A Pilot Study of Circulating miRNAs as Potential Biomarkers of Early Stage Breast Cancer

To date, there are no highly sensitive and specific minimally invasive biomarkers for detection of breast cancer at an early stage. The occurrence of circulating microRNAs (miRNAs) in blood components (including serum and plasma) has been repeatedly observed in cancer patients as well as healthy controls. Because of the significance of miRNA in carcinogenesis, circulating miRNAs in blood may be unique biomarkers for early and minimally invasive diagnosis of human cancers. The objective of this pilot study was to discover a panel of circulating miRNAs as potential novel breast cancer biomarkers.Using microarray-based expression profiling followed by Real-Time quantitative Polymerase Cycle Reaction (RT-qPCR) validation, we compared the levels of circulating miRNAs in plasma samples from 20 women with early stage breast cancer (10 Caucasian American (CA) and 10 African American (AA)) and 20 matched healthy controls (10 CAs and 10 AAs). Using the significance level of p<0.05 constrained by at least two-fold expression change as selection criteria, we found that 31 miRNAs were differentially expressed in CA study subjects (17 up and 14 down) and 18 miRNAs were differentially expressed in AA study subjects (9 up and 9 down). Interestingly, only 2 differentially expressed miRNAs overlapped between CA and AA study subjects. Using receiver operational curve (ROC) analysis, we show that not only up-regulated but also down-regulated miRNAs can discriminate patients with breast cancer from healthy controls with reasonable sensitivity and specificity. To further explore the potential roles of these circulating miRNAs in breast carcinogenesis, we applied pathway-based bioinformatics exploratory analysis and predicted a number of significantly enriched pathways which are predicted to be regulated by these circulating miRNAs, most of which are involved in critical cell functions, cancer development and progression.Our observations from this pilot study suggest that the altered levels of circulating miRNAs might have great potential to serve as novel, noninvasive biomarkers for early detection of breast cancer

The JNK Inhibitor XG-102 Protects against TNBS-Induced Colitis

The c-Jun N-terminal kinase (JNK)-inhibiting peptide D-JNKI-1, syn. XG-102 was tested for its therapeutic potential in acute inflammatory bowel disease (IBD) in mice. Rectal instillation of the chemical irritant trinitrobenzene sulfonic acid (TNBS) provoked a dramatic acute inflammation in the colon of 7–9 weeks old mice. Coincident subcutaneous application of 100 µg/kg XG-102 significantly reduced the loss of body weight, rectal bleeding and diarrhoea. After 72 h, the end of the study, the colon was removed and immuno-histochemically analysed. XG-102 significantly reduced (i) pathological changes such as ulceration or crypt deformation, (ii) immune cell pathology such as infiltration and presence of CD3- and CD68-positive cells, (iii) the production of tumor necrosis factor (TNF)-α in colon tissue cultures from TNBS-treated mice, (iv) expression of Bim, Bax, FasL, p53, and activation of caspase 3, (v) complexation of JNK2 and Bim, and (vi) expression and activation of the JNK substrate and transcription factor c-Jun. A single application of subcutaneous XG-102 was at least as effective or even better depending on the outcome parameter as the daily oral application of sulfasalazine used for treatment of IBD

Serum MicroRNA Expression Profile Distinguishes Enterovirus 71 and Coxsackievirus 16 Infections in Patients with Hand-Foot-and-Mouth Disease

Altered circulating microRNA (miRNA) profiles have been noted in patients with microbial infections. We compared host serum miRNA levels in patients with hand-foot-and-mouth disease (HFMD) caused by enterovirus 71 (EV71) and coxsackievirus 16 (CVA16) as well as in other microbial infections and in healthy individuals. Among 664 different miRNAs analyzed using a miRNA array, 102 were up-regulated and 26 were down-regulated in sera of patients with enteroviral infections. Expression levels of ten candidate miRNAs were further evaluated by quantitative real-time PCR assays. A receiver operating characteristic (ROC) curve analysis revealed that six miRNAs (miR-148a, miR-143, miR-324-3p, miR-628-3p, miR-140-5p, and miR-362-3p) were able to discriminate patients with enterovirus infections from healthy controls with area under curve (AUC) values ranged from 0.828 to 0.934. The combined six miRNA using multiple logistic regression analysis provided not only a sensitivity of 97.1% and a specificity of 92.7% but also a unique profile that differentiated enterovirial infections from other microbial infections. Expression levels of five miRNAs (miR-148a, miR-143, miR-324-3p, miR-545, and miR-140-5p) were significantly increased in patients with CVA16 versus those with EV71 (p<0.05). Combination of miR-545, miR-324-3p, and miR-143 possessed a moderate ability to discrimination between CVA16 and EV71 with an AUC value of 0.761. These data indicate that sera from patients with different subtypes of enteroviral infection express unique miRNA profiles. Serum miRNA expression profiles may provide supplemental biomarkers for diagnosing and subtyping enteroviral HFMD infections

Serological Markers for Inflammatory Bowel Disease in AIDS Patients with Evidence of Microbial Translocation

Background: Breakdown of the gut mucosal barrier during chronic HIV infection allows translocation of bacterial products such as lipopolysaccharides (LPS) from the gut into the circulation. Microbial translocation also occurs in inflammatory bowel disease (IBD). IBD serological markers are useful in the diagnosis of IBD and to differentiate between Crohn's disease (CD) and ulcerative colitis (UC). Here, we evaluate detection of IBD serological markers in HIV-infected patients with advanced disease and their relationship to HIV disease markers.Methods IBD serological markers (ASCA, pANCA, anti-OmpC, and anti-CBir1) were measured by ELISA in plasma from AIDS patients (n = 26) with low CD4 counts (<300 cells/$\mu$l) and high plasma LPS levels, and results correlated with clinical data. For meta-analysis, relevant data were abstracted from 20 articles. Results: IBD serological markers were detected in approximately 65% of AIDS patients with evidence of microbial translocation. An antibody pattern consistent with IBD was detected in 46%; of these, 75% had a CD-like pattern. Meta-analysis of data from 20 published studies on IBD serological markers in CD, UC, and non-IBD control subjects indicated that IBD serological markers are detected more frequently in AIDS patients than in non-IBD disease controls and healthy controls, but less frequently than in CD patients. There was no association between IBD serological markers and HIV disease markers (plasma viral load and CD4 counts) in the study cohort. Conclusions: IBD serological markers may provide a non-invasive approach to monitor HIV-related inflammatory gut disease. Further studies to investigate their clinical significance in HIV-infected individuals are warranted

Endogenous antigen presenting cell-derived IL-10 inhibits T lymphocyte responses to commensal enteric bacteria

Interleukin-10 deficient (IL-10-/-) mice develop chronic T cell-mediated colitis when colonized with normal commensal bacteria, but germ-free (GF) IL-10-/- mice remain disease-free. Antigen presenting cells (APC) secrete regulatory cytokines that help determine T lymphocyte activation or tolerance. CD4+ T cells from the mesenteric lymph nodes of inflamed IL-10-/- mice secrete more IFNγ and IL-17 when cultured with cecal bacterial lysate-pulsed splenic APC from IL-10-/- mice than when cultured with normal control APC. GF IL-10-/- APC induce similar IFNγ and IL-17 responses; therefore, the functional difference between normal and IL-10 deficient APC is inherent to the lack of IL-10 and not secondary to inflammation. Bacterial lysate-pulsed normal APC cultured with CD4+ cells from colitic IL-10-/- mice or with exogenous IFNγ secrete higher amounts of IL-10 compared to the same APC cultured with naïve T cells. APC enriched for CD11c+ cells are potent activators of IFNγ and IL-17 production by CD4+ cells from IL-10-/- mice. These APC also produce IL-12/IL-23 p40 and IL-10. Recombinant IL-10 suppressed and anti-IL-10 receptor antibody increased IFNγ, IL-17 and IL-12/IL-23 p40 production in bacterial lysate-pulsed APC and plus CD4+ T cell co-cultures. Taken together, our results show that endogenous IL-10 produced by APC inhibits responses to commensal bacteria and influences the ability of APC to stimulate IFNγ-producing effector lymphocytes, which reciprocally, induce IL-10 production by APC. Cytokines produced by APC are an important determinant of pathogenic versus protective mucosal immune responses to colonic bacterial stimulation

Inflammatory bowel disease: past, present, and future

Crohn’s disease and ulcerative colitis, collectively known as the inflammatory bowel diseases (IBD), are largely diseases of the twentieth century, and are associated with the rise of modern, Westernized industrial society. Although the causes of these diseases remain incompletely understood, the prevailing model is that the intestinal flora drives an unmitigated intestinal immune response and inflammation in the genetically susceptible host. A review of the past and present of these diseases shows that detailed description preceded more fundamental elucidation of the disease processes. Working out the details of disease pathogenesis, in turn, has yielded dividends in more focused and effective therapy for IBD. This article highlights the key descriptions of the past, and the pivotal findings of current studies in disease pathogenesis and its connection to medical therapy. Future directions in the IBD will likely explicate the inhomogeneous causes of these diseases, with implications for individualized therapy

Selective Release of MicroRNA Species from Normal and Malignant Mammary Epithelial Cells

MicroRNAs (miRNAs) in body fluids are candidate diagnostics for a variety of conditions and diseases, including breast cancer. One premise for using extracellular miRNAs to diagnose disease is the notion that the abundance of the miRNAs in body fluids reflects their abundance in the abnormal cells causing the disease. As a result, the search for such diagnostics in body fluids has focused on miRNAs that are abundant in the cells of origin. Here we report that released miRNAs do not necessarily reflect the abundance of miRNA in the cell of origin. We find that release of miRNAs from cells into blood, milk and ductal fluids is selective and that the selection of released miRNAs may correlate with malignancy. In particular, the bulk of miR-451 and miR-1246 produced by malignant mammary epithelial cells was released, but the majority of these miRNAs produced by non-malignant mammary epithelial cells was retained. Our findings suggest the existence of a cellular selection mechanism for miRNA release and indicate that the extracellular and cellular miRNA profiles differ. This selective release of miRNAs is an important consideration for the identification of circulating miRNAs as biomarkers of disease

MicroRNA-mediated drug resistance in breast cancer

Chemoresistance is one of the major hurdles to overcome for the successful treatment of breast cancer. At present, there are several mechanisms proposed to explain drug resistance to chemotherapeutic agents, including decreased intracellular drug concentrations, mediated by drug transporters and metabolic enzymes; impaired cellular responses that affect cell cycle arrest, apoptosis, and DNA repair; the induction of signaling pathways that promote the progression of cancer cell populations; perturbations in DNA methylation and histone modifications; and alterations in the availability of drug targets. Both genetic and epigenetic theories have been put forward to explain the mechanisms of drug resistance. Recently, a small non-coding class of RNAs, known as microRNAs, has been identified as master regulators of key genes implicated in mechanisms of chemoresistance. This article reviews the role of microRNAs in regulating chemoresistance and highlights potential therapeutic targets for reversing miRNA-mediated drug resistance. In the future, microRNA-based treatments, in combination with traditional chemotherapy, may be a new strategy for the clinical management of drug-resistant breast cancers