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1.
Cell Prolif ; : e12744, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31840352

RESUMO

OBJECTIVES: Mixed lineage leukaemia protein-1 (MLL1) mediates histone 3 lysine 4 (H3K4) trimethylation (me3) and plays vital roles during early embryonic development and hematopoiesis. In our previous study, we found its expression was positively correlated with embryonic myogenic ability in pigs, indicating its potential roles in mammalian muscle development. The present work aimed to explore the roles and regulation mechanisms of MLL1 in myogenesis. MATERIALS AND METHODS: The expression of MLL1 in C2C12 cells was experimentally manipulated using small interfering RNAs (siRNA). 5-ethynyl-2'-deoxyuridine (EdU) assay, cell cycle assay, immunofluorescence, qRT-PCR and Western blot were performed to assess myoblast proliferation and differentiation. Chromatin immunoprecipitation assay was conducted to detect H3K4me3 enrichment on myogenic factor 5 (Myf5) promoter. A cardiotoxin (CTX)-mediated muscle regeneration model was used to investigate the effects of MLL1 on myogenesis in vivo. RESULTS: MLL1 was highly expressed in proliferating C2C12 cells, and expression decreased after differentiation. Knocking down MLL1 suppressed myoblast proliferation and impaired myoblast differentiation. Furthermore, knockdown of MLL1 resulted in the arrest of cell cycle in G1 phase, with decreased expressions of Myf5 and Cyclin D1. Mechanically, MLL1 transcriptionally regulated Myf5 by mediating H3K4me3 on its promoter. In vivo data implied that MLL1 was required for Pax7-positive satellite cell proliferation and muscle repair. CONCLUSION: MLL1 facilitates proliferation of myoblasts and Pax7-positive satellite cells by epigenetically regulating Myf5 via mediating H3K4me3 on its promoter.

2.
Cell Death Differ ; 2019 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-31685980

RESUMO

Zinc finger protein 422 (Zfp422) is a widely expressed zinc finger protein that serves as a transcriptional factor to regulate downstream gene expression, but until now, little is known about its roles in myogenesis. We found here that Zfp422 plays a critical role in skeletal muscle development and regeneration. It highly expresses in mouse skeletal muscle during embryonic development. Specific knockout of Zfp422 in skeletal muscle impaired embryonic muscle formation. Satellite cell-specific Zfp422 deletion severely inhibited muscle regeneration. Myoblast differentiation and myotube formation were suppressed in Zfp422-deleted C2C12 cells, isolated primary myoblasts, and satellite cells. Chromatin Immunoprecipitation Sequencing (ChIP-Seq) revealed that Zfp422 regulated ephrin type-A receptor 7 (EphA7) expression by binding an upstream 169-bp DNA sequence, which was proved to be an enhancer of EphA7. Knocking EphA7 down in C2C12 cells or deleting Zfp422 in myoblasts will inhibit cell apoptosis which is required for myoblast differentiation. These results indicate that Zfp422 is essential for skeletal muscle differentiation and fusion, through regulating EphA7 expression to maintain proper apoptosis.

3.
BMC Genet ; 20(1): 72, 2019 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477002

RESUMO

BACKGROUND: Myogenic Differentiation 1 (MyoD) is a crucial master switch in regulating muscle-specific gene transcription. Forced expression of myoD is equipped to induce several cell lineages into myoblast, which then differentiate and fuse into myotube. Pig is one of the most significant livestock supplying meat, and has been classified into lean, fat and miniature pig breeds. However, the mechanisms underlying muscle mass variation among different pig breeds have remained unclear. Considering the important effect of MyoD on muscle development, it remains to be investigated whether the difference in muscle mass is caused by its single nucleotide polymorphisms (SNPs) which are the major differences among pig breeds at DNA level. RESULTS: In this study, we identified the locations of porcine myoD regulatory regions including proximal regulatory region (PRR), distal regulatory region (DRR), and core enhancer (CE) region. There are 8 SNPs in the regulatory regions and 6 SNPs in gene body region, which were identified from lean, fat and miniature pig populations. However, these SNPs have no effects on its temporal expression and transcriptional activity which might lead to the distinction in postnatal muscle mass. In addition, overexpression of myoD clones across from amphibious to mammals including xenopus tropicalis, chicken, mouse and pig whose gene identities vary from 68 to 84%, could promote myogenesis in NIH3T3 fibroblasts cells. CONCLUSIONS: These results proved that myoD nucleotide variations from different pig populations have no effect on muscle mass, suggesting that the function of myoD is highly conserved not only among different pig breeds, but also across different species. Thus, it would be futile to discover SNPs affecting muscle mass in pig populations with normal muscle development.

4.
J Anim Sci ; 97(5): 1967-1978, 2019 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-31222274

RESUMO

Pig is one of the major dietary protein sources for human consumption, from which muscle is the largest protein origin. However, molecular mechanisms concerning early porcine embryonic muscle development distinctions between pig breeds are still unclear. In this study, an integrated analysis of transcriptome and miRNAome was conducted using longissimus dorsi muscle of 4 early embryonic stages around the primary myofiber formation time (18-, 21-, 28-, and 35-d post coitus) from 2 pig breeds (Landrace [LR] and Wuzhishan [WZS]) differing in meat mass. The global miRNA/mRNA expression profile showed that WZS prepared for myogenic developmental processes earlier than LR. After identifying and analyzing the interaction network of top 100 up-/down-regulated miRNA and their target genes, we were able to find 3 gene clusters: chromatin modification-related (Chd2, H3f3a, Chd6, and Mll1), myogenesis-related (Pax3, Pbx1, Mef2a, and Znf423), and myosin component-related (Mylk, Myo5a, Mylk4, Myh9, and Mylk2) gene clusters. These genes may involve in miRNA-gene myogenic regulatory network that plays vital role in regulating distinct early porcine embryonic myogenic processes between LR and WZS. In summary, our study reveals an epigenetic-mediated myogenic regulatory axial that will help us to decipher molecular mechanisms concerning early porcine embryonic muscle development distinctions between pig breeds.


Assuntos
Epigênese Genética , Regulação da Expressão Gênica no Desenvolvimento/genética , MicroRNAs/genética , RNA Mensageiro/genética , Suínos/genética , Transcriptoma , Animais , Desenvolvimento Embrionário/genética , Feminino , Perfilação da Expressão Gênica/veterinária , Redes Reguladoras de Genes , Masculino , Desenvolvimento Muscular/genética , Especificidade da Espécie , Suínos/embriologia , Suínos/crescimento & desenvolvimento
5.
FASEB J ; 33(8): 9638-9655, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31145867

RESUMO

Here, we performed whole-genome bisulfite sequencing of longissimus dorsi muscle from Landrace and Wuzhishan (WZS) miniature pigs during 18, 21, and 28 d postcoitum. It was uncovered that in regulatory regions only around transcription start sites (TSSs), gene expression and methylation showed negative correlation, whereas in gene bodies, positive correlation occurred. Furthermore, earlier myogenic gene demethylation around TSSs and earlier hypermethylation of myogenic genes in gene bodies were considered to trigger their earlier expression in miniature pigs. Furthermore, by analyzing the methylation pattern of the myogenic differentiation 1(MyoD) promoter and distal enhancer, we found that earlier demethylation of the MyoD distal enhancer in WZSs contributes to its earlier expression. Moreover, DNA demethylase Tet1 was found to be involved in the demethylation of the myogenin promoter and promoted immortalized mouse myoblast cell line (C2C12) and porcine embryonic myogenic cell differentiation. This study reveals that earlier demethylation of myogenic genes contributes to precocious terminal differentiation of myoblasts in miniature pigs.-Zhang, X., Nie, Y., Cai, S., Ding, S., Fu, B., Wei, H., Chen, L., Liu, X., Liu, M., Yuan, R., Qiu, B., He, Z., Cong, P., Chen, Y., Mo, D. Earlier demethylation of myogenic genes contributes to embryonic precocious terminal differentiation of myoblasts in miniature pigs.

6.
Biochem Biophys Res Commun ; 503(2): 970-976, 2018 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-29932923

RESUMO

MicroRNAs are a class of highly conserved ∼20 nucleotides non-coding RNAs that post-transcriptionally regulate gene expression. Many miRNAs were studied in the development of skeletal muscle, such as miR-1, miR-206, and miR-133. In our previous study, miR-127-3p was found highly expressed in porcine fetal skeletal muscle, whereas the detailed functions of miR-127-3p in muscle development is still unclear. In this study, we detected that miR-127-3p also highly expressed in skeletal muscle, cardiac muscle of adult mice and proliferative C2C12 cell lines. Overexpression of miR-127-3p almost has no effects on differentiation of C2C12 cell lines. However, miR-127-3p significantly inhibited the cell proliferation of C2C12 cells. Moreover, we identified KMT5a as a target gene that was down-regulated in both mRNA and protein level when miR-127-3p mimics were introduced. Furthermore, KMT5a overexpression in miR-127-3p treated cells rescued the influence of miR-127-3p on C2C12 proliferation. In brief, our data reveals that miR-127-3p regulates the proliferation of myocytes through KMT5a.


Assuntos
Proliferação de Células , Regulação para Baixo , Histona-Lisina N-Metiltransferase/genética , MicroRNAs/genética , Células Musculares/citologia , Animais , Linhagem Celular , Células HEK293 , Humanos , Camundongos , Células Musculares/metabolismo , Regulação para Cima
7.
J Cell Physiol ; 233(3): 2537-2548, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28777437

RESUMO

The protein encoded by double sex and mab-3 related transcription factor 1 (Dmrt1) gene contains a double sex/mab-3 domain, which was considered as one of the most conservative structures in sex determination. However, its effect on spermatogenesis of dairy goat spermatogonial stem cells (SSCs) remains to be clarified. For the first time, the roles of Dmrt1 in spermatogenesis of livestock are highlighted. Here, we investigated the expression pattern of Dmrt1 in the testes of dairy goats. Dmrt1 primarily located in undifferentiated SSCs. Moreover, Dmrt1 enhanced differentiation and proliferation of mGSCs. On the contrary, the level of meiosis was down-regulated, as Dmrt1 determines whether SSCs undergo mitosis and spermatogonial differentiation or meiosis. In the busulfan-treated mice testes, Dmrt1 repair germ cell damage was emphasized as well. Our results exposed that Dmrt1 maintenance mGSCs in two ways: facilitating proliferation and self-renewal of SSCs; and reducing the inflammatory response caused by reproductive injury. These findings identify a central role for Dmrt1 in controlling population stability and injury restoring of SSCs.


Assuntos
Células-Tronco Germinativas Adultas/metabolismo , Diferenciação Celular , Proliferação de Células , Autorrenovação Celular , Indústria de Laticínios , Cabras/metabolismo , Espermatogênese , Fatores de Transcrição/metabolismo , Células-Tronco Germinativas Adultas/efeitos dos fármacos , Células-Tronco Germinativas Adultas/patologia , Animais , Bussulfano/toxicidade , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Autorrenovação Celular/efeitos dos fármacos , Células Cultivadas , Regulação da Expressão Gênica no Desenvolvimento , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Cabras/genética , Fator Estimulador de Colônias de Macrófagos/metabolismo , Masculino , Meiose , Mitose , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Transdução de Sinais , Espermatogênese/efeitos dos fármacos , Fatores de Transcrição/genética , Transfecção , Tretinoína/farmacologia
8.
Cell Prolif ; 49(4): 503-11, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27374854

RESUMO

OBJECTIVES: Previous studies have shown that adipose mesenchymal stem cells (AMSCs) share the potency of typical bone marrow mesenchymal stem cells (MSCs); however, there is little information concerning characteristics of canine AMSCs (CAMSCs); it has not previously been made clear whether CAMSCs would be able to differentiate into other cell types. MATERIALS AND METHODS: In this study, typical AMSC lines were established, and their characteristics including morphology, typical markers and differentiation potentiality were tested. RESULTS: The cells exhibited typical MSC morphology and were positive for CD90, CD44 and CD166, considered to be MSCs surface markers. They were negative for CD34 and CD45. The CAMSCs also exhibited embryonic stem cell (ESC) markers, including Oct4 and Sox2, at passage 2. In an appropriate microenvironment, CAMSCs differentiated into EBs and were able to produce cells of the three germ layers. These results indicate that established cells were putative adipocyte-derived MSCs, which also displayed properties of ESCs. Moreover, when the CAMSCs were induced by bone morphogenetic protein 4 (BMP4), they differentiated into PGC-like cells (PGCLCs) and male germ-like cells, which were positive for PR domain-containing 1 (Prdm1), PR domain-containing 14 (Prdm14), doublesex and mab-3 related transcription factor (Dmrt1), as well as for promyelocytic leukaemia zinc finger (Plzf). Quantitative real-time PCR (qRT-PCR) and western blotting analysis verified higher expression levels of these markers. CONCLUSION: This study provides an efficient approach to study germ cell development using CAMSCs.


Assuntos
Tecido Adiposo/citologia , Diferenciação Celular , Células Germinativas/citologia , Células-Tronco Mesenquimais/citologia , Adipogenia , Tecido Adiposo/metabolismo , Animais , Proteína Morfogenética Óssea 4/metabolismo , Ciclo Celular , Proliferação de Células , Separação Celular , Células Cultivadas , Condrogênese , Cães , Células Germinativas/metabolismo , Masculino , Células-Tronco Mesenquimais/metabolismo , Osteogênese
9.
J Phys Ther Sci ; 26(7): 1133-7, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25140112

RESUMO

[Purpose] The aim of the present study was to seek evidence for the effectiveness of Tai Chi for patients with knee osteoarthritis (KOA). [Subjects and Methods] Systematic searches were conducted of the China Journals Full-text Database, Pubmed, Medline, Science Direct-Online Journals and CINAHL for studies published between 2000 and 2012. Studies were evaluated based on following inclusion criteria: 1) design: randomized control, clinical trial; 2) subjects: patients with a knee osteoarthritis diagnosis; 3) intervention: exercise involving Tai Chi; 4) studies published in English or Chinese. [Results] Six randomized control studies involving Tai Chi and knee osteoarthritis were found. [Conclusion] Tai Chi was an effective way of relieving pain and improving physical function. Further randomized controlled trials with large sample sizes and long training period are needed to compare groups who perform Tai Chi training with other groups who undergo other forms of physical exercise in order to confirm the efficacy of Tai Chi.

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