Tusheng Song

MicroRNA profiling of human gastric cancer

MicroRNAs are a group of small non-coding RNAs that modulate gene expression. The de-regulation of microRNA expression has been found in several types of cancer. To study the role of microRNAs in gastric cancer (GC), we analyzed the expression profile of 847 microRNAs in GC from Chinese patients. Total RNA was used for hybridization on the miRCURY LNA Array (v. 11.0), which contains probes specific for 847 human microRNAs. The results from the miRNA microarray analysis were validated by real-time RT-PCR. A total of 24 miRNAs with a more than 2-fold change were differentially expressed between normal gastric tissue and GC. Of these, 22 miRNAs (miR-223, miR-106b, miR-147, miR-34a, miR-130b*, miR-106a, miR-18a, miR-17, miR-98, miR-616*, miR-181a-2*, miR-185, miR-1259, miR-601, miR-196a*, miR-221*, miR-302f, miR-340*, miR-337-3p, miR-520c-3p, miR-575 and miR-138) were significantly up-regulated in GC (P<0.05), whereas only miR-638 and miR-378 were significantly down-regulated in GC (P<0.05) compared to normal gastric tissue. The expression of miR-185 and miR-638, as measured by miRNA microarray analysis, was in agreement with the expression level of these microRNAs found by real-time RT-PCR in the same samples. Our results show that microRNAs are de-regulated in GC, suggesting the involvement of these genes in the development and progression of gastric cancer.

MicroRNA-302b suppresses cell proliferation by targeting EGFR in human hepatocellular carcinoma SMMC-7721 cells

BackgroundMicroRNAs are regulators that can play an essential role in tumorigenesis. Although miR-302 families have been suggested to be tumor repressors in human cancer, the mechanism by which they suppress tumor development remains to be defined. In this study, we discover that miR302b suppresses tumor proliferation may due to directly targeting EGFR in human hepatocellular carcinoma (HCC).MethodsQRT-PCR was used to assess miR-302b and EGFR expression in 27 pairs of clinical hepatocellular carcinoma tissues and their corresponding adjacent nontumorous liver tissues. MTT, colony formation, immunofluorescence staining, and cell cycle assays were used to examine the tumor suppressor role of miR302b in cell proliferation. Luciferase assays were performed to assess the EGFR was a novel target of miR-302b. Western blot assay was used to validate the protein expression level.ResultsWe demonstrated that miR-302b was frequently down-regulated, whereas EGFR was up-regulated in 27 pairs of clinical HCC and non-tumorous counterparts. The dual-luciferase reporter assays revealed that EGFR was a novel target of miR-302b. Re-expression of miR-302b resulted in the inhibition of proliferation in hepatocellular carcinoma SMMC-7721 cells. The silencing of EGFR by miR-302b or siEGFR led to down-regulation of proliferation-related proteins, such as AKT2, CCND1, and CDK2.ConclusionmiR-302b suppresses HCC growth may due to targeting the EGFR/AKT2/CCND1 pathway.

miR-338-3p Suppresses Gastric Cancer Progression through a PTEN-AKT Axis by Targeting P-REX2a

Results from recent studies suggest that aberrant microRNA expression is common in numerous cancers. Although miR-338-3p has been implicated in hepatocellular carcinoma, its role in gastric cancer is unknown. To this end, we report that miR-338-3p is downregulated in both gastric cancer tissue and cell lines. Forced expression of miR-338-3p inhibited cell proliferation and clonogenicity and induced a G1–S arrest as well as apoptosis in gastric cancer cells. Furthermore, P-Rex2a (PREX2) was identified as a direct target of miR-338-3p, and silencing P-Rex2a resulted in the same biologic effects of miR-338-3p expression in gastric cancer cells. Furthermore, both enforced expression of miR-338-3p or silencing of P-Rex2a resulted in activation of PTEN, leading to a decline in AKT phosphorylation. Also, miR-338-3p markedly inhibited the in vivo tumorigenicity in a nude mouse xenograft model system. These results demonstrate that miR-338-3p affects gastric cancer progression through PTEN—AKT signaling by targeting P-Rex2a in gastric cancer cells, which posits miR-338-3p as a novel strategy for gastric cancer treatment. Implications: miR-338-3p acts as a novel tumor suppressor that blocks the growth of gastric cancer cells through PTEN—PI3K signaling by targeting P-Rex2a. Mol Cancer Res; 12(3); 313–21. ©2013 AACR.

A novel chronic stress-induced shift in the Th1 to Th2 response promotes colon cancer growth.

Epidemiological data have shown that stress and other psychological factors might influence cancer onset and progression. However, to date, the mechanisms are not well understood. In the present study, we used chronic exposure to a scream as a novel form of sound stress to explore the influence of the chronic stress burden on colon cancer progression, and changes in the immune system were observed. Chronic exposure to scream sound stress induced freezing behavior in the mice and decreased the bodyweight gain. It also caused changes in the adrenal gland and increased serum corticosterone and norepinephrine levels. Cytokine microarray analysis showed changes in the levels of Th1 and Th2 cytokines. The chronic scream sound stress caused a shift from the Th1 to the Th2 response both in the circulation and in tumor-infiltrated lymphocytes, and it promoted colon cancer progression significantly. Taken together, chronic scream sound stress can be conveniently used as a novel chronic stress model. Chronic stress contributes to colon cancer progression and induces a Th1/Th2 imbalance in the mouse immune system, which is considered critical during cancer progression.

Serum peptidome profiling in patients with gastric cancer

To identify discriminating protein patterns in serum samples between gastric cancer patients (early and advanced stages) and healthy controls. We used magnetic bead-based separation followed by matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) to identify patients with gastric cancer. In total, serum samples from 62 gastric cancer patients (32 in the training set and 30 in the test set; 19 of which had early-stage tumors and 43 of which had advanced-stage tumors) and 64 healthy controls (32 in the training set and 32 in the test set) were analyzed. The mass spectra, analyzed using ClinProTools software, distinguished between cancer patients and healthy individuals based on three different algorithm models. In the training set, patients with gastric cancer could be identified with a mean sensitivity of 94.7% and a mean specificity of 99%. Similar results were obtained with the test set, showing 79.3% sensitivity and 86.5% specificity. Our study demonstrates the high sensitivity and specificity of screening serum protein patterns using MALDI-TOF MS for the identification of patients with gastric cancer.

A new stress model, a scream sound, alters learning and monoamine levels in rat brain

Most existing animal models for stress involve the simultaneous application of physical and psychological stress factors. In the current study, we described and used a novel psychological stress model (scream sound stress). To study the validity of it, we carried out acute and chronic scream sound stress. First, adult Sprague-Dawley (SD) rats were randomly divided into white noise, stress and background groups. The white noise group and stress group were treated with white noise and scream sound for 4h in the morning respectively. Compared with white noise and background groups, exposure to acute scream sound increased corticosterone (CORT) level and decreased latency in Morris water maze (MWM) test. The levels of noradrenaline (NE), dopamine (DA), 5-hydroxytryptamine (5-HT), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA) were altered in the striatum, hypothalamus and hippocampus of stress rats. Second, adult SD rats were randomly divided into background and stress groups, which were treated with scream sound for three weeks. Exposure to chronic scream sound suppressed body weight gain, increased corticosterone (CORT) level, influenced the morphology of adrenal gland, improved spleen and thymus indices, and decreased latency in MWM test. NE, DA, DOPAC, HVA and 5-HIAA levels were also altered in the brain of stress rats. Our results suggested that scream sound, as a novel stressor, facilitated learning ability, as well as altered monoamine levels in the rat brain. Moreover, scream sound is easy to apply and can be applied in more animals at the same time.

miRNA-302b Suppresses Human Hepatocellular Carcinoma by Targeting AKT2

miRNAs (miR) play a critical role in human cancers, including hepatocellular carcinoma. Although miR-302b has been suggested to function as a tumor repressor in other cancers, its role in hepatocellular carcinoma is unknown. This study investigated the expression and functional role of miR-302b in human hepatocellular carcinoma. The expression level of miR-302b is dramatically decreased in clinical hepatocellular carcinoma specimens, as compared with their respective nonneoplastic counterparts, and in hepatocellular carcinoma cell lines. Overexpression of miR-302b suppressed hepatocellular carcinoma cell proliferation and G1–S transition in vitro, whereas inhibition of miR-302b promoted hepatocellular carcinoma cell proliferation and G1–S transition. Using a luciferase reporter assay, AKT2 was determined to be a direct target of miR-302b. Subsequent investigation revealed that miR-302b expression was inversely correlated with AKT2 expression in hepatocellular carcinoma tissue samples. Importantly, silencing AKT2 recapitulated the cellular and molecular effects seen upon miR-302b overexpression, which included inhibiting hepatocellular carcinoma cell proliferation, suppressing G1 regulators (Cyclin A, Cyclin D1, CDK2) and increasing p27Kip1 phosphorylation at Ser10. Restoration of AKT2 counteracted the effects of miR-302b expression. Moreover, miR-302b was able to repress tumor growth of hepatocellular carcinoma cells in vivo. Implications: Taken together, miR-302b inhibits HCC cell proliferation and growth in vitro and in vivo by targeting AKT2. Mol Cancer Res; 12(2); 190–202. ©2013 AACR.

MicroRNA Profiling in Human Colon Cancer Cells during 5-Fluorouracil-Induced Autophagy

Autophagy modulation is now recognized as a potential therapeutic approach for cancer (including colorectal cancer), yet the molecular mechanisms regulating autophagy in response to cellular stress are still not well understood. MicroRNAs (miRNAs) have been found to play important roles in controlling many cellular functions, including growth, metabolism and stress response. The physiological importance of the miRNA-autophagy interconnection is only beginning to be elucidated. MiRNA microarray technology facilitates analysis of global miRNA expression in certain situations. In this study, we explored the expression profile of miRNAs during the response of human colon cancer cells (HT29s) to 5-FU treatment and nutrient starvation using miRNA microarray analysis. The alteration of miRNA expression showed the same pattern under both conditions was further testified by qRT-PCR in three human colon cancer cell lines. In addition, bioinformatic prediction of target genes, pathway analysis and gene network analysis were performed to better understand the roles of these miRNAs in the regulation of autophagy. We identified and selected four downregulated miRNAs including hsa-miR-302a-3p and 27 upregulated miRNAs under these two conditions as having the potential to target genes involved in the regulation of autophagy in human colon cancer cells. They have the potential to modulate autophagy in 5-FU-based chemotherapy in colorectal cancer.

Identification of Novel Low Molecular Weight Serum Peptidome Biomarkers for Non-Small Cell Lung Cancer (NSCLC)

To identify discriminating protein patterns in serum samples among non‐small cell lung cancer (NSCLC), chronic obstructive pulmonary disease (COPD), pneumonia, and healthy controls. To discover specific low molecular weight (LMW) serum peptidome biomarkers and establish a diagnostic pattern for NSCLCby using proteomic technology.

MicroRNA‑101 suppresses progression of lung cancer through the PTEN/AKT signaling pathway by targeting DNA methyltransferase 3A

It is well established that transcriptional silencing of critical tumor suppressor genes by DNA methylation is a fundamental process in the initiation of lung cancer. However, the involvement of microRNAs (miRNAs) in restoring abnormal DNA methylation patterns in lung cancer is not well understood. Therefore, and since miRNA-101 is complementary to the 3′-untranslated region of DNA methyltransferase 3A (DNMT3A), we investigated whether miRNA-101 could restore normal DNA methylation patterns in lung cancer cell lines. Bioinformatics has indicated that DNMT3A is a major target of miR-101. In addition, the overexpression of miR-101 downregulates DNMT3A. Using a methylation-specific polymerase chain reaction assay, we demonstrated that methylation of the phosphatase and tensin homolog (PTEN) promoter was reduced in A549 cells transfected with miR-101, but not in the transfected control. Furthermore, overexpression of miR-101 and silencing of DNMT3A suppressed lung cell proliferation and S/G2 transition, and increased apoptosis through the PTEN/AKT pathway in vitro. Furthermore, we observed the opposite phenomenon in A549 cells transfected with a miR-101 inhibitor. Subsequent investigation revealed that overexpression of miR-101 significantly inhibited the tumorigenicity of A549 cells in a nude mouse xenograft model. These results demonstrate that miR-101 affects lung cancer progression through the PTEN/AKT signaling pathway by targeting DNMT3A in lung cells, suggesting that miR-101 may be a novel potential therapeutic strategy in lung cancer treatment.