Gynura segetum induces hepatic sinusoidal obstruction syndrome in mice by impairing autophagy

ABSTRACT Purpose: To investigate the underlying mechanism of hepatic sinusoidal obstruction syndrome (HSOS) induced by Gynura segetum by measuring autophagy in mouse models. Methods: The model group was administered G. segetum (30 g/kg/d) by gavage, while the normal control group was administered an equal volume of saline daily for five weeks. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), hepatic histopathological examinations, and Masson staining were performed to evaluate liver injury. Liver intercellular adhesion molecule-1 (ICAM-1) and P-selectin were evaluated by immunohistochemistry. Hepatocellular apoptosis was assessed using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Protein expression levels of autophagy markers were measured using Western blot analysis. Results: Gynura segetum was found to significantly induce liver injury compared with control mice, as evidenced by the increase of serum transaminases, a decrease in triglyceride levels, and histopathological changes in mice. Gynura segetum remarkably induced hepatocellular apoptosis and upregulated the expressions of ICAM-1 and P-selectin and also downregulated the protein expression levels of LC3, Atg12 and cytoplasmic polyadenylation element binding protein. Conclusions: Our results suggested that G. segetum induced liver injury with HSOS, and it was partly due to its ability to impair the autophagy pathway.

Controle molecular do acúmulo, tradução e degradação de mRNA em oócitos e embriões / Molecular control of accumulation, translation and degradation of mRNA in oocytes and embryos

A competência oocitária para suportar os posteriores estágios do desenvolvimento depende não somenteda correta segregação cromossômica e de transformações citoesqueléticas, mas, principalmente, da adequadatranscrição e estoque de mRNAs cruciais ao desenvolvimento e à viabilidade celular. Com a retomada dameiose, no entanto, embora o oócito mantenha a capacidade de tradução gênica e de síntese proteica, suaatividade transcricional é interrompida, sendo restabelecida apenas com a ativação do genoma embrionário.Deste modo, todo mRNA materno mobilizado durante maturação, expansão do cumulus, fertilização eembriogênese inicial deve ser sintetizado e estocado, em sua forma traducionalmente inativa, nos oócitosmantidos no estágio diplóteno da prófase I. Complexos mecanismos regulatórios, os quais envolvem apoliadenilação e a desadenilação do mRNA, estão implicados no processo de ativação e silenciamento tantotranscricional quanto traducional. Assim, dada à relevância do tema, esta revisão se propõe a abordar osprincipais eventos moleculares envolvidos no controle da expressão, do estoque, da tradução e da degradação detranscritos maternos imprescindíveis ao desenvolvimento oocitário e embrionário.(AU)

The oocyte competence to support the later stages of development depends not only on correctchromosome segregation and cytoskeletal changes, but mainly, the proper transcription and storage of mRNAscritical to the cellular development and viability. However, with the resumption of meiosis, although the oocytekept the ability of gene translation and protein synthesis, its transcriptional activity is interrupted and restoredonly with embryonic genome activation. Thus, all maternal mRNA mobilized during maturation, cumulusexpansion, fertilization and early embryogenesis must be synthesized and stored, in its translationally inactiveform, in oocytes kept in the diplotene stage of prophase I. Complex regulatory mechanisms which involvedeadenylation and polyadenylation of mRNA are involved in this process of activation and silencing astranscriptional as translational. So, due to the importance of the topic, this review proposes to discuss the mainmolecular events involved in the control of expression, storage, translation and degradation of maternaltranscript essential to the oocyte and embryo development.(AU)

Controle molecular do acúmulo, tradução e degradação de mRNA em oócitos e embriões / Molecular control of accumulation, translation and degradation of mRNA in oocytes and embryos

A competência oocitária para suportar os posteriores estágios do desenvolvimento depende não somenteda correta segregação cromossômica e de transformações citoesqueléticas, mas, principalmente, da adequadatranscrição e estoque de mRNAs cruciais ao desenvolvimento e à viabilidade celular. Com a retomada dameiose, no entanto, embora o oócito mantenha a capacidade de tradução gênica e de síntese proteica, suaatividade transcricional é interrompida, sendo restabelecida apenas com a ativação do genoma embrionário.Deste modo, todo mRNA materno mobilizado durante maturação, expansão do cumulus, fertilização eembriogênese inicial deve ser sintetizado e estocado, em sua forma traducionalmente inativa, nos oócitosmantidos no estágio diplóteno da prófase I. Complexos mecanismos regulatórios, os quais envolvem apoliadenilação e a desadenilação do mRNA, estão implicados no processo de ativação e silenciamento tantotranscricional quanto traducional. Assim, dada à relevância do tema, esta revisão se propõe a abordar osprincipais eventos moleculares envolvidos no controle da expressão, do estoque, da tradução e da degradação detranscritos maternos imprescindíveis ao desenvolvimento oocitário e embrionário.

The oocyte competence to support the later stages of development depends not only on correctchromosome segregation and cytoskeletal changes, but mainly, the proper transcription and storage of mRNAscritical to the cellular development and viability. However, with the resumption of meiosis, although the oocytekept the ability of gene translation and protein synthesis, its transcriptional activity is interrupted and restoredonly with embryonic genome activation. Thus, all maternal mRNA mobilized during maturation, cumulusexpansion, fertilization and early embryogenesis must be synthesized and stored, in its translationally inactiveform, in oocytes kept in the diplotene stage of prophase I. Complex regulatory mechanisms which involvedeadenylation and polyadenylation of mRNA are involved in this process of activation and silencing astranscriptional as translational. So, due to the importance of the topic, this review proposes to discuss the mainmolecular events involved in the control of expression, storage, translation and degradation of maternaltranscript essential to the oocyte and embryo development.

Molecular characterization of SINES integrated in endogenous retrovirus sequences from Leopardus geoffroyi and Puma concolor / Molecular characterization of SINES integrated in endogenous retrovirus sequences from Leopardus geoffroyi and Puma concolor

Background: Short interspersed nuclear elements (SINEs) are transposable elements which are transcribed by RNA polymerase III and widespread in mammalian genomes. Can-SINE is a family of SINE sequences specific to carnivores, predominant in their genomes and present in high copy numbers. The aim of this study was to characterize sequences of Can-SINEs integrated into sequences of endogenous retroviruses (ERVs) from Brazilian wild cats Puma concolor and Leopardus geoffroyi. Additionally, these sequences are considered from some perspectives of their evolution. Material, Methods and Results: By using PCR and sequencing to screen for ERVs within the genomes of L. geoffroyi and P. concolor, two new ERV sequences were amplified with an insertion around 220 nucleotides long, similar to published carnivore SINEs. The sequences were further identified and characterized using a combination of BLAST, BLAT searches and phylogenetic analyses. The results showed that SINE sequences integrated into the ERV from P. concolor (SINE_Pco) and L. geoffroyi (SINE_Lg) are lysine-tRNA derived. These sequences presented a typical RNA polymerase III-specific internal promoter sequence followed by a microsatellite region (TC)n and by an A/T-rich tail with the polyadenylation signal AATAAA. BLAST searches using the whole sequence of L. geoffroyi clone as query (ERV plus SINE) detected two sequences which

Background: Short interspersed nuclear elements (SINEs) are transposable elements which are transcribed by RNA polymerase III and widespread in mammalian genomes. Can-SINE is a family of SINE sequences specific to carnivores, predominant in their genomes and present in high copy numbers. The aim of this study was to characterize sequences of Can-SINEs integrated into sequences of endogenous retroviruses (ERVs) from Brazilian wild cats Puma concolor and Leopardus geoffroyi. Additionally, these sequences are considered from some perspectives of their evolution. Material, Methods and Results: By using PCR and sequencing to screen for ERVs within the genomes of L. geoffroyi and P. concolor, two new ERV sequences were amplified with an insertion around 220 nucleotides long, similar to published carnivore SINEs. The sequences were further identified and characterized using a combination of BLAST, BLAT searches and phylogenetic analyses. The results showed that SINE sequences integrated into the ERV from P. concolor (SINE_Pco) and L. geoffroyi (SINE_Lg) are lysine-tRNA derived. These sequences presented a typical RNA polymerase III-specific internal promoter sequence followed by a microsatellite region (TC)n and by an A/T-rich tail with the polyadenylation signal AATAAA. BLAST searches using the whole sequence of L. geoffroyi clone as query (ERV plus SINE) detected two sequences which

Molecular characterization of SINEs integrated in endogenous retrovirus sequences from Leopardus geoffroyi and Puma concolor

Background: Short interspersed nuclear elements (SINEs) are transposable elements which are transcribed by RNA polymerase III and widespread in mammalian genomes. Can-SINE is a family of SINE sequences specific to carnivores, predominant in their genomes and present in high copy numbers. The aim of this study was to characterize sequences of Can-SINEs integrated into sequences of endogenous retroviruses (ERVs) from Brazilian wild cats Puma concolor and Leopardus geoffroyi. Additionally, these sequences are considered from some perspectives of their evolution. Material, Methods and Results: By using PCR and sequencing to screen for ERVs within the genomes of L. geoffroyi and P. concolor, two new ERV sequences were amplified with an insertion around 220 nucleotides long, similar to published carnivore SINEs. The sequences were further identified and characterized using a combination of BLAST, BLAT searches and phylogenetic analyses. The results showed that SINE sequences integrated into the ERV from P. concolor (SINE_Pco) and L. geoffroyi (SINE_Lg) are lysine-tRNA derived. These sequences presented a typical RNA polymerase III-specific internal promoter sequence followed by a microsatellite region (TC)n and by an A/T-rich tail with the polyadenylation signal AATAAA. BLAST searches using the whole sequence of L. geoffroyi clone as query (ERV plus SINE) detected two sequences which were highly similar to the cougar (P. concolor) and the domestic cat. However, the SINE from Leopardus geoffroyi is not present in these related sequences. On the other hand, during searches using the whole sequence of the P. concolor clone as query, we found the same SINE insertion in a very similar ERV from domestic cat. All insertions occurred in the RT domain, but SINE_Lg was integrated in a distinct site when compared to SINE_Pco. Another interesting difference between these SINE sequences was that the statistics reported in BLAST searches recovered a much higher number of hits from the domestic cat genome using SINE_Lg as seed than in searches for sequences related to SINE_Pco. The phylogenetic tree based on the SINE fragment grouped these new SINE sequences with Can-SINEs from felids. Within this major clade SINE_Lg and SINE_Pco are related to different lineages of felids Can-SINEs. Discussion: In this study we showed that two different sequences from felid endogenous retrovirus harbor Can-SINE sequences. These insertions are not surprising taking account that ~11% of domestic cat genome is composed of SINE sequences and they are ubiquitous in felid genomes. Furthermore, the insertions of SINEs into the ERV sequences reported here are not unique events. However, they are curious insertions representing genomic fossils and a little piece of felid history. Based on the results of the phylogenetic analyses and position of the integration sites, we suggest that SINE_Lg and SINE_Pco represent independent integration events originated by derived copies from different progenitors. We hypothesized that SINE_Lg is a "young" integration due to the absence of highly similar ERVs from Puma concolor and Felis catus. This lineage may be recently active in felid genomes given that we found very similar MegaBLAST hits at EST database from domestic cat. Instead, SINE_Pco seems to be "old", sharing an identical insertion site to ERVs from domestic cat and its lineage could be inactive in felids considering that any MegaBLAST hits resulted from EST database searches. The latter suggests an integration event in an ancestor species at least 6.7 million years ago, which represents the split between puma and domestic cat lineages.

Molecular characterization of SINES integrated in endogenous retrovirus sequences from Leopardus geoffroyi and Puma concolor / Molecular characterization of SINES integrated in endogenous retrovirus sequences from Leopardus geoffroyi and Puma concolor

Background: Short interspersed nuclear elements (SINEs) are transposable elements which are transcribed by RNA polymerase III and widespread in mammalian genomes. Can-SINE is a family of SINE sequences specific to carnivores, predominant in their genomes and present in high copy numbers. The aim of this study was to characterize sequences of Can-SINEs integrated into sequences of endogenous retroviruses (ERVs) from Brazilian wild cats Puma concolor and Leopardus geoffroyi. Additionally, these sequences are considered from some perspectives of their evolution. Material, Methods and Results: By using PCR and sequencing to screen for ERVs within the genomes of L. geoffroyi and P. concolor, two new ERV sequences were amplified with an insertion around 220 nucleotides long, similar to published carnivore SINEs. The sequences were further identified and characterized using a combination of BLAST, BLAT searches and phylogenetic analyses. The results showed that SINE sequences integrated into the ERV from P. concolor (SINE_Pco) and L. geoffroyi (SINE_Lg) are lysine-tRNA derived. These sequences presented a typical RNA polymerase III-specific internal promoter sequence followed by a microsatellite region (TC)n and by an A/T-rich tail with the polyadenylation signal AATAAA. BLAST searches using the whole sequence of L. geoffroyi clone as query (ERV plus SINE) detected two sequences which

Background: Short interspersed nuclear elements (SINEs) are transposable elements which are transcribed by RNA polymerase III and widespread in mammalian genomes. Can-SINE is a family of SINE sequences specific to carnivores, predominant in their genomes and present in high copy numbers. The aim of this study was to characterize sequences of Can-SINEs integrated into sequences of endogenous retroviruses (ERVs) from Brazilian wild cats Puma concolor and Leopardus geoffroyi. Additionally, these sequences are considered from some perspectives of their evolution. Material, Methods and Results: By using PCR and sequencing to screen for ERVs within the genomes of L. geoffroyi and P. concolor, two new ERV sequences were amplified with an insertion around 220 nucleotides long, similar to published carnivore SINEs. The sequences were further identified and characterized using a combination of BLAST, BLAT searches and phylogenetic analyses. The results showed that SINE sequences integrated into the ERV from P. concolor (SINE_Pco) and L. geoffroyi (SINE_Lg) are lysine-tRNA derived. These sequences presented a typical RNA polymerase III-specific internal promoter sequence followed by a microsatellite region (TC)n and by an A/T-rich tail with the polyadenylation signal AATAAA. BLAST searches using the whole sequence of L. geoffroyi clone as query (ERV plus SINE) detected two sequences which