Egzosomni faktor miR-433 regulira izlučivanje upalnih faktora u epitelnim stanicama endometrija mliječnih krava s endometritisom

Endometritis is a common postpartum reproductive system disease, which causes reproductive disorders and even infertility in dairy cows. High-throughput sequencing revealed that exosomal miR-433 is differentially expressed in the peripheral blood of dairy cows with endometritis. However, the specific roles of miR-433 in the occurrence and development of endometritis in dairy cows are still unclear. Therefore, a miR-433-labeled probe was used initially and found that miR-433 is mainly located in uterine cavity epithelial cells and stromal cells. The fluorescence expression of miR-433 in the uterine cavity epithelium and stromal tissues of dairy cows with endometritis is significantly weaker than that in the healthy tissues. The qRT-PCR results showed that the expressions of miR-433 in the uterine tissues of dairy cows with endometritis, LPS stimulated endometrial epithelial cells (EECs), and their derived exosomes were significantly lower than those in the uterine tissues of healthy dairy cows, EECs and their derived exosomes. Exosomal miR-433 derived from EECs transports into neighboring LPS-stimulated EECs by exosome vesicles fusion, regulating the secretion of inflammatory factors within the endometrial epithelium of dairy cows with endometritis, further influencing the development of endometritis. In conclusion, the intensity of miR-433 expression decreased in the epithelial and stromal cells of the uterine lumen and exosomes derived from endometrial epithelium in dairy cows during the occurrence of endometritis, which to some extent promotes the development and progression of endometritis in dairy cows.Endometritis je česta poslijeporođajna bolest reproduktivnog sustava, koja uzrokuje poremećaje, pa čak i neplodnost u mliječnih krava. Sekvenciranje visoke propusnosti otkrilo je da je u perifernoj krvi mliječnih krava s endometritisom egzosomni miR-433 diferencijalno eksprimiran. Još uvijek, međutim, nisu razjašnjene specifične uloge miR-433 u nastanku i razvoju endometritisa u mliječnih krava. S obzirom na to, inicijalno je upotrijebljena sonda označena kao miR-433 te je ustanovljeno da se miR-433 uglavnom nalazi u epitelnim i stromalnim stanicama maternične šupljine. Ekspresija miR-433 dobivena metodom fluorescencije iz epitela i stromalnog tkiva maternične šupljine mliječnih krava s endometritisom znakovito je manja nego u zdravim tkivima. Rezultati dobiveni qRT-PCR-om pokazali su da su ekspresija molekule miR-433 u materničnom tkivu mliječnih krava s endometritisom, endometrijske epitelne stanice (EEC) stimulirane lipopolisaharidom (LPS) i njihovi egzomi bili znakovito niži od onih u materničnom tkivu zdravih mliječnih krava. Egzosomni miR-433 dobiven iz EEC-a prevodi se u susjedne EEC-ove stimulirane LPS-om fuzijom egzosomskih vezikula, čime se regulira izlučivanje upalnih čimbenika u epitelu endometrija mliječnih krava s endometritisom, što dodatno utječe na razvoj endometritisa. Zaključno, intenzitet ekspresije miR-433 smanjio se u epitelnim i stromalnim stanicama maternične šupljine i egzomima dobivenim iz epitela endometrija u mliječnih krava s endometritisom, što u određenoj mjeri potiče razvoj i progresiju endometritisa u mliječnih krava

Akt/PKB suppresses DNA damage processing and checkpoint activation in late G2

Using chemical genetics to reversibly inhibit Cdk1, we find that cells arrested in late G2 are unable to delay mitotic entry after irradiation. Late G2 cells detect DNA damage lesions and form γ-H2AX foci but fail to activate Chk1. This reflects a lack of DNA double-strand break processing because late G2 cells fail to recruit RPA (replication protein A), ATR (ataxia telangiectasia and Rad3 related), Rad51, or CtIP (C-terminal interacting protein) to sites of radiation-induced damage, events essential for both checkpoint activation and initiation of DNA repair by homologous recombination. Remarkably, inhibition of Akt/PKB (protein kinase B) restores DNA damage processing and Chk1 activation after irradiation in late G2. These data demonstrate a previously unrecognized role for Akt in cell cycle regulation of DNA repair and checkpoint activation. Because Akt/PKB is frequently activated in many tumor types, these findings have important implications for the evolution and therapy of such cancer