Differential expression and release of exosomal miRNAs by human islets under inflammatory and hypoxic stress

Abstract

Aims/hypothesis Pancreatic islets produce non-coding microRNAs (miRNAs) that regulate islet cell function and survival. Our earlier investigations revealed that human islets undergo significant damage due to various types of stresses following transplantation and release miRNAs. Here, we sought to identify and validate exosomal miRNAs (exo-miRNAs) produced by human islets under conditions of cellular stress, preceding loss of cell function and death. We also aimed to identify islet stress signalling pathways targeted by exo-miRNAs to elucidate potential regulatory roles in islet cell stress. Methods Human islets were subjected to proinflammatory cytokine and hypoxic cell stress and miRNA from exosomes was isolated for RNA sequencing and analysis. Stress-induced exo-miRNAs were evaluated for kinetics of expression and release by intact islets for up to 48 h exposure to cytokines and hypoxia. A subset of stress-induced exo-miRNAs were assessed for recovery and detection as biomarkers of islet cell stress in a diabetic nude mouse xenotransplant model and in patients undergoing total pancreatectomy with islet auto-transplantation (TPIAT). Genes and signalling pathways targeted by stress-induced exo-miRNAs were identified by Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and direct interactions of miRNAs with downstream signalling targets were validated in human islet cells using the miRNA Tests for Read Analysis and Prediction (MirTrap) system. Results Global exo-miRNA sequencing revealed that 879 miRNA species were released from human islets and 190 islet exo-miRNAs were differentially expressed in response to proinflammatory cytokines, hypoxia or both. Release of exo-miRNAs hsa-miR-29b-3p and hsa-miR-216a-5p was detected within 6 h of exposure to cytokines and hypoxia. The remaining subset of stressinduced exo-miRNAs, including hsa-miR-148a-3p and islet cell damage marker hsa-miR-375, showed delayed release at 24-48 h, correlating with apoptosis and cell death. Stress and damage exo-miRNAs were significantly elevated in the circulation in human-to-mouse xenotransplant models and in human transplant recipients. Elevated blood exo-miRNAs negatively correlated with post-transplant islet function based on comparisons of stress and damage exo-miRNA indices with Secretory Unit of Islet Transplant Objects (SUITO) indices. KEGG analysis and further validation of exo-miRNA targets by MirTrap analysis revealed significant enrichment of islet mRNAs involved in phosphoinositide 3-kinase/Akt and mitogen-activated protein kinase signalling pathways. Conclusions/interpretation The study identifies exo-miRNAs differentially expressed and released by islets in response to damage and stress. These exo-miRNAs could serve as potential biomarkers for assessing islet damage and predicting outcomes in islet transplantation. Notably, exo-miRNAs 29b-3p and 216a-5p could be detected in islets prior to damage-released miRNAs and indicators of cellular apoptosis and death. Thus, these stress-induced exo-miRNAs may have potential diag...

MicroRNA Signatures as Future Biomarkers for Diagnosis of Diabetes States

“…These conflicting observations are not surprising for at least three reasons: (1) the bulk of beta-cell death in T1D precedes clinical diagnosis, and hence any increase in circulating miR-375 is possibly missed at the time of sampling; (2) the degree, extent, and dynamics of beta-cell death differ between individuals; and (3) islet compensatory mechanisms and ongoing autoimmunity may influence circulating levels even after disease diagnosis. For instance, our observations in mice suggest that miR-375 levels are significantly elevated in circulation immediately (3-6 h) after streptozotocin administration, subsiding to undetectable levels within 24 h. In the context of islet transplantation, we observed that miR-375 levels increase markedly during islet autotransplantation in patients receiving total pancreatectomy with islet auto transplantation, but normalized to baseline levels 7 days after transplantation [35]. In mice, we also observed elevated miR-375 levels 24 h after human islet transplantation, indicating islet inflammation and damage in the peritransplant period [35].…”

“…For instance, our observations in mice suggest that miR-375 levels are significantly elevated in circulation immediately (3-6 h) after streptozotocin administration, subsiding to undetectable levels within 24 h. In the context of islet transplantation, we observed that miR-375 levels increase markedly during islet autotransplantation in patients receiving total pancreatectomy with islet auto transplantation, but normalized to baseline levels 7 days after transplantation [35]. In mice, we also observed elevated miR-375 levels 24 h after human islet transplantation, indicating islet inflammation and damage in the peritransplant period [35]. Thus, miR-375 is a useful marker for islet cell damage during transplantation.…”

MicroRNA-29b-3p Promotes Human Retinal Microvascular Endothelial Cell Apoptosis via Blocking SIRT1 in Diabetic Retinopathy

“…Overexpression of miR-29b increased the expression of cleaved caspase-3, which is a typical apoptotic factor (Yuan et al, 2018). Saravanan et al (2019) reported that miR-29b-3p was selectively released in exosomes from inflammation and hypoxia induced islets before apoptosis and cell death, which coincided with activation of endoplasmic reticulum (ER) stress response markers IRE-1α, XBP1, HIF-1α, and CHOP. They also detected the selective release of miR-29b-3p in plasma exosomes after these cells were transplanted into streptozotocin (STZ) diabetic nude mice.…”

“…They also detected the selective release of miR-29b-3p in plasma exosomes after these cells were transplanted into streptozotocin (STZ) diabetic nude mice. Therefore, miR-29b-3p might be an early indicator of human islet cell apoptosis during prediabetic conditions (Saravanan et al, 2019). MiR-29b-3p was obviously upregulated in high glucose-induced endothelial cells (Silambarasan et al, 2016), and overexpression of miR-29b-3p could cause insulin resistance in mice (Su et al, 2019).…”

Preeclampsia: The Interplay between Oxygen-Sensitive miRNAs and Erythropoietin

“…The increased secretion of extracellular vesicles of trophoblastic origin containing miRNAs into the maternal blood flow due to placental dysfunction in PE can contribute to systemic biological changes. Among the miRNAs circulating in blood plasma, we identified (let-7g-5p, miR-26a-5p, miR-28 (-28-3p, -151a-3p), -30a-5p, -148a-3p, -181 (-181a-5p), -183-5p, -186-5p, -188 (-532-5p), -192-5p, and -320 (-320a), which change their expression in response to oxygen deprivation [18][19][20], oxidative stress (miR-140-3p, -143-3p, -378a-3p) [46], and are associated with immune and inflammatory responses (miR-375) [47], as well as ischemia/reperfusion (miR-423-3p) [48]. In general, our results are consistent with the data of other authors.…”