The role of apoptosis in antibody-dependent cell-mediated cytotoxicity against monolayers of human squamous cell carcinoma of the head and neck targets

Myung Whun Sung, Shigeki Nagashima, Jonas T. Johnson, G. A.M.S. Van Dongen, Theresa L. Whiteside

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Antibody-dependent cellular cytotoxicity (ADCC) against squamous cell carcinoma of the head and neck (SCCHN) targets in the presence of human/mouse chimeric monoclonal antibodies (cMAbs), SF-25 and 323/A3, is mediated by natural killer (NK) cells. In 4-hr 51Cr-release assays with SSCHN targets in suspension, ADCC was always significantly better (P < 0.01) than that measured in parallel with the same target cells in monolayers. No differences were observed in the level of expression of the relevant antigens recognized by cMAbs on these targets. To better explain the difference, 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) monolayer and [3H]thymidine-release assays were used. Cytostasis and cell death measured in monolayer MTT assays and DNA fragmentation measured in [3H]thymidine-release assays were significantly higher (P = 0.028) than cytotoxicity determined using 51Cr-labeled SCCHN monolayers. Cell death observed in monolayer MTT assays was blocked by pretreating SCCHN targets with cycloheximide or actinomycin-D or by paraformaldehyde fixation of effector cells. The presence of apoptotic cells in monolayers co-incubated with effector cells was demonstrated in situ by labeling fragmented ends of DNA with fluorescein-conjugated dUTP and terminal deoxynucleotidyl transferase and also by flow cytometry of target cells obtained from such monolayers. Our results indicate that NK cells preferentially utilize membrane lysis (necrosis) in ADCC with tumor cell targets in single-cell suspensions. However, necrosis is not efficient in monolayers. In the presence of cMAbs, apoptosis is the primary mechanism of NK cell-mediated killing in monolayers of SCCHN targets, which were found to express receptors for tumor necrosis factor and fas ligand.

Original languageEnglish
Pages (from-to)20-29
Number of pages10
JournalCellular Immunology
Volume171
Issue number1
DOIs
Publication statusPublished - 10 Jul 1996

Cite this

@article{1f705562645e4f9db59d336b260fadff,
title = "The role of apoptosis in antibody-dependent cell-mediated cytotoxicity against monolayers of human squamous cell carcinoma of the head and neck targets",
abstract = "Antibody-dependent cellular cytotoxicity (ADCC) against squamous cell carcinoma of the head and neck (SCCHN) targets in the presence of human/mouse chimeric monoclonal antibodies (cMAbs), SF-25 and 323/A3, is mediated by natural killer (NK) cells. In 4-hr 51Cr-release assays with SSCHN targets in suspension, ADCC was always significantly better (P < 0.01) than that measured in parallel with the same target cells in monolayers. No differences were observed in the level of expression of the relevant antigens recognized by cMAbs on these targets. To better explain the difference, 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) monolayer and [3H]thymidine-release assays were used. Cytostasis and cell death measured in monolayer MTT assays and DNA fragmentation measured in [3H]thymidine-release assays were significantly higher (P = 0.028) than cytotoxicity determined using 51Cr-labeled SCCHN monolayers. Cell death observed in monolayer MTT assays was blocked by pretreating SCCHN targets with cycloheximide or actinomycin-D or by paraformaldehyde fixation of effector cells. The presence of apoptotic cells in monolayers co-incubated with effector cells was demonstrated in situ by labeling fragmented ends of DNA with fluorescein-conjugated dUTP and terminal deoxynucleotidyl transferase and also by flow cytometry of target cells obtained from such monolayers. Our results indicate that NK cells preferentially utilize membrane lysis (necrosis) in ADCC with tumor cell targets in single-cell suspensions. However, necrosis is not efficient in monolayers. In the presence of cMAbs, apoptosis is the primary mechanism of NK cell-mediated killing in monolayers of SCCHN targets, which were found to express receptors for tumor necrosis factor and fas ligand.",
author = "Sung, {Myung Whun} and Shigeki Nagashima and Johnson, {Jonas T.} and {Van Dongen}, {G. A.M.S.} and Whiteside, {Theresa L.}",
year = "1996",
month = "7",
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The role of apoptosis in antibody-dependent cell-mediated cytotoxicity against monolayers of human squamous cell carcinoma of the head and neck targets. / Sung, Myung Whun; Nagashima, Shigeki; Johnson, Jonas T.; Van Dongen, G. A.M.S.; Whiteside, Theresa L.

In: Cellular Immunology, Vol. 171, No. 1, 10.07.1996, p. 20-29.

Research output: Contribution to journalArticleAcademicpeer-review

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T1 - The role of apoptosis in antibody-dependent cell-mediated cytotoxicity against monolayers of human squamous cell carcinoma of the head and neck targets

AU - Sung, Myung Whun

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AU - Whiteside, Theresa L.

PY - 1996/7/10

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N2 - Antibody-dependent cellular cytotoxicity (ADCC) against squamous cell carcinoma of the head and neck (SCCHN) targets in the presence of human/mouse chimeric monoclonal antibodies (cMAbs), SF-25 and 323/A3, is mediated by natural killer (NK) cells. In 4-hr 51Cr-release assays with SSCHN targets in suspension, ADCC was always significantly better (P < 0.01) than that measured in parallel with the same target cells in monolayers. No differences were observed in the level of expression of the relevant antigens recognized by cMAbs on these targets. To better explain the difference, 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) monolayer and [3H]thymidine-release assays were used. Cytostasis and cell death measured in monolayer MTT assays and DNA fragmentation measured in [3H]thymidine-release assays were significantly higher (P = 0.028) than cytotoxicity determined using 51Cr-labeled SCCHN monolayers. Cell death observed in monolayer MTT assays was blocked by pretreating SCCHN targets with cycloheximide or actinomycin-D or by paraformaldehyde fixation of effector cells. The presence of apoptotic cells in monolayers co-incubated with effector cells was demonstrated in situ by labeling fragmented ends of DNA with fluorescein-conjugated dUTP and terminal deoxynucleotidyl transferase and also by flow cytometry of target cells obtained from such monolayers. Our results indicate that NK cells preferentially utilize membrane lysis (necrosis) in ADCC with tumor cell targets in single-cell suspensions. However, necrosis is not efficient in monolayers. In the presence of cMAbs, apoptosis is the primary mechanism of NK cell-mediated killing in monolayers of SCCHN targets, which were found to express receptors for tumor necrosis factor and fas ligand.

AB - Antibody-dependent cellular cytotoxicity (ADCC) against squamous cell carcinoma of the head and neck (SCCHN) targets in the presence of human/mouse chimeric monoclonal antibodies (cMAbs), SF-25 and 323/A3, is mediated by natural killer (NK) cells. In 4-hr 51Cr-release assays with SSCHN targets in suspension, ADCC was always significantly better (P < 0.01) than that measured in parallel with the same target cells in monolayers. No differences were observed in the level of expression of the relevant antigens recognized by cMAbs on these targets. To better explain the difference, 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) monolayer and [3H]thymidine-release assays were used. Cytostasis and cell death measured in monolayer MTT assays and DNA fragmentation measured in [3H]thymidine-release assays were significantly higher (P = 0.028) than cytotoxicity determined using 51Cr-labeled SCCHN monolayers. Cell death observed in monolayer MTT assays was blocked by pretreating SCCHN targets with cycloheximide or actinomycin-D or by paraformaldehyde fixation of effector cells. The presence of apoptotic cells in monolayers co-incubated with effector cells was demonstrated in situ by labeling fragmented ends of DNA with fluorescein-conjugated dUTP and terminal deoxynucleotidyl transferase and also by flow cytometry of target cells obtained from such monolayers. Our results indicate that NK cells preferentially utilize membrane lysis (necrosis) in ADCC with tumor cell targets in single-cell suspensions. However, necrosis is not efficient in monolayers. In the presence of cMAbs, apoptosis is the primary mechanism of NK cell-mediated killing in monolayers of SCCHN targets, which were found to express receptors for tumor necrosis factor and fas ligand.

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