This study attempted to investigate the mechanism of miR-204-5p and its downstream gene in regulating bio-functions of esophageal cancer (EC).
Bioinformatics analysis was performed to select the mature miRNAs, mRNAs, and clinical data of EC. The miRNA-mRNA regulatory axis was predicted through bioinformatics and used Dual-luciferase analysis to verify the interaction between miR-204-5p and APLN. qRT-PCR was applied to analyze expression of miR-204-5p and APLN mRNA. Western blot was utilized to detect APLN protein expression. Functional assays like CCK-8, wound healing, Transwell, and stem cell sphere formation assays were launched to confirm proliferative, migratory, invasive and stemness of cells in different treatment groups.
MiR-204-5p was lowly expressed while its target gene APLN was highly expressed in tumor tissues. Besides, miR-204-5p overexpression hindered proliferation, invasion, migration, and stemness of EC cells. Additionally, dual-luciferase assay verified the interaction of miR-204-5p and APLN. MiR-204-5p could downregulate APLN level and its overexpression reduced the effect of APLN on EC cell functions.
Dysregulation of miR-204-5p/APLN axis was linked with malignant progression of EC. MiR-204-5p/APLN may be an underlying candidate for the design of anticarcinogens.
Lung adenocarcinoma (LUAD) is featured in high morbidity and mortality. Aberrant activation of the histone methyltransferase EZH2 has close association with cancer progression. This research aimed to deeply dive into the role and possible molecular mechanisms of EZH2 and its downstream genes in malignant progression and DNA damage repair of LUAD cells.
Expression of EZH2 in LUAD cells was analyzed by qRT-PCR, and the effects of EZH2 on proliferation, and apoptosis of LUAD cells were examined by CCK-8, colony formation and flow cytometry assays. The downstream targets of EZH2 were predicted by bioinformatics analysis. Then, the targeting relationship between EZH2 and RAI2 was examined by CHIP and luciferase reporter assays. Rescue assay were used to further validate the effect of EZH2/RAI2 on the malignant progression of LUAD cells. The expression levels of EZH2, RAI2 and p53 were examined by Western blot.
Upregulation of EZH2 was identified in LUAD tissues and cells. RAI2 was a downstream target gene of EZH2, and the two were negatively correlated. Silencing EZH2 suppressed proliferation of LUAD cells, promoted expression of p53, cell cycle arrest and apoptosis. While silencing RAI2 could reverse the above-mentioned effects caused by EZH2 silencing.
These results demonstrated that EZH2 promoted malignant progression and DNA damage repair of LUAD cells by targeting and negatively regulating RAI2.
Biological mechanism of miR-210-3p in endometrial carcinoma (EC) remains unclear. Here, our purpose is to study effects of miR-210-3p on malignant progression of EC.
Bioinformatics analysis showed miRNA and mRNA are abnormally expressed in EC tissues. Quantitative real-time fluorescence polymerase chain reaction (qRT-PCR) was utilized to compare miR-210-3p mRNA level in EC cells and tissues. qRT-PCR and western blot were used to measure RUNX1T1 and NCAM1 at mRNA and protein levels, and western blot for p-AKT and AKT proteins related to PI3K/AKT signaling pathway. Furthermore, EC cell behaviors were assayed via Cell Counting Kit-8, cell colony formation assay, wound healing, transwell and flow cytometry experiments. Interaction between RUNX1T1 and miR-210-3p was verified through dual-luciferase assay. Immunohistochemistry was used to analyze RUNX1T1 expression in clinical samples
MiR-210-3p was considerably upregulated and RUNX1T1 was significantly under-expressed in EC. Overexpression of miR-210-3p stimulated cell proliferation, migration, invasion, and restrained cell apoptosis in EC. Dual-luciferase assay proved that RUNX1T1 was a target gene of miR-210-3p. The level of RUNX1T1 in EC was downregulated after overexpressing miR-210-3p. Rescue assay showed that overexpression of RUNX1T1 had an inhibitory impact on tumor-relevant cell behaviors, whereas overexpression of miR-210-3p rescued such inhibition. Overexpression of RUNX1T1 reduced p-AKT expression, which was restored with concomitantly overexpressed miR-210-3p.
In general, miR-210-3p behaves as an oncogene in EC by down-regulating the expression of RUNX1T1. This study elucidates a new functional mechanism in EC, and indicates miR-210-3p an underlying target.
Increasing evidence shows that Transmembrane 4 L6 family member 1(TM4SF1) exerts a critical role in mediating the progression of various tumors. Nevertheless, the exact mechanism of TM4SF1 in gastric cancer (GC) remains unclear.
Bioinformatics analysis was utilized to analyze TM4SF1 expression in GC tissues. Also, MiRWalk and starBase databases were used to predict the upstream microRNAs which could regulate TM4SF1 expression. Gene set enrichment analysis (GSEA) for TM4SF1 was conducted to screen the potentially involved pathways. Dysregulation of microRNA-501–3p/TM4SF1 was implemented to investigate the regulatory roles of these genes in GC. qRT-PCR and western blot were employed to measure the expression changes of microRNA-501–3p, TM4SF1, and epithelial-mesenchymal transition (EMT) signaling pathway-associated proteins. CCK-8, colony formation, and transwell assays were introduced to examine the biological functions of GC cell lines.
TM4SF1 presented a significantly low level in mRNA and protein in GC cells. MicroRNA-501–3p could target TM4SF1 and reduce its expression. Cell function experiments revealed that microRNA-501–3p facilitated cell proliferation, migration, and invasion, while inhibiting cell apoptosis in GC by targeting TM4SF1. EMT-associated proteins were altered by changing microRNA-501–3p/TM4SF1 axis.
MicroRNA-501–3p regulated EMT signaling pathway by down-regulating TM4SF1 expression and therefore facilitated the malignant progression of GC, which may provide a new potential therapeutic target for the treatment of GC patients.
Rheumatoid arthritis (RA), which is driven by persistent activation of the immune system, primarily affects the joints. Several reports have estimated the risk of gonadal disruptions in arthritic patients, with potential attributable risk factors such as treatments with the disease-modifying antirheumatic drugs and the influence of the disease itself. The FDA approved rituximab, a therapy for non-Hodgkin's lymphoma, for management of RA in February 2006. However, the influence of repeated treatment with rituximab on gonadal function in RA has not been reported yet. Thus, the aim of the presents study is to evaluate whether repeated treatment with the clinically relevant dose of rituximab may change the gonadal disruptions in collagen-induced arthritis in male DBA/1 J mouse, a model of RA. Testicular disruptions, as determined by the sperm DNA strand breaks, spermatocyte chromosomal analysis and spermiogram examination have been conducted by the use of standard techniques. Additionally, we aimed to test whether the anti-rheumatic effect of rituximab also decreases the cellular oxidant-antioxidant imbalance in arthritic male DBA/1 J mice. Repeated treatment of naïve control DBA/1 J mice with rituximab did not exhibit any significant deleterious effects. Moreover, repeated administration of rituximab to the arthritic DBA/1 J mice suppressed disease severity and decreased testicular disruptions. Rituximab treatment also diminished gonadal oxidative stress, through decreasing reactive oxygen species generation and restoring the reduced glutathione level in arthritic DBA/1 J mice. In conclusion, rituximab is a safe therapeutic agent and can mitigate gonadal disruptions induced by arthritis, which insinuates the importance for arthritic patients especially at reproductive age.
8-Oxo-7,8-dihydroguanine (8-hydroxyguanine, G°) is a major oxidized base that is considered to play pivotal roles in the pathogenesis of various diseases, including cancer. G° induces G:C → T:A transversions at the damage site and untargeted (action-at-a-distance) mutations of G bases at 5′-GpA sequences. In this study, we examined the distribution of the action-at-a-distance mutations and the effects of the replication origin position relative to G° on the untargeted mutagenesis. The G° base was introduced into two shuttle plasmids, each with the SV40 replication origin at a different position with respect to the supF gene. The oxidized base was located at an upstream or downstream site (outside of the gene), or the center of the region encoding the pre-tRNA sequence of the gene, in the sense strand. These shuttle plasmids were introduced into human U2OS cells. The action-at-a-distance mutations were more frequently induced when the G° base was located downstream of the supF gene than upstream of the gene. In addition, more action-at-a-distance mutations were observed when the SV40 origin was present on the 5′-side of the G° base. These results indicated that the action-at-a-distance mutations are predominantly induced on the 5′-side of the lesion and occurred more frequently when the damaged base was located on the lagging strand template.
UV-induced mutagenesis is, to greater extent, a phenomenon dependent on translesion synthesis (TLS) and regulated by the SOS response in bacteria. Caulobacter crescentus, like many bacterial species, employs the ImuABC (ImuAB DnaE2) pathway in TLS. To have a better understanding of the characteristics of UV-induced mutagenesis in this organism, we performed a whole genome analysis of mutations present in survivors after an acute UVC exposure (300 J/m2). We found an average of 3.2 mutations/genome in irradiated samples, distributed in a mutational spectrum consisting exclusively of base substitutions, including tandem mutations. Although limited in conclusions by the small number of mutations identified, our study points to the feasibility of using whole-genome sequencing to study mutagenesis occurring in experiments involving a single acute exposure to genotoxic agents.
Lamin proteins which constitute the nuclear lamina in almost all higher eukaryotes, are mainly of two types A & B encoded by LMNA and LMNB1/B2 genes respectively. While lamin A remains the principal product of LMNA gene, variants like lamin C, C2 and A∆10 are also formed as alternate splice products. Role of lamin A in aging has been highlighted in recent times due to its association with progeroid or premature aging syndromes which is classified as a type of laminopathy. Progeria caused by accelerated accumulation of lamin A Δ50 or progerin occurs due to a mutation in this LMNA gene leading to defects in post translational modification of lamin A. One of the most common and severe symptoms of progeroid laminopathy is accelerated cellular senescence or aging along with bone resorption, muscle weakness, lipodystrophy and cardiovascular disorders. On the other hand, progerin accumulation and telomere dysfunction merge as common traits in the process of chronological aging. Two major hallmarks of physiological aging in humans include loss of genomic integrity and telomere attrition which can result from defective laminar organization leading to deformed nuclear architecture and culminates into replicative senescence. This also adversely affects epigenetic landscape, mitochondrial dysfunction and several signalling pathways like DNA repair, mTOR, MAPK, TGFβ. In this review, we discuss the telomere-lamina interplay in the context of physiological aging and progeria.
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