1,721,017 research outputs found
Discovering new lncRNAs in Zebrafish: characterization of LOC100535512 in the developing and adult central nervous system
No full textCYYR1: discovery of a new role in muscle and heart development trought generation of null mutant
No full textCYYR1 gene involved in zebrafish skeletal muscle development and neuromasts differentiation: potential role in rhabdomyosarcoma disease
No full textUp-regulation of LINC00520 in in vitro models of Parkinson's disease: searching for a physiological role and dysregulation in neurodegeneration
No full textThe central nervous system (CNS) expresses a large number of long non-coding RNAs (lncRNAs), non-protein coding transcripts longer than 200 nucleotides, which are important for numerous physiological processes like brain development and neural functioning. Notably, their dysregulation has been demonstrated in several neurodegenerative disorders, including Parkinson's disease (PD). The hallmark of PD is the depletion of dopaminergic neurons in the substantia nigra pars compacta (SNpc) and the accumulation of cytoplasmic inclusion of a-synuclein (Lewy bodies). The neurotoxic phenotype of activated microglia contributes to PD pathogenesis by worsening inflammatory processes and exposing neurons to oxidative stress. Interestingly, we identified a disrupted expression of some lncRNAs through a meta-analysis by comparing transcriptomic data of PD patients vs control brain samples. In particular, LINC00520, which had not previously been connected to PD, showed a marked up-regulation. Our aim is to validate the results of the meta-analysis and characterize its pathophysiological role in different in vitro models. We looked at LINC00520 expression in the human neuroblastoma SH-SY5Y cells, treated with 6-OHDA to elicit an oxidative stress response, and in human monocytic THP-1 cells differentiated towards microglial-like fate with PMA and activated with LPS. Then, we included the human microglia cell line HMC3 that had been exposed to 6-OHDA or Rotenone as well as IFNg-boosted by glucose to increase the NF-kB inflammatory pathway. Our results showed a significant upregulation of LINC00520 in all in vitro PD models. Moreover, preliminary results of RNAi assays suggested that LINC00520 was involved in oxidative stress-related pathways. We also identified a potential LINC00520 ortholog in zebrafish, ZFLNCG09760, whose sequencing and functional characterization are still ongoing
Sex-Specific Transcriptome Differences in Human Adipose Mesenchymal Stem Cells
Full text linkIn humans, sexual dimorphism can manifest in many ways and it is widely studied
in several knowledge fields. It is increasing the evidence that also cells differ according to sex,
a correlation still little studied and poorly considered when cells are used in scientific research.
Specifically, our interest is on the sex-related dimorphism on the human mesenchymal stem cells
(hMSCs) transcriptome. A systematic meta-analysis of hMSC microarrays was performed by using
the Transcriptome Mapper (TRAM) software. This bioinformatic tool was used to integrate and
normalize datasets from multiple sources and allowed us to highlight chromosomal segments and
genes differerently expressed in hMSCs derived from adipose tissue (hADSCs) of male and female
donors. Chromosomal segments and differentially expressed genes in male and female hADSCs
resulted to be related to several processes as inflammation, adipogenic and neurogenic dierentiation
and cell communication. Obtained results lead us to hypothesize that the donor sex of hADSCs
is a variable influencing a wide range of stem cell biologic processes. We believe that it should be
considered in biologic research and stem cell therapy
Transcriptomic and morphological alterations in hTNPO3 MUT-microinjected Zebrafish embryos modelling Limb Girdle Muscular Dystrophy D2 in vivo
No full textTransportin-3, encoded by the TNPO3 gene, is physiologically involved in the translocation to the nucleus of many different proteins, such as SR proteins, which are associated with RNA metabolism. Mutations in the termination codon of the TNPO3 gene have been proved to cause Limb Girdle Muscular Dystrophy D2 (LGMD D2). More specifically, a single nucleotide deletion leads to the production of a protein with a 15 amino acid extension of its C-terminal domain. This project aimed to investigate the cause of LGMD D2, utilizing an in vivo approach, focusing on the role of TNPO3 on muscle development and to identify any potential molecular pathways linked to the disorder. This procedure involved the microinjection of mRNAs encoding the mutated form of human TNPO3 in Zebrafish embryos, to follow its effects on the myogenic processes during development. The subsequent analysis revealed changes in the gene expression profiles of Myogenic Regulatory Factors (MRFs) and muscle-specific proteins. The transcriptomic alterations are reflected on a phenotypical level by an aberrant organization of muscle fibres, as demonstrated by immunofluorescence staining. The methods we employed enabled us to gain an initial understanding of the role of Transportin-3 in muscle development and the pathogenic mechanism of its mutated form underlying LGMD D2, which has yet to be determined
CYYR1 gene and Hh-mediated myogenesis during zebrafish development: potential role in rhabdomyosarcoma disease
No full textCYYR1 (Cysteine/tyrosine-rich 1) cloned on HC21 defines a family of highly conserved
vertebrate-specific genes. The human locus is characterized by a multitranscript-system
including at least six alternative spliced isoforms. To date, the function of the CYYR1 product
is still unknown even if original results suggest its possible involvement in myogenesis
differentiation with a putative role in the tumorigenic process related to the Hh pathway.
Zebrafish cyyr1 orthologue is present in single copy and the predicted protein maintains
almost 58% of identity with human protein. By WISH approach, we show a cyyr1 broad
expression in central nervous system (CNS), somites and muscles during development. The
protein seems to localize in plasma and even nuclear membranes.
We perform cyyr1 knock-down with two different approaches: microinjection of morpholino
oligos targeting the ATG and the first splice-site of the transcript, and the generation of cyyr1
null fish through CRISPr/Cas9 technique. Defects in heart and muscle development with a
significant rescue in embryo co-injected with cyyr1 mRNA, were observed in morphants,
while cyyr1 mutants analyses confirmed morphological and molecular weakness in heart.
Dysregulation of Nodal and/or Hedgehog (Hh) pathways in zebrafish decreased cyyr1
expression and the injection of cyyr1 mRNA was able to partially rescue Hh-defective
phenotype in embryos. In order to verify a putative role of CYYR1 in the process caused by
dysfunction of cell differentiation we performed experiments in rhabdomyosarcoma cell
lines showing an inverse correlation between CYYR1 expression and the range of
differentiating capabilities of these cells. Interestingly, treatments with inhibitors of Hh allow
us to confirm a correlation between CYYR1 and this pathway
Exposure of Zebrafish Embryos to Urea Affects NOS1 Gene Expression in Neuronal Cells
Full text linkNitrogen-based fertilizers represent the most common fertilization tools, particularly used in crop food agriculture, despite the low cost-efficiency and the high negative environmental impact. At present, there is still inadequate information available about the effects of urea on human health; nevertheless, previous studies in animals observed that high urea concentration exposure can damage different tissues, including the brain. In several vertebrates, a crucial factor involved in neuronal cell formation is represented by the gas molecule, nitric oxide (NO), derived from the conversion of arginine to citrulline through the enzymatic activity of nitric oxide synthases (NOS). In zebrafish, three different isoforms of the NOS gene are known: nos1, nos2a, and nos2b. In the present study we show that nos1 represents the unique isoform with a stable high expression in the brain and spinal cord during all the embryonic stages of zebrafish development. Then, by using a specific transgenic zebrafish line, Tg(HuC:GFP), to mark neuronal cells, we observed nos1 to be specifically expressed in neurons. Interestingly, we observed that urea exposure at sub-lethal doses affected cell proliferation and the number of nos1-expressing cells, inducing apoptosis. Consistently, brain NO levels were observed to be reduced in urea-treated animals compared to untreated ones. This finding represents the first evidence that urea exposure affects the expression of a key gene involved in neuronal cell formation during embryonic development
Characterization of the lncRNA LINC00520 in the developing and adult central nervous system and its correlation to neurodegenerative processes
No full textUnderstanding the mechanisms underlying human brain development and its dysfunctions has always been challenging due to the involvement of complex and dynamically regulated processes. Over the last few years, long non-coding RNAs (lncRNAs) have emerged as new players in fundamental biological processes, specifically in CNS development, neuronal proliferation, differentiation, homeostasis and synaptic plasticity. Given their importance, even a slight alteration in their expression levels has been linked to a wide range of neurological disorders, including Parkinson's disease (PD). Notably, by comparing the transcriptome of brain samples from PD patients and controls, we were able to determine that the expression of several lncRNAs, previously unrelated to the disease, was altered. We focused on LINC00520, since it was one of the most significantly up-regulated. To better define its physiopathological role, we employed several human in vitro models, ranging from neuronal to microglial cell lines. More specifically, SH-SY5Y, THP-1 and
HMC3 cell lines were treated with different compounds (6-OHDA, Rotenone, LPS, IFN
glucose) to induce oxidative and inflammatory stress responses and mimic PD conditions. Additionally, we performed RNAi assays directed against LINC00520 using the same experimental settings. Upon treatments, silencing or the combination of the two, our analyses revealed fluctuations in the expression levels of specific stress-related markers depending on LINC00520 modulation. An accurate examination of genomic traits neighbouring LINC00520 led to the identification of a syntenic region on Zebrafish chromosome 17, harbouring LOC100535512 as a potential orthologue. Interestingly, gene expression studies on Zebrafish embryos and adult tissues suggest a meaningful role of the lncRNA, especially during the gastrulation period and in the adult CNS
Interplay between CYYR1 gene and Sonic hedgehog pathway: from zebrafish myogenesis to a potential role in rhabdomyosarcoma disease
No full textCYYR1 (Cysteine/tyrosine-rich 1) cloned on human chromosome 21 defines a family of highly conserved vertebrate-specific genes. The human locus is characterized by a multitranscript-system including at least six alternative spliced isoforms and one ncRNA gene overlapping CYYR1 in antisense orientation (CYYR1-AS1). To date, the function of the CYYR1 product is still unknown even if original results suggest its possible involvement in myogenesis differentiation with a putative role in the tumorigenic process related to the Hh pathway. Zebrafish cyyr1 orthologue is present in single copy and the predicted protein maintains almost 58% of identity with human protein. By WISH approach, we show a cyyr1 broad expression in central nervous system (CNS), somites and muscles during development. The protein seems to localize in plasma membrane and even in nuclear envelope. We perform cyyr1 knock-down with two different approaches: microinjection of morpholino oligos targeting the ATG and the first splice-site of the transcript, and the generation of cyyr1 null fish through CRISPr/Cas9 technique. Defects in heart and muscle development with a significant rescue in embryo co-injected with cyyr1 mRNA, were observed in morphants, while cyyr1 mutants analyses confirmed morphological and molecular weakness in heart. Dysregulation of Nodal and/or Hh pathways in zebrafish decreased cyyr1 expression and the injection of cyyr1 mRNA was able to partially rescue Hh-defective phenotype in embryos. In order to verify a putative role of CYYR1 in process caused by dysfunction of cell differentiation we performed experiments in rhabdomyosarcoma cell lines showing an inverse correlation between CYYR1 expression and the range of differentiating capabilities of these cells. Interestingly, treatments with inhibitors of Shh allow us to confirm a correlation between CYYR1 and this pathway
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