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1.
Genet Mol Res ; 14(2): 4331-7, 2015 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-25966205

RESUMO

The mitochondrion is a crucial intracellular organelle responsible for regulating cellular energy metabolism, producing free radicals, initiating and executing the apoptotic pathways. Previous studies have shown that somatic mutations in mitochondrial DNA are associated with various tumors, which may be involved during carcinogenesis and tumor progression. To examine the mutation pattern in cancer, 625 reported somatic mutations in the mitochondrial DNA genome were analyzed. We found that, except for deletions and insertions, most somatic mutations were point mutations, accounting for 89.44% of somatic mutations. Transition was the predominant form of somatic mutation in the entire mitochondrial DNA genome, accounting for 87.12% of point mutations, most of which were homoplastic. Frequency statistics analysis of point mutations indicated that, except for 3 tRNA genes, the mutations were distributed on all resting genes and in the D-loop region, with the latter showing the highest frequency of somatic mutation (19.34%), followed by the tRNA leucine 2 gene and non-coding regions between base pairs 5892 and 5903, while 13 coding-region genes and 2 rRNA genes showed a relatively lower frequency of somatic point mutations. Nonsynonymous mutations and terminal amino acid changes were the primary point somatic mutations detected from 13 coding-region genes, which may cause mitochondrial dysfunction in cancer cells. We found that the somatic mutations may affect the mitochondrial DNA genome; the non-coding region should be examined to identify somatic mutations as potential diagnostic biomarkers for early detection of cancer.


Assuntos
Análise Mutacional de DNA , DNA Mitocondrial/genética , Genoma Mitocondrial/genética , Mitocôndrias/genética , Sequência de Bases/genética , Humanos , Mutação/genética
2.
Rev. bras. pesqui. méd. biol ; Braz. j. med. biol. res;42(6): 506-514, June 2009. ilus, tab, graf
Artigo em Inglês | LILACS | ID: lil-512771

RESUMO

Mesenchymal stem cells (MSCs) have been reported to secrete a variety of cytokines and growth factors acting as trophic suppliers, but little is known regarding the effects of conditioned medium (CM) of MSCs isolated from femurs and tibias of mouse on the artificial activation of mouse oocytes and on the developmental competence of the parthenotes. In the current study, we investigated the effect of CM on the events of mouse oocyte activation, namely oscillations of cytosolic calcium concentration ([Ca²+]i), meiosis resumption, pronucleus formation, and parthenogenetic development. The surface markers of MSCs were identified with a fluorescence-activated cell sorter. The dynamic changes of the spindle and formation of pronuclei were examined by laser-scanning confocal microscopy. Exposure of cumulus-oocyte complexes to CM for 40 min was optimal for inducing oocyte parthenogenetic activation and evoking [Ca²+]i oscillations similar to those evoked by sperm (95 vs 100 percent; P > 0.05). Parthenogenetically activated oocytes immediately treated with 7.5 µg/mL cytochalasin B (CB), which inhibited spindle rotation and second polar body extrusion, were mostly diploid (93 vs 6 percent, P < 0.01) while CB-untreated oocytes were mostly haploid (5 vs 83 percent, P < 0.01). Consequently, the blastocyst rate was higher in the CB-treated than in the CB-untreated oocytes. There was no significant difference in developmental rate between oocytes activated with CM and 7 percent ethanol (62 vs 62 percent, P > 0.05), but the developmental competence of the fertilized oocytes was superior to that of the parthenotes (88 vs 62 percent, P < 0.05). The present results demonstrate that CM can effectively activate mouse oocytes, as judged by the generation of [Ca²+]i oscillations, completion of meiosis and parthenogenetic development.


Assuntos
Animais , Masculino , Camundongos , Cálcio/metabolismo , Meios de Cultivo Condicionados/farmacologia , Citocalasina B/farmacologia , Células-Tronco Mesenquimais , Oócitos/efeitos dos fármacos , Partenogênese/efeitos dos fármacos , Microscopia Confocal , Oócitos/fisiologia , Partenogênese/fisiologia
3.
Braz J Med Biol Res ; 42(6): 506-14, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19448898

RESUMO

Mesenchymal stem cells (MSCs) have been reported to secrete a variety of cytokines and growth factors acting as trophic suppliers, but little is known regarding the effects of conditioned medium (CM) of MSCs isolated from femurs and tibias of mouse on the artificial activation of mouse oocytes and on the developmental competence of the parthenotes. In the current study, we investigated the effect of CM on the events of mouse oocyte activation, namely oscillations of cytosolic calcium concentration ([Ca(2)+]i), meiosis resumption, pronucleus formation, and parthenogenetic development. The surface markers of MSCs were identified with a fluorescence-activated cell sorter. The dynamic changes of the spindle and formation of pronuclei were examined by laser-scanning confocal microscopy. Exposure of cumulus-oocyte complexes to CM for 40 min was optimal for inducing oocyte parthenogenetic activation and evoking [Ca(2)+]i oscillations similar to those evoked by sperm (95 vs 100%; P > 0.05). Parthenogenetically activated oocytes immediately treated with 7.5 microg/mL cytochalasin B (CB), which inhibited spindle rotation and second polar body extrusion, were mostly diploid (93 vs 6%, P < 0.01) while CB-untreated oocytes were mostly haploid (5 vs 83%, P < 0.01). Consequently, the blastocyst rate was higher in the CB-treated than in the CB-untreated oocytes. There was no significant difference in developmental rate between oocytes activated with CM and 7% ethanol (62 vs 62%, P > 0.05), but the developmental competence of the fertilized oocytes was superior to that of the parthenotes (88 vs 62%, P < 0.05). The present results demonstrate that CM can effectively activate mouse oocytes, as judged by the generation of [Ca(2)+]i oscillations, completion of meiosis and parthenogenetic development.


Assuntos
Cálcio/metabolismo , Meios de Cultivo Condicionados/farmacologia , Citocalasina B/farmacologia , Células-Tronco Mesenquimais , Oócitos/efeitos dos fármacos , Partenogênese/efeitos dos fármacos , Animais , Masculino , Camundongos , Microscopia Confocal , Oócitos/fisiologia , Partenogênese/fisiologia
4.
Braz J Med Biol Res ; 41(11): 978-85, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19039379

RESUMO

Mesenchymal stem cells (MSCs) secrete a variety of cytokines and growth factors in addition to self-renewal and multiple forms of differentiation. Some of these secreted bioactive factors could improve meiotic maturation in vitro and subsequent embryo developmental potential. The aim of the present study was to determine whether in vitro maturation (IVM) of mouse oocyte with or without cumulus cells could be improved by contact with conditioned medium (CM) of MSCs as well as the efficiency of CM to support follicular growth and oocyte maturation in the ovarian organ of mice cultured on soft agar. The developmental potential of matured oocyte was assessed by blastocyst formation after in vitro fertilization (IVF). Germinal vesicle stage oocytes with or without cumulus cells were subjected to IVM in either CM, Dulbecco's modified Eagle's medium (DMEM), alpha-minimum essential medium (alpha-MEM) or human tubal fluid (HTF). Approximately 120 oocytes were studied for each medium. CM produced a higher maturation rate (91.2%) than DMEM (54.7%), alpha-MEM (63.5%) and HTF (27.1%). Moreover, CM improved embryo development to blastocyst stage significantly more than DMEM and HTF (85 vs 7% and 41.7%, respectively) but there was no significant difference compared with alpha-MEM (85 vs 80.3%). The behavior of cortical granules of IVM oocytes cultured in CM revealed cytoplasmic maturation. Moreover, CM also supported preantral follicles growth well in organotypic culture on soft agar resulting in the maturation of 60% of them to developmentally competent oocytes. The production of estrogen progressively increased approximately 1-fold every other day during organ culture, while a dramatic 10-fold increase in progesterone was observed 17 h after human chorionic gonadotropin stimulus at the end of culture. Thus, CM is an effective medium for preantral follicle growth, oocyte maturation, and sequential embryo development.


Assuntos
Meios de Cultivo Condicionados/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Oócitos/crescimento & desenvolvimento , Animais , Células do Cúmulo/citologia , Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário/fisiologia , Feminino , Fertilização in vitro , Meiose/fisiologia , Camundongos , Folículo Ovariano/crescimento & desenvolvimento , Gravidez
5.
Rev. bras. pesqui. méd. biol ; Braz. j. med. biol. res;41(11): 978-985, Nov. 2008. ilus, graf
Artigo em Inglês | LILACS | ID: lil-500368

RESUMO

Mesenchymal stem cells (MSCs) secrete a variety of cytokines and growth factors in addition to self-renewal and multiple forms of differentiation. Some of these secreted bioactive factors could improve meiotic maturation in vitro and subsequent embryo developmental potential. The aim of the present study was to determine whether in vitro maturation (IVM) of mouse oocyte with or without cumulus cells could be improved by contact with conditioned medium (CM) of MSCs as well as the efficiency of CM to support follicular growth and oocyte maturation in the ovarian organ of mice cultured on soft agar. The developmental potential of matured oocyte was assessed by blastocyst formation after in vitro fertilization (IVF). Germinal vesicle stage oocytes with or without cumulus cells were subjected to IVM in either CM, Dulbecco's modified Eagle's medium (DMEM), alpha-minimum essential medium (alpha-MEM) or human tubal fluid (HTF). Approximately 120 oocytes were studied for each medium. CM produced a higher maturation rate (91.2%) than DMEM (54.7%), alpha-MEM (63.5%) and HTF (27.1%). Moreover, CM improved embryo development to blastocyst stage significantly more than DMEM and HTF (85 vs 7% and 41.7%, respectively) but there was no significant difference compared with alpha-MEM (85 vs 80.3%). The behavior of cortical granules of IVM oocytes cultured in CM revealed cytoplasmic maturation. Moreover, CM also supported preantral follicles growth well in organotypic culture on soft agar resulting in the maturation of 60% of them to developmentally competent oocytes. The production of estrogen progressively increased approximately 1-fold every other day during organ culture, while a dramatic 10-fold increase in progesterone was observed 17 h after human chorionic gonadotropin stimulus at the end of culture. Thus, CM is an effective medium for preantral follicle growth, oocyte maturation, and sequential embryo development.


Assuntos
Animais , Feminino , Camundongos , Gravidez , Meios de Cultivo Condicionados/farmacologia , Células-Tronco Mesenquimais , Oócitos/crescimento & desenvolvimento , Células do Cúmulo/citologia , Embrião de Mamíferos/embriologia , Desenvolvimento Embrionário/fisiologia , Fertilização in vitro , Meiose/fisiologia , Folículo Ovariano/crescimento & desenvolvimento
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