GROWTH FACTORS IN REPRODUCTIVE TISSUES
7:30 AM-10:00 AM
(198) BOVINE COLONY STIMULATING FACTOR-1: CLONING AND EXPRESSION IN THE BOVINE UTERUS, CONCEPTUS AND PLACENTOMES.
Lee, Rita1, Li, Ning1, Ledgard, Anita1, Pollard, Jeffery2, 1 AgResearch, Hamilton, NZ2 Dept of Developmental and Molecular Biology, Bronx, NY
ABSTRACT- Colony stimulating factor-1 (CSF-1) is a hematopoetic cytokine that also plays an important role in placental physiology in the mouse. We report here the cloning of the bovine cDNA and the study of its expression in cattle pregnancy. Tissue samples were collected from cows at various gestational stages after transfer at day 7 of a single in vitro produced embryo into the uterus and the expression of CSF-1 in uterine and placental tissues studied using Northerns and in situ hybridisation. The bovine CSF-1 cDNA was cloned by sequential RT-PCR of overlapping fragments and 3′-RACE. Two splice variants were cloned, one equivalent to the secreted form of CSF-1 found in the mouse, the other, equivalent to the putative membrane-bound splice variant first cloned in the human endometrium. Bovine CSF-1 was expressed mainly as two alternatively spliced transcripts of 3 and 4 kb but 1.4 and 0.8 kb mRNAs were also detected. In the non-pregnant uterus, both the 4 and 3 kb mRNA were present throughout the estrous cycle. The ratio of the 4 kb:3 kb transcripts changed from ~ 0.8 on day 1 of the cycle, to ~ 0.3 between day 14 and 17 ( bovine estrous cycle is ~ 21 days), suggesting there was a shift of expression to more of the putative membrane-bound form. In pregnant animals, expression of the 3 kb mRNA declined as the abundance of the 4 kb transcript increased. By day 29 of gestation, the 4 kb:3 kb ratio was ~2.3. This change in the ratio of the two forms was detected as early as day 14; thus, the bovine uterus was able to detect the presence of an elongating blastocyst and alter the expression of the splice variants. As pregnancy progressed, the 4 kb mRNA increased progressively and accounted for almost all of the CSF-1 expressed. CSF-1 mRNA was also detected in the yolk sac, allantois and trophoblast of preimplantation conceptuses and in the fetal cotyledons after placentation. In situ hybridisation showed that CSF-1 mRNA was present in the uterine luminal and glandular epithelia and in the stromal cells of the caruncles in the non-gravid and early gravid uteri. After implantation, CSF-1 was detected in maternal caruncular tissue as well as in the epithelial cells of the fetal villi. The expression of CSF-1 in pre-implantation conceptuses, caruncular tissue and the placentomes is consistent with its proposed role in placental physiology. (FRST C10X0018, New Zealand)
KEY WORDS: CSF-1, bovine, placentome