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1.
Yi Chuan ; 41(4): 285-292, 2019 Apr 20.
Artigo em Chinês | MEDLINE | ID: mdl-30992250

RESUMO

Histone methylation is a modification which occurs in the N-terminal peptide chains of the histone nucleosome. The 4th, 9th, 27th, 36th and 79th lysines in N-terminal peptide chain of histone H3 are hot spots for this modification, including mono-, di-, and tri-methylation. H3K27me3 is the tri-methylation modification on histone H3 lysine 27, which mainly functions as a transcriptional repressor regulating skeletal muscle development. Studies have shown that H3K27me3 can finely regulate skeletal muscle proliferation, including the level and duration of skeletal muscle development by specifically binding to myogenic regulatory factors (e.g., MyoD, MyoG, etc.), cell cycling regulators, and epigenetic regulators including lncRNA and miRNA. In this review, we introduce the types and mechanisms of histone methylation and de-methylation of H3K27. We also summarize how H3K27me3 functions in the proliferation and differentiation of skeletal muscle cell. This review will contribute to the comprehension of the function of H3K27me3 in regulating skeletal muscle development and provide reference for further improving our understanding of mammalian muscle.


Assuntos
Histonas/fisiologia , Desenvolvimento Muscular , Músculo Esquelético/crescimento & desenvolvimento , Animais , Proliferação de Células , Lisina/química , Mamíferos , Metilação , Células Musculares/citologia , Nucleossomos/química
2.
Yi Chuan ; 40(9): 749-757, 2018 Sep 20.
Artigo em Chinês | MEDLINE | ID: mdl-30369478

RESUMO

Non-homologous end-joining (NHEJ) is the predominant DNA double-strand break (DSB) repair pathway in mammalian cells. It inhibits the efficiency of homologous recombination (HR) by competing for DSB targets. To improve the efficiency of HR in porcine fetal fibroblasts (PFFs), several RNA interference (RNAi) systems were designed to knockdown NHEJ key molecules, such as polynucleotide kinase/phosphatase (PNKP), DNA ligase IV (LIG4) and NHEJ1. The results show that siRNA significantly knocked down LIG4, PNKP and NHEJ1 expression. Suppression of PNKP dramatically increased the efficiency of single-strand annealing (SSA), double-strand DNA (dsDNA) and single-strand DNA (ssODN) mediated homology-directed repair (HDR) by 55.7%, 37.4% and 73.1% after transfected with the SSA-GFP reporter, HDR-GFP system or ssODN-GFP system, respectively; whereas knockdown of LIG4 and NHEJ1 repair factors significantly increased dsDNA or ssODN-mediated HDR efficiency by 37.5% and 76.9%, respectively.


Assuntos
Reparo do DNA por Junção de Extremidades , Recombinação Homóloga , Interferência de RNA , Suínos/genética , Animais , DNA Ligase Dependente de ATP/genética , DNA Ligase Dependente de ATP/metabolismo , DNA de Cadeia Simples/genética , DNA de Cadeia Simples/metabolismo , Feminino , Fibroblastos/metabolismo , Técnicas de Silenciamento de Genes , Masculino , Reparo de DNA por Recombinação , Suínos/embriologia , Suínos/metabolismo
3.
Yi Chuan ; 38(12): 1081-1089, 2016 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-28034840

RESUMO

Somatic cell nuclear transfer technique has great applications in livestock breeding, production of genetically modified animals, rescue of endangered species and treatment of human diseases. However, the currently low efficiency in animals cloning, an average of less than 5%, greatly hindered the rapid development of this technique. Among many factors which affect the efficiency of cloning pigs, X chromosome inactivation is an important one. Moreover, Xist gene is closely related to X chromosome inactivation, suggesting that it may directly or indirectly affects cloning efficiency. In this study, multiple sgRNAs were designed based on the CRISPR/Cas system, and two sites (Target 3 and Target 4) whose mutation efficiency were 1% and 3% at the cellular level were selected. We successfully knocked out Xist with 100% efficiency by microinjecting sgRNAs for Target 3 and Target 4 in embryo. Finally, 6 cloning piglets were born including two Xist-fully-knockout piglets. The follow-up studies on increasing cloning efficiency can be carried out based on the Xist-knockout model.


Assuntos
RNA Longo não Codificante/metabolismo , Animais , Sistemas CRISPR-Cas/genética , Sistemas CRISPR-Cas/fisiologia , Técnicas de Inativação de Genes , RNA Guia/genética , RNA Longo não Codificante/genética , Suínos
4.
Yi Chuan ; 38(5): 402-10, 2016 05.
Artigo em Chinês | MEDLINE | ID: mdl-27232488

RESUMO

The cloning technique, also called somatic cell nuclear transfer (SCNT), has been successfully established and gradually applied to various mammalian species. However, the developmental rate of SCNT mammalian embryos is very low, usually at 1% to 5%, which limits the application of SCNT. Placental developmental defects are considered as the main cause of SCNT embryo development inhibition. Almost all of SCNT-derived mammalian placentas exhibit various abnormalities, such as placental hyperplasia, vascular defects and umbilical cord malformation. Mechanistically, these abnormalities result from failure of establishment of correct epigenetic modification in the trophectoderm genome, which leads to erroneous expression of important genes for placenta development-related, particularly imprinted genes. Consequently, aberrant imprinted gene expression gives rise to placental morphologic abnormalities and functional defects, therefore decreases developmental competence of cloned embryos. Currently, although numerous methods that can improve the developmental ability of SCNT-derived embryos have been reported, most of them are unable to substantially enhance the success rate of SCNT due to failure to eliminate the placental development defects. In this review, we summarize placental abnormalities and imprinted gene expression in mammalian cloning, and propose directions for the future research aiming to improve the cloning efficiency.


Assuntos
Técnicas de Transferência Nuclear , Placenta/anormalidades , Animais , Embrião de Mamíferos , Desenvolvimento Embrionário , Feminino , Impressão Genômica , Placenta/irrigação sanguínea , Gravidez , Cordão Umbilical/anormalidades
5.
Neural Regen Res ; 11(3): 493-501, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27127492

RESUMO

Exogenous substance P accelerates wound healing in diabetes, but the mechanism remains poorly understood. Here, we established a rat model by intraperitoneally injecting streptozotocin. Four wounds (1.8 cm diameter) were drilled using a self-made punch onto the back, bilateral to the vertebral column, and then treated using amniotic membrane with epidermal stem cells and/or substance P around and in the middle of the wounds. With the combined treatment the wound-healing rate was 100% at 14 days. With prolonged time, type I collagen content gradually increased, yet type III collagen content gradually diminished. Abundant protein gene product 9.5- and substance P-immunoreactive nerve fibers regenerated. Partial nerve fiber endings extended to the epidermis. The therapeutic effects of combined substance P and epidermal stem cells were better than with amniotic membrane and either factor alone. Our results suggest that the combination of substance P and epidermal stem cells effectively contributes to nerve regeneration and wound healing in diabetic rats.

6.
Zhonghua Shao Shang Za Zhi ; 29(4): 374-7, 2013 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-24351538

RESUMO

MicroRNAs are endogenous noncoding RNA molecules with 19-22 nucleotides in length. MicroRNAs can post-transcriptionally regulate gene and (or) protein expression by binding to their target messenger RNAs (mRNAs), leading to mRNA degradation or suppression of translation. As a huge family that regulates gene expression, microRNAs has recently been shown to not only participate in the normal healing processes of wounds but also closely related to pathologic wound healing, and formation of hypertrophic scars and keloids. This review focuses on the biogenesis of microRNA and its role in wound healing.


Assuntos
MicroRNAs , Cicatrização/genética , Animais , Humanos
7.
Zhonghua Zheng Xing Wai Ke Za Zhi ; 29(6): 406-12, 2013 Nov.
Artigo em Chinês | MEDLINE | ID: mdl-24624876

RESUMO

OBJECTIVE: To observe the effect of tetrandine on gene expression of collagen type I, collagen type III, transformation growth factor-beta1 and to investigate the inhibitory effect of tetrandine on the scar tissue hyperplasia in rabbits' ears. METHODS: After the scar model was formed on the rabbits' ears, the rabbits were divided into 4 groups to receive intro-lesion injection with saline, or prednisolone (Pre) or tetrandrine in low concentration (L-Tet, 1.0 mg/ml) or tetrandrine in high concentration (H-Tet, 7.5 mg/ml). The morphological changes of scar tissue were observed. The changes of fibroblasts quantity and collagen expression were observed with HE and Masson staining. Immunohistochemical study was used to observe the expression level of collagen type I and collagen type III and TGF-beta1. Collagen type I and collagen type III and TGF-beta1, and signal factor Smad 3 mRNA were detected with RT-PCR. RESULTS: (1) 24 days after injury, all the wounds healed completely with formation of red, tough and hypertrophic scar. HE and Masson staining showed significant increase of fibroblasts and collagen density with irregularly arrangement. (2) Compared with that in saline group, the scar in other groups became softer, lighter and thinner, especially in H-Tet group. (3) HE and Masson staining shows the scar in Tet and Pre groups contained less fibroblasts and lower collagen dentsity with comparatively regular arrangement than that in saline group (P < 0.01), especially in H-Tet group. (4) According to the immunohistochemical study, the expression of collage type I and III and TGF-beta was positive in all the groups, but the positive rate and the ratio of collagen density I to III decreased in the order of saline, L-Tet, H-Tet and Pre groups (P < 0.01). (5) PT-PCR detection results showed that the amplification bands brightness of collagen type I and III and TGF-beta1 and signal molecular Smad 3 mRNA in scar tissue were obviously different. Compared with that in saline group, the expression of collagen type I and III and TGF-beta1 and Smad 3 mRNA decreased in Tet and Pre groups (P < 0.01). H-Tet group showed the most obvious reduce in the expression of type I collagen and TGF-beta1 and Smad 3 mRNA. Conclusions Tetrandine can significantly suppress the expression of collagen type I and collagen type III and TGF-beta1 on hypertrophic scar of rabbit ears, and reduce signal factor Smad 3 mRNA' s expression. It may be one of the important mechanism for its inhibitory effect on scar hyperplasia.


Assuntos
Benzilisoquinolinas/farmacologia , Cicatriz Hipertrófica/tratamento farmacológico , Cicatriz Hipertrófica/genética , Colágeno Tipo III/genética , Colágeno Tipo I/genética , Medicamentos de Ervas Chinesas/farmacologia , Expressão Gênica , Fator de Crescimento Transformador beta1/genética , Animais , Cicatriz Hipertrófica/patologia , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Orelha , Fibroblastos , Masculino , RNA Mensageiro/metabolismo , Coelhos , Proteína Smad3/genética , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
8.
Zhonghua Shao Shang Za Zhi ; 28(4): 274-7, 2012 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-23248961

RESUMO

OBJECTIVE: To investigate the feasibility of differentiation of human induced pluripotent stem cells (iPSCs) into epidermal-like stem cells. METHODS: (1) Human strain of iPSCs were plated on-to trophoblast of inactivated Fb strain of mouse embryos and cultured in complete medium of embryonic stem cells, iPSCs were subcultured by collagenase IV digestion method. The morphology and growth of iPSCs were observed under inverted phase contrast microscope, and the cells were stained with alkaline phosphatase (AKP). iPSCs were cultured in incomplete medium of embryonic stem cells to observe the ability of embryoid body formation. (2) Human iPSCs were inoculated onto 6-well plate covered with human amniotic membrane to culture as induction group. Other iPSCs were cultured on 6-well plate without human amniotic membrane as control group. Morphological changes in iPSCs in two groups were observed. Expressions of integrin beta1 and CK19 of iPSCs in two groups were determined by immunocytochemical staining. RESULTS: Human iPSCs showed a typical stem cell clone-like growth with a clear boundary, and they proliferated vigorously in complete medium of embryonic stem cells. These cells were AKP-positive. iPSCs formed embryoid body in trophoblast-free and suspension culture conditions. After 4 days of co-culture, stem cell clones were formed on the surface of amniotic membrane in induction group, and part of the cells were integrin beta1 and CK19 positive. Most of the cells died, and no integrin beta1 and CK19 positive cells were found in control group. CONCLUSIONS: Human iPSCs can be differentiated into epidermal-like stem cells by amniotic membrane induction, and it lays an experimental basis for providing new source of seed cells of skin tissue engineering.


Assuntos
Técnicas de Cultura de Células , Diferenciação Celular , Células-Tronco Pluripotentes Induzidas/citologia , Animais , Células Cultivadas , Células Epidérmicas , Humanos , Camundongos
9.
Zhonghua Yi Xue Za Zhi ; 92(10): 692-4, 2012 Mar 13.
Artigo em Chinês | MEDLINE | ID: mdl-22781298

RESUMO

OBJECTIVE: To explore the miRNA differential expression profiles of hyperplastic scar and normal skin so as to further elucidate the pathogenesis of hyperplastic scar and search for new therapeutic targets. METHODS: The total RNA was extracted from 5 human hyperplastic scar and normal skin tissues by Trizol. The specimens were collected from the First Affiliated Hospital of Nanchang University from November 2010 to May 2011, and purified by mirVana(TM) miRNA Isolation Kit and then labeled and hybridized by miRNA Complete Labeling and Hyb Kit. The images of hybridization were analyzed by the Feature Extraction (v10.7) software and the microarray results confirmed by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: In hyperplastic scar, 92 miRNA genes were up-regulated and 13 down-regulated. The most significantly up-regulated miRNAs were hsa-miR-564 and hsa-miR-936, etc. while hsa-miR-451, hsa-miR-223, hsa-miR-363 and hsa-miR-29b-1* became significantly down-regulated. The findings of RT-PCR on hsa-miR-21 and hsa-miR-451 of regulation were in a high concordance with the microarray results. CONCLUSION: Distinct differences of miRNA expression between human hyperplastic scar and normal skin, it may be closely correlated with the formation, development and evolution of hyperplastic scar.


Assuntos
Cicatriz Hipertrófica/genética , MicroRNAs/genética , Pele/metabolismo , Transcriptoma , Adolescente , Adulto , Estudos de Casos e Controles , Criança , Pré-Escolar , Cicatriz Hipertrófica/metabolismo , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Adulto Jovem
10.
Zhonghua Shao Shang Za Zhi ; 28(1): 25-31, 2012 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-22490536

RESUMO

OBJECTIVE: To observe the effect of sensory neuropeptide substance P combined with epidermal stem cells (ESC) on wound healing and nerve regeneration in diabetic rats. METHODS: ESC that had been isolated from SD rats were identified and cultured in vitro, and they were inoculated onto nourishing layer of amniotic membrane to construct amniotic membrane-ESC. Four full-thickness skin wounds were produced on the back of each of 48 diabetic rats. The resulted 192 wounds were randomly divided into ESC + substance P group, ESC group, substance P group, and control group according to the lottery method, with 48 wounds in each group. Wounds in ESC + substance P group and ESC group were transplanted with amniotic membrane-ESC, and those in substance P group and control group were transplanted with amniotic membrane. After transplantation, 250 µL substance P in the concentration of 1 × 10(-7) mol/L was injected around and into the middle of the wounds in ESC + substance P group and substance P group, 2 times a day, and continued for 4 days, while 250 µL PBS solution was injected in the above-mentioned position in ESC group and control group as control, 2 times a day, and continued for 4 days. On post injury day (PID) 4, 7, 10, 14, 17, and 23, the wound healing rate (with 8 wounds at each time point) was observed and determined, and changes in wound tissue structure were observed with HE staining. On PID 4, 7, and 10, collagen distribution in wound tissue was observed with Masson staining, and type I and type III collagen deposition in wound tissue was respectively observed after immunohistochemical staining. The distribution of protein gene product 9.5 (PGP 9.5) and regeneration of substance P positive nerve fibers in wound tissue were observed with immunohistochemical staining on PID 14 and 23. Data were processed with one-way analysis of variance and t test. RESULTS: (1) The wound healing rate in ESC + substance P group reached 100.0% on PID 14, which was obviously earlier than that in ESC group, substance P group, and control group, healing was respectively observed on PID 17, 17, and 23. The wound healing quality in ESC + substance P group was better than that in the other three groups as shown by HE staining. (2) On PID 10, collagen that was darkly stained and widely distributed was observed in wound tissue of ESC + substance P group and substance P group, while collagen in the other two groups was lightly stained and narrowly distributed. Deposition quantity of type I collagen gradually increased, and that of type III collagen gradually decreased in the wounds of each group over time. On PID 4, 7, and 10, distribution amount of type I collagen in wound tissue of ESC + substance P group was significantly higher than that in ESC group (with t value respectively 32.72, 118.21, 26.71, P values all below 0.01) and control group (with t value respectively 44.37, 22.76, 30.32, P values all below 0.01), while there was no significance between ESC + substance P group and substance P group. On PID 4, 7, and 10, distribution amount of type III collagen in wound tissue of ESC + substance P group was significantly higher than that in ESC group (with t value respectively 32.27, 28.68, 14.51, P values all below 0.01) and control group (with t value respectively 35.68, 22.52, 22.24, P values all below 0.01). (3) A large amount of PGP 9.5 and regeneration of substance P positive nerve fibers, and some peripheral nerve fibers in deep wound extending to epidermis were observed in wound tissue of ESC + substance P group and substance P group. A small amount of PGP 9.5 and regeneration of substance P positive nerve fibers without peripheral nerve fibers extending to epidermis were observed in deep wound tissue of ESC group and control group. On PID 14, 23, ratios of area of PGP 9.5 positive nerve fiber in the wounds of ESC + substance P group were (3.86 ± 0.25)% and (7.03 ± 0.28)%, and they were significantly higher than those of ESC group [(1.48 ± 0.30)%, (3.01 ± 0.43)%, with t value respectively 23.95, 30.27, P values all below 0.01] and control group [(1.46 ± 0.23)%, (2.84 ± 0.29)%, with t value respectively 27.35, 40.32, P values all below 0.01]. On PID 14, 23, ratios of substance P positive nerve fiber area in the wounds of ESC + substance P group were (2.01 ± 0.14)% and (1.19 ± 0.11)%, which were obviously higher than those of ESC group [(0.85 ± 0.17)%, (1.34 ± 0.21)%, with t value respectively 20.50, 2.60, P < 0.05 or P < 0.01] and control group [(0.74 ± 0.15)%, (1.30 ± 0.17)%, with t value respectively 23.98, 2.41, P < 0.05 or P < 0.01]. CONCLUSIONS: Joint application of substance P and ESC can effectively promote healing of wound and nerve regeneration in diabetic rats.


Assuntos
Diabetes Mellitus Experimental , Regeneração Nervosa , Células-Tronco/citologia , Substância P/farmacologia , Cicatrização , Animais , Diabetes Mellitus Experimental/patologia , Células Epiteliais/citologia , Ratos , Ratos Sprague-Dawley , Substância P/uso terapêutico
11.
Zhonghua Shao Shang Za Zhi ; 27(1): 26-31, 2011 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-21591337

RESUMO

OBJECTIVE: To analyze expression characteristics of human skin epidermal stem cell at different developmental stages, and to explore its biological significance. METHODS: Health skin samples from 28-32 w fetuses (F group), 4-12 y children (C group), and 35-55 y adult (A group) were harvested, with 10 cases in each group. Epidermis were separated using trypsin digestion and EDTA, and human epidermal stem cells were isolated and purified with type IV collagen attachment method. The monoclonal antibody of integrin beta1 and keratin 19 were used for detection and identification of epidermal stem cells by immunohistochemical staining. Total RNA was extracted from above cells by Trizol one-step method, and were detected by formaldehyde denaturing agarose gel electrophoresis. Probes were prepared and hybridized into cDNA microarray for scanning fluorescent signals and analysis of images, with two-fold differential expression value for screening. Significantly up/down-regulated genes were selected for verification by real time RT-PCR. RESULTS: By comparing expression profile between A and C groups, a total of 1808 genes with differential expression were detected, including 1089 up-regulated genes and 719 down-regulated genes, and they were classified into 128 categories. Among them, 1462 genes were known (found in GeneBank), 346 genes were unknown. A total of 4534 genes with differential expression were detected between C and F groups, in which 1783 genes were up-regulated and 2751 genes were down-regulated, and they were classified into 216 categories. Among them, 3577 genes were known (found in GeneBank), and 957 genes were unknown. There were 1104 genes with differential expression consistently detected in F, C and A groups, which were classified into 32 categories according to gene function. Among them, 94 genes were consistently up-regulated and 75 genes consistently down-regulated. Test results of real time RT-PCR were in accordance with above-mentioned results. CONCLUSIONS: Gene expression profiles of epidermal stem cells cultured in vitro, harvested from fetuses, children, and adult, exhibit obvious difference. This may be closely related to different stages of proliferation and differentiation of human epidermal stem cell and self-repair ability of wound at different developmental stages.


Assuntos
Epiderme/crescimento & desenvolvimento , Perfilação da Expressão Gênica , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Células-Tronco/citologia , Adulto , Diferenciação Celular , Criança , Pré-Escolar , Células Epidérmicas , Células Epiteliais/citologia , Feto/citologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Pessoa de Meia-Idade , Transcriptoma
12.
Mol Med Rep ; 4(2): 377-81, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-21468580

RESUMO

The healing of diabetic wounds represents a formidable clinical challenge, and the molecular mechanisms involved in diabetic wound healing are far from clear. In this study, we investigated the expression of ß-catenin and cyclin D1 in the epidermal stem cells (ESCs) of diabetic rats, and explored whether the reduction of ß-catenin and its downstream target in ESCs, cyclin D1, lead to poor wound healing in diabetes mellitus (DM). We found that, compared to the controls, the ESCs of diabetic rats were markedly reduced, the clone formation efficiency of the ESCs was markedly lower, and the mRNA and protein expression of ß-catenin and cyclin D1 was significantly decreased. These findings suggest that the low expression of ß-catenin and cyclin D1 may reduce the activity of ESCs from diabetic rats, which might be one of the important mechanisms of delayed wound healing in DM.


Assuntos
Ciclina D1/metabolismo , Diabetes Mellitus Experimental/metabolismo , Epiderme/patologia , Células-Tronco/metabolismo , beta Catenina/metabolismo , Animais , Western Blotting , Forma Celular , Ensaio de Unidades Formadoras de Colônias , Ciclina D1/genética , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Regulação da Expressão Gênica , Imuno-Histoquímica , Masculino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Células-Tronco/patologia , beta Catenina/genética
13.
Zhonghua Shao Shang Za Zhi ; 25(4): 261-4, 2009 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-19951543

RESUMO

OBJECTIVE: To establish an effective method of transfecting human marrow mesenchymal stem cells (MSC) with human vascular endothelial growth factor 165 (VEGF 165) gene. METHODS: MSCs isolated and cultured in vitro were divided into transfection group (pShuttle-CMV/VEGF 165 plasmid was transfected into MSCs through liposome-mediating method), empty plasmid group (pShuttle-CMV vehicle was transfected into MSCs as control), liposome group (liposome was transfected into MSCs as control) and control group (normal culture). Expressions of mRNA and protein of MSCs were determined by RT-PCR, enzyme-linked immunosorbent assay and Western Blot. Sensitivity to MSCs on VEGF plasmid transfection was detected by MTT test. RESULTS: Expression level of VEGF 165 gene mRNA in transfection group, empty plasmid group, liposome group, and control group was respectively 0.89 +/- 0.03, 0.34 +/- 0.04, 0.40 +/- 0.03, and 0.30 +/- 0.03, and the difference between transfection group and the other three groups was statistically significant (P < 0.01). Content of VEGF protein in transfection group, empty plasmid group, liposome group, and control group was respectively (778 +/- 35), (543 +/- 24), (561 +/- 28), (571 +/- 23) pg/mL, and the difference between transfection group and the other three groups was statistically significant (P < 0.01). In the transfection group, expression level of VEGF protein peaked on 7(th) day after transfection, which was decreased gradually later. In transfection group, expression level of VEGF 165 protein was obviously higher than that of the other three groups (P < 0.01), and no inhibitory effect of VEGF plasmid transfection on MSCs proliferation was found. CONCLUSIONS: The method for transfecting human VEGF 165 gene into MSCs is established in this research, through which target gene and protein can express effectively.


Assuntos
Células da Medula Óssea/citologia , Células-Tronco Mesenquimais/citologia , Transfecção , Fator A de Crescimento do Endotélio Vascular/genética , Técnicas de Cultura de Células , Células Cultivadas , Humanos
14.
Zhonghua Shao Shang Za Zhi ; 24(4): 275-7, 2008 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-19102984

RESUMO

OBJECTIVE: To investigate the feasibility of transfection of recombinant human endothelial nitric oxide synthase (eNOS) into human hypertrophic scar fibroblasts (HSFbs), and to observe NO secretion and the synthesis of collagen I and III. METHODS: Recombinant human eNOS with karyocyte expressive vector was constructed in vitro, then was transfected into HSFbs which was isolated from hypertrophic scar tissues and cultured in vitro (T group). The HSFbs untransfected (normal culture) or transfected with empty-vector was used as control group and empty-vector group respectively. The mRNA expression of eNOS, collagen I and III was determined by Realtime PCR. The content of NO was determined by NO assay kit. RESULTS: The expression of eNOS mRNA in T group was 5.92 +/- 0.21, which was obviously higher than that in empty-vector group (0.98 +/- 0.13, P < 0.05). The expression of collagen I mRNA (0.76 +/- 0.15), and collagen III (0.79 +/- 0.08) in T group was significantly lower than those in empty-vector group (0.98 +/- 0.15, 1.02 +/- 0.12, P < 0.05, respectively). The content of NO in T group (36.1 +/- 0.8 micromol/L) was obviously higher than that in empty-vector group (28.4 +/- 1.0 micromol/L, P < 0.01) and control group (27.7 +/- 1.3 micromol/L, P < 0.01). CONCLUSION: HSFbs can be the target cells for eNOS gene transfection. The transfected cells can express eNOS and produce NO, which inhibit the synthesis of collagen.


Assuntos
Cicatriz Hipertrófica/metabolismo , Fibroblastos/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Transfecção , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Colágeno Tipo III/metabolismo , Humanos , Técnicas In Vitro , Óxido Nítrico/metabolismo , RNA Mensageiro/metabolismo
15.
Yi Chuan Xue Bao ; 33(8): 711-6, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16939005

RESUMO

To study the potential use of estrogen receptor gene (ESR) as a genetic marker to improve the reproductive traits of pigs, the genotypes of the ESR PCR product digested by Pvu II were determined in 2,239 litters from 612 Landrace sows. The data of the first, second, and later parities were separately evaluated. Although the frequency of the B allele was much lower than that of the A allele, likelihood ratio test showed that the gene frequencies were in Hardy-Weinberg equilibrium. The effects of ESR of different parities were not equal. In summary, the sows with the BB genotype showed better performance for total number of piglets born (TNB) and number of piglets born alive (NBA), but had a lower average piglet weight at birth (AWB). It was concluded that ESR could be used as a marker for the selection of litter size in the Landrace population.


Assuntos
Receptores Estrogênicos/genética , Reprodução/genética , Sus scrofa/genética , Animais , Peso Corporal/genética , Estrogênios/metabolismo , Feminino , Tamanho da Ninhada de Vivíparos/genética , Gravidez
16.
Zhonghua Shao Shang Za Zhi ; 22(6): 440-4, 2006 Dec.
Artigo em Chinês | MEDLINE | ID: mdl-17438691

RESUMO

OBJECTIVE: To investigate the efficacy of early fluid resuscitation on hepatic steatosis in rats after severe scald. METHODS: One hundred and forty-four Sprague-Dawley rats were enrolled in the study. In thirty-six rats skin of 30% TBSA was treated with cold water to serve as sham injury group. All other rats were inflicted with 30% full-thickness scald, and they were subdivided into 3 groups, i. e. scald group(S, without resuscitation), delayed resuscitation group ( DR, with Ringer's solution at 6 post-scald hour(PSH) ) and early resuscitation group( ER, with Ringer's solution immediately after scald). The hepatic tissues of the rats were harvested at 0.5, 1.0,2.0,3.0,7.0 post-scald hour( PSH) and on 21.0 PSD for the observation of pathological changes with light-microscope and transmission electron microscope. The serum contents of TC, TG, HDL, ALP were determined at the same time-points. Body weight of each rat was measured before blood sampling, and total liver weight after blood sampling. Liver weight/body weight ratio was recorded. RESULTS: Compared with sham injury group, the fat denaturation degree of hepatic tissue in ER group was obviously less than that in S and DR group . The serum level of high density lipoprotein (TC) , triglyceride ( TG) , and alkaline phosphatase (ALP) after scald increased ranking as S > DR > ER, while the level of HDL decreased in that order. The liver weight/body weight ratio of the rats in DR group on 1.0 PSD was obviously elevated compared with that in ER group( P <0. 05) , and there exhibited significant difference of liver weight/body weight ratio between DR and ER groups on 7. 0 PSD ( P < 0. 01). The liver steatosis had obvious negative correlation with HDL content after scald( r = -0. 37, P <0.01) , but it had positive correlation with the ALP content( r = 0. 45, P <0. 01), TG content( r = 0. 25, P <0. 01) and liver weight/body weight ratio( r = 0. 440, P <0. 01). The remaining parameters showed no correlation with the liver steatosis. CONCLUSION: Fluid resuscitation immediately after scald can ameliorate hepatic fatty degeneration, reduce its incidence, and beneficial to recovery of liver damage to a certain extent.


Assuntos
Queimaduras/terapia , Fígado Gorduroso/terapia , Hidratação , Animais , Queimaduras/complicações , Queimaduras/patologia , Modelos Animais de Doenças , Fígado Gorduroso/etiologia , Feminino , Fígado/patologia , Masculino , Ratos , Ratos Sprague-Dawley
17.
Yi Chuan Xue Bao ; 31(12): 1361-8, 2004 Dec.
Artigo em Chinês | MEDLINE | ID: mdl-15633641

RESUMO

With F2 design, 16 Chinese Lantang sows crossbred with eight highly improved Landrace boars to establish a resource population including 40 F1 sows, eight F1 boars, and 232 F2 pigs. Genetic analysis of the resource population showed that the 32 performance traits displayed some degree of variation, and coefficients of variation of the majority of economic traits exceeded 10%. The variance component analysis revealed that the ratios of additive genetic variance to phenotypic variance of the majority of economic traits were high. Of the 22 microsatellite DNA markers, only 12 microsatellite DNA markers are polymorphisic in this populations. The average heterozygosity of these markers and PIC were 0.53 and 0.46 respectively. The results showed that these markers can provide enough information for QTL mapping. In conclusion,the F2 pigs were sufficiently segregated, and were capable of serving as a resource population for QTL mapping.


Assuntos
Locos de Características Quantitativas , Suínos/genética , Animais , Feminino , Heterozigoto , Masculino , Repetições de Microssatélites
18.
Yi Chuan Xue Bao ; 30(9): 835-9, 2003 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-14577375

RESUMO

A Landrace x Lantang resource population (LL-SCAU) including 216 F2 pigs was founded by F2 design for analysis of IGF-1 gene polymorphism by PCR-RFLP. The least square means of divergent IGF-1 genotypes for the measured traits were estimated with the fixed model. The genetic effects of IGF-1 gene were estimated with a mixed model and the additive and dominant effects of IGF-1 gene were accordingly calculated. The results of the fixed model and the mixed model showed that IGF-1 locus significantly affected average daily gain after weaning. Average daily gain after weaning for AA genotype on IGF-1 locus was 20.58 g (P = 0.0347) higher than for AB. The additive and dominant effects attributed to IGF-1 were 1.78 g and -18.81 g respectively. IGF-1 locus also significantly affected carcass composition. Bone percentages for AA and AB on IGF-1 locus were lower than for BB by 5.22% (P = 0.0008) and 5.19% (P = 0.0007) respectively, and the additive and dominant effects were -2.61% and -2.58% respectively. Amount of carcass lean for AA on IGF-1 locus was less than for AB by 0.45 kg (P = 0.0264), and the additive and dominant effects were 0.16 kg and 0.61 kg respectively. Skin and fat percentage for AA and AB on IGF-1 locus was higher than for BB by 8.81% (P = 0.0206) and 7.64% (P = 0.0431) respectively, and the additive and dominant effects were 4.41% and 3.24% respectively. The genetic analysis of IGF-1 gene showed that IGF-1 locus significantly affected average daily gain after weaning, bone percentage, carcass lean, skin and fat percentage. The estimated additive and dominant effects showed that only the additive effect of IGF-1 locus on skin and fat percentage was much higher than dominant effect, and IGF-1 gene can be used as a major gene for effective selection of these traits.


Assuntos
Fator de Crescimento Insulin-Like I/genética , Suínos/genética , Animais , Animais Recém-Nascidos , Peso Corporal/genética , Cruzamento , DNA/genética , DNA/isolamento & purificação , Feminino , Frequência do Gene , Genótipo , Masculino , Polimorfismo Genético , Suínos/crescimento & desenvolvimento , Ganho de Peso/genética
19.
Zhonghua Zheng Xing Wai Ke Za Zhi ; 19(1): 51-3, 2003 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-12778798

RESUMO

OBJECTIVE: To investigate the role of endothelin (ET) in the proliferation and collagen synthesis of human scar-derived fibroblasts and the modulation of its antagonists such as nitric oxide (NO), tetrandrine (Tet). METHODS: With the cultured fibroblasts from the scarring tissue, the cell proliferation was determined by [3H]-TdR incorporation, while the collagen synthesis was evaluated by [3H]-proline incorporation. RESULTS: The ET-1 was significantly increasing the proliferation and collagen synthesis of human scar-derived fibroblasts. The values of [3H]-TdR absorption in the 2.5 ng/ml, 25 ng/ml and 100 ng/ml of ET-1 groups were 1.8 times, 4 times and 4.9 times more than in the control group, respectively (P < 0.01), while the values of the [3H]-proline incorporation were 1.1 times, 3.1 times and 3.8 times respectively (P < 0.01). The fibroblasts, treated with 50 micrograms/ml of S-nitroso-N-acetyl penicillamine(SNAP), were no detectable effect on the basal level of DNA synthesis, but produced decreasing effect on the [3H]-TdR absorption (the rate of inhibition was 22.89%, P < 0.05). It was found that the SNAP inhibited the [3H]-proline incorporation in cultured fibroblasts, but the rate of [3H]-proline incorporation induced by ET-1 was unaltered. The Tet with 3 micrograms/ml, in which does not inhibit the basal level of DNA synthesis, was significantly decreasing the collagen synthesis and decreasing the ET-mediated DNA synthesis (the rate of inhibition was 33.21% (P < 0.01). CONCLUSION: These results indicate that the ET can obviously increase the proliferation and collagen synthesis of human scar-derived fibroblasts, but it can be partially antagonized by NO and Tet.


Assuntos
Proliferação de Células/efeitos dos fármacos , Cicatriz/patologia , Colágeno/biossíntese , Endotelinas/farmacologia , Fibroblastos/efeitos da radiação , Benzilisoquinolinas/farmacologia , Células Cultivadas , DNA/biossíntese , Endotelinas/antagonistas & inibidores , Fibroblastos/citologia , Humanos , Óxido Nítrico/metabolismo , Óxido Nítrico/farmacologia , Prolina/metabolismo , S-Nitroso-N-Acetilpenicilamina/farmacologia
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