Exosomes, which are one of the smallest extracellular vesicles released from cells, have been shown to carry different nucleic acids, including microRNAs (miRNAs). miRNAs significantly regulate cell ...growth and metabolism by posttranscriptional inhibition of gene expression. The rapidly changing understanding of exosomes' formation and function in delivering miRNAs from cell to cell has prompted us to review current knowledge in exosomal miRNA secretion mechanisms as well as possible therapeutic applications for personalized medicine.
Cellular replacement therapy has emerged as a novel strategy for the treatment of heart failure. The aim of our study was to determine the fate of injected mesenchymal stem cells (MSCs) and whole ...bone marrow (BM) cells in the infarcted heart. MSCs were purified from BM of transgenic mice and characterized using flow cytometry and in vitro differentiation assays. Myocardial infarctions were generated in mice and different cell populations including transgenic MSCs, unfractionated BM cells, or purified hematopoietic progenitors were injected. Encapsulated structures were found in the infarcted areas of a large fraction of hearts after injecting MSCs (22 of 43, 51.2%) and unfractionated BM cells (6 of 46, 13.0%). These formations contained calcifications and/or ossifications. In contrast, no pathological abnormalities were found after injection of purified hematopoietic progenitors (0 of 5, 0.0%), fibroblasts (0 of 5, 0.0%), vehicle only (0 of 30, 0.0%), or cytokine-induced mobilization of BM cells (0 of 35, 0.0%). We conclude that the developmental fate of BM-derived cells is not restricted by the surrounding tissue after myocardial infarction and that the MSC fraction underlies the extended bone formation in the infarcted myocardium. These findings seriously question the biologic basis and clinical safety of using whole BM and in particular MSCs to treat nonhematopoietic disorders.
NF-κB signal transduction is a potential therapeutic target in many malignant tumors. We have recently shown, in malignant renal proximal tumor cells, that a transcription complex, consisting of ...NF-κB p65 and mitogen-activated protein kinase p38α, joined by protein kinase C (PKC) α, transmigrates into the nucleus. There, PKCα suppresses the nuclear release of primary microRNA (pri-miRNA) 15a. Induced by endothelin (ET)-1, a decrease in PKCα levels leads to increased miRNA 15a (miR-15A) expression. An identical system can be identified in renal carcinomas, in which, after nuclear transmigration, PKCα binds directly to pri-miRNA 15a in the nucleus. However, the pattern of PKC isoforms differs between malignant renal cell carcinoma (RCC) and benign oncocytoma. PKCα, a component of the transcription complex in tumors, is up-regulated in benign oncocytoma but down-regulated in RCC. Conversely, miRNA 15a is up-regulated in RCC and down-regulated in oncocytoma. A similar behavior is observed in chromophobe carcinoma, whereas differences are less pronounced in papillary RCC (type I): NF-κB target gene expression (ie, ET-1, ET-A and ET-B receptors, vascular cell adhesion molecule-1, IL-6, and fractalkine) is particularly high in malignant RCCs. Up-regulated miRNA 15a can be measured in urine from patients with RCC but is nearly undetectable in oncocytoma, other tumors, and urinary tract inflammation. Thus, the up-regulation of miRNA 15a may be an important marker to help identify malignant clear-cell RCCs in both biopsy and urine samples.
Circulating biomarkers such as microRNAs (miRNAs), short noncoding RNA strands, represent prognostic and diagnostic indicators for a variety of physiological disorders making their detection and ...quantification an attractive approach for minimally invasive early disease diagnosis. However, highly sensitive and selective detection methods are required given the generally low abundance of miRNAs in body fluids together with the presence of large amounts of other potentially interfering biomolecules. Although a variety of miRNA isolation and detection methods have been established in clinics, they usually require trained personnel and often constitute labor-, time- and cost-intensive approaches. During the last years, nanoparticle-based biosensors have received increasing attention due to their superior detection efficiency even in very low concentration regimes. This is based on their unique physicochemical properties in combination with their high surface area that allows for the immobilization of multiple recognition sites resulting in fast and effective recognition of analytes. Among various materials, magnetic nanoparticles have been identified as useful tools for the separation, concentration, and detection of miRNAs. Here, we review state-of-the-art technology with regard to magnetic particle-based miRNA detection from body fluids, critically discussing challenges and future perspective of such biosensors while comparing their handling, sensitivity as well as selectivity against the established miRNA isolation and detection methods.
This review highlights new developments in miRNA as diagnostic and surveillance tools in diseases damaging the renal proximal tubule mediated by endothelin in the field of renal carcinoma, ...proteinuric kidney disease and tubulotoxicity. A new mechanism in the miRNA regulation of proteins leads to the binding of the miRNA directly to the DNA with premature transcriptional termination and hence the formation of truncated protein isoforms (Mxi2, Vim3). These isoforms are mediated through miRNA15a or miRNA 498, respectively. ET‐1 can activate a cytoplasmic complex consisting of NF‐κB p65, MAPK p38α, and PKCα. Consequently, PKCα does not transmigrate into the nucleus, which leads to the loss of suppression of a primiRNA15a, maturation of this miRNA in the cytoplasm, tubular secretion and detectability in the urine. This mechanism has been shown in renal cell carcinoma and in proteinuric disease as a biomarker for the activation of the signalling pathway. Similarly, ET‐1 induced miRNA 498 transmigrates into the nucleus to form the truncated protein Vim3, which is a biomarker for the benign renal cell tumour, oncocytoma. In tubulotoxicity, ET‐1 induced miRNa133a down‐regulating multiple‐drug‐resistant related protein‐2, relevant for proteinuric and cisplatin/cyclosporine A toxicity. Current advantages and limitations of miRNAs as urinary biomarkers are discussed.
Review: This review highlights the use of miRNAs as urinary markers for proximal tubular damage via proteinuria, and in renal cancer (miRNA15a) and in nephrotoxicity (miRNA 133a). All three conditions exert their effect by endothelin‐1 stimulation leading to a specific signalling cascade. In nephrotoxicity, the classical protein repression of an ATP‐binding cassette (ABC) transporter, the multidrug resistance associated protein 2 (MRP2), through miRNA133a leads to tubular damage by retention of CyA and Cisplatin. In proteinuria and in renal cancer, protein kinase C α is down‐regulated and no longer able to suppress miRNA15a maturation. This is achieved by a novel mechanism: miRNA re‐entering the nucleus, binding directly to DNA. This leads to premature transcriptional termination, forming the truncated gene, Mxi‐2, which further accelerates miRNA 15a maturation. Meanwhile, the activation of NF‐κB is ongoing unrestrictedly.
Cellular cardiomyoplasty is an attractive option for the treatment of severe heart failure. It is, however, still unclear and controversial which is the most promising cell source. Therefore, we ...investigated and examined the fate and functional impact of bone marrow (BM) cells and embryonic stem cell (ES cell)-derived cardiomyocytes after transplantation into the infarcted mouse heart. This proved particularly challenging for the ES cells, as their enrichment into cardiomyocytes and their long-term engraftment and tumorigenicity are still poorly understood. We generated transgenic ES cells expressing puromycin resistance and enhanced green fluorescent protein cassettes under control of a cardiac-specific promoter. Puromycin selection resulted in a highly purified (>99%) cardiomyocyte population, and the yield of cardiomyocytes increased 6-10-fold because of induction of proliferation on purification. Long-term engraftment (4-5 months) was observed when co-transplanting selected ES cell-derived cardiomyocytes and fibroblasts into the injured heart of syngeneic mice, and no teratoma formation was found (n = 60). Although transplantation of ES cell-derived cardiomyocytes improved heart function, BM cells had no positive effects. Furthermore, no contribution of BM cells to cardiac, endothelial, or smooth muscle neogenesis was detected. Hence, our results demonstrate that ES-based cell therapy is a promising approach for the treatment of impaired myocardial function and provides better results than BM-derived cells.
To date, there are no preoperative and quantitative dynamics in clinical practice that can reliably differentiate between a benign and malignant renal cell carcinoma (RCC). For monitoring different ...analytes in body fluids, more than 40 different molecular biomarkers have been identified, however, they are associated with limited clinical sensitivity and/or non-optimal specificity due to their leaky nature. Previous work on RCC demonstrated the miRNA15a to be reliable and novel biomarker with 98.1% specificity and 100% sensitivity. Despite the high potential of miRNA15a biomarker, its clinical application is considerably hampered by the insensitive nature of the detection methods and low concentration of biomarker in samples that is aggravated by the high level of contamination due to other solutes present in body fluids. In this work, a non-invasive quantitative approach is demonstrated to overcome such diagnostics issues through biotin-streptavidin binding and fluorescence active magnetic nanocarriers that ensured prompt isolation, enrichment and purification of the biomarker miRNA15a from urine. The study demonstrates that detectable low levels of these miRNAs through miRNA capturing nanocarriers can potentially function as advanced diagnostic markers for the non-invasive investigation and early detection of renal cancer.
The dynamic interplay between metabolism and immune responses in health and disease, by which different immune cells impact on metabolic processes, are being increasingly appreciated. However, the ...potential of master regulators of metabolism to control innate immunity are less understood. Here, we studied the cross-talk between leptin signaling and macrophage function in the context of bacterial infections. We found that upon infection with Gram-negative pathogens, such as Salmonella Typhimurium, leptin receptor (Lepr) expression increased in both mouse and human macrophages. Unexpectedly, both genetic Lepr ablation in macrophages and global pharmacologic leptin antagonization augmented lysosomal functions, reduced S. Typhimurium burden, and diminished inflammation in vitro and in vivo. Mechanistically, we show that leptin induction activates the mTORC2/Akt pathway and subsequently down-regulates Phlpp1 phosphatase, allowing for phosphorylated Akt to impair lysosomal-mediated pathogen clearance. These data highlight a link between leptin signaling, the mTORC2/Phlpp1/Akt axis, and lysosomal activity in macrophages and have important therapeutic implications for modulating innate immunity to combat Gram-negative bacterial infections.
MicroRNAs (miRNAs) are small non-coding RNA molecules involved in the expressional regulation of genes by inhibiting gene translation. MicroRNAs are recruited and incorporated into the miRISC, ...ribonucleoprotein complex, targeting specific mRNAs through mechanisms specific for a miRNA sequence. Here we review the biogenesis, regulation, and monitoring of miRNAs, as well as the current evidence for potential roles of miRNAs in human diseases associated with activation of the endothelin system. These diseases include cancer, kidney disease, cardiovascular diseases, inflammatory diseases, infectious diseases, and blood diseases, that may all be aggravated by aberrant miRNA expression. In this review we will also discuss regulatory mechanisms determining production of miRNA as well as measuring or targeting miRNAs as potential novel approaches for diagnosis and treatment. Targeting miRNAs possibly will allow one to detect diseases or to interfere with the progression of diseases associated with activation of the endothelin system.
Principles of positive and negative miRNA biogenesis. On the left hand (blue site) all proteins visualized bind in the nucleus to the pri-miRNA and therefore inhibit the processing step to the pre-miRNA.
On the right hand (red site) all proteins listed bind to the pri-miRNA and promote the processing to pre-miRNA. Due to the binding of Exportin-5 the pre-miRNA migrates to the cytoplasm where a further processing step takes place. The double stranded pre-miRNA is separated by a processing complex. One strand (indicated by miRNA*) is degraded whereas the other is either important for mRNA cleavage or P-body storage of the target mRNAs. Display omitted
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