Project description:PCD is a highly organised process that is involved in development and in an organisms response to biotic stresses (toxins and avirulent pathogens) and abiotic stresses (such as temperature, water availability, etc.). It is a genetically regulated form of cellular suicide, however in plants the underlying process is poorly understood. Although PCD may occur in response to different stimuli; we believe once it is triggered, one core mechanism is responsible for the cellular demise. It is our aim to identify the elements of this mechanism. We will do this by expanding on the work of a previous user of GARNet's GeneChip microarray facility, Dr. Jodi Swidzinski. She utilised an Arabidopsis cell suspension system; performing microarray analysis on both senescing, and heat shock induced PCD samples. This resulted in data showing that a large number of genes were upregulated (or downregulated) in response to both treatments. We are working under the premise that some of the genes that were similarly regulated under both treatments must be core PCD genes. However because of the large amount of data generated it is difficult to choose appropriate candidate genes (with any degree of confidence that they are core genes) for further study. We propose to use a third PCD-inducing treatment, involving a mycotoxin, to generate another population of microarray results. The mycotoxin we intend to use is Fumonisin B1 (FB1). This is an extremely potent compound that induces PCD by disrupting ceramide synthesis. We have found Arabidopsis protoplasts to be much more sensitive than cells to the toxin at low concentrations. Protoplasts are treated with 20mM FB1 and RNA is extracted at time points when 0%, 20% and 40% of protoplasts have died. This RNA is then pooled. RNA from methanol treated protoplasts is used as a control. We intend for these RNA samples to be subjected to GeneChip microarray analysis. This would identify genes differentially regulated due to FB1 treatment, which would be interesting in itself. However, by combining this data with that from previous work by Swidzinski (2002) we will be able to decrease the pool of possible core genes further, and increase the chances of selecting an appropriate candidate gene. This will both improve upon existing work and add value to an existing GARNet data set. Experimenter name = Mark Diamond Experimenter phone = 017162251 Experimenter fax = 017161153 Experimenter address = Botany Department Experimenter address = University College Dublin Experimenter address = Belfield Experimenter zip/postal_code = Dublin 4 Experimenter country = Ireland Keywords: compound_treatment_design

Project description:PCD is a highly organised process that is involved in development and in an organisms response to biotic stresses (toxins and avirulent pathogens) and abiotic stresses (such as temperature, water availability, etc.). It is a genetically regulated form of cellular suicide, however in plants the underlying process is poorly understood. Although PCD may occur in response to different stimuli; we believe once it is triggered, one core mechanism is responsible for the cellular demise. It is our aim to identify the elements of this mechanism. We will do this by expanding on the work of a previous user of GARNet's GeneChip microarray facility, Dr. Jodi Swidzinski. She utilised an Arabidopsis cell suspension system; performing microarray analysis on both senescing, and heat shock induced PCD samples. This resulted in data showing that a large number of genes were upregulated (or downregulated) in response to both treatments. We are working under the premise that some of the genes that were similarly regulated under both treatments must be core PCD genes. However because of the large amount of data generated it is difficult to choose appropriate candidate genes (with any degree of confidence that they are core genes) for further study. We propose to use a third PCD-inducing treatment, involving a mycotoxin, to generate another population of microarray results. The mycotoxin we intend to use is Fumonisin B1 (FB1). This is an extremely potent compound that induces PCD by disrupting ceramide synthesis. We have found Arabidopsis protoplasts to be much more sensitive than cells to the toxin at low concentrations. Protoplasts are treated with 20mM FB1 and RNA is extracted at time points when 0%, 20% and 40% of protoplasts have died. This RNA is then pooled. RNA from methanol treated protoplasts is used as a control. We intend for these RNA samples to be subjected to GeneChip microarray analysis. This would identify genes differentially regulated due to FB1 treatment, which would be interesting in itself. However, by combining this data with that from previous work by Swidzinski (2002) we will be able to decrease the pool of possible core genes further, and increase the chances of selecting an appropriate candidate gene. This will both improve upon existing work and add value to an existing GARNet data set. Experimenter name = Mark Diamond; Experimenter phone = 017162251; Experimenter fax = 017161153; Experimenter address = Botany Department; Experimenter address = University College Dublin; Experimenter address = Belfield; Experimenter zip/postal_code = Dublin 4; Experimenter country = Ireland Experiment Overall Design: 8 samples were used in this experiment

Project description:Background: Atrial fibrillation (AF), a critical health and economic issue is common in patients with heart failure (HF). Meanwhile, HF is a leading complication of nonvalvular atrial fibrillation (NVAF), and the presence of both conditions is associated with worsen outcomes. Accumulating evidence has indicated that messenger RNA (mRNA), microRNA (miRNA) and circular RNA (circRNA) play vital roles in the occurrence and progression of HF and AF. However, the underlying molecular mechanism of HF with AF remains elusive. We aimed to investigate the role of circRNA/miRNA/mRNA regulatory network in HF with AF through High-throughput sequencing and bioinformatics analysis. Methods: Peripheral blood mononuclear cells (PBMCs) were obtained from 4 patients who had heart failure with atrial fibrillation (HF /AF group) and 4 matched healthy subjects. RNA sequencing was performed to get circRNAs and mRNAs. miRNAs were obtained through miRanda and HMDD databases. qRT-PCR was used to verify our data. R software was employed for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. The PPI network was completed using STRING. Immune infiltration analysis was performed using cibersort method. The ceRNA network of circRNA/miRNA/mRNA was constructed by Cytoscape. Results: Differential expression analysis showed that circRNAs (137 upregulated and 31 downregulated) and mRNAs (534 upregulated and 255 downregulated) were considered as significantly different between HF with AF group and control group. Immune infiltration analysis demonstrated that macrophages were obviously upregulated and mast cells were significantly downregulated in PBMCs of the HF/AF group. GO and KEGG analyses showed that HF with AF-related mRNAs were abundant in the pathway of programmed cell death (PCD) and myocardial remodeling. So we confirm that hsa_circ_0047870/hsa-miR-34a-5p/SMPD1, hsa_circ_0127340/hsa-miR-192-3p/FBXW5, hsa_circ_0002484/hsa-miR-26b-3p/PYCARD, hsa_circ_0047870/hsa-miR-20b-3p/HTRA2 and hsa_circ_0004027/hsa-miR-26a-5p/HIF1A ceRNA axes could promote the development of HF with AF through the PCD-related pathway. Conclusions: We suggested that the imbalance of immune cell composition in PBMCs may play an initial role in the progression of HF with AF. The PCD-related ceRNA regulatory network may be the potential therapeutic target in the occurrence and progression of HF with AF. The circRNAs could serve as novel diagnostic markers.

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PCD

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