Plasma miRNA Expression Profiles Research Articles

Circulating miR-25-3p and miR-451a May Be Potential Biomarkers for the Diagnosis of Papillary Thyroid Carcinoma.

ObjectiveThere is no effective and reliable biomarker to distinguish benign thyroid nodules from papillary thyroid carcinomas (PTC). This study aimed at examining the levels of plasma miRNAs in patients with PTC or benign nodules to explore the potential miRNA biomarkers for PTC.Patients and MethodsGenome-wide plasma miRNA expression profiles were determined by the miRNA Microarray and the significantly higher levels of miRNAs were validated in plasma and tissues by quantitative RT-PCR. The levels of two miRNAs were further tested in seven patients before and after tumor excision and the potential values for the diagnosis of PTC were evaluated by receiver operating characteristic curve (ROC).ResultsIn comparison with that in the patients with benign nodules, eight significantly higher and three lower levels of plasma miRNAs were detected in the PTC patients. Further validation indicated that the levels of plasma miR-25-3p, miR-451a, miR-140-3p and let-7i were significantly higher in the PTC cases than in those with benign nodules or the healthy controls. Significantly higher levels of miR-25-3p and miR-451a were detected in the thyroid tissues from the PTC patients. The levels of plasma miR-25-3p and miR-451a in seven patients significantly decreased after tumor excision. ROC analyses revealed that the levels of plasma miR-25-3p at cut-off 1.41 and miR-451a at 1.38 had sensitivity of 92.8% and 88.9%, and specificity of 68.8% and 66.7% for distinguishing PTC from benign nodules, respectively.ConclusionOur findings suggest that the levels of plasma miR-25-3p and miR-451a may be valuable for the diagnosis of PTC.

Expression profile of plasma microRNAs in premature infants with respiratory distress syndrome.

As well-known regulators of gene expression, microRNAs (miRNAs) are important not only in cell proliferation and differentiation, but also in tumorigenesis and organ development. It has been estimated that miRNAs may be responsible for regulating the expression of almost one third of the human genome. Simultaneously, with advances in neonatal care in the clinic, an increased number of premature infants are being saved and, thus, respiratory distress syndrome (RDS) has become more common. However, previous non-miRNA studies have suggested their connection with RDS. In the present study, a miRNA microarray, including >1,891 capture probes was used to compared the expression profiles of plasma miRNAs between RDS and control groups. miRNAs, which were observed to have consistent fold-changes (fold-change ≥ 1.3) between the two groups were selected and validated using reverse transcription-quantitative polymerase chain reaction. As a result, 171 differentially expressed miRNAs were identified, including two upregulated and seven downregulated miRNAs. Of these miRNAs, four were selected as having higher fold-changes between the two groups. This is the first time, to the best of our knowledge, that these nine miRNAs have been reported in RDS. It was hypothesized that these novel miRNAs may be important in RDS, and may provide meaningful biomarkers for the diagnosis of RDS.

A five-microRNA panel in plasma was identified as potential biomarker for early detection of gastric cancer

Background:Circulating microRNAs (miRNAs) have been implicated as novel biomarkers for gastric cancer (GC) diagnosis. However, the mixture of GC subtypes may have led to the inconsistent circulating miRNA profiles, and the clinical performance of circulating miRNAs has not yet been evaluated independently on early detection of GC.Methods:A four-phase study was designed with a total of 160 cancer-free controls, 124 patients with gastric non-cardia adenocarcinoma (GNCA) and 36 patients diagnosed gastric cardia adenocarcinoma (GCA). In the discovery phase, we screened the miRNA expression profile in plasma of 40 GNCA patients (stage I) and 40 matched controls by TaqMan low density array (TLDA) chips with pooled samples. Differentially expressed miRNAs were further validated in individual sample using quantitative reverse-transcriptase PCR (qRT–PCR) in the training phase. Subsequently, in an independent validation phase, the identified miRNAs were evaluated in 48 GNCA patients (stage I) and 102 matched controls. Finally, the identified miRNAs were further assessed in an external validation phase including advanced GNCA and GCA patients. Additionally, the expression levels of identified miRNAs were measured in the media of BGC823 and MGC803 cell lines.Results:Five miRNAs (miR-16, miR-25, miR-92a, miR-451 and miR-486-5p) showed consistently elevated levels in plasma of the GC patients as compared with controls, and were identified to be potential markers for GNCA with area under the receiver operating characteristic (ROC) curves (AUCs) ranging from 0.850 to 0.925 and 0.694 to 0.790 in the training and validation phases, respectively. The five-miRNA panel presented a high diagnostic accuracy for the early-stage GNCA (AUCs=0.989 and 0.812 for the training and validation phases, respectively). Three miRNAs (miR-16, miR-25 and miR-92a) were excreted into the culture media of GC cell lines.Conclusions:The five-miRNA panel in plasma may serve as a potential non-invasive biomarker in detecting the early-stage GC.

Diagnostic Potential of Plasmatic MicroRNA Signatures in Stable and Unstable Angina

PurposeWe examined circulating miRNA expression profiles in plasma of patients with coronary artery disease (CAD) vs. matched controls, with the aim of identifying novel discriminating biomarkers of Stable (SA) and Unstable (UA) angina.MethodsAn exploratory analysis of plasmatic expression profile of 367 miRNAs was conducted in a group of SA and UA patients and control donors, using TaqMan microRNA Arrays. Screening confirmation and expression analysis were performed by qRT-PCR: all miRNAs found dysregulated were examined in the plasma of troponin-negative UA (n=19) and SA (n=34) patients and control subjects (n=20), matched for sex, age, and cardiovascular risk factors. In addition, the expression of 14 known CAD-associated miRNAs was also investigated.ResultsOut of 178 miRNAs consistently detected in plasma samples, 3 showed positive modulation by CAD when compared to controls: miR-337-5p, miR-433, and miR-485-3p. Further, miR-1, -122, -126, -133a, -133b, and miR-199a were positively modulated in both UA and SA patients, while miR-337-5p and miR-145 showed a positive modulation only in SA or UA patients, respectively. ROC curve analyses showed a good diagnostic potential (AUC ≥ 0.85) for miR-1, -126, and -483-5p in SA and for miR-1, -126, and -133a in UA patients vs. controls, respectively. No discriminating AUC values were observed comparing SA vs. UA patients. Hierarchical cluster analysis showed that the combination of miR-1, -133a, and -126 in UA and of miR-1, -126, and -485-3p in SA correctly classified patients vs. controls with an efficiency ≥ 87%. No combination of miRNAs was able to reliably discriminate patients with UA from patients with SA.ConclusionsThis work showed that specific plasmatic miRNA signatures have the potential to accurately discriminate patients with angiographically documented CAD from matched controls. We failed to identify a plasmatic miRNA expression pattern capable to differentiate SA from UA patients.

Circulating microRNA Profile throughout the Menstrual Cycle

Normal physiological variables, such as age and gender, contribute to alterations in circulating microRNA (miRNA) expression levels. The changes in the female body during the menstrual cycle can also be reflected in plasma miRNA expression levels. Therefore, this study aimed to determine the plasma miRNA profile of healthy women during the menstrual cycle and to assess which circulating miRNAs are derived from blood cells. The plasma miRNA expression profiles in nine healthy women were determined by quantitative real time PCR using Exiqon Human Panel I assays from four time-points of the menstrual cycle. This platform was also used for studying miRNAs from pooled whole blood RNA samples at the same four time-points. Our results indicated that circulating miRNA expression levels in healthy women were not significantly altered by the processes occurring during the menstrual cycle. No significant differences in plasma miRNA expression levels were observed between the menstrual cycle time-points, but the number of detected miRNAs showed considerable variation among the studied individuals. miRNA analysis from whole blood samples revealed that majority of miRNAs in plasma are derived from blood cells. The most abundant miRNA in plasma and blood was hsa-miR-451a, but a number of miRNAs were only detected in one or the other sample type. In conclusion, our data suggest that the changes in the female body during the menstrual cycle do not affect the expression of circulating miRNAs at measurable levels.

GW24-e2497 Circulating MicroRNAs as Potential Biomarkers of Coagulation Dysfunction in Patients with Vulnerable Coronary Artery Disease

ObjectivesThe activation of coagulation and fibrinolysis plays a critical role in the incidence of coronary events. MicroRNAs (miRNAs) are small non-coding ribonucleic acids (RNAs) that negatively regulate gene expression on...

Plasma MicroRNAs in Ovarian Cancer—Response

We are pleased that our research (1) has generated interest in the scientific community, and we are happy to provide our reply to the comments submitted by Dr. Haifeng Qiu.The major concern addressed by Dr. Qiu is that the upregulation of plasma miR-21 level in endometriosis-associated ovarian cancer (EAOC) cases is due to loss of estrogen possibly present in this group of patients. Dr. Qiu's argument is based on two lines of reasoning: (i) miR-21 expression is repressed by estrogen, and (ii) our patients with EAOC are significantly older than the healthy controls and those in the endometriosis group, and therefore they are likely postmenopausal women with low estrogen levels. In addition to miR-21, Dr. Qiu also has concerns about the increased expression of miR-16 and miR-195, which have also been previously reported as estrogen targets.Although the estrogen/progesterone imbalance is a well-documented pathogenic factor in ovarian cancer and endometriosis, the role of estrogen in regulating miR-16, 21, and 195 expressions remains inconclusive. Studies have shown miR-21 being either repressed (2, 3) or induced by estrogen (4). Furthermore, evidence from several studies on the effect of estrogen on miRNA expression (5) suggests that miR-21 was not a consistent estrogen-repressed miRNA, and miR-16 and 195 only were shown to be repressed by estrogen in one study (6) among 19 studies reviewed (5).Second, in our study the averaged fold change of miR-21 expression between EAOC and healthy controls is dramatic (fold change = 147.4) compared with the reported repressive effect of estrogen to miR-21 expression (ranging from 50%–70% repression), making it difficult to assume that estrogen has a big effect on our experimental results.Third, the average age of our healthy control group and endometriosis group is comparable (38.75 and 36.24 years, respectively). Although estrogen levels were not measured in our study, the patients with endometriosis are likely to have higher estrogen levels that were also suggested by Dr. Qiu. Thus, if miR-21 expression is repressed by estrogen, miR-21 levels in patients with endometriosis should be lower than that in the healthy controls. However, this was not what we found. Instead, patients with endometriosis displayed a much higher plasma miR-21 expression compared with healthy controls (fold change = 10.65).Fourth, differences in the above miRNAs were also present in comparisons with age-matched serous versus EAOC cases (all menopausal) and with age-matched female mice (tumor bearing vs. controls), further suggesting that factors other than estrogen may have impacted these changes.Finally, as we have shown in our study, plasma and tissue samples have distinct miRNA expression profiles. Currently, the effect of estrogen levels on plasma miRNA expression profile remains unclear.In summary, based on our study results and the above arguments, we do not believe that estrogen plays a significant role in regulating the plasma miRNA signature we have identified, which distinguishes EAOC patients with endometriosis from healthy controls.See the original Letter to the Editor, p. 3325No potential conflicts of interest were disclosed.

P-064 Analysis and validation of the miRNA expression profile in plasma from patients diagnosed with myelodysplastic syndrome

P-064 Analysis and validation of the miRNA expression profile in plasma from patients diagnosed with myelodysplastic syndrome

Clinical relevance of plasma miR-106b levels in patients with chronic obstructive pulmonary disease

Chronic obstructive pulmonary disease (COPD) is characterized by both chronic inflammation in the airway and systemic inflammation; however, the molecular mechanism of COPD has not been fully elucidated. By measuring microRNA (miRNA) expression in the plasma of COPD subjects, we aimed to identify the clinical relevance of plasma miRNA levels in these patients. Blood samples were obtained from COPD patients and age-matched normal controls. We initially produced plasma miRNA expression profiles using TaqMan low-density array screening. For further validation, individual qRT-PCRs were performed in 40COPD patients and 20 healthy subjects. TaqMan low-density array screening showed that 9miRNAs (miR-29b, miR-483-5p, miR-152, miR-629, miR-26b, miR-101, miR-106b, miR-532-5p and miR-133b) were significantly downregulated in the plasma from COPD patients when compared with normal smokers. Among these miRNAs, we focused on miR-106b. A reduction in the plasma miR-106b levels was evident in COPD ex-smokers and COPD current smokers compared with levels in smokers. There was a negative correlation between the plasma miR-106b level and the duration of disease since diagnosis in COPD ex-smokers and the duration of smoking in COPD current smokers. These findings support the concept that progressive reduction in the plasma miR-106b level may reflect persistent and systemic changes even after the discontinuation of smoking in COPD patients. miR-106b may therefore play an important role in the pathogenesis of COPD.

Two Panels of Plasma MicroRNAs as Non-Invasive Biomarkers for Prediction of Recurrence in Resectable NSCLC

The diagnosis of non-small cell lung carcinoma (NSCLC) at an early stage, as well as better prediction of outcome remains clinically challenging due to the lack of specific and robust non-invasive markers. The discovery of microRNAs (miRNAs), particularly those found in the bloodstream, has opened up new perspectives for tumor diagnosis and prognosis. The aim of our study was to determine whether expression profiles of specific miRNAs in plasma could accurately discriminate between NSCLC patients and controls, and whether they are able to predict the prognosis of resectable NSCLC patients. We therefore evaluated a series of seventeen NSCLC-related miRNAs by quantitative real-time (qRT)-PCR in plasma from 52 patients with I-IIIA stages NSCLC, 10 patients with chronic obstructive pulmonary disease (COPD) and 20-age, sex and smoking status-matched healthy individuals. We identified an eleven-plasma miRNA panel that could distinguish NSCLC patients from healthy subjects (AUC = 0.879). A six-plasma miRNA panel was able to discriminate between NSCLC patients and COPD patients (AUC = 0.944). Furthermore, we identified a three-miRNA plasma signature (high miR-155-5p, high miR-223-3p, and low miR-126-3p) that significantly associated with a higher risk for progression in adenocarcinoma patients. In addition, a three-miRNA plasma panel (high miR-20a-5p, low miR-152-3p, and low miR-199a-5p) significantly predicted survival of squamous cell carcinoma patients. In conclusion, we identified two plasma miRNA expression profiles that may be useful for predicting the outcome of patients with resectable NSCLC.

Maternal plasma miRNAs expression in preeclamptic pregnancies.

Objective. Preeclampsia (PE) is a pregnancy-specific syndrome and one of the leading causes of maternal and fetal morbidity and mortality. The pathophysiological mechanisms of PE remain poorly known. Recently, circulating miRNAs are considered as potential useful noninvasive biomarkers. The aim of this study was to identify differentially expressed plasma miRNAs in preeclamptic pregnancies compared with normal pregnancies. Methods. Maternal plasma miRNA expression profiles were detected by SOLiD sequencing. Differential expressions between mPE/sPE and control group were found. Next, four differentially expressed plasma miRNAs were chosen to validate their expression in other large scale samples by real-time PCR. Results. In terms of sequencing results, we identified that 51 miRNAs were differentially expressed. Four differentially expressed plasma miRNAs (miR-141, miR-144, miR-221, and miR-29a) were selected to validate the sequencing results. RT-PCR data confirmed the reliability of sequencing results. The further statistical analysis showed that maternal plasma miR-141 and miR-29a are significantly overexpressed in mPE (P < 0.05). Maternal plasma miR-144 is significantly underexpressed in mPE and sPE (P < 0.05). Conclusions. Results showed that there were differentially expressed maternal plasma miRNAs in patients with preeclampsia. These plasma miRNAs might be used as notable biomarkers for diagnosis of preeclampsia.

Circulating MicroRNAs as Biomarkers of Colorectal Cancer: Results From a Genome-Wide Profiling and Validation Study

Circulating microRNAs (miRNAs/miRs) might be used as biomarkers for the diagnosis of cancer and other diseases. Noninvasive approaches are needed to complement and improve upon current strategies for colorectal cancer (CRC) screening. We investigated whether plasma levels of miRNA can differentiate patients with CRC from healthy individuals. We also investigated whether plasma samples from patients with premalignant neoplastic lesions, such as advanced adenomas (AAs), also had a different expression pattern of miRNAs. We analyzed 196 plasma samples from 123 patients newly diagnosed with sporadic colorectal neoplasia (63 with CRC and 60 with AAs) and 73 healthy individuals (controls) seen at 2 tertiary medical centers in Spain. An initial set of samples was analyzed using a genome-wide miRNA expression profiling assay (n = 61). Quantitative reverse-transcription PCR was used to validate the expression of selected miRNAs in an independent cohort (n = 135). Patients with CRC or AAs had plasma miRNA expression profiles that differed significantly from those of controls. We selected a group of 13 miRNAs for validation in an independent cohort of patients; 6 (miR18a, miR19a, miR19b, miR15b, miR29a, and miR335) were confirmed to be significantly up-regulated in patients with CRC, differentiating patients with CRC from controls with area under the receiver operating characteristic curve values ranging from 0.80 (95% confidence interval [CI], 0.71-0.89) to 0.70 (95% CI, 0.59-0.80). Only miR18a was confirmed to be significantly up-regulated in patients with AAs, compared with controls; the area under the receiver operating characteristic curve value was 0.64 (95% CI, 0.52-0.75). Patients with CRC have significantly different patterns of miRNA expression than healthy individuals. These patterns might be developed as biomarkers for CRC, although they have limited value in identifying patients with premalignant neoplastic lesions.

206 EFFECT OF OVARIAN HYPERSTIMULATION ON CIRCULATORY microRNA PROFILE IN BOVINE FOLLICULAR FLUID AND BLOOD PLASMA

MicroRNAs (miRNA) are noncoding small RNA that are known to play a role in posttranscriptional regulation of genes involved in various physiological processes including disease and reproduction. The circulatory forms of miRNA, which are present in body fluids, are reported to be used as biomarkers for disease and pregnancy. This study was conducted to investigate the effect of ovarian hyperstimulation on the expression pattern of circulatory miRNA in blood plasma and follicular fluid. For this, one group of Simmental heifers (n = 6) was hyper-stimulated using a superovulation (SO) protocol of 8 consecutive FSH injections over 4 days in decreasing doses, and others (n = 6) were synchronized (SY) using a standard synchronization protocol. Following this, the blood samples were collected at Day 0 (onset of oestrus) and Day 7, and follicular fluid was collected from both groups at Day 0 of the oestrous cycle. Total RNA, including small RNA, was then isolated from follicular fluid and blood plasma using a miRNeasy mini kit (Qiagen, Valencia, CA, USA) and subsequently used for circulatory miRNA expression studies using the human miRCURY LNA™ Universal RT miRNA PCR array system (Exiqon, Woburn, MA, USA). Following the miRNA PCR array run, data analysis was performed using a delta threshold cycle (ΔCT) method. Results showed that 23 miRNAs were found to be differentially expressed in blood plasma (fold change ≥2 and P ≤ 0.05) between SO and SY groups. Among these, 8 miRNAs including miR-127-3p, miR-494, miR-147, miR-134, and miR-153 were downregulated and 15 miRNAs including miR-34a, miR-103, miR-181c, miR-24-2-3p, miR-18a-3p, miR-20b-3p, and miR-708-3p were found to be upregulated in SO groups. Similarly, in follicular fluid, 71 miRNAs were found to be differentially expressed between the SO and SY groups. Of these, 33 miRNA including miR-100, miR-877, miR-659, miR-200c, miR-29b-2-3p, miR-361-5p, and miR-145 were downregulated, whereas 38 miRNAs including miR-374a, miR-720, miR-155-3p, miR-202-3p, miR-33b, miR-12-3p, and miR-224-3p were upregulated in follicular fluid aspirated from SO cows compared with the SY animals. Ingenuity pathway analysis of predicted target genes of miRNA, which are dysregulated due to ovarian hyperstimulation, showed the dominance of pathways related to neuregulin signalling, axonal guidance signalling, GNRH signalling, and Wnt β-catenin signalling pathways. In conclusion, this study revealed alternation in circulatory miRNA expression profile both in blood plasma and follicular fluid as a result of ovarian hyperstimulation.

Reply to G. Cai et al

We would like to thank Cai et al for their comments on our article. In the study by Gao et al, microRNA (miRNA) was extracted from formalin-fixed, paraffin-embedded dysplastic nodules, hepatocellular carcinomas (HCCs), and the adjacent nontumorous liver tissues. This study showed that miRNA deregulation at the tissue level was an early event and accumulated throughout the various steps of hepatitis B virus–associated hepatocarcinogenesis. In our study, the panel with seven miRNAs (miR-122, miR-192, miR-21, miR-223, miR-26a, miR-27a, and miR-801) was identified in the plasma of patients with HCC. The discrepancy between the two studies would be primarily a result of the different categories of the study specimens (tissue v plasma). To our knowledge, the expression profiles of plasma miRNA are different from that of tissue miRNA. Plasma circulating miRNA was reported to have diverse origins. It is well known that most hepatitis B virus–related HCCs involve two disease processes: the HCC as well as liver damage as a result of either hepatitis or cirrhosis. Thus, liver damage, tumor immune response, and/or hepatitis may also influence the miRNA profiles in the plasma of patients with HCC. Therefore, the expression profiles of plasma miRNAs in patients with HCC may not trend according to the expression profiles in HCC tissue. For example, studies by Wang et al and Starkey Lewis et al found miR-122 and miR-192 to be associated with liver damage, and the two miRNAs expressed opposite regulations in the tissue and plasma specimens. This indicates that the miRNA profiles in the plasma specimens are more complicated compared with those in pure tumor tissue in patients with HCC. At the tissue level, two studies revealed the differences in miRNA dysregulation in early versus advanced HCCs. Such differences have not been reported in the current plasma studies of miRNA profiles. It would be interesting to further investigate the differences in miRNA spectrum and expression level between early and advanced HCCs in the plasma of patients with HCC.

Abstract LB-476: A universal method for elimination of haemolyzed plasma samples that improves miRNA signature performance for early detection of colorectal cancer

Abstract microRNAs (miRNAs) are short (∼21 bp) non-coding RNAs that regulate gene expression through post-transcriptional interactions with target messenger RNAs. miRNAs are exported to extracellular fluids by both malignant cells and cells of the immune system. Blood plasma and serum miRNAs are stable under standard clinical sampling protocols and have therefore attracted considerable interest for their potential as minimally invasive biomarkers for diverse pathological conditions, including various cancers. Although blood plasma contains as little as 1-20 ng of RNA per mL, plasma miRNAs are easily and reliably quantified using the highly sensitive and specific miRCURY LNA™ Universal RT microRNA PCR platform. However, the relative scarcity of miRNA in plasma creates a potential for contamination of the plasma miRNA expression profile by miRNAs from the cellular constituents in blood. One common cause of plasma sample rejection in clinical chemistry is haemolysis. If not identified, haemolysis can lead to erroneous results in a number of standard clinical laboratory tests, including blood potassium and lactate dehydrogenase (LDH) levels. Here we investigate the effect of sample haemolysis on plasma miRNA profiles. Using isolated red blood cells (RBCs) and genome-wide miRNA PCR profiling, we define the miRNome of RBCs. By spiking RBC extract into a non-haemolyzed plasma sample, we demonstrate that as little as 0.05% contamination of plasma can be detected as an increased expression level of a subset of miRNAs. We also define a set of plasma miRNAs that are not affected by haemolysis. Finally, we discover a miRNA qPCR QC signature that can be used to eliminate haemolyzed plasma samples. In a project aimed at defining a signature for the early detection of colorectal cancer (CRC) from a plasma sample, we screened 325 plasma samples from CRC patients and colonoscopy-verified CRC-negative controls. The samples were part of a clinical trial conducted in 7 different Danish hospitals, and were examined for the expression of 378 miRNAs previously detected in plasma. We show that haemolysis in this sample set correlates with hospital ID, and with the utilization of specific blood sample collection vials. Using our haemolysis signature, we eliminated haemolyzed samples and demonstrated that this step leads to a major improvement of CRC detection (ROC AUC increase from 0.67 to &amp;gt;0.80). We conclude that haemolysis can be a cause of plasma miRNA profile contamination, and that elimination of haemolyzed samples using our miRNA haemolysis QC signature overcomes this clinical problem and leads to an increase in plasma miRNA biomarker performance. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr LB-476. doi:1538-7445.AM2012-LB-476

A comprehensive survey of maternal plasma miRNAs expression profiles using high-throughput sequencing

ObjectiveRecently, microRNAs (miRNAs) had been shown as potential important regulators in pregnancy. Circulating miRNAs are considered as potentially useful non-invasive biomarkers for the diagnosis of pregnancy-related disease and congenital disorders, but maternal peripheral blood miRNAs expression profile in pregnancy remains less investigated. We thus set out to investigate maternal plasma miRNAs expression profile using genome-wide sequencing. MethodsMaternal plasma miRNA expression profiles of different pregnancy stages were detected by SOLiD sequencing. We observed the expression level of the most abundant miRNAs in maternal plasma during pregnancy process. We examined functional relationships of targets of pregnancy-relative miRNAs by enrichment analyzing of signaling pathways. Results147 miRNAs were sequenced from maternal plasma in this study, among them, 90 types of miRNAs were found in all of the samples, while 136 miRNAs in the first trimester gestation, 108 in second trimester gestation, and 99 miRNAs in the third trimester gestation, respectively. The varieties and the expression level of maternal plasma miRNAs were changing during pregnancy. The expression level of miRNA cluster members was changing with the same trend during pregnancy. The function and functional relationship analysis of target genes of pregnancy-relative miRNAs showed that genetic disorder, immunological disease, cell signaling, cancer, and cell cycle were the enriched pathways. ConclusionsMaternal plasma miRNA expression profiles are dynamically changing during pregnancy. The results of function analysis suggested that miRNAs may play an important role in regulating pregnancy process, which can help us understand the refine regulation mechanism in pregnancy. Moreover, the results of this present study may be the basis for a further study to find useful prenatal diagnosis biomarkers.

Plasma miRNA as Biomarkers for Assessment of Total-Body Radiation Exposure Dosimetry

The risk of radiation exposure, due to accidental or malicious release of ionizing radiation, is a major public health concern. Biomarkers that can rapidly identify severely-irradiated individuals requiring prompt medical treatment in mass-casualty incidents are urgently needed. Stable blood or plasma-based biomarkers are attractive because of the ease for sample collection. We tested the hypothesis that plasma miRNA expression profiles can accurately reflect prior radiation exposure. We demonstrated using a murine model that plasma miRNA expression signatures could distinguish mice that received total body irradiation doses of 0.5 Gy, 2 Gy, and 10 Gy (at 6 h or 24 h post radiation) with accuracy, sensitivity, and specificity of above 90%. Taken together, these data demonstrate that plasma miRNA profiles can be highly predictive of different levels of radiation exposure. Thus, plasma-based biomarkers can be used to assess radiation exposure after mass-casualty incidents, and it may provide a valuable tool in developing and implementing effective countermeasures.

506. MENSTRUAL CYCLE VARIATIONS IN PLASMA microRNA EXPRESSION PROFILES

microRNAs are short, single-stranded RNAs that regulate gene expression at the post-transcriptional level. Plasma and serum microRNAs correlate closely with microRNA profiles of diseased tissue and have been explored as blood-based biomarkers for human diseases, including steroid-driven malignancies. However, reproductive steroid signalling regulates the expression of specific microRNAs and this could impact the utility of microRNA biomarkers in reproductive aged women. We hypothesised that microRNA expression profiles are altered by steroid hormone fluctuations associated with the menstrual cycle. To test this hypothesis, plasma microRNA expression was measured in healthy women at 3 stages of a 28 day menstrual cycle; ie menstrual (day 3-5), follicular (day 9–13) and implantation window/secretory phase (day 18–22). Total RNA was extracted from plasma, multiplex reverse transcription was performed, and the cDNA pre-amplified prior to expression analysis of 667 microRNAs on Taqman low density PCR arrays (n=6 women). Preliminary data shows that up to 200 microRNAs may be detected with this methodology, and that at least 14 of these are differentially expressed (fold change ≥±1.5) at follicular and secretory phase, as compared to menstrual phase. We plan to confirm these findings with standard Taqman microRNA assays (n=10 women). Our findings suggest that plasma miRNA expression profiles change over the menstrual cycle, and that this could confound microRNA-based diagnostic tests. We hope to demonstrate the most appropriate cycle phase for blood-based miRNA profiling, facilitating the development of plasma microRNA-based diagnostic tests and providing valuable information to researchers studying circulating microRNA profiles in reproductive aged women.