Small RNA Sequencing across Diverse Biofluids Identifies Optimal Methods for exRNA Isolation.

Poor reproducibility within and across studies arising from lack of knowledge regarding the performance of extracellular RNA (exRNA) isolation methods has hindered progress in the exRNA field. A systematic comparison of 10 exRNA isolation methods across 5 biofluids revealed marked differences in the complexity and reproducibility of the resulting small RNA-seq profiles. The relative efficiency with which each method accessed different exRNA carrier subclasses was determined by estimating the proportions of extracellular vesicle (EV)-, ribonucleoprotein (RNP)-, and high-density lipoprotein (HDL)-specific miRNA signatures in each profile. An interactive web-based application (miRDaR) was developed to help investigators select the optimal exRNA isolation method for their studies. miRDar provides comparative statistics for all expressed miRNAs or a selected subset of miRNAs in the desired biofluid for each exRNA isolation method and returns a ranked list of exRNA isolation methods prioritized by complexity, expression level, and reproducibility. These results will improve reproducibility and stimulate further progress in exRNA biomarker development.

exRNA Atlas Analysis Reveals Distinct Extracellular RNA Cargo Types and Their Carriers Present across Human Biofluids.

To develop a map of cell-cell communication mediated by extracellular RNA (exRNA), the NIH Extracellular RNA Communication Consortium created the exRNA Atlas resource (https://exrna-atlas.org). The Atlas version 4P1 hosts 5,309 exRNA-seq and exRNA qPCR profiles from 19 studies and a suite of analysis and visualization tools. To analyze variation between profiles, we apply computational deconvolution. The analysis leads to a model with six exRNA cargo types (CT1, CT2, CT3A, CT3B, CT3C, CT4), each detectable in multiple biofluids (serum, plasma, CSF, saliva, urine). Five of the cargo types associate with known vesicular and non-vesicular (lipoprotein and ribonucleoprotein) exRNA carriers. To validate utility of this model, we re-analyze an exercise response study by deconvolution to identify physiologically relevant response pathways that were not detected previously. To enable wide application of this model, as part of the exRNA Atlas resource, we provide tools for deconvolution and analysis of user-provided case-control studies.

Hypoadrenocorticism in a Dog Following Recovery from Alpha-Amanitin Intoxication.

A 10-year-old, female spayed Labrador Retriever was referred for acute hepatopathy and urinary retention. Blood work from the initial presentation (day 0) revealed a severe, mixed hepatopathy. Over the course of the patient's hospitalization, the patient developed liver insufficiency. Urine was submitted for toxicological screening and revealed detection of a trace concentration of alpha-amanitin. The patient was treated supportively for alpha-amanitin intoxication and was discharged from the hospital on day 8, with most biochemical parameters being markedly improved. The patient was persistently hyporexic at the time of discharge. On day 15, at a recheck appointment, the patient had lost 2.4 kg and liver enzymology revealed improved values. On day 24, the patient was presented for anorexia and vomiting and had lost another 2.3 kg. Blood work and endocrinological testing at that time were consistent with hypoadrenocorticism. The patient was started on glucocorticoids and mineralocorticoids. At day 106, the patient was doing well clinically while receiving monthly mineralocorticoids and daily glucocorticoids. This case report is the first to describe the chronological association between alpha-amanitin-induced liver dysfunction and the subsequent development of adrenal insufficiency in a dog.