High-Density Lipoprotein-Associated miR-223 Is Altered after Diet-Induced Weight Loss in Overweight and Obese Males

Background and Aims

microRNAs (miRNAs) are small, endogenous non-coding RNAs that regulate metabolic processes, including obesity. The levels of circulating miRNAs are affected by metabolic changes in obesity, as well as in diet-induced weight loss. Circulating miRNAs are transported by high-density lipoproteins (HDL) but the regulation of HDL-associated miRNAs after diet-induced weight loss has not been studied. We aim to determine if HDL-associated miR-16, miR-17, miR-126, miR-222 and miR-223 levels are altered by diet-induced weight loss in overweight and obese males.

Methods

HDL were isolated from 47 subjects following 12 weeks weight loss comparing a high protein diet (HP, 30% of energy) with a normal protein diet (NP, 20% of energy). HDL-associated miRNAs (miR-16, miR-17, miR-126, miR-222 and miR-223) at baseline and after 12 weeks of weight loss were quantified by TaqMan miRNA assays. HDL particle sizes were determined by non-denaturing polyacrylamide gradient gel electrophoresis. Serum concentrations of human HDL constituents were measured immunoturbidometrically or enzymatically.

Results

miR-16, miR-17, miR-126, miR-222 and miR-223 were present on HDL from overweight and obese subjects at baseline and after 12 weeks of the HP and NP weight loss diets. The HP diet induced a significant decrease in HDL-associated miR-223 levels (p = 0.015), which positively correlated with changes in body weight (r = 0.488, p = 0.032). Changes in miR-223 levels were not associated to changes in HDL composition or size.

Conclusion

HDL-associated miR-223 levels are significantly decreased after HP diet-induced weight loss in overweight and obese males. This is the first study reporting changes in HDL-associated miRNA levels with diet-induced weight loss.     

Regulation of microRNAs miR-30a and miR-143 in cerebral vasculature after experimental subarachnoid hemorrhage in rats

Background

microRNAs (miRNAs) are important regulators of translation and have been implicated in the pathogenesis of a number of cardiovascular diseases, including stroke, and suggested as possible prognostic biomarkers. Our aim was to identify miRNAs that are differentially regulated in cerebral arteries after subarachnoid hemorrhage (SAH), using a rat injection model of SAH and a qPCR-based screen of 728 rat miRNAs. Additionally, serum was analyzed for a possible spill-over to the circulation of regulated miRNAs from the vessel walls.

Results

We identified 482 different miRNAs expressed in cerebral arteries post-SAH. Two miRNAs, miR-30a and miR-143, were significantly upregulated in cerebral arteries after SAH when compared to sham-operated animals. However, none of these exhibited significantly altered serum levels after SAH versus post-sham surgery. The most robust upregulation was seen for miR-143, which has several predicted targets and is a strong regulator of vascular morphology. We hypothesize that miR-30a and miR-143 may play a role in the vascular wall changes seen after SAH.

Conclusions

We report that miR-30a and miR-143 in the cerebral arteries show significant changes over time after SAH, but do not differ from sham-operated rats at 24 h post-SAH. Although this finding suggests interesting novel possible mechanisms involved in post-SAH cerebrovascular changes, the lack of regulation of these miRNAs in serum excludes their use as blood-borne biomarkers for cerebrovascular changes following SAH.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1341-7) contains supplementary material, which is available to authorized users.     

Circulating MicroRNA-150 Serum Levels Predict Survival in Patients with Critical Illness and Sepsis

Background and Aims

Down-regulation of miR-150 was recently linked to inflammation and bacterial infection. Furthermore, reduced serum levels of miR-150 were reported from a small cohort of patients with sepsis. We thus aimed at evaluating the diagnostic and prognostic value of miR-150 serum levels in patients with critically illness and sepsis.

Methods

miR-150 serum levels were analyzed in a cohort of 223 critically ill patients of which 138 fulfilled sepsis criteria and compared to 76 healthy controls. Results were correlated with clinical data and extensive sets of routine and experimental biomarkers.

Results

Measurements of miR-150 serum concentrations revealed only slightly reduced miR-150 serum levels in critically ill patients compared to healthy controls. Furthermore miR-150 levels did not significantly differ in critically ill patients with our without sepsis, indicating that miR-150 serum levels are not suitable for diagnostic establishment of sepsis. However, serum levels of miR-150 correlated with hepatic or renal dysfunction. Low miR-150 serum levels were associated with an unfavorable prognosis of patients, since low miR-150 serum levels predicted mortality with high diagnostic accuracy compared with established clinical scores and biomarkers.

Conclusion

Reduced miR-150 serum concentrations are associated with an unfavorable outcome in patients with critical illness, independent of the presence of sepsis. Besides a possible pathogenic role of miR-150 in critical illness, our study indicates a potential use of circulating miRNAs as a prognostic rather than diagnostic marker in critically ill patients.     

Five microRNAs in plasma as novel biomarkers for screening of early-stage non-small cell lung cancer

Background

In order to find novel noninvasive biomarkers with high accuracy for the screening of early-stage non-small cell lung cancer (NSCLC), we investigate the predictive power of 5 microRNAs (miR-20a, miR-145, miR-21, miR223 and miR-221) as potential biomarkers in early-stage NSCLC.

Methods

In training set, 25 early-stage NSCLC patients and 25 matched healthy controls are included to assess the miRNA expression profile between early-stage NSCLC patients and healthy controls by real-time RT-PCR. We found that five of these miRNAs (miR-20a, miR-223, miR-21, miR-221 and miR-145) levels in NSCLC patients were significantly dysregulated compared with the healthy groups and thus were selected to validation set. Therefore, a validation experiment was further performed to investigate the potential predictive power of these five miRNAs based on 126 early-stage NSCLC patients, 42 NCPD patients and 60 healthy controls. The receiver operating characteristic (ROC) curves were generated for the five miRNAs.

Results

ROC curve analyses suggested that these five plasma miRNAs could be promising biomarkers for NSCLC, with relatively high AUC values as follows: miR-20a, 0.89 with 95% CI of [0.85-0.93]; miR-223, 0.94 with 95% CI of [0.91-0.96]; miR-21, 0.77 with 95% CI of [0.71-0.83]; miR-155, 0.92 with 95% CI of [0.89-0.96]; miR-145, 0.77 with 95% CI of [0.71-0.83]. Stratified analyses indicated that plasma miR-20a, miR-223, miR-21 and miR-145 showed better predictive value in smokers than in non-smokers, while miR-155 might be more suitable for non-smokers. In addition, all of these five miRNAs could differentiate NSCLC from controls with a higher accuracy in advanced stage and squamous carcinoma subgroups.

Conclusions

In conclusion, our study suggested that five plasma miRNAs (miR-20a, miR-145, miR-21, miR-223 and miR-221) can be used as promising biomarkers in early screening of NSCLC. Nevertheless, further validation and optimizing improvement should be performed on larger sample to confirm our results.     

Localization of the Sodium-Taurocholate cotransporting polypeptide in membrane rafts and modulation of its activity by cholesterol in vitro

Background

The relevance of discrete localization of hepatobiliary transporters in specific membrane microdomains is not well known.

Aim

To determine whether the Na⁺/taurocholate cotransporting polypeptide (Ntcp), the main hepatic sinusoidal bile salt transporter, is localized in specific membrane microdomains.

Methods

Presence of Ntcp in membrane rafts obtained from mouse liver was studied by immunoblotting and immunofluorescence. HEK-293 cells stably transfected with rat Ntcp were used for in vitro studies. Expression, localization and function of Ntcp in these cells were assessed by immunoblotting, immunofluorescence and biotinylation studies and Na⁺-dependent taurocholate uptake assays, respectively. The effect of cholesterol depletion/repletion assays on Ntcp function was also investigated.

Results

Ntcp localized primarily to membrane rafts in in vivo studies and localized partially in membrane rafts in transfected HEK-293 cells. In these cells, membrane cholesterol depletion resulted in a shift of Ntcp localization into non-membrane rafts, which correlated with a 2.5-fold increase in taurocholate transport. Cholesterol repletion shifted back part of Ntcp into membrane rafts, and normalized taurocholate transport to values similar to control cells.

Conclusion

Ntcp localizes in membrane rafts and its localization and function are regulated by membrane cholesterol content. This may serve as a novel regulatory mechanism of bile salt transport in liver.     

Circulating miR-30a,miR-195 and let-7b Associated with Acute Myocardial Infarction

Background

MicroRNAs (miRNAs) play key roles in diverse biological and pathological processes, including the regulation of proliferation, apoptosis, angiogenesis and cellular differentiation. Recently, circulating miRNAs have been reported as potential biomarkers for various pathologic conditions. This study investigated miR-30a, miR-195 and let-7b as potential of biomarker for acute myocardial infarction (AMI).

Methods and Results

Plasma samples from 18 patients with AMI and 30 healthy adults were collected. Total RNA was extracted from plasma with TRIzol LS Reagent. MiRNA levels and plasma cardiac troponin I (cTnI) concentrations were measured by quantitative real-time PCR and ELISA assay, respectively. Results showed that circulating miR-30a in AMI patients was highly expressed at 4 h, 8 h and 12 h after onset of AMI, and miR-195 was highly expressed at 8 h and 12 h. However, let-7b was lower in AMI patients than in controls throughout the whole time points. Interestingly, in these patients, circulating miR-30a, miR-195 and let-7b all reached their expression peak at 8 h. By the receiver operating characteristic (ROC) curve analyses, these plasma miRNAs were of significant diagnostic value for AMI. The combined ROC analysis revealed the an AUC value of 0.93 with 94% sensitivity and 90% specificity at 8 h after onset, and an AUC value of 0.92 with 90% sensitivity and 90% specificity at 12 h after onset, in discriminating the AMI patients from healthy controls.

Conclusions

Our results imply that the plasma concentration of miR-30a, miR-195 and let-7b can be potential indicators for AMI.     

Consumption of green tea or green tea products: Is there an evidence for antioxidant effects from controlled interventional studies?

Purpose

Epidemiological data suggest that green tea (GT) consumption may protect against cardiovascular diseases (CVDs) and different types of cancer. This effect is attributed primarily to the antioxidant properties of flavanols from GT. This review provides an overview of controlled intervention studies investigating the effect of GT consumption on antioxidant effects ex vivo and in vivo.

Methods

The Medline and Cochrane databases were searched independently by two investigators for controlled intervention studies (English) on GT consumption and antioxidant effects published up to June 2010. Thirty-one studies investigating antioxidant effects ex vivo [plasma antioxidant capacity (AC), DNA's resistance against oxidative induced damage) or in vivo (lipid and protein oxidation, DNA damage] met the criteria. Results were compared by considering the participants, the dose of GT, the amount of ingested flavanols, the duration of supplementation and the investigated biomarkers.

Results

The comparison between the studies was difficult as relevant data, e.g., on flavanol concentration in plasma (10 of 31 studies) or on major antioxidants contributing to AC, were often missing. Lipid peroxidation and DNA damage were commonly investigated. Data on protein oxidation are scarce. An antioxidant effect of at least one parameter (increase in AC or reduction of oxidative stress marker) was observed in 15 out of 22 studies by daily consumption of GT, primarily in participants exposed to oxidative stress (smokers or mixed collectives of smokers and non-smokers and physical activity) and in 6 out of 9 studies investigating the bolus consumption of GT.

Conclusion

There is limited evidence that regular consumption of GT in amounts of at least 0.6-1.5 l/day may increase AC and reduce lipid peroxidation (especially oxidation of LDL). This may contribute to the protection against CVDs and different types of cancer. Beneficial effects seem to be more likely in participants exposed to oxidative challenge.     

Circulating miR-19a and miR-205 in Serum May Predict the Sensitivity of Luminal A Subtype of Breast Cancer Patients to Neoadjuvant Chemotherapy with Epirubicin Plus Paclitaxel

Background

The luminal A subtype of breast cancer has a good prognosis and is sensitive to endocrine therapy but is less sensitive to chemotherapy. It is necessary to identify biomarkers to predict chemosensitivity and avoid over-treatment. We hypothesized that miRNAs in the serum might be associated with chemosensitivity.

Methods

Sixty-eight breast cancer patients received neoadjuvant chemotherapy with epirubicin plus paclitaxel. The serum of the patients was collected before chemotherapy and stored at −80°C. The samples were classified into two groups in term of the chemosensitivity. We identified the differential expression patterns of miRNAs between the chemotherapy sensitive and resistant groups using microRNA profiling. Four miRNAs that were differentially expressed between the two groups were further validated in another 56 samples. We created a model fitting formula and a receiver operating characteristics (ROC) curve using logistic regression analysis to evaluate the prediction potency.

Results

We identified 8 miRNAs differentially expressed between the two groups: 6 miRNAs were up-regulated, and 2 miRNAs were down-regulated in the resistant group compared with the sensitive group. The expression of miR-19a and miR-205 were determined to have significant differences between the two groups (P<0.05). A predictive model of these two miRNAs was created by the logistic regression analysis. The probability of this model was 89.71%. Based on the ROC curve, the specificity was 75.00%, and the sensitivity was 81.25%.

Conclusions

The combination of miR-19a and miR-205 in the serum may predict the chemosensitivity of luminal A subtype of breast cancer to epirubicin plus paclitaxel neoadjuvant chemotherapy.     

Association study of microRNA polymorphisms with risk of idiopathic recurrent spontaneous abortion in Korean women

Aim

The aim of this study was to investigate the association of microRNA polymorphisms (miR-146aC>G, miR-149T>C, miR-196a2T>C, and miR-499A>G) in Korean patients with recurrent spontaneous abortion (RSA).

Methods

We conducted a case-control study of 564 Korean women: 330 patients with at least two unexplained consecutive pregnancy losses and 234 healthy controls with at least one live birth and no history of pregnancy loss.

Results

RSA patients exhibited significantly different frequencies of the miR-196a2CC (TT+TC vs. CC; adjusted odds ratio [AOR], 1.587; 95% confidence interval [CI], 1.042-2.417) and miR-499AG+GG genotypes (AOR, 1.671; 95% CI, 1.054-2.651) compared with the control group. The combination of miR-196a2CC and miR-499AG+GG showed synergistic effects (AOR, 3.541; 95% CI, 1.645-7.624).

Conclusion

miR-196a2CC, miR-499AG+GG, and the miR-196a2CC/miR-499AG+GG combination are significantly associated with idiopathic RSA in Korean women.     

The peripheral blood mononuclear cell microRNA signature of coronary artery disease

Background

MicroRNAs are being used in the oncology field to characterize tumors and predict the survival of cancer patients. Here, we explored the potential of microRNAs as biomarkers for coronary artery disease (CAD) and acute coronary syndromes.

Methods and results

Using real-time PCR-based profiling, we determined the microRNA signature of peripheral blood mononuclear cells (PBMCs) from stable and unstable CAD patients and unaffected controls. 129 of 157 microRNAs measured were expressed by PBMCs and low variability between separate PBMC pools was observed. The presence of CAD in general coincided with a marked 5-fold increase (P < 0.001) in the relative expression level of miR-135a, while the expression of miR-147 was 4-fold decreased (P < 0.05) in PBMCs from CAD patients as compared to controls, resulting in a 19-fold higher miR-135a/miR-147 ratio (P < 0.001) in CAD. MicroRNA/target gene/biological function linkage analysis suggested that the change in PBMC microRNA signature in CAD patients is probably associated with a change in intracellular cadherin/Wnt signaling. Interestingly, unstable angina pectoris patients could be discriminated from stable patients based upon their relatively high expression level of a cluster of three microRNAs including miR-134, miR-198, and miR-370, suggesting that the microRNA signatures can be used to identify patients at risk for acute coronary syndromes.

Conclusions

The present study is the first to show that microRNA signatures can possibly be utilized to identify patients exhibiting atherosclerotic CAD in general and those at risk for acute coronary syndromes. Our findings highlight the importance of microRNAs signatures as novel tool to predict clinical disease outcomes.     

High expression of miR-210 predicts poor survival in patients with breast cancer: A meta-analysis

Background

Many microRNAs (miRNAs) exhibit altered expression levels in cancers, and they may be considered as valuable prognostic biomarkers for cancers. Here we aimed to summarize the recent advances in miR-210 involvement in human breast cancer and analyze the predicting role of miR-210 for survival.

Methods

A meta-analysis was performed by searching PubMed, Cochrane Library, and Science Direct databases. Data were extracted from studies comparing survival in patients with breast cancer having higher expression of miR-210 with those having lower expression. Pooled hazard ratios (HRs) and 95% confidence intervals (CI) were calculated.

Results

A total of 511 cases of breast cancer were involved for this global meta-analysis. For post-operational survival, the HR of higher miR-210 expression in breast cancer tissue was 3.39 (95% CI: 2.04–5.63, P < 0.05), which could significantly predict poorer survival.

Conclusions

High expression of miR-210 might predict poor survival in patients with breast cancer.     

Circulating miR-17, miR-20a,miR-29c,and miR-223 Combined as Non-Invasive Biomarkers in Nasopharyngeal Carcinoma

Background

MicroRNAs have been considered as a kind of potential novel biomarker for cancer detection due to their remarkable stability in the blood and the characteristics of their expression profile in many diseases.

Methods

We performed microarray-based serum miRNA profiling on the serum of twenty nasopharyngeal carcinoma patients at diagnosis along with 20 non-cancerous individuals as controls. This was followed by a real-time quantitative Polymerase Chain Reaction (RT-qPCR) in a separate cohort of thirty patients with nasopharyngeal carcinoma and thirty age- matched non-cancerous volunteers. A model for diagnosis was established by a conversion of mathematical calculation formula which has been validated by analyzing 74 cases of patients with nasopharyngeal carcinoma and 57 cases of non-cancerous volunteers.

Results

The profiles showed that 39 and 17 miRNAs are exclusively expressed in the serum of non-cancerous volunteers and of patients with nasopharyngeal carcinoma respectively. 4 miRNAs including miR-17, miR-20a, miR-29c, and miR-223 were found to be expressed differentially in the serum of NPC compared with that of non-cancerous control. Based on this, a diagnosis equation with Ct difference method has been established to distinguish NPC cases and non-cancerous controls and validated with high sensitivity and specificity.

Conclusions

We demonstrate that the serum miRNA-based biomarker model become a novel tool for NPC detection. The circulating 4-miRNA-based method may provide a novel strategy for NPC diagnosis.     

Age and sex differences in kidney microRNA expression during the life span of F344 rats

Background

Growing evidence suggests that epigenetic mechanisms of gene regulation may play a role in susceptibilities to specific toxicities and adverse drug reactions. MiRNAs in particular have been shown to be important regulators in cancer and other diseases and show promise as predictive biomarkers for diagnosis and prognosis. In this study, we characterized the global kidney miRNA expression profile in untreated male and female F344 rats throughout the life span. These findings were correlated with sex-specific susceptibilities to adverse renal events, such as male-biased renal fibrosis and inflammation in old age.

Methods

Kidney miRNA expression was examined in F344 rats at 2, 5, 6, 8, 15, 21, 78, and 104 weeks of age in both sexes using Agilent miRNA microarrays. Differential expression was determined using filtering criteria of ≥1.5 fold change and ANOVA or pairwise t-test (FDR <5%) to determine significant age and sex effects, respectively. Pathway analysis software was used to investigate the possible roles of these target genes in age- and sex-specific differences.

Results

Three hundred eleven miRNAs were found to be expressed in at least one age and sex. Filtering criteria revealed 174 differentially expressed miRNAs in the kidney; 173 and 34 miRNAs exhibiting age and sex effects, respectively. Principal component analysis revealed age effects predominated over sex effects, with 2-week miRNA expression being much different from other ages. No significant sexually dimorphic miRNA expression was observed from 5 to 8 weeks, while the most differential expression (13 miRNAs) was observed at 21 weeks. Potential target genes of these differentially expressed miRNAs were identified.

Conclusions

The expression of 56% of detected renal miRNAs was found to vary significantly with age and/or sex during the life span of F344 rats. Pathway analysis suggested that 2-week-expressed miRNAs may be related to organ and cellular development and proliferation pathways. Male-biased miRNA expression at older ages correlated with male-biased renal fibrosis and mononuclear cell infiltration. These miRNAs showed high representation in renal inflammation and nephritis pathways, and included miR-214, miR-130b, miR-150, miR-223, miR-142-5p, miR-185, and miR-296*. Analysis of kidney miRNA expression throughout the rat life span will improve the use of current and future renal biomarkers and inform our assessments of kidney injury and disease.

Electronic supplementary material

The online version of this article (doi:10.1186/s13293-014-0019-1) contains supplementary material, which is available to authorized users.     

Evaluation of miR-21 and miR-150 expression in immune thrombocytopenic purpura pathogenesis: a case-control study

Background

Immune thrombocytopenic purpura (ITP) is a common autoimmune disorder diagnosed with thrombocytopenia and bleeding symptoms due to production of autoantibodies (Abs) against platelets. Nowadays, microRNAs are known as novel biomarkers for diagnosis of diseases. The aim of this study was to investigate the expression levels of miR-21 and miR-150 in ITP patients and determine the role of these miRNAs in ITP pathogenesis.

Materials and Methods

Thirty newly diagnosed patients with acute ITP and 30 healthy subjects( age and sex matched) as controls were enrolled in this study. The expression level of miR-21 and miR-150 was investigated using Real-time-PCR. Comparison of demographic characteristics of the cases was done using independent t-test and chi-square test. Comparison of the expression level of miR-21 and miR-150 with the related parameters was done using independent t-test or Mann–Whitney and Kruskal–Wallis test. Spearman rho correlation coefficient was used to investigate the relationship between the expression of miR-21 and miR-150 with demographic characteristics.

Results

The expression of miR-21, 150 in the patients was not different compared with the control group in general. A significant relationship between the expression of miR-21 with hemoglobin, hematocrit and red blood cell hemoglobin concentration was observed.

Discussion

Expression of miR-21 and miR-150 is not associated with pathogenesis of acute ITP and can involve the synergistic role of other miRNAs. Investigation of miR-21 and miR-150 expression along with other miRNAs and cytokines can be helpful in diagnosis and pathogenesis of ITP.

     

MicroRNA 144 impairs insulin signaling by inhibiting the expression of insulin receptor substrate 1 in type 2 diabetes mellitus

Background

Dysregulation of microRNA (miRNA) expression in various tissues and body fluids has been demonstrated to be associated with several diseases, including Type 2 Diabetes mellitus (T2D). Here, we compare miRNA expression profiles in different tissues (pancreas, liver, adipose and skeletal muscle) as well as in blood samples from T2D rat model and highlight the potential of circulating miRNAs as biomarkers of T2D. In parallel, we have examined the expression profiles of miRNAs in blood samples from Impaired Fasting Glucose (IFG) and T2D male patients.

Methodology/Principal Findings

Employing miRNA microarray and stem-loop real-time RT-PCR, we identify four novel miRNAs, miR-144, miR-146a, miR-150 and miR-182 in addition to four previously reported diabetes-related miRNAs, miR-192, miR-29a, miR-30d and miR-320a, as potential signature miRNAs that distinguished IFG and T2D. Of these microRNAs, miR-144 that promotes erythropoiesis has been found to be highly up-regulated. Increased circulating level of miR-144 has been found to correlate with down-regulation of its predicted target, insulin receptor substrate 1 (IRS1) at both mRNA and protein levels. We could also experimentally demonstrate that IRS1 is indeed the target of miR-144.

Conclusion

We demonstrate that peripheral blood microRNAs can be developed as unique biomarkers that are reflective and predictive of metabolic health and disorder. We have also identified signature miRNAs which could possibly explain the pathogenesis of T2D and the significance of miR-144 in insulin signaling.