L Tseng1, J Zhang, M S Goligorsky. 1. Department of Obstetrics, Gynecology, State University of New York at Stony Brook 11794-8091, USA.
Abstract
OBJECTIVES: Nitric oxide synthase (NOS), responsible for the synthesis of nitric oxide (NO), is the key enzyme that regulates the NO-mediated biologic functions in various types of tissue. We believed that NO may be involved in regulating the development of the microvascular system and vascular tone in human endometrium. However, the biology of NOS in this system remains poorly understood. To determine the paracrine action of NOS derived from endometrial cells on vascular blood flow, the first task is to identify NOS in endometrial cells. This study was undertaken to examine whether human endometrial cells would express endothelial or inducible NOS (eNOS or iNOS). In addition, we examined the cell-specific expression of eNOS. METHODS: Beta nicotinamide adenine dinucleotide reduced from (NADPH)-diaphorase histochemistry assay was carried out in human endometrial specimens (n = 4). Nitric oxide synthase mRNA expression was studied in intact tissue (n = 4), isolated epithelial glands (n = 25), and stromal cells (n = 10). Northern blot and solution hybridization/ribonuclease protection assay with specific RNA probes transcribed from human eNOS and iNOS cDNA were used to identify NOS mRNAs. RESULTS: NADPH-diaphorase histochemistry selectively labeled epithelial glands in the late secretory phase. Northern blot analysis revealed that glandular cells expressed a single size of eNOS mRNA (4.5 kb) but no detectable iNOS. Endothelial NOS mRNA was expressed in epithelial glands, but it was not detectable in stromal cells. The relative amount of eNOS in epithelial glands isolated from specimens (n = 23) at various stages of menstrual cycle were analyzed using a solution hybridization assay. The levels of eNOS mRNA varied among specimens. Epithelial glands from early secretory endometrium showed greater expression of eNOS mRNA. The highest eNOS expression was found in the glands of late secretory endometrium. In contrast, iNOS was detected only in the epithelial glands of a menstrual endometrium. CONCLUSION: This study demonstrated that NADPH-diaphorase activity is highly concentrated in the epithelial glands of late secretory endometrium. Expression of eNOS mRNA is enhanced in the glands of early secretory endometrium. The strongest expression resides in the epithelial glands of late secretory endometrium. This observation suggests that regulation of eNOS may be a physiologic response to steroid hormones and locally produced peptide hormones in the endometrial environment. It also suggests that NO may play a role in the onset of menses.
OBJECTIVES:Nitric oxide synthase (NOS), responsible for the synthesis of nitric oxide (NO), is the key enzyme that regulates the NO-mediated biologic functions in various types of tissue. We believed that NO may be involved in regulating the development of the microvascular system and vascular tone in human endometrium. However, the biology of NOS in this system remains poorly understood. To determine the paracrine action of NOS derived from endometrial cells on vascular blood flow, the first task is to identify NOS in endometrial cells. This study was undertaken to examine whether human endometrial cells would express endothelial or inducible NOS (eNOS or iNOS). In addition, we examined the cell-specific expression of eNOS. METHODS:Beta nicotinamide adenine dinucleotide reduced from (NADPH)-diaphorase histochemistry assay was carried out in human endometrial specimens (n = 4). Nitric oxide synthase mRNA expression was studied in intact tissue (n = 4), isolated epithelial glands (n = 25), and stromal cells (n = 10). Northern blot and solution hybridization/ribonuclease protection assay with specific RNA probes transcribed from humaneNOS and iNOS cDNA were used to identify NOS mRNAs. RESULTS:NADPH-diaphorase histochemistry selectively labeled epithelial glands in the late secretory phase. Northern blot analysis revealed that glandular cells expressed a single size of eNOS mRNA (4.5 kb) but no detectable iNOS. Endothelial NOS mRNA was expressed in epithelial glands, but it was not detectable in stromal cells. The relative amount of eNOS in epithelial glands isolated from specimens (n = 23) at various stages of menstrual cycle were analyzed using a solution hybridization assay. The levels of eNOS mRNA varied among specimens. Epithelial glands from early secretory endometrium showed greater expression of eNOS mRNA. The highest eNOS expression was found in the glands of late secretory endometrium. In contrast, iNOS was detected only in the epithelial glands of a menstrual endometrium. CONCLUSION: This study demonstrated that NADPH-diaphorase activity is highly concentrated in the epithelial glands of late secretory endometrium. Expression of eNOS mRNA is enhanced in the glands of early secretory endometrium. The strongest expression resides in the epithelial glands of late secretory endometrium. This observation suggests that regulation of eNOS may be a physiologic response to steroid hormones and locally produced peptide hormones in the endometrial environment. It also suggests that NO may play a role in the onset of menses.