Detection of function-associated molecules on rat leukemic NK cells

Activation by monoclonal antibody or phorbol ester

D. L. Evans, David T. Harris, D. L. Staton, L. Jaso-Friedmann

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

In the present study a rat leukemia NK cell line designated CRC- (derived from RNK-16 cells) was shown to spontaneously transform into a noncytolytic (NL) line referred to as CRC-/NL cells. CRC- and CRC-/NL cells were utilized to study pathways of NK activation by phorbol esters, calcium ionophore (A23187), and monoclonal antibody (mAb). 10-6-10-7 M phorbol myristate acetate (PMA) but not phorbol didecanoate or 4-beta-phorbol activated CRC-/NL to lyse YAC-1 targets. Activated CRC-/NL cells produced 20-90% specific cytotoxicity compared to 0-5% for nonactivated cells. 10-7 M PMA inhibited normal CRC- cytotoxicity. The optimum concentration of PMA for activation was 106-10-7 M and 3-6 h treatment time. Augmentation of cytotoxicity by PMA occurred at different E:T ratios. The time required to reverse the PMA activation of CRC-/NL cells was approximately 9-10 h posttreatment. In an effort to attempt to differentiate pathways which initiated activation, CRC-/NL cells were treated with FAM binding mAb, or with combinations of mAb and ionophore, mAb and PMA, or PMA and A23187. mAb singly or in combination with 10-7 M PMA increased cytotoxicity. However, A23187 either singly or when combined with PMA or mAb did not produce an augmented lysis of YAC-1 target cells. Additional experiments were conducted to determine if PMA activation was associated with FAM binding. This was accomplished by analyzing redirected killing of various FAM mAb-producing myeloma cells in the presence of 10-7 M PMA. PMA treatment of the CRC-/NL cells caused a significant increase in the lysis of myeloma/mAb-producing cells compared to control cells. Further evidence that FAM binding was associated with cytotoxicity was presented by demonstrating specific inhibition of redirected lysis by homologous mAb. Phenotype analysis of CRC- and CRC-/NL cells demonstrated that OX-7 and OX-1 expression on CRC-/NL cells was increased by 71.8 and 86.8% respectively compared to CRC-. FAM expression (78-83% positives) by CRC- and CRC-/NL cells was not different. These experiments indicated at the functional level that rat NK cells can be activated for increased cytotoxicity by FAM-specific mAb binding and/or by treatment with the diacylglycerol analogue PMA. This implies that protein kinase C mobilization either singly or in concert with inositol-1,4,5-triphsophate activation following FAM mAb binding may play important roles in NK cell cytotoxicity.

Original languageEnglish (US)
Pages (from-to)353-365
Number of pages13
JournalNatural Immunity and Cell Growth Regulation
Volume9
Issue number6
StatePublished - 1990
Externally publishedYes

Fingerprint

Phorbol Esters
Tetradecanoylphorbol Acetate
Natural Killer Cells
Rats
Chemical activation
Monoclonal Antibodies
Molecules
Cytotoxicity
Calcimycin
Antibody-Producing Cells
Calcium Ionophores
Diglycerides
Ionophores
Inositol
Protein Kinase C
Leukemia
Experiments
Cells

ASJC Scopus subject areas

  • Immunology
  • Clinical Biochemistry

Cite this

Detection of function-associated molecules on rat leukemic NK cells : Activation by monoclonal antibody or phorbol ester. / Evans, D. L.; Harris, David T.; Staton, D. L.; Jaso-Friedmann, L.

In: Natural Immunity and Cell Growth Regulation, Vol. 9, No. 6, 1990, p. 353-365.

Research output: Contribution to journalArticle

@article{d3328ae1479b4ca9b917abac1fd082e3,
title = "Detection of function-associated molecules on rat leukemic NK cells: Activation by monoclonal antibody or phorbol ester",
abstract = "In the present study a rat leukemia NK cell line designated CRC- (derived from RNK-16 cells) was shown to spontaneously transform into a noncytolytic (NL) line referred to as CRC-/NL cells. CRC- and CRC-/NL cells were utilized to study pathways of NK activation by phorbol esters, calcium ionophore (A23187), and monoclonal antibody (mAb). 10-6-10-7 M phorbol myristate acetate (PMA) but not phorbol didecanoate or 4-beta-phorbol activated CRC-/NL to lyse YAC-1 targets. Activated CRC-/NL cells produced 20-90{\%} specific cytotoxicity compared to 0-5{\%} for nonactivated cells. 10-7 M PMA inhibited normal CRC- cytotoxicity. The optimum concentration of PMA for activation was 106-10-7 M and 3-6 h treatment time. Augmentation of cytotoxicity by PMA occurred at different E:T ratios. The time required to reverse the PMA activation of CRC-/NL cells was approximately 9-10 h posttreatment. In an effort to attempt to differentiate pathways which initiated activation, CRC-/NL cells were treated with FAM binding mAb, or with combinations of mAb and ionophore, mAb and PMA, or PMA and A23187. mAb singly or in combination with 10-7 M PMA increased cytotoxicity. However, A23187 either singly or when combined with PMA or mAb did not produce an augmented lysis of YAC-1 target cells. Additional experiments were conducted to determine if PMA activation was associated with FAM binding. This was accomplished by analyzing redirected killing of various FAM mAb-producing myeloma cells in the presence of 10-7 M PMA. PMA treatment of the CRC-/NL cells caused a significant increase in the lysis of myeloma/mAb-producing cells compared to control cells. Further evidence that FAM binding was associated with cytotoxicity was presented by demonstrating specific inhibition of redirected lysis by homologous mAb. Phenotype analysis of CRC- and CRC-/NL cells demonstrated that OX-7 and OX-1 expression on CRC-/NL cells was increased by 71.8 and 86.8{\%} respectively compared to CRC-. FAM expression (78-83{\%} positives) by CRC- and CRC-/NL cells was not different. These experiments indicated at the functional level that rat NK cells can be activated for increased cytotoxicity by FAM-specific mAb binding and/or by treatment with the diacylglycerol analogue PMA. This implies that protein kinase C mobilization either singly or in concert with inositol-1,4,5-triphsophate activation following FAM mAb binding may play important roles in NK cell cytotoxicity.",
author = "Evans, {D. L.} and Harris, {David T.} and Staton, {D. L.} and L. Jaso-Friedmann",
year = "1990",
language = "English (US)",
volume = "9",
pages = "353--365",
journal = "Natural Immunity and Cell Growth Regulation",
issn = "0254-7600",
publisher = "S. Karger AG",
number = "6",

}

TY - JOUR

T1 - Detection of function-associated molecules on rat leukemic NK cells

T2 - Activation by monoclonal antibody or phorbol ester

AU - Evans, D. L.

AU - Harris, David T.

AU - Staton, D. L.

AU - Jaso-Friedmann, L.

PY - 1990

Y1 - 1990

N2 - In the present study a rat leukemia NK cell line designated CRC- (derived from RNK-16 cells) was shown to spontaneously transform into a noncytolytic (NL) line referred to as CRC-/NL cells. CRC- and CRC-/NL cells were utilized to study pathways of NK activation by phorbol esters, calcium ionophore (A23187), and monoclonal antibody (mAb). 10-6-10-7 M phorbol myristate acetate (PMA) but not phorbol didecanoate or 4-beta-phorbol activated CRC-/NL to lyse YAC-1 targets. Activated CRC-/NL cells produced 20-90% specific cytotoxicity compared to 0-5% for nonactivated cells. 10-7 M PMA inhibited normal CRC- cytotoxicity. The optimum concentration of PMA for activation was 106-10-7 M and 3-6 h treatment time. Augmentation of cytotoxicity by PMA occurred at different E:T ratios. The time required to reverse the PMA activation of CRC-/NL cells was approximately 9-10 h posttreatment. In an effort to attempt to differentiate pathways which initiated activation, CRC-/NL cells were treated with FAM binding mAb, or with combinations of mAb and ionophore, mAb and PMA, or PMA and A23187. mAb singly or in combination with 10-7 M PMA increased cytotoxicity. However, A23187 either singly or when combined with PMA or mAb did not produce an augmented lysis of YAC-1 target cells. Additional experiments were conducted to determine if PMA activation was associated with FAM binding. This was accomplished by analyzing redirected killing of various FAM mAb-producing myeloma cells in the presence of 10-7 M PMA. PMA treatment of the CRC-/NL cells caused a significant increase in the lysis of myeloma/mAb-producing cells compared to control cells. Further evidence that FAM binding was associated with cytotoxicity was presented by demonstrating specific inhibition of redirected lysis by homologous mAb. Phenotype analysis of CRC- and CRC-/NL cells demonstrated that OX-7 and OX-1 expression on CRC-/NL cells was increased by 71.8 and 86.8% respectively compared to CRC-. FAM expression (78-83% positives) by CRC- and CRC-/NL cells was not different. These experiments indicated at the functional level that rat NK cells can be activated for increased cytotoxicity by FAM-specific mAb binding and/or by treatment with the diacylglycerol analogue PMA. This implies that protein kinase C mobilization either singly or in concert with inositol-1,4,5-triphsophate activation following FAM mAb binding may play important roles in NK cell cytotoxicity.

AB - In the present study a rat leukemia NK cell line designated CRC- (derived from RNK-16 cells) was shown to spontaneously transform into a noncytolytic (NL) line referred to as CRC-/NL cells. CRC- and CRC-/NL cells were utilized to study pathways of NK activation by phorbol esters, calcium ionophore (A23187), and monoclonal antibody (mAb). 10-6-10-7 M phorbol myristate acetate (PMA) but not phorbol didecanoate or 4-beta-phorbol activated CRC-/NL to lyse YAC-1 targets. Activated CRC-/NL cells produced 20-90% specific cytotoxicity compared to 0-5% for nonactivated cells. 10-7 M PMA inhibited normal CRC- cytotoxicity. The optimum concentration of PMA for activation was 106-10-7 M and 3-6 h treatment time. Augmentation of cytotoxicity by PMA occurred at different E:T ratios. The time required to reverse the PMA activation of CRC-/NL cells was approximately 9-10 h posttreatment. In an effort to attempt to differentiate pathways which initiated activation, CRC-/NL cells were treated with FAM binding mAb, or with combinations of mAb and ionophore, mAb and PMA, or PMA and A23187. mAb singly or in combination with 10-7 M PMA increased cytotoxicity. However, A23187 either singly or when combined with PMA or mAb did not produce an augmented lysis of YAC-1 target cells. Additional experiments were conducted to determine if PMA activation was associated with FAM binding. This was accomplished by analyzing redirected killing of various FAM mAb-producing myeloma cells in the presence of 10-7 M PMA. PMA treatment of the CRC-/NL cells caused a significant increase in the lysis of myeloma/mAb-producing cells compared to control cells. Further evidence that FAM binding was associated with cytotoxicity was presented by demonstrating specific inhibition of redirected lysis by homologous mAb. Phenotype analysis of CRC- and CRC-/NL cells demonstrated that OX-7 and OX-1 expression on CRC-/NL cells was increased by 71.8 and 86.8% respectively compared to CRC-. FAM expression (78-83% positives) by CRC- and CRC-/NL cells was not different. These experiments indicated at the functional level that rat NK cells can be activated for increased cytotoxicity by FAM-specific mAb binding and/or by treatment with the diacylglycerol analogue PMA. This implies that protein kinase C mobilization either singly or in concert with inositol-1,4,5-triphsophate activation following FAM mAb binding may play important roles in NK cell cytotoxicity.

UR - http://www.scopus.com/inward/record.url?scp=0025609830&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0025609830&partnerID=8YFLogxK

M3 - Article

VL - 9

SP - 353

EP - 365

JO - Natural Immunity and Cell Growth Regulation

JF - Natural Immunity and Cell Growth Regulation

SN - 0254-7600

IS - 6

ER -