventional Ad-gp100-transduced mBM-DCs. The improvement in antimelanoma effects upon immunization with AdRGD-gp100/mBM-DCs correlated with enhanced cytotoxic activities of natural killer (NK) cells and B16BL6-specific cytotoxic T lymphocytes (CTLs). Furthermore, in vivo depletion analysis demonstrated that CD8(+) CTLs and NK cells were the predominant effector cells responsible for the anti-B16BL6 immunity induced by vaccination with AdRGD-gp100/mBM-DCs, and that helper function of CD4(+) T cells was necessary for sufficiently eliciting effector activity. These findings clearly revealed that highly efficient MAA gene transduction to DCs by AdRGD could greatly improve the efficacy of DC-based immunotherapy against melanoma.

Efficient antigen gene transduction using Arg-Gly-Asp fiber-mutant adenovirus vectors can potentiate antitumor vaccine efficacy and maturation of murine dendritic cells.

Dendritic cells (DCs), the most effective antigen-presenting cells, are being studied as adjuvants or antigen delivery vehicles for eliciting T-cell-mediated antitumor immunity. Gene delivery to DCs provides an intracellular source of antigen for efficient and persistent loading to MHC class I molecules capable of activating CD8(+) CTLs, which play a central role in antitumor immunity. We previously reported that the fiber-mutant adenovirus vector (Ad) harboring the Arg-Gly-Asp (RGD) sequence in the HI loop of its fiber knob could more efficiently transduce the LacZ gene into both murine DC lines and normal human DCs than conventional Ad. In the present study, we compared immunological properties and vaccine efficacy of DC2.4 cells, an immature murine DC line, transduced with an ovalbumin (OVA) gene by fiber-mutant Ad (Ad-RGD-OVA) or conventional Ad (Ad-OVA). Ad-RGD-OVA-infected DC2.4 cells could more efficiently present OVA peptides via MHC class I molecules in a vector particle-dependent manner and induce OVA-specific CTL response by vaccination than Ad-OVA-infected DC2.4 cells. This result was correlated with the efficiency of gene transduction into DC2.4 cells by both types of Ad. Moreover, vaccination with Ad-RGD-OVA-infected DC2.4 cells could achieve an equal or greater antitumor effect against challenge with E.G7-OVA tumor cells with lower doses of Ad on infection or fewer cells for immunization than the vaccination procedure using Ad-OVA-infected DC2.4 cells. In addition, the maturation of DC2.4 cells was promoted by efficient expression of the antigen gene by the Arg-Gly-Asp fiber-mutant Ad. Flow cytometric analysis indicated enhanced expression of MHC class I and II molecules as well as CD80, CD86, CD40, and CD54, and reverse transcription-PCR analysis revealed increased levels of interleukin 12 p40 mRNA. However, infection by Ad-OVA or Ad that did not contain the cDNA of interest (Ad-Null and Ad-RGD-Null) contributed little to phenotypical changes in DC2.4 cells. On the basis of these results, we propose that DC manipulation using the Arg-Gly-Asp fiber-mutant Ad system could advance the development of more effective vaccines and allow for more convenient administration of DC-based gene immunotherapy.

Effects of lipofectin-antigen complexes on major histocompatibility complex class I-restricted antigen presentation pathway in murine dendritic cells and on dendritic cell maturation.

We previously reported that exogenous antigens complexed with the cationic liposome lipofectin (LF) were efficiently presented via major histocompatibility complex (MHC) class I molecules on pulsed dendritic cells (DCs) in vitro. In the present study, we demonstrated that MHC class I-restricted antigen presentation on DC2.4 cells, a murine immature DC line, treated with LF-antigen complexes was remarkably suppressed through the inhibition of endocytosis, proteasome catalysis, and Golgi transport. We also found that LF did not influence expression of interleukin-12 p40 mRNA, MHC molecules, or co-stimulatory molecules in DC2.4 cells. These findings suggest that an antigen-loading procedure using LF could enhance delivery of exogenous antigens to the classical MHC class I pathway in DCs, but it does not initiate DC maturation.

Effect of the angiotensin converting enzyme inhibitor, captopril, on proteinuria in chronic glomerular disease.

The antiproteinuric effect of the angiotensin-I-converting enzyme inhibitor, captopril, was studied in 14 patients (10 men and 4 women, age range of 24 to 60 years) with chronic glomerulonephritis in whom IgA nephritis had been confirmed by renal biopsy. Eight of the 14 patients had received antihypertensive drugs such as calcium channel blockers, diuretics or beta-blockers. Captopril was added to these regimens at 25 mg twice daily in 3 patients, and 37.5 mg in 11 patients. Proteinuria decreased from 2.55 +/- 0.48 g/day to 1.58 +/- 0.35 g/day within three months after the start of administration. In 4 patients (28.6%), the extent of reduction was over 50%, and in 8 patients (57.1%), over 25%. Blood pressure, creatinine clearance and serum creatinine were not changed significantly. There was a positive linear correlation between the extent of reduction of proteinuria and the increase in plasma renin activity (r = 0.93, p < 0.001). We conclude that captopril reduces proteinuria in some patients with IgA nephritis whose plasma renin activity responds to the drug.

[Significance of the bioclean room during the treatment of lung cancer].

The effectiveness of a vertical air flow room using HEPA filter (class 5000) was studied in relation to the prevention of infection during induction chemotherapy for primary lung cancer and metastatic pulmonary tumors. From November 1982 to July 1983, 12 patients (11 primary lung cancer: 9 small cell carcinoma, 2 non-small cell carcinoma and 1 metastatic pulmonary tumor) were treated in the bioclean room, while 14 patients (1 small cell carcinoma, 13 non-small cell carcinoma) were treated in conventional room during this period as control. Partial response was observed in 6 patients with small cell carcinoma treated in the bioclean room and 4 patients with non-small cell carcinoma treated in the conventional room. In patients treated in the bioclean room, the median day of stay in the room was 21 ranged 9 to 43. The leukocyte counts in the peripheral blood less than 1500/mm3 and 1000/mm3 were observed in 6 and 4 cases, respectively. Decrease in granulocyte counts less than 500/mm3 was observed in 6 cases. Only one patient with leukocyte count of 800 (granulocyte count 104) experienced fever higher than 38 C lasting 8 days. On the other hand, in 14 patients treated in the conventional room, leukocytopenia less than 1500/mm3 and 1000/mm3 was observed in 7 and 0 cases, respectively. Granulocytopenia less than 500/mm3 was observed in 8 cases, and 7 patients experienced fever higher than 38 C. Number of non-viable particles in the bioclean room ranged from 2.9 to 6.8 X 10(2)/ft3. Organisms isolated from the bioclean room and from the skin of patients were normal flora, and the numbers of colonies detected from the skin of patients decreased 1-2 weeks after the beginning of treatment in the bioclean room. The usefulness of a vertical air flow room during chemotherapy of lung cancer, especially of small cell carcinoma, was suggested.