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Using methodology recently developed to assess gene-specific DNA repair, we have demonstrated that it is possible not only to study mitochondrial DNA repair, but also directly to compare mitochondrial and nuclear DNA repair in the same biological sample. Complex enzymatic mechanisms recognize and repair nuclear DNA damage, but it has long been thought that there was no DNA repair in mitochondria. Therefore, in an attempt to delineate more clearly which DNA repair mechanisms, if any, are functioning in mitochondria, we have investigated the repair of several specific DNA lesions in mitochondrial DNA. They include cyclobutane dimers, cisplatin intrastrand adducts, cisplatin interstrand crosslinks and alkali-labile sites. We find that pyrimidine dimers and complex alkylation damage are not repaired in mitochondrial DNA, and that there is minimal repair of cisplatin intrastrand crosslinks. In contrast, there is efficient repair of cisplatin interstrand crosslinks as evidenced by approximately 70% of the lesions being removed by 24 h. Additionally, there is efficient repair of N-methylpurines following exposure to methylnitrosourea with approximately 70% of the lesions being removed by 24 h. The results of these studies reveal that repair capacity of mitochondrial DNA damage depends upon the type of lesion produced by the damaging agent. We speculate that a process similar to the base excision mechanism for nuclear DNA exists for mitochondrial DNA but that there is no nucleotide excision repair mechanism to remove more bulky lesions in this organelle.

We have explored the induction of DNA repair synthesis in monocyte/B- and T-lymphocyte enriched cell fractions from 12 different human mononuclear blood cell populations. Unscheduled DNA synthesis was measured in monocyte/B- and T-cells after exposure to the DNA-damaging agents dimethylsulfate (DMS) and N-acetoxy-N-acetylaminofluorene in vitro. Also, the binding of DMS to DNA was measured. An increased DNA repair synthesis was measured in monocyte/B-lymphocytes after induction of the two different types of DNA lesions, whereas no induction of unscheduled DNA synthesis was observed in T-lymphocytes. A significantly higher DMS-DNA binding was also observed in monocyte/B-lymphocytes when compared with T-lymphocytes. Specific characterization of mononuclear blood cell populations used in biomonitoring of DNA adducts and repair is recommended.

Unlabelled: Significant impairment of lung volumes and gas exchange in HIV-infected patients with acute Pneumocystis carinii pneumonia (PCP) has been reported, whereas little is known about lung function compromise following successful therapy. In 9 patients with acute PCP and 9 patients 1-5 month after PCP lung function testing including spirometry, diffusing capacity for carbon monoxide and exercise blood gas analysis were performed serially at monthly intervals. The results were summarized in a total score. The mean period of follow-up for each patient was 7.2 +/- 2 months. A decrease in lung volumes (FEV1 67 +/- 14.2% pred.norm., VC 72.7 +/- 11.9% pred.norm.) and gas exchange (CO-transfer factor 53.2 +/- 18.5% pred.norm., CO-transfer coefficient 67.8 +/- 14.2 pred. norm.) was observed in all 9 patients with acute PCP. Post-PCP lung volumes normalized within 1 month, whereas disorders in gas exchange persisted for 1-3 months. The total score normalized in 16/18 patients. One of the remaining patients with persisting functional impairment had chronic obstructive airway disease, whereas in the other dysfunction was even observed prior to PCP and no diagnosis could be obtained. 2-6 months following acute disease a second period of decreased lung function occurred in 6 pts. In 3 of the 6 there were no clinical signs of infection (rebronchoscopy refused), in 1 patient infection with cytomegalovirus was suspected. In the other 2 patients a relapse of PCP was diagnosed by bronchoscopy., Conclusions: Acute PCP compromises lung mechanics and gas exchange. During recovery deficits in gas exchange persist longer than diminished lung volumes.(ABSTRACT TRUNCATED AT 250 WORDS)

A biosurvey in the Danish metal industry measured the genotoxic exposure from stainless steel welding. The study comprised measurements of chromosomal aberrations (CA), sister-chromatid exchanges (SCE), unscheduled DNA synthesis (UDS) in peripheral lymphocytes and serum immunoglobulin G. Environmental monitoring of welding fumes and selected metal oxides, biomonitoring of chromium and nickel in serum and urine and mutagenic activity in urine, and evaluation of semen quality were also done. Manual metal arc (MMA) welding and tungsten inert gas (TIG) welding were the dominant welding processes. A higher frequency of chromosomal aberrations, classified as translocations, double minutes, exchanges and rings, was observed in stainless steel welders than in non-welders. SCE was lower in welders working with both MMA and TIG welding than in reference persons. N-Acetoxy-N-acetylaminofluorene (NA-AAF)-induced UDS was lower in 23 never-smoking welders than in 19 unexposed never-smokers. Smoking was a confounding factor resulting in significantly higher CA, SCE, NA-AAF binding to DNA and mutagenic activity in urine. Age was also a confounder: CA, SCE, NA-AAF binding to DNA and UDS increased significantly with age. No significant correlation between SCE and CA or between CA and UDS was found. UDS decreased significantly with increasing lymphocyte count and a higher lymphocyte count was seen in MMA welders than in reference persons and in smokers than in non-smokers. Differences in the composition among lymphocytes in exposed persons compared with non-exposed are suggested. MMA welding gave the highest exposure to chromium, an increased number of chromosomal aberrations and a decrease in SCE when compared with TIG welding. Consequently improvements in the occupational practice of stainless steel welding with MMA is recommended.

Thirty patients with a history of asthma and ten patients with suspected bronchial hyperreactivity underwent nonspecific provocation testing. The control group consisted of ten normal volunteers without a history of lung disease. The patients' baseline FEV1 (percent predicted) revealed mild obstructive disease (72.9 +/- 8.9 percent and 74.6 +/- 7.7 percent) compared with controls (87.2 +/- 8.5 percent, p less than 0.001). The mean volume of trapped gas (D) (ie, TLCB-TLCHe) was not significantly different between groups (0.11 +/- 0.49 L vs 0.15 +/- 0.4 L vs 0.18 +/- 0.45 L), and no correlation was established with any of the remaining lung function data. Bronchial hyperreactivity in response to inhaling acetylcholine could be observed in the asthma group only. Their mean D increased significantly from 0.11 +/- 0.49 L to 0.62 +/- 0.66 L (p less than 0.001), and returned to baseline (0.26 +/- 0.55, NS) subsequent to inhaling salbutamol. D changes induced by acetylcholine correlated weakly with concurrent changes of FEV1 (r = -0.44, p = 0.01), RV (r = 0.59, p less than 0.001), and Rs (r = 0.59, p less than 0.001). In response to bronchodilating doses of salbutamol, however, D was changed in close correlation with FEV1 (r = -0.82, p less than 0.0001), RV (r = 0.85, p less than 0.0001), and Rs (r = 0.76, p less than 0.0001). Provided that D is a valid parameter of small airways function, these data may give a clue to the site of action of both drugs. Acetylcholine affects small and large airways alike with no clear-cut preference, whereas salbutamol's predominant target appears to be the small airways. These conclusions are only partially supported by the pertinent literature.

The kind of relation of central lung cancer (c) to the walls of the central pulmonary arteries (PA) and the aorta is an important information prior to operative or interventional (laser/afterloading) therapy. As computed tomography (CT) and angiography are often inaccurate in the assessment of PA-infiltration, we assessed the diagnostic value of transesophageal echography (TEE) in the staging of LC. 16 patients (pts.) were investigated using TEE in addition to CT or magnetic resonance imaging (MRI). Eleven pts. had central LC, 3 peripheral LC, 1 anterior mediastinal mass and 1 central pneumonia (cancer excluded). 2 pts. with central LC were unable to swallow the probe. In 9/9 pts. with central LC, 1/3 pts. with peripheral LC and 1 pt. with enlarged anterior mediastinum the tumour mass could be visualized. In the pt. with a centrally located infiltrate on chest radiogram TEE demonstrated enlarged hilar lymph nodes, but excluded a central tumour. Main PA branches could be identified in all 14/14 pts. Central left or right PA were compressed slightly in 3 pts. and severely in 2 pts., with a near total occlusion in one (confirmed by MRI/CT). TEE revealed PA-infiltration in 2 pts. and aortic wall infiltration in 2 other pts. Despite adjacent tumour mass aortic wall infiltration was excluded in 2 pts. Enlarged hilar lymph nodes could be demonstrated in 2/9 pts. with central LC, whereas CT/MRI showed enlarged mediastinal lymph nodes in 7/9 pts. In conclusion, TEE is able to visualize central lung cancer and gives useful additional informations about the kind of relation to central PA and the aorta.(ABSTRACT TRUNCATED AT 250 WORDS)

We surveyed 11 Burkitt's lymphoma cell lines for chemosensitivity to nitrogen mustard (HN2) in order to determine whether any simple correlates to cytotoxic response might be revealed. The lines tested varied over a 5-fold range in concentration of HN2 required to inhibit tumor cell growth by 50%. Drug sensitivity correlated neither with continental origin of tumor, growth fraction, presence of Epstein-Barr virus, nor with the precise locations of (8;14) translocation breakpoints. Furthermore, contrary to experience with other cell lines, no simple correlation was found between the HN2 sensitivity of the four most divergent lines (low sensitivity, CA46 and MC116 cells; high sensitivity, Namalwa and JLP119 cells) and exposure to DNA cross-links (area under the DNA cross-linking-versus-time curve). In addition, we found similar extents of gene-specific HN2-induced damage in the native and translocated c-myc alleles of CA46 and JLP119 cells. At equimolar HN2 treatment, CA46 cells exhibited a profound arrest in G2M phase, while JLP119 cells exhibited prolonged S-phase delay. This suggested that despite similar DNA cross-link exposure, JLP119 cells were less able to complete DNA replication while repair was in progress. As cell cycle distribution returned to near normal, JLP119 cells exhibited DNA degradation characterized by oligonucleosome-sized DNA fragments prior to cell membrane disintegration. Our findings indicate that HN2-sensitive Burkitt's lymphoma cells may be more susceptible to delay in S phase for a given frequency of DNA cross-links and that prolongation of S phase correlated with apoptotic cell death.

The effect of combinations of the anthracyclines aclarubicin and daunorubicin was investigated in a clonogenic assay using the human small cell lung cancer cell line OC-NYH and a multidrug-resistant (MDR) murine subline of Ehrlich ascites tumor (EHR2/DNR+). It was found that the cytotoxicity of daunorubicin in OC-NYH cells was antagonized by simultaneous exposure to nontoxic concentrations of aclarubicin. Coordinately, aclarubicin inhibited the formation of daunorubicin-induced protein-concealed DNA single-strand breaks and DNA-protein cross-links in OC-NYH cells when assayed by the alkaline elution technique. Aclarubicin had no influence on the accumulation of daunorubicin in these cells. In contrast, the accumulation of daunorubicin in EHR2/DNR+ cells was enhanced by more than 300% when the cells were simultaneously incubated with the MDR modulator verapamil, aclarubicin, or the two agents combined. Yet the cytotoxicity of daunorubicin was potentiated significantly only by verapamil. The increased cytotoxicity of daunorubicin in the presence of verapamil was completely antagonized when aclarubicin was used together with the MDR modulator. Finally, the effect of daunorubicin on the DNA cleavage activity of purified topoisomerase II in the presence and absence of aclarubicin was examined. It was found that daunorubicin stimulated DNA cleavage by topoisomerase II at specific DNA sites. The addition of aclarubicin completely inhibited the daunorubicin-induced stimulation of DNA cleavage. Taken together, these data indicate that aclarubicin-mediated inhibition of daunorubicin-induced cytotoxicity is due mainly to a drug interaction with the nuclear enzyme topoisomerase II. This antagonism at the nuclear level explains why aclarubicin is a poor modulator of daunorubicin resistance even though aclarubicin is able to increase the intracellular accumulation of daunorubicin in a MDR cell line.

In order to determine the possible effect of aerosolized pentamidine on the cellular composition of the bronchoalveolar lavage fluid in HIV-infected patients, differential counts of 22 consecutive patients who had been rebronchoscopied after 3-19 months were reviewed. Eleven patients were started on pentamidine prophylaxis subsequent to their first presentation. Eleven patients had never taken pentamidine or had discontinued the prophylactic regimen. Compared to first bronchoscopy, the bronchoalveolar lavage (BAL) from patients on regular prophylaxis revealed a significant increase in absolute alveolar macrophage (AM) counts at second presentation (20.8 +/- 11.2 to 50.3 +/- 39.4 x 10(5) cells/100 ml BAL; P less than 0.01). The AM counts of those without pentamidine remained essentially unchanged. Lymphocytes, including CD4 and CD8 subtypes, and neutrophils did not change over time in either group. The results of this retrospective analysis suggest that, in addition to its antimicrobial action, pentamidine may modulate local lung defence mechanisms, particularly by increasing the absolute number of AM.

Liblomycin (LBM), a novel bleomycin analogue, and bleomycin A2 (BLM A2) were compared with respect to their relative potential to inhibit growth in a human head and neck squamous carcinoma cell line and to produce DNA damage within cellular DNA and nuclei DNA and against isolated naked DNA. Against the BLM-sensitive cell line 183A, the concentration of LBM that inhibits cell growth by 50% was 1.1 microM for a 30-min drug exposure, while it was 23 microM for BLM A2. Drug-mediated DNA double-strand cleavage within cells was compared with the relative ability of these drugs to produce DNA cleavage in isolated 183A cell nuclei. Though 30-min exposures of cells to equimolar concentrations of both drugs resulted in 4-fold greater cellular DNA damage by LBM than BLM A2, the two drugs were nearly equipotent in producing DNA injury within isolated nuclei. Against Simian virus 40 DNA, however, LBM was 10-fold less effective than BLM A2 in producing Forms II and III DNA from Form I DNA. Radioactivity from either [3H]BLM A2 or 125I-LBM found associated with cells after a 30-min incubation period was also assessed in the 183A cell line. The exposure of cells to radiolabeled drug (1 microM) resulted in a 71-fold greater amount of cell-associated radioactivity for LBM than for BLM A2. The relative abilities of the 183A cell line to partially reseal LBM- or BLM A2-mediated DNA double-strand breaks were also assessed. No preferential repair of overall drug-mediated DNA injury, however, was observed. Finally, drug-mediated specific cleavage sites on pBR322 DNA were determined. At doses that gave the same extent of DNA cleavage, both BLM A2 and LBM gave similar patterns of strand scission, although minor differences were observed. Taken together, these data demonstrate that the greater efficacy of LBM against the BLM-sensitive head and neck squamous cell line is due mainly to LBM's greater association with cells over a defined time period, even though the DNA cleaving ability of LBM is relatively lower than that of BLM A2.

Doxorubicin (DX) was found to inhibit the incorporation of [1-14C]linoleic acid and [1(3)-3H]glycerol into the major membrane phosphoglycerides, phosphatidylcholine and phosphatidylethanolamine of cultured myocardial cells in a dose-dependent manner (0.16-16 microM). It is suggested that DX affects de novo biosynthesis of these lipids. In contrast, DX-treatment of the cells stimulated incorporation of [1-14C]linoleic acid into triacylglycerol. The effects of DX on lipid metabolism were only demonstrable 20-24 hr after a 1 hr exposure of the cells to the drug indicating that DX exerts little or no direct effect on the enzymes participating in lipid synthesis and that the alterations in lipid metabolism induced by DX probably are secondary to inhibition of protein synthesis and progressive cell injury. Extensive peroxidative decomposition of membrane lipids appeared not to take place in the DX-treated cells as judged from fatty acid analysis of total membrane phosphoglyceride.

Cytotoxicity, extent of DNA double-strand breaks, and stimulation of poly(adenosine diphosphoribose)-synthesis were measured in two established human head and neck squamous carcinoma cell lines (183A and 1483) following x-irradiation. The 1483 cell line was 15-fold more resistant to x-ray-mediated cytotoxicity than was the 183A cell line. X-ray-mediated DNA strand cleavage also differed in these two cell lines with the absolute frequency of DNA double-strand breaks in the sensitive cells 183A cells being twice that in the resistant 1483 cell line. No detectable stimulation of poly(adenosine diphosphoribose)-synthesis was measured in the sensitive 183A cells whereas a marked increase in incorporation of [3H]-nicotinamide adenine dinucleotide was readily detected following x-irradiation of the resistant 1483 cells. These findings suggest a possible role of altered poly(adenosine diphosphoribose)-synthesis in the sensitivity of human head and neck squamous carcinoma cells to ionizing radiation.

The potential mechanisms of the extremely potent anthracycline analogue 3'-deamino-3'-(3-cyano-4-morpholinyl)doxorubicin (MRA-CN) have been compared with those of doxorubicin (DOX) by examination of drug effects on colony formation, macromolecular synthesis, DNA integrity, and ultrastructure of human leukemia cells in vitro. Following a 1-h exposure, MRA-CN was found to be 1400-fold more cytocidal than DOX which correlated with the drugs' inhibitory effects on DNA and total RNA synthesis. Treatment with MRA-CN resulted in a dose-dependent production of DNA interstrand cross-links as quantified by alkaline elution. One-h treatments with DOX or 3'-deamino-3'-(4-morpholinyl) doxorubicin (the non-cyano-containing analogue of MRA-CN) produced no DNA-DNA cross-links; rather they produced protein-concealed DNA single-strand breaks. After removal of MRA-CN, the DNA of KBM-3 cells displayed time-dependent fragmentation as indicated by rapid DNA filter elution during the pH 10 lysis step which preceded pH 12 elution. Within 4 h of MRA-CN exposure (10 nM, 1 h), 50% of the cellular DNA was in the lysis fraction. By 24 h, all the cellular DNA was in this fraction. MRA-CN (10 nM), 3'-deamino-3'-(4-morpholinyl)doxorubicin (1 microM), and actinomycin D (1 microM), but not DOX (3 mircroM), each produced distinctive nucleolar macrosegregation, indicating an effect on rRNA synthesis. The alpha-CN substituent on the morpholinyl moiety of MRA-CN appears to be responsible for the unique antitumor potency of this anthracycline. Nucleolar macrosegregation is probably associated with the morpholinyl moiety and is independent of the alpha-CN substituent.

On the basis of comprehensive clinicopathological evidence 6 patients presenting with unilateral left-sided hyperlucent lungs are evaluated for pathogenesis, function and postoperative (n = 5) course. The common denominator in all proves to be moderate to severe hypoperfusion and overinflation of the respective lungs. Pulmonary function is characterized by a combined restrictive-obstructive pattern. In 4 patients overinflation is due to a central check-valve mechanism (tumor: n = 3; central airways collapse: n = 1), whereas in 2 increased translucency results from some sort of peripheral obstruction (Swyer-James syndrome: n = 1; congenital cystic bronchiectasis: n = 1). We consider the origin of hypoperfusion to be alveolar distension and hypoxic precapillary vasoconstriction, both participating in diminished blood flow to the check-valve obstructed lung. In Swyer-James syndrome reduced vascularity is an additional feature. Preoperative and long-term postoperative lung function data of 5 pneumonectomized patients are compared. On the whole, FEV1 and IVC remain unchanged, whereas the obstructive profile (RV, RV/TLC, sRAW) improves. From these data it is concluded that the affected hyperlucent lung is 'amputated' even before operation - irrespective of the nature of tissue damage. On the other hand postoperative relief of airways obstruction is supposed to be due to both antiobstructive medication and the removal of a diseased lung.

The antineoplastic activity of the anthracycline antibiotic doxorubicin (Adriamycin) differs in its cytotoxic effectiveness against different types of human tumors. In the present study the effect of doxorubicin on the growth of two human lung carcinomas and one human mammary carcinoma transplanted into athymic mice was correlated with the pharmacokinetics of doxorubicin in the same tumors after intraperitoneal administration. Doxorubicin produced a greater inhibition of tumor growth in the lung carcinomas than in the mammary carcinoma. Furthermore, the pharmacokinetic characteristics of doxorubicin differed widely within the three human solid tumors. No apparent correlation was found to exist between the different tumor growth sensitivities to doxorubicin and the pharmacokinetic parameters of doxorubicin within the tumor tissue. It is suggested that the differences in the demonstrated antitumor effectiveness of doxorubicin may be due to differences in the "intrinsic sensitivity" of the three human solid tumors.

Upon incubation of cultured mammalian cells with the new anthracycline analogues cyanomorpholinyldoxorubicin and morpholinyldoxorubicin, nucleoli irreversibly segregate into their substructures which form individual portions of the nucleolar mass and characteristic electron-dense components adjacent to the nucleolonema; these changes in nucleolar ultrastructure are similar to those produced by actinomycin D (AMD). In the present study we have examined the effects of anthracycline analogues on RNA synthesis, localization of RNA polymerase I in situ, and activity of RNA polymerases in vitro, and compared these effects with those of the parent compound doxorubicin (DOX) and AMD. The results show that, following treatment with cyanomorpholinyldoxorubicin, morpholinyldoxorubicin, and AMD, but not DOX, RNA polymerase I-containing transcription complexes were reduced, reflecting the transcriptional activity of the rRNA genes. The residual RNA polymerase-containing entities were redistributed into cap-like aggregates at the nucleolar periphery. Within 30 min of exposure to cyanomorpholinyldoxorubicin, morpholinyldoxorubicin, and AMD, but not DOX, a 75-90% inhibition of RNA polymerase I activity in situ and in vitro was observed. At this early time there was no significant inhibition of nucleoplasmic RNA labeling in situ or RNA polymerases II and III activities in vitro. At later times following reincubation in drug-free medium, inhibition of all three polymerases was observed. Impairment of RNA synthesis appeared to result from drug interaction with the DNA template rather than an interaction with RNA polymerase I itself. We conclude that the morpholinyl derivatives of DOX are preferential inhibitors of ribosomal gene transcription and that they may have a mechanism of action similar to that of AMD on rRNA synthesis.

The effect of intravenous infusion of dopamine (10 and 25 micrograms X kg-1 X min-1 consecutively) on visceral blood flow distribution was examined in anesthetized cats using the microsphere technique and electromagnetic flowmetry. Arterial blood pressure did not change in response to dopamine infusion, but blood flow through the superior mesenteric artery, and blood flow in the mucosa-submucosa of the gastric antrum and various gut segments increased significantly. During infusion of the high dose the increase was most marked in the mucosa-submucosa of the antrum (+355%) and distal colon (+371%). By contrast, blood flow decreased in the muscularis-serosa of the gut segments investigated, in the spleen, pancreas, and the hepatic arterial bed. The increase in blood flow through the superior mesenteric artery was blocked by the dopamine antagonist bulbocapnine (10 mg/kg i.v.). The results suggest that the receptors mediating the dopamine-induced vasodilation in the gastrointestinal tract are located in the resistance vessels of the mucosa-submucosa.