Search

Background: Mesenteric lymph (ML) has been implicated in the development of multiple organ dysfunction syndrome in critical illness. Extracellular RNAs play a role in cell-to-cell communication during physiological and disease processes but they are rarely studied in ML. We aimed at examining the RNA profiles of peripheral plasma, ML, and ML's extracellular vesicle (ML-EV) and triglyceride-rich lipoprotein (ML-TRL) fractions, obtained from rodent models of critical illness. Methods and Results: We collected ML for 5 hours from rodent models of critical illness [Acute Pancreatitis, Cecal Ligation and Incision (CLI), Gut Ischemia-Reperfusion (IR)] and matching Sham control rats. ML-EV and ML-TRL fractions were also isolated. RNA sequencing was performed on the RNA extracted from ML, ML-EV, ML-TRL, and plasma by using the Ion Torrent Personal Genome Machine platform. RNA sequences were searched using the Basic Local Alignment Search Tool against rat genome and RefSeq, microRNA (miRNA), genomic tRNA, functional RNA, and Genbank nucleotide databases, and the read counts were analyzed. Each sample type had a distinct RNA profile. ML contained more RNA per volume and a larger proportion of tRNA fragments than plasma. ML-EVs were enriched with miRNA, whereas ML-TRLs contained low absolute amounts of RNA. The RNA size profiles for CLI and Gut IR were different from Sham. ML carried intestinal RNAs and in a CLI model it was significantly enriched with bacterial RNA sequences. Conclusions: We found the distinct but diverse RNA profiles of ML and its compartments, and their different profiles in critical illness. Intestinal-derived small RNAs in ML may have a direct role in critical illness and utility as potential biomarkers.

In human beings the immature brain is highly plastic and depending on the stage of gestation is particularly vulnerable to a range of insults that if sufficiently severe, can result in long-term motor, cognitive and behavioral impairment. With improved neonatal care, the incidence of major motor deficits such as cerebral palsy has declined with prematurity. Unfortunately, however, milder forms of injury characterized by diffuse non-cystic white matter lesions within the periventricular region and surrounding white matter, involving loss of oligodendrocyte progenitors and subsequent axonal hypomyelination as the brain matures have not. Existing therapeutic options for treatment of preterm infants have proved inadequate, partly owing to an incomplete understanding of underlying post-injury cellular and molecular changes that lead to poor neurodevelopmental outcomes. This has reinforced the need to improve our understanding of brain plasticity, explore novel solutions for the development of protective strategies, and identify biomarkers. Compelling evidence exists supporting the involvement of microRNAs (miRNAs), a class of small non-coding RNAs, as important post-transcriptional regulators of gene expression with functions including cell fate specification and plasticity of synaptic connections. Importantly, miRNAs are differentially expressed following brain injury, and can be packaged within exosomes/extracellular vesicles, which play a pivotal role in assuring their intercellular communication and passage across the blood–brain barrier. Indeed, an increasing number of investigations have examined the roles of specific miRNAs following injury and regeneration and it is apparent that this field of research could potentially identify protective therapeutic strategies to ameliorate perinatal brain injury. In this review, we discuss the most recent findings of some important miRNAs in relation to the development of the brain, their dysregulation, functions and regulatory roles following brain injury, and discuss how these can be targeted either as biomarkers of injury or neuroprotective agents. [ABSTRACT FROM AUTHOR]

Bacteria secrete RNAs, some of which have effects on other cells and on other species as signalling RNAs. Prokaryotic membrane vesicles (MVs) contain a range of RNA types. The MV structure offers protection from degradation as well as receptors to facilitate delivery to target cells. Microscopic imaging and molecular biology analyses have provided evidence to demonstrate that bacterial MVs deliver their RNA into eukaryotic cells. Moreover, in some cases the RNA cargo is demonstrably functional and phenotypic changes can be identified in MV-RNA treated target cells. The challenge now is to dissect the effect of MV-RNA on target cells away from the effects of non-RNA components of the MV such as lipopolysaccharide that can co-purify with RNA. [ABSTRACT FROM AUTHOR]

Background: Bacterium-to-host signalling during infection is a complex process involving proteins, lipids and other diffusible signals that manipulate host cell biology for pathogen survival. Bacteria also release membrane vesicles (MV) that can carry a cargo of effector molecules directly into host cells. Supported by recent publications, we hypothesised that these MVs also associate with RNA, which may be directly involved in the modulation of the host response to infection. Methods and Results: Using the uropathogenic Escherichia coli (UPEC) strain 536, we have isolated MVs and found they carry a range of RNA species. Density gradient centrifugation further fractionated and characterised the MV preparation and confirmed that the isolated RNA was associated with the highest particle and protein containing fractions. Using a new approach, RNA-sequencing of libraries derived from three different ‘size’ RNA populations (<50nt, 50-200nt and 200nt+) isolated from MVs has enabled us to now report the first example of a complete bacterial MV-RNA profile. These data show that MVs carry rRNA, tRNAs, other small RNAs as well as full-length protein coding mRNAs. Confocal microscopy visualised the delivery of lipid labelled MVs into cultured bladder epithelial cells and showed their RNA cargo labelled with 5-EU (5-ethynyl uridine), was transported into the host cell cytoplasm and nucleus. MV RNA uptake by the cells was confirmed by droplet digital RT-PCR of csrC. It was estimated that 1% of MV RNA cargo is delivered into cultured cells. Conclusions: These data add to the growing evidence of pathogenic bacterial MV being associated a wide range of RNAs. It further raises the plausibility for MV-RNA-mediated cross-kingdom communication whereby they influence host cell function during the infection process. [ABSTRACT FROM AUTHOR]

Background: The differential expression pattern of microRNAs (miRNAs) during mammary gland development might provide insights into their role in regulating the homeostasis of the mammary epithelium. Our aim was to analyse these regulatory functions by deriving a comprehensive tissue-specific combined miRNA and mRNA expression profile of post-natal mouse mammary gland development. We measured the expression of 318 individual murine miRNAs by bead-based flow-cytometric profiling of whole mouse mammary glands throughout a 16-point developmental time course, including juvenile, puberty, mature virgin, gestation, lactation, and involution stages. In parallel whole-genome mRNA expression data were obtained. Results: One third (n = 102) of all murine miRNAs analysed were detected during mammary gland development. MicroRNAs were represented in seven temporally co-expressed clusters, which were enriched for both miRNAs belonging to the same family and breast cancer-associated miRNAs. Global miRNA and mRNA expression was significantly reduced during lactation and the early stages of involution after weaning. For most detected miRNA families we did not observe systematic changes in the expression of predicted targets. For miRNA families whose targets did show changes, we observed inverse patterns of miRNA and target expression. The data sets are made publicly available and the combined expression profiles represent an important community resource for mammary gland biology research. Conclusion: MicroRNAs were expressed in likely co-regulated clusters during mammary gland development. Breast cancer-associated miRNAs were significantly enriched in these clusters. The mechanism and functional consequences of this miRNA co-regulation provide new avenues for research into mammary gland biology and generate candidates for functional validation. [ABSTRACT FROM AUTHOR]