Is RNA in Serum Bound to Nucleoprotein Complexes?

Sisco, Kenneth L.

doi:10.1093/clinchem/47.9.1744

Cited by 28 publications

(13 citation statements)

References 10 publications

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“…cirRNA in blood is protected from ribonucleases. For example, it was shown that cirRNA can be packaged in apoptotic bodies separately from DNA, 4 it can be contained within nucleoprotein complexes, 5 lipids, proteolipids, phospholipids, 6,7 proteins, 8 and it can be associated with circulating particles 9 . Such complexed forms of cirRNA provide stability of RNA in blood and can reflect the origin of specific RNA that depends on its intracellular localization and function.…”

Section: Introductionmentioning

confidence: 99%

Concentrations of Circulating RNA from Healthy Donors and Cancer Patients Estimated by Different Methods

Rykova

Wünsche

Brizgunova

et al. 2006

Annals of the New York Academy of Sciences

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Circulating RNA (cirRNA) was isolated from plasma and cell surface-bound fractions of blood of healthy women and breast cancer patients. RNA samples were DNase treated and quantified by a SYBR Green II assay. Concentrations of RNA sequences of GAPDH, Ki-67 mRNA, and 18S rRNA were measured by real-time quantitative PCR (RT-qPCR) after reverse transcription with random hexamer primers. The obtained data spread over three orders of magnitude for GAPDH and Ki-67 mRNA signals and two orders of magnitude for the copy number of 18S rRNA in blood fractions in both groups. In blood of healthy donors, no correlation was found between the copy number of GAPDH, Ki-67 mRNA, and 18S rRNA and RNA concentrations measured by the SYBR Green II assay. Within the group of breast cancer patients, the concentration GAPDH and Ki-67 mRNA correlated with the concentration of total RNA only in the cell surface-bound fraction; whereas the concentration of 18S rRNA correlated with total RNA in both, the cell surface-bound fraction and blood. The copy number of Ki-67 mRNA correlated with copy numbers of GAPDH mRNA in all fractions of cirRNA of healthy donors and breast cancer patients. A correlation between copy numbers of Ki-67 mRNA and 18S rRNA was found only in cell surface-bound fraction of breast cancer patients. The data described here demonstrate the necessity of searching for more suitable RNA markers in order to estimate total cirRNA concentrations by RT-qPCR, although mRNA of GAPDH could be used for normalization of the level of cancer-specific mRNA among patients.

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Section: Introductionmentioning

confidence: 99%

Concentrations of Circulating RNA from Healthy Donors and Cancer Patients Estimated by Different Methods

Rykova

Wünsche

Brizgunova

et al. 2006

Annals of the New York Academy of Sciences

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“…Because of the presence of potent ribonucleases, most investigators doubted that extracellular RNA could survive in the blood [30]. Up to now, many studies have documented circulating a large number of serum miRs remain stable and consistent in severe conditions [31][32][33][34]. It is very possible to detect serum miRs which help to diagnose this disease.…”

mentioning

confidence: 99%

Serum miR-21 may be a Potential Diagnostic Biomarker for Diabetic Nephropathy

Wang

Duan

Tian

et al. 2015

Exp Clin Endocrinol Diabetes

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MiRNAs play important roles in initiation and progress of many pathologic processes. MiR-21 was closely associated with diabetic nephropathy (DN). However, whether serum miR-21 was as a potential diagnostic biomarker for DN and the relationship between serum miR-21 and tissue miR-21 remained unclear. In this study, real-time RT-PCR, cell transfection, luciferase reporter gene assays, western blot and confocal microscope were used, respectively. Here, we found that serum and renal tissue miR-21 was significantly elevated with the progress of DN. Moreover, luciferase reporter gene assays showed that smad7 was a validated miR-21 target, cell transfection showed that miR-21 overexpression downregulated target smad7 expression. Interestingly, serum miR-21 was significantly consistent with the alterations of tissue miR-21 with the development of DN. In addition, serum miR-21 was also positively correlated with GBM, GA, ACR and CCF, while negatively correlated with Ccr. Importantly, antagomiR-21 not only alleviated GBM, GA, ACR and CCF, but also ameliorated Ccr by increasing target smad7. In conclusion, our data demonstrated that serum miR-21 was closely associated with renal structure and function, and serum miR-21 may be regarded as a potential diagnostic biomarker of DN.

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“…A possible reason for this may be that Tsui et al 6 looked at the housekeeping gene glyceraldehyde 3-phosphate dehydrogenase (GAPDH), which may be more stable than β-actin mRNA in plasma. Proposed mechanisms for the stability of RNA in cell-free plasma include formations of RNA with proteins/lipids, 1 RNA bound with DNA in nucleosomes, 7 and protection in apoptotic bodies. 8 Thus, our data indicate that mRNA for β-actin and GAPDH may be protected by different mechanisms.…”

Section: Resultsmentioning

confidence: 99%

Stability of β‐Actin mRNA in Plasma

Holford

Sandhu

Thakkar

et al. 2008

Annals of the New York Academy of Sciences

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It has long been thought that mRNA is labile and easily prone to degradation. However, a recent study demonstrated that GAPDH mRNA in cell-free plasma may remain stable up to 24 hours after blood collection. As there are no other independent studies, we attempted to reproduce the findings of that study. In our study, blood was collected from a healthy male volunteer into Vacutainer tubes containing EDTA. Blood samples were placed on ice and plasma separated by double-centrifugation at times 0, 1, 2, and 5 hours after blood collection. mRNA was extracted from four aliquots of the blood sample by means of the QIAamp Viral RNA kit. Extracted mRNA was converted to cDNA by reverse transcription before real time quantitative PCR measurement of the housekeeping beta-actin gene. Plasma beta-actin mRNA at 2 hours (0.012; 0.0031-0.0297, median and range) was significantly lower (P= 0.022) than at 0 hours (0.12; 0.057-0.165) (P= 0.016). The levels decreased further at 5 hours (0.0037; 0.0024-0.011) (P= 0.004). The results show that plasma beta-actin mRNA levels decrease with time after blood collection and that this is likely to be due to degradation in vitro.

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Is RNA in Serum Bound to Nucleoprotein Complexes?

Cited by 28 publications

References 10 publications

Concentrations of Circulating RNA from Healthy Donors and Cancer Patients Estimated by Different Methods

Concentrations of Circulating RNA from Healthy Donors and Cancer Patients Estimated by Different Methods

Serum miR-21 may be a Potential Diagnostic Biomarker for Diabetic Nephropathy

Stability of β‐Actin mRNA in Plasma

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