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Frontiers in Oncology 2024Pervasive transcription of the eukaryotic genome generates noncoding RNAs (ncRNAs), which regulate messenger RNA (mRNA) stability and translation. MicroRNAs...
BACKGROUND
Pervasive transcription of the eukaryotic genome generates noncoding RNAs (ncRNAs), which regulate messenger RNA (mRNA) stability and translation. MicroRNAs (miRNAs/miRs) represent a group of well-studied ncRNAs that maintain cellular homeostasis. Thus, any aberration in miRNA expression can cause diseases, including carcinogenesis. According to microRNA microarray analyses, intronic miR-617 is significantly downregulated in oral squamous cell carcinoma (OSCC) tissues compared to normal oral tissues.
METHODS
The miR-617-mediated regulation of is established by performing experiments on OSCC cell lines, patient samples, and xenograft nude mice model. Overexpression plasmid constructs, bisulphite sequencing PCR, bioinformatics analyses, RT-qPCR, Western blotting, dual-luciferase reporter assay, and cell-based assays are utilized to delineate the role of miR-617 in OSCC.
RESULTS
The present study shows that miR-617 has an anti-proliferative role in OSCC cells and is partly downregulated in OSCC cells due to the hypermethylation of its independent promoter. Further, we demonstrate that miR-617 upregulates gene by interacting with its promoter in a dose-dependent and sequence-specific manner, and this interaction is found to be biologically relevant in OSCC patient samples. Subsequently, we show that miR-617 regulates cell proliferation, apoptosis, and anchorage-independent growth of OSCC cells by modulating DDX27 levels. Besides, our study shows that miR-617 exerts its effects through the PI3K/AKT/MTOR pathway via regulating DDX27 levels. Furthermore, the OSCC xenograft study in nude mice shows the anti-tumorigenic potential of miR-617.
CONCLUSION
miR-617-mediated upregulation of DDX27 is a novel mechanism in OSCC and underscores the therapeutic potential of synthetic miR-617 mimics in cancer therapeutics. To the best of our knowledge, miR-617 is the 15th example of a miRNA that upregulates the expression of a protein-coding gene by interacting with its promoter.
PubMed: 38939334
DOI: 10.3389/fonc.2024.1411539 -
Molecular Therapy. Nucleic Acids Sep 2024Effective therapeutics are necessary for managing severe COVID-19 disease despite the availability of vaccines. Small interfering RNA (siRNA) can silence viral genes and...
Effective therapeutics are necessary for managing severe COVID-19 disease despite the availability of vaccines. Small interfering RNA (siRNA) can silence viral genes and restrict SARS-CoV-2 replication. Cell-penetrating peptides is a robust method for siRNA delivery, enhancing siRNA stability and targeting specific receptors. We developed a peptide HE25 that blocks SARS-CoV-2 replication by various mechanisms, including the binding of multiple receptors involved in the virus's internalization, such as ACE2, integrins and NRP1. HE25 not only acts as a vehicle to deliver the SARS-CoV-2 RNA-dependent RNA polymerase siRNA into cells but also facilitates their internalization through endocytosis. Once inside endosomes, the siRNA is released into the cytoplasm through the Histidine-proton sponge effect and the selective cleavage of HE25 by cathepsin B. These mechanisms effectively inhibited the replication of the ancestral SARS-CoV-2 and the Omicron variant BA.5 . When HE25 was administered , either by intravenous injection or inhalation, it accumulated in lungs, veins and arteries, endothelium, or bronchial structure depending on the route. Furthermore, the siRNA/HE25 complex caused gene silencing in lung cells . The SARS-CoV-2 siRNA/HE25 complex is a promising therapeutic for COVID-19, and a similar strategy can be employed to combat future emerging viral diseases.
PubMed: 38939051
DOI: 10.1016/j.omtn.2024.102227 -
Journal of Extracellular Biology Sep 2023Extracellular vesicles (EVs) recently emerged as important players in the pathophysiology of parasitic infections. While the protist parasite can produce EVs, their...
Extracellular vesicles (EVs) recently emerged as important players in the pathophysiology of parasitic infections. While the protist parasite can produce EVs, their role in giardiasis remains obscure. can disrupt gut microbiota biofilms and transform commensal bacteria into invasive pathobionts at sites devoid of colonizing trophozoites via unknown mechanisms. We hypothesized that EVs could modify gut bacterial behaviour via a novel mode of trans-kingdom communication. Our findings indicate that EVs exert bacteriostatic effects on HB101 and TW1, increasing their swimming motility. EVs also decreased the biofilm-forming ability of HB101 but not by TW1, supporting the hypothesis that these effects are, at least in part, bacteria-selective. HB101 and TW1 exhibited increased adhesion/invasion onto small intestine epithelial cells when exposed to EVs. EVs labelled with PKH67 revealed colocalization with HB101 and TW1 bacterial cells. Small RNA sequencing revealed a high abundance of ribosomal RNA (rRNA)- and transfer RNA (tRNA)-derived small RNAs, short-interfering RNAs (siRNAs) and micro-RNAs (miRNAs) within EVs. Proteomic analysis of EVs uncovered the presence of RNA chaperones and heat shock proteins that can facilitate the thermal stability of EVs and its sRNA cargo, as well as protein-modifying enzymes. In vitro, RNase heat-treatment assays showed that total RNAs in EVs, but not proteins, are responsible for modulating bacterial swimming motility and biofilm formation. small RNAs of EVs, but not proteins, were responsible for the increased bacterial adhesion to intestinal epithelial cells induced upon exposure to EVs. Together, the findings indicate that EVs contain a heat-stable, RNase-sensitive cargo that can trigger the development of pathobiont characteristics in Enterobacteria, depicting a novel trans-kingdom cross-talk in the gut.
PubMed: 38938375
DOI: 10.1002/jex2.109 -
Journal of Digestive Diseases Jun 2024We aimed to disclose the molecular mechanism of snail1 in liver fibrosis.
OBJECTIVE
We aimed to disclose the molecular mechanism of snail1 in liver fibrosis.
METHODS
Carbon tetrachloride (CCl) was used to induce a liver fibrosis model in mice whereby serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were evaluated, and liver pathological alternations were assessed. Rat hepatic stellate cells (HSC-T6) were irritated with transforming growth factor (TGF)-β1, followed by assessment of cell viability and migration. The levels of snail1, ALKBH5, and lysine specific demethylase 4C (KDM4C) were quantified by immunohistochemistry, western blot, or reverse transcription-quantitative polymerase chain reaction, in addition to α-smooth muscle actin (SMA), anti-collagen type I α1 (COL1A1), vimentin, and E-cadherin. Photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation and RNA stability were evaluated to determine the relationship between ALKBH5 and snail1. Changes in KDM4C-bound ALKBH5 promoter and enrichment of histone H3 lysine 9 trimethylation (H3K9me3) at the ALKBH5 promoter were determined using chromatin immunoprecipitation.
RESULTS
In fibrosis mice, snail1 was upregulated while ALKBH5 and KDM4C were downregulated. KDM4C overexpression reduced serum ALT and AST levels, liver injury, and α-SMA, COL1A1 and VIMENTIN expressions but increased E-cadherin expression. However, the aforementioned trends were reversed by concurrent overexpression of snail1. In HSC-T6 cells exposed to TGF-β1, ALKBH5 overexpression weakened cell viability and migration, downregulated α-SMA, COL1A1 and VIMENTIN, upregulated E-CADHERIN, and decreased m6A modification of snail1 and its mRNA stability. KDM4C increased ALKBH5 expression by lowering H3K9me3 level, but inhibited HSC-T6 cell activation by regulating the ALKBH5/snail1 axis.
CONCLUSION
KDM4C decreases H3K9me3 methylation to upregulate ALKBH5 and subsequently inhibits snail1, ultimately impeding liver fibrosis.
PubMed: 38938016
DOI: 10.1111/1751-2980.13291 -
Journal of Microbiology and... Jun 2024The gastrointestinal (GI) tract of shrimp, which is comprised of the stomach, hepatopancreas, and intestine, houses microbial communities that play crucial roles in...
The gastrointestinal (GI) tract of shrimp, which is comprised of the stomach, hepatopancreas, and intestine, houses microbial communities that play crucial roles in immune defense, nutrient absorption, and overall health. While the intestine's microbiome has been well-studied, there has been limited research investigating the stomach and hepatopancreas. The present study addresses this gap by profiling the bacterial community in these interconnected GI segments of Pacific whiteleg shrimp. To this end, shrimp samples were collected from a local aquaculture farm in South Korea, and 16S rRNA gene amplicon sequencing was performed. The results revealed significant variations in bacterial diversity and composition among GI segments. The stomach and hepatopancreas exhibited higher Proteobacteria abundance, while the intestine showed a more diverse microbiome, including Cyanobacteria, Actinobacteria, Bacteroidetes, Firmicutes, Chloroflexi, and Verrucomicrobia. Genera such as , , , , and dominated the intestine, while , , and prevailed in the stomach and hepatopancreas. It is particularly notable that , which is associated with nitrate reduction and pollutant degradation, was prominent in the hepatopancreas. Overall, this study provides insights into the microbial ecology of the Pacific whiteleg shrimp's GI tract, thus enhancing our understanding of shrimp health with the aim of supporting sustainable aquaculture practices.
Topics: Animals; Penaeidae; Hepatopancreas; RNA, Ribosomal, 16S; Bacteria; Gastrointestinal Microbiome; Republic of Korea; Intestines; Phylogeny; Stomach; Biodiversity; Aquaculture; DNA, Bacterial
PubMed: 38938005
DOI: 10.4014/jmb.2403.03039 -
Cell Communication and Signaling : CCS Jun 2024Tumor cells release extracellular vesicles (EVs) that contribute to the polarization of macrophages towards tumor-associated macrophages (TAMs). High expression levels...
BACKGROUND
Tumor cells release extracellular vesicles (EVs) that contribute to the polarization of macrophages towards tumor-associated macrophages (TAMs). High expression levels of the RNA binding protein IGF2BP2/IMP2 are correlated with increased tumor cell proliferation, invasion, and poor prognosis in the clinic. However, there is a lack of understanding of whether IMP2 affects the cargo of cancer cell-derived EVs, thereby modulating macrophage polarization.
METHODS
EVs were isolated from IMP2-expressing HCT116 parental cells (WT) and CRISPR/Cas9 IMP2 knockout (KO) cells. EVs were characterized according to MISEV guidelines, microRNA cargo was assessed by microRNA-Seq, and the protein cargo was analyzed by proteomics. Primary human monocyte-derived macrophages (HMDMs) were polarized by EVs, and the expression of genes and surface markers was assessed using qPCR and flow cytometry, respectively. Morphological changes of macrophages, as well as the migratory potential of cancer cells, were assessed by the Incucyte system and macrophage matrix degradation potential by zymography. Changes in the metabolic activity of macrophages were quantified using a Seahorse analyzer. For in vivo studies, EVs were injected into the yolk sac of zebrafish larvae, and macrophages were isolated by fluorescence-activated cell sorting.
RESULTS
EVs from WT and KO cells had a similar size and concentration and were positive for 25 vesicle markers. The expression of tumor-promoting genes was higher in macrophages polarized with WT EVs than KO EVs, while the expression of TNF and IL6 was reduced. A similar pattern was observed in macrophages from zebrafish larvae treated in vivo. WT EV-polarized macrophages showed a higher abundance of TAM-like surface markers, higher matrix degrading activity, as well as a higher promotion of cancer cell migration. MicroRNA-Seq revealed a significant difference in the microRNA composition of WT and KO EVs, particularly a high abundance of miR-181a-5p in WT EVs, which was absent in KO EVs. Inhibitors of macropinocytosis and phagocytosis antagonized the delivery of miR-181a-5p into macrophages and the downregulation of the miR-181a-5p target DUSP6. Proteomics data showed differences in protein cargo in KO vs. WT EVs, with the differentially abundant proteins mainly involved in metabolic pathways. WT EV-treated macrophages exhibited a higher basal oxygen consumption rate and a lower extracellular acidification rate than KO EV-treated cells.
CONCLUSION
Our results show that IMP2 determines the cargo of EVs released by cancer cells, thereby modulating the EVs' actions on macrophages. Expression of IMP2 is linked to the secretion of EVs that polarize macrophages towards a tumor-promoting phenotype.
Topics: Humans; Extracellular Vesicles; RNA-Binding Proteins; Animals; Zebrafish; Tumor-Associated Macrophages; HCT116 Cells; MicroRNAs; Cell Movement; Macrophages
PubMed: 38937789
DOI: 10.1186/s12964-024-01701-y -
Cancer Cell International Jun 2024Triple negative breast cancer (TNBC) is a type of cancer that lacks receptor expression and has complex molecular mechanisms. Recent evidence shows that the...
Triple negative breast cancer (TNBC) is a type of cancer that lacks receptor expression and has complex molecular mechanisms. Recent evidence shows that the ubiquitin-protease system is closely related to TNBC. In this study, we obtain a key ubiquitination regulatory substrate-ABI2 protein by bioinformatics methods, which is also closely related to the survival and prognosis of TNBC. Further, through a series of experiments, we demonstrated that ABI2 expressed at a low level in TNBC tumors, and it has the ability to control cell cycle and inhibit TNBC cell migration, invasion and proliferation. Molecular mechanism studies proved E3 ligase CBLC could increase the ubiquitination degradation of ABI2 protein. Meanwhile, RNA-seq and IP experiments indicated that ABI2, acting as a crucial factor of tumor suppression, can significantly inhibit PI3K/Akt signaling pathway via the interaction with Rho GTPase RAC1. Finally, based on TNBC drug target ABI2, we screened and found that FDA-approved drug Colistimethate sodium(CS) has significant potential in suppressing the proliferation of TNBC cells and inducing cell apoptosis, making it a promising candidate for impeding the progression of TNBC.
PubMed: 38937761
DOI: 10.1186/s12935-024-03407-0 -
BMC Infectious Diseases Jun 2024When COVID-19 hit the world in 2019, an enhanced focus on diagnostic testing for SARS-CoV-2 was essential for a successful pandemic response. Testing laboratories...
INTRODUCTION
When COVID-19 hit the world in 2019, an enhanced focus on diagnostic testing for SARS-CoV-2 was essential for a successful pandemic response. Testing laboratories stretched their capabilities for the new coronavirus by adopting different test methods. The necessity of having external quality assurance (EQA) mechanisms was even more critical due to this rapid expansion. However, there was a lack of experience in providing the necessary SARS-CoV-2 EQA materials, especially in locations with constrained resources.
OBJECTIVE
We aimed to create a PT (Proficiency testing) programme based on the Dried Tube Specimens (DTS) method that would be a practical option for molecular based SARS-CoV-2 EQA in Low- and Middle-Income Countries.
METHODS
Based on previous ISO/IEC 17043:2010 accreditation experiences and with assistance from the US Centers for Disease Control and Prevention, The Supranational Reference Laboratory of Uganda (adapted the DTS sample preparation method and completed a pilot EQA program between 2020 and 2021. Stability and panel validation testing was conducted on the designed materials before shipping to pilot participants in six African countries. Participants received a panel containing five SARS-CoV-2 DTS samples, transported at ambient conditions. Results submitted by participants were compared to validation results. Participants were graded as satisfactory (≥ 80%) or unsatisfactory (< 80%) and performance reports disseminated.
RESULTS
Our SARS-CoV-2 stability experiments showed that SARS-CoV-2 RNA was stable (-15 to -25 °C, 4 to 8 °C, (18 to 28 °C) room temperature and 35 to 38 °C) as well as DTS panels (4 to 8 °C, 18 to 28 °C, 35 to 38 °C and 45 °C) for a period of 4 weeks. The SARS-CoV-2 DTS panels were successfully piloted in 35 test sites from Zambia, Malawi, Mozambique, Nigeria, and Seychelles. The pilot results of the participants showed good accuracy, with an average of 86% (30/35) concordance with the original SARS CoV-2 expectations.
CONCLUSION
The SARS-CoV-2 DTS PT panel is reliable, stable at ambient temperature, simple to prepare and requires minimal resources.
Topics: Humans; COVID-19; SARS-CoV-2; Specimen Handling; Laboratory Proficiency Testing; Developing Countries; COVID-19 Testing; Uganda; Pilot Projects
PubMed: 38937708
DOI: 10.1186/s12879-024-09555-y -
The Role of N6-methyladenosine Modification in Gametogenesis and Embryogenesis: Impact on Fertility.Genomics, Proteomics & Bioinformatics Jun 2024The most common epigenetic modification of messenger ribonucleic acids (mRNAs) is N6-methyladenosine (m6A), which is mainly located near the 3' untranslated region of...
The most common epigenetic modification of messenger ribonucleic acids (mRNAs) is N6-methyladenosine (m6A), which is mainly located near the 3' untranslated region of mRNAs, near the stop codons, and within internal exons. The biological effect of m6A is dynamically modified by methyltransferases (writers), demethylases (erasers), and m6A-binding proteins (readers). By controlling post-transcriptional gene expression, m6A has a significant impact on numerous biological functions, including RNA transcription, translation, splicing, transport, and degradation. Hence, m6A influences various physiological and pathological processes, such as spermatogenesis, oogenesis, embryogenesis, placental function, and human reproductive system diseases. During gametogenesis and embryogenesis, genetic material undergoes significant changes, including epigenomic modifications such as m6A. From spermatogenesis and oogenesis to the formation of an oosperm and early embryogenesis, m6A changes occur at every step. m6A abnormalities can lead to gamete abnormalities, developmental delays, impaired fertilization, and maternal-to-zygotic transition blockage. Both mice and humans with abnormal m6A modifications exhibit impaired fertility. In this review, we discuss the dynamic biological effects of m6A and its regulators on gamete and embryonic development and review the possible mechanisms of infertility caused by m6A changes. We also discuss the drugs currently used to manipulate m6A and provide prospects for the prevention and treatment of infertility at the epigenetic level.
PubMed: 38937660
DOI: 10.1093/gpbjnl/qzae050 -
Scientific Reports Jun 2024There is no treatment for acute aortic dissection (AAD) targeting inflammatory cells. We aimed to identify the new therapeutic targets associated with inflammatory...
There is no treatment for acute aortic dissection (AAD) targeting inflammatory cells. We aimed to identify the new therapeutic targets associated with inflammatory cells. We characterized the specific distribution of myeloid cells of both human type A AAD samples and a murine AAD model generated using angiotensin II (ANGII) and β-aminopropionitrile (BAPN) by single-cell RNA sequencing (scRNA-seq). We also examined the effect of an anti-interleukin-1β (IL-1β) antibody in the murine AAD model. IL1B inflammatory macrophages and classical monocytes were increased in human AAD samples. Trajectory analysis demonstrated that IL1B inflammatory macrophages differentiated from S100A8/9/12 classical monocytes uniquely observed in the aorta of AAD. We found increased infiltration of neutrophils and monocytes with the expression of inflammatory cytokines in the aorta and accumulation of inflammatory macrophages before the onset of macroscopic AAD in the murine AAD model. In blocking experiments using an anti-IL-1β antibody, it improved survival of murine AAD model by preventing elastin degradation. We observed the accumulation of inflammatory macrophages expressing IL-1β in both human AAD samples and in a murine AAD model. Anti-IL-1β antibody could improve the mortality rate in mice, suggesting that it may be a treatment option for AAD.
Topics: Aortic Dissection; Interleukin-1beta; Animals; Humans; Macrophages; Mice; Disease Models, Animal; Male; Aminopropionitrile; Angiotensin II; Inflammation; Monocytes; Aorta; Mice, Inbred C57BL; Female
PubMed: 38937528
DOI: 10.1038/s41598-024-65931-3