LL-37-induced caspase-independent apoptosis is associated with plasma membrane permeabilization in human osteoblast-like cells

• Published in: Peptides (New York, N.Y. : 1980) Vol. 135; p. 170432
• Main Authors: Bankell, Elisabeth, Dahl, Sara, Gidlöf, Olof, Svensson, Daniel, Nilsson, Bengt-Olof
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.01.2021
• Subjects: Antimicrobial peptides (AMP); Apoptosis; Basic Medicine; Cathelicidin; Cell and Molecular Biology; Cell- och molekylärbiologi; Cytotoxicity; Innate immunity; Medical and Health Sciences; Medicin och hälsovetenskap; Medicinska och farmaceutiska grundvetenskaper; Cathelicidin; Cytotoxicity; Innate immunity; Antimicrobial peptides (AMP); Apoptosis
• ISSN: 0196-9781; 1873-5169; 1873-5169
• DOI: 10.1016/j.peptides.2020.170432

•Treatment with LL-37 reduces MG63 cell viability and enhances LDH release.•LL-37 reduces cell number and causes cellular shrinkage typical for apoptotic cells.•LL-37 induces apoptosis assessed by TUNEL and Annexin V flow cytometry.•LL-37 does not promote either caspase-3 or PARP cleavage.•LL-37 triggers caspase-independent apoptosis and membrane permeabilization. The host defense peptide LL-37 is active against both gram-positive and gram-negative bacteria, but it has also been shown to reduce human host cell viability. However, the mechanisms behind LL-37-induced human host cell cytotoxicity are not yet fully understood. Here, we assess if LL-37-evoked attenuation of human osteoblast-like MG63 cell viability is associated with apoptosis, and if the underlying mechanism may involve LL-37-induced plasma membrane permeabilization. MG63 cell viability and plasma membrane permeabilization were investigated by using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method and by measuring lactate dehydrogenase (LDH) release, respectively. Apoptosis was assessed by the terminal deoxynucleotidyl dUTP nick end labeling (TUNEL) assay and Annexin V flow cytometry, and caspase-3 and poly (ADP-ribose) polymerase (PARP) cleavage were determined by Western blot. LL-37 (4 and 10 μM) reduced both cell number and cell viability, and these effects were associated with a pro-apoptotic effect demonstrated by positive TUNEL staining and Annexin V flow cytometry. LL-37-induced apoptosis was not coupled to either caspase-3 or PARP cleavage, suggesting that LL-37 causes caspase-independent apoptosis in MG63 cells. Both LL-37 and the well-known plasma membrane permeabilizer Triton X-100 reduced cell viability and stimulated LDH release. Triton X-100-treated cells showed positive TUNEL staining, and the detergent accumulated cells in late apoptosis/necrosis. Similar to LL-37, Triton X-100 caused no PARP cleavage. We conclude that LL-37 promotes caspase-independent apoptosis, and that this effect seems coupled to plasma membrane permeabilization in human MG63 cells.