Whole Blood Holding Time Prior to Plasma Processing Alters microRNA Expression Profile

MicroRNAs (miRNAs) can exhibit aberrant expression under different physiological and pathological conditions. Therefore, differentially expressed circulating miRNAs have been a focus of biomarker discovery research. However, the use of circulating miRNAs comes with challenges which may hinder the reliability for their clinical application. These include varied sample collection protocols, storage times/conditions, sample processing and analysis methods. This study focused on examining the effect of whole blood holding time on the stability of plasma miRNA expression profiles. Whole blood samples were collected from healthy pregnant women and were held at 4°C for 30 min, 2 h, 6 h or 24 h prior to processing for plasma isolation. Plasma RNA was extracted and the expression of 179 miRNAs were analyzed. Unsupervised principal component analysis demonstrated that whole blood holding time was a major source of variation in miRNA expression profiles with 53 of 179 miRNAs showing significant changes in expression. Levels of specific miRNAs previously reported to be associated with pregnancy-associated complications such as hsa-miR-150-5p, hsa-miR-191-5p, and hsa-miR-29a-3p, as well as commonly used endogenous miRNA controls, hsa-miR-16-5p, hsa-miR-25-3p, and hsa-miR-223-3p were significantly altered with increase in blood holding time. Current protocols for plasma-based miRNA profiling for diagnostics describe major differences in whole blood holding periods ranging from immediately after collection to 26 h after. Our results demonstrate holding time can have dramatic effects on analytical reliability and reproducibility. This highlights the importance of standardization of blood holding time prior to processing for plasma in order to minimize introduction of non-biological variance in miRNA profiles.

Circulating miRNAs in Type 2 Diabetic Patients with and without Albuminuria in Malaysia

<b><i>Introduction:</i></b> Diabetic kidney disease (DKD) remains the leading cause of chronic kidney disease. Dysregulation of circulating miRNAs has been reported, suggesting their pathological roles in DKD. This study aimed to investigate differentially expressed miRNAs in the sera of type 2 diabetes mellitus (T2DM) patients with and without albuminuria in a selected Malaysian population. <b><i>Method:</i></b> Forty-one T2DM patients on follow-up at a community clinic were divided into normo-(NA), micro-(MIC), and macroalbuminuria (MAC) groups. Differential levels of miRNAs in 12 samples were determined using the pathway-focused (human fibrosis) miScript miRNA qPCR array and was validated in 33 samples, using the miScript custom qPCR array (CMIHS02742) (Qiagen GmbH, Hilden, Germany). <b><i>Results:</i></b> Trends of upregulation of 3 miRNAs in the serum, namely, miR-874-3p, miR-101-3p, and miR-145-5p of T2DM patients with MAC compared to those with NA. Statistically significant upregulation of miR-874-3p (<i>p</i> = 0.04) and miR-101-3p (<i>p</i> = 0.01) was seen in validation cohort. Significant negative correlations between the estimated glomerular filtration rate (eGFR) and miR-874-3p (<i>p</i> = 0.05), miR-101-3p (<i>p</i> = 0.03), and miR-145-5p (<i>p</i> = 0.05) as well as positive correlation between miR-874-3p and age (<i>p</i> = 0.03) were shown by Pearson’s correlation coefficient analysis. <b><i>Conclusion:</i></b> Upregulation of previously known miRNA, namely, miR-145-5p, and possibly novel ones, namely, miR-874-3p and miR-101-3p in the serum of T2DM patients, was found in this study. There was a significant correlation between the eGFR and these miRNAs. The findings of this study have provided encouraging evidence to further investigate the putative roles of these differentially expressed miRNAs in DKD.

A Differential Signature of Circulating miRNAs and Cytokines Between COVID-19 and Community-Acquired Pneumonia Uncovers Novel Physiopathological Mechanisms of COVID-19

Coronavirus Disease 2019 (COVID-19) pneumonia is a life-threatening infectious disease, especially for elderly patients with multiple comorbidities. Despite enormous efforts to understand its underlying etiopathogenic mechanisms, most of them remain elusive. In this study, we compared differential plasma miRNAs and cytokines profiles between COVID-19 and other community-acquired pneumonias (CAP). A first screening and subsequent validation assays in an independent cohort of patients revealed a signature of 15 dysregulated miRNAs between COVID-19 and CAP patients. Additionally, multivariate analysis displayed a combination of 4 miRNAs (miR-106b-5p, miR-221-3p, miR-25-3p and miR-30a-5p) that significantly discriminated between both pathologies. Search for targets of these miRNAs, combined with plasma protein measurements, identified a differential cytokine signature between COVID-19 and CAP that included EGFR, CXCL12 and IL-10. Significant differences were also detected in plasma levels of CXCL12, IL-17, TIMP-2 and IL-21R between mild and severe COVID-19 patients. These findings provide new insights into the etiopathological mechanisms underlying COVID-19.

Bone-Regulating MicroRNAs and Resistance Exercise: A Mini-Review

MicroRNAs (miRNA) are a class of short noncoding RNA that play important roles in controlling gene expression. Many miRNAs have been identified as being important regulators of bone cell function, thus affecting the bone remodeling processes. In addition to being expressed in specific tissues and exerting intracellular effects, miRNAs can enter the blood where they can be taken up by other tissues. These circulating miRNAs (c-miRNA) also have clinical significance as biomarkers of musculoskeletal diseases as they are tissue-specific, are stable and easily detectable, and require minimally invasive procedures. This mini-review discusses miRNAs with regulatory roles in bone metabolism and c-miRNA responses to acute bouts of resistance exercise. MiRNA responses (e.g., upregulation/downregulation of expression) vary depending on the resistance exercise protocol characteristics and the age of the participants. There are gaps in the literature that need to be addressed as most of the resistance exercise studies focused on miRNAs that regulate skeletal muscle in male participants.

The Landscape of Circulating miRNAs in the Post-Genomic Era

In the past decade, there has been an epochal change in the way that diseases are investigated and diagnosed [...]

Insight into microRNAs-Mediated Communication between Liver and Brain: A Possible Approach for Understanding Acute Liver Failure?

Acute liver failure (ALF) is a life-threatening consequence of hepatic function rapid loss without preexisting liver disease. ALF may result in a spectrum of neuropsychiatric symptoms that encompasses cognitive impairment, coma, and often death, collectively defined as acute hepatic encephalopathy (HE). Micro RNAs are small non-coding RNAs that modulate gene expression and are extensively verified as biomarker candidates in various diseases. Our systematic literature review based on the last decade’s reports involving a total of 852 ALF patients, determined 205 altered circulating miRNAs, of which 25 miRNAs were altered in the blood, regardless of study design and methodology. Selected 25 miRNAs, emerging predominantly from the analyses of samples obtained from acetaminophen overdosed patients, represent the most promising biomarker candidates for a diagnostic panel for symptomatic ALF. We discussed the role of selected miRNAs in the context of tissue-specific origin and its possible regulatory role for molecular pathways involved in blood–brain barrier function. The defined several common pathways for 15 differently altered miRNAs were relevant to cellular community processes, indicating loss of intercellular, structural, and functional components, which may result in blood-brain barrier impairment and brain dysfunction. However, a causational relationship between circulating miRNAs differential expression, and particular clinical features of ALF, has to be demonstrated in a further study.

The Association between Bisphenol A, Steroid Hormones, and Selected MicroRNAs Levels in Seminal Plasma of Men with Infertility

Bisphenol A (BPA), the most common endocrine-disrupting chemical, has been associated with male reproductive dysfunctions. Recently, it has been shown that BPA may also affect miRNAs expression. Herein, we aimed to evaluate the association of BPA levels with steroid hormone concentration and circulating miRNAs levels to investigate the potential direct effect of BPA on homeostasis in the testis environment. The level of BPA in the seminal plasma of azoospermic men was significantly higher compared to the healthy control. The concentrations of estradiol (E2) and androstenedione (A) were significantly decreased in the seminal plasma of azoospermic men compared to the normospermic men. The levels of miR-let-7a, miR-let-7b, and miR-let-7c were significantly up-regulated, and the level of miR-518f was significantly down-regulated in the seminal plasma of the azoospermic men compared to the healthy control. The level of BPA correlated negatively with sperm concentration and normal semen morphology. A significant positive correlation was found between BPA levels and miR-let-7a and miR-let-7c levels, whereas BPA negatively correlated with miR-518f levels. Our results suggest that BPA may negatively affect sperm quality. Moreover, BPA correlated with the miR-let-7a, miR-let-7c, and miR-518f levels in seminal plasma, which suggests that BPA may act directly in seminal plasma, affecting the testicular environment.