Rebreathing K(CO) and single breath K(CO) measured at the same mean alveolar volume reveal a physiological relationship between unequal diffusion and unequal ventilation in normal subjects.

BACKGROUND: The single breath (SB) method for determining the transfer factor for carbon monoxide is influenced by an unequal distribution of diffusion and ventilation. The rebreathing (RB) method is thought not to be influenced by these inequalities. Therefore, a comparison of the results of the two methods enables us to investigate unequal ventilation and diffusion. We have previously shown that even in normal subjects unequal ventilation influences the relation between the transfer factor as measured by the RB method and by the SB method. OBJECTIVES: To investigate to what extent unequal diffusion influences the relation between the RB and SB transfer factor in normal subjects. METHODS: Measurements were performed in 8 normal subjects. Differences in alveolar volume as measured by the RB and SB method were prevented from influencing the results by comparing the RB and SB transfer factor per liter of alveolar volume, i.e. the RB and SB Krogh factor, instead of the RB and SB transfer factor. The effect of the known dependence of the Krogh factor on inspired volume was prevented by measuring the SB Krogh factor at an inspired volume equal to the mean inspired volume of the RB measurement. This SB Krogh factor was compared with the RB Krogh factor calculated as the quotient of the RB transfer factor and the mean alveolar volume during the RB measurement. RESULTS: This RB Krogh factor was always larger than the corresponding SB Krogh factor. The difference was significant (p = 0.006). A high percentage (80%) of the variance in the quotient of the two Krogh factors, which can be considered as a parameter for unequal diffusion, was explained by the variance in the quotient of the mean RB volume and the corresponding SB volume. The latter quotient is a measure of unequal ventilation. CONCLUSIONS: The finding suggests a physiological relation between unequal diffusion and unequal ventilation. A plausible explanation was found in the decrease of the Krogh factor with an increasing ratio of inspired volume to residual volume for each part of the lung separately. The consequence is that in the absence of unequal ventilation the RB transfer factor is equal to the SB transfer factor measured at an inspired volume equal to the mean RB inspired volume.

Re-breathing vs single-breath TLCO in patients with unequal ventilation and diffusion.

The single-breath (SB) method for determining the transfer factor for carbon monoxide (TLCO) is of limited value for the detection of diffusion disorders on the alveolar level, because the results are influenced by unequal distribution of ventilation and diffusion. The rebreathing method (RB) is thought not to be influenced by these inequalities. To the authors' knowledge, no study has measured both TLCORB and TLCOSB systematically and compared them with regard to the influence of unequal ventilation and diffusion. Therefore, the present study measured total lung capacity (TLC) as well as TLCO, both with the RB vital capacity method and the SB method, using the same apparatus in 10 healthy subjects and in 35 patients with chronic obstructive pulmonary disease (COPD). These patients are known to have increased unequal ventilation and diffusion in comparison with healthy subjects. In the healthy subjects, a small difference was found between TLC measured with the RB method (TLCRB) divided by the predicted value (TLCRB/pred) and TLCSB/pred (mean difference 0.07; SE = 0.02); no significant difference was found between TLCORB divided by the predicted value of TLCOSB (TLCORB/pred) and TLCOSB/pred. In the COPD patients, however, TLCRB/pred was larger than TLCSB/pred (mean difference 0.17; SE = 0.02) and TLCORB/pred was larger than TLCOSB/pred (mean difference 0.23; SE = 0.05). Multiple regression analysis revealed that in the COPD patients, 54% of the variance of the difference between TLCRB/pred and TLCSB/pred, and 76% of the variance of the difference between TLCORB/pred and TLCOSB/pred, were explained by parameters related to unequal ventilation and diffusion. In 25 of the 35 COPD patients, TLCOSB/pred was less than 0.8, whereas in 11 of these 25 patients, TLCORB/pred was more than 0.8. This difference was significant (P = 0.0005). In these 11 patients, the SB measurement resulted in the incorrect diagnosis of a diffusion disorder on the alveolar level. The RB method, however, never resulted in the diagnosis of a diffusion disorder when TLCOSB/pred was larger than 0.8. It is concluded that in a significant number of COPD patients, TLCOSB is below the normal range, whereas TLCORB is not below the normal range. This difference between TLCORB and TLCOSB is related to the combined effect of unequal ventilation and diffusion, and is of clinical importance for the detection of a diffusion disorder on the alveolar level.

Rebreathing TLCO versus single breath TLCO at different degrees of unequal ventilation.

The influence of unequal ventilation on the differences between the rebreathing (RB) and the single breath (SB) measurement of the transfer factor of the lung for carbon monoxide (TLCO) was investigated. An apparatus was developed which measured both TLCORB and TLCOSB. Unequal ventilation increases with lung volume, caused by the combined effect of increasing ratio of total volume to RV from apex to basis, the fact that apex inspires before basis and asymmetric intra-acinar branching patterns. Therefore measurements were done at three different values of inspired volume (VI) in 10 normal subjects. To separate the effects of gas mixing and diffusion, residual volume (RV) was measured by He dilution in addition to TLCO. We found that RV and TLCO increase with increasing VI. The increase is larger with RB. We also found that RVRB is larger than RVSB at the three values of VI. TLCORB is smaller than TLCOSB except at vital capacity (VC). We concluded from the different behavior of RVRB and RVSB as a function of VI that the SB measurement increasingly underestimated RV with increasing unequal ventilation. This is also reflected in the measurement of TLCOSB. From the different behavior of TLCORB and TLCOSB as a function of VI we concluded that TLCORB was smaller than TLCOSB when unequal ventilation was minimal. This is caused by the smaller mean alveolar volume during RB. But increasing unequal ventilation causes an increasing underestimation with the SB method, resulting in TLCOSB being equal to TLCORB at VC.

Extracellular and intracellular magnesium concentrations in asthmatic patients.

Magnesium deficiency is associated with increased contractility of smooth muscle cells. Since contractility of bronchial smooth muscle is important in patients with asthma, magnesium deficiency could negatively influence the clinical condition. We wanted to assess whether magnesium deficiency exists in patients with asthma. Extracellular (plasma) and intracellular (erythrocytes and mononuclear leucocytes) concentrations of magnesium were determined in 20 mildly symptomatic patients with asthma and compared to 20 healthy controls. In asthmatic patients, the mean +/- SD magnesium level in plasma was 0.81 +/- 0.05 mmol.l-1, in erythrocytes 0.20 +/- 0.02 fmol.cell-1, and in mononuclear leucocytes 5.10 +/- 2.55 fmol.cell-1; these values did not differ significantly from those of the healthy controls: 0.79 +/- 0.06 mmol.l-1, 0.19 +/- 0.02 fmol.cell-1, and 4.61 +/- 1.75 fmol.cell-1, respectively. No evidence for the existence of a magnesium deficit needing chronic magnesium supplementation was, thus, found in these patients.

Occupancy of platelet receptors for platelet-activating factor in asthmatic patients during an allergen-induced bronchoconstrictive reaction.

The administration of platelet-activating factor (PAF) to human subjects triggers asthma-like responses. We investigated whether a bronchoconstrictive reaction was accompanied by the release of PAF in the circulation of allergic asthmatics. The appearance of PAF was assessed by measuring the number of freely accessible PAF-receptors on platelets in vitro, assuming that the contact between platelets and PAF in vivo would reduce the receptor binding of [3H]PAF in vitro. 16 asthmatics were challenged twice, first with buffer and the next day with allergen. A comparison between receptor binding after provocation with the data of the same patients after allergen challenge revealed a significant difference in PAF binding (P = 0.034), with an average decrease of 14% immediately after allergen challenge followed by a return to control values after about 4 h and a transient increase of 9% at 7 h after provocation. The decrease in accessible PAF receptors was accompanied by a slight decrease in platelet count in peripheral blood between 30 min and 4 h after allergen challenge. The platelet counts recovered to the original values afterwards. These data support the concept that in patients with allergic asthma PAF is secreted in the circulation. The contact between PAF and the platelets may trigger the transient sequestration of platelets, possibly in the lung. Thus, PAF and platelets may contribute to the pathogenesis of allergic asthma.

Down modulation of L-Selectin expression on eosinophils recovered from bronchoalveolar lavage fluid after allergen provocation.

In allergic asthma eosinophils infiltrate into the lung after allergen challenge. The mechanism of this cellular infiltration is not fully understood. L-Selectin is involved in leucocyte-endothelial cell recognition and participates in homing of leucocytes into sites of inflammation. To find indications for a role of L-Selectin in the migration of eosinophils to the bronchoalveolar space we measured L-Selectin expression on eosinophils in peripheral blood and bronchoalveolar lavage fluid (BAL) 4 hr after the early allergic reaction after allergen challenge. Nine patients with allergic asthma participated in the study. An eosinophil specific high depolarization signal enabled us to measure L-Selectin expression on eosinophils in a FACS analysis without isolation of these cells. Eosinophils recovered from BAL showed a strong decrease of L-Selectin expression compared to blood eosinophils. This decrease in L-Selectin expression can be induced in vitro by activation of eosinophils with PMA or FMLP whereas priming of eosinophils during several hours with GM-CSF did not influence L-Selectin expression. Our results are a first indication that L-Selectin may play a role during homing of eosinophils in the lung in asthma after allergen challenge. Moreover, the low expression of L-Selectin on eosinophils in the lung is a further indication that these cells exhibit an activated phenotype.

Modulation and induction of eosinophil chemotaxis by granulocyte-macrophage colony-stimulating factor and interleukin-3.

Eosinophilia and eosinophil function are regulated by cytokines such as granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-5. We have investigated the modulatory role of GM-CSF and IL-3 on the platelet-activating factor (PAF)-, neutrophil-activating factor (NAF/IL-8)-, leukotriene B4 (LTB4)-, N-formyl-methionyl-leucyl-phenylalanine (FMLP)-, and human complement factor C5a-induced chemotaxis of eosinophils from normal individuals. These eosinophils show a chemotactic response toward PAF, LTB4, and C5a, but not to NAF/IL-8 and FMLP. Preincubation of the eosinophils with picomolar concentrations of GM-CSF caused a significant increase in the response toward LTB4 and induced a significant chemotactic response toward NAF/IL-8 and FMLP. Preincubation of the eosinophils with picomolar concentrations of IL-3 also induced a chemotactic response toward NAF/IL-8 and FMLP, and enhanced the PAF-induced chemotaxis response toward C5a was not influenced by both cytokines. Nanomolar concentrations of GM-CSF or IL-3 caused a significant inhibition of the C5a-induced chemotaxis. The LTB4-induced chemotaxis was also significantly inhibited in case of GM-CSF. At these concentrations both GM-CSF and IL-3 acted as chemotaxins for eosinophils were washed after pretreatment with GM-CSF and IL-3 the potentiation of the chemotactic response remained, whereas the inhibitory mode of action disappeared. Our data indicate that at picomolar concentrations the cytokines GM-CSF and IL-3 can modulate eosinophil chemotaxis and at nanomolar concentrations these cytokines can act as chemotaxins for eosinophils.

Inhibition of neutrophil and eosinophil induced chemotaxis by nedocromil sodium and sodium cromoglycate.

1. Neutrophils and eosinophils infiltrate the airways in association with the allergen-induced late phase asthmatic reaction. Mobilization of these cells takes place via lipid-like and protein-like chemotactic factors. In this study platelet-activating factor (PAF), leukotriene B4 (LTB4), zymosan-activated serum (ZAS) and N-formyl-methionyl-leucyl-phenylalanine (FMLP) were used as illustrative examples of both groups. Chemotaxis was studied in human neutrophils and eosinophils. The inhibitory effects of nedocromil sodium and sodium cromoglycate were evaluated. 2. All chemotactic factors tested attracted neutrophils with the following rank order of activity: ZAS greater than PAF identical to FMLP identical to LTB4. Eosinophils were only mobilized by PAF, LTB4 and ZAS with the following rank order of activity: ZAS greater than PAF greater than LTB4. 3. Nedocromil sodium and sodium cromoglycate were equally active as the PAF antagonist BN 52021 in inhibiting the PAF-induced chemotaxis of neutrophils (IC50 approximately 10(-8) M). Both drugs were also equally active in inhibiting the chemotaxis of neutrophils induced by ZAS (IC50 approximately 10(-7)-10(-6) M), FMLP (IC50 approximately 10(-7) M) and LTB4 (IC50 approximately 10(-6) M). 4. Nedocromil sodium significantly inhibited the chemotaxis of eosinophils induced by PAF (IC50 approximately 10(-6) M) and LTB4 (IC50 approximately 10(-7) M). The inhibitory potency of BN 52021 was similar to that of nedocromil sodium on the PAF-induced chemotaxis of eosinophils. Sodium cromoglycate was incapable of eliciting significant inhibition of these chemotactic responses. However, sodium cromoglycate significantly inhibited the chemotaxis of eosinophils induced by ZAS (IC50 approximately 10(-7) M), whereas nedocromil sodium was ineffective.

[The probability of survival of patients with bronchial carcinoma]. / De overlevingskansen van patiënten met bronchuscarcinoom.

A study was made of the effect of the discontinuation of population screening for tuberculosis, on January 1, 1982, on the referral pattern and therapeutic results in patients with bronchial carcinoma. In 1981 only a few patients were referred because of lesions detected at the screening. Accordingly, discontinuing the screening had no demonstrable effect. Also, there was no difference between patients referred because of a fortuitous finding and those who consulted because of symptoms. Nevertheless the results, especially the 5-year survival rates, were far better for those whose tumours had favourable TNM classifications. This applied in particular if resection of parts of the lung was feasible and operation revealed no metastases in mediastinal lymph nodes and no infiltrating growth. For this group of patients the 5-year survival rate amounted to 59.4%. With only symptomatic treatment the rate was 16.7% and for patients with small-cell anaplastic carcinomas given chemotherapy, it was 5.6%.

Nedocromil sodium inhibits the A23187- and opsonized zymosan-induced leukotriene formation by human eosinophils but not by human neutrophils.

1. Inflammatory cells such as eosinophils and neutrophils are thought to contribute actively to the pathogenesis of asthma by the release of bronchoconstrictor mediators including leukotrienes. Previous studies have revealed the almost exclusive synthesis of leukotriene C4 (LTC4) by human eosinophils and of leukotriene B4 (LTB4), 20-OH-LTB4 and the non-enzymatically formed LTB4-isomers by neutrophils when stimulated in vitro with the calcium ionophore A23187 or opsonized zymosan (OZ). In this study we have investigated whether nedocromil sodium, a new anti-asthma drug, was capable of inhibiting A23187- and OZ-induced leukotriene formation by these cells. 2. Nedocromil sodium inhibited A23187- and OZ-induced LTC4 formation by eosinophils in a concentration-dependent manner (mean IC30 for A23187: 5.6 X 10(-5) M; mean IC30 for OZ: 6.3 X 10(-5) M), whereas it did not inhibit A23187- and OZ-induced LTB4 formation by neutrophils. 3. Extension of the preincubation time of the cells with the drug did not alter the observed inhibitory capacity. The optimal preincubation time was 5 min. 4. The in vitro inhibition of LTC4 formation by eosinophils by nedocromil sodium may be a valuable property of this drug in the treatment of asthma.

Inhibitory effects of nedocromil sodium on the in vitro induced migration and leukotriene formation of human granulocytes.

Inflammatory cells, such as neutrophils and eosinophils, are thought to actively contribute to the pathogenesis of asthma since they infiltrate into the lung tissue and may be activated locally to release bronchoconstrictor mediators. In this study we provide evidence that nedocromil sodium is capable of effectively inhibiting the platelet-activating factor (PAF) and zymosan-activated serum (ZAS)-induced chemotaxis of polymorphonuclear granulocytes (PMN) [IC50 approximately 1 nmol/L and 0.1 mumol/L respectively]. The same inhibitory potency was obtained with sodium cromoglycate. Thus, nedocromil sodium may effectively inhibit the mobilisation of inflammatory cells in the lung. Furthermore, nedocromil sodium is capable of inhibiting the formation of the bronchoconstrictor mediator leukotriene-C4 (LTC4) by eosinophils in a concentration-dependent way [IC30 for A23187: 5.6 10(-5) mol/L; IC30 for opsonised zymosan (OZ): 6.3 10(-5) mol/L], whereas this drug is not capable of inhibiting leukotriene-B4 (LTB4) formation by neutrophils. These findings indicate that nedocromil sodium inhibits the release of bronchoconstrictor mediators not only from mast cells but also from eosinophils.

Patients with chronic bronchitis differ in their mast cell subtypes as compared with normal subjects.

Mast cells were isolated by enzymatic digestion from lung tissue obtained from patients with chronic obstructive lung disease and from normal subjects. Two mast cell subtypes could be demonstrated in human lung tissue. Mast cell subtypes were differentiated in formalin-sensitive and formalin-insensitive mast cells. It appeared that compared with normal individuals, patients suffering from chronic bronchitis had increased numbers of mast cells of the formalin-sensitive type, whereas patients with emphysema had reduced numbers, but the same ratio, of both mast cell subtypes.

Mast cell subtypes from human lung tissue: their identification, separation, and functional characteristics.

The contribution of mast cell subtypes and their different mediators to the pathogenesis of chronic obstructive lung diseases (COLD) has not yet been established. In the present study, enzymatic digestion, centrifugal elutriation and Percoll gradient centrifugation were used to obtain two populations of mast cell subtypes from human lung tissue. Mast cell subtypes were challenged with anti-human IgE, propranolol, compound 48/80, or opsonized zymosan. Both subtypes were able to release histamine, but differed in the amount of the amine release. Only the formalin-sensitive and alcian blue-positive type (FS-AB) released histamine on challenge with opsonized zymosan. The same subtype was able to release leukotriene C4 (LTC4) after challenge with anti-human IgE. The other subtype, the formalin-insensitive and alcian blue-positive type (FI-AB), did not respond to opsonized zymosan and did not release LTC4 after challenge with anti-human IgE. Stimulation with propranolol or compound 48/80 did not release histamine from the FS-AB mast cells while the FI-AB mast cells released only about 10% of their histamine content upon challenge with these secretagogues.

The isolation of human lung mast cells by affinity chromatography.

A method of isolation has been developed to purify mast cells from human lung tissue. The purification steps are: (1) dispersion of human lung tissue in single-cell suspensions by enzymatic digestion, (2) partial purification by counterflow centrifugal elutriation, (3) Percoll gradient centrifugation, and (4) enrichment of the mast cells by affinity chromatography using anti-human IgE-Sepharose. Enzymatic dispersion yielded 0.6 +/- 0.2 x 10(6) mast cells per gram wet tissue with purities of 3.3 +/- 1.0% (mean +/- SEM n = 3). Elutriation and gradient centrifugation yielded 0.36 +/- 0.05 x 10(6) mast cells per gram lung tissue in fractions with purities of 30.8 +/- 10.7%. Enriched mast cell fractions were combined, and disposed of contaminating cells by affinity chromatography, thereby yielding 0.25 +/- 0.03 x 10(6) mast cells per gram lung tissue, and improving the purity to 75.3 +/- 8.3%. The purified mast cells were intact and vitality exceeded 95%. In this way from 1 g wet lung tissue 0.25 +/- 0.03 x 10(6) mast cells may be isolated with a mean recovery of 41.7 +/- 2.4% and a mean purity of 75.3 +/- 8.3%.

Terbutaline sustained-release treatment during one week 5 mg b.i.d. compared to one week placebo: terbutaline plasma levels, c-AMP plasma levels, lung function and tremor measurement.

In a group of ten patients with chronic asthmatic bronchitis, a good improvement of the lung function was achieved by a sustained-release terbutaline preparation (Bricanyl Retard) in a dosage of 5 mg twice daily. Tremor measurements and c-AMP plasma level showed a statistically significant increase. A good patient compliance was achieved with this dosage of terbutaline two times daily. The sustained-release preparation caused a rather constant and low terbutaline plasma level. We think that this relative low terbutaline plasma level is the most important reason for the lack of side effects.

Combination therapy of theophylline and terbutaline as sustained-release preparations in patients with asthmatic bronchitis.

In a group of ten patients with chronic asthmatic bronchitis, the effect of monotherapy with theophylline and terbutaline and the combination of both drugs, as sustained-release tablets, was compared to placebo, in a double-blind cross-over, triple dummy at random study. Lung function parameters, tremor measurements, c-AMP plasma levels, theophylline- and terbutaline-plasma levels were determinated. Monotherapy with theophylline and terbutaline in a relatively low dosage achieved a good lung function improvement without severe side effects, as compared to placebo. The combination of both drugs in the same dosage achieved a statistically significant enhancement of the lung function improvement, as compared to monotherapy, without an increase of side effects. An additional effect of theophylline and terbutaline on bronchodilation is demonstrated.