Ontogeny, immunocytochemical localization, and biochemical properties of the pregnancy-associated uterine elastase/cathepsin-G protease inhibitor, antileukoproteinase (ALP): monospecific antibodies to a synthetic peptide recognize native ALP.
Expression of the mRNA encoding the elastase/cathepsin-G protease inhibitor, antileukoproteinase (ALP), is highest in pig uterus during mid- and late pregnancy, suggesting a stage of pregnancy-dependent role for ALP in feto-maternal interactions. To elucidate a function for ALP in these events, immunogenic probes were developed to localize sites of ALP expression in the environment of the developing fetus. Monospecific antibodies raised against a 16-mer synthetic peptide corresponding to residues 21-36 (ALP 16P) of the deduced amino acid sequence of pig uterine ALP were generated by active immunization of sheep. ALP 16P conjugated to keyhole limpet hemocyanin elicited high titer antibodies that were specific to ALP. The antipeptide antibodies were used to characterize pig uterine ALP from allantoic fluids. Uterine ALP has an approximate mol wt of 14,000 and a pI of 8.2 and exhibits elastase inhibitor activity. Amino-terminal amino acid sequencing of uterine ALP indicated the sequence AENALKGGACPPRKIVQC, which has 44% identity with the corresponding region in human bronchial ALP. RIA for ALP, developed using ALP 16P as standard and iodinated tracer, demonstrated the presence of immunoreactive ALP in early, mid-, and late pregnant endometrium and myometrium, placenta, allantoic fluids, fetal cord blood, and fetal liver. ALP was undetectable in the maternal circulation. The ALP levels in endometrium, allantoic fluids, and fetal cord blood changed with the stage of pregnancy; however, ALP content in placenta, myometrium, and fetal liver, although different among tissues, remained invariant during gestation. By immunocytochemical analyses, ALP was localized in the glandular epithelium of the uterus, in placenta, and in fetal liver, consistent with the presence of immunoreactive ALP as measured by RIA. The localization of uterine ALP in placenta and its corresponding transport to fetal circulation provide strong evidence to support a physiological function for the protease inhibitor in the biological mechanisms controlling fetal development in utero.
Simmen
R C
RC
Department of Animal Science, University of Florida, Gainesville 32611.
Michel
F J
FJ
Fliss
A E
AE
Smith
L C
LC
Fliss
M F
MF
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D005260:Female; D007150:Immunohistochemistry; D008431:Maternal-Fetal Exchange; D008969:Molecular Sequence Data; D011247:Pregnancy; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins; D012333:RNA, Messenger / Q000032:analysis; D015842:Serine Proteinase Inhibitors / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D012756:Sheep; D013552:Swine; D014599:Uterus / Q000737:chemistry
D000906:Antibodies; D053491:Proteinase Inhibitory Proteins, Secretory; D011506:Proteins; D012333:RNA, Messenger; D015842:Serine Proteinase Inhibitors