RESUMO
Background: Numerous changes in maternal physiology occur during pregnancy that are critical in controlling and maintaining the maternal metabolic adaptations and fetal development. The placenta is the key source through which the fetus receives nutrients, blood, and oxygen for growth. The human placenta releases several molecules into maternal circulation that include hormones, proteins, RNA, and DNA throughout the course of pregnancy. Additionally, extracellular vesicles (EVs) originating from the placenta have been found in the maternal circulation. Methods: In this review, we discuss the role of EVs in maternal-fetal communication during pregnancy. Results: EVs originating from the placenta can be divided into 3 categories based on their size and/or origin: exosomes (50 to 150 nm), microvesicles (nm to several µm), and apoptotic bodies or syncytial nuclear aggregates (>1 µm). The cellular microenvironment-such as oxygen tension and glucose concentration-have been found to control EV release from the placenta and their bioactivity on target cells. Furthermore, maternal EVs can stimulate cytokine release from endothelial cells and are involved in several physiologic and pathologic events in pregnancy. Conclusion: Exosomes provide a way to identify the function and metabolic state of cell origin through their ability to reflect the microenvironment that they are released from. Further understanding of how EVs regulate key events in pregnancy may help elucidate how maternal-fetal communication is established in both normal and pathologic conditions.
RESUMO
There is an emerging need for viral gene specific quantitative PCR (qPCR) assays that validate and complement whole transcriptome level technologies, including microarray and next generation sequencing. Therefore, a compilation of qPCR assays that represented the breadth of the entire Herpes simplex virus type 1 (HSV-1) genome were developed and evaluated. SYBR Green-I-based quantitation of each of the 74 HSV-1 lytic genes enabled accurate and reproducible detection of viral genes using a minimal number of reaction conditions. The amplification specificity of these assays for HSV-1 target genes was confirmed by amplicon size and purity determination on agarose gels, melt temperature dissociation curve analysis, and direct DNA sequencing of amplified products. Analysis of representative target genes demonstrated that these assays accurately and reproducibly quantified target gene expression across a wide and linear range of detection. In addition, minimal intra- and inter-assay variability was observed with significant well-to-well and plate-to-plate/assay-to-assay precision. To evaluate the utility of the developed qPCR assay system, kinetic profiles of viral gene expression were determined for an array of representative genes from all HSV-1 transcriptional gene classes. Collectively, these data demonstrate that the compiled optimized qPCR assays is a scalable and cost-effective method to assess HSV-1 gene expression with broad application potential, including investigation of pathogenesis and antiviral therapies. In addition, they can be employed to validate and complement evolving technologies for genome-wide transcriptome analysis.
