Human interferon-inducible protein 10: expression and purification of recombinant protein demonstrate inhibition of early human hematopoietic progenitors.
Human interferon-inducible protein 10 (IP-10), a member of the family of the small secreted proteins called intercrine cytokines or chemokines, is secreted by interferon gamma-stimulated T cells, monocytes, endothelial cells, and keratinocytes. We have begun to explore the biological properties of IP-10 by cloning and overexpression in baculovirus and in bacterial protein expression systems. A 9.9-kD protein was secreted by infected insect cells, which on sodium dodecyl sulfate-polyacrilamide gel electrophoresis comigrated with keratinocyte IP-10 and with f(22-98), a bacterial recombinant fragment lacking the signal sequence but containing all other residues of IP-10. All three reacted with antibodies recognizing residues 10-98 (alpha IP-10) and 77-98 of IP-10 (alpha 22), demonstrating that it is secreted by keratinocytes and insect cells after removal of the signal sequence but without proteolysis of the COOH-terminal end. Purified rIP-10 suppresses in vitro colony formation by early human bone marrow progenitor cells which need r-steel factor (rSLF) and rGM-CSF or rSLF and r-erythropoeitin (rEPO). The inhibition is dose dependent, is complete at concentrations > or = 50 ng/ml, is prevented by preincubation of rIP-10 with alpha IP-10, but not by alpha 22, and is seen with highly purified CD34+ cells, suggesting direct effect of rIP-10 on the progenitors. Combination of rIP-10 and other chemokines at inactive concentrations inhibited colony formation in a synergistic manner. rIP-10 did not affect colony formation in the absence of any growth factors or in the presence of rEPO or rGM-CSF but in absence of rSLF. The effects of IP-10 may be relevant to normal marrow function and might be harnessed to protect human hematopoietic progenitors from the cytotoxic effects of chemotherapy.
Sarris
A H
AH
Lymphoma Service, Memorial Sloan-Kettering Cancer Center, New York 10021.
Broxmeyer
H E
HE
Wirthmueller
U
U
Karasavvas
N
N
Cooper
S
S
Lu
L
L
Krueger
J
J
Ravetch
J V
JV
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D002478:Cells, Cultured; D054357:Chemokine CXCL10; D019743:Chemokines, CXC; D003001:Cloning, Molecular; D016207:Cytokines / Q000502:physiology*; D006410:Hematopoiesis / Q000187:drug effects*; D006412:Hematopoietic Stem Cells / Q000187:drug effects*; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009036:Moths; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D011994:Recombinant Proteins / Q000494:pharmacology
D054357:Chemokine CXCL10; D019743:Chemokines, CXC; D016207:Cytokines; D009838:Oligodeoxyribonucleotides; D011994:Recombinant Proteins