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Exosomas/metabolismo , Linfedema/terapia , Células Madre Mesenquimatosas/metabolismo , Animales , Exosomas/trasplante , Proteínas de Homeodominio/metabolismo , Humanos , Linfangiogénesis , Células Madre Mesenquimatosas/citología , Ratones , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Transducción de Señal , Cola (estructura animal)/patología , Trasplante Heterólogo , Proteínas Supresoras de Tumor/metabolismo , Proteínas de Transporte Vesicular/antagonistas & inhibidores , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismo , Gelatina de Wharton/citologíaRESUMEN
This study is to investigate the effects of puerarin on the proliferation and differentiation of umbilical cord mesenchymal stem cells (MSCs) into osteoblasts. Umbilical cord MSCs were cultured by tissue adherence and the third passage of cells was used in the experiment. The effect of puerarin on proliferation of umbilical cord MSCs was measured with MTT. The effects of puerarin on umbilical cord MSCs were evaluated by ALP immunohistochemisty and von kossa staining. The OD value decreased with the increase of puerarin concentration. On 7th day, ALP expression of puerarin group was higher than that of control group. On 14th day, ALP staining showed that the positive rate of puerarin group was higher than that of control group. Von kossa staining showed the quantity of calcium nodules was higher in puerarin group than that of control group. Puerarin can promote the umbilical cord MSCs to differentiate into osteoblasts and has an effect on the proliferation of umbilical cord MSCs.
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Humanos , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Isoflavonas , Farmacología , Células Madre Mesenquimatosas , Biología Celular , Osteoblastos , Biología Celular , Osteogénesis , Plantas Medicinales , Química , Pueraria , Química , Cordón Umbilical , Biología CelularRESUMEN
<p><b>OBJECTIVE</b>The purpose of this study was to assess the feasibility of fluorescent 2-deoxyglucose analog, 2-[N-(7-nitrobenz-2-oxa-1, 3-diaxol-4-yl)amino]-2-deoxyglucose (2-NBDG), that could be taken up by breast cancer cells highly expressing glucose transporter 1 (GLUT-1). The purpose of this study was to clarify if a fluorescent 2-deoxyglucose analog, 2-[N-(7-nitrobenz-2-oxa-1, 3-diaxol-4-yl)amino]-2-deoxyglucose (2-NBDG), can be taken up by breast cancer cells highly expressing glucose transporter 1 (GLUT-1), and to assess whether it can be used as a targeting imaging agent.</p><p><b>METHODS</b>The expressions of GLUT-1 mRNA and protein in breast cancer MDA-MB-231 cells were detected by RT-PCR and immunohistochemistry, respectively. The difference of GLUT-1 protein expression between breast cancer MDA-MB-231 cells and MCF-7 cells was compared by Western blot. Secondly, MDA-MB-231 cells which were grown in 6-well plates were incubated with 2-NBDG, and the result of 2-NBDG uptake was analyzed by fluorescence microscopy and flow cytometry. The difference of 2-NBDG absorption in MDA-MB-231 and MCF-7 cells was compared by flow cytometry.</p><p><b>RESULTS</b>The results of RT-PCR and immunohistochemistry confirmed that MDA-MB-231 cells highly expressed GLUT-1. Furthermore, Western blot revealed that GLUT-1 expression of MDA-MB-231 cells (0.946 ± 0.007) was higher than that in the MCF-7 cells (0.833 ± 0.010). Fluorescence microscopic and flow cytometric analysis showed that 2-NBDG was uptaken rapidly by MDA-MB-231 cells. Addition of 50 mmol/L D-glucose to the media with 2-NBDG reduced its uptake by 46.0%. Moreover, flow cytometry indicated that the fluorescence intensity of MDA-MB-231 cells (25.10 ± 0.57) was higher than that of MCF-7 cells (10.12 ± 0.62) when incubated with 2-NBDG for 20 minutes.</p><p><b>CONCLUSION</b>The preliminary data clearly demonstrate that 2-NBDG is taken up and accumulated in breast cancer cells that highly express GLUT-1, and may be used as an optical probe for glucose uptake in hypermetabolic malignant cells.</p>
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Femenino , Humanos , 4-Cloro-7-nitrobenzofurazano , Farmacocinética , Western Blotting , Neoplasias de la Mama , Metabolismo , Patología , Línea Celular Tumoral , Desoxiglucosa , Farmacocinética , Citometría de Flujo , Transportador de Glucosa de Tipo 1 , Genética , Metabolismo , Inmunohistoquímica , ARN Mensajero , Metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The purpose of this study was to investigate the growth characteristics and the expression level of integrin mRNA of the cultured bone marrow mesenchymal stem cells (BMMSCs) from patients with chronic myeloid leukemia (CML) in myeloid crisis (MC), and explore the role of BMMSCs in pathogenesis of CML. Five CML patients were enrolled in experimental group, five healthy persons were used as control. BMMSCs were cultured in vitro. The morphology of BMMSCs was observed every day and the growth curve were portrayed, and the ability of cell proliferation were detected according to the daily results of cell counting. Total RNA was extracted from third and fourth passages of BMMSCs, The expression of integrins mRNA of BMMSCs were measured by real-time PCR. The results showed that the BMMSCs of experimental and control groups had no difference in growth characterisctics, but the expression of integrins mRNA of the BMMSCs was higher in CML patients than in normal control group (p < 0.05). It is concluded that the abnormally high expression of integrins of BMMSC from the CML patients take part in pathogenesis of CML.
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Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Crisis Blástica , Metabolismo , Células de la Médula Ósea , Metabolismo , Patología , Proliferación Celular , Integrinas , Genética , Metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva , Metabolismo , Patología , Células Madre Mesenquimatosas , Metabolismo , Patología , ARN Mensajero , Genética , Metabolismo , Células Tumorales CultivadasRESUMEN
<p><b>OBJECTIVE</b>To study the methylation status of fragile histidine triad (FHIT) gene promoter in patients with myelodysplastic syndrome (MDS) and its clinical relevance.</p><p><b>METHODS</b>Methylation-specific PCR (MSP) was used to detect FHIT promoter methylation in bone marrow samples from 54 MDS cases.</p><p><b>RESULTS</b>Hypermethylation of FHIT promoter was detected in 26 cases (48.1%). Association was not found between FHIT gene hypermethylation and sex, hematologic parameters and chromosomal abnormalities of MDS patients, but found between FHIT gene hypermethylation and age of the MDS cases. Although significant difference was not observed in the frequencies of FHIT gene hypermethylation among patients with refractory anemia/refractory anemia with ringed sideroblasts (RA/RAS) (1/6, 16.7%), refractory anemia/refractory anemia with ringed sideroblasts (RCMD) and refractory cytopenia with multilineage dysplasia with ringed blasts (RCMD-RS) (6/19, 31.6%), refractory anemia with excess blasts-1 (RAEB-1) (7/11, 63.6%), refractory anemia with excess blasts-2 (RAEB-2) (4/7, 57.1%) and refractory anemia with excess blasts in transformation/acute myeloid leukemia (RAEBt/AML) (8/11, 72.7%)(chi-square=8.417, P=0.077), it was observed in patients in early stages (RA/RAS and RCMD) (7/25, 28.0%), advanced stages (RAEB-1 and RAEB-2)(11/18, 61.1%) and RAEBt/AML (8/11, 72.7%) (chi-square=7.938, P=0.019). Furthermore, there was a positive correlation between the frequency of FHIT gene hypermethylation and different IPSS groups (chi-square=10.110, P=0.018).</p><p><b>CONCLUSION</b>FHIT gene hypermethylation might be one of the molecular events involved in the disease progression of MDS.</p>
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Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Ácido Anhídrido Hidrolasas , Genética , Factores de Edad , Secuencia de Bases , Metilación de ADN , Datos de Secuencia Molecular , Síndromes Mielodisplásicos , Clasificación , Genética , Patología , Proteínas de Neoplasias , Genética , Reacción en Cadena de la Polimerasa , Regiones Promotoras Genéticas , GenéticaRESUMEN
<p><b>AIM</b>To investigate the expression of recombinant human phosphodiesterase 3A (HPDE3A) using baculovirus expression system in Tn cell line.</p><p><b>METHODS</b>The HPDE3A cDNA was recombined with baculovirus, and then the recombinant was transfected into Tn cell line. The expression of HPDE3A in Tn cell line was detected and identified by the RT-PCR, SDS-PAGE and Western blotting.</p><p><b>RESULTS</b>The recombinant HPDE3A protein was stably expressed in Tn cell line and detected by the distinct morphological changes of Tn cell, RT-PCR, SDS-PAGE and Western blotting using polyclonal antibody. The M(w) of the recombinant protein was about 120 kD.</p><p><b>CONCLUSION</b>Recombinant HPDE3A can be expressed in Tn cell line using the baculovirus expression system, and thus provided the basic material for studying its bioactivity and application in screening for HPDE3A inhibitor.</p>
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Animales , 3',5'-AMP Cíclico Fosfodiesterasas , Genética , Metabolismo , Baculoviridae , Genética , Línea Celular , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 3 , Electroforesis en Gel de Poliacrilamida , Mariposas Nocturnas , Biología Celular , Metabolismo , ARN Mensajero , Genética , Metabolismo , Proteínas Recombinantes , Genética , Metabolismo , TransfecciónRESUMEN
Objective To establish a method of labeling human mesenchymal stem cells (MSCs) with PKH26 in vitro.Methods MSCs were cultured and labeled with PKH26 according to the manufacturer's instruction.The growth,fluorescence intensity and serial subcuhivation of labeled MSCs were analyzed with the confocal laser microscope and the flow cytometry.The biological characteristics of labeled MSCs were investigated by RT-PCR.Results The labeled MSCs appeared red fluorescence and the labeling rate was 100 percent.During serial subcuhivation of labeled MSC from passage 1 to passage 7,the fluorescence intensity and the labeling rate of MSCs were gradually decreased.The biological features such as morphology,growth,expression level of nucleostemin and GAPDH gene and capability of differentiation into osteoblast in vitro were not affected by labeling.Conclusion Labeling the human MSCs with PKH26 is an effective and practical method,which can be used as an important tool in the study on the homing, plasticity and transplantation of MSCs.