[Serum levels of matrix metalloproteinase 9 and 19 in the patients with pneumoconiosis].

OBJECTIVE: To measure peripheral serum levels of matrix metalloproteinase 9 (MMP9) and matrix metalloproteinase 19 (MMP19) in patients with pneumoconiosis, and to investigate their feasibility as potential biomarkers for pneumoconiosis. METHODS: Ninety-eight male patients with pneumoconiosis (49 patients in phase I, 36 patients in phase II, and 13 patients in phase III) were enrolled as subjects, which included 41 patients with silicosis and 57 patients with coal workers' pneumoconiosis. Ninety-eight healthy male physical examinees were used as controls. A fasting blood sample (3 ml) was collected from the peripheral venous blood of each patient or control, and the serum was separated from the blood sample. The expression levels of MMP9 and MMP19 in serum were measured by enzyme-linked immunosorbent assay. RESULTS: Serum levels of MMP9 and MMP19 in patients with silicosis or coal workers' pneumoconiosis were significantly lower than those in the control group (P < 0.05). Serum levels of MMP19 in patients with silicosis were significantly higher than those in patients with coal workers' pneumoconiosis (P < 0.05). Serum levels of MMP19 in patients exposed to dust for less than 7 years were significantly higher than those in patients exposed to dust for more than 20 years (P < 0.05). There were no significant differences in serum levels of MMP9 and MMP19 between patients with different levels of pulmonary function impairment (P > 0.05). Serum expression levels of MMP9 and MMP19 were positively correlated with each other in both patients with pneumoconiosis and those in the control group (P < 0.05). The serum expression level of MMP9 was negatively correlated with the stage of pneumoconiosis (P < 0.05). CONCLUSION: Serum MMP9 and MMP19 may be used as potential biomarkers for pneumoconiosis.

[Evaluation of matrix metalloproteinases (pro-MMP-1, MMP-2,8) and their inhibitor (TIMP-1) contents in patients with occupational lung diseases].

Studies of matrix metalloproteinases in patients with occupational bronchopulmonary diseases and in individuals exposed to asbestos dust revealed hyperactivated protease system--lower level of MMP-1 proenzyme and increased production of TIMP-1 (metalloproteinases inhibitor)--in all the examinees groups. Patients with pneumoconiosis and occupational dust bronchitis demonstrated increased neutrophilic elastase that is activator of metalloproteinases inducing sclerotic changes and pulmonary fibrosis.

Antioxidant response at early stages and low grades of simple coal worker's pneumoconiosis diagnosed by high resolution computed tomography.

In miners exposed to coal dusts, coal worker's pneumoconiosis (CWP) can occur. The purpose of the present study is to better understand the relations between coal dust exposure and activities of blood plasma antioxidant enzymes, namely, superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and lipid peroxidation end product malondialdehyde (MDA) concentration in coal workers with early and low grade simple CWP diagnosed by high resolution computed tomography (HRCT). Forty-three coal workers who had profusions of 0/1-2/2 according to ILO 1980 chest X-ray (CXR) classification, 43 coal workers without CWP (control group 1) and 44 healthy subjects (control group 2) who were randomly selected from the population register or recruited from the hospital staff were enrolled. Coal workers were reevaluated by HRCT (Hosoda-Shida classification) due to its higher sensitivity than standard CXR. Then, blood plasma SOD and GSH-Px activities and MDA plasma levels were measured. CWP was found positive in 46 of 89 coal workers by HRCT evaluation. Profusion 0 (P0, CWP not present), profusion 1 (P1, early CWP) and profusion 2 (P2, low grade CWP) were found in 43, 23 and 19 of patients found to have CWP by HRCT, respectively. We had no worker with profusion 3 (P3). Complicated CWP was shown in four of 46 patients and thesecases were excluded as the study was restricted to early and low-grade pneumoconiosis. In respect to the plasma levels of MDA and plasma activities of SOD and GSH-Px, statistically significant differences were found between CWP cases and control groups (p < 0.01, p < 0.01, p < 0.001 respectively). Statistical differences were also obtained for the plasma activities of SOD and GSH-Px and levels of MDA in relation to HRCT profusions (p < 0.05). In conclusion, these findings suggest an oxidative stress due to increased free radicals and reactive oxygen metabolite production in early stages and low grades of simple CWP diagnosed by HRCT.

The presence of promatrix metalloproteinase-3 and its relation with different categories of coal workers' pneumoconiosis.

Extracellular matrix formation (ECM) and remodeling are critical events related to the pathogenesis of pulmonary fibrosis. Matrix metalloproteinases play an essential role in degrading and remodeling the ECM. In this study, we tried to show the presence and correlation of promatrix metalloproteinase-3 (proMMP-3) (the inactive form of metalloproteinase-3) levels in coal workers' pneumoconiosis (CWP) with different categories. The study population consisted of 44 coal miners with CWP (pos CWP). Coal miners without CWP (neg CWP, n = 24) and non-underground personnel (controls, n = 17) were taken as controls. All coal miners were stable and had no systemic infection or disease. Standard posterio-anterior chest radiographs and pulmonary function tests were performed to exclude any diseases other than CWP. Serum proMMP-3 was analysed using the sandwich enzyme-linked immunosorbent assay according to the manufacturer's instructions (The Binding Site, Birmingham, UK). Mean proMMP-3 values of the all three groups were compared and a significant statistical difference obtained (p < 0.001). In addition, a statistically significant difference was found between categories of the disease and proMMP-3 values (p < 0.05). The effects of age, exposure duration and cigarette smoking on proMMP-3 values in coal miners with CWP were investigated. There were no correlations between age, smoking and proMMP-3 values. However, a positive correlation was found between exposure duration and proMMP-3 values (r = 0.447, p = 0.008). In conclusion, proMMP-3 (prostromelysin 1) may play an essential role in degrading and remodeling the ECM in workers with pneumoconiosis. ProMMP-3 may also reflect the stage of pneumoconiosis disease.

[Impaired balance of the lung proteolytic system in patients with pneumoconiosis and chronic dust-induced bronchitis]. / Narushenie ravnovesiia sistemy proteoliza legkikh u bol'nykh pnevmokoniozom i khronicheskim pylevym bronkhitom.

The activity of neutrophilic elastase and the level of its major inhibitor--an alpha 1-inhibitor of proteinases were studied to evaluate the protease-antiprotease system in the bronchoalveolar lavage fluid (BALF) of patients with chronic dust-induced bronchitis (CDB) and pneumoconiosis (PC). It was found that CDB, as compared with PC, was characterized by a higher elastase activity in the BALF (70%) and that there were a larger number of patients with elastase activity (40%), which correlated with the detection rate of emphysema. Free elastase activity and relative proteinase alpha 1-inhibitor deficiency suggest that the BALF protease-antiprotease system is impaired in patients with CDB and PC, which is more pronounced in patients with CDB.

Relations between occupational exposure to coal mine dusts, erythrocyte catalase and Cu++/Zn++ superoxide dismutase activities, and the severity of coal workers' pneumoconiosis.

OBJECTIVES: To better understand the relations between occupational exposure, blood antioxidant enzyme activities, total plasma antioxidant concentration, and the severity of coal workers' pneumoconiosis (CWP). METHODS: Blood samples were obtained from miners without CWP exposed to low dust concentrations for > or = 4 years at the time of the study (n = 105), or exposed to high dust concentrations for > or = 14 years at the time of the study (n = 58), and from retired miners with CWP (n = 19). Miners without CWP were classified into three subgroups according to their estimated cumulative exposure to dust. Chest x ray films were obtained for each miner. Miners were classified in five subgroups according to their International Labour Organisation (ILO) profusion grades. Univariate tests were completed by multiple linear regression analyses. RESULTS: The estimated cumulative exposure to dust was strongly positively related to erythrocyte catalase activity and strongly negatively related to Cu++/Zn++ SOD activity only in miners exposed to high dust concentrations for > or = 14 years at the time of the study (F tests p = 0.006 and p = 0.004 respectively). Moreover, catalase activity was strongly related to the severity of CWP expressed as five subgroups of ILO profusion grades (F test p = 0.003); the greatest difference in the mean values was found between the group of 1/1 to 1/2 ILO profusion grades and the group of 2/1 to 3/3 ILO profusion grades. CONCLUSION: These results are in good agreement with the hypothesis that production of reactive oxygen species may be an important event in the exposure to coal mine dusts and the severity of CWP. Erythrocyte catalase and Cu++/Zn++ SOD activities are more closely related to recent exposure to high dust concentrations than to cumulative exposure, and could be considered as biological markers of exposure rather than as markers of early adverse biological effect.

Comparison of inducible nitric oxide synthase gene expression and lung inflammation following intratracheal instillation of silica, coal, carbonyl iron, or titanium dioxide in rats.

The pulmonary toxicity of the respirable dusts silica, coal, carbonyl iron, and titanium dioxide on alveolar macrophage (AM) and neutrophil (PMN) inducible nitric oxide synthase (iNOS) gene expression and nitric oxide (NO) production was investigated. Rats were intratracheally instilled with 5 mg/100 g body weight of silica, coal, carbonyl iron, or titanium dioxide. The dust particles averaged less than 5 microns in diameter. Bronchoalveolar lavage was performed 24 h later. Bronchoalveolar lavage cell (BALC) differentials, iNOS gene expression and NO production by BALC (measured indirectly as NO-dependent chemiluminescence), and lavageable lung protein levels were measured. Analyzed on an equal mass basis, silica, coal, and titanium dioxide dusts increased the production of iNOS-dependent NO by AM. Silica and titanium dioxide both increased the levels of iNOS mRNA while carbonyl iron and coal did not. Each dust caused an increase in PMN, indicating an inflammatory response. Carbonyl iron and titanium dioxide decreased the numbers of AM. Levels of acellular lavageable lung protein were increased by silica, carbonyl iron, and titanium dioxide. When exposure was normalized for an equal number of particles, the pneumotoxic dusts, silica and coal, caused more inflammation and NO production than the nuisance dusts, carbonyl iron and titanium dioxide. Therefore, it appears that particle number is a more appropriate metric of exposure than mass when comparing the relative pathogenicity of dusts of different sizes. Furthermore, since the potency of these dusts (on a particle number basis) to increase iNOS gene expression reflects their inflammatory and pathogenic potential, it is proposed that NO may contribute to the early inflammatory damage observed in the lung following dust exposure.

Characterization of a soluble form of neutral endopeptidase-24.11 (EC 3.4.24.11) in human serum: enhancement of its activity in serum of underground miners exposed to coal dust particles.

A previous epidemiological study has reported the elevation of a serum metalloendopeptidase activity for underground coalminers exposed to chronic inhalation of coal mine dust particles. In this work, we have unambiguously characterized this activity as neutral endopeptidase (EC 3.4.24.11) using five different criteria. The apparent molecular weight of 100,000 g mol-1 calculated for the serum peptidase using Western blots or direct binding of the neutral endopeptidase 24.11 inhibitor [125I]-RB104 to the enzyme in acrylamide gels, suggests that the soluble form of this ectoenzyme is not generated by a post-translational cleavage of the membrane-bound form, as is the case for the closely related ectoenzyme, angiotensin-converting enzyme. The circulating endopeptidase very likely results from a shedding process. The increase in serum neutral endopeptidase 24.11 activity of underground miners compared with surface miners (5.7-fold), P < 0.01) is not correlated with systemic inflammation parameters, but seems to reflect the chronic pulmonary inflammatory state induced by coalmine dust exposure, and so may be a marker of lung injury.

Increased expression of manganese-containing superoxide dismutase in rat lungs after inhalation of inflammatory and fibrogenic minerals.

Steady-state mRNA levels and immunoreactive protein for manganese-containing superoxide dismutase (MnSOD) were assayed in rat lungs after subchronic inhalation of the fibrogenic silicon dioxide, cristobalite, or preparations of titanium dioxide (TiO2) of different inflammatory and fibrogenic potential. Total and differential cell counts recoverable by bronchoalveolar lavage (BAL) were also measured to ascertain whether induction of certain antioxidant enzymes (AOE) correlated with inflammatory responses. Inhalation of cristobalite and ultra-fine TiO2, a particle causing pulmonary inflammation and fibrosis, caused dramatic increases in MnSOD mRNA levels in rat lung which correlated with increases in MnSOD immunoreactive protein. Increases in gene expression of other AOE [catalase, glutathione peroxidase (GPX), copper-zinc containing superoxide dismutase (CuZnSOD)] were less striking and did not correlate precisely with inflammatory potential of minerals. Inflammatory changes in BAL correlated directly with steady-state MnSOD mRNA levels in lung. Inhalation of TiO2-F, a noninflammatory, nonfibrogenic mineral, failed to induce MnSOD or mRNAs for other AOE. Our data suggest that particles causing inflammation and pulmonary fibrosis increase expression of AOE in lung, most notably MnSOD. Thus, elevations of MnSOD mRNA levels in lung or BAL may be predictive of lung disease.

Decreased glutathione content and glutathione S-transferase activity in red blood cells of coal miners with early stages of pneumoconiosis.

Blood samples of miners heavily exposed to coal dust were examined for changes in glutathione S-transferase (GST) activity. Decreased GST activity was found in red blood cells of subjects with early stages of coal workers' pneumoconiosis (International Labour Office classification 0/1-1/2) when compared with control miners. At further progression of coal workers' pneumoconiosis (> or = 2/1), the activity of GST was not different from controls. In the same group with moderate coal workers' pneumoconiosis a decrease in GSH in red blood cells occurred. Decreases in GST activity in early stages of coal workers' pneumoconiosis, as well as the decreases in glutathione peroxidase (GPx) activity and in GSH concentrations reported earlier, may originate from damage caused by reactive oxygen species. These changes might imply an impairment of the detoxification capacity for electrophilic and oxidative compounds during this stage of the disease.

Decreased sialidase activity in alveolar macrophages of guinea pigs exposed to coal mine dust.

The origin of immune dysfunctions that are observed in pneumoconiotic miners still remains unknown. There is evidence that the carbohydrate moiety of membrane glycoconjugates is of primary importance in many functions of immunocompetent cells. The glycosylation, and especially the sialylation level of membrane components of various lymphocyte and macrophage subsets, vary depending on the state of cellular differentiation and activation. Sialidases, which may regulate the amount of sialic acids exposed on the cell membrane, can thus be considered as immunoregulatory enzymes. In this report, the sialidase activity has been measured in alveolar macrophages (AM) and in cell-free bronchoalveolar lavage fluid (BALF) from guinea pigs exposed for 4 months to coal mine dust at a concentration of 300 mg/m3. The samples were collected by bronchoalveolar lavage 2 months after cessation of exposure. The sialidase activity in the cell-free fluid and in the purified alveolar macrophages showed a 10-fold decrease (p less than 0.001). Kinetic parameters of the enzyme such as Km and optimum pH did not change. This changed activity was specific for sialidase, as two other lysosomal glycosidases, beta-galactosidase and N-acetylglucosaminidase, showed unchanged activities. These results suggest the possibility that, by inducing a decreased sialidase activity, exposure to coal mine dust may lead to a modified expression of AM membrane-associated sialic acids giving rise to altered immune functions (i. e., phagocytosis, antigen processing, response to cytokines, etc.).

Serum angiotensin-converting enzyme is elevated in association with underground coal mining.

Serum angiotensin-converting enzyme activity (SACE) and lysozyme activity were measured in a group of 40 underground coal miners and two control groups, 20 subjects with sarcoidosis and 15 normal non-dust-exposed volunteers. The miners were grouped first according to whether they had recent exposure (still actively mining or retired three years or less prior to measurement) or temporally more distant exposure (retired more than three years prior to measurement). Secondly, they were grouped as to whether or not they had coal workers' pneumoconiosis (CWP). The subjects with sarcoidosis were grouped according to disease activity. As expected, the subjects with active sarcoidosis had elevated SACE activity compared with normal subjects. The coal miners as a group did not have elevation of their SACE activity. However, the coal miners with recent exposure had elevated SACE activity (57.1 +/- 3.9 U/ml) compared with normal controls (43.8 +/- 1.5 U/ml, p = 0.007). The SACE activity in miners without recent exposure was not elevated (39.8 +/- 1.3 U/ml) compared with the normal controls. No increase in SACE activity was found when the miners were grouped according to the presence or absence of CWP. In contrast, the miners' serum lysozyme activity was not elevated. Since alveolar macrophages are a potential source of SACE, elevation of SACE activity in underground coal miners may reflect alveolar macrophage activation caused by increased pulmonary mixed coal mine dust burden. Furthermore, since both SACE and serum lysozyme are elevated in association with silicosis, these findings may confirm that the macrophage responses to inhaled silica and coal dust differ.

[Changes in adenosine deaminase and antioxidative enzyme activity in patients with lung diseases of dust etiology]. / Izmenenie aktivnosti adenozindezaminazy i antiokislitel'nykh fermentov u bol'nykh s zabolevaniiami legkikh "pylevoi" étiologii.

Activity of purine metabolism enzyme, adenosine deaminase (ADA) in lymphocytes and erythrocytes from patients with pneumoconiosis and chronic bronchitis was determined. ADA activity decrease was found in lymphocytes and erythrocytes for both decreases. The observed decrease of ADA activity indicates that after the long term work in mineral dust industry damage of purine metabolism takes place, that is similar to the disturbance observed under secondary immunodeficiency. Changes in activity of superoxide dismutase and catalase show some decrease of functional activity of anti-radical protection. Therefore the results obtained can be used for the selection of a group with highest risk of predisposition to these diseases.

Pulmonary function in Pi M and MZ grainworkers.

Twenty-eight men with the Pi MZ phenotype who have been employed in the Saskatchewan country grain elevators and thus regularly exposed to high levels of grain dust, were case matched for age, years of employment, employment status, smoking status, and smoking history with grainworkers of type Pi M. Individuals answered a questionnaire, had a chest roentgenogram, skin tests, and performed a battery of pulmonary function tests. There were no differences between the two groups in prevalence of symptoms or atopy. Although not statistically significant, the MZ group had three times as many individuals with abnormal roentgenograms suggestive of COPD as the M group. The Pi MZ grainworkers had consistently poorer mean results for the pulmonary function tests with significantly lower mean values for FEV1, FEV1/FVC, MMFR, and Vmax50, leading us to suggest that Pi MZ individuals may be at higher risk of COPD than Pi M individuals, but only in the presence of other risk factors such as grain dust exposure.

Serum angiotensin-converting enzyme and lysosomal enzymes in tobacco workers.

The activities of angiotensin-converting enzyme (ACE) in people with extrinsic allergic alveolitis (EAA) have been both increased and decreased. These observations suggest that pulmonary macrophages or endothelial cells participate in the disease process. Exposure to molds in the tobacco industry has recently been suspected to be associated with chronic extrinsic allergic alveolitis. In the present study, we analyzed the serum activities of ACE and two lysosomal enzymes, beta-N-acetylglycosaminidase (NAG) and beta-glucuronidase (GLU), among 57 tobacco workers. The tobacco workers not exposed had serum ACE levels similar to those of the reference population workers not occupationally exposed to dust (N = 127). The tobacco workers' serum levels of NAG (16.0 +/- 2.0 units/L and GLU (2.4 +/- 0.7 units/L) were higher than among the referents (NAG, 9.1 +/- 2.0 units/L; GLU, 1.0 +/- 0.6 units/L; p less than 0.01). Fifteen tobacco workers with respiratory symptoms compatible with pulmonary diseases caused by organic dust had a trend toward increased ACE, NAG, and GLU levels. The mean level of ACE in serum was higher among the workers with (25.8 +/- 4.5 units/L) than among those without pulmonary fibrosis (20.7 +/- 7.5 units/L; p less than 0.025). The mean ACE level was also higher among workers with the highest exposure to molds (24.6 +/- 7.1 units/L) compared to those with the mildest exposure (18.3 +/- 5.7 units/L; p less than 0.05). Tobacco workers with or without antibodies against one or more microbes had similar mean levels of ACE, NAG, and GLU. All of these findings indicate that raw tobacco dust and its contaminants may cause allergic or toxic reactions or both, reflected by the serum levels of ACE, NAG, and GLU.

[Diagnostic, prognostic and developmental value of serum angiotensin converting enzyme in sarcoidosis]. / Intérêt diagnostigue, pronostigue et évolutif de l'enzyme de conversion de l'angiotensine sérique dans la sarcoïdose.

Two hundred ninety three patients with mediastinal and pulmonary sarcoidosis were assayed one or more than one time for seric angiotensin converting enzyme (SACE) using the method of Cushman and Cheung (substrate hippuryl-histidyl-leucin). Seventy one normal subjects and 163 patients with various broncho-pulmonary diseases excluding sarcoidosis were used as control. SACE is elevated in 67.2 p. 100 of the patients with sarcoidosis and reflects the intra and extrathoracic extend of the granuloma. Elevated levels of SACE in pneumoconiosis diminishes the diagnostic value of this test (as well as the presence of a normal SACE level in some sarcoid patients). There is no correlation between SACE and the percentage of lymphocytes in bronchoalveolar lavage fluid. SACE returns to normal in cases with spontaneous radiological improvement, and reaches more elevated levels in cases with worsening. An initial low level of SACE is usually a sign of a future good evolution. An initial high level is an argument for starting on steroid treatment. Repeated dosages of SACE are useful for monitoring the steroid posology at the end of the treatment and for deciding to stop it. Persistence of low levels allows to stop the treatment. Re-elevation of SACE may correspond to a radiological and clinical relapse or to an isolated and resolvent rebound.