Mee-Sun Tsai

Expression and Regulation of Macrophage Inflammatory Protein-2 Gene by Vanadium in Mouse Macrophages

Environmental and occupational exposure to vanadium (V) dusts results in inflammation mainly confined to the respiratory tract. Macrophages apparently play an important role in mediating the inflammation via the production of many chemokines. In the current study, we investigated whether vanadium can regulate the gene expression of a CXC chemokine macrophage inflammatory protein-2 (MIP-2), and to determine the molecular mechanisms controlling MIP-2 gene expression. A mouse macrophage cell line RAW 264.7 was treated with sodium metavanadate (NaVO3) at the dose of 0.5, 5, or 10 μg/ml V. Northern blot analysis showed that induction of MIP-2 mRNA expression was in a dose-dependent manner. To define the time course of the inflammatory response, RAW 264.7 cells were exposed to 5 μg/ml V, MIP-2 mRNA in macrophages increased markedly as early as 1 h after treatment, maximally induced at 4 h and reduced to 2-fold above control levels by 6 and 8 h. The protein levels of MIP-2 in conditioned media, measured by enzyme-linked immunosorbent assay (ELISA), was well correlated with the levels of MIP-2 mRNA following all of the treatments in the study. In addition, the increase in MIP-2 mRNA expression by vanadium was attenuated by co-treatment with the antioxidant N-acetylcysteine (NAC), at the doses of 10 and 20 mM, suggesting that the induction of MIP-2 mRNA is mediated via the generation of reactive oxygen species (ROS). To further investigate transcriptional regulation of the MIP-2 gene expression by vanadium, we performed RNA decay assay by measuring the half-life of MIP-2 mRNA. Co-treatment of macrophages with the transcriptional inhibitor actinomycin D at 5 μg/ml following exposure to 5 μg/ml V for 4 h revealed complete stabilization of vanadium-induced MIP-2 mRNA and no sign of mRNA degradation, at least, for 6 h, in comparison to the half-life of MIP-2 mRNA was approximately 2.5 h by bacterial lipopolysaccharide (LPS) treatment, supporting post-transcriptional stabilization as the predominant role of MIP-2 gene expression. In conclusion, these observations demonstrate that in vitro vanadium can induce MIP-2 mRNA expression, mediating, at least in part, via the production of ROS. In addition, the increase in MIP-2 mRNA level involves, most likely, post-transcriptional control via increased mRNA stability.

Serum hepatocyte growth factor levels in patients with inflammatory lung diseases.

HGF is a pulmotrophic factor in the regeneration of an injured lung. However, the physiological role of HGF in vivo remains largely unknown. We studied HGF in patients with inflammatory lung diseases to investigate the clinical significance of HGF and compared with C-reactive protein (CRP) in inflammatory lung diseases. Forty-seven patients with inflammatory lung diseases (16 tuberculosis, 18 pneumonia, and 13 chronic obstructive pulmonary disease (COPD)) were studied. Fifty normal, healthy individuals were analyzed as normal control subjects. Serum HGF levels were measured by enzyme-linked immunosorbent assays (ELISA). Serum CRP levels were also performed. The mean +/- SE numbers of serum HGF levels in the patients with inflammatory lung diseases (4.33 +/- 0.41 ng/ml) were significantly elevated when compared with those in normal control subjects (0.36 +/- 0.02 ng/ml) (p < 0.0001). Serum HGF levels in patients with COPD was significantly lower than those were with tuberculosis or pneumonia (p < 0.05). There was a significant correlation between serum HGF levels and CRP in inflammatory pulmonary diseases (r = 0.48, p = 0.00087). The significantly decreased serum HGF levels in patients with improved inflammatory lung diseases were also observed subsequently. Our results suggest that secreted HGF may play an important role in bronchial epithelium reconstruction during respiratory inflammation.

Cytokeratin fragment 19 (CYFRA 21-1) as a tumor marker in non-small cell lung cancer.

To evaluate the diagnostic value of the serum cytokeratin 19 fragment (CYFRA 21-1) in bronchogenic carcinoma, we investigated the sera of 138 patients (58 with benign pulmonary disease and 80 with non-small cell lung cancer (NSCLC)) using immunoradiometric assay. The mean (SD) value of serum CYFRA 21-1 in NSCLC (13.26 (16.54)) was significantly higher than in benign lung diseases (1.74 (1.55)) (p < 0.0001). Sensitivity for CYFRA 21-1 (using 3.5 ng/ml, a cut-off value corresponding to a 95% specificity for benign pulmonary disease) in NSCL was 62%. Positive CYFRA 21-1 levels were significantly higher in 75% of patients with squamous cell carcinoma (n = 36) than in 53% with other NSCLC (n = 44) (p < 0.05). CYFRA 21-1 levels were significantly different between squamous cell carcinoma (17.28 (19.94)) and the other NSCLC (9.96 (12.44)) (P < 0.05). Elevated CYFRA 21-1 levels in patients with stage III and IV disease (n = 64, 18.19 (26.51)) were significantly higher than in stage I and II (n = 16, 4.41 (5.76)) (p < 0.02). The positive rate of CYFRA 21-1 in tumor stage I and II was only 37%. Our results indicate that CYFRA 21-1 may be a useful tumor marker in NSCLC, especially in squamous cell carcinoma. However, CYFRA 21-1 cannot be used for the diagnosis of early stage disease of NSCLC. CYFRA 21-1 may also contribute to the monitoring of NSCLC.

Clinical study of Mycoplasma pneumoniae pneumonia

From 1982 to 1991, we experienced 76 patients with Mycoplasma pneumoniae pneumonia which were confirmed by serologic tests. There were 32 (42%) male and 44 (58%) female patients. One patient had underlying disease of diabetes mellitus while the other patients were in good health. The age ranged from 9 months old to 72 years old. All the patients complained of fever and coughing; 63% had dry cough and 37% had sputum production. Upper respiratory tract complaints such as rhinorrhea, sore throat, or earache were noted in 57% of the patients. Fifty-five percent of the patients had GI symptoms of anorexia, nausea, vomiting, or diarrhea. Other complaints included myalgia/arthralgia (29%), headache (30%), and general malaise (32%). Dyspnea (17%) and chest pain (20%) were occasional complaints. Seventy-one percent of the patients had WBC counts < 10000/cu mm and 29% > 10000/cu mm. The mean value of C-reactive protein (CRP) was 53.1 micrograms/ml, while 16% of the patients had a CRP value above 100 micrograms/ml. Thirty-one percent of the patients were noted to have a transient elevation of serum transaminase. Four different patterns of infiltration were seen in chest radiographic manifestation: 1) peribronchial and perivascular interstitial infiltrates (18.4%), 2) nonhomogeneous patchy consolidations (22.4%), 3) homogeneous acinar consolidations (27.6%), and 4) mixed interstitial and alveolar infiltrates (27.6%). Interstitial infiltration was more commonly seen in pediatric than adult patients (46% vs 20%). Other features of the radiologic manifestation were as follows: unilateral lesions in 80% of patients, single lobe lesions in 77%, lower lobe predominant in 69%, pleural effusion in 7%, and radiographic deterioration in 10%. Mycoplasmal pneumonia should be considered in the differential diagnosis of community-acquired pneumonias.

Asbestos related pleural plaques in retired boiler room workers.

Occupational disease is often underestimated and only a few formal reports have been published in Taiwan. This study reports of a group of workers with asbestos-induced-disease, pleural plaque in Taiwan. Pleural plaque is a marker of exposure to asbestos. The disease was found in chest radiographs of five boiler room workers in a sugar refining factory. The chest radiographs of 248 current workers in that plant were reviewed, and none of them was found to have pleural plaques. The storage of asbestos and the long-time use of mixed asbestos cement for insulation of the inner wall of the stove and pipes were found in the factory. The authors believe that the pleural plaques might be resulted from occupational exposure to asbestos. It is suggested that the use of asbestos should be prohibited, step by step, and regular follow-up of the workers with an asbestos exposure history is required.

Comparison of Pulmonary Tuberculosis in Younger and Elderly Patients

Pulmonary tuberculosis remains a significant clinical and public health problem in the elderly population. To describe age-related differences in disease manifestations, a comparison of the clinical features, predisposing factors, diagnostic approaches and radiographic findings in cases of pulmonary tuberculosis among 52 young and 62 elderly patients was performed. The elderly patients had a higher number of underlying disease than younger patients (p less than 0.05). Prior to admission, symptoms occurring with equal frequency in both younger and elderly patients included coughing, malaise, and weight loss. Elderly patients had significantly higher incidences of negative reactions to the PPD test (p less than 0.05). Radiographic findings revealed that upper lung field infiltrates were still common in both groups, but the elderly had more severe lung field involvement (two or more lobes affected), and more frequent pleural reactions than younger patients (p less than 0.05). Since there were non-specific clinical features, false negative skin test and complex radiographic manifestations, tuberculosis was frequently not suspected in the differential diagnosis, especially among elderly patients with multiple medical problems. We suggest that physicians need to have a high level of suspicion and awareness of varied manifestations for tuberculosis, especially elderly patients.

Application of Staining of Nucleolar Organizer Regions in Cytological Smears of the Bronchus

An argyrophil technique for the staining nucleolar organizer regions (AgNOR) was applied to cytological preparations obtained from bronchoscopic brushing materials. The number of AgNOR has been thought to be related to cellular activation. To differentiate malignant cells from non-malignant atypical cells, this study was carried out in 20 cases of adenocarcinoma of the lung (mean AgNOR: 18.34), 12 cases of pulmonary inflammatory diseases (mean AgNOR: 6.54) and 10 normal bronchial epithelial specimens for control (mean AgNOR: 2.07). On the basis of AgNOR number, we could differentiate between the three groups. The differences observed were statistically highly significant (p less than 0.0001). Moreover, the nucleolar organizer regions (NOR) in cancer cells were found the more irregularly distributed and more variable in sized than those in atypical and normal bronchial columnar cells. We suggest that the AgNOR technique will find increasing application as a complementary test in diagnostic cytopathology.

Flow Cytometric DNA Analysis of Pleural Effusions

A total of 71 cases of pleural effusion in patients with and without cancer were analyzed by conventional cytology and flow cytometry (FCM) in order to detect cells with an abnormal DNA content (aneuploidy). For cytologic examination, the samples were prepared using standard techniques. Sample for FCM analysis were centrifuged and exposed to hypotonic solution containing detergent and propidium iodide. Thirty-eight patients had pleural effusion due to benign disease, whilst 33 patients had primary lung cancer. All 38 patients with benign pleural effusions showed FCM diploidy. There were 17 aneuploidy (52%) and 16 diploidy (48%) in the 33 patients with lung cancer by FCM analysis. Four of these 33 effusions were cytologically negative, however, FCM showed aneuploidy in 2 of these 4 patients. Based on these results, FCM analysis combined with conventional cytopathology yielded 100% specificity, 94% sensitivity and 100% predictive value of positive result. There were no false-positive results but 2 false-negative results. These findings suggest that FCM is a rapid and useful technique in the analysis of pleural effusion and can be a very useful adjunct to conventional cytopathology.

Nucleolar Organizer Regions in Small Cell Carcinoma of Lung

Agyrophilic staining of nucleolar organizer regions (NOR) has been used to differentiate between cells of small cell carcinoma and lymphocytes in tissue specimens. We used cytologic smears which were previously Papanicolaou-destained to study the one-step agyrophilic staining technique for nucleolar organizer regions (AgNOR) in cell of small cell carcinoma and lymphocyte. The purposes of this study were to assess the feasibility and usefulness of AgNOR staining in diagnostic cytology and to try to set up a procedure that could be used on prestained smears for retrospective study. While the NOR of each lymphocyte appeared to be one round dot after AgNOR staining, the NOR of cell of small cell carcinoma showed dots, slightly variable in size and shape. The mean number of NOR was significantly higher (p < 0.01) in cells of small cell carcinoma (4.7 +/- 0.7) than in lymphocytes (1.4 +/- 0.4). In conclusion, AgNOR staining was demonstrated to be a useful method to differentiate between cells of small cell carcinoma and lymphocyte in Papanicolaou-destained smears.

Nucleolar organizer regions in smears of pleural effusion.

Agyrophil staining was applied to nucleolar organizer regions (NOR) to differentiate cells of adenocarcinoma and histiomesotheliosis in pleural effusion. The smears were either nuclearly unstained, but cytoplasmically counterstained by Papanicolaou method or Papanicolaou-destained before agyrophil staining of nucleolar organizer regions (AgNOR). The purposes of this study were to assess the feasibility and usefulness of AgNOR staining in diagnostic cytology and to try to set up a procedure that could be used on prestained smears for retrospective study. All smears showed good background and cellular outline. The distribution of NOR was either intranuclearly or in the nucleoplasm diffusely. In previously nuclearly unstained smears, NOR showed granular to powder-like appearance. The mean number of NOR in adenocarcinoma (38.4 +/- 12.5) was significantly higher than that in histiomesotheliosis (15.6 +/- 2.9). In Papanicolaou-destained smears, the NOR showed confluent dots. The mean number of NOR was much lower as compared to that of previously nuclearly unstained smears. Furthermore, the mean number of NOR in adenocarcinoma (3.6 +/- 1.4) showed no significant difference with that of histiomesotheliosis (2.7 +/- 0.8). In conclusion, AgNOR staining is one of the methods to differentiate benign from malignant cells, but not in Papanicolaou-destained smears.