A prerequisite for implantation in humans is differentiation (decidualization) of stromal cells in the endometrium, believed to be stimulated by progesterone (P) and/or cAMP. In the current study, advances in microarray technology have allowed us to investigate genes differentially expressed in human endometrial stromal cells decidualized in vitro in response to P or cAMP, compared to nondecidualized cells. Endometrial stromal cells were isolated from endometrial biopsy tissue and cultured without steroid hormones, with 1 microM P (after E2 priming), or 1 mM 8-bromo-cAMP. Total RNA was isolated and reverse transcribed to synthesize 32P-labeled cDNA probes using primers corresponding to genes represented on the Clontech Human Atlas cDNA Expression Array. After hybridization, signals were quantified by phosphor imaging densitometry and were normalized to GAPDH and ubiquitin. Of the 588 genes screened, marked upregulation was observed of cytokines, growth factors, nuclear transcription factors, members of the cyclin family, and mediators of the cAMP signal transduction pathway. Additional mRNAs expressed unexpectedly and regulated by P and cAMP, include the insulin receptor, some neurotransmitter receptors, neuromodulators, the FSH receptor, inhibin/activin betaA subunit, inhibin alpha, and TNF-related apoptosis-inducing ligand (TRAIL). Expression of previously unrecognized genes regulated in decidualized human endometrial stromal cells suggests mechanisms not yet appreciated in the endometrium during decidualization. In addition, marked upregulation of cytokines, chemokines, growth factors, apoptosis modulators, and their receptors in decidualized stromal cells supports a major role for paracrine interactions between the stroma and other endogenous and transient cell populations within the endometrium and during early pregnancy.