SUMMARYHuman lung cancer expresses cell membrane complement inhibitory proteins (CIP). We investigated whether human lung cancer cell lines also express cell-membrane CIP molecules and whether the biology of CIP molecules in these cell lines differs from that of CIP in normal human respiratory epithelium in culture. The cell lines ChaGo K-1 and NCI-H596 were compared with normal human nasal epithelium in primary cultures in respect to the level of cell membrane CIP expression of membrane cofactor protein (MCP; CD46), decay-accelerating factor (DAF; CD55) and CD59, in respect to the level of cell resistance to complement-mediated lysis, and in respect to the contribution of cell membrane CIP to cell resistance against complement-mediated lysis. We found, using flow cytometry, that both human lung cancer cell lines expressed MCP, DAF and CD59, as did normal nasal epithelial cells. However, normal cells showed a large subpopulation of low DAF-expressing cells (60% of all cells) and a smaller subpopulation of high DAF-expressing cells (40%), while the lung cancer cell lines showed only one cell population, of high DAF expression. In addition, both lung cancer cell lines expressed higher MCP levels, and NCI-H596 cells showed higher levels of CD59. Cell resistance to complement-mediated lysis of both lung cancer cell lines was much higher than that of normal cells. Fifty percent normal human serum, under the same concentrations of complement activators, induced lysis of less than a mean of 10% of lung cancer cells, while lysing up to a mean of 50% of nasal epithelial cells. Lung cancer cell resistance to complement was due to its ability to prevent significant activation of complement upon its cell membrane, as manifested by a failure of complement activators to increase cell membrane deposition of C3-related fragments. The exact mechanism for this resistance remains obscure. Unexpectedly, neutralizing antibodies, anti-MCP and anti-DAF were entirely ineffective and anti-CD59 was only slightly effective (18% mean cell lysis) in increasing the susceptibility of the lung cancer cell lines to complement, while the same antibodies were very effective in facilitating complementmediated lysis of the normal nasal epithelial cells (50% mean cell lysis with CD59 MoAb). On the other hand, detachment of DAF and CD59 by phosphatidylinositol-specific phospholipase C (PIPLC) from the lung cancer cell lines abrogated their resistance to lysis. We suggest that the biology of cell membrane CIP molecules in human lung cancer cell lines is different from that of CIP in normal respiratory epithelial cells. Human lung cancer cell lines are able to prevent significant complement activation upon its cell membrane and are therefore especially resistant to complement-mediated lysis. Complement resistance may serve this common and highly lethal human cancer as an escape mechanism from the body's immunosurveillance and prevent effective immunotherapy with tumour-specific MoAbs.Keywords CD46 CD55 CD59 complement inactivators nasal epithelium lung...
Background-The interrelationship between human airway epithelium and complement proteins may aVect airway defence, airway function, and airway epithelial integrity. A study was undertaken to determine (1) whether unstimulated human bronchial epithelium generates complement proteins and expresses cell membrane complement inhibitory proteins (CIP) and (2) (Thorax 2000;55:364-369)
Human lung cancers overexpress several cell-membrane complement inhibitory proteins (CIP). These complement inhibitory proteins are membrane cofactor protein (CD46), decay-accelerating factor (DAF; CD55), and CD59 (protectin). These cell-membrane proteins have a wide normal tissue distribution, are known to protect normal host cells from homologous complement-mediated lysis, and are thought to facilitate tumor escape from immunosurveillance. To study whether proinflammatory cytokines that are involved in cancer growth can modulate cell-membrane CIP expression in lung cancer cells, we studied the effect of interleukin (IL)-1alpha, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma on two human lung cancer cell lines. ChaGo K-1 and NCI-H596 cell lines, undifferentiated carcinoma and lung adenosquamous carcinoma, respectively, were stimulated with different cytokines, and the effects of incubation time and cytokine concentration on cell-membrane CIP expression were studied. Cell-membrane CIP expression was evaluated using flow cytometry and cytokine effect was calculated as percent change in mean fluorescence intensity of each CIP molecule from its untreated control. We found that DAF was the lung cancer cell-membrane CIP molecule that was the most responsive to cytokine stimulation. Maximal stimulatory effect was usually noted 72 h after a cytokine was introduced. In ChaGo K-1 and NCI-H596 lung cancer cell lines, IL-1alpha and TNF-alpha increased DAF expression. IL-1alpha (100 U/ml/72 h) increased DAF expression up to a maximal mean of 45 and 48%, respectively, in comparison with untreated cells. TNF-alpha (1, 000 U/ml/72 h) increased DAF expression up to a mean of 131 and 46%, respectively. IFN-gamma (1 U/ml/72 h) increased DAF expression in NCI-H596 cells up to a mean of 100%, but had a slight inhibitory effect on DAF expression in ChaGo K-1 cells, decreasing expression by a mean of 17% in comparison with untreated cells. We conclude that cell-membrane DAF expression in the studied human lung cancer cell lines is modulated by IL-1alpha, TNF-alpha, and IFN-gamma, and speculate that cytokine-mediated modulation of cell-membrane DAF in human lung cancer cells might affect lung cancer cell biology.
Complement in the respiratory tract protects the host from invading micoorganisms and other inhaled insults, but may damage normal tissue. Recently we reported that human respiratory epithelium from the nose to the alveoli expresses three cell-membrane regulators of complement activation: membrane cofactor protein (MCP, CD46), decay accelerating factor (DAF; CD55), and CD59. In this study we investigated whether two of these complement-regulatory proteins, DAF and CD59, protect human nasal epithelial cells from complement-mediated lysis. Treatment of nasal epithelial cells in suspension with 50% or 100% normal human serum (NHS) lysed small percentages of cells (8% and 16%, respectively). Addition of complement activators, rabbit serum antinasal epithelial cells (anti-NEC), or lipopolysaccharide (LPS) increased cell lysis in the presence of 50% NHS in a dose-dependent manner up to 50% and 35% lysis, respectively. Human serum deficient in C3 or C7 did not lyse nasal epithelial cells even in the presence of anti-NEC. To assay the contribution of DAF and CD59 to cell protection against lysis, nasal epithelial cells in suspension were treated with appropriate blocking antibodies. Both anti-DAF and anti-CD59 markedly increased the susceptibility of human nasal epithelial cells to lysis by complement. At 50% NHS, anti-DAF and anti-CD59 antibodies increased epithelial cell lysis from 8% to 24% and 67%, respectively. A similar pattern of response to complement was demonstrated by monolayers of substrate-anchored cultured cells. These results indicate that DAF and CD59 protect human nasal epithelial cells from complement-mediated lysis; however, intense activation of complement may overcome this protection, leading to cell death and tissue injury. We speculate that imbalance between complement regulation and complement activation in the human respiratory tract in disease may result in tissue injury and impaired tissue function.
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