Xia Shan

miR‐181b modulates multidrug resistance by targeting BCL2 in human cancer cell lines

MicroRNAs (miRNAs) are short noncoding RNA molecules, which posttranscriptionally regulate genes expression and play crucial roles in diverse biological processes, such as development, differentiation, apoptosis and proliferation. Here, we investigated the possible role of miRNAs in the development of multidrug resistance (MDR) in human gastric and lung cancer cell lines. We found that miR‐181b was downregulated in both multidrug‐resistant human gastric cancer cell line SGC7901/vincristine (VCR) and multidrug‐resistant human lung cancer cell line A549/cisplatin (CDDP), and the downregulation of miR‐181b in SGC7901/VCR and A549/CDDP cells was concurrent with the upregulation of BCL2 protein, compared with the parental SGC7901 and A549 cell lines, respectively. In vitro drug sensitivity assay demonstrated that overexpression of miR‐181b sensitized SGC7901/VCR and A549/CDDP cells to anticancer drugs, respectively. The luciferase activity of a BCL2 3′‐untranslated region‐based reporter construct in SGC7901/VCR and A549/CDDP cells suggests that a new target site in the 3′UTR of BCL2 of the mature miR‐181s (miR‐181a, miR‐181b, miR‐181c and miR‐181d) was found. Enforced miR‐181b expression reduced BCL2 protein level and sensitized SGC7901/VCR and A549/CDDP cells to VCR‐induced and CDDP‐induced apoptosis, respectively. Taken together, our findings suggest that miR‐181b could play a role in the development of MDR in both gastric and lung cancer cell lines, at least in part, by modulation of apoptosis via targeting BCL2.

−765G>C and 8473T>C polymorphisms of COX-2 and cancer risk: a meta-analysis based on 33 case–control studies

Cyclooxygenase-2 (COX-2) is an inducible enzyme converting arachidonic acid to prostaglandins and playing important roles in cancer etiology. The −765G>C and 8473T>C polymorphisms have been implicated in cancer risk. However, the results on the association between the two COX-2 polymorphisms and cancer risk are conflicting. To derive a more precise estimation of the association between them, we performed a meta-analysis of 8,090 cancer cases and 11,010 controls concerning −765G>C polymorphism and 14,283 cancer cases and 15,489 controls concerning 8473T>C polymorphism from 33 case–control studies. We used odds ratios (ORs) with 95% confidence intervals (CIs) to assess the strength of the association. Overall, individuals with the −765GC or GC/CC genotypes were associated with higher cancer risk than those with the −765GG genotype and in the stratified analysis this effect maintained in colorectal carcinoma or esophageal cancer of Asian descents. Overall, no significant cancer risk of 8473T>C polymorphism was found. Stratified by cancer types, the variant 8473CC was associated with a decreased risk in breast cancer, compared with the TT or TC/TT genotypes and in lung cancer subgroup after sensitive analysis, there was a decreased risk in CC versus TT, TC versus TT and the dominant models. Moreover, a decreased risk of lung cancer was observed among smokers in the dominant model. In summary, this meta-analysis suggesting that −765G>C may cause an increased risk of colorectal carcinoma and esophageal cancer in Asian descents while 8473T>C polymorphism may cause a decreased risk of breast and lung cancer.

A six-microRNA panel in plasma was identified as a potential biomarker for lung adenocarcinoma diagnosis

Differently expressed microRNAs (miRNAs) in the plasma of lung adenocarcinoma (LA) patients might serve as biomarkers for LA detection. MiRNA expression profiling was performed using Exiqon panels followed by the verification (30 LA VS. 10 healthy controls (HCs)) with quantitative reverse transcription polymerase chain reaction (qRT-PCR) in the screening phase. Identified miRNAs were confirmed through training (42 LA VS. 32 HCs) and testing stages (66 LA VS. 62 HCs) by using qRT-PCR based absolute quantification methods. A total of six up-regulated plasma miRNAs (miR-19b-3p, miR-21-5p, miR-221-3p, miR-409-3p, miR-425-5p and miR-584-5p) were identified. The six-miRNA panel could discriminate LA patients from HCs with areas under the receiver operating characteristic curve of 0.72, 0.74 and 0.84 for the training, testing and the external validation stage (33 LA VS. 30 HCs), respectively. All the miRNAs identified except miR-584-5p were significantly up-regulated in LA tissues. MiR-19-3p, miR-21-5p, miR-409-3p and miR-425-5p showed high expression in arterial plasma with borderline significance. Additionally, miR-19-3p, miR-21-5p and miR-221-3p were significantly up-regulated in exosomes extracted from LA peripheral plasma samples. In conclusion, we identified a six-miRNA panel in peripheral plasma which might give assistance to the detection of LA at least for Asian population to a certain extent.

Involvement of miR-143 in cisplatin resistance of gastric cancer cells via targeting IGF1R and BCL2

We investigated the possible role of miR-143 in the development of cisplatin resistance in human gastric cancer cell line. miR-143 was detected by quantitative real-time PCR. Cisplatin resistance changes of cells was tested via MTT assay. Target genes of miR-143 were verified by dual-luciferase activity assay. Immunohistochemistry, immunofluorescence staining, Western blot, cell proliferation, and clonogenic and apoptosis assay were used to elucidate the mechanism of miR-143 in cisplatin resistance formation. miR-143 was downregulated in gastric cancer tissues and cell lines. It was also downregulated in cisplatin-resistant gastric cancer cell line SGC7901/cisplatin (DDP), which was concurrent with the upregulation of IGF1R and BCL2, compared with the parental SGC7901 cell line, respectively. Overexpressed miR-143 sensitized SGC7901/DDP cells to cisplatin. The luciferase activity suggested that IGF1R and BCL2 were both target genes of miR-143. Enforced miR-143 reduced its target proteins, inhibited SGC7901/DDP cells proliferation, and sensitized SGC7901/DDP cells to DDP-induced apoptosis. Our findings suggested that hsa-miR-143 could modulate cisplatin resistance of human gastric cancer cell line at least in part by targeting IGF1R and BCL2.

A panel of microRNA signature in serum for colorectal cancer diagnosis

Dysregulated expression of specific microRNAs (miRNAs) in serum has been recognised as promising diagnostic biomarkers for colorectal cancer (CRC). In the initial screening phase, a total of 32 differentially expressed miRNAs were selected by quantitative reverse transcription polymerase chain reaction (qRT-PCR) based Exiqon panel with 3 CRC pool samples and 1 normal control (NC) pool. Using qRT-PCR, selected serum miRNAs were further confirmed in training (30 CRC VS. 30 NCs) and testing stages (136 CRC VS. 90 NCs). We identified that serum levels of miR-19a-3p, miR-21-5p and miR-425-5p were significantly higher in patients with CRC than in NCs. The areas under the receiver operating characteristic (ROC) curve of the three-miRNA panel were 0.86, 0.74 and 0.87 for the training, testing and the external validation stages (30 CRC VS. 18 NCs), respectively. Significantly, elevated expression of the three miRNAs was also observed in CRC tissues (n = 24). Furthermore, the expression levels of the three miRNAs were significantly elevated in exosomes from CRC serum samples (n = 10). In conclusion, we identified a serum three-miRNA panel for the diagnosis of CRC.

A novel serum microRNA signature to screen esophageal squamous cell carcinoma

Circulating microRNAs (miRNAs) have been used as promising diagnostic biomarkers for esophageal squamous cell carcinoma (ESCC). We performed miRNA expression profiling using quantitative reverse transcription polymerase chain reaction (qRT‐PCR) based Exiqon panels from three ESCC pools and one normal control (NC) pool samples. Using qRT‐PCR, identified serum miRNAs were further confirmed in training (32 ESCC vs. 32 NCs) and testing stages (108 ESCC vs. 96 NCs). Consequently, five serum miRNAs (miR‐20b‐5p, miR‐28‐3p, miR‐192‐5p, miR‐223‐3p, and miR‐296‐5p) were significantly overexpressed in ESCC compared with NCs. The diagnostic value of the 5‐miRNA signature was validated by an external cohort (60 ESCC vs. 60 NCs). The areas under the receiver operating characteristic curve (ROC) of the 5‐miRNA signature were 0.753, 0.763, and 0.966 for the training, testing, and the external validation stages, respectively. The expression levels of the miRNAs were also determined in tissues, arterial serum, and exosomes. MiR‐20b‐5p, miR‐28‐3p, and miR‐192‐5p were significantly upregulated in ESCC tissues, while miR‐296‐5p was overexpressed in ESCC serum exosomes. In conclusion, we identified a 5‐miRNA signature in serum for the detection of ESCC.

A six-microRNA signature in plasma was identified as a potential biomarker in diagnosis of esophageal squamous cell carcinoma

The differential expression of microRNAs (miRNAs) in plasma of esophageal squamous cell carcinoma (ESCC) patients may serve as a diagnostic biomarker. A four-stage study was conducted to identify plasma miRNAs with potential in detecting ESCC. Exiqon panels (2 ESCC pools vs. 1 normal control (NC) pool) were applied in the screening phase to obtain miRNA profiles. The identified miRNAs were further evaluated through training (36 ESCC VS. 42 NCs) and testing stages (101 ESCC VS. 113 NCs) with qRT-PCR assays. A six-miRNA signature including up-regulated miR-106a, miR-18a, miR-20b, miR-486-5p, miR-584 and down-regulated miR-223-3p in ESCC was identified. The signature could accurately discriminate ESCC patients from NCs with areas under the receiver operating characteristic curve of 0.935, 0.959 and 0.966 for the training, testing and the additional validation stage (41 ESCC VS. 50 NCs), respectively. MiR-106a and miR-584 were significantly up-regulated in tumor tissues with qRT-PCR assays. And miR-584 was also up-regulated in ESCC tissues from TCGA database. In addition, exosomal miR-223-3p and miR-584 were consistently dysregulated with those in plasma and could also act as biomarkers in diagnosis of ESCC. In conclusion, we identified a six-miRNA signature in plasma which could act as a non-invasive biomarker in detection of ESCC.

MiR-28-3p as a potential plasma marker in diagnosis of pulmonary embolism

OBJECTIVES Circulating miRNAs have been reported to have potential in detecting various diseases. However, few studies explored differentially expressed miRNAs in plasma of patients with pulmonary embolism (PE). Our study is to identify plasma miRNAs which can serve as potential biomarkers of PE. MATERIALS AND METHODS Exiqon miRCURY Ready-to-Use PCR Human panel I+II V1.M was conducted to identify differently expressed miRNAs in pooled plasma samples of PE patients compared with normal controls. Expressions of identified miRNAs were assessed in 37 PE patients as well as matched normal individuals followed by validation on six Beagle dogs by quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS Twelve miRNAs were identified from the screening phase. Moreover, miR-134, previously reported related with PE, and hypoxia-induced miR-210 were also submitted to the validation phase. Only miR-28-3p was found significantly elevated in the plasma of PE patients. Compared with the level of plasma miR-28-3p of the dogs before PE, the elevated miR-28-3p did not alter significantly at 1, 2, 4 and 6h after PE. The area under the receiver operating characteristic (ROC) curve of plasma miR-28-3p was 0.792 (95% confidence interval: 0.689-0.896). KEGG pathway analysis showed that miR-28-3p might involve in PE related pathways such as inositol phosphate metabolism and phosphatidylinositol signaling system. CONCLUSION Our study indicated that elevated plasma miR-28-3p could be used as a non-invasive and stable biomarker in the detection of PE. Further researches on the miRNA are warranted.

A three-microRNA signature for lung squamous cell carcinoma diagnosis in Chinese male patients

Various studies have demonstrated the diagnostic value of microRNA (miRNA) for lung cancer, but miRNA signatures varied between different subtypes. Whether serum miRNAs could be used as biomarkers in lung squamous cell carcinoma (SCC) remains unknown. Using quantitative real-time polymerase chain reaction (qRT-PCR) based Exiqon panel, 38 differentially expressed miRNAs were identified from 3 male lung SCC pool samples and 1 normal control (NC) pool in the initial screening phase. After the training (24 SCC VS. 15 NCs), testing (44 SCC VS. 57 NCs) and external validation (34 SCC VS. 36 NCs VS. 10 pulmonary hamartoma) processes via qRT-PCR, we identified a three-miRNA panel ((miR-106a-5p, miR-20a-5p and miR-93-5p) to be a potential diagnostic marker for male lung SCC patients. The areas under the receiver operating characteristic (ROC) curve of the three-miRNA panel for the training, testing and validation phases were 0.969, 0.881 and 0.954 respectively. In addition, this signature could also differentiate lung SCC from pulmonary hamartoma (AUC=0.900). The 3 miRNAs were consistently up-regulated in lung SCC tissues (23 SCC VS. 23 NCs) and serum exosomes (17 SCC VS. 24 NCs). Moreover, expression of the 3 miRNAs was decreased in arterial serum (n = 3). In conclusion, we established a three-miRNA signature in the peripheral serum with considerable clinical value in the diagnosis of male lung SCC patients.

Differential expression levels of plasma microRNA in Hashimoto's disease

BACKGROUND The altered expression of circulating miRNAs has been discovered in many autoimmune diseases (ADs). With rare existing research, it is still unclear in Hashimotos thyroiditis (HT). We detected plasma miRNA expression of HT patients in this three-stage designed study. METHODS Differently expressed miRNAs (4 HT pools vs. 1 normal control pool) were identified using quantitative reverse transcription polymerase chain reaction (qRT-PCR) based Exiqon panel (miRCURY-Ready-to-Use- PCR-Human- panel-I+II-V1.M) in the initial discovery stage. These miRNAs were then confirmed in the training stage and further validated in the testing stage using qRT-PCR with 64 (32 HT vs. 32 NCs) and 136 samples (68 HT vs. 68 NCs), respectively. RESULTS A total of 10 miRNAs showed differential expression through the training stage. For further validation in the testing stage, expression of 6 miRNAs (miR-205, miR-20a-3p, miR-375, miR-296, miR-451, miR-500a) were consistent with those in the training stage. Combination results showed that these 6 miRNAs were significantly up-regulated in peripheral plasma of HT patients compared with normal controls (P<0.05). In addition, the six-miRNA signature was evaluated to be a potential diagnostic marker of HT. The areas under the receiver operating characteristic curve of the signature were 0.80, 0.75 and 0.69 for the training, testing and the combined stages, respectively. Three miRNAs were associated with TSH levels in HT patients (miR-451, P=0.043; miR-375, P=0.043; miR-500a, P=0.043). Additionally, miR-20a-3p was related with TgAb level (P=0.046). CONCLUSIONS We identified a miRNA signature including six dysregulated plasma miRNAs which could act as a diagnostic marker in plasma of HT, providing more evidence and better understanding for the association between circulating miRNAs and autoimmune diseases.