Lung fibroblasts from patients with emphysema show markers of senescence in vitro.

BACKGROUND: The loss of alveolar walls is a hallmark of emphysema. As fibroblasts play an important role in the maintenance of alveolar structure, a change in fibroblast phenotype could be involved in the pathogenesis of this disease. In a previous study we found a reduced in vitro proliferation rate and number of population doublings of parenchymal lung fibroblasts from patients with emphysema and we hypothesized that these findings could be related to a premature cellular aging of these cells. In this study, we therefore compared cellular senescence markers and expression of respective genes between lung fibroblasts from patients with emphysema and control patients without COPD. METHODS: Primary lung fibroblasts were obtained from 13 patients with moderate to severe lung emphysema (E) and 15 controls (C) undergoing surgery for lung tumor resection or volume reduction (n = 2). Fibroblasts (8E/9C) were stained for senescence-associated beta-galactosidase (SA-beta-Gal). In independent cultures, DNA from lung fibroblasts (7E/8C) was assessed for mean telomere length. Two exploratory 12 k cDNA microarrays were used to assess gene expression in pooled fibroblasts (3E/3C). Subsequently, expression of selected genes was evaluated by quantitative PCR (qPCR) in fibroblasts of individual patients (10E/9C) and protein concentration was analyzed in the cell culture supernatant. RESULTS: The median (quartiles) percentage of fibroblasts positive for SA-beta-Gal was 4.4 (3.2;4.7) % in controls and 16.0 (10.0;24.8) % in emphysema (p = 0.001), while telomere length was not different. Among the candidates for differentially expressed genes in the array (factor > or = 3), 15 were upregulated and 121 downregulated in emphysema. qPCR confirmed the upregulation of insulin-like growth factor-binding protein (IGFBP)-3 and IGFBP-rP1 (p = 0.029, p = 0.0002), while expression of IGFBP-5, -rP2 (CTGF), -rP4 (Cyr61), FOSL1, LOXL2, OAZ1 and CDK4 was not different between groups. In line with the gene expression we found increased cell culture supernatant concentrations of IGFBP-3 (p = 0.006) in emphysema. CONCLUSION: These data support the hypothesis that premature aging of lung fibroblasts occurs in emphysema, via a telomere-independent mechanism. The upregulation of the senescence-associated IGFBP-3 and -rP1 in emphysema suggests that inhibition of the action of insulin and insulin-like growth factors could be involved in the reduced in vitro-proliferation rate.

Comparison of exhaled nitric oxide analysers.

Currently no published data are available concerning the comparability of different types of NO analysers, making inter-laboratory comparisons difficult. In two sets of experiments we compared 4 and 5 NO analysers, respectively, from 3 different manufacturers using different calibration regimes: calibration with (1) a separate recommended calibration gas for each analyser, (2) a single low concentration for all (394 ppb), and (3) a single high concentration (12.8 ppm). We measured three subjects with known low (L), moderate (M) and high (H) bronchial exhaled nitric oxide concentrations as well as standard gases (SG). In the first set of experiments, calibration regime 1 resulted in the largest differences between analysers (coefficient of variation (CV) for L, M, H, SG: 0.42, 0.22, 0.20, 0.14). The lowest CV between analysers was observed after calibration 2 (0.34, 0.19, 0.12, 0.02). Very similar results were obtained in the second set of comparisons. Thus, differences between analysers existed, but were mainly due to differences in recommended calibration gases/procedures. Only a small part was explainable by deviations from target flow. These differences need to be taken into account when comparing data between laboratories or replacing the calibration gas of an analyser, as well as for the establishment and interpretation of normal values.

[Detection of tartrate-resistant isoenzyme 5 of acid phosphatase in the white blood cells by disc electrophoresis in a case of hairy cell leukemia]. / Diskelektrophoretischer Nachweis des tartratresistenten Isoenzym 5 der sauren Phosphatase in den weissen Blutzellen bei einem Fall von Haarzell-Leukämie.

In a case of hairy cell leukemia the evidence of tartrate-resistant isoenzyme 5 of phosphatase (TRISP 5) developed to findings important for the diagnosis. The evidence was achieved by means of disc electrophoresis by separating the cell lysate of white blood cells. The percentage of TRISP 5 in the total activity of acid phosphatase in leukocytes amounted to 17.6%. In cytochemical respect the evidence of TRISP 5 proved to be negative. The classification of these findings and their differential-diagnostic significance are discussed.

[Modification of special blood-level forming lipid metabolites by long-distance running in men 20-60 years of age]. / Beeinflussung spezieller blutspiegelbildender Lipidmetaboliten bei Männern im Alter von 20-60 Jahren durch Langstreckenlauf.

The lipid metabolism and the lipoprotein metabolism was examined in 44 long-distance runners (people's athletics) and a group of 65 untrained persons as well as in 32 runners before and immediately after a 38-km and a 40-km-run, respectively. High density lipoprotein cholesterol (HDLC) was significantly increased in the runners, whereas very low density lipoprotein cholesterol (VLDLC) was found decreased. The quotient LDLC/HDLC (quotient of arteriosclerosis) is clearly decreased in runners. Total cholesterol and low density lipoprotein cholesterol (LDLC) of runners (people's athletics) and untrained persons were statistically not different. The post-heparin-lipase (pHL) and the rate of lipolysis pHL appeared clearly increased in persons participating in people's athletics. There were parallelities between the lipid values and the disc-electrophoretically established percental proportions of lipoproteins. The proportion of alpha-lipoprotein (Lp) of the runners was increased, the beta-Lp unchanged and the pre-beta-Lp decreased. The increase of the alpha-Lp was concomitant with a shift of the spectre of the alpha-Lp-subfractions. After the running (38 and 40 km) the triglycerides and VLDLC were increased, the pHL-activity and the rate of lipolysis pHL were decreased. In the other cholesterol parameters no significant shifts appeared after load. While Apo A does not change under the influence of the running, like HDLC, Apo B clearly increased. The results were discussed, taking into consideration the results of other authors.

[Effect of physical conditioning on fat metabolism--examinations performed on middle-aged long distance runners]. / Einfluss physischer Konditionierung auf den Fettstoffwechsel--Untersuchungen an Langstreckenläufern im mittleren Lebensalter.

The lipid and lipoprotein metabolism was examined in fortyfour long distance runners and in a cohort of sixty-five untrained subjects as well as in thirty-two runners before and immediately after 38- and 40-kilometer runs respectively. There was observed a significant increase of high density lipoprotein cholesterol in the runners, whereas very low density lipoprotein cholesterol showed a marked decrease. The low density lipoprotein cholesterol/high density lipoprotein cholesterol quotient (arteriosclerotic quotient) was found to be markedly reduced in the runners. There was no statistical difference in total cholesterol and low density lipoprotein cholesterol between long distance runners following sports as a pastime and untrained subjects. Post-heparin lipase and the rate of lipolysis were markedly increased in subjects following sports as a recreation. Parallelisms were observable between lipid values and lipoprotein percentages determined by disk electrophoresis. The proportions of alpha-lipoprotein, beta-lipoprotein, and pre-beta-lipoprotein in runners were found to be increased, unchanged, and reduced respectively. The increase in alpha-lipoprotein was accompanied by a shift in the spectrum of alpha-lipoprotein subfractions. After the 38- and 40-kilometer run, respectively, there were determined an increase in TGL and very low density lipoprotein cholesterol and a decrease in both post-heparin lipase and rate of lipolysis. The other cholesterol parameters showed no significant variations after straining. Whereas Apo A, like high density lipoprotein cholesterol, did not change during running, Apo B showed a marked increase. The results were discussed with due consideration of those reported by other authors.

Lung fibroblasts from patients with emphysema show a reduced proliferation rate in culture.

Emphysema is characterised by a loss of alveolar structure, as reflected in elastic recoil and gas exchange. As fibroblasts play a key role in the maintenance of structure, the current authors hypothesised that their proliferation might be constitutively impaired in lung emphysema. Using explant cultures, lung fibroblasts were obtained from resected lungs of 10 patients with emphysema (median forced expiratory volume in one second (FEV1) 40% predicted) and 10 control patients (FEV1, 95% pred). The doubling time (DT) was measured over 4 days under standard conditions (10% foetal calf serum) prior and after cryopreservation. Additionally, in seven samples per group the total population doubling level (PDL) was determined. In emphysema, mean+/-sem DT was 33.6+/-2.8 h compared with 24.8+/-1.4 h in controls. The differences in DT were preserved after cryopreservation. Groups also differed in the initial slope of the PDL plot during long-term culture (up to 35 days). However, the median (range) maximum PDL did not differ significantly between groups (13.8 (7.4-22.6) versus 20.2 (11.2-25.5)). The current authors, therefore, suggest that the reduced proliferation rate in vitro of lung fibroblasts from patients with emphysema reflects a persistent, intrinsic failure of cellular replacement and maintenance in this disease, possibly in relation to pre-term aging.