Temporal and Spatial Expression of neu Differentiation Factor (NDF) in the Mouse Uterus during the Periimplantation Period. • 286

NDF is a 44 kD soluble glycoprotein that exists as membranebound and secreted forms. It was originally isolated as the factor which stimulated tyrosine phosphorylation of neu (ErbB2,

neu/HER2), a RTK of the EGF receptor family. NDF belongs to the family of EGFlike ligands, which includes EGF, TGFa, HBEGF, amphiregulin, betacellulin and epiregulin. Several of these are

expressed in the mouse periimplantation uterus in a temporal and cell type specific manner. However, the expression pattern of betacellulin, epiregulin and NDF during this period is unknown.

Here, we examined the expression of NDF in the uterus during the periimplantation period. RTPCR was performed on uterine tissues from d1d8 pregnant CD1 mice (d1= vaginal plug). Primers were

derived from the rat NDFa2c sequence and were designed to amplify a region that is conserved among the 10 known NDF isoforms. The expected 330 bp product was detected in all d1d8 uterine and

in the control (adult brain) tissues. This PCR fragment was cloned and sequenced and was found to differ from the rat sequence at only a single nucleotide. This cDNA and a 1.4 kb cDNA

fragment of rat NDFa2c were used to generate 32P labeled cRNA probes. Northern analysis revealed expression of multiple transcripts in the uterus and brain RNA samples. There was sequential

up or down regulation of several RNA transcripts over d1d8, but no band that was predominant at d4d5, around the time of implantation (d4.5). Insitu hybridization of uterine tissues was then

performed, and discrete localization of NDF mRNAs was detected in the stroma adjacent to the implanting blastocyst on the evening of d4 at 2300 hours, at the time of attachment reaction.

This pattern of expression is unique and differs from HBEGF and amphiregulin. The expression of HBEGF and amphiregulin is restricted to the luminal epithelium of the uterus at the time of

attachment reaction, which may suggest an independent role for NDF in the implantation process. In addition, this pattern of expression suggests that embryouterine cell signaling may be

highly localized and coordinated with the temporally and spatially restricted expression of other EGF receptors and their ligands.

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