Ginseng mediates its anticancer activity by inhibiting the expression of DNMTs and reactivating methylation-silenced genes in colorectal cancer.

Developing safe and effective therapeutic modalities remains a critical challenge for improving the prognosis of patients with colorectal cancer (CRC). In this regard, targeting epigenetic regulation in cancers has recently emerged as a promising therapeutic approach. Since several natural compounds have recently been shown to be important epigenetic modulators, we hypothesized that Ginseng might exert its anticancer activity by regulating DNA methylation alterations in CRC. In this study, a series of cell culture studies were conducted, followed by their interrogation in patient-derived 3D organoid models to evaluate Ginseng's anticancer activity in CRC. Genome-wide methylation alterations were interrogated by undertaking MethylationEpic BeadChip microarrays. First, 50% inhibitory concentrations (IC50) were determined by cell viability assays, and subsequent Ginseng treatment demonstrated a significant anticancer effect on clonogenicity and cellular migration in CRC cells. Treatment with Ginseng potentiated cellular apoptosis through regulation of apoptosis-related genes in CRC cells. Furthermore, Ginseng treatment downregulated the expression of DNA methyltransferases (DNMTs) and decreased the global DNA methylation levels in CRC cells. The genome-wide methylation profiling identified Ginseng-induced hypomethylation of transcriptionally silenced tumor suppressor genes. Finally, cell culture-based findings were successfully validated in patient-derived 3D organoids. In conclusion, we demonstrate that Ginseng exerts its antitumorigenic potential by regulating cellular apoptosis via the downregulation of DNMTs and reversing the methylation status of transcriptionally silenced genes in CRC.

The Nonproteolytic Intracellular Domain of Membrane-Type 1 Matrix Metalloproteinase Coordinately Modulates Abdominal Aortic Aneurysm and Atherosclerosis in Mice-Brief Report.

BACKGROUND: MT1-MMP (membrane-type 1 matrix metalloproteinase, MMP-14) is a transmembrane-anchored protein with an extracellular proteinase domain and a cytoplasmic tail devoid of proteolytic functions but capable of mediating intracellular signaling that regulates tissue homeostasis. MT1-MMP extracellular proteolytic activity has been shown to regulate pathological remodeling in aortic aneurysm and atherosclerosis. However, the role of the nonproteolytic intracellular domain of MT1-MMP in vascular remodeling in abdominal aortic aneurysms (AAA) is unknown. METHODS: We generated a mutant mouse that harbors a point mutation (Y573D) in the MT1-MMP cytoplasmic domain that abrogates the MT1-MMP signaling function without affecting its proteolytic activity. These mice and their control wild-type littermates were subjected to experimental AAA modeled by angiotensin II infusion combined with PCSK9 (proprotein convertase subtilisin/kexin type 9) overexpression and high-cholesterol feeding. RESULTS: The mutant mice developed more severe AAA than the control mice, with concomitant generation of intraaneurysmal atherosclerotic lesions and dramatically increased macrophage infiltration and elastin degradation. Aortic lesion-associated and bone marrow-derived macrophages from the mutant mice exhibited an enhanced inflammatory state and expressed elevated levels of proinflammatory Netrin-1, a protein previously demonstrated to promote both atherosclerosis and AAA. CONCLUSIONS: Our findings show that the cytoplasmic domain of MT1-MMP safeguards from AAA and atherosclerotic plaque development through a proteolysis-independent signaling mechanism associated with Netrin-1 expression. This unexpected function of MT1-MMP unveils a novel mechanism of synchronous onset of AAA and atherogenesis and highlights its importance in the control of vascular wall homeostasis.

A blood-based circulating microbial metagenomic panel for early diagnosis and prognosis of oesophageal adenocarcinoma.

BACKGROUND: Emerging evidence indicates the potential clinical significance of specific microbial signatures as diagnostic and prognostic biomarkers, in multiple cancers. However, to date, no studies have systematically interrogated circulating metagenome profiling in oesophageal adenocarcinoma (EAC) patients, particularly as novel non-invasive, early detection, surveillance and prognostic classifiers. METHODS: Metagenome sequencing was performed on 81 serum specimens collected across EAC spectrum, with sequencing reads classified using Bracken and MetaPhlAn3. Followed by the Linear Discriminant Analysis effect size (LEfSe) method to identify microbial profiles between groups. Logistic regression and Kaplan-Meier analyses were used to build classifiers. RESULTS: A significant loss of alpha and beta diversity was identified in serum specimens from EAC patients. We observed a shift in microbial taxa between each group-at the phylum, genus, and species level-with Lactobacillus sakei as the most prominent species in gastroesophageal reflux (GERD) vs other patient groups. Interestingly, LEfSe analysis identified a complete loss of Lactobacillus (L. Sakei and L. Curvatus), Collinsella stercoris and Bacteroides stercoris but conversely a significant increase in Escherichia coli in patients with EAC. Finally, we developed a metagenome panel that discriminated EAC from GERD patients with an AUC value of 0.89 (95% CI: 0.78-0.95; P < 0.001) and this panel in conjunction with the TNM stage was a robust predictor of overall survival (≥24 months; AUC = 0.84 (95% CI: 0.66-0.92; P = 0.006)). CONCLUSION: This study firstly describes unique blood-based microbial profiles in patients across EAC carcinogenesis, that are further utilised to establish a novel circulating diagnostic and prognostic metagenomic signature for EAC. TRANSLATIONAL RELEVANCE: Accumulating data indicates the clinical relevance of specific microbial signatures as diagnostic and prognostic biomarkers, in multiple cancers. However, to date, no studies have systematically interrogated circulating metagenome profiling in patients with oesophageal adenocarcinoma (EAC). Herein, we performed metagenome sequencing in serum specimens from EAC patients 81 collected across EAC spectrum and observed a significant loss of alpha and beta diversity, with a shift in microbial taxa between each group-at the phylum, genus, and species level-with Lactobacillus sakei as the most prominent species in gastroesophageal reflux (GERD) vs other patient groups. Interestingly, LEfSe analysis identified a complete loss of Lactobacillus (L. Sakei and L. Curvatus), Collinsella stercoris and Bacteroides stercoris but conversely a significant increase in Escherichia coli in patients with EAC. Finally, we developed a metagenome panel that discriminated EAC from GERD patients with an AUC value of 0.89 and this panel, in conjunction with the TNM stage, was a robust predictor of overall survival. This study for the first time describes unique blood-based microbial profiles in patients across EAC carcinogenesis, that are further utilised to establish a novel circulating diagnostic and prognostic metagenomic signature for EAC.

Mapping the unicellular transcriptome of the ascending thoracic aorta to changes in mechanosensing and mechanoadaptation during aging.

Aortic stiffening is an inevitable manifestation of chronological aging, yet the mechano-molecular programs that orchestrate region- and layer-specific adaptations along the length and through the wall of the aorta are incompletely defined. Here, we show that the decline in passive cyclic distensibility is more pronounced in the ascending thoracic aorta (ATA) compared to distal segments of the aorta and that collagen content increases in both the medial and adventitial compartments of the ATA during aging. The single-cell RNA sequencing of aged ATA tissues reveals altered cellular senescence, remodeling, and inflammatory responses accompanied by enrichment of T-lymphocytes and rarefaction of vascular smooth muscle cells, compared to young samples. T lymphocyte clusters accumulate in the adventitia, while the activation of mechanosensitive Piezo-1 enhances vasoconstriction and contributes to the overall functional decline of ATA tissues. These results portray the immuno-mechanical aging of the ATA as a process that culminates in a stiffer conduit permissive to the accrual of multi-gerogenic signals priming to disease development.

Sequence analysis of SARS-CoV-2 Delta variant isolated from Makassar, South Sulawesi, Indonesia.

Introduction: This study aimed to perform mutation and phylogenetic analyses of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Delta variants and analyze the characteristic signs and symptoms of patients infected with SARS-CoV-2 Delta variant originated from Makassar during the Delta outbreak.Methods: We collected samples from patients who were infected with coronavirus disease 2019 (COVID-19) between June and October 2021. We selected the Quantitative Reverse Transcription-Polymerase Chain Reaction (PCR)-positive samples with a cycle threshold value of <30 for whole genome sequencing. Total viral ribonucleic acid (RNA) was isolated from 34 PCR-positive nasopharyngeal swab samples, and whole genome sequencing was performed using the Oxford Nanopore GridlON sequencer. Phylogenetic and maximum clade credibility analyses were performed using the Bayesian Markov chain Monte Carlo method. Results: It was found that 33 patients were infected with the SARS-CoV-2 Delta variant in this cohort study, among whom 63.6% (21) patients were female. According to the clinical data, 24 (72.7%), 7 (21.2%), and 2 (6.1%) patients had mild, moderate, and severe COVID-19 infections. Phylogenetic analysis based on the spike and RNA-dependent RNA polymerase (RdRp) genes showed that the collected samples were clustered in the main lineage of B.1.617.2 (Delta variant). The Delta variants had a high frequency of distinct mutations in the spike protein region, including T19R (94.12%), L452R (88.23%), T478K (91.17%), D614G (97%), P681R (97%), and D950 N (97%). Other unique mutations found in a smaller frequency in our samples were present in the N-terminal domain, including A27T (2.94%) and A222V (14.70%), and in the receptor-binding domain, including Q414K (5.88%), G446V (2.94%), and T470 N (2.94%). Conclusion: This study revealed the unique mutations in the S protein region of Delta variants. T19R, L452R, T478K/T478R, D614G, P681R, and D950 N were the most common substitutions in Makassar's Delta variant.

Multiplexed shRNA-miRs as a candidate for anti HIV-1 therapy: strategies, challenges, and future potential.

The spread of HIV is on the rise and has become a global issue, especially for underdeveloped and developing countries. This is due to the fact that HIV majorly occurs asymptomatically and is implausible for early diagnosis. Recent advances in research and science have enabled the investigation of a new potential treatment involving gene-based therapy, known as RNA interference (RNAi) that will direct gene silencing and further compensate for natural variants and viral mutants. Several types of small regulatory RNA are discussed in this present study, including microRNA (miRNA), small interfering RNA (siRNA), and short hairpin RNA (shRNA).This paper examines the mechanism of RNAi as a viable HIV therapy, using a minimum of four shRNAs to target both dispensable host components (CCR5) and viral genes (Gag, Env, Tat, Pol I, Pol II and Vif). Moreover, a multiplexed mechanism of shRNAs and miRNA is known to be effective in preventing viral escape due to mutation as the miRNA develops a general polycistronic platform for the expression of a large amount of shRNA-miRs. Several administration methods as well as the advantages of this RNAi treatment are also discussed in this study. The administration methods include (1) ex vivo delivery with the help of viral vectors, nanoparticles, and electroporation, (2) nonspecific in vivo delivery using non-viral carriers including liposomes, dendrimers and aptamers, as well as (3) targeted delivery that uses antibodies, modified nanoparticles, nucleic acid aptamers, and tissue-specific serotypes of AAV. Moreover, the advantages of this treatment are related to the effectiveness in silencing the HIV gene, which is more compatible compared to other gene therapy treatments, such as ZFN, TALEN, and CRISPR/Cas9.

Biomarkers for the Detection and Management of Hepatocellular Carcinoma in Patients Treated with Direct-Acting Antivirals.

Hepatocellular carcinoma (HCC) is the sixth-most common type of cancer worldwide and chronic Hepatitis C virus (HCV) represents the main etiological factor in developed countries. HCV promotes hepatocarcinogenesis through persistent liver inflammation and dysregulation of cell signaling pathways. The introduction of direct-acting antivirals (DAAs) resulted in a significant improvement in the eradication of the virus, with an expected reduction of HCC incidence. However, the risk of HCC development can persist after DAA treatment. Recent studies have investigated the potential use of molecular biomarkers that predict HCC occurrence or recurrence helping the stratification of patients under surveillance. This review aimed to summarize all pre-clinical exploration of predictive biomarkers to identify DAA-treated patients at risk for HCC development. Dysregulated microRNAs, lncRNAs, histone modifications, cytokines, proteins, and sphingolipids represent various classes of HCC risk predictors identified in two different biological sources (tissue and serum). The non-invasive serum markers can provide a more accessible means to perform clinical monitoring and predict the risk of HCC. In addition, conditions like cirrhosis, predisposing to HCC, strongly correlate with most of the molecular predictors identified, supporting the value of these molecules as possible biomarkers of HCC in DAA-treated patients.

Mechanosignals in abdominal aortic aneurysms.

Cumulative evidence has shown that mechanical and frictional forces exert distinct effects in the multi-cellular aortic layers and play a significant role in the development of abdominal aortic aneurysms (AAA). These mechanical cues collectively trigger signaling cascades relying on mechanosensory cellular hubs that regulate vascular remodeling programs leading to the exaggerated degradation of the extracellular matrix (ECM), culminating in lethal aortic rupture. In this review, we provide an update and summarize the current understanding of the mechanotransduction networks in different cell types during AAA development. We focus on different mechanosensors and stressors that accumulate in the AAA sac and the mechanotransduction cascades that contribute to inflammation, oxidative stress, remodeling, and ECM degradation. We provide perspectives on manipulating this mechano-machinery as a new direction for future research in AAA.

Linking single nucleotide polymorphisms to signaling blueprints in abdominal aortic aneurysms.

Abdominal aortic aneurysms (AAA) is a multifactorial complex disease with life-threatening consequences. While Genome-wide association studies (GWAS) have revealed several single nucleotide polymorphisms (SNPs) located in the genome of individuals with AAA, the link between SNPs with the associated pathological signals, the influence of risk factors on their distribution and their combined analysis is not fully understood. We integrated 86 AAA SNPs from GWAS and clinical cohorts from the literature to determine their phenotypical vulnerabilities and association with AAA risk factors. The SNPs were annotated using snpXplorer AnnotateMe tool to identify their chromosomal position, minor allele frequency, CADD (Combined Annotation Dependent Depletion), annotation-based pathogenicity score, variant consequence, and their associated gene. Gene enrichment analysis was performed using Gene Ontology and clustered using REVIGO. The plug-in GeneMANIA in Cytoscape was applied to identify network integration with associated genes and functions. 15 SNPs affecting 20 genes with a CADD score above ten were identified. AAA SNPs were predominantly located on chromosome 3 and 9. Stop-gained rs5516 SNP obtained high frequency in AAA and associated with proinflammatory and vascular remodeling phenotypes. SNPs presence positively correlated with hypertension, dyslipidemia and smoking history. GO showed that AAA SNPs and their associated genes could regulate lipid metabolism, extracellular matrix organization, smooth muscle cell proliferation, and oxidative stress, suggesting that part of these AAA traits could stem from genetic abnormalities. We show a library of inborn SNPs and associated genes that manifest in AAA. We uncover their pathological signaling trajectories that likely fuel AAA development.

microRNA-221 and tamoxifen resistance in luminal-subtype breast cancer patients: A case-control study.

BACKGROUND: Around 70% of breast cancers (BCs) are estrogen receptor-α (ERα)-positive. Adjuvant endocrine therapy is used to reduce estrogen levels and inhibit signal transduction through the ER. The anti-estrogen drugs that are most commonly used in endocrine therapy belong to the selective ER modulator (SERM) class and include tamoxifen. Although it has been used for three decades in cases of early-stage and ERα-positive BC, resistance to tamoxifen is a common problem. microRNAs (miRNAs) have a potential role in demonstrating BC resistance to tamoxifen therapy. Hence, there is a need to investigate the expression of miRNA-221 (miR-221) in luminal-subtype BC patients receiving tamoxifen therapy. METHODS: This case-control study investigated luminal-subtype BC patients who had undergone endocrine therapy for at least 1 year. The case group comprised patients with local or metastatic recurrence, and the control group comprised patients without local or metastatic recurrence. RESULTS: There was a significant difference in miR-221 expression (p = 0.005) between the case and control groups. There were no significant differences between the groups that were positive and negative for the progesterone receptor (PR) (p = 0.25), had high and low marker of proliferation Ki-67 levels (p = 0.60), were positive and negative for lymphovascular invasion (p = 0.14), and had stage 2 and stage 3 cancer (p = 0.25). CONCLUSION: miR-221 expression was higher in tamoxifen-resistant BC cases. miR-221 is a potential biomarker of tamoxifen resistance.

The Role of microRNAs in the Cisplatin- and Radio-Resistance of Cervical Cancer.

Cervical cancer is the fourth leading cause of cancer-related death among women worldwide. The chemotherapeutical agent cisplatin, a small platinum-based compound, is considered as the standard therapy for locally advanced cervical cancer or recurrent cancers, sometimes in combination with radiotherapy or other drugs. However, drug resistance and radio-resistance phenomena could reduce the life expectancy of cervical cancer patients. Resistance mechanisms are complex and often involve multiple cellular pathways in which microRNAs (miRNAs) play a fundamental role. miRNAs are a class of endogenous non-coding small RNAs responsible for post-transcriptional gene regulation. Convincing evidence demonstrates that several deregulated miRNAs are important regulators in the onset of drug and radioresistance in cervical cancer, thus underlying their potential applications in a clinical setting. In this review, we summarized the mechanisms by which miRNAs affect both cisplatin and radioresistance in cervical cancer. We also described the regulatory loops between miRNAs and lncRNAs promoting drug resistance. Besides, we reported evidence for the role of miRNAs in sensitizing cancer cells to cisplatin-based chemotherapy, and provided some suggestions for the development of new combined therapies for cervical cancer.

HCV Proteins Modulate the Host Cell miRNA Expression Contributing to Hepatitis C Pathogenesis and Hepatocellular Carcinoma Development.

Hepatitis C virus (HCV) genome encodes for one long polyprotein that is processed by cellular and viral proteases to generate 10 polypeptides. The viral structural proteins include the core protein, and the envelope glycoproteins E1 and E2, present at the surface of HCV particles. Non-structural (NS) proteins consist of NS1, NS2, NS3, NS4A, NS4B, NS5a, and NS5b and have a variable function in HCV RNA replication and particle assembly. Recent findings evidenced the capacity of HCV virus to modulate host cell factors to create a favorable environment for replication. Indeed, increasing evidence has indicated that the presence of HCV is significantly associated with aberrant miRNA expression in host cells, and HCV structural and non-structural proteins may be responsible for these alterations. In this review, we summarize the recent findings on the role of HCV structural and non-structural proteins in the modulation of host cell miRNAs, with a focus on the molecular mechanisms responsible for the cell re-programming involved in viral replication, immune system escape, as well as the oncogenic process. In this regard, structural and non-structural proteins have been shown to modulate the expression of several onco-miRNAs or tumor suppressor miRNAs.

Weighted miRNA co-expression networks analysis identifies circulating miRNA predicting overall survival in hepatocellular carcinoma patients.

The weighted gene co-expression network analysis (WGCNA) has been used to explore gene expression datasets by constructing biological networks based on the likelihood expression profile among genes. In recent years, WGCNA found application in biomarker discovery studies, including miRNA. Serum samples from 20 patients with hepatocellular carcinoma (HCC) were profiled through miRNA 3.0 gene array and miRNAs biomarker candidates were identified through WGCNA. Results were validated by qRT-PCR in 102 HCC serum samples collected at diagnosis. WGCNA identified 16 miRNA modules, nine of them were significantly associated with the clinical characteristics of the patient. The Red module had a significant negative correlation with patients Survival (- 0.59, p = 0.007) and albumin (- 0.52, p = 0.02), and a positive correlation with PCR (0.61, p = 0.004) and alpha-fetoprotein (0.51, p = 0.02). In the red module, 16 circulating miRNAs were significantly associated with patient survival. MiR-3185 and miR-4507 were identified as predictors of patient survival after the validation phase. At diagnosis, high expression of circulating miR-3185 and miR-4507 identifies patients with longer survival (HR 2.02, 95% CI 1.10-3.73, p = 0.0086, and HR of 1.75, 95% CI 1.02-3.02, p = 0.037, respectively). Thought a WGCNA we identified miR-3185 and miR-4507 as promising candidate biomarkers predicting a longer survival in HCC patients.

The Crosstalk between Tumor Cells and the Microenvironment in Hepatocellular Carcinoma: The Role of Exosomal microRNAs and their Clinical Implications.

The communication between hepatocellular carcinoma (HCC) cells and their microenvironment is an essential mechanism supporting or preventing tumor development and progression. Recent evidence has identified extracellular vesicles (EVs) as one of the mechanisms mediating paracrine signaling between cells. Exosomes, the most described class of EVs, deliver proteins, mRNAs, noncoding RNAs, DNA, and lipids to recipient cells, also at remote distances. MicroRNAs (miRNAs), as part of the non-coding RNA exosomal cargo, have an important role in regulating cellular pathways in targeted cells, regulating several processes related to tumor progression invasion and metastasis, such as angiogenesis, immune-escape, epithelial-to-mesenchymal transition, invasion, and multi-drug resistance. Accumulating evidence suggests exosomal miRNAs as relevant players in the dynamic crosstalk among cancerous, immune, and stromal cells in establishing the tumorigenic microenvironment. In addition, they sustain the metastasic niche formation at distant sites. In this review, we summarized the recent findings on the role of the exosome-derived miRNAs in the cross-communication between tumor cells and different hepatic resident cells, with a focus on the molecular mechanisms responsible for the cell re-programming. In addition, we describe the clinical implication derived from the exosomal miRNA-driven immunomodulation to the current immunotherapy strategies and the molecular aspects influencing the resistance to therapeutic agents.

Circulatory miRNA as a Biomarker for Therapy Response and Disease-Free Survival in Hepatocellular Carcinoma.

The clinical outcome of hepatocellular carcinoma (HCC) treatment remains unsatisfactory, contributing to the high mortality of HCC worldwide. Circulating miRNAs have the potential to be a predictor of therapy response. Microarray profiling was performed in serum samples of 20 HCC patients before treatment. Circulating miRNAs associated with treatment response were validated in 86 serum HCC samples using the qRT-PCR system. Patients were treated either with curative treatments (resection or radiofrequency) or trans-arterial chemoembolization (TACE), and grouped according to therapy response in complete responders (CR) and partial responders or progressive disease (PRPD), following mRECIST criteria. Four miRNA candidates from the discovery phase (miR-4443, miR-4454, miR-4492, and miR-4530) were validated. Before therapy, miR-4454 and miR-4530 were up-regulated in CR to curative treatments (2.83 fold, p = 0.02 and 2.33 fold, p = 0.008, respectively) and were able to differentiate CR from PRPD (area under the curve (AUC) = 0.74, sens/spec 79/63% and AUC = 0.77, sens/spec 72/73%). On the contrary, miR-4443 was 1.95 times down-regulated in CR (p = 0.05) with an AUC of 0.72 (sens = 70%, spec = 60%) in distinguishing CR vs. PRPD). The combination of the three miRNAs was able to predict the response to curative treatment with an AUC of 0.84 (sens = 72%, spec = 75%). The higher levels of miR-4454 and miR-4530 in were associated to longer overall survival (HR = 2.79, p = 0.029 and HR = 2.97, p = 0.011, respectively). Before TACE, miR-4492 was significantly up-regulated in CR patients (FC = 2.67, p = 0.01) and able to differentiate CR from PRPD (AUC = 0.84, sens/spec 84.6/71%). We demonstrated that different miRNAs predictors can be used as potential prognostic circulating biomarkers according to the selected treatment for HCC.

Selection and validation of miR-1280 as a suitable endogenous normalizer for qRT-PCR Analysis of serum microRNA expression in Hepatocellular Carcinoma.

Normalization procedures for the qRT-PCR analysis of miRNA in biological samples are recommended to reduce the variability caused by pre-analytical factors. Since there is no universal standardized normalization strategy for miRNA qRT-PCR studies, we conducted a throughout study to evaluate a panel of small non-coding RNAs (sncRNAs) as reference gene candidate for biomarker studies in serum samples of patients with hepatocellular carcinoma (HCC). Five sncRNAs (miR-1280, miR-1275, SNORD-116, SNORD-68, and U6) were chosen as candidate of reference genes. This study included 122 patients with HCC and was organized into a "pilot phase" consisting of 20 serum samples of HCC patients, and a "validation phase" of 102 patients. Expression level of these candidates were analyzed by qRT-PCR. Assessment of gene stability was performed using four different integrative platforms (geNorm NormFinder, Bestkeeper, and the Delta Ct method). To determine the gene stability during the follow-up of the patient, we extend the analysis of the validation cohort at T1 (1 month after treatment) and T2 (6 month after treatment). MiR-1280 was identified as the most stably expressed reference gene in both pilot and validation phase also during the follow-up. MiR-1280 appears a reliable reference gene candidate in biomarker studies.

Circulating microRNA Associated to Different Stages of Liver Steatosis in Prader-Willi Syndrome and Non-Syndromic Obesity.

BACKGROUND: Prader-Willi syndrome (PWS) is a rare and poorly characterized disease. Recent genomic and transcriptomic studies contributed to elucidate the molecular bases of the syndrome. In this study, we characterized the expression of circulating miRNAs in patients with PWS compared to those with non-syndromic obesity in association with liver steatosis. METHODS: MiRNAs were studied by qRT-PCR in serum samples from 30 PWS and 30 non-syndromic obese subjects. RESULTS: MiRNA expression was associated with the presence of the syndrome and to the grade of liver steatosis. MiR-122-5p, miR-151a, miR-92a-3p were up-regulated in obese (4.38-fold, p < 0.01; 2.72-fold, p < 0.05; 1.34-fold p < 0.05, respectively) and were able to differentiate obese from PWS (AUC = 0.81, sens/spec 78/71%). When stratifying groups according to the presence of steatosis, the expression of miR-151a-5p, miR-92a-3p, miR-106b-5p, and miR-93-5p were lower in PWS with steatosis grade 1. Within the group with steatosis grade 1, miR-151a-5p was significantly distinguished PWS from obese (AUC = 0.85, sens/spec 80/85%) and the combination of miR-106b-5p and miR-93-5p showed higher performances in discriminating different grades of steatosis in PWS (AUC = 0.84, sens/spec 93/74%). CONCLUSIONS: MiRNAs represent a tool to better classify and characterize PWS, providing new information about the clinical picture and the extent of steatosis.

The role of microRNA in the resistance to treatment of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the second most common cause of cancer-related death with a limited efficacy of treatment for intermediate and advanced stages of the disease. Several therapeutic approaches such as trans-arterial chemoembolization (TACE) with anthracyclines, cisplatin and multikinase inhibitor sorafenib have been appealing choices of treatments yet failed to reach a satisfactory outcome mainly due to the numerous mechanisms that influence patient's response. MicroRNAs (miRNAs) are key regulators of many intracellular processes related to drug resistance. This phenomenon has been linked to the modulation of several complex pathways, ranging from the loss of ability of drug accumulation, protective mechanism of autophagy, adaptive mechanism of cancer cells towards the drugs-induced environment, decrease DNA damage and suppression of downstream events that transduce its signal into apoptosis. We summarize the recent findings on the involvement of miRNAs in various drug resistance-related mechanisms in the development of resistance to anthracyclines, cisplatin and sorafenib therapies. Furthermore, we describe the possible application of miRNAs as circulating biomarkers predicting therapy response in HCC. Thus, the undeniable potential and paramount role of miRNA in drug resistance may eventually lead to improved clinical strategies and outcomes for HCC patients.

Serum miRNA Are Promising Biomarkers for the Detection of Early Hepatocellular Carcinoma after Treatment with Direct-Acting Antivirals.

Direct antiviral agents (DAAs) have excellent efficacy against chronic hepatitis C virus (HCV) infection. Despite this strength, recent studies raised concerns about an unexpected hepatocellular carcinoma (HCC) occurrence rate after DAA therapy. In this exploratory case-control study, we evaluated the potential use of miRNAs as serum biomarkers for the detection of early HCC in DAA-treated patients. In the discovery phase, the circulating miRNome was assessed in 10 matched patients with (HCC+) or without HCC (HCC-) occurrence. Microarray analysis was performed before (T0) and after one month of the DAA therapy (T1). MiRNAs discriminating HCC+ and HCC- patients were validated in 60 samples by means of RT-qPCR. We estimated the time-averaged difference of a given miRNA between HCC+ and HCC- patients using a bootstrapped random-effect generalized least square regression model (RE-GLS). At T0, miR-1207-5p, miR-1275, miR-3197, miR-4443, miR-3178, miR-483-5p, miR-4706, miR-4793-3p and miR-1246 discriminated HCC+ from HCC- patients (p < 0.05). At T1, only miR-1180-3p, miR-1228-3p, miR-4329 and miR-4484 (p < 0.05) discriminated HCC+ from HCC- patients. The subsequent validation phase identified miR-3197 as changing with both disease and time. Our results suggest that patients might be already committed to HCC occurrence before DAA therapy. MiR-3197 shows some potential for the identification of patients at risk of HCC during DAA treatments.

Evaluation of rapid GeneXpert MTB/RIF method using DNA tissue specimens of vertebral bones in patients with suspected spondylitis TB.

PURPOSE: To detect Mycobacterium tuberculosis DNA and rifampicin resistance in vertebral bone tissue specimens from spondylitis TB suspects. METHODS AND RESULTS: The rapid GeneXpert MTB/RIF and MGIT 960 liquid culture methods have been used in the specimens. Results from 70 suspects with spondylitis TB shown that 31.42% identified positive for spondylitis TB using culture method, while 88.57% shown positive results using rapid GeneXpert MTB/RIF method. The validity of GeneXpert MTB/RIF shown sensitivity value of 100%, specificity value of 16.6%, PPV of 35.48%, and NPV of 100%. CONCLUSION: GeneXpert has a high sensitivity but low specificity value in this study.