ResearchPad - cellular-molecular-and-developmental-genetics https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Combined use of mitochondrial and nuclear genetic markers further reveal immature marine turtle hybrids along the South Western Atlantic]]> https://www.researchpad.co/article/Ne2fdf233-7780-4122-85eb-65f194ef3974 Marine turtle hybridization is usually sporadic and involves reports of only a few individuals; however, Brazilian populations have high hybridization rates. Here we investigated the presence of hybrids in morphologically identified immature hawksbills (Eretmochelys imbricata) along the South Western Atlantic (SWA). We sequenced one mitochondrial (D-Loop) and three nuclear DNA (RAG1, RAG2, and CMOS) markers to better understand the patterns and characteristics of hybrids. We identified 22 hybrids (n = 270), 11 of them at the extreme South of the SWA. Uruguay had the highest hybrid frequency in the SWA (~37.5%) followed by southern Brazil with 30%. These are common areas for loggerheads (Caretta caretta) but uncommon for hawksbills, and these hybrids may be adopting the behavior of loggerheads. By analyzing nuclear markers, we can infer that 50% of the sampled hybrids are first generation (F1) and 36% are the result of backcrosses between hybrids and pure E. imbricata (> F1). We also report for the first time immature E. imbricata x Lepidochelys olivacea hybrids at the Brazilian coast. Considering the high frequency of hybrids in the SWA, continuous monitoring should be performed to assess the fitness, genetic integrity, and extent of changes in the gene pools of involved populations.

]]>
<![CDATA[Role of the MAPK pathway in human lung epithelial-like A549 cells apoptosis induced by paraquat]]> https://www.researchpad.co/article/N44f347c1-caa4-47d7-85ae-20a476d40b39 This study aims to investigate the value of mitogen-activated protein kinases (MAPKs) for paraquat (PQ)-induced apoptosis in human lung epithelial-like A549 cells and the specific mechanism. A549 cell apoptosis were induced by PQ. These cells were divided into six groups: control group (cells were cultured in RPMI-1640 medium); SP600125 group (cells were preconditioned with SP600125); SB203580 group (cells were preconditioned with SB203580); PQ group (cells were treated with PQ); SP600125+PQ group (cells were preconditioned with SP600125 following PQ); SB203580+PQ group (cells were preconditioned with SB203580 following PQ). The cell survival rate, apoptosis rate, and activities of caspase-3 and -9 were detected. When compared with the control group, both SP600125 and SB203580 groups had no significant difference in the detected indicators. When compared with PQ group, the cells in both SP600125+PQ group and SB203580+PQ group had significantly increased viability and level of anti-apoptotic protein Bcl-2; and had decreased apoptotic rates, decreased levels of caspase-3 and -9, and decreased level of pro-apoptotic protein Bax. The ratio of p-JNK/JNK protein expression in the SP600125+PQ group significantly decreased, while the ratio of the p-P38/P38 protein expression in the SB203580+PQ group decreased. PQ induced A549 cell apoptosis through the MAPKs pathway.

]]>
<![CDATA[PACT/PRKRA and p53 regulate transcriptional activity of DMRT1]]> https://www.researchpad.co/article/N58863942-2148-4851-b4d8-d9b58ffe339c The transcription factor DMRT1 (doublesex and mab-3 related transcription factor) has two distinct functions, somatic-cell masculinization and germ-cell development in some vertebrate species, including mouse and the African clawed frog Xenopus laevis. However, its transcriptional regulation remains unclear. We tried to identify DMRT1-interacting proteins from X. laevis testes by immunoprecipitation with an anti-DMRT1 antibody and MS/MS analysis, and selected three proteins, including PACT/PRKRA (Interferon-inducible double-stranded RNA dependent protein kinase activator A) derived from testes. Next, we examined the effects of PACT/PRKRA and/or p53 on the transcriptional activity of DMRT1. In transfected 293T cells, PACT/PRKRA and p53 significantly enhanced and repressed DMRT1-driven luciferase activity, respectively. We also observed that the enhanced activity by PACT/PRKRA was strongly attenuated by p53. Moreover, in situ hybridization analysis of Pact/Prkra mRNA in tadpole gonads indicated high expression in female and male germline stem cells. Taken together, these findings suggest that PACT/PRKRA and p53 might positively and negatively regulate the activity of DMRT1, respectively, for germline stem cell fate.

]]>
<![CDATA[Combining canine mesenchymal stromal cells and hyaluronic acid for cartilage repair]]> https://www.researchpad.co/article/Nf7ef01e7-764c-4c58-b13c-a5787e0ce074 Cell therapy and tissue engineering have been intensively researched for repair of articular cartilage. In this study, we investigated the chondrogenic potential of canine adipose-derived mesenchymal stromal cells (ASCs) combined to high molecular weight hyaluronic acid (HA) in vitro, and their therapeutic effect in dogs with chronic osteoarthritis (OA) associated with bilateral hip dysplasia. Canine ASCs were characterized after conventional 2D culture or 3D culture in HA, showing adequate immunophenotype, proliferation and trilineage differentiation, as well as chondrogenesis after cultivation in HA. ASC/HA constructs were used to treat 12 dogs with OA, sequentially assigned to control, ASC and ASC/HA groups. Animals were examined for clinical, orthopedic and radiological parameters. Lameness at walk and pain on manipulation were reduced in the ASC group and mainly in the ASC/HA group. Range of motion and detection of crepitus on hip rotation and abduction improved similarly in all groups. For articular edema, muscle atrophy, Norberg angle values and radiographic analyses, there were no variations throughout the period. These results indicate that ASC/HA constructs are safe and may be an effective therapeutic tool in treating canine chronic osteoarthritis, which should be confirmed with larger studies and additional clinical parameters.

]]>
<![CDATA[The perturbed expression of m6A in parthenogenetic mouse embryos]]> https://www.researchpad.co/article/N95e5aa1a-ae10-4e92-b4ce-6dc2974ae772

Abstract

Parthenogenetically activated oocytes cannot develop to term in mammals owing to abnormal epigenetic modifications. Methylation of the N6 position of adenosine (m6A) is a post-transcriptional epigenetic modification of RNA. To investigate the role of m6A methylation in parthenogenetic (PA) embryonic development, we analyzed METTL3, METTL14, FTO, ALKBH5, YTHDF2, IGF2BP1, and IGF2BP2 expression by quantitative real-time PCR. These genes were found dynamically expressed during the 2-cell, 4-cell, 8-cell, and blastocyst stages of the embryo. Compared to normally fertilized embryos, the expression of these genes was perturbed in PA embryos, especially at the 8-cell stage. Furthermore, immunofluorescence was used to detect m6A expression. The results demonstrated that m6A expression decreased in the 2-cell stage, whereas it increased in the 8-cell stage of PA embryos. Taken together, these results suggest that the expression of RNA methylation-related genes was perturbed, leading to abnormal m6A modification during early development in PA embryos.

]]>
<![CDATA[Individual expression features of GPX2, NQO1 and SQSTM1 transcript variants induced by hydrogen peroxide treatment in HeLa cells]]> https://www.researchpad.co/article/5b409c04463d7e5a04eefaf6

Abstract

Pathway activity assessment-based approaches are becoming highly influential in various fields of biology and medicine. However, these approaches mostly rely on analysis of mRNA expression, and total mRNA from a given locus is measured in the majority of cases. Notably, a significant portion of protein-coding genes produces more than one transcript. This biological fact is responsible for significant noise when changes in total mRNA transcription of a single gene are analyzed. The NFE2L2/AP-1 pathway is an attractive target for biomedical applications. To date, there is a lack of data regarding the agreement in expression of even classical target genes of this pathway. In the present paper we analyzed whether transcript variants of GPX2, NQO1 and SQSTM1 were characterized by individual features of expression when HeLa cells were exposed to pro-oxidative stimulation with hydrogen peroxide. We found that all the transcripts (10 in total) appeared to be significantly individually regulated under the conditions tested. We conclude that individual transcripts, rather than total mRNA, are best markers of pathway activation. We also discuss here some biological roles of individual transcript regulation.

]]>
<![CDATA[Stable silencing of β-lactoglobulin (BLG) gene by lentivirus-mediated RNAi in goat fetal fibroblasts]]> https://www.researchpad.co/article/5ac64495463d7e310640cd10

β-lactoglobulin (BLG), a dominant allergen in goat milk, is difficult to remove by traditional biochemical methods. Its elimination from goat milk by genetic modification therefore poses a major challenge for modern goat breeders. A shRNA targeting BLG mRNA with high interference efficiency was identified, with which lentiviral vectors were used for mediating stable shRNA interference in goat-fetal fibroblast cells. Apart from high efficiency in the knockdown of BLG expression in these cells, lentivector-mediated RNAi manifested stable integration into the goat genome itself. Consequently, an in vitro model for goat BLG-content control was compiled, and a goat-cell line for accompanying transgenetic goat production created.

]]>