Evidence for the existence of cytochrome b558 in fish neutrophils by polyclonal anti-peptide antibody.

When western blotting analysis of Japanese eel neutrophil lysate was performed using antibody against the synthetic peptide corresponding to the carboxyl terminal region of human cytochrome b558 large subunit, a broad band was specifically detected at approximately 90 kDa. The antibody recognized the epitope present in eel neutrophils only after the cells were permeabilized with detergent. These results indicate that the large subunit of cytochrome b exists in fish neutrophils and the epitope is exposed to the cytoplasmic side of membrane, and suggest that cytochrome b is conserved across species.

Kinetics of oxygen metabolism during respiratory burst in Japanese eel neutrophils.

Two oxygen radicals, superoxide and hydrogen peroxide, were measured by ferricytochrome C reduction and scopoletin fluorescent assays to investigate oxygen metabolism during the respiratory burst in Japanese eel neutrophils. Maximal superoxide production was obtained using 0.1 microgram/mL of PMA. Oxygen consumption was almost equivalent to superoxide production, indicating that the consumed oxygen was almost fully converted to superoxide in eel neutrophils. Hydrogen peroxide production was approximately half that of oxygen consumption or superoxide production in neutrophils elicited by killed bacteria or casein. The amounts of superoxide and hydrogen peroxide production, by neutrophils elicited by irritants, were about 11.5 and 5.0 nM/10(7) cells/ min, respectively, which were significantly higher than those of unelicited neutrophils (2.4 and 0.3 nM/10(7) cells/min, respectively).

Human alveolar macrophages of anergic patients with lung cancer lack the responsiveness to recombinant interferon gamma.

We studied the effect of recombinant interferon gamma (rIFN-gamma) on the phagocytic and bactericidal actions in human alveolar macrophages (AM) of patients with pulmonary tuberculosis and lung cancer. Treatment with 100 or 1,000 U/ml of rIFN-gamma for 24 hours resulted in an increased percentage of AM ingesting bacillus Calmette-Guérin (BCG) and an increased number of ingested BCG in individual AM in tuberculin-positive patients with lung cancer and pulmonary tuberculosis. The rIFN-gamma treatment also showed increased killing activity of AM in tuberculin-positive patients. However, rIFN-gamma treatment of AM in tuberculin-negative anergic patients with lung cancer did not induce the increase of phagocytic and killing activities against BCG. These results suggest that responsiveness and activation of AM to rIFN-gamma are inhibited in the anergic environment.

Establishment and characterization of resistant cells to etoposide (VP16) from a mouse breast cancer cell line, FM3A.

We established 4 cell lines resistant to VP16 from a mouse breast cancer cell line, FM3A. The IC50 values of all 4 resistant strains were approximately 2 micrograms/ml as measured by colony formation in soft agar; about 40 times higher than that of parent cell (0.05 microgram/ml). These cells showed a cross-resistance to VM26, a compound related to VP16, but not to a variety of other antitumor drugs including adriamycin, mitomycin C, cis-platinum, 5-fluorouracil, bleomycin, vincristine, 4-hydroperoxycyclophosphamide, methotrexate and cytosine arabinoside. Topoisomerase II, the putative target of VP16, was partially purified from cells, and was assayed using knotted P4 phage DNA as a substrate. However, no significant difference was observed between enzymes from resistant cells and from the parent cells in either activity per cell or sensitivity to VP16. On the other hand, the resistance of these cell lines to VP16 was greatly reduced by adding a calcium antagonist, verapamil, to the soft agar at a concentration as low as 5 microM, at which the viability of cells was hardly affected. A similar verapamil-induced reduction in the resistance of the cells to VM26 was also observed. These results suggest that the acquired resistance may be largely due to an altered membrane permeability to drugs, which may be overcome by verapamil, rather than to an altered topoisomerase II.

[Adriamycin, cyclophosphamide, vincristine and methotrexate combination chemotherapy of small cell lung cancer].

Twenty-two patients who had not received previous chemotherapy for small cell lung cancer were treated with a combination of adriamycin (30 mg/m2, i.v., on day 1), cyclophosphamide (500 mg/m2, i.v., on day 1, and 350 mg/m2, i.v., on day 8), vincristine (1 mg/m2, i.v., on day 1) and methotrexate (20 mg/m2, i.v., on days 1 and 8). This chemotherapy regimen was repeated at 3- or 4-week intervals. Among 20 evaluable patients, none showed complete response but 14 (70%) showed a partial response, with a median response duration of 8 weeks (range, 4-20 weeks). The median survival time for the 20 evaluable patients was 28 weeks. Toxicity included mild to moderate hematologic toxicity, alopecia and nausea and vomiting. This combination chemotherapy appears to be suboptimal for the treatment of small cell lung cancer, with no complete responses and a relatively short median survival time.

Diagnostic value of cancer antigen 15-3 (CA15-3) detected by monoclonal antibodies (115D8 and DF3) in exudative pleural effusions.

Pleural fluid levels of the cancer antigen 15-3 (CA15-3) detected by monoclonal antibodies (115D8 and DF3) were determined in 40 patients with carcinomatous pleural effusions and in 41 patients with tuberculous pleural effusions. Using a cut off level of 16 U/ml, 15 of the 40 carcinomatous fluids but none of the 41 tuberculous fluids were positive. Pleural fluid levels of CA15-3 were not correlated with those of carcinoembryonic antigen (CEA) or carbohydrate antigen 19-9 (CA19-9). Combined assay of CEA and CA15-3, or CA19-9 and CA15-3, increased the positive rate from 79 to 82% and from 67 to 73%, respectively. Measurement of pleural fluid CA15-3 levels are less useful in separating carcinomatous from tuberculous effusions than is measurement of CEA or CA19-9.

Occurrence of ditaurobilirubin, bilirubin conjugated with two moles of taurine, in the gallbladder bile of yellowtail, Seriola quinqueradiata.

An unknown bilirubin conjugate was detected by HPLC analysis in the bile of yellowtail, Seriola quinqueradiata. The retention time of this unknown conjugate was identical with that of authentic ditaurobilirubin, a bilirubin conjugated with 2 mol of taurine. The retention times of the ethyl anthranilate diazo derivatives of both bilirubin conjugates were the same. The azo derivatives of this unknown conjugate were purified by Amberlite XAD-2 treatment and preparative TLC and hydrolyzed with 6 N HCl at 105 degrees C for 12 h. The hydrolyzates obtained were analyzed with an amino acid analyzer. The ninhydrin-positive substance in the hydrolyzates was purified by column chromatography and analyzed by TLC and mass spectrometry. The results of these analyses indicated that this ninhydrin-positive substance was taurine. Therefore, this unknown bilirubin conjugate was identified as ditaurobilirubin.

[Cisplatin, peplomycin, mitomycin C, and vincristine combination chemotherapy of non-small cell carcinoma of the lung].

Twenty patients with unresectable non-small cell lung cancer were treated with a combination chemotherapy of cisplatin (30 mg/body i.v., days 1-5), peplomycin (5 or 8 mg/body continuous infusion, days 1-5), mitomycin C (4 mg/body i.v., day 1), and vincristine (2 mg/body i.v., day 1), of 15 patients evaluable for response (9 with squamous cell carcinoma, 2 with adenocarcinoma, nd 4 with large cell carcinoma), the overall response rate was 46.7% with 7 partial responses. The median survival period for responders. Toxicity included hair loss, nausea and/or vomiting, mild to moderate myelosuppression, nephrotoxicity, and pulmonary toxicity, all of which was manageable. This four-drug combination chemotherapy is concluded to be effective for non-small cell lung cancer.