Ovine Fetal Immune Response to Cache Valley Virus Infection

Cache Valley virus (CVV)-induced malformations have been previously reproduced in ovine fetuses. To evaluate the development of the antiviral response by the early, infected fetus, before the development of immunocompetency, ovine fetuses at 35 days of gestation were inoculated in utero with CVV and euthanized at 7, 10, 14, 21, and 28 days postinfection. The antiviral immune response in immature fetuses infected with CVV was evaluated. Gene expression associated with an innate, immune response was quantified by real-time quantitative PCR. The upregulated genes in infected fetuses included ISG15, Mx1, Mx2, IL-1, IL-6, TNF-α, TLR-7, and TLR-8. The amount of Mx1 protein, an interferon-stimulated GTPase capable of restricting growth of bunyaviruses, was elevated in the allantoic and amniotic fluid in infected fetuses. ISG15 protein expression was significantly increased in target tissues of infected animals. B lymphocytes and immunoglobulin-positive cells were detected in lymphoid tissues and in the meninges of infected animals. These results demonstrated that the infected ovine fetus is able to initiate an innate and adaptive immune response much earlier than previously known, which presumably contributes to viral clearance in infected animals

Exosomal miR-940 maintains SRC-mediated oncogenic activity in cancer cells: a possible role for exosomal disposal of tumor suppressor miRNAs

Exosomes have emerged as important mediators of diverse biological functions including tumor suppression, tumor progression, invasion, immune escape and cell-to-cell communication, through the release of molecules such as mRNAs, miRNAs, and proteins. Here, we identified differentially expressed exosomal miRNAs between normal epithelial ovarian cell line and both resistant and sensitive ovarian cancer (OC) cell lines. We found miR-940 as abundant in exosomes from SKOV3-IP1, HeyA8, and HeyA8-MDR cells. The high expression of miR-940 is associated with better survival in patients with ovarian serous cystadenocarcinoma. Ectopic expression of miR-940 inhibited proliferation, colony formation, invasion, and migration and triggered G0/G1 cell cycle arrest and apoptosis in OC cells. Overexpression of miR-940 also inhibited tumor cell growth in vivo. We showed that proto-oncogene tyrosine-protein kinase (SRC) is directly targeted by miR-940 and that miR-940 inhibited SRC expression at mRNA and protein levels. Following this inhibition, the expression of proteins downstream of SRC, such as FAK, paxillin and Akt was also reduced. Collectively, our results suggest that OC cells secrete the tumor-suppressive miR-940 into the extracellular environment via exosomes, to maintain their invasiveness and tumorigenic phenotype

CELLULAR AND MOLECULAR MECHANISMS REGULATING POSTNATAL DEVELOPMENT OF THE UTERUS

The uterus is essential for mammalian reproduction as it provides environment for conceptus implantation and subsequent development. Morphogenesis of the uterus is initiated in fetal life, however tissue-specific histoarchitecture is only completed postnatally including formation of endometrial glands. Endometrial glands synthesize and secrete or transport substances critical for conceptus survival and development; however, little is known about the mechanisms regulating development as well as adult function of the uterine glands. Aims of this dissertation included: identification of biological pathways involved in postnatal uterine morphogenesis and determination of the role of uterine glands in establishment of pregnancy in mice. Those objectives were addressed by: (1) conditional deletion of fibroblast growth factor receptor two (FGFR2) in the mouse uterus; (2) generation of a progesterone-induced uterine gland knockout (PUGKO) mouse model and investigation of mechanisms that underlie their infertility; (3) transcriptome profiling of microdissected luminal (LE) and glandular epithelial (GE) cells of the developing neonatal and adult mouse uterus; and (4) identification of molecular networks regulated by forkhead transcription factor box A2 (FOXA2) in the neonatal and adult uterine glands. Results of these studies established that: (1) Fgfr2 is dispensable for proper differentiation of uterine glands but plays a role in LE integrity in the adult; (2) neonatal progesterone exposure can be used as an effective method to discover pathways regulating endometrial adenogenesis and function; (3) uterine glands and their secretions have important biological roles in development of other cell types in the uterus and can affect blastocyst implantation and stromal cell decidualization; (4) FOXA2 positively regulates signaling pathways that are essential for glandular epithelial cell differentiation and development, whereas it may suppress pathways that could trigger carcinogenesis. Collectively, results of these studies provide a framework for elaborating gene regulatory networks governing development and function of uterine epithelia. Knowledge of those networks is required to facilitate the discovery of biomarkers to diagnose and treat uterine-based infertility and uterine diseases including endometriosis and endometrial cancer

Fibroblast Growth Factor Receptor Two (FGFR2) Regulates Uterine Epithelial Integrity and Fertility in Mice

Abstract Fibroblast growth factors (FGFs) and their receptors (FGFRs) regulate luminal epithelial (LE) cell proliferation in the adult mouse uterus. This study tested the hypothesis that FGFR2 has a biological role in postnatal development and function of the uterus by conditionally deletingFgfr2 after birth using progesterone receptor (Pgr)-Cre mice. AdultFgfr2 mutant female mice were initially subfertile and became infertile with increasing parity. No defects in uterine gland development were observed in conditionalFgfr2 mutant mice. In the adult,Fgfr2 mutant mice possessed a histologically normal reproductive tract with the exception of the uterus. The LE of theFgfr2 mutant uterus was stratified, but no obvious histological differences were observed in the glandular epithelium, stroma, or myometrium. Within the stratified LE, cuboidal basal cells were present and positive for basal cell markers (KRT14 and TRP63). Nulliparous bredFgfr2 mutants contained normal numbers of blastocysts on Day 3.5 postmating, but the number of embryo implantation sites was substantially reduced on Day 5.5 postmating. These results support the idea that loss of FGFR2 in the uterus after birth alters its development, resulting in LE stratification and peri-implantation pregnancy loss

Exosomal Non-Coding RNAs: Diagnostic, Prognostic and Therapeutic Applications in Cancer

Non-coding RNAs, such as microRNAs and long non-coding RNAs, are important regulatory molecules which are corrupted in cancer, often in a tissue and stage specific manner. Accumulated data suggests that these promising biomarkers, may also form the basis of novel targeted therapeutic strategies. The role of exosomes in cancer development and metastasis pathways is also increasingly well described. These endosome derived extracellular vesicles which are trafficked horizontally between tumor cells, and vertically between tumor cells and the surrounding microenvironment, carry bioactive cargos, which can reprogram the phenotype of recipient cells with important oncogenic consequences. Exosomes are enriched with non-coding RNA content. Within exosomes, non-coding RNAs are secreted into the peripheral circulation and other bodily fluids where they are protected from enzymatic degradation by the surrounding phospholipid membrane. Exosomes are therefore a highly promising source of diagnostic and prognostic material in cancer. Furthermore, as exosomes are natural ncRNA carriers, they may be adapted for the purpose of drug delivery by the introduction of exogenous ncRNAs or by manipulating their endogenous ncRNA content. In the current review, we will explore these highly clinically relevant themes by examining the roles of exosomal ncRNAs in cancer diagnostics, prognostics and therapy