RESUMEN
Human mesenchymal stem cells (MSCs) can be isolated from various tissues. However, the cytokine profile in different MSC types remains unclear. In this study, MSCs were extracted from adipose, umbilical cord, and placental tissues. The surface marker expression, multilineage differentiation potential, and cytokine secretion of these cells were compared. The isolated MSCs exhibited similar morphology and surface marker expression. However, they differed with regard to their differentiation potential. Adipose-MSCs (A-MSCs) exhibited a higher potential for adipogenesis and osteogenic differentiation compared with umbilical cord-MSCs (UC-MSCs) and placental-MSCs (P-MSCs). The expression levels of 80 cytokines were detected, and the data demonstrated that the three MSC types abundantly secreted insulin-like growth factor-binding protein (IGFBP)-4, IGFBP-3, tissue inhibitor of metalloproteinase (TIMP)-1, TIMP-2, IGFBP-6, monocyte chemoattractant protein-1, and granulocyte colony-stimulating factor. However, the expression levels of vascular endothelial growth factor, tumor necrosis factor alpha, interleukin (IL)-6 receptor, and IL-13 in A-MSCs were higher compared with those of UC-MSCs and P-MSCs. Moreover, the expression levels of intercellular adhesion molecule-1 and growth differentiation factor 15 were lower in A-MSCs. Kyoto Encyclopedia of Genes and Genomes analysis indicated that the "adipocytokine" and the "PI3K/Akt pathways" were enriched in A-MSCs. Taken together, the results demonstrated that MSCs from different sources exhibited differences in the secretion of specific factors. A-MSCs were associated with the expression of several proangiogenic factors and may be an improved source for angiogenesis and tissue regeneration.
Asunto(s)
Células Madre Mesenquimatosas , Osteogénesis , Diferenciación Celular , Células Cultivadas , Citocinas , Femenino , Humanos , Fosfatidilinositol 3-Quinasas , Placenta , Embarazo , Cordón Umbilical , Factor A de Crecimiento Endotelial VascularRESUMEN
Transplantation of mesenchymal stromal cells (MSCs) improves functional recovery in experimental models of spinal cord injury (SCI), but the mechanism is not fully understood. Activation of procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2), a collagen-modifying enzyme, reportedly follows MSC transplantation in an SCI animal model. We investigated the regulation of PLOD2 expression and its potential contribution to the neuroprotective effects of adipose-derived stromal cells (ADSCs) following mechanical injury to neurons in vitro and SCI in vivo. ADSCs enhanced wound healing in vitro and promoted functional recovery after their implantation near injury sites in a rat SCI model. These effects correlated with upregulation of PLOD2, MAP2, NSE and GAP43, and downregulation of GFAP, which is indicative of improved neuronal survival and axonal regeneration as well as reduced glial scar formation. The neurorestorative effect of ADSCs was weakened after inhibition of PLOD2 expression. ADSCs appeared to induce PLOD2 upregulation via TGF-ß1 secretion, as ADSC-mediated PLOD2 expression, neuronal survival, and functional recovery after SCI were largely prevented by SB431542, a TGF-(1 receptor inhibitor. These findings indicate that ADSCs reduce lesion size and promote functional recovery after SCI mainly through activation of a TGF-ß1/P-Samd3/PLOD2 pathway in spinal cord neurons.
Asunto(s)
Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Neuronas/metabolismo , Procolágeno-Lisina 2-Oxoglutarato 5-Dioxigenasa/genética , Recuperación de la Función , Proteína smad3/genética , Traumatismos de la Médula Espinal/genética , Factor de Crecimiento Transformador beta1/genética , Adipocitos , Adipogénesis , Animales , Técnicas In Vitro , Osteocitos , Osteogénesis , Células PC12 , Procolágeno-Lisina 2-Oxoglutarato 5-Dioxigenasa/metabolismo , Ratas , Proteína smad3/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/terapia , Factor de Crecimiento Transformador beta1/metabolismoRESUMEN
BACKGROUND: Neonatal hypoxia ischemia causes severe brain damage. Stem cell therapy is a promising method for treating neuronal diseases. Clinical translation of human umbilical cord-derived mesenchymal stem cells (UC-MSCs) for the recovery of neurons after hypoxic ischemic encephalopathy (HIE) may represent an effective therapy. METHODS: Primary neurons were exposed to oxygen-glucose deprivation (OGD) and subsequently cocultured with UC-MSCs. Apoptosis was examined by Annexin V-FITC-PI. Genes related to apoptosis were detected using RT-PCR and western-blot analyses. Using an in vivo model, HIE was induced in postnatal day 7 mice, and UC-MSCs were transplanted via the intraventricular route. UC-MSC migration was investigated by immunofluorescence, and lesion volumes were measured by TTC staining. Apoptosis in injured brain cells was detected by the TUNEL assay. RT-PCR and ELISA were used to detect the expression of inflammatory factors in cells and animal tissues. RESULTS: Flow cytometry analysis revealed that apoptosis in injured neurons was inhibited by UC-MSCs. The RT-PCR and western blot results indicated that coculture inhibited the expression of proapoptotic genes and upregulated expression of antiapoptotic genes. In the animal model, transplanted UC-MSCs migrated toward the cerebral lesion site and decreased the lesion extent in HIE. TUNEL staining showed that the MSC group exhibited significantly reduced numbers of TUNEL-positive cells. RT-PCR and ELISA showed that UC-MSCs inhibited the upregulation of TNF-α and IL-1ß in response to hypoxic ischemic injury. CONCLUSION: These results indicate that UC-MSCs exert neuroprotective effects against hypoxic ischemic injury by inhibiting apoptosis, and the mechanism appears to be through alleviating the inflammatory response.
Asunto(s)
Apoptosis/fisiología , Hipoxia-Isquemia Encefálica/etiología , Células Madre Mesenquimatosas/fisiología , Neuroprotección , Animales , Animales Recién Nacidos , Células Cultivadas , Femenino , Masculino , Ratones , Ratas , Ratas Wistar , Cordón Umbilical/citologíaRESUMEN
This study was aimed to investigate the change of cell phenotype and the expression of hematopoiesis associated cytokines in umbilical cord mesenchymal stem cells (UC-MSC) in three-dimensional (3-D) system. MSC were isolated from umbilical cord, and then cultured in 2-D and 3-D system respectively. The phenotype of MSC was detected by flow cytometry; the angiogenic capability of MSC cultured in 2-D and 3-D syitem was assessed using in vitro capillary formation assay. The cytokine expression of MSC in two kinds of culture conditions was measured by real-time PCR. The results showed that MSC were successfully isolated from umbilical cord. Flow cytometry showed that the percentage of CD31, CD133 and CD271 expressed in endothelial cells, endothelial progenitor cells and primitive mesenchymal stem cells increased significantly in 3-D culture conditions, as compared to 2-D system. Capillary formation assay showed that the angiogenic capability of UC-MSC was greatly enhanced. Quantitative PCR showed that the expression of ß-actin was upregulated in 3-D system. The expression of some cytokines associated with hematopoiesis, such as G-CSF, LIF, SCF, IL-1α, IL-1ß, IL-3, IL-7 and IL-11, increased, especially for LIF, IL-3, IL-7. The expression of IL-10 associated with immune regulation also increased. The expression of SDF-1, IL-6 slightly decreased, but without significant difference. It is concluded that expression of CD31, CD133 and CD271 increases in 3-D system, the angiogenic capability of UC-MSC enhances and the expression of hematopoiesis-associated cytokines in UC-MSC increases in 3-D system.
Asunto(s)
Citocinas/biosíntesis , Células Madre Mesenquimatosas/metabolismo , Cordón Umbilical/metabolismo , Actinas , Células Cultivadas , Citometría de Flujo , Hematopoyesis , Humanos , Fenotipo , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: Gastric cancer is a leading causes of cancer-related deaths ,but the underlying molecular mechanisms of its progression are largely unknown. Differentiated embryonic chondrocyte-expressed gene 1 (DEC1), is an important transcription factor involved in the progression of tumors and has recently been identified to be strongly inducible by hypoxia. Little is known about the contribution of DEC1 to the intracellular hypoxia and proliferation signaling events in gastric cancer. METHODS: Immunohistochemistry was used to detect the expression of DEC1, hypoxia-inducible factor 1(HIF-1α) and Ki67 in 173 human gastric cancer samples and adjacent non-tumor tissues samples. The relationship between DEC1, HIF-1α and Ki67 was evaluated. RESULTS: DEC1 protein was persistently expressed in the nucleus and cytoplasm of gastric cancer tissue. The protein expression of DEC1 and HIF-1α in tumour tissues was 83.8% and 54.3%, respectively, and was significantly higher than that in adjacent normal tissues (83.8% vs 23.7%, P < 0.001; 54.3% vs 12.7%, P < 0.001). The expression of DEC1 and HIF-1α was associated with poor histological differentiation. (P < 0. 01). Furthermore, DEC1 level was positively correlated with HIF-1α (P < 0. 01, r=0.290) and Ki67 expression (P < 0. 01, r=0.249). CONCLUSION: The upregulation of DEC1 may play an important role in hypoxia regulation and cell proliferation in gastric cancer. The relevant molecular mechanism requires further investigation.
Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/análisis , Proteínas de Homeodominio/análisis , Subunidad alfa del Factor 1 Inducible por Hipoxia/análisis , Antígeno Ki-67/análisis , Neoplasias Gástricas/química , Adulto , Anciano , Anciano de 80 o más Años , Diferenciación Celular , Hipoxia de la Célula , Núcleo Celular/química , Proliferación Celular , Citoplasma/química , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Neoplasias Gástricas/patologíaRESUMEN
OBJECTIVE: To investigate the genetic polymorphisms of 19 STR Loci in Shandong Han population in order to provide the genetic data for paternity testing. METHODS: The genotypes of 205 unrelated individuals in Shandong Han population were typed by Goldeneye 20A kit to get the allele frequencies and population genetic parameters of 19 STR loci. Four kits, Identifiler kit, SinoFiler kit, PowerPlex 16 kit, and Goldeneye 20A kit, were compared with each other and used in the analysis of a special paternity test case. RESULTS: The population genetic parameters of 19 STR loci in Shandong Han Population were obtained. The cumulative discrimination power (CDP) and cumulative probability of exclusion (CPE) ranked from high to low were Goldeneye 20A kit, SinoFiler kit, PowerPlex 16 kit and Identifiler kit, respectively. As duo case, the result of the real case showed that Identifiler kit had no excluding loci, and none of the SinoFiler kit, PowerPlex 16 kit or Goldeneye 20A kit could exclude fatherhood. CONCLUSION: Compared with Identifiler kit, SinoFiler kit, and PowerPlex 16 kit, Goldeneye 20A kit shows the higher efficiency than the others, but is not completely satisfied for duo cases.
Asunto(s)
Pueblo Asiatico/genética , Genética de Población , Repeticiones de Microsatélite , Paternidad , Polimorfismo Genético/genética , China , Genética Forense/métodos , Frecuencia de los Genes , Sitios Genéticos/genética , Genotipo , Humanos , MasculinoRESUMEN
OBJECTIVE: To evaluate the potential usefulness of a multivariable model in predicting the response to immunosuppressive therapy (IST) in patients with aplastic anemia (AA), and its application to the clinical practice. METHODS: PB T cells subpopulation and BM T cells intracellular IFN-gamma and IL-4 were serially analyzed by flow cytometry (FCM) before and during treatment. HLA-DRB1 * 1501 phenotype was analyzed by PCR-SSP. The predictive potentials of different parameter combinations for clinical responsiveness were statistically assessed. RESULTS: In all evaluated parameters, CD8+ cell intracellular IFN-gamma had the relatively best diagnostic value with sensitivity and specificity of 94.3% and 62.5%, and positive and negative predictive value of 84.6% and 83.3% respectively. Positive CD8+ cell intracellular IFN-gamma plus Tc1/Tc2 < 50 could increase the positive predictive value to 92.3%. A multivariable model consisting of absolute neutrophil count (ANC), BM T cell intracellular IFN-gamma, Tc1/Tc2 ratio and HLA-DRB * 1501 phenotype of the patients was finally established. CONCLUSION: The multivariable model is superior to each of the single parameters in terms of predictive power of IST therapeutic outcome, and its higher accuracy and the clinical application make it potentially useful in practice.
Asunto(s)
Anemia Aplásica/tratamiento farmacológico , Terapia de Inmunosupresión , Modelos Estadísticos , Adolescente , Adulto , Anciano , Anemia Aplásica/inmunología , Niño , Estudios de Factibilidad , Femenino , Antígenos HLA-DR/inmunología , Humanos , Inmunosupresores/uso terapéutico , Masculino , Persona de Mediana Edad , Subgrupos de Linfocitos T/inmunología , Linfocitos T/inmunología , Resultado del TratamientoRESUMEN
OBJECTIVE: To evaluate the clinical implication of combined measurement of bone marrow (BM) T lymphocyte intracellular IFNgamma with HLA-DRB1*1501 in predicting the response to immunosuppressive therapy (IST) in patients with aplastic anemia (AA). METHODS: Enrolled into the present study were 51 idiopathic AA patients treated with cyclosporine A (CsA) based IST. BM CD(8)(+) T lymphocyte intracellular IFNgamma was determined with flow cytometry and HLA-DRB1*1501 detected with PCR-sequence specific primer before treatment. The relationship between laboratory indices and clinical response were investigated and the potential usefulness of parameters in predicting the response to IST for AA was evaluated. RESULTS: These HLA-DRB1*1501 shows sensitivity of 45.7% (16/35) and specificity of 87.5% (14/16) respectively. Intracellular IFNgamma has sensitivity of 94.3% (33/35) and specificity of 62.5% (10/16), respectively. With combination of intracellular IFNgamma with HLA-DRB1*1501, the parallel test increases the sensitivity of 97.1% (34/35) and the negative predictive value of 90.0% (9/10). On the other hand, the serial test improves the specificity and positive predictive value which both achieve 93.7% (15/16). It could be calculated through a logistic regression equation that the probabilities of prediction of four subgroups of patients whose results are both positive reaction, a positive intracellular IFNgamma plus negative HLA-DRB1*1501, a negative intracellular IFNgamma plus positive HLA-DRB1*1501 and both negative reaction are 89.0%, 77.4%, 34.5% and 18.2%, respectively. CONCLUSIONS: Combination of BM T cells intracellular IFNgamma stain and HLA-DRB1*1501 phenotype can be a useful predictor for AA patients in immunosuppressive therapy. The patients with both positive results of the two tests may have more possibilities to response to IST. It may have an important implication for the majority of AA patients whose intracellular IFNgamma stain has a positive reaction.
Asunto(s)
Anemia Aplásica/tratamiento farmacológico , Células de la Médula Ósea/efectos de los fármacos , Antígenos HLA-DR/análisis , Inmunosupresores/uso terapéutico , Interferón gamma/análisis , Adolescente , Adulto , Anciano , Anemia Aplásica/sangre , Células de la Médula Ósea/metabolismo , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/metabolismo , Niño , Ciclosporina/uso terapéutico , Femenino , Citometría de Flujo , Antígenos HLA-DR/genética , Cadenas HLA-DRB1 , Humanos , Modelos Logísticos , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Valor Predictivo de las Pruebas , Resultado del TratamientoRESUMEN
To investigate the pathophysiological mechanisms for platelet-neutrophils cross talk mediated by platelet-activating factor (PAF) and to lay a foundation for clinical application, ginkgolides B (GB), a PAF receptor antagonist, was added in the whole blood to block the effects of PAF on activation of platelet-neutrophil; PAF and ADP were respectively added in the whole blood to monitor the expression of CD62P on platelet by flow cytometry; PAF and ADP were added in the whole blood to monitor the expression of CD11b on neutrophil by flow cytometry; PAF and ADP were added in the whole blood to monitor the platelet-leucocyte aggregates (PLA) which were PLA in the total leucocyte population (PLA/L) and the mean fluorescence intensity (MFI) of CD42b. Outcomes were analyzed by t-test, and the differences were statistically significant (P < 0.05). The results showed that the expression of CD62P on platelats, the expression of CD11b on neutrophils and PLA formation were all increased by PAF and ADP; the PAF receptor antagonists (GB) could obviously inhibit the expression of CD62P, CD11b and PLA formation induced by PAF, but could not completely inhibit the activation of platelet and neutrophil, and the platelet-neutrophil cross talk; GB could inhibit the expression of CD62P and CD11b induced by ADP, but could not conpletely inhibit the activation of platelet and neutrophli; GB could not obviously inhibit the platelet-leucocyte aggregates mediated by ADP. It is concluded that the multiligand-receptor systems involved in PLA formation and platelet-netrophils cross talk seem to be regulated by complex mechanisms; the PAF receptor antagonists (GB) obviously inhibit the effect of PAF, and may be widely utilized in the therapy of thrombosis and inflammation.
Asunto(s)
Plaquetas/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Factor de Activación Plaquetaria/farmacología , Adenosina Difosfato/farmacología , Adulto , Plaquetas/citología , Plaquetas/fisiología , Antígeno CD11b/sangre , Adhesión Celular/efectos de los fármacos , Adhesión Celular/fisiología , Comunicación Celular/efectos de los fármacos , Comunicación Celular/fisiología , Femenino , Citometría de Flujo , Ginkgólidos/farmacología , Humanos , Masculino , Persona de Mediana Edad , Neutrófilos/citología , Neutrófilos/fisiología , Selectina-P/sangre , Activación Plaquetaria/efectos de los fármacos , Complejo GPIb-IX de Glicoproteína Plaquetaria/análisis , Glicoproteínas de Membrana Plaquetaria/antagonistas & inhibidores , Glicoproteínas de Membrana Plaquetaria/fisiología , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/fisiologíaRESUMEN
To discuss the false-positive of serological diagnostic testing for coronavirus antibody in patients with systemic lupus erythematosus(SLE), 66 normal individual and 31 SLE with non-SARS patients were detected for SARS-associated coronavirus (SARS-CoV) antibody and RNA by enzymelinked immunosorbent assays(ELISA) and reverse transcriptase-polymerase chain reaction(RT-PCR). The result showed 2/66 cases(3.0%) were positive of SARS-CoV-IgG antibody and 66 cases were negative of SARS-CoV-IgM antibody in the 66 cases healthy controls; in 31 cases with SLE, positive rates of SARS-CoV-IgG and IgM antibody were 58.1% (18/31) and 29% (9/31), respectively, in which 7 cases(22.6%) were positive of both SARS-CoV-IgG and IgM antibody. All samples of positive SARS-CoV-IgG and IgM antibody were negative by RT-PCR. The ELISA kit coated by non-purification antigen may induce the false-positive of SARS-CoV antibody in patients with SLE. This result suggested that the specificity of ELISA tests for SARS was excellent and has low false-positive rates when using SARS-CoV-IgG and IgM antibody tests. A possible cause of false-positive of SARS-CoV-IgG and IgM antibody in SLE patients is coated antigens with SARS-CoV and Vero-E6 cells in ELISA methods.