Transcriptome analysis of mesenchymal stromal cells of the large and small intestinal smooth muscle layers reveals a unique gastrontestinal stromal signature.

Mesenchymal stromal cells in the muscle layer of the large intestine are essential for the regulation of intestinal motility. They form electrogenic syncytia with the smooth muscle and interstitial cells of Cajal (ICCs) to regulate smooth muscle contraction. Mesenchymal stromal cells are present in the muscle layer throughout the gastrointestinal tract. However, their area-specific characteristics remain ambiguous. In this study, we compared mesenchymal stromal cells from the large and small intestinal muscle layers. Histological analysis using immunostaining showed that the cells in the large and small intestines were morphologically distinct. We established a method to isolate mesenchymal stromal cells from wild-type mice with platelet-derived growth factor receptor-alpha (PDGFRα) as a marker on the cell surface and performed RNAseq. Transcriptome analysis revealed that PDGFRα+ cells in the large intestine exhibited increased expression levels of collagen-related genes, whereas PDGFRα+ cells in the small intestine exhibited increased expression levels of channel/transporter genes, including Kcn genes. These results suggest that mesenchymal stromal cells differ morphologically and functionally depending on gastrointestinal tract. Further investigations of the cellular properties of mesenchymal stromal cells in the gastrointestinal tract will aid in optimizing methods for the prevention and treatment of gastrointestinal diseases.

Estrogen-dependent uterine secretion of osteopontin activates blastocyst adhesion competence.

Embryo implantation is a highly orchestrated process that involves blastocyst-uterine interactions. This process is confined to a defined interval during gestation referred to as the "window of embryo implantation receptivity". In mice this receptive period is controlled by ovarian estrogen and involves a coordination of blastocyst adhesion competence and uterine receptivity. Mechanisms coordinating the acquisition of blastocyst adhesion competence and uterine receptivity are largely unknown. Here, we show that ovarian estrogen indirectly regulates blastocyst adhesion competence. Acquisition of blastocyst adhesion competence was attributed to integrin activation (e.g. formation of adhesion complexes) rather than de novo integrin synthesis. Osteopontin (OPN) was identified as an estrogen-dependent uterine endometrial gland secretory factor responsible for activating blastocyst adhesion competence. Increased adhesion complex assembly in OPN-treated blastocysts was mediated through focal adhesion kinase (FAK)- and phosphatidylinositol 3-kinase (PI3K)-dependent signaling pathways. These findings define for the first time specific regulatory components of an estrogen-dependent pathway coordinating blastocyst adhesion competence and uterine receptivity.

Expression of mesenchymal-related genes by the bovine trophectoderm following conceptus attachment to the endometrial epithelium.

In the course of experiments to identify and characterize the factors that function in bovine conceptuses during peri-attachment periods, various transcripts related to the epithelial-mesenchymal transition (EMT) were found. In this study, RNA was extracted from different sets of days 17, 20, and 22 (day 0=day of estrous) bovine conceptuses and subjected to real-time PCR analysis as well as Western blotting, from which abundances of N-cadherin (CDH2), vimentin, matrix metalloproteinase 2 (gelatinase A, 72 kDa gelatinase, 72 kDa type IV collagenase) (MMP2), and matrix metallopeptidase 9 (gelatinase B, 92 kDa gelatinase, 92 kDa type IV collagenase) (MMP9) mRNAs were determined on day 22, concurrent with (CDH1) mRNA and protein downregulation. Transcription factors in EMT processes were then analyzed and changes in snail homolog 2 (Drosophila) (SNAI), zinc finger E-box binding homeobox 1 (ZEB1), zinc finger E-box binding homeobox 2 (ZEB2), twist homolog 1 (Drosophila) (TWIST1), twist homolog 2 (Drosophila) (TWIST2), and Kruppel-like factor 8 (KLF8) transcripts were found in day 22 conceptuses, while confirming SNAI2 expression by Western blotting. Immunohistochemical analysis revealed that the day 22 trophectoderm expressed the mesenchymal markers N-cadherin and vimentin as well as the epithelial marker cytokeratin. In attempts to identify the molecular mechanisms by which the trophectoderm expressed EMT-related genes, growth factor receptors associated with EMT were analyzed. Upregulation of the growth factor receptor transcripts, fibroblast growth factor receptor 1 (FGFR1), platelet-derived growth factor receptor, alpha polypeptide (PDGFRA), platelet-derived growth factor receptor, beta polypeptide (PDGFRB), and transforming growth factor, beta receptor II (70/80 kDa) (TGFBR2) mRNAs, was found on day 22. The analysis was extended to determine the integrin (ITG) transcripts and found high levels of integrin, alpha 4 (antigen CD49D, alpha 4 subunit of VLA-4 receptor) (ITGA4), integrin, alpha 8 (ITGA8), integrin, beta 3 (platelet glycoprotein IIIa, antigen CD61) (ITGB3), and integrin, beta 5 (ITGB5) mRNAs on day 22. These observations indicate that after the conceptus-endometrium attachment, EMT-related transcripts as well as the epithelial marker cytokeratin were present in the bovine trophectoderm and suggest that the implantation process for noninvasive trophoblasts requires not only extracellular matrix expression but also partial EMT.