Gene heterogeneity between CAFs and NFs.

<p>A. HeatMap of gene expression data obtained by Agilent micro-arrays analysis from CAF and NF isolated from all six cases (BCa 1, 2, 3, 4, 5, 6) were subjected to unsupervised cluster analysis. B. The mRNA levels of 9 genes selected randomly were analyzed by qRT-PCR in CAF and NF from 3 patients with breast cancer. The data were shown as fold change in CAF vs. NF. C. Signaling pathway analysis of enriched processes and signaling pathway in CAFs vs. NFs.</p

The potential interaction of CAFs and tumor cells on cell invasion.

<p>A, C. MDA-MB-231 invasion ability was tested by Transwell assay cultured in the medium with FBS or FBS free (A), and migrated cells were shown by histogram (C). B, D. The migrated MDA-MB-231 cells were checked by Transwell assay (B) in the co-culture system using condition medium derived from CAFs and NFs. And (D) the migrated cells shown by histogram. E, F. The invasive potential of CAFs and NFs were determined by Transwell assay (E) in the co-culture system using condition medium derived from MDA-MB-231 cells. The histogram to show the average migrated cells each view (F). Data is representative of 5 views (**<i>P</i><0.01), (magnification: A, B 100×; E 200×).</p

Biological Characteristics and Genetic Heterogeneity between Carcinoma-Associated Fibroblasts and Their Paired Normal Fibroblasts in Human Breast Cancer

<div><p>Background</p><p>The extensional signals in cross-talk between stromal cells and tumor cells generated from extracellular matrix molecules, soluble factor, and cell-cell adhesion complexes cooperate at the extra- and intracellular level in the tumor microenvironment. CAFs are the primary type of stromal cells in the tumor microenvironment and play a pivotal role in tumorigenesis and development. Hitherto, there is hardly any systematic analysis of the intrinsic relationship between CAFs function and its abnormal signaling pathway. The extreme complexity of CAFs’ features and their role in tumor development are needed to be further investigated.</p> <p>Methodology/Principal Findings</p><p>We primary cultured CAFs and NFs from early stages of breast cancer tissue and identified them using their biomarker by immunohistochemistry for Fibronectin, α-SMA and FAP. Microarray was applied to analyze gene expression profiles of human breast CAFs and the paired NFs. The Up-regulated genes classified by Gene Ontology, signal pathways enriched by DAVID pathway analysis. Abnormal signaling pathways in breast cancer CAFs are involved in cell cycle, cell adhesion, signal transduction and protein transport being reported in CAFs derived from other tumors. Significantly, the altered ATM signaling pathway, a set of cell cycle regulated signaling, and immune associated signaling are identified to be changed in CAFs.</p> <p>Conclusions/Significance</p><p>CAFs have the vigorous ability of proliferation and potential of invasion and migration comparing with NFs. CAFs could promote breast cancer cell invasion under co-culture conditions through up-regulated CCL18 and CXCL12. Consistently with its biologic behavior, the gene expression profiling analyzed by microarray shows that some of key signaling pathways, such as cell cycle, cell adhesion, and secreting factors play an important role in CAFs. The altered ATM signaling pathway is abnormally active in the early stage of breast cancer. The set of immune associated signaling may be involved in tumor cell immune evasion.</p> </div

Characterization of fibroblasts isolated from human breast tissue samples.

<p>A. Representative cell morphology of CAFs and NFs. B. Identification of CAFs using fibroblast biomarker fibronectin, and CAFs specific biomarker α-SMA and FAP by immunofluorescence staining. C. The biomarker gene expression in CAFs and NFs was re-proved by qTR-PCR, and its relative fold change in CAFs and NFs was displayed. The CAFs and NFs are positive for fibroblast biomarker fibronectin, CAFs specific biomarker α-SMA and FAP are high expression in CAFs (magnification 100× (cell morphology); 200× (Biomarker immunofluorescence staining).</p

The migration and invasion of CAFs compared with NFs.

<p>A. Cell migration capability was determined by wound healing assay. B. The distance of wound closure (compared with control at 0 h) was measured in four-independent wound sites each group after 24 h. C. The invasion capability of CAFs and NFs was determined by Transwell assay. D. The migrated cells of CAFs and NFs shown by a histogram. Data were shown as mean±SD of 5 repeats. (**<i>P</i><0.01), (magnification A. 100×; C. 200×).</p

Significant difference of proliferation between CAFs and NFs.

<p>A. Cell proliferation determined by MTT assay for three of CAFs and NFs. B. Representative DNA content of CAFs and NFs were tested by flow cytometry. C. The percentages of cells in each of cell cycle phases shown by histogram for CAFs vs. NFs. The data were shown as mean±SD for N≥3 separate experiments (**<i>P</i><0.01).</p

The common cytokines-CCL18 was obtained by compared with published microarray data.

<p>The number of dysregulated difference genes and shared genes from current work and two other studies in breast cancer associated CAF versus paired NF was displayed in each group. Dysregulated Cytokines in each study were listed. Only the common cytokines-CCL18 was obtained compared local data with Basik’s microarray data (GSE29270).</p

Potential Role of microRNA-21 in the Diagnosis of Gastric Cancer: A Meta-Analysis

<div><p>Introduction</p><p>Accumulating evidences indicate that microRNA-21(miR-21) show significant high concentration in plasma of gastric cancer (GC) patients compared to normal individuals, suggesting that it may be a useful novel diagnostic biomarker for gastric cancer. Therefore, we aimed to assess the potential diagnostic value of miR-21 for gastric cancer in this study.</p><p>Methods</p><p>Literature database including PubMed, Embase, the Cochrane Library, Web of Science, Ovid, SciVerse, Science Direct, Scopus, BioMed Central, Biosis previews,Chinese Biomedical Literature Database (CBM), Chinese National Knowledge Infrastructure (CNKI), Technology of Chongqing (VIP), and Wan Fang DATA were searched for publications concerning the diagnostic value of miR-21 for GC without language restriction. The quality of each study was scored with the Quality Assessment of Diagnostic Accuracy Studies (QUADAS). Then, data were retrieved from any qualified article hits and subject to meta-analysis. Receiver operating characteristic curves (ROC) were used to check the overall test performance. Evidence of heterogeneity was evaluated using the Chi-square and <i>I</i>² test.</p><p>Results</p><p>Five studies with a total 251 GC patients and 184 control individuals were included in this meta-analysis. All of the included studies are of high quality (QUADAS score$13). The summary estimates revealed that the pooled sensitivity is 66.5% (95% confidence interval (CI): 55.0%–76.3%) and the specificity is 83.1% (95% CI: 69.4%–91.5%). In addition, the area under the summary ROC curve (AUC) is 0.80.</p><p>Conclusion</p><p>The current evidence suggests that miR-21 has potential diagnostic value with a moderate sensitivity and specificity for GC. More prospective studies on the diagnostic value of miR-21 for GC are needed in the future.</p></div

CAFs promote proliferation of MDA-MB-231 cells in co-culture system compared with NFs.

<p>A. Cell growth curve determined by cell count. B. Representative DNA content of MDA-MB-231 cultured with normal medium and with conditioned medium derived from CAFs and NFs determined by flow cytometry. C. The percentages of cells in each of cell cycle phases shown by histogram for MDA-MB-231. The data were shown as mean±SD for 3 separate experiments (*<i>P</i><0.05; **<i>P</i><0.01).</p

Summary of studies using miR-21 as a biomarker of GC and study quality assessment.

<p>Note: *Calculated from independent patient data (IPD) QUADAS = quality assessment for studies of diagnostic accuracy. AUC = Area under the curve of a receiver operator curve. NR = not report. Se = sensitivity. Sp = specificity.</p