Regulation of the Activity of Tissue Plasminogen Activator and Plasminogen Activator Inhibitor-1 by Zinc in Rat Primary Astrocytes

Tissue-type plasminogen activator (tPA) is a serine proteinase which plays important roles in functional and structural synaptic plasticity, neural migration, as well as excitotoxic injuries in several pathological situations including ischemic stroke, seizure and Alzheimer's disease (AD). It has been suggested that a divalent cation zinc also plays pathological roles in ischemia and seizure. Interestingly, it has been suggested that zinc and tPA may negatively regulate the activity or the level of each other by mechanism involving physical interaction between the two. In the present study, we investigated the effect of zinc in tPA activity and expression in rat primary astrocyte. Astrocytes were transiently exposed to 20~200micrometer Zn2+ for 2 h and then were recovered for 24 h. In the culture supernatants, zinc treatment concentration-dependently inhibited the activity of tPA which was determined by casein-plasminogen zymography. There was only marginal changes, if any, in the level of tPA mRNA and protein. On the other hand, the activity of an endogenous inhibitor of tPA, plasminogen activator inhibitor-1 (PAI-1) as well as its expression was increased by zinc treatment in a concentration-dependant manner. These results suggest that zinc-induced decrease in tPA activity was also, at least in part, regulated by indirect way by regulating the level of PAI-1. The decrease in tPA activity may be a part of body's plan to reduce excitotoxic neural injury in a condition of elevated zinc in the brain.

Cellular analysis and measurement of mucin in sputum of chronic airway disease / 결핵

BACKGROUND: In chronic airway disease, mucus secretion in increased, but extraction of mucin, which is the main component of mucus secretion, is a very complicated and limited in clinical use. Recently, monoclonal antibody for mucin was developed for possible clinical use. In this study, cellular analysis and quantification of respiratory mucin in sputum of patients with chronic airway diseases were performed. METHOD: Sputum was collected from patients with asthma(n=33), bronchiectasis(n=8) or chronic bronchitis(n=13) by spontaneous expectoration or by hypertonic saline induction, Collected sputums was treated by 0.1% dithiotreitol to dissociate the disulfide bond of the mucus and filtered through a nylon gauze. Total cell count, viability and differential count were measured. For detection of mucin, collected samples were treated with sodium dodoecyl sulfate polyacrylamide gel electrophoresis and then with monoclonal antibody(HMO2), as the primary antibody, and PAS stain. The amount of mucin was measured with ELISA by HMO2. Correlation with clinical information, cellular analysis, and amount of measured mucin were analyzed. RESULTS: Total cell counts of sputum were significantly increased in patients with bronchiectasis but viability remained the same. Eosinophils were significantly increased in patients with asthma, neutrophils in bronchiectasis chronic bronchitis, respectively (p<0.05). The results of Western blotting and PAS staining confirmed the presence of glycoproteins and matched? with mucin. The amounts of mucin measured by ELISA were not significantly different among the disease groups. Significant correlation was identified between the amount of mucin and viability(r=-0.482, p<0.05). CONCLUSION: Inflammatory cells in the sputum of those with chronic airway disease were different for each disease type. Measurement of mucin by ELISA via monoclonal antibodies may be a simple method for the evaluation of chronic airway disease.

Measurement of Mucin Amounts Using RTO3 in Patients with Chronic Airway Disease / 결핵

BACKGROUND: It has been anticipated that the amount and composition of mucin are changed in patients with chronic airway diseases. We evaluated whether RTO3 (mAb against rat tracheal mucins) could que ntify the amount of mucin from the airway in the patients with chronic airway diseases. METHODS AND RESULTS: 1) RTO3 was bound to high molecular weight of mucin based on Western blot in sputum and BALF from patients with chronic airway diseases. 2) The goblet cells and submucosal glands in main bronchus from human were observed by PAS stain. And immunohistochemical stain with RTO3 showed immunoreactivity on some goblet cells. 3) The amount of mucin was more increased in patients with chronic airway diseases compared to those in normal subjects. 4) In the exacerbation of asthmatics, mucin amounts were more increased than stable asthmatics. CONCLUSION: We suggested that secreted mucin in chronic airway diseases can be quantified by ELISA with RTO3.