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1.
Cancer Lett ; 459: 100-111, 2019 Sep 10.
Artigo em Inglês | MEDLINE | ID: mdl-31158430

RESUMO

The mixed lineage kinase domain-like protein (MLKL) has emerged as a critical mediator of necroptosis, which results in the release of cellular damage-associated molecular patterns (DAMPs). However, its physiological role in regulating inflammation is not fully understood. We herein showed that Mlkl-/- mice were highly susceptible to colitis and colitis-associated tumorigenesis (CAT), which was associated with massive leukocyte infiltration and increased inflammatory responses. Moreover, we used bone marrow transplantation to reveal that MLKL in inflammatory cells is crucial for its role on colitis. Intestinal mucosal tissue and polyps isolated from Mlkl-/- mice exhibited increased ERK activation and elevated expression of genes associated with inflammation and cancer. Mechanistically, enhanced inflammation in Mlkl-/- mice was due to MEK/ERK activation particularly in dendritic cells (DCs). Our results demonstrate the role of MLKL in maintaining intestinal homeostasis and protecting against colitis and tumorigenesis.

2.
Cell Rep ; 19(4): 798-808, 2017 04 25.
Artigo em Inglês | MEDLINE | ID: mdl-28445730

RESUMO

RIPK3 mediates cell death and regulates inflammatory responses. Although genetic studies have suggested that RIPK3-MLKL-mediated necroptosis leads to embryonic lethality in Fadd or Caspase-8-deficient mice, the exact mechanisms are not fully understood. Here, we generated Ripk3 mutant mice by altering the RIPK3 kinase domain (Ripk3Δ/Δ mice), thus abolishing its kinase activity. Ripk3Δ/Δ cells were resistant to necroptosis stimulation in vitro, and Ripk3Δ/Δ mice were protected from necroptotic diseases. Although the Ripk3Δ/Δ mutation rescued embryonic lethality in Fadd-/- embryos, Fadd-/-Ripk3Δ/Δ mice died within 1 day after birth due to massive inflammation. These results indicate that Ripk3 ablation rescues embryonic lethality in Fadd-deficient mice by suppressing two RIPK3-mediating processes: necroptosis during embryogenesis and inflammation during postnatal development in Fadd-/- mice.


Assuntos
Apoptose , Proteína de Domínio de Morte Associada a Fas/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Apoptose/efeitos dos fármacos , Caspase 8/genética , Caspase 8/metabolismo , Ceruletídeo/toxicidade , Quimiocinas/metabolismo , Citocinas/análise , Citocinas/metabolismo , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Proteína de Domínio de Morte Associada a Fas/deficiência , Células HEK293 , Humanos , Inflamação , Lipopolissacarídeos/toxicidade , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Mutagênese , Necrose , Oligopeptídeos/farmacologia , Pancreatite/induzido quimicamente , Pancreatite/patologia , Fosforilação , Proteínas Quinases/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/genética
3.
J Exp Ther Oncol ; 11(1): 41-9, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26259389

RESUMO

Pristimerin (PM) is a quinonemethide triterpenoid present in various plant species with strong antiprolifertive and proapoptotic activities in cancer cells. The effect of PM on telomerase which is reactivated in most cancers including carcinoma of the prostate (CaP) has not been studied. We investigated the effect of PM on the expression of human telomerase reverse transcriptase (hTERT) gene that codes for the catalytic subunit of the telomerase holoenzyme complex in prostate cancer cell lines LNCaP and PC-3 cells. The inhibition of cell proliferation and induction of apoptosis by PM in both cell lines was associated with the inhibition of hTERT mRNA expression, suppression of native and phosphorylated hTERT protein and hTERT telomerase activity. The ablation of hTERT expression increased the sensitivity of cancer cells to PM. In addition, results also revealed that the inhibition of hTERT expression is attributed to the inhibition of transcription factors SP1, c-Myc and STAT3 and protein kinase B/Akt which regulate hTERT transcriptionally and post-translationally, respectively. These data provide evidence that telomerase is a potential target of PM in prostate cancer.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Neoplasias da Próstata/enzimologia , Telomerase/antagonistas & inibidores , Triterpenos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Fosforilação , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Interferência de RNA , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Telomerase/genética , Telomerase/metabolismo , Transcrição Genética/efeitos dos fármacos , Transfecção
4.
Appl Microbiol Biotechnol ; 99(12): 5317-26, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25620371

RESUMO

Aniline is of great environmental concern with regards to widespread occurrence in water and soil and increasing threat into the life forms. Bioremediation involving the use of degrading bacterium in the removal of aniline is the most promising process, yet inhibited under low temperature usually. In the present study, a new psychrotrophic bacterial strain isolated from groundwater, designated AN-1, was shown to be capable of aniline degradation in a concentration range of 135-2202 mg L(-1) within 72 h at 10 °C. Strain AN-1 was proposed to be a Pseudomonas migulae group of bacteria based on the evolutionary relationship and the morphological and biochemical characteristics. The pH, NaH2PO4, and aniline concentration were used as independent variables to optimize the aniline removal by AN-1 at 10 °C, and a statistically significant (R (2) = 0.9230, p < 0.005) quadratic polynomial mathematical model was suggested. Moreover, an efficient biocomposite by assembling Fe3O4 nanoparticles onto the surface of AN-1 cells was constructed. Compared with free cells, the microbial cell/Fe3O4 biocomposite had the same biodegradation activity but exhibited remarkable reusability. This study highlights AN-1 might be a promising candidate for aniline removal from wastewater at low temperatures.


Assuntos
Compostos de Anilina/metabolismo , Água Subterrânea/microbiologia , Pseudomonas/metabolismo , Biodegradação Ambiental , Temperatura Baixa , Magnetismo , Pseudomonas/química , Pseudomonas/genética , Pseudomonas/crescimento & desenvolvimento , Pseudomonas/isolamento & purificação , Poluentes do Solo/metabolismo , Poluentes Químicos da Água/metabolismo
5.
Transgenic Res ; 24(3): 537-47, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25487040

RESUMO

This study aims to investigate the relative plant growth and reproduction of insect-resistant and susceptible plants following the introgression of an insect-resistance Bt-transgene from Brassica napus, oilseed rape, to wild Brassica juncea. The second backcrossed generation (BC2) from a single backcross family was grown in pure and mixed stands of Bt-transgenic and non-transgenic siblings under two insect treatments. Various proportions of Bt-transgenic plants were employed in mixed stands to study the interaction between resistant and susceptible plants. In the pure stands, Bt-transgenic BC2 plants performed better than non-transgenic plants with or without insect treatments. In mixed stands, Bt-transgenic BC2 plants produced fewer seeds than their non-Bt counterparts at low proportions of Bt-transgenic BC2 plants in the absence of insects. Reproductive allocation of non-transgenic plants marginally increased with increasing proportions of Bt-transgenic plants under herbivore pressure, which resulted in increased total biomass and seed production per stand. The results showed that the growth of non-transgenic plants was protected by Bt-transgenic plants under herbivore pressure. The Bt-transgene might not be advantageous in mixed stands of backcrossed hybrids; thus transgene introgression would not be facilitated when herbivorous insects are not present. However, a relatively large initial population of Bt-transgenic plants might result in transgene persistence when target herbivores are present.


Assuntos
Fluxo Gênico , Mostardeira/crescimento & desenvolvimento , Mostardeira/genética , Plantas Geneticamente Modificadas , Brassica napus/genética , Herbivoria , Hibridização Genética , Sementes/genética , Sementes/crescimento & desenvolvimento , Transgenes
6.
Int J Oncol ; 45(4): 1735-41, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25175770

RESUMO

Pristimerin (PM), a quinonemethide triterpenoid, is a promising anticancer agent with potent antiproliferative and apoptosis-inducing activities against cancer cell lines. However, the anticancer activity and mechanisms of PM in prostate cancer cells have not been adequately investigated. Here we report that the degradation of survivin plays an important role in the antiproliferative and proapoptotic effects of PM in carcinoma of the prostate (CaP) cell lines. Treatment with PM inhibited proliferation and induced apoptosis in LNCaP and PC-3 cells as characterized by the loss of cell viability and an increase in Annexin V-binding and cleavage of PARP-1, respectively. The antiproliferative and apoptosis-inducing effects of PM were associated with the inhibition of cell cycle regulatory proteins, antiapoptotic survivin and members of the Bcl-2 family. Data showed that response to PM is regulated by survivin since overexpression of survivin rendered CaP cells resistant to PM. Furthermore, downregulation of survivin by PM was mediated through the ubiquitin-proteasomal degradation. Together, these data demonstrate that pristimerin inhibits proliferation and induces apoptosis in CaP cells by abolishing survivin through the ubiquitin-proteasome pathway.


Assuntos
Proteínas Inibidoras de Apoptose/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Triterpenos/farmacologia , Ubiquitina/metabolismo , Apoptose , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Survivina
7.
Int J Oncol ; 44(5): 1707-15, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24603988

RESUMO

Lack of effective therapeutics for pancreatic cancer at the present time underscores the dire need for safe and effective agents for the treatment of this malignancy. In the present study, we have evaluated the anticancer activity and the mechanism of action of pristimerin (PM), a quinonemethide triterpenoid, against MiaPaCa-2 and Panc-1 pancreatic ductal adenocarcinoma (PDA) cell lines. Treatment with PM inhibited the proliferation and induced apoptosis in both cell lines as characterized by the increased Annexin V-binding and cleavage of PARP-1 and procaspases -3, -8 and -9. PM also induced mitochondrial depolarization and the release of cytochrome c from the mitochondria. The induction of apoptosis by PM was associated with the inhibition of the pro-survival Akt, NF-κB and mTOR signaling proteins and their downstream intermediaries such as Foxo-3α and cyclin D1 (Akt); Cox-2 and VEGF (NF-κB); p-S6K1 and p-4E-BP1 (mTOR) as well as PKCε. Treatment with PM also inhibited the expression of anti-apoptotic Bcl-2 and survivin but not Bcl-xL. The downregulation of Bcl-2 by PM was not due to proteasomal or lysosomal proteolytic degradation of Bcl-2, since treatment with PM in the presence of proteasomal inhibitors MG132 or lactacystin (LAC) or calpain inhibitor MG101 failed to block the downregulation of Bcl-2 by PM. On the other hand, RT-PCR analysis showed the inhibition of Bcl-2 mRNA by PM in a dose-related manner, indicating that inhibition of Bcl-2 by PM is mediated through the suppression of Bcl-2 gene expression. Thus, the mechanistic understanding of the antitumor activity of pristimerin could facilitate in vivo efficacy studies of pristimerin for pancreatic cancer.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neoplasias Pancreáticas/patologia , Transdução de Sinais/efeitos dos fármacos , Triterpenos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Técnicas In Vitro , Mitocôndrias/fisiologia , NF-kappa B/genética , NF-kappa B/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
8.
Ying Yong Sheng Tai Xue Bao ; 25(9): 2745-55, 2014 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-25757330

RESUMO

Agricultural biodiversity is a key part of the ecosystem biodiversity, but it receives little concern. The monoculture, environmental pollution and habitat fragmentation caused by agricultural activities have threatened agricultural biodiversity over the past 50 years. To optimize agricultural management measures for crop production and environmental protection, we reviewed the effects of agricultural activities, including cultivation patterns, plastic mulching, chemical additions and the cultivation of transgenic crops, on agricultural biodiversity. The results showed that chemical pesticides and fertilizers had the most serious influence and the effects of transgenic crops varied with other factors like the specific transgene inserted in crops. The environmental risk of transgenic crops should be assessed widely through case-by-case methods, particularly its potential impacts on agricultural biodiversity. It is important to consider the protection of agricultural biodiversity before taking certain agricultural practices, which could improve agricultural production and simultaneously reduce the environmental impacts.


Assuntos
Agricultura , Biodiversidade , Produtos Agrícolas , Plantas Geneticamente Modificadas , Conservação dos Recursos Naturais , Ecossistema , Poluição Ambiental , Fertilizantes , Praguicidas
9.
J Carcinog Mutagen ; Suppl 6: 005, 2013 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-24877026

RESUMO

Pristimerin is a quinonemethide triterpenoid with the potential of a promising anticancer agent. Pristimerin (PM) has shown anticancer activity against a range of cancer cell lines, but its activity for prostate cancer has not been adequately investigated. In the present study we have examined the underlying mechanisms of the apoptotic response of the hormone-sensitive (LNCaP) and hormone-refractory (PC-3) prostate cancer cell lines to PM. Treatment with PM induced apoptosis in both cell lines as characterized by increased annexin V-binding and cleavage of PARP-1 and procaspases-3 and -9. It also induced mitochondrial depolarization, cytochrome c release from mitochondria and generation of reactive oxygen species (ROS). Response to PM is regulated by Bcl-2 since it down-regulated Bcl-2 expression and overexpression of Bcl-2 rendered prostate cancer cells resistant to PM. ROS plays a role in down-regulation of Bcl-2, since treatment with PM in the presence of various ROS modulators, e.g., n-acetylcysteine (NAC), a general purpose antioxidant; diphenylene iodonium (DPI), a NADPH inhibitor; rotenone (ROT), a mitochondrial electron transport chain interrupter rotenone or MnTBAP, a O2 scavenger, attenuated the down-regulation of Bcl-2. Furthermore, ROS is also involved in the ubiquitination and proteasomal degradation of Bcl-2 as both of these events were blocked by O 2- scavenger MnTBAP. Thus, pristimerin induces apoptosis in prostate cancer cells predominately through the mitochondrial apoptotic pathway by inhibiting antiapoptic Bcl-2 through a ROS-dependent ubiquitin-proteasomal degradation pathway.

10.
J Exp Ther Oncol ; 10(1): 51-64, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22946344

RESUMO

Oleanolic acid-derived synthetic triterpenoids are broad spectrum antiproliferative and antitumorigenic agents. In this study, we investigated the role of reactive oxygen species (ROS) in induction of apoptosis and inhibition of prosurvival Akt, NF-kappaB and mTOR signaling pro-teins by methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) in pancreatic cancer cells. Micromolar concentrations of CDDO-Me inhibited proliferation and induced apoptosis in MiaPaCa-2 and Panc-1 pancreatic cancer cells. Treatment with CDDO-Me caused the generation of hydrogen peroxide and superoxide anion and pretreatment of cells with NADPH oxidase inhibitor diphylene iodonium (DPI) or respiratory chain complex 1 inhibitor rotenone prevented ROS generation. Pretreatment with N-acetylcysteine (NAC) or overexpression of glutathione peroxidase (GPx) or superoxide dismutase-1 (SOD-1) blocked the antiproliferative effects of CDDO-Me. Likewise, NAC prevented the induction of apoptosis (annexin V-FITC binding and cleavage of PARP-1 and procaspases-3,-8 and -9) and reversed the loss of mitochondrial membrane potential and release of cytochrome c from mitochondria by CDDO-Me. CDDO-Me down-regulated p-Akt, p-mTOR and NF-kappaB (p65) but increased the activation of Erk1/2 and NAC blocked the modulation of these cell signaling proteins by CDDO-Me. Thus, the results of this study indicate that the antiproliferative and apoptosis inducing effects of CDDO-Me are mediated through a ROS-dependent mechanism and the role of ROS in modulation of signaling proteins by CDDO-Me warrants further investigation.


Assuntos
Acetilcisteína/farmacologia , Ácido Oleanólico/análogos & derivados , Neoplasias Pancreáticas/metabolismo , Espécies Reativas de Oxigênio , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glutationa Peroxidase/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Potencial da Membrana Mitocondrial , Ácido Oleanólico/administração & dosagem , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1 , Superóxidos/metabolismo
11.
Ying Yong Sheng Tai Xue Bao ; 23(6): 1713-20, 2012 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-22937665

RESUMO

China is one of the origin centers of rice (Oryza sativa), with a distribution of abundant wild rice germplasm resources. Since the biosafety certificates were issued to Bt transgenic rice containing cry1Ac gene against lepidopteran pests, concerns are raised on the gene flow from transgenic rice to its wild relatives. This paper reviewed the occurrence of transgenic rice gene flow and its potential ecological consequences. Most studies suggested that transgenic rice could successfully hybridize with cultivated rice, wild rice (O. rufipogon), red rice (O. sativa f. spontanea), and barnyard grass (Echinochloa caudata), and the gene flow from transgenic rice had a lower occurrence frequency and was highly variable. Once the gene flow successfully occurred, the transgenes could introgress into a wild population within only a few generations because of the fitness advantage of transgenic rice due to the new selective traits of transgenes. When the transgenic plants invaded and persisted in a wild population, the dynamic changes of the mixed population would depend on the relative fitness and competition capability. To study the effects of the gene flow of transgenic rice and its long-term ecological consequences would have significance in rationally protecting and utilizing wild rice germplasm resources in China.


Assuntos
Ecossistema , Fluxo Gênico , Hibridização Genética , Oryza/genética , Plantas Geneticamente Modificadas/genética , Echinochloa/genética , Oryza/crescimento & desenvolvimento
12.
PLoS One ; 7(6): e39705, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22745814

RESUMO

BACKGROUND: Seed size has significant implications in ecology, because of its effects on plant fitness. The hybrid seeds that result from crosses between crops and their wild relatives are often small, and the consequences of this have been poorly investigated. Here we report on plant performance of hybrid and its parental transgenic oilseed rape (Brassica napus) and wild B. juncea, all grown from seeds sorted into three seed-size categories. METHODOLOGY/PRINCIPAL FINDINGS: Three seed-size categories were sorted by seed diameter for transgenic B. napus, wild B. juncea and their transgenic and non-transgenic hybrids. The seeds were sown in a field at various plant densities. Globally, small-seeded plants had delayed flowering, lower biomass, fewer flowers and seeds, and a lower thousand-seed weight. The seed-size effect varied among plant types but was not affected by plant density. There was no negative effect of seed size in hybrids, but it was correlated with reduced growth for both parents. CONCLUSIONS: Our results imply that the risk of further gene flow would probably not be mitigated by the small size of transgenic hybrid seeds. No fitness cost was detected to be associated with the Bt-transgene in this study.


Assuntos
Brassica napus/crescimento & desenvolvimento , Brassica napus/fisiologia , Mostardeira/crescimento & desenvolvimento , Mostardeira/fisiologia , Sementes/crescimento & desenvolvimento , Sementes/fisiologia , Brassica napus/anatomia & histologia , Hibridização Genética/fisiologia , Mostardeira/anatomia & histologia , Sementes/anatomia & histologia
13.
Zhonghua Zheng Xing Wai Ke Za Zhi ; 25(3): 213-6, 2009 May.
Artigo em Chinês | MEDLINE | ID: mdl-19803206

RESUMO

OBJECTIVE: To detect the effect of tunicamycin on Fas protein expression and Fas monoclonal antibody (FasMcAb)-induced apoptosis of fibroblast from hypertrophic scar and keloid. METHODS: The expression of Fas protein was detected by immunostaining in 5 cases of keloid, 5 cases of hypertrophic scar and 5 cases of normal skin as control. The fibroblasts were cultured and treated with tunicamycin. The Fas protein expression and the fibroblast apoptosis rate were assessed by Western Blot and flow cytometry. RESULTS: It revealed that Fas protein was detectable in all the three groups. The Fas glycosylation level was highest in hypertrophic scar, but lowest in normal skin. The FasMcAb-induced apoptosis had a positive relationship with the Fas glycosylation. Tunicamycin had a significant inhibitory effect on the Fas glycosylation in keloid and hypertrophic scar, but not in normal skin. CONCLUSIONS: The FasMcAb-induced apoptosis has a positive relationship with the Fas glycosylation. Tunicamycin has a significant inhibitory effect on the Fas glycosylation in keloid and hypertrophic scar.


Assuntos
Cicatriz Hipertrófica/metabolismo , Fibroblastos/metabolismo , Queloide/metabolismo , Tunicamicina/farmacologia , Receptor fas/metabolismo , Adolescente , Adulto , Anticorpos Monoclonais/farmacologia , Apoptose , Criança , Cicatriz Hipertrófica/patologia , Feminino , Glicosilação , Humanos , Queloide/patologia , Masculino , Transdução de Sinais , Adulto Jovem
14.
Mol Biol Rep ; 36(8): 2221-8, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19116772

RESUMO

Methylation is one of epigenetic mechanisms regulating gene expression. The methylation pattern is determined during embryogenesis and passed over to differentiating cells and tissues. Beginning with the ESTs which were highly expressed in undifferentiated human ES cells and using homology research in mouse dbEST database, we cloned two novel putative (N (5))-glutamine methyltransferase (Hemk) splice variants termed mHemk1 and mHemk2 (Genbank accession number AY456393 and AY583759). Sequence analysis revealed that mHemk1 and mHemk2 cDNAs are 1,792 bp and 1,696 bp in length respectively. The deduced proteins have 214 amino acid residues (mHemk1) and 138 residues (mHemk2) in length and both share significant homology with (N (5))-glutamine methyltransferase (Hemk proteins) in database. Northern blot and RT-PCR analysis showed that mHemk mRNAs were abundantly expressed in undifferentiated ES cells, testis and brain, weakly expressed in differentiated ES cells and kidney, and not expressed in muscle, heart, placenta, pancreas, lung and stomach. Immunohistochemical analysis further revealed that the protein was most abundant in undifferentiated ES cells. The green fluorescent protein produced by pEGFP-C3/mHemk1 was detected mainly in the nucleus of COS7 cell lines after 24 h post-transfection. RNA interference (RNAi)-mediated knock-down method was established. Cell cycle analysis suggests that the cell proliferation decreases after RNAi with mHemk1. In vitro bioactivity assay showed that no evidence for a DNA adenine-methyltransferase activity was detected. The accumulating functional information from Hemk homology proteins in bacteria and yeast suggests that it may be an uncharacterized new mammalian N(5)-glutamine methyltransferase.


Assuntos
Células-Tronco Embrionárias/metabolismo , Metiltransferases de Proteína/genética , Animais , Células COS , Ciclo Celular/fisiologia , Processos de Crescimento Celular/fisiologia , Linhagem Celular , Cercopithecus aethiops , Clonagem Molecular , Metilação de DNA , Células-Tronco Embrionárias/citologia , Técnicas de Silenciamento de Genes , Glutamina/metabolismo , Imuno-Histoquímica , Espaço Intracelular/metabolismo , Camundongos , Microscopia de Fluorescência , Isoformas de Proteínas , Metiltransferases de Proteína/biossíntese , Metiltransferases de Proteína/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
15.
Int Wound J ; 5(3): 453-63, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18593394

RESUMO

Bone marrow stem cells participate in tissue repair processes and may have a role in wound healing. Diabetes is characterised by delayed and poor wound healing. We investigated the potential of bone marrow-derived mesenchymal stromal cells (BMSCs) to promote healing of fascial wounds in diabetic rats. After manifestation of streptozotocin (STZ)-induced diabetic state for 5 weeks in male adult Sprague-Dawley rats, healing of fascial wounds was severely compromised. Compromised wound healing in diabetic rats was characterised by excessive polymorphonuclear cell infiltration, lack of granulation tissue formation, deficit of collagen and growth factor [transforming growth factor (TGF-beta), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), platelet-derived growth factor PDGF-BB and keratinocyte growth factor (KGF)] expression in the wound tissue and significant decrease in biomechanical strength of wounds. Treatment with BMSC systemically or locally at the wound site improved the wound-breaking strength (WBS) of fascial wounds. The improvement in WBS was associated with an immediate and significant increase in collagen levels (types I-V) in the wound bed. In addition, treatment with BMSCs increased the expression of growth factors critical to proper repair and regeneration of the damaged tissue moderately (TGF-beta, KGF) to markedly (EGF, VEGF, PDGF-BB). These data suggest that cell therapy with BMSCs has the potential to augment healing of the diabetic wounds.


Assuntos
Transplante de Medula Óssea/métodos , Diabetes Mellitus Experimental/complicações , Modelos Animais de Doenças , Células Estromais/transplante , Cicatrização/fisiologia , Ferimentos Penetrantes/terapia , Análise de Variância , Animais , Fenômenos Biomecânicos , Células da Medula Óssea , Colágeno/análise , Colágeno/fisiologia , Diabetes Mellitus Experimental/induzido quimicamente , Ensaio de Imunoadsorção Enzimática , Tecido de Granulação/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/análise , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Masculino , Mesoderma/citologia , Ratos , Ratos Sprague-Dawley , Estreptozocina , Resistência à Tração , Regulação para Cima , Ferimentos Penetrantes/etiologia , Ferimentos Penetrantes/patologia
16.
J Invest Surg ; 21(5): 270-9, 2008 Sep-Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19160135

RESUMO

Bone marrow-derived stromal cells (BMSCs) exhibit extraordinary degree of plasticity and growth factor repertoire for which they have been investigated for repair and regeneration of damaged tissues, but have not been adequately examined for wound healing. The ability of BMSCs to accelerate healing of surgically inflicted cutaneous and fascial wounds was tested in vivo in rats and in vitro using a fibroblast monolayer wound model. Intravenous treatment with BMSCs augmented healing of both cutaneous and fascial wounds as determined by an increase in the biomechanical strength of wounds. In vitro experiments showed that incorporation of BMSCs in fibroblast monolayers accelerates the closure of mechanically disrupted monolayers, which was attributed to the enhanced migration of fibroblasts onto the denuded surfaces. Furthermore, culture medium conditioned by activated BMSCs promoted the closure of defects in monolayers and enhanced the proliferation/growth and directional migration (chemotaxis) of fibroblasts. This study demonstrates that BMSCs significantly augment healing of cutaneous and fascial wounds in vivo at least in part through interaction with fibroblasts in which BMSCs promote growth and chemotaxis of fibroblasts.


Assuntos
Células da Medula Óssea/patologia , Cicatrização/fisiologia , Animais , Células da Medula Óssea/fisiologia , Transplante de Medula Óssea , Linhagem Celular , Proliferação de Células , Fatores Quimiotáticos/metabolismo , Quimiotaxia , Técnicas de Cocultura , Meios de Cultivo Condicionados , Modelos Animais de Doenças , Fáscia/lesões , Fáscia/patologia , Fibroblastos/patologia , Masculino , Ratos , Ratos Sprague-Dawley , Pele/lesões , Pele/patologia , Células Estromais/patologia , Células Estromais/fisiologia , Células Estromais/transplante
17.
Wound Repair Regen ; 14(4): 471-8, 2006 Jul-Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16939576

RESUMO

Bone marrow-derived mesenchymal stromal cells (BMSCs) are multipotential stem cells capable of differentiation into numerous cell types, including fibroblasts, cartilage, bone, muscle, and brain cells. BMSCs also secrete a large number of growth factors and cytokines that are critical to the repair of injured tissues. Because of the extraordinary plasticity and the ability of syngeneic or allogeneic BMSCs to secrete tissue-repair factors, we investigated the therapeutic efficacy of BMSCs for healing of fascial and cutaneous incisional wounds in Sprague-Dawley rats. Systemic administration of syngeneic BMSCs (2 x 10(6)) once daily for 4 days or a single treatment with 5 x 10(6) BMSCs 24 hours after wounding significantly increased the wound bursting strength of fascial and cutaneous wounds on days 7 and 14 postwounding. Wound healing was also significantly improved following injection of BMSCs locally at the wound site. Furthermore, allogeneic BMSCs were as efficient as syngeneic BMSCs in promoting wound healing. Administration of BMSCs labeled with iron oxides/1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate fluorescent dye revealed that systemically administered BMSCs engraft to the wound. The increase in the tensile strength of wounds treated with BMSCs was associated with increased production of collagen in the wound. In addition, BMSC treatment caused more rapid histologic maturation of wounds compared with untreated wounds. These data suggest that cell therapy with BMSCs has the potential to augment healing of surgical and cutaneous wounds.


Assuntos
Fáscia/lesões , Mesoderma/citologia , Pele/lesões , Células Estromais/fisiologia , Cicatrização/fisiologia , Ferimentos Penetrantes/fisiopatologia , Animais , Células da Medula Óssea/fisiologia , Colágeno/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Resistência à Tração , Ferimentos Penetrantes/metabolismo
18.
World J Gastroenterol ; 9(11): 2419-23, 2003 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-14606068

RESUMO

AIM: To construct subtracted cDNA libraries of human vascular endothelial cells (VECs) related to gastrocarcinoma using suppression substractive hybridization (SSH) and to analyze cDNA libraries of gastrocarcinoma and VECs in Cancer Gene Anatomy Project (CGAP) database. METHODS: Human VECs related to gastric adenocarcinoma and corresponding normal tissue were separated by magnetic beads coupled with antibody CD31 (Dynabeads CD31). A few amount of total RNA were synthesized and amplified by SMART PCR cDNA Synthesis Kit. Then, using SSH and T/A cloning techniques, cDNA fragments of differentially expressed genes in human VECs of gastric adenocarcinoma were inserted into JM109 bacteria. One hundred positive bacteria clones were randomly picked and identified by colony PCR method. To analyze cDNA libraries of gastrocarcinoma and VECs in CGAP database, the tools of Library Finder, cDNA xProfiler, Digital GENE Expression Displayer (DGED), and Digital Differential Display (DDD) were used. RESULTS: Forward and reverse subtraction cDNA libraries of human VECs related to gastrocarcinoma were constructed successfully with SSH and T/A cloning techniques. Analysis of CGAP database indicated that no appropriate library of VECs related to carcinoma was constructed. CONCLUSION: Construction of subtraction cDNA libraries of human VECs related to gastrocarcinoma was successful and necessary, which laid a foundation for screening and cloning new and specific genes of VECs related to gastrocarcinoma.


Assuntos
Endotélio Vascular/fisiologia , Regulação Neoplásica da Expressão Gênica , Biblioteca Gênica , Neoplasias Gástricas/genética , DNA Complementar/análise , Desoxirribonucleases de Sítio Específico do Tipo II , Endotélio Vascular/citologia , Humanos , Técnicas de Amplificação de Ácido Nucleico , Reação em Cadeia da Polimerase
19.
Zhonghua Zheng Xing Wai Ke Za Zhi ; 19(4): 258-60, 2003 Jul.
Artigo em Chinês | MEDLINE | ID: mdl-14628411

RESUMO

OBJECTIVE: To detect gene mutations of p53 gene (exon 4-6) in fibroblasts. METHODS: Samples of keloids were taken from 15 patients. The mutations of p53 gene were detected using polymerase chain reaction, the single-strand conformational polymorphism(SSCP) analysis and DNA sequencing. RESULTS: Gene mutations in p53 gene exon 4, 5, and 6 were identified in all the patients with keloids. CONCLUSION: Gene mutations resulted in keloid p53 protein losing its functions of suppressing cell processes and conducting apoptosis.


Assuntos
Genes p53 , Queloide/genética , Mutação , Polimorfismo Conformacional de Fita Simples , Apoptose , Sequência de Bases , Éxons/genética , Fibroblastos , Humanos , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNA , Proteína Supressora de Tumor p53/genética
20.
Di Yi Jun Yi Da Xue Xue Bao ; 22(1): 61-3, 2002 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-12390849

RESUMO

OBJECTIVE: To detect Fas DNA mutations (exon 1-6) in the fibroblasts of patients with keloids, thereby to understand the clinicopathological implications of altered structure of the keloids. METHODS: PCR followed by single-strand conformational polymorphism analysis and direct DNA sequencing were used to detect Fas gene mutations in 15 patients with keloids. RESULTS: Insertion and point mutations were identified on the boundary between intron 5 and exon 6 in two patients, while no Fas mutations were found in the fibroblasts derived from normal skin samples of any of the patients. CONCLUSION: Nonfunction of Fas protein may be related to aberrant gene structure that codes for the transmembrane domain.


Assuntos
Queloide/genética , Receptor fas/genética , Adulto , DNA/análise , Éxons/genética , Feminino , Humanos , Masculino , Mutação , Reação em Cadeia da Polimerase , Polimorfismo Conformacional de Fita Simples , Receptor fas/análise
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