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Escherichia coli endotoxin (0.1 to 1000 micrograms/ml) stimulated the release of neutrophil chemotactic activity (P < 0.001) and induced bronchial epithelial cell (BEC) cytotoxicity assessed by lactate dehydrogenase release (P < 0.001). Endotoxin (100 micrograms/ml) inhibited BEC accumulation (P < 0.001). In the present study, we investigated the role of proteolytic activity of BECs per se in response to endotoxin. Several structurally and functionally different antiproteases, alpha 1 protease inhibitor, soybean trypsin inhibitor, two chloromethyl ketone derivatives (N-tosyl-L-lysine chloromethyl ketone and methoxysuccinyl-Ala-Ala-Pro-Val chloromethyl ketone), and L-658,758, a neutrophil elastase inhibitor, attenuated the release of neutrophil chemotactic activity and lactate dehydrogenase (P < 0.01). alpha 1-Protease inhibitor and N-tosyl-L-lysine chloromethyl ketone attenuated the inhibition of BEC accumulation by endotoxin (P < 0.001). The proteolytic enzyme activity measured by synthetic substrates revealed that endotoxin significantly augmented the serine proteolytic activity in the cell layers. Culture supernatant fluids and cell lysates of BECs in the presence of endotoxin solubilized 14C-labeled casein. These data suggest that responses of BECs to endotoxin may involve activation of cellular proteolytic activity.

Polymorphonuclear neutrophils (PMNs) accumulate within the airways during acute and chronic bronchitis and can adhere to bronchial epithelium. Substance P (SP), a neuropeptide released from the primary afferent nerve endings, has been shown to have a proinflammatory effect on PMNs. To test the hypothesis that SP could modulate PMN bronchial epithelial cell adherence, bovine bronchial epithelial cells (BBECs) were isolated and cultured, and the capacity of SP to modulate PMN-BBEC adherence was evaluated. SP interacted with BBECs to induce an increase in PMN adhesion (14.7 +/- 1.2% vs 5.3 +/- 0.7% adherence, p < 0.01). The effect of SP was both time- and dose-dependent with maximal responses at 6 h and 10(-10) M. The effect was reproduced by the carboxyl-terminal sequence of the molecule (SP 6-11). Importantly, pretreatment of the BBECs with the tachykinin SP receptor (NK1) antagonist, CP-96,345, significantly reduced the increase in adhesion induced by SP (p < 0.01). Furthermore, treatment of the BBECs with antibodies against CD11a (LFA-1), CD11b (MAC-1), or ICAM-1 significantly decreased SP-induced adherence (p < 0.01 all comparisons). Conversely, SP stimulation of PMN induced a dose-dependent, rapid (within 5 min) increase in adherence. This effect was also mediated by the carboxy end of the molecule and was decreased by CP-96,345, again suggesting that this effect was NK1 mediated. These data demonstrate the SP has the capacity for modulating PMN-BBEC interactions and suggest an important role for SP in modulating airway inflammation.

Laminin and Type IV collagen are both components of basement membrane. Antibodies to these two proteins, when injected into mice, were found to accumulate in all basement membranes examined, but at highest levels in kidney, liver, and spleen. An acute respiratory distress syndrome was noted shortly after injection. A transient segmental proliferative glomerulonephritis was observed both in the heterologous (early) and autologous (late) phase. The glomerular basement membrane of mice injected with anti-Type IV collagen antibodies was observed to be thickened and to contain dense deposits in the lamina rara externa and lamina rara interna.

For a study of the evolution of interstitial and intraalveolar fibrosis, ultrastructural and immunohistochemical observations were made of the lungs of 16 cynomolgous monkeys given 1 or 2 injections of 10 mg/kg of paraquat and sacrificed 2 days to 8 weeks later. At 2-3 days, alveolar epithelial cells were denuded in many areas, and fibronectin was conspicuous in alveolar spaces. At 1 week, fibroblasts and inflammatory cells were migrating through gaps in the denuded epithelial basement membranes; Type II cells were regenerating in some areas. At 3-4 weeks, alveoli developing intraalveolar fibrosis contained many myofibroblasts, collagen fibrils, and small elastic fibers; fibrotic alveolar walls were lined by metaplastic squamous cells and bronchiolar epithelial cells. Spiraling collagen fibrils were found in interstitium but not in alveolar spaces, which suggests that they were formed from breakdown of collagen. Newly formed intraalveolar collagen was mainly Type I. At 8 weeks, intraalveolar fibrosis had led to extensive remodeling, with new glandlike alveoli lined by Type II cells; alveoli without intraalveolar fibrosis had more normal architecture. Thus, intraalveolar fibrosis in paraquattreated lung is mediated by intraalveolar migration of interstitial cells, through gaps in the epithelial basement membranes, after epithelial injury. This is followed by connective tissue synthesis on the luminal side of the epithelial basement membrane, by differentiation of interstitial cells into myofibroblasts and smooth-muscle cells, by incorporation of areas of intraalveolar fibrosis into the interstitium, and by coalescence of alveolar walls. Intraalveolar fibrosis is more important than interstitial fibrosis in the structural remodeling that occurs in paraquattreated lung, because it results in obliteration of alveoli, coalescence of alveolar walls, and loss of functional alveolar-capillary units.