Crohn’s disease (CD) is frequently complicated by strictures that can be either inflammatory or fibrostenotic. This distinction is important for deciding the best treatment course, but it can be ...difficult to determine clinically, sometimes even by advanced imaging techniques. We performed miRNA PCR panel screening on pooled samples of ileum with CD fibrostenosis or inflammatory stenosis. Eight miRNAs with profibrotic (miR-93-5p, miR-376c-3p and miR-424-5p), or fibroprotective (miR-133a-3p, miR-133b, miR-193a-5p, miR-335-5p and miR-378a-3p) functions described in the literature were selected for validation on 20 samples each of CD with fibrostenosis or inflammatory stenosis, with a separate sampling of the submucosa and subserosa. The results showed significant differences between the groups in subserosal samples, with upregulation of profibrotic miRNAs and downregulation of fibroprotective miRNAs in fibrostenosis compared to inflammatory stenosis. Only miR-424-5p showed a significant difference in the submucosa. There were significant differences in miRNA expression between subserosa and submucosa. Our results provide further evidence that the major differences between fibrostenosis and inflammatory stenosis are located in the subserosa, which is inaccessible to endoscopic sampling, highlighting the need for cross-sectional imaging or serological markers. We identify several miRNAs previously not connected to fibrosis in CD, which could potentially serve as biomarkers of fibrostenosis.
NANOG is an important stem cell transcription factor involved in human development and cancerogenesis. Its expression is complex and regulated on different levels. Moreover, NANOG protein might ...regulate hundreds of target genes at the same time. NANOG is crucial for preimplantation development phase and progressively decreases during embryonic stem cells differentiation, thus regulating embryonic and fetal development. Postnatally, NANOG is undetectable or expressed in very low amounts in the majority of human tissues. NANOG re-expression can be detected during cancerogenesis, already in precancerous lesions, with increasing levels of NANOG in high grade dysplasia. NANOG is believed to enable cancer cells to obtain stem-cell like properties, which are believed to be the source of expanding growth, tumor maintenance, metastasis formation, and tumor relapse. High NANOG expression in cancer is frequently associated with advanced stage, poor differentiation, worse overall survival, and resistance to treatment, and is therefore a promising prognostic and predictive marker. We summarize the current knowledge on the role of NANOG in cancerogenesis and development, including our own experience. We provide a critical overview of NANOG as a prognostic and diagnostic factor, including problems regarding its regulation and detection.
Impact statement
NANOG has emerged as a key stem cell transcription factor in normal development and cancerogenesis. It is generally regarded as a good prognostic and predictive factor in various human cancers. It is less known that it is expressed already at precancerous stages in various organs, suggesting that finally an ideal candidate diagnostic marker has been discovered, enabling to distinguish between true dysplasia and reactive atypia. NANOG regulation is complex, and new insights into our understanding of its regulation might provide important information for future development in a broad field of two entirely different processes, i.e. normal development and cancerogenesis, showing how a physiologic mechanism can be used and abused, transforming itself into a key mechanism of disease development and progression.
Cancer stem cells (CSC) play one of the crucial roles in the pathogenesis of various cancers, including colorectal cancer (CRC). Although great efforts have been made regarding our understanding of ...the cancerogenesis of CRC, CSC involvement in CRC development is still poorly understood. Using bioinformatics and RNA-seq data of normal mucosa, colorectal adenoma, and carcinoma (n = 106) from GEO and TCGA, we identified candidate CSC genes and analyzed pathway enrichment analysis (PEI) and protein–protein interaction analysis (PPI). Identified CSC-related genes were validated using qPCR and tissue samples from 47 patients with adenoma, adenoma with early carcinoma, and carcinoma without and with lymph node metastasis and were compared to normal mucosa. Six CSC-related genes were identified: ANLN, CDK1, ECT2, PDGFD, TNC, and TNXB. ANLN, CDK1, ECT2, and TNC were differentially expressed between adenoma and adenoma with early carcinoma. TNC was differentially expressed in CRC without lymph node metastases whereas ANLN, CDK1, and PDGFD were differentially expressed in CRC with lymph node metastases compared to normal mucosa. ANLN and PDGFD were differentially expressed between carcinoma without and with lymph node metastasis. Our study identified and validated CSC-related genes that might be involved in early stages of CRC development (ANLN, CDK1, ECT2, TNC) and in development of metastasis (ANLN, PDGFD).
Malignant liver tumors include primary malignant liver tumors and liver metastases. They are among the most common malignancies worldwide. The disease has a poor prognosis and poor overall survival, ...especially with liver metastases. Therefore, early detection and differentiation between malignant liver tumors are critical for patient treatment selection. The detection of cancer and the prediction of its origin is possible with a DNA methylation profile of the tumor DNA compared to that of normal cells, which reflects tissue differentiation and malignant transformation. New technologies enable the characterization of the tumor methylome in circulating tumor DNA (ctDNA), providing a variety of new ctDNA methylation biomarkers, which can provide additional information to clinical decision-making. Our review of the literature provides insight into methylation changes in ctDNA from patients with common malignant liver tumors and can serve as a starting point for further research.
Epithelial-mesenchymal transition (EMT) plays a pivotal role in carcinogenesis, influencing cancer progression, metastases, stemness, immune evasion, metabolic reprogramming and therapeutic ...resistance. EMT in most carcinomas, including colorectal carcinoma (CRC), is only partial, and can be evidenced by identification of the underlying molecular drivers and their regulatory molecules. During EMT, cellular reprogramming is orchestrated by core EMT transcription factors (EMT-TFs), namely
,
,
(
) and
(
). While microRNAs have been clearly defined as regulators of EMT, the role of long non-coding RNAs (lncRNAs) in EMT is poorly defined and controversial. Determining the role of lncRNAs in EMT remains a challenge, because they are involved in a number of cellular pathways and are operating through various mechanisms. Adding to the complexity, some lncRNAs have controversial functions across different tumor types, acting as EMT promotors in some tumors and as EMT suppressors in others. The aim of this review is to summarize the role of lncRNAs involved in the regulation of EMT-TFs in human CRC. Additional candidate lncRNAs were identified through a bioinformatics analysis.
Fibrosis is an important complication in inflammatory bowel diseases. Previous studies suggest an important role of matrix Gla protein (MGP) and thrombospondin 2 (THBS2) in fibrosis in various ...organs. Our aim was to analyse their expression together with regulatory miRNAs in submucosal and subserosal fibroblasts in ulcerative colitis (UC) and Crohn’s disease (CD) using immunohistochemistry and qPCR. Digital pathology was used to compare collagen fibre characteristics of submucosal and subserosal fibrosis. Immunohistochemistry showed expression of MGP, but not THBS2 in submucosa in UC and CD. In the subserosa, there was strong staining for both proteins in CD but not in UC. qPCR showed significant upregulation of THBS2 and MGP genes in CD subserosa compared to the submucosa. Digital pathology analysis revealed higher proportion of larger and thicker fibres that were more tortuous and reticulated in subserosal fibrosis compared to submucosal fibrosis. These results suggest distinct fibroblast populations in fibrostenosing CD, and are further supported by image analysis showing significant differences in the morphology and architecture of collagen fibres in submucosal fibrosis in comparison to subserosal fibrosis. Our study is the first to describe differences in submucosal and subserosal fibroblast populations, contributing to understanding of the pathogenesis of fibrostenosis in CD.
Verrucous carcinoma is a low-grade variant of squamous cell carcinoma with specific morphologic, cytokinetic and clinical features. Despite low mitotic activity and slow growth, it can infiltrate ...adjacent tissues in advanced stages but does not metastasize. The most frequently affected site is the oral cavity. The following article provides latest updates in the etiology, clinical presentation, diagnostics and treatment options in oral verrucous carcinoma and discusses the existing dilemmas linked to this unique malignancy.
Oral verrucous carcinoma must be differentiated from conventional squamous cell carcinoma due to its less aggressive behaviour with a more favourable prognosis. Close communication between clinician and pathologist is mandatory for making a correct diagnosis. Primary surgery with negative surgical margins seems to be the most successful treatment. However, management recommendations are not uniform since they are mostly based on case reports and small retrospective case series. Prospective and pooled multi-institutional studies are therefore needed.
MicroRNAs (miRNAs) are noncoding single-stranded RNA molecules that regulate gene expression in physiological functions, development and disease. In recent studies, three miRNAs have been described ...as muscle or cardiac specific: miR-1, miR-133, and miR-208, being involved in heart development and disease; but there are limited data on their role in human myocardial infarction (MI). We therefore analyzed their expression in human MI.
Autopsy samples of infarcted heart tissue from 50 patients with MI, 8 healthy trauma victims and 9 fetuses that died in utero were included. miRNAs miR-1, miR-133a/b and miR-208 were analyzed using quantitative real-time polymerase chain reaction.
miR-208 was upregulated, whereas miR-1 and miR-133a were downregulated in MI compared to healthy adult and fetal hearts. All four tested miRNAs were downregulated in fetal hearts compared to healthy adult hearts.
Our study showed the involvement of muscle- and/or cardiac-specific miRNAs miR-1, miR-133a/b and miR-208 in human MI. The most significant finding was upregulation of miR-208 and downregulation of miR-1 and miR-133a in MI compared to healthy adult hearts. Some patterns of miRNA expression were similar in MI and fetal hearts, supporting the concept of cardiac gene reprogramming in the remodeling of the heart.
Cardiac sarco(endo)plasmic reticulum calcium ATPase-2 (SERCA2) plays one of the central roles in myocardial contractility. Both, SERCA2 mRNA and protein are reduced in myocardial infarction (MI), but ...the correlation has not been always observed. MicroRNAs (miRNAs) act by targeting 3'-UTR mRNA, causing translational repression in physiological and pathological conditions, including cardiovascular diseases. One of the aims of our study was to identify miRNAs that could influence SERCA2 expression in human MI.
The protein SERCA2 was decreased and 43 miRNAs were deregulated in infarcted myocardium compared to corresponding remote myocardium, analyzed by western blot and microRNA microarrays, respectively. All the samples were stored as FFPE tissue and in RNAlater. miRNAs binding prediction to SERCA2 including four prediction algorithms (TargetScan, PicTar, miRanda and mirTarget2) identified 213 putative miRNAs. TAM and miRNApath annotation of deregulated miRNAs identified 18 functional and 21 diseased states related to heart diseases, and association of the half of the deregulated miRNAs to SERCA2. Free-energy of binding and flanking regions (RNA22, RNAfold) was calculated for 10 up-regulated miRNAs from microarray analysis (miR-122, miR-320a/b/c/d, miR-574-3p/-5p, miR-199a, miR-140, and miR-483), and nine miRNAs deregulated from microarray analysis were used for validation with qPCR (miR-21, miR-122, miR-126, miR-1, miR-133, miR-125a/b, and miR-98). Based on qPCR results, the comparison between FFPE and RNAlater stored tissue samples, between Sybr Green and TaqMan approaches, as well as between different reference genes were also performed.
Combing all the results, we identified certain miRNAs as potential regulators of SERCA2; however, further functional studies are needed for verification. Using qPCR, we confirmed deregulation of nine miRNAs in human MI, and show that qPCR normalization strategy is important for the outcome of miRNA expression analysis in human MI.
There is emerging evidence suggesting that epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) play an important role in colorectal carcinoma (CRC), but their exact ...role remains controversial. Our aim was to analyze the miR-200 family as EMT markers and their target genes expression at invasive tumor front and in nodal and liver metastases. Sixty-three formalin-fixed paraffin-embedded tissue samples from 19 patients with CRC were included. Using a micropuncture technique, tissue was obtained from central part and invasive front of the primary tumor, and nodal and liver metastases. Expression of the
miR-200
family and their target genes
CDKN1B, ONECUT2, PTPN13, RND3, SOX2, TGFB2
and
ZEB2
was analyzed using real-time PCR. We found
miR-200
family down-regulation at invasive front compared to central part, and up-regulation of
miRNA-200a/b/c
and
miR-429
in metastases compared to invasive front. At invasive front,
TGFB2
was the only gene with inverse expression to the miR-200 family, whereas in metastases inverse expression was found for
ONECUT2
and
SOX2. CDKN1B, PTPN13
and
ZEB2
were down-regulated at invasive front and up-regulated in metastases. Our results suggest the involvement of partial EMT at invasive tumor front, and partial MET in metastases in CRC, based on miR-200 family and its target genes expression.
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