Expression and activation of caspase-3/CPP32 in CD34+ cord blood cells is linked to apoptosis after growth factor withdrawal

Li Sheng Wang, Hong Jun Liu, Zhen Biao Xia, Hal E. Broxmeyer, Li Lu

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Objective. Caspase-3/CPP32, a member of the interleukin-1 converting enzyme (ICE) family, is considered an executioner protease in mammalian cells during apoptosis. Although expression and activation of caspase-3/CPP32 protein have been studied in many tissues and leukemia cell lines, this has not been explored in primitive hematopoietic CD34+ cells. In this study, we evaluated expression and activation of caspase-3/CPP32 protein in CD34+ cells from cord blood (CB) during apoptosis induced by growth factor deprivation. Methods. Reverse transcriptase-polymerase chain reaction (RT- PCR), Western blot, and flow cytometry analysis were used in this study to determine the expression of caspase-3/CPP32 in CD34+ CB cells during apoptosis. Results. Our results demonstrated that caspase-3/CPP32 mRNA was constitutively expressed at a very low level in freshly isolated CD34+ cells. Expression of caspase-3/CPP32 mRNA and protein was upregulated when these cells were first expanded in suspension culture with growth factors for 3 days. However, only the 32 kDa inactive caspase-3/CPP32 proenzyme was detected in the freshly isolated CD34+ cells and after 3 days expansion with cytokines. Within 12 hours after growth factor withdrawal from expanded cells caspase-3/CPP32 was activated and a cleavage 20 kDa protein was detected; a poly(ADP-ribose) polymerase (PARP) was cleaved by activated caspase-3/CPP32. Activation of caspase-3/CPP32 and apoptosis upon growth factor withdrawal were inhibited/reduced by the caspase inhibitors, z-VAD-fmk and DEVD-CHO. Conclusion. These results demonstrate that caspase-3/CPP32 is involved in apoptosis of primitive CB CD34+ cells but may not be the only mechanism involved. (C) 2000 International Society for Experimental Hematology.

Original languageEnglish (US)
Pages (from-to)907-915
Number of pages9
JournalExperimental Hematology
Volume28
Issue number8
DOIs
StatePublished - Aug 1 2000

Fingerprint

Fetal Blood
Caspase 3
Blood Cells
Intercellular Signaling Peptides and Proteins
Apoptosis
Proteins
Caspase 1
Messenger RNA
Enzyme Precursors
Caspase Inhibitors
Poly(ADP-ribose) Polymerases
Reverse Transcriptase Polymerase Chain Reaction
Suspensions
Flow Cytometry
Leukemia
Peptide Hydrolases
Western Blotting
Cytokines
Cell Line

Keywords

  • Apoptosis
  • Caspase-3/CPP32
  • CD34 cells
  • Cord blood

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

Cite this

Expression and activation of caspase-3/CPP32 in CD34+ cord blood cells is linked to apoptosis after growth factor withdrawal. / Wang, Li Sheng; Liu, Hong Jun; Xia, Zhen Biao; Broxmeyer, Hal E.; Lu, Li.

In: Experimental Hematology, Vol. 28, No. 8, 01.08.2000, p. 907-915.

Research output: Contribution to journalArticle

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abstract = "Objective. Caspase-3/CPP32, a member of the interleukin-1 converting enzyme (ICE) family, is considered an executioner protease in mammalian cells during apoptosis. Although expression and activation of caspase-3/CPP32 protein have been studied in many tissues and leukemia cell lines, this has not been explored in primitive hematopoietic CD34+ cells. In this study, we evaluated expression and activation of caspase-3/CPP32 protein in CD34+ cells from cord blood (CB) during apoptosis induced by growth factor deprivation. Methods. Reverse transcriptase-polymerase chain reaction (RT- PCR), Western blot, and flow cytometry analysis were used in this study to determine the expression of caspase-3/CPP32 in CD34+ CB cells during apoptosis. Results. Our results demonstrated that caspase-3/CPP32 mRNA was constitutively expressed at a very low level in freshly isolated CD34+ cells. Expression of caspase-3/CPP32 mRNA and protein was upregulated when these cells were first expanded in suspension culture with growth factors for 3 days. However, only the 32 kDa inactive caspase-3/CPP32 proenzyme was detected in the freshly isolated CD34+ cells and after 3 days expansion with cytokines. Within 12 hours after growth factor withdrawal from expanded cells caspase-3/CPP32 was activated and a cleavage 20 kDa protein was detected; a poly(ADP-ribose) polymerase (PARP) was cleaved by activated caspase-3/CPP32. Activation of caspase-3/CPP32 and apoptosis upon growth factor withdrawal were inhibited/reduced by the caspase inhibitors, z-VAD-fmk and DEVD-CHO. Conclusion. These results demonstrate that caspase-3/CPP32 is involved in apoptosis of primitive CB CD34+ cells but may not be the only mechanism involved. (C) 2000 International Society for Experimental Hematology.",
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T1 - Expression and activation of caspase-3/CPP32 in CD34+ cord blood cells is linked to apoptosis after growth factor withdrawal

AU - Wang, Li Sheng

AU - Liu, Hong Jun

AU - Xia, Zhen Biao

AU - Broxmeyer, Hal E.

AU - Lu, Li

PY - 2000/8/1

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N2 - Objective. Caspase-3/CPP32, a member of the interleukin-1 converting enzyme (ICE) family, is considered an executioner protease in mammalian cells during apoptosis. Although expression and activation of caspase-3/CPP32 protein have been studied in many tissues and leukemia cell lines, this has not been explored in primitive hematopoietic CD34+ cells. In this study, we evaluated expression and activation of caspase-3/CPP32 protein in CD34+ cells from cord blood (CB) during apoptosis induced by growth factor deprivation. Methods. Reverse transcriptase-polymerase chain reaction (RT- PCR), Western blot, and flow cytometry analysis were used in this study to determine the expression of caspase-3/CPP32 in CD34+ CB cells during apoptosis. Results. Our results demonstrated that caspase-3/CPP32 mRNA was constitutively expressed at a very low level in freshly isolated CD34+ cells. Expression of caspase-3/CPP32 mRNA and protein was upregulated when these cells were first expanded in suspension culture with growth factors for 3 days. However, only the 32 kDa inactive caspase-3/CPP32 proenzyme was detected in the freshly isolated CD34+ cells and after 3 days expansion with cytokines. Within 12 hours after growth factor withdrawal from expanded cells caspase-3/CPP32 was activated and a cleavage 20 kDa protein was detected; a poly(ADP-ribose) polymerase (PARP) was cleaved by activated caspase-3/CPP32. Activation of caspase-3/CPP32 and apoptosis upon growth factor withdrawal were inhibited/reduced by the caspase inhibitors, z-VAD-fmk and DEVD-CHO. Conclusion. These results demonstrate that caspase-3/CPP32 is involved in apoptosis of primitive CB CD34+ cells but may not be the only mechanism involved. (C) 2000 International Society for Experimental Hematology.

AB - Objective. Caspase-3/CPP32, a member of the interleukin-1 converting enzyme (ICE) family, is considered an executioner protease in mammalian cells during apoptosis. Although expression and activation of caspase-3/CPP32 protein have been studied in many tissues and leukemia cell lines, this has not been explored in primitive hematopoietic CD34+ cells. In this study, we evaluated expression and activation of caspase-3/CPP32 protein in CD34+ cells from cord blood (CB) during apoptosis induced by growth factor deprivation. Methods. Reverse transcriptase-polymerase chain reaction (RT- PCR), Western blot, and flow cytometry analysis were used in this study to determine the expression of caspase-3/CPP32 in CD34+ CB cells during apoptosis. Results. Our results demonstrated that caspase-3/CPP32 mRNA was constitutively expressed at a very low level in freshly isolated CD34+ cells. Expression of caspase-3/CPP32 mRNA and protein was upregulated when these cells were first expanded in suspension culture with growth factors for 3 days. However, only the 32 kDa inactive caspase-3/CPP32 proenzyme was detected in the freshly isolated CD34+ cells and after 3 days expansion with cytokines. Within 12 hours after growth factor withdrawal from expanded cells caspase-3/CPP32 was activated and a cleavage 20 kDa protein was detected; a poly(ADP-ribose) polymerase (PARP) was cleaved by activated caspase-3/CPP32. Activation of caspase-3/CPP32 and apoptosis upon growth factor withdrawal were inhibited/reduced by the caspase inhibitors, z-VAD-fmk and DEVD-CHO. Conclusion. These results demonstrate that caspase-3/CPP32 is involved in apoptosis of primitive CB CD34+ cells but may not be the only mechanism involved. (C) 2000 International Society for Experimental Hematology.

KW - Apoptosis

KW - Caspase-3/CPP32

KW - CD34 cells

KW - Cord blood

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