Down-regulation of DNMT3b in PC3 cells effects locus-specific DNA methylation, and represses cellular growth and migration

<p>Abstract</p> <p>Background</p> <p>Aberrations in DNA methylation patterns promote changes in gene expression patterns and are invariably associated with neoplasia. DNA methylation is carried out and maintained by several DNA methyltransferases (DNMTs) among which DNMT1 functions as a maintenance methylase while DNMT3a and 3b serve as de novo enzymes. Although DNMT3b has been shown to preferentially target the methylation of DNA sequences residing in pericentric heterochromatin whether it is involved in gene specific methylation remains an open question. To address this issue, we have silenced the expression of DNMT3b in the prostate-derived PC3 cells through RNA interference and subsequently studied the accompanied cellular changes as well as the expression profiles of selected genes.</p> <p>Results</p> <p>Our results demonstrate that DNMT3b depletion results in increased apoptosis and reduced migration of PC3 cells compared to the untransfected control cells. Reduced DNMT3b expression resulted in hypomethylation of retinoblastoma (Rb), retinoic-acid receptor β (RAR-β), and adenomatous polyposis coli (APC) gene promoters, and also culminated in increased expression of <it>CDKN3 </it>and <it>cytochrome b5</it>. Although DNMT3b silenced cells were found to have reduced growth and migratory potential, there was no apparent changes in their invasive ability compared to the parental PC3 cell line.</p> <p>Conclusion</p> <p>Our findings reveal that DNMT3b preferentially targets certain gene promoters in PC3 cells and that its depletion significantly reduces growth and migration of PC3 cells.</p

Use of integrated clinical scenarios in neuroanatomy laboratory sessions a strategy to foster students’ learning

Objective: Combinations of various teaching strategies have been employed to enhance neuroanatomy teaching, to little success. Herein, we describe the use of integrated clinical scenarios in anatomy laboratory sessions as an educational strategy to improve the learning process for students undertaking neuroscience course. Methods: We assessed the perception and performance of students who utilised the integrated clinical resources in the neuroanatomy labs. We also compared the performance of the students on their neuroscience course with the performance of the previous year\u27s students (who did not use the clinical scenarios). Results: A total of (130) 88% of the registered student both male and female participated by filling out a questionnaire. A majority (80%) of students supported introducing clinical cases at this early stage and agreed that cases covered learning objectives well. Students (81%) preferred a decreased faculty participation and 68% strongly agreed that cases were well integrated with other disciplines and assisted critical thinking and conceptual understanding. Most of them (90%) approved using plastic models and pictures as resource-material. The average mark obtained for all block together between the two cohorts did not differ significantly, while student performance was significantly improved in neuroscience block of the cohort which had access to the integrated clinical scenarios. Conclusions: Collectively, or specifically designed neuroanatomy lab sessions provided students with an empowering experience to help them apply critical thinking and use their basic neuroscience knowledge to solve clinical problems

Frequent hypomethylation of PTGS2 gene promoter in human term placenta

Background: Gene expression profiles of several tumor suppressor genes are regulated by the methylation and demethylation of their promoters. Here, we aim to identify and quantify the methylation status of four tumor suppressor genes from placentas at term and compare them with the maternal white-blood-cells. Methods: In order to achieve this objective, DNA enriched from twenty placentas at term and maternal white blood cells was bisulfite-converted and amplified using quantitative real-time methyl-light polymerase chain reaction for the four-genes studied (RASSF1A, APC, RAR-beta, and PTGS2). Results: Among the four genes examined, RASSF1A, APC and RAR-beta promoter regions were hypermethylated in all the placental samples compared with maternal WBCs. Strikingly, PTGS2 was found to be hypomethylated in the placentas compared to the maternal cells. Conclusion: Since placental DNA represents fetal methylation profile and it is an established fact that there is certain amount of cell free circulating DNA in human plasma/serum, these data strongly suggest that hypermethylation of RASSF1A, APC and RAR-beta can be used as gender independent biomarkers to distinctly identify placental DNA in maternal blood. In addition, this is the first report which demonstrates hypomethylation of PTGS2 locus which may have important clinical implications e.g. placental abnormalities

Silencing of MBD1 and MeCP2 in prostate-cancer-derived PC3 cells produces differential gene expression profiles and cellular phenotypes

Alterations in genomic CpG methylation patterns have been found to be associated with cell transformation and neoplasia. Although it is recognized that methylation of CpG residues negatively regulates gene expression, how the various MBPs (methyl-binding proteins) contribute to this process remains elusive. To determine whether the two well characterized proteins MeCP2 (methyl-CpG-binding protein 2) and MBD1 (methyl-CpG-binding domain 1) have distinct or redundant functions, we employed RNAi (RNA interference) to silence their expression in the prostate cancer-derived PC3cell line, and subsequently compared cell growth, invasion and migration properties of these cell lines in addition to their respective mRNA-expressionprofiles. Cells devoid of MeCP2 proliferated more poorly compared with MBD1-deficient cells and the parental PC3 cells. Enhanced apoptosis was observed in MeCP2-deficient cells, whereas apoptosis in parental and MBD1-deficient cells appeared to be equivalent. Boyden chamber invasion and wound-healing migration assays showed that MBD1-silenced cells were both more invasive and migratory compared with MeCP2-silenced cells. Finally, gene chip microarray analyses showed striking differences in the mRNA-expression profiles obtained from MeCP2- and MBD1-depleted cellsrelative to each other as well as when compared with control cells. The results of the present study suggest that MeCP2 and MBD1 silencing appear to affect cellular processes independently in vivo and that discrete sets of genes involved in cellular proliferation, apoptosis, invasion and migration are targeted by each protein

Tackling the glial scar in spinal cord regeneration: new discoveries and future directions

Axonal regeneration and functional recovery are poor after spinal cord injury (SCI), typified by the formation of an injury scar. While this scar was traditionally believed to be primarily responsible for axonal regeneration failure, current knowledge takes a more holistic approach that considers the intrinsic growth capacity of axons. Targeting the SCI scar has also not reproducibly yielded nearly the same efficacy in animal models compared to these neuron-directed approaches. These results suggest that the major reason behind central nervous system (CNS) regeneration failure is not the injury scar but a failure to stimulate axon growth adequately. These findings raise questions about whether targeting neuroinflammation and glial scarring still constitute viable translational avenues. We provide a comprehensive review of the dual role of neuroinflammation and scarring after SCI and how future research can produce therapeutic strategies targeting the hurdles to axonal regeneration posed by these processes without compromising neuroprotection

Corrigendum: Reprogramming the immunosuppressive tumor microenvironment: exploiting angiogenesis and thrombosis to enhance immunotherapy

This review focuses on the immunosuppressive effects of tumor angiogenesis and coagulation on the tumor microenvironment (TME). We summarize previous research efforts leveraging these observations and targeting these processes to enhance immunotherapy outcomes. Clinical trials have documented improved outcomes when combining anti-angiogenic agents and immunotherapy. However, their overall survival benefit over conventional therapy remains limited and certain tumors exhibit poor response to anti-angiogenic therapy. Additionally, whilst preclinical studies have shown several components of the tumor coagulome to curb effective anti-tumor immune responses, the clinical studies reporting combinations of anticoagulants with immunotherapies have demonstrated variable treatment outcomes. By reviewing the current state of the literature on this topic, we address the key questions and future directions in the field, the answers of which are crucial for developing effective strategies to reprogram the TME in order to further the field of cancer immunotherapy

Reprogramming the immunosuppressive tumor microenvironment: exploiting angiogenesis and thrombosis to enhance immunotherapy

This review focuses on the immunosuppressive effects of tumor angiogenesis and coagulation on the tumor microenvironment (TME). We summarize previous research efforts leveraging these observations and targeting these processes to enhance immunotherapy outcomes. Clinical trials have documented improved outcomes when combining anti-angiogenic agents and immunotherapy. However, their overall survival benefit over conventional therapy remains limited and certain tumors exhibit poor response to anti-angiogenic therapy. Additionally, whilst preclinical studies have shown several components of the tumor coagulome to curb effective anti-tumor immune responses, the clinical studies reporting combinations of anticoagulants with immunotherapies have demonstrated variable treatment outcomes. By reviewing the current state of the literature on this topic, we address the key questions and future directions in the field, the answers of which are crucial for developing effective strategies to reprogram the TME in order to further the field of cancer immunotherapy

Urinary biomarkers for lupus nephritis: a systems biology approach

Systemic lupus erythematosus (SLE) is the prototypical systemic autoimmune disorder. Kidney involvement, termed lupus nephritis (LN), is seen in 40–60% of patients with systemic lupus erythematosus (SLE). After the diagnosis, serial measurement of proteinuria is the most common method of monitoring treatment response and progression. However, present treatments for LN—corticosteroids and immunosuppressants—target inflammation, not proteinuria. Furthermore, subclinical renal inflammation can persist despite improving proteinuria. Serial kidney biopsies—the gold standard for disease monitoring—are also not feasible due to their inherent risk of complications. Biomarkers that reflect the underlying renal inflammatory process and better predict LN progression and treatment response are urgently needed. Urinary biomarkers are particularly relevant as they can be measured non-invasively and may better reflect the compartmentalized renal response in LN, unlike serum studies that are non-specific to the kidney. The past decade has overseen a boom in applying cutting-edge technologies to dissect the pathogenesis of diseases at the molecular and cellular levels. Using these technologies in LN is beginning to reveal novel disease biomarkers and therapeutic targets for LN, potentially improving patient outcomes if successfully translated to clinical practice

Cerebrospinal fluid microRNAs as potential biomarkers in Alzheimer’s disease

Alzheimer’s disease (AD) is the leading form of dementia worldwide, but its early detection and diagnosis remain a challenge. MicroRNAs (miRNAs) are a group of small endogenous RNA molecules that regulate mRNA expression. Recent evidence suggests miRNAs play an important role in the five major hallmarks of AD pathophysiology: amyloidogenesis, tauopathy, neuroinflammation, synaptic dysfunction, and neuronal death. Compared to traditional biomarkers of AD, miRNAs display a greater degree of stability in cerebrospinal fluid. Moreover, aberrant changes in miRNA expression can be measured over time to monitor and guide patient treatment. Specific miRNA profiles and combinations may also be used to distinguish AD subjects from normal controls and other causes of dementia. Because of these properties, miRNAs are now being considered as promising and potential biomarkers of AD. This review comprehensively summarizes the diagnostic potential and regulatory roles miRNAs play in AD

Unraveling the epigenetic fabric of type 2 diabetes mellitus: pathogenic mechanisms and therapeutic implications

Type 2 diabetes mellitus (T2DM) is a rapidly escalating global health concern, with its prevalence projected to increase significantly in the near future. This review delves into the intricate role of epigenetic modifications - including DNA methylation, histone acetylation, and micro-ribonucleic acid (miRNA) expression - in the pathogenesis and progression of T2DM. We critically examine how these epigenetic changes contribute to the onset and exacerbation of T2DM by influencing key pathogenic processes such as obesity, insulin resistance, β-cell dysfunction, cellular senescence, and mitochondrial dysfunction. Furthermore, we explore the involvement of epigenetic dysregulation in T2DM-associated complications, including diabetic retinopathy, atherosclerosis, neuropathy, and cardiomyopathy. This review highlights recent studies that underscore the diagnostic and therapeutic potential of targeting epigenetic modifications in T2DM. We also provide an overview of the impact of lifestyle factors such as exercise and diet on the epigenetic landscape of T2DM, underscoring their relevance in disease management. Our synthesis of the current literature aims to illuminate the complex epigenetic underpinnings of T2DM, offering insights into novel preventative and therapeutic strategies that could revolutionize its management