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1.
Stem Cell Res ; 64: 102915, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36166871

RESUMO

Induced pluripotent stem cells (iPSCs) obtained from genetically characterized patients benefit the biological study of bipolar disorder (BD). Here, we present iPSC lines from three-generation patients with BD and recurrent depressive disorder (RDD) and a healthy control sibling in a family. All patients shared the specified haplotype in the 1p36-35, previously reported as the susceptibility locus of mood disorders. iPSCs were generated with the reprogramming factors OTC3/4, l-MYC, LIN28, SOX2, KLF4, and p53 shRNA through non-integrated episomal vectors. All iPSC lines strongly expressed pluripotency markers and proved the ability to differentiate into three germ lineages in vitro.


Assuntos
Transtorno Bipolar , Transtorno Depressivo , Células-Tronco Pluripotentes Induzidas , Humanos , Irmãos , Haplótipos/genética , Transtorno Bipolar/genética , RNA Interferente Pequeno , Proteína Supressora de Tumor p53
2.
Brain ; 136(Pt 3): 828-43, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23364351

RESUMO

A multiplex analysis for profiling the expression of candidate genes along with epigenetic modification may lead to a better understanding of the complex machinery of neuropathic pain. In the present study, we found that partial sciatic nerve ligation most remarkably increased the expression of monocyte chemotactic protein 3 (MCP-3, known as CCL7) a total of 33 541 genes in the spinal cord, which lasted for 4 weeks. This increase in MCP-3 gene transcription was accompanied by the decreased trimethylation of histone H3 at Lys27 at the MCP-3 promoter. The increased MCP-3 expression associated with its epigenetic modification observed in the spinal cord was almost abolished in interleukin 6 knockout mice with partial sciatic nerve ligation. Consistent with these findings, a single intrathecal injection of recombinant proteins of interleukin 6 significantly increased MCP-3 messenger RNA with a decrease in the level of Lys27 trimethylation of histone H3 at the MCP-3 promoter in the spinal cord of mice. Furthermore, deletion of the C-C chemokine receptor type 2 (CCR2) gene, which encodes a receptor for MCP-3, failed to affect the acceleration of MCP-3 expression in the spinal cord after partial sciatic nerve ligation. A robust increase in MCP-3 protein, which lasted for up to 2 weeks after surgery, in the dorsal horn of the spinal cord of mice with partial sciatic nerve ligation was seen mostly in astrocytes, but not microglia or neurons. On the other hand, the increases in both microglia and astrocytes in the spinal cord by partial sciatic nerve ligation were mostly abolished in interleukin 6 knockout mice. Moreover, this increase in microglia was almost abolished by CCR2 gene deletion, whereas the increase in astrocytes was not affected in nerve-ligated mice that lacked the CCR2 gene. We also found that either in vivo or in vitro treatment with MCP-3 caused robust microglia activation. Under these conditions, intrathecal administration of MCP-3 antibody suppressed the increase in microglia within the mouse spinal cord and neuropathic pain-like behaviours after nerve injury. With the use of a functional magnetic resonance imaging analysis, we demonstrated that a single intrathecal injection of MCP-3 induced dramatic increases in signal intensity in pain-related brain regions. These findings suggest that increased MCP-3 expression associated with interleukin 6 dependent epigenetic modification at the MCP-3 promoter after nerve injury, mostly in spinal astrocytes, may serve to facilitate astrocyte-microglia interaction in the spinal cord and could play a critical role in the neuropathic pain-like state.


Assuntos
Comunicação Celular/fisiologia , Quimiocina CCL7/biossíntese , Epigênese Genética/fisiologia , Interleucina-6/metabolismo , Neuralgia/fisiopatologia , Ativação Transcricional/fisiologia , Animais , Astrócitos/metabolismo , Axotomia , Western Blotting , Quimiocina CCL7/genética , Imunoprecipitação da Cromatina , Dor Crônica/genética , Dor Crônica/metabolismo , Dor Crônica/fisiopatologia , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatologia , Imuno-Histoquímica , Interleucina-6/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Camundongos Knockout , Análise em Microsséries , Microglia/metabolismo , Neuralgia/genética , Neuralgia/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Nervo Isquiático/lesões , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia
3.
Stem Cells Dev ; 20(9): 1587-99, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21126169

RESUMO

Nonhuman primate embryonic stem (ES) cells have vast promise for preclinical studies. Genetic modification in nonhuman primate ES cells is an essential technique for maximizing the potential of these cells. The common marmoset (Callithrix jacchus), a nonhuman primate, is expected to be a useful transgenic model for preclinical studies. However, genetic modification in common marmoset ES (cmES) cells has not yet been adequately developed. To establish efficient and stable genetic modifications in cmES cells, we inserted the enhanced green fluorescent protein (EGFP) gene with heterotypic lox sites into the ß-actin (ACTB) locus of the cmES cells using gene targeting. The resulting knock-in ES cells expressed EGFP ubiquitously under the control of the endogenous ACTB promoter. Using inserted heterotypic lox sites, we demonstrated Cre recombinase-mediated cassette exchange (RMCE) and successfully established a monomeric red fluorescent protein (mRFP) knock-in cmES cell line. Further, a herpes simplex virus-thymidine kinase (HSV-tk) knock-in cmES cell line was established using RMCE. The growth of tumor cells originating from the cell line was significantly suppressed by the administration of ganciclovir. Therefore, the HSV-tk/ganciclovir system is promising as a safeguard for stem cell therapy. The stable and ubiquitous expression of EGFP before RMCE enables cell fate to be tracked when the cells are transplanted into an animal. Moreover, the creation of a transgene acceptor locus for site-specific transgenesis will be a powerful tool, similar to the ROSA26 locus in mice.


Assuntos
Actinas/genética , Callithrix/genética , Células-Tronco Embrionárias/metabolismo , Marcação de Genes , Técnicas de Transferência de Genes , Proteínas Recombinantes de Fusão/genética , Actinas/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Diferenciação Celular , Células Cultivadas , Clonagem Molecular , Técnicas de Cocultura , Células-Tronco Embrionárias/transplante , Loci Gênicos , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Cariótipo , Camundongos , Camundongos Endogâmicos NOD , Dados de Sequência Molecular , Mutagênese Insercional , Moléculas de Adesão de Célula Nervosa/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Teratoma/metabolismo , Teratoma/patologia , Teratoma/prevenção & controle , Vimentina/metabolismo , alfa-Fetoproteínas/metabolismo
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