title: Apoptosis of lung epithelial cells in response to TNF-alpha requires angiotensin II generation de novo.
abstract: Recent work from this laboratory demonstrated that apoptosis of pulmonary alveolar epithelial cells (AEC) in response to Fas requires angiotensin II (ANGII) generation de novo and binding to its receptor (Wang et al., 1999b, Am J Physiol Lung Cell Mol Physiol 277:L1245-L1250). These findings led us to hypothesize that a similar mechanism might be involved in the induction of AEC apoptosis by TNF-alpha. Apoptosis was detected by assessment of nuclear and chromatin morphology, increased activity of caspase 3, binding of annexin V, and by net cell loss inhibitable by the caspase inhibitor ZVAD-fmk. Purified human TNF-alpha induced dose-dependent apoptosis in primary type II pneumocytes isolated from rats or in the AEC-derived human lung carcinoma cell line A549. Apoptosis in response to TNF-alpha was inhibited in a dose-dependent manner by the nonselective ANGII receptor antagonist saralasin or by the nonthiol ACE inhibitor lisinopril; the inhibition of TNF-induced apoptosis was maximal at 50 microgram/ml saralasin (101% inhibition) and at 0.5 microgram/ml lisinopril (86% inhibition). In both cell culture models, purified TNF-alpha caused a significant increase in the mRNA for angiotensinogen (ANGEN), which was not expressed in unactivated cells. Transfection of primary cultures of rat AEC with antisense oligonucleotides against ANGEN mRNA inhibited the subsequent induction of TNF-stimulated apoptosis by 72% (P < 0.01). Exposure to TNF-alpha increased the concentration of ANGII in the serum-free extracellular medium by fivefold in A549 cell cultures and by 40-fold in primary AEC preparations; further, exposure to TNF-alpha for 40 h caused a net cell loss of 70%, which was completely abrogated by either the caspase inhibitor ZVAD-fmk, lisinopril, or saralasin. Apoptosis in response to TNF-alpha was also completely inhibited by neutralizing antibodies specific for ANGII (P < 0.01), but isotype-matched nonimmune immunoglobulins had no significant effect. These data indicate that the induction of AEC apoptosis by TNF-alpha requires a functional renin/angiotensin system (RAS) in the target cell. They also suggest that therapeutic control of AEC apoptosis in response to TNF-alpha is feasible through pharmacologic manipulation of the local RAS.
authors.affiliation: The Cardiovascular Institute, Michael Reese Hospital and Medical Center, Chicago, Illinois, USA.
authors.forename: R
authors.initials: R
authors.lastname: Wang
authors.forename: G
authors.initials: G
authors.lastname: Alam
authors.forename: A
authors.initials: A
authors.lastname: Zagariya
authors.forename: C
authors.initials: C
authors.lastname: Gidea
authors.forename: H
authors.initials: H
authors.lastname: Pinillos
authors.forename: O
authors.initials: O
authors.lastname: Lalude
authors.forename: G
authors.initials: G
authors.lastname: Choudhary
authors.forename: D
authors.initials: D
authors.lastname: Oezatalay
authors.forename: B D
authors.initials: BD
authors.lastname: Uhal
mesh_terms: D000804:Angiotensin II / Q000096:biosynthesis* / Q000378:metabolism; D057911:Angiotensin Receptor Antagonists; D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D000808:Angiotensinogen / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000502:physiology; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D017706:Lisinopril / Q000494:pharmacology; D008168:Lung / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D011499:Protein Processing, Post-Translational / Q000502:physiology; D012333:RNA, Messenger / Q000378:metabolism; D011945:Receptors, Angiotensin / Q000378:metabolism; D012504:Saralasin / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D057911:Angiotensin Receptor Antagonists; D000806:Angiotensin-Converting Enzyme Inhibitors; D012333:RNA, Messenger; D011945:Receptors, Angiotensin; D014409:Tumor Necrosis Factor-alpha; D000808:Angiotensinogen; D000804:Angiotensin II; D017706:Lisinopril; D012504:Saralasin