Expression of the c-fms proto-oncogene and of the cytokine, CSF-1, during mouse embryogenesis

The c-fms gene encodes the cell surface receptor of the colony-stimulating factor, CSF-1. CSF-1 has recently been shown to be expressed in the maternal uterine endometrium of pregnant mice. The ontogenetic and spatial patterns of expression of the murine proto-oncogene c-fms were analyzed in the developing mouse placenta by the technique of in situ hybridization. c-fms expression was not detected in fetally derived tissues until 9.5 days postcoitum (pc) when expression first appeared in the mural trophoblast giant cells. Expression persisted at high levels in trophoblast cells throughout gestation. In the mature placenta from 13.5 days pc on, c-fms was expressed chiefly in the spongiotrophoblast layer and, to a lesser extent, in the labyrinthine trophoblast. CSF-1 expression was first detectable in the uterine epithelium at 8.5 days pc which loosely correlated with the appearance at 7.5 days of c-fms in the decidual cells around the developing egg cylinder. The time course and spatial pattern of expression of these two genes suggest a functional role for the c-fms receptor and its ligand, CSF-1, in trophoblast development and differentiation.     

Expression of c-fos in parietal endoderm, amnion and differentiating F9 teratocarcinoma cells

The expression of the cellular proto-oncogene, c-fos, in extra-embryonic tissues of the mouse was investigated using a v-fos DNA probe and an affinity-purified antiserum raised against a C-terminal synthetic peptide. At 13.5 days of development, parietal endoderm--a tissue not previously studied using these methods--was found to express c-fos RNA at a higher level than the amnion or placenta. The previously reported dramatic increase in c-fos RNA levels in extra-embryonic membranes during gestation was found to be confined to the amnion. The antipeptide serum specifically recovered proteins with Mr values of 46,000 and 39,000 from extracts of parietal endoderm and amnion cells labelled for 15 min with 35S-methionine. On sodium-dodecyl-sulphate/polyacrylamide gel electrophoresis these proteins co-migrated with proteins immunoprecipitated using serum from rats inoculated with FBJ-MuSV-transformed cells (tumour-bearing rat serum). Pulse-chasing and 32P-labelling experiments showed that the protein with an Mr of 46,000 was rapidly converted into higher-molecular-weight phosphorylated derivatives. F9 teratocarcinoma stem cells differentiated into parietal-endoderm-like cells in response to treatment with retinoic acid and dibutyryl cyclic AMP. However, this differentiation was not accompanied by any large transient increase in c-fos RNA expression.     

Temporal expression of tissue inhibitors of metalloproteinases in mouse reproductive tissues during gestation

Tissue inhibitors of metalloproteinases (TIMPs) appear to play an important regulatory role in tissue remodelling and invasion by malignant cells. Since pregnancy involves morphological changes in existing maternal tissues, as well as a strictly controlled invasion by fetal trophoblasts, we have examined the temporal expression of TIMP-1, TIMP-2, and specific metalloproteinases in the mouse uterus, decidua, placenta, amnion, and ovaries throughout gestation by examining mRNA levels on northern and slot blots. Maximal levels of TIMP-1 mRNA were observed from day 6 to day 10 in the uterus, decidua, and placenta. In clear contrast to the early burst of TIMP-1 mRNA accumulation, the level of TIMP-2 mRNA increased steadily throughout gestation in the uterus, decidua, and amnion, while in the placenta it showed a sevenfold increase after day 14. In amnion, TIMP-1 was induced specifically on day 18. Interestingly, the normally high level of TIMP-1 mRNA seen in the ovaries of virgin mice was low during gestation, until day 18 and postpartum, when a sixfold increase over the levels in virgin ovary was observed. In contrast, ovarian TIMP-2 mRNA showed a marginal increase during gestation. The temporal pattern of 72 kDa gelatinase type A followed that of TIMP-2 in the decidua and ovary. Stromelysin-2 mRNA was detected at term only in ovary and decidua. Our data show that the temporal accumulation of TIMP-1 and TIMP-2 mRNA is precisely coordinated in each of the tissue compartments and is independently regulated during the in vivo remodelling of reproductive tissues in gestation. The peak of TIMP-1 mRNA levels in the uterus, decidua, and placenta at midgestation is associated with the most invasive period of embryo development. © 1993 Wiley-Liss, Inc.     

Localization and synthesis of the tumor protein oncomodulin in extraembryonic tissues of the fetal rat

The calcium-binding protein oncomodulin, previously found only in tumors, has been detected during rat development. Specific antisera to purified rat hepatoma oncomodulin (MW 11,500) were used to detect oncomodulin by radioimmunoassay (RIA) and by avidin-biotin-peroxidase complex (ABC) immunohistochemistry. Using RIA, oncomodulin was found to increase in placenta from below the limits of detection (2 ng/mg protein) on Day 13 to approximately 25 ng/mg on Day 16 of pregnancy, and to remain high through to the end of gestation. Determinations on separated inner and outer placenta showed the increase to be greater in the outer placenta (basal zone and decidua) than in the inner placenta (labyrinth). The ABC technique on paraffin sections produced positive staining for oncomodulin throughout the placenta, with the most intense staining occurring in the outer placenta (cytotrophoblast and giant cells of the basal zone). Parietal and visceral yolk sac, and amnion also stained positively, while fetal organs did not. Oncomodulin synthesis measured by [35S]methionine incorporation into immunoprecipitates occurred in isolated inner and outer placenta, whole placenta, the separated trophectoderm and endoderm of the parietal yolk sac, and amnion. No oncomodulin synthesis could be measured in visceral yolk sac, fetal liver, or 16-day embryo. This occurrence in developing and transformed tissues demonstrates that oncomodulin is an oncodevelopmental protein.     

Differential calmodulin gene expression in fetal, adult, and neoplastic tissues of rodents

Differential expression during rat development of three genes for calmodulin (CaM I-III) was examined in amnion, decidua, embryo, liver, placenta, parietal and visceral yolk sacs and uterus. CaMI expression was constant except for increasing activity in VYS during gestation. CaMII expression increased in all tissues except for a decrease in embryo. CaMIII did not change dramatically. Differential expression was also found in chemically or virally induced rat tumors, and in metastatic lung nodules of mouse mammary carcinoma. CaMII was the major gene expressed in all these neoplastic tissues.     

Expression and genomic imprinting of DCN, PON2 and PEG3 genes in porcine placenta

Imprinted genes play vital roles in the placental development and fetal growth in eutherian mammals. DCN (decorin), PON2 (paraoxonase 2) and PEG3 (paternally expressed 3) genes have been identified as imprinted genes in the mouse. Here, we detected the imprinting status of three genes in the porcine placenta on DG90 (day 90 of gestation) and the expression differences in Yorkshire and Meishan placenta on DG26, DG55 and DG90. The results indicated that the DCN and PON2 genes were not imprinted genes, while the PEG3 gene showed paternal monoallelic expression in porcine placenta. The expression of the DCN gene increased from DG26 to DG90 in both Yorkshire and Meishan pig placenta. However, this gene expression was greater in Yorkshire than Meishan pig on DG55. The expression of the PON2 gene was greater in Meishan pig than that in Yorkshire on DG26 and DG90. The PEG3 gene expression was not affected by day of pregnancy or breed. Data from the present study contribute to function genomic of porcine placental development.     

Localization of placental lactogen-I in trophoblast giant cells of the mouse placenta

The purpose of this investigation was to identify the cellular origin of placental lactogen-I (PL-I) expression in the mouse placenta and to cytologically define the transition from PL-I to PL-II expression during gestation. PL-I mRNA expression was assessed by in situ hybridization, and expression of PL-I and PL-II protein was determined by immunocytochemical analysis. PL-I mRNA and protein were localized to trophoblast giant cells. Trophoblast giant cells ceased producing PL-I at midgestation and began expressing PL-II. PL-I immunoreactivity was present in trophoblast giant cells on Days 9 and 10 of gestation but was not detectable in trophoblast giant cells on Day 11 of gestation. Immunoreactive PL-II-producing giant cells were detected first on Day 10 of gestation, continuing on Day 11 of gestation. Expression of PL-I and PL-II signals a significant functional transition in trophoblast giant cells of the developing mouse placenta.     

Appearance of functional EGF receptor kinase during rodent embryogenesis.

Mouse and rat embryonic tissues at various stages of development were examined for epidermal growth factor (EGF) receptor kinase activity. The phosphorylated EGF receptor from embryonic tissues appeared as a band of mol. wt. 170 000 daltons on SDS gels. It was clearly demonstrable in the developing mouse fetus from 10 days of gestation onwards. The distribution of the EGF receptor kinase was studied in various tissues of 13 day mouse fetuses. The activity was apparent in the skin, developing skeletal muscles and various internal organs but was notably absent in the liver and brain. The amnion was found to be one of the richest sources of activity while the yolk sac was negative, and the placenta was weakly positive. In 16 day rat fetuses the distribution was quite similar to that of the 13 day mouse fetus. The liver acquired EGF receptor kinase activity by 18 days of gestation and had high activity in neonates. Phosphoamino acid analysis revealed that phosphotyrosine was the major labelled amino acid residue in the embryonic tissues. Thus, the EGF receptor of fetal tissues as studied by immune precipitation and phosphorylation appears to be a similar entity to that found in adult mammalian tissues. This functional EGF receptor kinase activity could first be detected at the time of onset of organogenesis.