RESUMO
Relapse remains the leading cause of death in acute myeloid leukemia (AML) patients following allogeneic hematopoietic stem-cell transplantation (allo-HSCT), limiting the efficacy of allo-HSCT. Thus, the ability to identify high-risk patients in a manner that permits early intervention has the potential to improve survival outcomes. We retrospectively enrolled 414 younger patients (aged 14-60 years) with AML who received allo-HSCT between January 2014 and May 2020. From June 2020 to June 2021, 110 consecutive patients were included prospectively in the validation cohort. The primary outcome was early relapse (relapse within 1 year). The cumulative incidence of early relapse after allo-HSCT was 11.8%. The overall survival rate for patients who relapsed within 1-year was 4.1% at 3 years after relapse. After multivariable adjustment, statistically significant associations between primary resistance, pre-transplantation measurable residual disease, DNMT3A mutation, or white blood cell count at diagnosis and early relapse were observed. An early relapse prediction model was developed based on these factors and the model performed well. Patients deemed to have a high risk or a low risk of early relapse had early relapse rates of 26.2% and 6.8%, respectively (P < 0.001). The prediction model could be used to help identify patients at risk for early relapse and to guide personalized relapse prevention.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Humanos , Estudos Retrospectivos , Transplante Homólogo , Doença Crônica , Recidiva , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Leucemia Mieloide Aguda/genéticaRESUMO
BACKGROUND: Minimal residual disease (MRD) is an important prognostic factor for survival in adults with acute leukemia. The role of pretransplantation MRD status in myelodysplastic syndrome with excess blasts (MDS-EB) is unknown. This study retrospectively analyzed the relationship between pretransplantation MRD status and long-term survival. MATERIALS AND METHODS: Patients with MDS-EB who underwent allogeneic hematopoietic stem cell transplantation (allo-HSCT) from March 5, 2005, to November 8, 2020, were included. The relationship between pretransplantation MRD status and long-term survival was analyzed using univariate and multivariate logistic regression models. RESULTS: Of 220 patients with MDS-EB who underwent allo-HSCT, 198 were eligible for inclusion in this multicenter, retrospective cohort study. Complete remission was attained in 121 (61.1%) patients, and 103 patients underwent detection of MRD pretransplantation, with 67 patients being MRD-positive and 36 patients being MRD-negative. The median follow-up time was 16 months, the median age was 41 years (6-65 years), and 58% of the patients were men. The 3-year disease-free survival (DFS) and overall survival (OS) probabilities for all patients were 70.1% and 72.9%, respectively. For patients in complete remission, the 3-year DFS and OS probabilities were 72.2% and 74.8%, respectively. Further analysis found that the 3-year DFS rates of MRD-negative and MRD-positive patients were 85.6% and 66.5% (p = .045), respectively, whereas the 3-year OS rates were 91.3% and 66.4% (p = .035), respectively. Univariate and multivariate analyses showed that poor pretransplantation MRD clearance was an independent prognostic risk factor for DFS and OS. CONCLUSION: Poor pretransplantation MRD clearance is an independent prognostic risk factor for long-term survival after allo-HSCT for patients with MDS-EB. PLAIN LANGUAGE SUMMARY: Poor minimal residual disease clearance pretransplanation is an independent prognostic risk factor for long-term survival after allogeneic hematopoietic stem cell transplantation for patients with myelodysplastic syndrome with excess blasts.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Síndromes Mielodisplásicas , Adulto , Masculino , Humanos , Feminino , Prognóstico , Estudos Retrospectivos , Neoplasia Residual/diagnóstico , Síndromes Mielodisplásicas/terapia , Fatores de RiscoRESUMO
The use of Bcl-2 inhibitor Venetoclax (VEN) combined with hypomethylating agents or chemotherapy has shown efficacy in treating acute myeloid leukemia (AML) as frontline treatment and for relapse, allowing more patients to bridge to allogeneic hematopoietic stem cell transplantation (allo-HSCT). However, the influence of VEN-based therapy on the prognosis of subsequent allogeneic HSCT remains unknown. We retrospectively collected data from patients who proceeded to allo-HSCT between November 2018 and November 2020 after VEN-based therapy at five transplant centers in Zhejiang Province, China. A total of 39 patients were analyzed. Thirty-one patients were diagnosed with AML (28 de novo, 3 secondary to MDS), 6 with MDS, and 2 with CMML. The majority (74.4%) of patients received VEN-based therapy for the treatment of relapse (38.5%) or refractory disease (35.9%); 5 (12.8%) received it as an initial treatment, and 5 (12.8%) patients who were already in complete remission (CR) received VEN for further consolidation or deep remission before HSCT. Twenty-seven (69.2%) patients were in CR at the time of HSCT. Day + 100 cumulative incidences of grade I-IV acute graft-versus-host disease (aGVHD) and grade II-IV aGVHD were 43.6% and 15.4%, respectively. Of 34 evaluable patients, 6.4% and 25.6% developed chronic GVHD at 1 year and 2 years. The 100-day cytomegalovirus (CMV) reactivation occurred in 76.3% of patients and Epstein-Barr virus (EBV) reactivation occurred in 29.7% of patients. With a median follow-up of 14.7 months, overall survival, progression-free survival, relapse, and non-relapse mortality incidence at 1 year were 75.5%, 61.6%, 16.7%, and 21.7%, respectively. Both univariate and multivariate analysis revealed that relapsed/refractory (R/R) disease was associated with inferior PFS (HR 4.849, 95% CI 1.009-23.30; p = 0.049). Prior poor response to VEN was found to be a significant factor predicting higher risk of relapse (HR 4.37, 95% CI 1.130-16.9; p = 0.033). Our results showed that VEN-based regimen therapy followed by allo-HSCT in AML patients is feasible and does not increase the risk of transplant-related mortality and toxicity.
Assuntos
Infecções por Vírus Epstein-Barr , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Leucemia Mieloide Aguda , Humanos , Estudos Retrospectivos , Infecções por Vírus Epstein-Barr/complicações , Herpesvirus Humano 4 , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/complicações , Doença Enxerto-Hospedeiro/etiologia , RecidivaRESUMO
Minimal residual disease (MRD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT) heralds high risk of relapse. Whether preemptive recombinant interleukin-2 (pre-IL2) is effective for patients with late-onset MRD (LMRD) remains unknown. We retrospectively compared the efficacy and safety of pre-IL2 (n = 30) and pre-DLI (n = 25) for LMRD in patients receiving allo-HSCT for acute leukemia or myelodysplastic syndrome. The 1-year overall survival (OS) and disease-free survival (DFS) rates were 86.7% and 78.4% (P = 0.267), 83.3% and 75.6% (P = 0.329), the cumulative incidence of grades III-IV acute graft-versus-host disease (aGVHD) at 100 days post-preemptive intervention was 3.3% and 12.0% (P = 0.226) in the pre-IL2 group and pre-DLI group, respectively. The 1-year cumulative incidence of moderate/severe chronic GVHD (cGVHD), relapse (CIR), and non-relapse mortality (NRM) were 7.7% and 27.9% (P = 0.018), 13.6% and 20.0% (P = 0.561) and 3.3% and 5.5% (P = 0.321) in the two groups, respectively. No remarkable differences in CIR, OS, and DFS between the two intervention groups were found in multivariate analysis. The GVHD-free and relapse-free survival (GRFS) were better in the pre-IL2 group than in the pre-DLI group (HR = 0.31, 95% confidence interval (CI), 0.12-0.76; P = 0.011). In conclusion, preemptive low-dose IL2 and preemptive DLI yield comparable outcomes for patients with LMRD receiving allo-HSCT, in terms of aGVHD, NRM, relapse, OS, and DFS. However, preemptive low-dose IL2 has a lower incidence of moderate/severe cGVHD and a higher CRFS. Preemptive low-dose IL2 may be an alternative method for patients who develop LMRD after allo-HSCT, particularly for patients who cannot receive preemptive DLI.
Assuntos
Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Interleucina-2/administração & dosagem , Leucemia , Transfusão de Linfócitos , Síndromes Mielodisplásicas , Doença Aguda , Adolescente , Adulto , Aloenxertos , Criança , Doença Crônica , Intervalo Livre de Doença , Feminino , Doença Enxerto-Hospedeiro/sangue , Doença Enxerto-Hospedeiro/mortalidade , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Incidência , Leucemia/sangue , Leucemia/mortalidade , Leucemia/terapia , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/sangue , Síndromes Mielodisplásicas/mortalidade , Síndromes Mielodisplásicas/terapia , Neoplasia Residual , Estudos Retrospectivos , Taxa de SobrevidaRESUMO
BACKGROUND: A consensus has been reached that carbapenem-resistant Enterobacteriaceae (CRE) screening in immunosuppressed individuals can reduce the incidence of CRE bloodstream infection (BSI). METHODS: We retrospectively studied the clinical data of 395 consecutive HSCT patients from September 2017 to April 2019. From September 2017 to June 2018 (period 1), 200 patients received single CRE screening before transplantation. From July 2018 to April 2019 (period 2), 195 patients received continuous weekly CRE screening after admission. For patients colonized with CRE, targeted managements were received: (1) contact precautions and (2) preemptive CRE-targeted treatment if necessary. RESULTS: During period 1, 3 patients with CRE colonization were detected (1.5%). The CRE BSI rate was 2.0% (4 patients), and the related 30-day mortality was 50.0% (2 out of 4 patients). During period 2, 21 patients with CRE colonization were detected, and the detection rate was significantly higher than that in period 1 (P < 0.001). Of the 21 colonized patients, 4 (19.0%) patients were identified as positive for CRE at the first screening, 5 (23.8%) were identified at the second screening, and the remaining 12 (57.1%) were identified at the third or later screening. The CRE BSI rate decreased to 0.5% (1/195), and there were no CRE-related death. Fifteen colonized patients developed neutropenic fever. Thirteen colonizers were preemptively treated with tigecycline within 24 h of fever onset, and they achieved rapid temperature control. One colonizer received tigecycline later than 48 h after fever onset and ultimately survived due to the addition of polymyxin. The other received tigecycline later than 72 h after fever onset and died of septic shock. CONCLUSION: The increase in screening frequency contributed to the detection of patients with CRE colonization. Targeted managements for these colonized patients may contribute to reducing the incidence and mortality of CRE BSI, therefore improving the prognosis of patients.
Assuntos
Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , Infecções por Enterobacteriaceae/diagnóstico , Transplante de Células-Tronco Hematopoéticas/mortalidade , Polimixinas/uso terapêutico , Tigeciclina/uso terapêutico , Adolescente , Adulto , Idoso , Antibioticoprofilaxia , Criança , Diagnóstico Precoce , Infecções por Enterobacteriaceae/tratamento farmacológico , Infecções por Enterobacteriaceae/microbiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mortalidade , Admissão do Paciente , Estudos Retrospectivos , Resultado do Tratamento , Adulto JovemRESUMO
OBJECTIVE: To explore the mechanism of Shangke Jiegu Tablet (SJT)in repairing the mandibular defect. METHODS: Totally 72 healthy male New Zealand rabbits were randomly divided into the normal control group (n = 24), the model group (n = 24), and the SJT group (n = 24). Then the mandibular defect model was established. Animals in the normal control group and the model group were fed with normal forage, while those in the SJT group were fed with SJT forage. On the day 7, 14, 28, and 56 after model establishment, 6 rabbits were killed in each group. The bone was collected from the mandibular defect. The gene expressions of osteoprotegerin (OPG) and osteoprotegerin ligand (OPGL) were detected by means of RT-PCR. The positive dyeing strength and area of the bone tissue were detected by means of immunohistochemical technique. RESULTS: Compared with the normal control group, the degree of OPGmRNA expression was remarkably up-regulated on day 7 after model establishment (P < 0.05) and the degree of OPGLmRNA expression was remarkably up-regulated on day 14 after model establishment (P < 0.05) in the model group. Compared with the model group, the degree of OPGmRNA expression was remarkably up-regulated (P < 0.05), and the positive dyeing strength and area of bone tissue were stronger and broader on day 14, 28, and 56 after model establishment in the SJT group. The degree of OPGLmRNA expression was remarkably down-regulated (P < 0.05), and the positive dyeing strength and area of bone tissue were weaker and smaller on day 14 after model establishment in the SJT group. The ratio of OPGmRNA/OPGLmRNA was remarkably up-regulated on day 14, 28, and 56 after model establishment (P < 0.05). CONCLUSION: The effect mechanism of promoting mandibular defect repairing by SJT may be correlated to regulating the expressions of OPGmRNA and OPGLmRNA.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Traumatismos Mandibulares/metabolismo , Osteoprotegerina/metabolismo , Ligante RANK/metabolismo , Cicatrização/efeitos dos fármacos , Animais , Expressão Gênica/efeitos dos fármacos , Ligantes , Masculino , Traumatismos Mandibulares/genética , CoelhosRESUMO
OBJECTIVE: To investigate the effects of mycophenolic acid (MPA) on the proliferation and differentiation of human bone marrow-derived mesenchymal stem cells (MSCs). METHODS: MSCs were treated with MPA at the concentration of 1 µ mol/L, 10 µ mol/L, 50 µ mol/L, and 100 µ mol/L, respectively. Cell proliferation was analyzed using CCK-8 method. Apoptosis was detected by PI/Annexin V assay kit. The mRNA expression of inosine-5'-monophosphate dehydrogenase (IMPDH) in MSCs was analyzed by RT-PCR. Osteogenic differentiation was analyzed by Von Kossa staining, Ca(2+) quantification and real-time PCR. RESULTS: In the range of 1 µ mol/L to 100 µ mol/L, MPA caused a significant subdued proliferation rate of MSCs in a concentration-and time-dependent manner by guanosine depletion, and PI/Annexin staining showed no apoptosis induced by MPA. RT-PCR results showed that MSCs expressed both IMPDH I and IMPDH II. von Kossa staining and Ca(2+) quantification indicated that MPA inhibited osteogenic differentiation of MSCs, and real-time PCR detected a dose-dependent decrease in expression of Osteopontin and BMP-2. Further investigation showed that MPA down-regulated the expression of Runx2 and Osterix. CONCLUSION: MPA can inhibit the proliferation of MSCs by guanosine depletion in a concentration-and time-dependent manner and inhibit the osteogenic differentiation of MSCs by down-regulation of the expression of Runx2 and Osterix.
Assuntos
Células da Medula Óssea/citologia , Células-Tronco Mesenquimais/citologia , Ácido Micofenólico/farmacologia , Apoptose/efeitos dos fármacos , Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Humanos , Células-Tronco Mesenquimais/efeitos dos fármacos , Fator de Transcrição Sp7 , Fatores de Transcrição/metabolismoRESUMO
OBJECTIVE: To investigate the effect of blocking the inhibitory receptors KIR2DL1 and KIR2DL2/2DL3 with monoclonal antibody on cytotoxic activity of human NK cells. METHODS: Human peripheral blood NK cells were isolated by Rosettesep NK sorting kit. The cytotoxic activity of NK cells against human leukemia NB4, K-562, Raji cells and allogeneic mature or dendritic cells (DCs) was detected before or after KIR2DL1 and KIR2DL2/2DL3 were blocked. The effect of NK cells on T lymphocyte proliferation was detected by mixed lymphocyte reaction and TGF-ß1 concentration in culture supernatant was measured. RESULTS: The cytotoxicity of NK cells to NB4 cells was augmented with increasing concentration of the antibody. Combination of both antibodies enhanced killing activity of NK cells. NK cells had strong cytotoxicity to K-562 cells, but were not enhanced by the blockade of inhibitory receptors. The cytotoxicity to Raji cells was not evidently augmented. The cytotoxicity of NK cells to mature DC was enhanced remarkably with the increase of concentration of the antibodies (2.20% ±1.10% compared with 37.59% ±5.06%, P<0.05). In mixed lymphocyte reaction, the blockade of two antibodies enhanced the inhibition effect of NK cells on T cell proliferation (77.85% ± 8.31% compared with 43.05% ± 5.95%, P<0.05) and the content of TGF-ß1 in the supernatant was increased. CONCLUSION: The cytotoxic effects of human NK cells against target cells were significantly enhanced with the blockade of inhibitory KIR receptor; and the cytokine TGF-ß1 secreted by NK cells further inhibits T cells proliferation.
Assuntos
Anticorpos Monoclonais/farmacologia , Células Matadoras Naturais/imunologia , Receptores KIR2DL1/efeitos dos fármacos , Receptores KIR2DL2/efeitos dos fármacos , Receptores KIR2DL3/efeitos dos fármacos , Anticorpos Monoclonais/imunologia , Linhagem Celular , Células Cultivadas , Citotoxicidade Imunológica/efeitos dos fármacos , Citotoxicidade Imunológica/imunologia , Células Dendríticas/imunologia , Humanos , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/metabolismo , Teste de Cultura Mista de Linfócitos , Receptores KIR2DL1/imunologia , Receptores KIR2DL2/imunologia , Receptores KIR2DL3/imunologia , Linfócitos T/imunologia , Fator de Crescimento Transformador beta1/metabolismoRESUMO
OBJECTIVE: To explore the relationship between tumor necrosis factor (TNF) gene polymorphisms in donors and recipients and the incidence and severity of acute graft-versus-host diseases (aGVHD) after unrelated allogeneic hematopoietic stem cell transplantation (allo-HSCT). METHODS: Single nucleotide polymorphisms (SNPs) of TNFalpha-238 (G/A), TNFalpha-857 (C/T), TNFalpha-863 (C/A), TNFalpha-1031 (T/C), TNFbeta + 252(A/G) were analyzed by Multiplex SNaPshot analysis in 76 pairs of donors and recipients. RESULTS: Transplantation involving donors with TNFalpha-857 CC genotype resulted in a higher incidence of grade II-IV aGVHD than donors with CT genotype (91.3% vs 8.7%, P = 0.039). In the 23 patients with grade II-IV aGVHD, no patients had TNFbeta + 252 AA genotype, 19 (82.6%) had GA genotype and 4 (17.4%) had GG genotype. There was a significant difference in the distribution pattern of the TNFbeta + 252 (AA, GA and GG) genotypes in these patients (P = 0.03). There was no significant association of TNFalpha-238 (G/A), TNFalpha-863 (C/A) and TNFalpha-1031(T/C) polymorphisms with the risk of aGVHD. CONCLUSION: These results suggest donor TNFalpha-857 CC genotype is related to a higher incidence of grade II-IV aGVHD, and patients with TNFbeta + 252 AA genotype have protection against the risk of grade II-IV aGVHD.
Assuntos
Doença Enxerto-Hospedeiro/genética , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Linfotoxina-alfa/genética , Polimorfismo de Nucleotídeo Único , Fator de Necrose Tumoral alfa/genética , Adolescente , Adulto , Criança , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Doadores de Tecidos , Transplante Homólogo , Adulto JovemRESUMO
AIM: To examine the ability of rapamycin to suppress growth and regulate telomerase activity in the human T-cell leukemia cell line Jurkat. METHODS: Cell proliferation was assessed after exposure to rapamycin by 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide assay. Cell cycle progression and apoptosis were determined by flow cytometry. The proteins important for cell cycle progression and Akt/mammalian target of rapamycin signaling cascade were assessed by Western blotting. Telomerase activity was quantified by telomeric repeat amplication protocol assay. The human telomerase reverse transcriptase (hTERT) mRNA levels were determined by semi-quantitative RT-PCR. RESULTS: Rapamycin inhibited the proliferation of Jurkat, induced G1 phase arrest, unregulated the protein level of p21 as well as p27, and downregulated cyclinD3, phospho-p70s6k, and phospho-s6, but had no effect on apoptosis. Treatment with rapamycin reduced telomerase activity, and reduced hTERT mRNA and protein expression. CONCLUSION: Rapamycin displayed a potent antileukemic effect in the human Tcell leukemia cell line by inhibition of cell proliferation through G1 cell cycle arrest and also through the suppression of telomerase activity, suggesting that rapamycin may have potential clinical implications in the treatment of some leukemias.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sirolimo/farmacologia , Telomerase/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Fase G1/efeitos dos fármacos , Humanos , Células Jurkat , Leucemia de Células T/tratamento farmacológico , RNA Mensageiro/metabolismoRESUMO
Human bone marrow mesenchymal stem cells (hMSCs) are a promising source for clinical stem cell transplantation. However, telomere regulation mechanisms, as one of the possible major mechanisms by which hMSCs sustain their stem cell characteristics, remain unknown. We isolated hMSCs by plastic adhesion and characterized these cells by morphology, immune phenotype and differentiation capacity. Telomerase was found negative in hMSCs, but slightly up-regulated in hMSC-derived adipocytes by the Telomeric Repeat Amplification Protocol (TRAP) assay. Moreover, hMSCs lack the alternative lengthening of telomeres (ALT) mechanism, because the hallmarks of ALT, such as very long and heterogeneous telomeres, extra-chromosome telomere repeat DNA (ECTR), and ALT-associated promyelocytic leukemia bodies (APBs), were not evident. However, when hMSCs were arrested in S phase with a combination of serum deprivation and aphidicolin, previously undetectable telomerase activity became predominantly positive. Meanwhile, the expression level of hTERT protein and mRNA increased, paralleled with the appearance of a large cohort of synchronized hMSCs at S phase. These findings provide a profile of telomere regulation by cell cycle dependent expression of telomerase in hMSCs and may lead to a better understanding of the stem cell nature of these cells.
Assuntos
Células da Medula Óssea/enzimologia , Ciclo Celular , Células-Tronco Mesenquimais/enzimologia , Telomerase/metabolismo , Telômero/metabolismo , Células da Medula Óssea/citologia , Diferenciação Celular , Células Cultivadas , Humanos , Células-Tronco Mesenquimais/citologia , Fenótipo , Telomerase/genéticaRESUMO
OBJECTIVE: To prepare monoclonal antibodies (McAbs) against human mesenchymal stem cells (hMSCs) and to study their biological characteristics. METHODS: BALB/C mice were immunized with pooled hMSCs. McAbs were prepared by hybridoma technique and their biological characteristics were analyzed by indirect immunofluorescence, immunohistochemistry and flow cytometry. RESULT: Five hybridoma cell lines were successfully established, which secret McAbs specifically against hMSCs. Investigations showed that all these McAb reacted only to hMSCs and had no cross-reaction to other human cells, the relative affinities of 5 McAbs were 1x10(6) (ZUB1), 1x10(5) (ZUB4), 1x10(6) (ZUC3), ZUE12 (1x10(5)) and 1x10(5) (ZUF10), respectively. Isotype analysis showed that ZUB1, ZUE12, ZUF10 against the same isotype, while ZUC3, ZUB4 against other two different isotypes alone. Flow cytometric analysis showed that the positive expression rate of cultured hMSCs was 87.39% (ZUB1), 88.07% (ZUB4), 88.12% (ZUC3), 69.89% (ZUE12) and 83.67% (ZUF10). CONCLUSION: The prepared five McAbs can specifically react against hMSCs, which can be used for selection and study of hMCSs.
Assuntos
Anticorpos Monoclonais/biossíntese , Células da Medula Óssea/imunologia , Imunoglobulina G/imunologia , Células-Tronco Mesenquimais/imunologia , Animais , Especificidade de Anticorpos , Células da Medula Óssea/citologia , Imunofluorescência/métodos , Células HL-60 , Humanos , Hibridomas/metabolismo , Imunoglobulina M/imunologia , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos BALB C , RatosRESUMO
OBJECTIVE: Detecting the expression and mutation of human telomeric repeat binding factor (hTRF1) in 10 malignant hematopoietic cell line cells on the base of determining its genomic structure and its four pseudogenes to clarify if hTRF1 mutation is one of the factors of the activation of telomerase. METHODS: hTRF1cDNA sequences were obtained from GenBank, its genome structure and pseudogenes were forecasted by BLAST and other biology information programs and then testified by sequencing. Real-time RT-PCR was used to detect the expression of hTRF1mRNA in 10 cell line cells, including myelogenous leukemia cell lines K562, HL-60, U-937, NB4, THP-1, HEL and Dami; lymphoblastic leukemia cell lines 6T-CEM, Jurkat and Raji. Telomerase activities of cells were detected by using telomeric repeat amplification (TRAP)-ELISA protocol. PCR and sequencing were used to detect mutation of each exon of hTRF1 in 10 cell line cells. RESULTS: hTRF1 gene, mapped to 8q13, was divided into 10 exons and spans 38.6 kb. Four processed pseudogenes of hTRF1 located on chromosome 13, 18, 21 and X respectively, was named as PsihTRF1-13, PsihTRF1-18, PsihTRF1-21 and PsihTRF1-X respectively. All cell line cells showed positive telomerase activity. The expression of hTRF1 was significantly lower in malignant hematopoietic cell lines cells (0.0338, 0.0108-0.0749) than in normal mononuclear cells (0.0493, 0.0369-0.128) (P=0.004). But no significant mutation was found in all exons of hTRF1 in 10 cell line cells. Four variants were found in part of intron 1, 2 and 8 of hTRF1. Their infection on gene function is unknown and needs further studies. CONCLUSION: hTRF1 mutation is probably not one of the main factors for telomerase activation in malignant hematopoietic disease.
Assuntos
Mapeamento Cromossômico/métodos , Análise Mutacional de DNA/métodos , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/metabolismo , Proteínas de Ligação a Telômeros/genética , Proteínas de Ligação a Telômeros/metabolismo , Sequência de Bases , Linhagem Celular Tumoral/metabolismo , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença/genética , Humanos , Dados de Sequência Molecular , Complexo ShelterinaRESUMO
OBJECTIVE: To study the expression of human telomere repeat binding factor 1 (TRF1) to investigate the correlation of telomerase activity with acute leukemia. METHODS: Leukemic cells were collected from 30 cases of acute leukemia. Realtime quantitative PCR with fluorescence probe hybridization was used to measure expression of TRF1 and hTERT mRNA in leukemic cells. RESULTS: TRF1 mRNA expression was 0.0126 (0.0127-0.0546) in acute non-lymphocytic leukemia (ANLL), which was lower than that in normal mononuclear cells [0.0457 (0.00839-0.262), P<0.001], but its expression in acute lymphoblastic leukemia (ALL) cells [0.0745 (1.92 x 10(-6)-0.193)] had no significant difference compared with that in normal mononuclear cells. TRF1 expression in ANLL cells was significantly lower than that in ALL cells (P=0.001). The expressions of TRF1 mRNA in AL cells and normal mononuclear cells had no significant correlation with expression of hTERT mRNA (r=-0.173, P=0.207). CONCLUSION: The expression of TRF1 is lower in ANLL cells, which indicates TRF1 may have some effect on telomerase activity by regulating telomere length in ANLL cells.
Assuntos
Leucemia Mieloide Aguda/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , Telomerase/metabolismo , Proteína 1 de Ligação a Repetições Teloméricas/biossíntese , Adolescente , Adulto , Idoso , Feminino , Humanos , Leucemia Mieloide Aguda/enzimologia , Masculino , Pessoa de Meia-Idade , Leucemia-Linfoma Linfoblástico de Células Precursoras/enzimologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Proteína 1 de Ligação a Repetições Teloméricas/genéticaRESUMO
OBJECTIVE: To study the expression of peptidyl-prolyl cis/trans isomerase (PPIase or Pin1) in malignant hematopoietic cells and its relation with cell cycle. METHODS: Realtime quantitative PCR with fluorescence probe hybridization was used to measure expression of Pin1 mRNA in malignant hematopoietic cell lines and normal mononuclear cells separated from bone marrow. HeLa cells were blocked with Thymidine and Nocodazole in different cell phases and then the expression of Pin1 mRNA and protein were detected by realtime-PCR and immunoblotting. RESULTS: The expression of Pin1 in malignant hematopoietic cell lines was significantly higher than that in normal controls (0.339 +/-0.093 compared with 0.038 +/-0.005, P<0.01). Its expression in myeloid malignant hematopoietic cell lines was significantly higher than that in normal controls (0.388 +/-0.115 compared with 0.038 +/-0.005, P<0.01) and so was the malignant lymphocytic cell lines (0.226 +/-0.166 compared with 0.038 +/-0.005, P<0.01). The expression of Pin1 was closely correlated with cell cycle. It was the highest in G1 phase and the lowest in S phase (110.762 +/-16.737 compared with 4.080 +/-0.634, P<0.01). CONCLUSION: Pin1 is overexpressed in malignant hematopoietic cell lines and its expression is different during cell cycle that is highest in G1 phase and lowest in S phase.