Cardamonin is a naturally occurring chalcone, majorly from the Zingiberaceae family, which includes a wide range of spices from India. Herein, we investigated the anti-inflammatory property of cardamonin using different in vitro and in vivo systems. In RAW 264.7 cells, treatment with cardamonin showed a reduced nitrous oxide production without affecting the cell viability and decreased the expression of iNOS, TNF-α, and IL-6, and inhibited NF-kB signaling which emphasizes the role of cardamonin as an anti-inflammatory molecule. In a mouse model of dextran sodium sulfate (DSS)-induced colitis, cardamonin treatment protected the mice from colitis. Subsequently, we evaluated the therapeutic potential of this chalcone in a colitis-associated colon cancer model. We performed microRNA profiling in the different groups and observed that cardamonin modulates miRNA expression, thereby inhibiting tumor formation. Together, our findings indicate that cardamonin has the potential to be considered for future therapy against colorectal cancer.

This study is an integrated analysis of the transcriptome profile microRNA (miRNA) and its experimentally validated mRNA targets differentially expressed in the tumorigenic stem-like fraction of oral squamous cell carcinoma (OSCC). We had previously reported the coexistence of multiple drug-resistant tumorigenic fractions, termed side population (SP1, SP2, and MP2), and a nontumorigenic fraction, termed main population (MP1), in oral cancer. These fractions displayed a self-renewal, regenerative potential and expressed known stemness-related cell surface markers despite functional differences. Flow cytometrically sorted pure fractions of SP1 and MP1 cells were subjected to differential expression analysis of both mRNAs and miRNAs. A significant upregulation of genes associated with inflammation, cell survival, cell proliferation, drug transporters, and antiapoptotic pathways, in addition to enhanced transcriptome reprogramming mediated by DNA-histone binding proteins and pattern recognition receptor-mediated signaling, was found to play a crucial role in the transformation of the nontumorigenic MP1 fraction to the tumorigenic SP1 fraction. We also identified several differentially expressed miRNAs that specifically target genes distinctive of tumorigenic SP1 fraction. miRNA-mediated downregulation of stemness-associated markers CD44 and CD147 and upregulation of CD151 may also account for the emergence and persistence of multiple tumorigenic stem cell fractions with varying degrees of malignancy. The phenotypic switch of cancer cells to stem-like OSCC cells mediated by transcriptomal regulation is effectual in addressing biological tumor heterogeneity and subsequent therapeutic resistance leading to a minimal residual disease (MRD) condition in oral cancer. A detailed study of the interplay of miRNAs, mRNA, and the cellular phases involved in the gradual transition of nontumorigenic cancer cells to tumorigenic stem-like cells in solid tumors would enable detection and development of a treatment regimen that targets and successfully eliminates multiple, drug-resistant fractions of cancer cells.

Colorectal cancer is currently the third leading cause of cancer related deaths. There is considerable interest in using dietary intervention strategies to prevent chronic diseases including cancer. Cardamonin is a spice derived nutraceutical and herein, for the first time we evaluated the therapeutic benefits of cardamonin in Azoxymethane (AOM) induced mouse model of colorectal cancer. Mice were divided into 4 groups of which three groups were given six weekly injections of AOM. One group served as untreated control and remaining groups were treated with either vehicle or Cardamonin starting from the same day or 16 weeks after the first AOM injection. Cardamonin treatment inhibited the tumor incidence, tumor multiplicity, Ki-67 and β-catenin positive cells. The activation of NF-kB signaling was also abrogated after cardamonin treatment. To elucidate the mechanism of action a global microRNA profiling of colon samples was performed. Computational analysis revealed that there is a differential expression of miRNAs between these groups. Subsequently, we extend our findings to human colorectal cancer and found that cardamonin inhibited the growth, induces cell cycle arrest and apoptosis in human colorectal cancer cell lines. Taken together, our study provides a better understanding of chemopreventive potential of cardamonin in colorectal cancer.

Next-generation sequencing (NGS) technologies are now well established and have become a routine analysis tool for its depth, coverage, and cost. RNA sequencing (RNA-Seq) has readily replaced the conventional array-based approaches and has become method of choice for qualitative and quantitative analysis of transcriptome, quantification of alternative spliced isoforms, identification of sequence variants, novel transcripts, and gene fusions, among many others. The current chapter discusses the multi-step transcriptome data analysis processes in detail, in the context of re-sequencing (where a reference genome is available). We have discussed the processes including quality control, read alignment, quantification of gene from read level, visualization of data at different levels, and the identification of differentially expressed genes and alternatively spliced transcripts. Considering the data that are freely available to the public, we also discuss The Cancer Genome Atlas (TCGA), as a resource of RNA-Seq data on cancer for selection and analysis in specific contexts of experimentation. This chapter provides insights into the applicability, data availability, tools, and statistics for a beginner to get familiar with RNA-Seq data analysis and TCGA.

Chromatin remodeling factor metastatic tumor protein 1 (MTA1), one of the most upregulated oncogene in human cancer, has an important role in gene expression, cell survival and promoting hypoxic response. Successful cancer progression is dependent on the ability of cells to utilize its survival pathways for adapting to hypoxic microenvironment. Although MTA1 is a stress-responsive gene, but whether hypoxia modulates its function and its role in engaging other core stress-responsive survival pathway(s) remains unknown. Here we have discovered that MTA1 is a novel corepressor of serum and glucocorticoid-inducible kinase 1 (SGK1). Surprisingly, this regulatory corepressive function of MTA1 is lost under hypoxia, allowing upregulation of SGK1 expression and engaging the MTA1–SGK1 axis for the benefit of the cell survival. The underlying mechanism of the noticed stimulation of SGK1 expression by hypoxia includes de-repression of SGK1 transcription because of hypoxia-triggered nucleus-to-cytoplasmic translocation of MTA1. In addition, the newly recognized cytoplasmic translocation of MTA1 was dependent on the chaperoning function of heat shock protein 90 (HSP90) and co-accompanied by the formation of MTA1, HSP90 and HIF1α complex under hypoxic condition but not under normoxic condition. Hypoxia-triggered redistribution of MTA1, SGK1 upregulation and cell survival functions were compromised by a pharmacological SGK1 inhibitor. In summary, for the first time, we report MTA1 regulation of SGK1 expression, hypoxia-dependent MTA1 translocation to the cytoplasm and de-repression of SGK1 transcription. These findings illustrate how cancer cells utilize a chromatin remodeling factor to engage a core survival pathway to support its cancerous phenotypes, and reveal new facets of MTA1–SGK1 axis by a physiologic signal in cancer progression.

Almost all eukaryotic mRNAs have a poly (A) tail at the 3'-end. Canonical PAPs (PAPα/γ) polyadenylate nuclear pre-mRNAs. The recent identification of the non-canonical Star-PAP revealed specificity of nuclear PAPs for pre-mRNAs, yet the mechanism how Star-PAP selects mRNA targets is still elusive. Moreover, how Star-PAP target mRNAs having canonical AAUAAA signal are not regulated by PAPα is unclear. We investigate specificity mechanisms of Star-PAP that selects pre-mRNA targets for polyadenylation. Star-PAP assembles distinct 3'-end processing complex and controls pre-mRNAs independent of PAPα. We identified a Star-PAP recognition nucleotide motif and showed that suboptimal DSE on Star-PAP target pre-mRNA 3'-UTRs inhibit CstF-64 binding, thus preventing PAPα recruitment onto it. Altering 3'-UTR cis-elements on a Star-PAP target pre-mRNA can switch the regulatory PAP from Star-PAP to PAPα. Our results suggest a mechanism of poly (A) site selection that has potential implication on the regulation of alternative polyadenylation.

Plasma level of cyclophilin A is a promising marker of vascular disease in patients with type 2 diabetes. Genetic variants in the peptidylprolyl isomerase A gene, encoding human cyclophilin may alter protein synthesis thus affecting its activity, function, and circulating plasma levels. We examined the effect of single-nucleotide polymorphisms (SNPs) within the PPIA gene on plasma levels of cyclophilin A and coupled this with status of vascular disease in patients with and without type 2 diabetes in 212 South Indian subjects. The regulatory region of PPIA gene was sequenced for SNPs. The association of SNPs with known blood markers of type 2 diabetes and coronary artery disease such as HbA1c, low- and high-density lipoproteins, triglycerides, fasting and postprandial blood sugar levels, and cyclophilin A were probed. We identified three SNPs namely, rs6850: A > G; (AG/−) c.*227_*228delAG and (−/T) c.*318_*319insT. Welchs two-sample t test indicated an association of SNP rs6850: A > G, located at the 5′ UTR region with increased plasma levels of cyclophilin A in patients with coronary artery disease and with coronary artery disease associated with diabetes. The presence of rs6850: A > G variant was significantly associated with coronary artery disease irrespective of whether the patients had diabetes or not. In silico analysis of the sequence using different tools and matrix libraries did not predict any significant differential binding sites for rs6850: A > G, c.*227_*228delAG and c.*318_*319insT. Our results indicate that the SNP rs6850: A > G is associated with increased risk for elevated plasma levels of cyclophilin A and coronary artery disease in patients with and without type 2 diabetes.

Chemokine (C-C motif) receptor 7 (CCR7), a class A subtype G-Protein Coupled Receptor (GPCR), is involved in the migration, activation and survival of multiple cell types including dendritic cells, T cells, eosinophils, B cells, endothelial cells and different cancer cells. Together, CCR7 signaling system has been implicated in diverse biological processes such as lymph node homeostasis, T cell activation, immune tolerance, inflammatory response and cancer metastasis. CCL19 and CCL21, the two well-characterized CCR7 ligands, have been established to be differential in their signaling through CCR7 in multiple cell types. Although the differential ligand signaling through single receptor have been suggested for many receptors including GPCRs, there exists no resource or platform to analyse them globally. Here, first of its kind, we present the cell-type-specific differential signaling network of CCL19/CCL21-CCR7 system for effective visualization and differential analysis of chemokine/GPCR signaling. Database URL: http:// www. netpath. org/ pathways? path_ id= NetPath_ 46.

Motor imagery is the recreation of a motor activity which can be used to design Brain Computer Interfaces (BCI). A BCI bypasses the neuromuscular system and provides a communication link, which directly connects the human brain to an external device. Each individual is able to control his EEG through imaginary motor act it supports to control devices. In this work for designing a BCI system five class motor imagery EEG is used. EEG recorded from the sensory motor cortex is analyzed using wavelet transform. Features extracted from the wavelet coefficients are classified using Support Vector Machine. In this work the motor imagery EEG signal is translated into control signal using a five class BCI to control the directional movement of a wheelchair.

DNA methylation, the highly studied epigenetic mechanism which is involved in the regulatory events of various cellular processes like chromatin structure modifications, chromosomal inactivation, gene expressional patterns, embriyonic developments and transcriptional modification etc. Various high throughput techniques evolved for direct detection of methylation actions as well as information across the entire region. However, despite high throughput technological advances in experimental field, the development of software tools that has been dedicated to the prediction of epigenetic information from specific genome sequences is warranted. To this end we developed a tissue specific classifier MethFinder based on the frequency of novel sequence patterns across nine human tissues that was capable of discriminating methylation prone and methylation resistant CpG islands with an overall accuracy of 93%.

Objectives: Classical diagnostic methods are not sensitive enough in detecting oral lesions that may progress to cancer and in assessing minimal residual disease (MRD) in oral surgical margins. Altered expression of microRNAs (miRNAs) contributes to human cancer, including oral cancer. Although there are many studies on microRNAs in oral cancer, there is no reported study comparing the expression of microRNAs during oral tumor progression and in oral surgical margins.Materials and methods: This study analyzed the expression of 72 miRNAs that were reported (till June 2011) to be differentially expressed in oral cancer, during phases of oral cancer progression and in oral surgical margins.Results: Of the 72 miRNAs analyzed, four (hsa-miR-125a, hsa-miR-184, hsa-miR16 and hsa-miR-96) showed a common pattern of expression in both sets of tissues. We further analyzed the downstream target genes of hsa-miR-16 BCL2 and CCND1. The in silico network analysis of these four microRNAs and their target genes revealed presence of genes involved in tumor progression and transcription factors.Conclusions: The findings suggest that the combinatorial regulation by these miRNAs and their target transcription factors might play a substantial role in oral tumorigenesis. Here we report for the first time that a decreased expression of hsa-miR-125a, hsa-miR-184 and hsa-miR-16 and an increased expression of hsa-miR-96 could be useful in predicting oral tumorigenesis and importantly in the detection of MRD and decision-making process for postoperative treatment modalities. (C) 2013 Elsevier Ltd. All rights reserved.

BACKGROUND:Experimentally validated co-expression correlations between miRNAs and genes are a valuable resource to corroborate observations about miRNA/mRNA changes after experimental perturbations, as well as compare miRNA target predictions with empirical observations. For example, when a given miRNA is transcribed, true targets of that miRNA should tend to have lower expression levels relative to when the miRNA is not expressed.METHODS:We processed publicly available human RNA-seq experiments obtained from NCBI's Sequence Read Archive (SRA) to identify miRNA-mRNA co-expression trends and summarized them in terms of their Pearson's Correlation Coefficient (PCC) and significance.RESULTS:We found that sequence-derived parameters from TargetScan and miRanda were predictive of co-expression, and that TargetScan- and miRanda-derived gene-miRNA pairs tend to have anti-correlated expression patterns in RNA-seq data compared to controls. We provide this data for download and as a web application available at http://wrenlab.org/mirCoX/.CONCLUSION:This database of empirically established miRNA-mRNA transcriptional correlations will help to corroborate experimental observations and could be used to help refine and validate miRNA target predictions.

Myocardial Ischemia is the most common heart disease. Stress ECG has been effectively used for analysis of the myocardial ischemia than normal ECG because of the reason that ischemic conditions will be dominated in stress conditions. According to the clinically proven facts, after taking a series of exercise, the chance of finding ischemia can rise up to 80%-90%. In ECG, the ST segment detection has close relationship with myocardial ischemia and myocardial infarction. Denoising of the stress ECG has done using filters. The key points of ECG signal like Q, R, S are found out using Pan Tompkins algorithm. Other key points like P, T, Ton, Toff, J, Iso-electric point are also found using window method. The feature of interest is ST segment. Based on R-R interval, heart rate was found out. By considering the age of the individual sub-maximal heart rate is fixed and let the patient to do exercise stress test which lasts until sub-maximal heart rate was reached. The ST segment analysis had been done in time domain as well as in frequency domain. ST trend was analyzed on and after the sub-maximal heart rate. According to the clinically proven facts, for a person having ischemia the ST level shows a depression for about two minutes or more during relaxing stage. The signals from MIT-BIH ST Change Database had been used to verify the algorithm in MATLAB software.

Hyperglycemia is widely recognized to be a potent stimulator of monocyte activity, which is a crucial event in the pathogenesis of atherosclerosis. We analyzed the monocyte proteome for potential markers that would enhance the ability to screen for early inflammatory status in Type 2 diabetes mellitus (T2DM), using proteomic technologies. Monocytic cells (THP‐1) were primed with high glucose (HG), their protein profiles were analyzed using 2DE and the downregulated differentially expressed spots were identified using MALDI TOF/MS. We selected five proteins that were secretory in function with the help of bioinformatic programs. A predominantly downregulated protein identified as cyclophilin A (sequence coverage 98%) was further validated by immunoblotting experiments. The cellular mRNA levels of cyclophilin A in various HG‐primed cells were studied using qRT‐PCR assays and it was observed to decrease in a dose‐dependent manner. LC‐ESI‐MS was used to identify this protein in the conditioned media of HG‐primed cells and confirmed by Western blotting as well as ELISA. Cyclophilin A was also detected in the plasma of patients with diabetes. We conclude that cyclophilin A is secreted by monocytes in response to HG. Given the paracrine and autocrine actions of cyclophilin A, the secreted immunophilin could be significant for progression of atherosclerosis in type 2 diabetes. Our study also provides evidence that analysis of monocyte secretome is a viable strategy for identifying candidate plasma markers in diabetes.

IntroductionFor close to a century, researchers have known that Papillomavirus infections in humans cause a variety of benign proliferations including warts, epithelial cysts, intraepithelial neoplasias, oral laryngeal, pharyngeal papillomas and other types of hyperkeratosis.However the molecular mechanisms involved are far from understood.HPV has been d e t e c t e d i n m o r e t h a n 9 0 % o f c e r v i c a l c a ncers and therefore implicated as the main etiological agents in cervical cancer.The pathogenesis of cervical cancer is well-known to involve a multi-step process that includes the transformation of normal cervical epithelium to pre-neoplastic cervical intraepithelial neoplasia that is subsequently transformed to invasive cervical cancer.Although the causal relationship between high-risk human papillomavirus (HPV) infection and cervical cancer has been well-documented in epidemiologic and functional studies, HPV infection alone is not sufficient to induce the malignant transformation of HPV-infected cells.Hence, other unidentified genetic alterations, such as microRNAs the master switches, are required.A class of molecules discovered quite recently, microRNAs (miRNAs), appear to play a significant role in cell proliferation and differentiation, and aberrant miRNAs are associated with several cancers.An new era focusing on micro RNAs, and the studies on HPV and host miRNA interactions will continue shedding more light on understanding of the HPV life cycle and the mechanistic underpinnings of HPV-induced oncogenesis.These small non-coding RNAs can contribute to the repertoire of host pathogen interactions during viral infection.This interplay has important consequences, both for the virus and the host.There has been reported evidence of host-cellular miRNAs modulating expression of various viral genes, thereby playing a pivotal role in the host-pathogen interaction network.In the hide-and-seek game between the pathogens and the infected host, viruses have evolved highly sophisticated gene-silencing mechanisms to evade host-immune response.Recent reports indicate that virus also encode miRNAs that protect them against cellular antiviral response.Furthermore, they may exploit the cellular miRNA pathway to their own advantage.This chapter aims to summarize our current knowledge about miRNA profiles in cervical cancer cell lines and tissues as well as recapitulate recent updates on miRNAinduced gene-silencing mechanism; modulating host-virus interactions of HPV integrated Cervical Carcinomas.

MicroRNAs control gene expression at the posttranscriptional level by base-pairing to the 3′-UTR of their target mRNAs, thus leading to mRNA degradation of protein fabrication. We hypothesize, SNPs within miRNAs and their targets could be of significance to an individual's risk of developing cancer. We analyzed in silico SNP information on cervical cancer associated aberrant alleles and further investigated this in a case-control study by examining eleven SNPs from different genes. It was observed that a C to T polymorphism in putative miRNA target site of BCL2 was significantly conspicuous for the aberrant SNP allele in cancer tissues as compared to controls. This study provides evidence that SNPs in miRNA-binding sites may play an important role in increasing risk of cancer. The results also paves way for future studies to validate these results in other well-characterized populations as well as to explore the biological significance of these particular SNPs.