Predictive three-biomarker panel in peripheral blood mononuclear cells for detecting hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) ranks among the most prevalent cancers and accounts for a significant proportion of cancer-associated deaths worldwide. This disease, marked by multifaceted etiology, often poses diagnostic challenges. Finding a reliable and non-invasive diagnostic method seems to be necessary. In this study, we analyzed the gene expression profiles of 20 HCC patients, 12 individuals with chronic hepatitis, and 15 healthy controls. Enrichment analysis revealed that platelet aggregation, secretory granule lumen, and G-protein-coupled purinergic nucleotide receptor activity were common biological processes, cellular components, and molecular function in HCC and chronic hepatitis B (CHB) compared to healthy controls, respectively. Furthermore, pathway analysis demonstrated that "estrogen response" was involved in the pathogenesis of HCC and CHB conditions, while, "apoptosis" and "coagulation" pathways were specific for HCC. Employing computational feature selection and logistic regression classification, we identified candidate genes pivotal for diagnostic panel development and evaluated the performance of these panels. Subsequent machine learning evaluations assessed these panels' performance in an independent cohort. Remarkably, a 3-marker panel, comprising RANSE2, TNF-α, and MAP3K7, demonstrated the best performance in qRT-PCR-validated experimental data, achieving 98.4% accuracy and an area under the curve of 1. Our findings highlight this panel's promising potential as a non-invasive approach not only for detecting HCC but also for distinguishing HCC from CHB patients.

Prediction of anti-TNF therapy failure in ulcerative colitis patients by ensemble machine learning: A prospective study.

Nowadays, anti-TNF therapy remarkably improves the medical management of ulcerative colitis (UC), but approximately 40 % of patients do not respond to this treatment. In this study, we used 79 anti-TNF-naive patients with moderate-to-severe UC from four cohorts to discover alternative therapeutic targets and develop a personalized medicine approach that can diagnose UC non-responders (UCN) prior to receiving anti-TNF therapy. To this end, two microarray data series were integrated to create a discovery cohort with 35 UC samples. A comprehensive gene expression and functional analysis was performed and identified 313 significantly altered genes, among which IL6 and INHBA were highlighted as overexpressed genes in the baseline mucosal biopsies of UCN, whose cooperation may lead to a decrease in the Tregs population. Besides, screening the abundances of immune cell subpopulations showed neutrophils' accumulation increasing the inflammation. Furthermore, the correlation of KRAS signaling activation with unresponsiveness to anti-TNF mAb was observed using network analysis. Using 50x repeated 10-fold cross-validation LASSO feature selection and a stack ensemble machine learning algorithm, a five-mRNA prognostic panel including IL13RA2, HCAR3, CSF3, INHBA, and MMP1 was introduced that could predict the response of UC patients to anti-TNF antibodies with an average accuracy of 95.3 %. The predictive capacity of the introduced biomarker panel was also validated in two independent cohorts (44 UC patients). Moreover, we presented a distinct immune cell landscape and gene signature for UCN to anti-TNF drugs and further studies should be considered to make this predictive biomarker panel and therapeutic targets applicable in the clinical setting.

Intestinal organoids: A versatile platform for modeling gastrointestinal diseases and monitoring epigenetic alterations.

Considering the significant limitations of conventional 2D cell cultures and tissue in vitro models, creating intestinal organoids has burgeoned as an ideal option to recapitulate the heterogeneity of the native intestinal epithelium. Intestinal organoids can be developed from either tissue-resident adult stem cells (ADSs) or pluripotent stem cells (PSCs) in both forms induced PSCs and embryonic stem cells. Here, we review current advances in the development of intestinal organoids that have led to a better recapitulation of the complexity, physiology, morphology, function, and microenvironment of the intestine. We discuss current applications of intestinal organoids with an emphasis on disease modeling. In particular, we point out recent studies on SARS-CoV-2 infection in human intestinal organoids. We also discuss the less explored application of intestinal organoids in epigenetics by highlighting the role of epigenetic modifications in intestinal development, homeostasis, and diseases, and subsequently the power of organoids in mirroring the regulatory role of epigenetic mechanisms in these conditions and introducing novel predictive/diagnostic biomarkers. Finally, we propose 3D organoid models to evaluate the effects of novel epigenetic drugs (epi-drugs) on the treatment of GI diseases where epigenetic mechanisms play a key role in disease development and progression, particularly in colorectal cancer treatment and epigenetically acquired drug resistance.

TGF-ß receptor I inhibitor may restrict the induction of EMT in inflamed intestinal epithelial cells.

Despite the extensive body of research, understanding the exact molecular mechanisms governing inflammatory bowel diseases (IBDs) still demands further investigation. Transforming growth factor-ß1 (TGF-ß1) signaling possesses a multifacial effect on a broad range of context-dependent cellular responses. However, long-term TGF-ß1 activity may trigger epithelial-mesenchymal transition (EMT), followed by fibrosis. This study aimed to determine the role of epithelial TGF-ß1 signaling in inflammatory bowel disease (IBD) pathogenesis. The expression of TGF-ß1 signaling components and EMT-related and epithelial tight junction markers was examined in IBD patients (n = 60) as well as LPS-induced Caco-2/RAW264.7 co-culture model using quantitative real-time polymerase chain reaction (qRT-PCR), Western blotting, and immunofluorescence staining. Furthermore, the effect of A83-01, as a TGF-ß receptor I (TßRI) inhibitor, on the inflamed epithelial cells was evaluated in vitro. To evaluate the cytotoxic effects of the TßRI inhibitor, a cell viability assay was performed by the MTS method. Considering the activation of canonical and non-canonical TGF-ß1 signaling pathways in IBD patients, expression results indicated that administering A83-01 in inflamed Caco-2 cells substantially blocked the expression level of TGF-ß1, SMAD4, and PI3K and the phosphorylation of p-SMAD2/3, p-AKT, and p-RPS6 as well as prevented downregulation of LncGAS5 and LncCDKN2B. Further analysis revealed that the inhibition of TGF-ß1 signaling in inflamed epithelial cells by the small molecule could suppress the EMT-related markers as well as improve the expression of epithelial adherens and tight junctions. Collectively, these findings indicated that the inhibition of the TGF-ß1 signaling could suppress the induction of EMT in inflamed epithelial cells as well as exert a protective effect on preserving tight junction integrity. There is a pressing need to determine the exact cellular mechanisms by which TGF-ß1 exerts its effect on IBD pathogenesis.

Effective role of Curcumin on expression regulation of EZH2 histone methyltransferase as a dynamic epigenetic factor in osteogenic differentiation of human mesenchymal stem cells.

BACKGROUND: Efficient differentiation of mesenchymal stem cells (MSCs) into a desired cell lineage remains challenging in cell-based therapy and regenerative medicine. Numerous efforts have been made to efficiently promote differentiation of MSCs into osteoblast lineage. Accordingly, epigenetic signatures emerge as a key conductor of cell differentiation. Among them, Enhancer of Zeste Homolog 2 (EZH2), a histone methyltransferase appears to suppress osteogenesis. Curcumin is an osteoinductive natural polyphenol compound which supposedly modulates epigenetic mechanisms. Hence, the current study aims to address the role of the EZH2 epigenetic factor in osteogenic activity of MSCs after Curcumin treatment. METHODS: The effect of Curcumin on viability and osteogenic differentiation was evaluated at different time points in vitro. The expression level of EZH2 was assessed using quantitative real-time polymerase chain reaction (qRT-PCR) after 14 and 21 days. RESULTS: MTT results showed no cytotoxic effects at concentrations of 10 and 15 µM of Curcumin and cells survived up to 70 % at all time-points. qRT-PCR results demonstrated that Curcumin significantly enhanced the expression levels of osteogenic markers that included Runx2, Osterix, Collagen type I, Osteopontin and Osteocalcin at day 21. CONCLUSIONS: Interestingly, we observed that the expression level of the EZH2 gene was downregulated in the presence of Curcumin compared to the control group during osteogenesis. This study confirmed that Curcumin acts as an epigenetic switch to regulate osteoblast differentiation specifically through the EZH2 suppression.

Carob extract induces spermatogenesis in an infertile mouse model via upregulation of Prm1, Plzf, Bcl-6b, Dazl, Ngn3, Stra8, and Smc1b.

ETHNOPHARMACOLOGICAL RELEVANCE: Ethnopharmacological studies for drug discovery from natural compounds play an important role for developing current therapeutical platforms. Plants are a group of natural sources which have been served as the basis in the treatment of many diseases for centuries. In this regard, Ceratonia siliqua (carob) is one of the herbal medicine which is traditionally used for male infertility treatments. But so far the main mechanisms for effects of carob are unknown. Here, we intend to investigate the ability of carob extract to induce spermatogenesis in an azoospermia mouse model and determine the mechanisms that underlie its function. AIM OF THE STUDY: This is a pre-clinical animal model study to evaluate the effect of carob extract in spermatogenesis recovery. METHODS: We established an infertile mouse model with the intent to examine the ability of carob extract as a potential herbal medicine for restoration of male fertility. Sperm parameters, as well as gene expression dynamics and levels of spermatogenesis hormones, were evaluated 35 days after carob administration. RESULTS: Significant enhanced sperm parameters (P < 0.05) showed that the carob extract could induce spermatogenesis in the infertile mouse model. Our data suggested an anti-apototic and inducer role in the expressions of cell cycle regulating genes. Carob extract improved the spermatogenesis niche by considerable affecting Sertoli and Leydig cells (P < 0.05). The carob-treated mice were fertile and contributed to healthy offspring that matured. Our data confirmed that this extract triggered the hormonal system, the spermatogenesis-related gene expression network, and signaling pathways to induce and promote sperm production with notable level (P < 0.05). We found that the aqueous extract consisted of a polar and mainly well water-soluble substance. Carob extract might upregulate spermatogenesis hormones via its amino acid components, which were detected in the extract by liquid chromatography-mass spectrometry (LC-MS). CONCLUSION: Our results strongly suggest that carob extract might be a promising future treatment option for male infertility. This finding could pave the way for clinical trials in infertile men. This is the first study that has provided reliable, strong pre-clinical evidence for carob extract as an effective candidate for fertility recovery in cancer-related azoospermia.

Inhibition of epithelial SHH signaling exerts a dual protective effect against inflammation and epithelial-mesenchymal transition in inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a debilitating and incurable inflammatory disorder. Despite its increasing prevalence, the underlying pathogenic mechanisms of IBD have not been fully clarified. In addition to the regulatory role of Sonic Hedgehog (SHH) signaling in the maintenance of gut homeostasis, its involvement in development of inflammatory disorders and organ fibrosis has also been reported. Here, we investigated the role of SHH signaling in IBD and examined the molecular mechanisms targeted by the SHH signaling blockade. In addition to increased inflammatory responses and induced Epithelial-mesenchymal transition (EMT) process, SHH signaling activity also increased in active lesions of IBD patients. These findings were similar to what was observed in the LPS-induced Caco2-RAW264.7 co-culture model. Inhibition of SHH signaling in the intestinal epithelial cells using SHH inhibitors influenced inflammatory responses through decreased expression of inflammatory cytokines. Moreover, treatment of differentiated Caco2 cells with SHH signaling inhibitors prevented the overexpression of EMT markers and downregulation of epithelial adherens and tight junctions in inflammatory conditions. This study demonstrated that the inhibition of SHH signaling by small molecules might have therapeutic benefit in IBD, and provided compelling experimental evidence that SHH signaling inhibitors can impose anti-inflammatory effects in intestinal epithelial cells while preserving their epithelial characteristics by restricting the induction of EMT.

Epidrugs: novel epigenetic regulators that open a new window for targeting osteoblast differentiation.

Efficient osteogenic differentiation of mesenchymal stem cells (MSCs) is a critical step in the treatment of bone defects and skeletal disorders, which present challenges for cell-based therapy and regenerative medicine. Thus, it is necessary to understand the regulatory agents involved in osteogenesis. Epigenetic mechanisms are considered to be the primary mediators that regulate gene expression during MSC differentiation. In recent years, epigenetic enzyme inhibitors have been used as epidrugs in cancer therapy. A number of studies mentioned the role of epigenetic inhibitors in the regulation of gene expression patterns related to osteogenic differentiation. This review attempts to provide an overview of the key regulatory agents of osteogenesis: transcription factors, signaling pathways, and, especially, epigenetic mechanisms. In addition, we propose to introduce epigenetic enzyme inhibitors (epidrugs) and their applications as future therapeutic approaches for bone defect regeneration.

The Importance of Stem Cell Senescence in Regenerative Medicine.

Mesenchymal stem cells (MSCs) are an interesting tool in regenerative medicine and a unique cell-based therapy to treat aging-associated diseases. Successful MSC therapy needs a large-scale cell culture, and requires a prolonged in vitro cell culture that subsequently leads to cell senescence. Administration of senescent MSCs results in inefficient cell differentiation in the clinical setting. Therefore, it is of utmost importance to enhance our knowledge about the aging process and methods to detect cell senescence in order to overcome this challenge. Numerous studies have addressed senescence in various aspects. Here, we review the characteristics of MSCs, how aging affects their features, mechanisms involved in aging of MSCs, and potential approaches to detect MSC senescence in vitro.

Contribution of NOTCH signaling pathway along with TNF-α in the intestinal inflammation of ulcerative colitis.

AIM: The aim of this study was to determine gene expression levels of TNF-α, NOTCH1, and HES1 in patients with UC. BACKGROUND: Intestinal inflammation and epithelial injury are the leading actors of inflammatory bowel disease (IBD), causing an excessive expression of pro-inflammatory cytokines such as TNF-α. Also, target genes of NOTCH signaling are involved in the regulation of intestinal homeostasis. Previous studies have demonstrated that TNF-α increases in ulcerative colitis (UC) patients, but the relationship between TNF-α and NOTCH signaling pathway in UC etiopathology needs further study. METHODS: Twelve active UC patients and twelve healthy controls were enrolled in this study. RNA was extracted and the mRNA expression levels of TNF-α, NOTCH1, and HES1 were examined using real-time PCR analyses. Further, transcriptome data deposited in Gene Expression Omnibus (GEO) database were analyzed to detect the differential expression of TNF superfamily and NOTCH1 gene in IBD patients. Finally, the interaction of TNF-α and NOTCH signaling was obtained from The SIGnaling Network Open Resource 2.0 (SIGNOR 2.0) database. RESULTS: The transcription levels of TNF-α, NOTCH1, and HES1 genes were significantly elevated in UC patients compared with control (p < 0.05). In addition, GEO results confirmed our expression results. SIGNOR analysis showed that TNF-α interacts with NOTCH signaling components. CONCLUSION: Based on our data, we observed that NOTCH1 and HES1 in co-operation of TNF-α, may play an important role in pathogenesis of UC. The members of NOTCH signaling pathway can be ideal candidates to target the therapy of IBD.