Roles of miR-200 family members in lung cancer: more than tumor suppressors.

miRNAs are a class of single-stranded noncoding RNAs, which have no coding potential, but modulate many molecular mechanisms including cancer pathogenesis. miRNAs participate in cell proliferation, differentiation, apoptosis, as well as carcinogenesis or cancer progression, and their involvement in lung cancer has been recently shown. They are suggested to have bidirectional functions on important cancer-related genes so as to enhance or attenuate tumor genesis. Epithelial-mesenchymal transition (EMT) is a fundamental process which contributes to integrity of organogenesis and tissue differentiation as well as tissue repair, organ fibrosis and the progression of carcinoma, and several miRNAs were suggested to form the network regulating EMT in lung cancer, among which, miR-200 family members (miR-200a, miR-200b, miR-200c, miR-429 and miR-141) play crucial roles in the suppression of EMT.

Microarray analysis reveals a potential role of LncRNAs expression in cardiac cell proliferation.

BACKGROUND: Long non-coding RNAs (LncRNAs) have been identified to play important roles in epigenetic processes that underpin organogenesis. However, the role of LncRNAs in the regulation of transition from fetal to adult life of human heart has not been evaluated. METHODS: Immunofiuorescent staining was used to determine the extent of cardiac cell proliferation. Human LncRNA microarrays were applied to define gene expression signatures of the fetal (13-17 weeks of gestation, n = 4) and adult hearts (30-40 years old, n = 4). Pathway analysis was performed to predict the function of differentially expressed mRNAs (DEM). DEM related to cell proliferation were selected to construct a lncRNA-mRNA co-expression network. Eight lncRNAs were confirmed by quantificational real-time polymerase chain reaction (n = 6). RESULTS: Cardiac cell proliferation was significant in the fetal heart. Two thousand six hundred six lncRNAs and 3079 mRNAs were found to be differentially expressed. Cell cycle was the most enriched pathway in down-regulated genes in the adult heart. Eight lncRNAs (RP11-119 F7.5, AX747860, HBBP1, LINC00304, TPTE2P6, AC034193.5, XLOC_006934 and AL833346) were predicted to play a central role in cardiac cell proliferation. CONCLUSIONS: We discovered a profile of lncRNAs differentially expressed between the human fetal and adult heart. Several meaningful lncRNAs involved in cardiac cell proliferation were disclosed.

Potent antitumor activity of HSP90 inhibitor AUY922 in adrenocortical carcinoma.

The objective of this study is to investigate the expression of HSP90 and the effect of HSP90 inhibitor AUY922 in ACC. The expression of HSP90 was measured in tissue samples from 36 human sporadic adrenocortical tumors by immunohistochemistry, Western blotting, and real-time PCR. The effect of AUY922 was tested on SW13 and H295R cells by evaluating cell viability and apoptosis in vitro. Transwell assay was performed to evaluate the migration of SW13 cells after different concentrations of AUY922. Western blot, real-time PCR, and immunohistochemistry revealed that both HSP90 mRNA and protein were obviously expressed in a higher degree in ACC tissues than ACA tissues and normal adrenal tissues (P < 0.01). Positive staining for HSP90 was found in 15 of 20 ACCs (75.00 %) and in 3 of 16 (18.75 %) ACAs. There existed the significant statistical difference (P < 0.001). AUY922 inhibited the proliferation of ACC cells in a time- and concentration-dependent manner, and increasing apoptosis was observed in tumor cells treated with the HSP90 inhibitor. Finally, migration of SW13 cells was distinctly suppressed after undergoing treatment with AUY922. Our data suggest that the specific HSP90 inhibitor AUY922 can play a therapeutic role in treatment of ACC and, thus, HSP90 could qualify as a promising new target in ACC.

[Expression and clinical significance of circadian gene Per2 in non-small cell lung cancer].

OBJECTIVE: To detect the expression of Per2 in non-small cell lung cancer (NSCLC), and analyze its clinical significance. METHODS: The expression of Per2 was determined in 60 NSCLC and 20 normal lung tissues by immunohistochemical assay, and the relationship between Per2 expression and clinicopathological features was analyzed. RESULTS: The positive expression rates of Per2 in NSCLC and normal lung tissues were 71.7% and 95.0%, respectively (P < 0.05). The expression of Per2 in NSCLC was correlated with pathological differentiation and TNM stage (P < 0.05). CONCLUSION: The expression of Per2 in NSCLC is decreased. The negative expression of Per2 may contribute to the development and invasion in NSCLC.

Central zone of myocardial infarction: a neglected target area for heart cell therapy.

The purpose of this study was to investigate the fate of transplanted cells in the central zone of myocardial infarction (MI), and to clarify the relationship between the injection-site impact and the efficacy of cell therapy. MI was created by coronary ligation in female rats. Three weeks later, 3-million labelled male bone marrow mesenchymal stem cells (BMSCs) were directly injected into the border (BZC group) or central zone (CZC group) of MI area. As a control, culture medium was injected into the same sites. Cell survival was evaluated by quantitative real-time polymerase chain reaction, and apoptosis was assayed with TUNEL and caspase-3 staining. Four weeks after transplantation, heart function and cardiac morphometry were evaluated by echocardiography and Masson's Trichrome staining, respectively. Angiogenesis and myogenesis were detected by immunofluorescence staining. After cell transplantation into the border or central zone, there was no cell migration between the different zones of MI. BMSCs in the CZC group exhibited no difference in apoptotic percentage, in the long-term survival, when compared with those in the BZC group. However, they did effectively promote angiogenesis and cellular myogenic differentiation. Although cell delivery in the central zone of MI had no effect on the recovery of heart function compared with the BZC group, the retained BMSCs could still increase the scar thickness, and subsequently exhibit a trend in the reverse remodelling of ventricular dilation. Hence, we concluded that the central zone of MI should not be ignored during cell-based therapy. Multiple site injection (border+central zone) is strongly recommended during the procedure of cell transplantation.

Unloading the infarcted heart affect MMPs-TIMPs axis in a rat cardiac heterotopic transplantation model.

Ventricular assist devices may function as a bridge to recovery or heart transplantation, however, little is known about its mechanisms. This study examined the role of matrix metalloproteinases (MMP)-tissue inhibitors of metalloproteinases (TIMP) axis in the process of recovery after unloading in a rat ischemic-induce heart failure (HF) model. Myocardial infarction model was created with the coronary artery ligation. The infarcted rats hearts were unloaded by heterotopic cardiac transplantation (n=14). 2 weeks later, the function of normal and infarcted hearts with or without loading was evaluated by Langendorff perfusion model. The hearts were then harvested and prepared for the study of expression of MMPs and TIMPs. Developed pressure in the unloading group was higher than the loading group (P=0.0074). Unloading increased the ratio of TIMP-1-MMP-1(1.38±0.11 vs. 0.76±0.09, P<0.05), TIMP-2-MMP-2 (1.06±0.10 vs. 0.33±0.07, P<0.01), TIMP-3-MMP-9(1.07±0.08 vs. 0.59±0.06, P<0.05). Although MMP-1, 2, 9 were downregulated (P<0.01, 0.01, 0.05, respectively), TIMP-2 and TIMP-3 upregulated (P<0.01, 0.05, respectively), MMP-7 and TIMP-1 was not affected significantly. The infarcted cardiac function could be improved by unloading. It was attributed to downregulation of MMP-1, 2 and 9, and upregulation of TIMP-2 and -3, and furthermore, the ratio of TIMPs to MMPs was increased, which might be more sensitive than sole MMPs or TIMPs for the judgment of myocardial matrix homeostasis.

SDF-1α inhibits hypoxia and serum deprivation-induced apoptosis in mesenchymal stem cells through PI3K/Akt and ERK1/2 signaling pathways.

Bone marrow-derived mesenchymal stem cells (BMSCs) have been demonstrated to be a promising cell sources for cardiac regeneration. Poor survival rate of transplanted BMSCs in infarcted myocardium attenuated its clinical application. It's reported that stromal-derived factor-1 (SDF-1) could protect progenitor cells including endothelial progenitor cells and embryonic stem cells from apoptosis. But little is known whether SDF-1α protein has the same protective effects on BMSCs under conditions of hypoxia and serum deprivation (hypoxia/SD). In present study, we verified that SDF-1α (0.50-2.0 µg/ml) inhibited hypoxia/SD induced apoptosis of BMSCs through mitochondrial pathway. After administration of SDF-1α, the loss of mitochondrial membrane potential and cytochrome c released from mitochondria to cytosol were significantly inhibited, and caspase 3 activity also declined. Furthermore, the effect of SDF-1α on mitochondrial pathway was neutralized by using PI3K inhibitor (Wortmannin) and ERK1/2 inhibitor (U0126). Our observations suggested that SDF-1α inhibits hypoxia/SD induced BMSCs apoptosis through PI3K/Akt and ERK1/2 signaling pathways. These data also imply that the anti-apoptotic effect mediated by SDF-1α may enhance cell survival after cell transplantation.

[miRNA changes in the reverse remodeling heart of rats].

OBJECTIVE: To establish a reverse remodeling heart model in rats and observe collagen and TGF-ß expression and relevant microRNAs changes during reverse remodeling. METHODS: Lewis rats were divided into four groups including sham (NL, n = 10), abdominal aortic constriction (AAC, n = 10), heterotopic transplantation of abdominal aortic constriction (AAC-HT, n = 9) and heterotopic transplantation of normal heart (HT, n = 8). Left ventricular wall thickness and LV cavity were measured by echocardiography. The cardiomyocyte cross-sectional area (CSA) was determined on HE stained sections. Immunohistochemical and qRT-PCR were used to detect collagen and TGF-ß expressions. miRNAs were detected by MicroRNA microarray. RESULTS: Heart weight, left ventricular wall thickness and CSA were significantly increased in AAC hearts compared to those in the NL and AAC-HT hearts. The collagen and TGF-ß were increased in AAC hearts and further increased in AAC-HT hearts. miRNA microarray evidenced more than two folds changes on 82 miRNAs compared to NL (10 in AAC, 32 in AAC-HT and 40 in HT). CONCLUSION: Rat abdominal aortic constriction and heterotopic transplantation could be used as a reverse remodeling heart model and significant collagen and TGF-ß as well microRNA expression changes were evidenced in this model.

Microarray analysis of long non-coding RNA expression profiles in Marfan syndrome.

Long non-coding RNAs (lncRNAs) serve a crucial role in every aspect of cell biological functions as well as in a variety of diseases, including cardiovascular disease, cancer and nervous system disease. However, the differential expression profiles of lncRNAs in Marfan syndrome (MFS) have not been reported. The aim of the present study was to identify potential target genes behind the pathogenesis of MFS by analyzing microarray profiles of lncRNA in aortic tissues from individuals with MFS and normal aortas (NA). The differentially expressed lncRNA profiles between MFS (n=3) and NA (n=4) tissues were analyzed using microarrays. Bioinformatics analyses were used to further investigate the candidate lncRNAs. Reverse transcription-quantitative (RT-qPCR) was applied to validate the results. In total, the present study identified 294 lncRNAs (245 upregulated and 49 downregulated) and 644 mRNAs (455 upregulated and 189 downregulated) which were differential expressed between MFS and NA tissues (fold change ≥1.5; P<0.05). Gene Ontology enrichment analysis indicated that the differentially expressed mRNAs were involved in cell adhesion, elastic fiber assembly, extracellular matrix (ECM) organization, the response to virus and the inflammatory response. Kyoto Encyclopedia of Gene and Genomes pathway analysis indicated that the differentially expressed mRNAs were mainly associated with focal adhesion, the ECM-receptor interaction, the mitogen-activated protein kinase signaling pathway and the tumor necrosis factor signaling pathway. The lncRNA-mRNA coexpression network analysis further elucidated the interaction between the lncRNAs and mRNAs. A total of five lncRNAs (uc003jka.1, uc003jox.1, X-inactive specific transcript, linc-lysophosphatidic acid receptor 1 and linc-peptidylprolyl isomerase domain and WD repeat containing 1) with the highest degree of coexpression were selected and confirmed using RT-qPCR. In the present study, expression profiles of lncRNA and mRNA in MFS were revealed using microarray analysis. These results provided novel candidates for further investigation of the molecular mechanisms and effective targeted therapies for MFS.

[Concurrent standard dose of cisplatin, paclitaxel, and radiotherapy followed by surgery in treatment of thoracic esophageal carcinoma].

OBJECTIVE: To investigate the curative effect of incorporation of the regimen of standard dose of paclitaxel combined with cisplatin into concurrent radiotherapy as pre-operative treatment for patients with esophageal carcinoma. METHODS: Twenty-six patients with primary diagnosis of esophageal carcinoma, 17 in stage II and 9 in stage III, underwent conventional fractionated radiotherapy with a total dosage of 40 Gy (2 Gy per day, 5 doses per week). Paclitaxel were given intravenously at a dose of 135 mg/m(2) for 3 h on day1 and day 22. Cisplatin was given intravenously at a dose of 20 mg/m(2) on D1-D3 and D22-24. 4 - 6 weeks after the completion of chemo-radiotherapy, left thoracic incision and transhiatal esophagectomy with anastomosis in the neck was performed. The patients were followed up for 42.28 months. Kaplan-Meier method was used to analyze the overall survival (OS) and disease-free survival (DFS), and Log-rank test was performed to assess the survival rates statistical significance among groups. RESULTS: The radical resection rate was 96.15%. The pathologic response to chemoradiotherapy were grade I in 9 patients, grade II in 6 patients, and grade III in 11 patients. The pathological complete remission (PCR) rate was 42.31% (11/26). Toxicity grade 3 - 4 included leucopenia (7.69%, 2/26), thrombocytopenia (7.69%, 2/26), and radiation esophagitis (11.54%, 3/26). Surgery-related complications included anastomotic leakage (3.85%, 1/26), recurrent laryngeal nerve injury (7.69%, 2/26), and chylothorax (3.85%, 1/26). The 3- and 5-year overall survival rates were 62.96% and 54.56% respectively. The 3- and 5-year disease-free survival rates were 59.94% and 55.65% respectively. The 3-year overall survival rates of the patients with different pathologic responses were 25.40% (for those of grade I), 60% (for grade II), and 90.91% (for grade III) respectively (P < 0.05). The 5-year overall survival rates were 0 (for grade I), 60% (for grade II), and 81.82% (for grade III) respectively (P < 0.05). CONCLUSION: Preoperative chemoradiotherapy containing full dose of paclitaxel and cisplatin increases the 5-year overall survival for the patients with postoperative pathologic response grade II and above, and does not increase the treatment-related complications.

[Carinal resection and reconstruction, and bronchoplasty and pulmonary arterioplasty in the treatment of central-type lung cancer].

BACKGROUND: Boonchoplasty can not only remove tumor but also reserve lung tissue maximally, and it becomes an alternative choice for patient with poor pulmonary function who could not accept pneumonectomy. The aim of this study is to summarize the experience of carinal resection and reconstruction, bronchoplasty and pulmonary arterioplasty in the treatment of central-type lung cancer. METHODS: From March, 1987 to March, 2005, A total of 79 patients with central-type lung cancer underwent operation. The operations included: left bronchoplasty (34 cases) combined with pulmonary arterioplasty in 10 cases and partial resection of left atrium in 3 cases; right bronchoplasty (45 cases) combined with carinal resection in 14 cases and segmentplasty in 5 cases, pulmonary arterioplasty in 5 cases, partial resection of superior vena cava wall in 5 cases. RESULTS: There were no perioperative deaths. Twenty-eight cases (35.4%) had postoperative complication. The 1-, 3-and 5-year survival rate were 86.1%, 55.2% and 32.1% respectively. CONCLUSIONS: Proper selection of carinal resection, bronchoplasty and pulmonary arterioplasty can expand the indications. They can reduce the ratio of pneumonectomy and improve the postoperative quality of life and the prognosis of lung cancer patients.

microRNA miR-10b inhibition reduces cell proliferation and promotes apoptosis in non-small cell lung cancer (NSCLC) cells.

Lung cancer is one of the most common and serious types of cancer. Till now, the treatment of lung cancer has been unsatisfactory, which is associated with poor prognosis and high mortality. Therefore, there is an urgent requirement to investigate the molecular mechanisms underlying lung tumorigenesis. To study the potential function of miR-10b involved in the regulation of lung tumors, we monitored NSCLC cell behaviour including proliferation, apoptosis and cell cycle using CCK-8 and flow cytometry analysis. Real-time PCR was used to detect the expression levels of miR-10b in 75 NSCLC patients' tissues and Western blot was also used to analyze the expression level of genes correlated with apoptosis in NSCLC cells. miR-10b expression levels were higher in NSCLC tissues compared with an adjacent normal tissue control. Silencing of miR-10b inhibited cancer cell progress by arresting cell cycle progression in the G0/G1 phase and promoted apoptosis in NSCLC cells. Western blot analysis of miR-10b-silenced cells revealed up-regulation of apoptosis-inducing members Fas, FasL, Bax and caspase 3, and down-regulation of apoptosis-inhibiting factors Bcl-2 and PCNA. And, a significant inverse correlation between the level of miR-10b and klotho was observed, which has been demonstrated to be a novel tumor suppressor gene. A further in vivo tumor formation study in nude mice indicated that inhibition of miR-10b in lung cancer cells delayed the progress of tumor formation. These findings indicated that miR-10b might serve as a useful potential target for treatment of NSCLC.

miR-33a levels in hepatic and serum after chronic HBV-induced fibrosis.

BACKGROUND: Chronic hepatitis B virus (HBV) infection, which can lead to hepatic disease, has become a critical national healthcare problem, and many people die each year as a result of HBV infection and its complications. Although microRNA-33a (miR-33a) is a novel modulator of lipid and cholesterol metabolism, the role of miR-33a in the hepatic fibrogenesis is still unknown. Here, we aimed to explore the roles and mechanisms of miR-33a in liver fibrosis. METHODS: miR-33a expression in whole liver and serum samples was measured from chronic hepatitis B (CHB) patients by quantitative real-time PCR (qRT-PCR). In addition, different murine hepatic fibrosis models were produced to consolidate the results in human tissue. Human and murine primary liver fibrosis-associated cells were isolated and treated with transforming growth factor-ß1 (TGF-ß1). RESULTS: miR-33a expression levels in liver tissue significantly increased with a fibrosis progression manner in the human liver. Furthermore, serum miR-33a levels associated positively with progressing process of hepatic fibrosis. miR-33a was in particular increased in hepatic stellate cells (HSC) than other liver fibrosis-associated cells. Stimulation of HSCs with TGF-ß1 leads to a critical increase of miR-33a. Increasing miR-33a levels increased (whereas inhibiting miR-33a weakened) the activation role of TGF-ß1 in LX-2 cells, which might be a potential mechanism through moderating Smad7 expression. CONCLUSIONS: miR-33a may be a novel marker for HSC activation and hepatic fibrosis progress, suggesting a new therapeutic target in liver fibrosis.

Acute dysfunction of mechanical aortic valve as electrocardiographic mimic of acute left main coronary artery occlusion.

We report three patients with acute dysfunction of mechanical aortic valve prosthesis resulting in chest pain, concomitant cardiogenic shock, and electrocardiographic changes mimicking acute occlusion of the left main coronary artery whereas emergency coronary angiography revealed normal coronary arteries. During operation, abnormal proliferation of subvalvular pannus overgrowth on the inflow aspect of the prosthesis was found to impede normal prosthesis closure. We discuss the possible underlying mechanisms of electrocardiographic presentations.

Generation of disease-specific induced pluripotent stem cells from patients with different karyotypes of Down syndrome.

INTRODUCTION: Down syndrome (DS), a major cause of mental retardation, is caused by trisomy of some or all of human chromosome 21 and includes three basic karyotypes: trisomy 21, translocation, and mosaicism. The derivation of DS-specific induced pluripotent stem cells (iPSCs) provides us novel DS models that can be used to determine the DS mechanism and to devise therapeutic approaches for DS patients. METHODS: In the present study, fibroblasts from patients with DS of various karyotypes were reprogrammed into iPSCs via the overexpression of four factors: OCT4, SOX2, KLF4, and c-MYC, by using lentiviral vectors. The abilities of the iPSC-DS in the self-renewal and pluripotency in vitro and in vivo were then examined. RESULTS: The iPSC-DS showed characteristics similar to those of human embryonic stem cells, particularly the morphology, surface marker (SSEA4, TRA-1-60, and TRA-1-81) expression, pluripotent-specific transcription-factor expression levels, and methylation status of the OCT4 promoter. The pluripotency of iPSC-DS was also tested in vitro and in vivo. Embryoid bodies were formed and showed the expression of differentiated markers for three germ layers. Furthermore, iPSC-DS formed classic teratomas when injected into nonobese diabetic-severe combined immunodeficient (NOD-SCID) mice. CONCLUSIONS: iPSCs were generated from patients with DS. The iPSCs derived from different types of DS may be used in DS modeling, patient-care optimization, drug discovery, and eventually, autologous cell-replacement therapies.

MicroRNA-155 silencing enhances inflammatory response and lipid uptake in oxidized low-density lipoprotein-stimulated human THP-1 macrophages.

It has been proposed that the inflammatory response of monocytes/macrophages induced by oxidized low-density lipoprotein (oxLDL) is a key event in the pathogenesis of atherosclerosis. MicroRNA-155 (miR-155) is an important regulator of the immune system and has been shown to be involved in acute inflammatory response. However, the function of miR-155 in oxLDL-stimulated inflammation and atherosclerosis remains unclear. Here, we show that the exposure of human THP-1 macrophages to oxLDL led to a marked up-regulation of miR-155 in a dose-dependent manner. Silencing of endogenous miR-155 in THP-1 cells using locked nucleic acid-modified antisense oligonucleotides significantly enhanced oxLDL-induced lipid uptake, up-regulated the expression of scavenger receptors (lectinlike oxidized LDL receptor-1, cluster of differentiation 36 [CD36], and CD68), and promoted the release of several cytokines including interleukin (IL)-6, -8, and tumor necrosis factor α (TNF-α). Luciferase reporter assay showed that targeting miR-155 promoted nuclear factor-kappa B (NF-κB) nuclear translocation and potentiated the NF-κB-driven transcription activity. Moreover, miR-155 knockdown resulted in a marked increase in the protein amount of myeloid differentiation primary response gene 88 (MyD88), an important adapter protein used by Toll-like receptors to activate the NF-κB pathway. Our data demonstrate that miR-155 serves as a negative feedback regulator in oxLDL-stimulated THP-1 inflammatory responses and lipid uptake and thus might have potential therapeutic implications in atherosclerosis.