Current advances in circular RNA detection and investigation methods: Are we running in circles?

Circular RNAs (circRNAs), characterized by their closed-loop structure, have emerged as significant transcriptomic regulators, with roles spanning from microRNA sponging to modulation of gene expression and potential peptide coding. The discovery and functional analysis of circRNAs have been propelled by advancements in both experimental and bioinformatics tools, yet the field grapples with challenges related to their detection, isoform diversity, and accurate quantification. This review navigates through the evolution of circRNA research methodologies, from early detection techniques to current state-of-the-art approaches that offer comprehensive insights into circRNA biology. We examine the limitations of existing methods, particularly the difficulty in differentiating circRNA isoforms and distinguishing circRNAs from their linear counterparts. A critical evaluation of various bioinformatics tools and novel experimental strategies is presented, emphasizing the need for integrated approaches to enhance our understanding and interpretation of circRNA functions. Our insights underscore the dynamic and rapidly advancing nature of circRNA research, highlighting the ongoing development of analytical frameworks designed to address the complexity of circRNAs and facilitate the assessment of their clinical utility. As such, this comprehensive overview aims to catalyze further advancements in circRNA study, fostering a deeper understanding of their roles in cellular processes and potential implications in disease. This article is categorized under: RNA Methods > RNA Nanotechnology RNA Methods > RNA Analyses in Cells RNA Methods > RNA Analyses In Vitro and In Silico.

MicroRNAs: Potential Biomarkers of Disease Severity in Chronic Rhinosinusitis with Nasal Polyps.

Background and Objectives: Chronic rhinosinusitis with nasal polyps (CRwNP) has multiple clinical presentations, and predictors of successful treatment are correlated to different parameters. Differentially expressed microRNAs in nasal polyps emerge as possible facilitators of precise endotyping in this disease. We aimed to evaluate the correlation between the clinical parameters of CRSwNP and two different microRNAs. Materials and Methods: The expression of miR-125b and miR-203a-3p in nasal polyps (n = 86) and normal nasal mucosa (n = 20) was determined through microarray analysis. Preoperative workup included CT scan, nasal endoscopy, blood tests, symptoms and depression questionnaires. Results: MiR-125b showed significant overexpression in NP compared to the normal nasal mucosa. miR-125b expression levels were positively and significantly correlated with blood eosinophilia (p = 0.018) and nasal endoscopy score (p = 0.021). Although high CT scores were related to miR-125b overexpression, the correlation did not reach statistical significance. miR-203a-3p was underexpressed in nasal polyps and was significantly underexpressed in CRSwNP patients with environmental allergies. Conclusions: Both miR-125b and miR-203a-3p are potential biomarkers in CRSwNP. miR-125b also correlates with the clinical picture, while miR-203a-3p could help identify an associated allergy.

A perspective on the applications of furin inhibitors for the treatment of SARS-CoV-2.

Currently, the world is facing a pandemic of the new coronavirus SARS-CoV-2 that causes COVID-19. Identifying key targets in the viral infection lifecycle is urgently needed for designing therapeutic strategies to combat the virus. Furin is a subtilisin-like proprotein convertase with diverse cellular functions. Emerging evidence suggests that furin plays a critical role in the activation and/or infectivity of SARS-CoV-2. In this perspective, we discuss the potential role of furin in the entry SARS-CoV-2 into host cells. Furthermore, we evaluate available peptide and non-peptide furin inhibitors and potential outcomes, including immune responses.

MicroRNA profiling in kidney in pigs fed ochratoxin A contaminated diet.

OTA is a toxic metabolite produced by fungus belonging to Aspergillus and Penicillium genera. Kidney is the main target of this toxin; OTA is considered as one of the etiological factors at the origin of the human Balkan endemic nephropathy. microRNA are short non-coding transcrips (18-22 nucleotides in length) regulating key cellular processes. Various miRNAs have been established to play important roles in development of renal carcinoma and urothelial cancer. The objective of this study is to analyse the miRNA profiling in the kidney of piglets experimentally intoxicated with feed contaminated with OTA. Fifteen piglets (five pigs/group) were randomly distributed into 3 groups, fed normal diet (Group 1: control), or diets contaminated with OTA in two concentrations: 50 µg OTA/kg feed (Group 2: 50 µg OTA/kg feed) or 200 µg OTA/kg feed (Group 3: 200 µg OTA/kg feed) for 28 days. At the end of the experiment blood samples were taken for serological analyses. Animals from control group and 200 µg OTA/kg feed were sacrificed and kidney samples were taken for histological and molecular analyses. As resulted from molecular profiling study there are 8 miRNA differentially expressed in OTA kidney vs control kidney, in which five miRNA were overexpressed in the kidney of OTA intoxicated animals: miR-497 (FC = 6.34), miR-133a-3p (FC = 5.75), miR-423-3p (FC = 5.48), miR-34a (FC = 1.68), miR-542-3p (1.65) while three miRNA were downregulated: miR-421-3p (FC = -3.96); miR-490 (FC = -3.87); miR-9840-3p (FC = -2.13). The altered miRNAs as effect of OTA are strongly connected to the engine of cancer, disturbing nodal points in different pathways, as TP53 signalling. This proof-of-concept study proves the actual utility of miRNAs as biomarkers of mycotoxin exposure, including OTA.

The emerging role of exosomes in multiple myeloma.

Multiple myeloma (MM), one of the most prevalent hematological malignancies, accounts for approximately 10% of all blood cancers. In spite of the recent advancements in MM therapy, this malignancy of terminally differentiated plasma cells (PCs) continues to remain a hard-to-cure disease due to the emergence of drug resistance and frequent relapses. It is now well-established that the tumor-supportive involvement of the bone marrow microenvironment (BMM) including the cellular and non-cellular elements are the major causes behind treatment failures of MM as well as its main complications such as osteolytic bone loss. Exosomes (EXs) are membranous structures that carry signaling molecules and have recently received a great deal of attention as important mediators of inter-cellular communication in health and disease. EXs involve in the growth and drug resistance of many tumors via delivering their rich contents of bioactive molecules including miRNAs, growth factors, cytokines, signaling molecules, etc. With regard to MM, many studies have reported that EXs are among the main culprits playing key roles in the vicious network within the BMM of these patients. The main producers of EXs that largely contribute to MM pathogenesis are bone marrow stromal cells (BMSCs) as well as MM cells themselves. These cell types produce large amounts of EXs that affect a variety of target cells including natural killer (NK) cells, osteoclasts (OCs) and osteoblasts (OBs) to the advantage of tumor survival and progression. These EXs contain a different profile of proteins and miRNAs from that of EXs obtained from their counterparts in healthy individuals. MM patients exhibit distinguishable elevations in some of their contents such as miR-21, miR-146a, let-7b and miR-18a, while some molecules like miR-15a are markedly downregulated in EXs of MM patients compared to healthy individuals. These findings make EXs desirable biomarkers for early prediction of disease progression and drug resistance in the context of MM. On the other hand, due to the tumor-supportive role of EXs, targeting these structures in parallel to the conventional therapeutic regimens may be a promising approach to a successful anti-MM therapy. In the present work, an extensive review of the literature has been carried out to highlight the recent advances in the field.

Isolation and Characterization of a Fetal-Maternal Microchimeric Stem Cell Population in Maternal Hair Follicles Long after Parturition.

Fetal-maternal microchimerism describes the acquisition of fetal stem cells (FSC) by the mother during pregnancy and their long-term persistence after parturition. FSC may engraft in a variety of maternal tissues especially if there is organ/tissue injury, but their role and mechanism of persistence still remains elusive. Clinical applications due to their pluripotency, immunomodulatory effects and accessibility make them good candidates for ex-vivo manipulation and autologous therapies. The hair follicles contain a distinctive niche for pluripotent stem cells (PSC). To date, there is no published evidence of fetal microchimerism in the hair follicle. In our study, follicular unit extraction (FUE) technique allowed easy stem cell cultures to be obtained while simple hair follicle removal by pull-out technique failed to generate stem cells in culture. We identified microchimeric fetal stem cells within the primitive population of maternal stem cells isolated from the hair follicles with typical mesenchymal phenotype, expression of PSC genes and differentiation potential towards osteocytes, adypocites and chondrocytes. This is the first study to isolate fetal microchimeric stem cells in adult human hair long after parturition. We presume a sanctuary partition mechanism with PSC of the mother deposited during early embryogenesis could explain their long-term persistence.

MicroRNAs as regulators of apoptosis mechanisms in cancer.

MicroRNAs or miRNAs are small non-coding RNAs that regulate gene expression. Their discovery has brought new knowledge in biological processes of cancer. Involvement of miRNAs in cancer development includes several major pathways from cell transformation to tumor cell development, metastasis and resistance to treatment. The first part of this review discusses miRNAs function in the intrinsic and extrinsic pathways of apoptosis. Due to the fact that many miRNAs that regulate apoptosis have been shown to play a major role in tumor cell resistance to treatment, in the second part of the review we aim at discussing miRNAs potential in becoming curative molecules.

Next generation sequencing applications for breast cancer research.

For some time, cancer has not been thought of as a disease, but as a multifaceted, heterogeneous complex of genotypic and phenotypic manifestations leading to tumorigenesis. Due to recent technological progress, the outcome of cancer patients can be greatly improved by introducing in clinical practice the advantages brought about by the development of next generation sequencing techniques. Biomedical suppliers have come up with various applications which medical researchers can use to characterize a patient's disease from molecular and genetic point of view in order to provide caregivers with rapid and relevant information to guide them in choosing the most appropriate course of treatment, with maximum efficiency and minimal side effects. Breast cancer, whose incidence has risen dramatically, is a good candidate for these novel diagnosis and therapeutic approaches, particularly when referring to specific sequencing panels which are designed to detect germline or somatic mutations in genes that are involved in breast cancer tumorigenesis and progression. Benchtop next generation sequencing machines are becoming a more common presence in the clinical setting, empowering physicians to better treat their patients, by offering early diagnosis alternatives, targeted remedies, and bringing medicine a step closer to achieving its ultimate goal, personalized therapy.

Efficient siRNA delivery system using carboxilated single-wall carbon nanotubes in cancer treatment.

Several functionalized carbon nanotubes have been designed and tested for the purpose of nucleic acid delivery. In this study, the capacity of SWNTC-COOH for siRNA deliverey were investigated delivery in parallel with an efficient commercial system. Hep2G cells were reverse-transfected with 50 nM siRNA (p53 siRNA, TNF-alphasiRNA, VEGFsiRNA) using the siPORT NeoFX (Ambion) transfection agent in paralel with SWNTC-COOH, functionalised with siRNA. The highest level of gene inhibition was observed in the cases treated with p53 siRNA gene; in the case of transfection with siPort, the NeoFX value was 33.8%, while in the case of SWNTC-COOH as delivery system for p53 siRNA was 37.5%. The gene silencing capacity for VEGF was 53.7%, respectively for TNF-alpha 56.7% for siPORT NeoFX delivery systems versus 47.7% (VEGF) and 46.5% (TNF-alpha) for SWNTC-COOH delivery system. SWNTC-COOH we have been showed to have to be an efficient carrier system. The results from the inhibition of gene expresion for both transfection systems were confirmed at protein level. Overall, the lowest mRNA expression was confirmed at protein level, especially in the case of p53 siRNA and TNF-alpha siRNA transfection. Less efficient reduction protein expressions were observed in the case of VEGF siRNA, for both transfection systems at 24 h; only at 48 h, there was a statistically significant reduction of VEGF protein expression. SWCNT-COOH determined an efficient delivery of siRNA. SWNTC-COOH, combined with suitable tumor markers like p53 siRNA, TNFalpha siRNA or VEGF siRNA can be used for the efficient delivery of siRNA.

Is interleukin-17 a proatherogenic biomarker?

The importance of chronic inflammation in atherogenesis and cytokine involvement in all stages of atherosclerotic plaque development is now obvious. Our approach of the significant cytokines involved in atherogenesis or cardiovascular diseases is based on a correlation between clinical research and experiments on animal models. The contribution of IL-17 in atherogenesis remains controversial. In this study we investigated the role of IL-17 in cardiovascular diseases and in atherosclerosis associated with pathological aging. We performed a case-control study, enrolling subjects aged over 65 years in both groups. We included 40 patients with cardiovascular disorders and 10 healthy volunteers. IL-17 levels were measured in the serum of patients and healthy controls, along with serum total cholesterol and triglycerides. Significantly higher levels of IL-17 were obtained in patients compared to healthy controls (p<0.001). The level of this biomarker correlated significantly with two biochemical parameters - serum total cholesterol and triglycerides (the Pearson coefficient showed statistical significance, p=0.033, respectively p=0.043). We did not find any correlation between IL-17 and these two parameters in the control group. Our study is useful in understanding the physiopathological implications of IL-17 in the atherogenesis process. This could represent a starting point for future studies, including research regarding the therapeutic potential of IL-17 in pathological aging.

The relationships between biological activities and structure of flavan-3-ols.

Flavan-3-ols are involved in multiple metabolic pathways that induce inhibition of cell proliferation. We studied the structure-activity relationship of gallic acid (GA) and four flavan-3-ols: epigallocatechin gallate (EGCG), epigallocatechin (EGC), catechin (C), and epicatechin (EC). We measured the cell viability by the MTT assay and we determined the concentration of testing compound required to reduce cell viability by 50% (IC(50)). All tested compounds showed a dose-dependent and time-dependent inhibitory antiproliferative effect on Hs578T cells; IC(50) values varying from the 15.81 to 326.8 µM. Intracellular ROS (reactive oxygen species) were quantified using a fluorescent probe 2',7'-dichlorofluorescin diacetate (DCFH-DA). Only the treatment with 10 µM EGC and EGCG was able to induce a significant decrease of ROS concentration and increased levels of ROS were registered for 100 µM EGCG, EGC and GA. Flavans-3-ols and GA induced apoptosis in a dose- and time-dependent manner, which indicated that the induction of apoptosis mediated their cytotoxic activity at least partially. The galloylated catechins have shown a stronger antiproliferative activity and apoptotic effect than the one produced by non galloylated catechins. The galloylated flavan-3-ols are potential therapeutic agents for patients with triple negative breast cancer via induction of apoptosis.

Clinical and pharmacokinetics study of oxaliplatin in colon cancer patients.

AIM: to evaluate the therapeutic efficacy of oxaliplatin and to analyze the pharmacokinetics of both ultrafiltrable (free) and protein-bound platinum in patients with metastatic colon cancer. METHOD: 60 patients with stage IV colon carcinoma received 4-6 (mean 4.5) cycles of oxaliplatin based combination chemotherapy. Response rate, progression-free survival (PFS) and toxicity were evaluated. The pharmacokinetics of oxaliplatin was evaluated in 8 patients who were given 85 mg/sqm or 130 mg/sqm using an infusion time of 2-4 h. Pharmacokinetic analysis was performed on blood, plasma and plasma ultrafiltrable by ICP-MS (Inductively Coupled Plasma Mass Spectrometry). RESULTS: Overall response rate (complete and partial) occurred in 33 (55%) patients. The median time of progression was 9.3 months. Cumulative neurotoxicity, vomiting and diarrhea, myelosuppression appeared in 32.3%, 21.3%, and 39.4% patients, respectively. The mean Cmax and AUC 0-24 of oxaliplatin increased in a dose-related manner. The pharmacokinetics of platinum after oxaliplatin administration was triphasic characterized by a short initial distribution phase and a long terminal elimination phase.The clearance of ultrafiltrable platinum was relatively high and the clearance of platinum from plasma and blood cells was relatively low, which is probably a reflection of the covalent binding of platinum to these matrices. CONCLUSION: Oxaliplatin is active and well tolerated in patients with advanced colon cancer. With a relatively low interpatient variability, it is eliminated triphasically and the mean Cmax and AUC 0-24 increases in a dose-related manner. These results provide a scientific basis for the safe and effective use of oxaliplatin in the clinic.