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Objective: To investigate the characteristic and prognostic significance of leukemia stem cells associated antigens expressions including CD34, CD38, CD123, CD96 and TIM-3 in t (8;21) AML. Methods: Bone marrow samples of 47 t (8;21) AML patients were collected at diagnosis from October 2015 to April 2018 in Peking University Peoples' Hospital, then flow cytometry method was performed to detect the expression frequencies of CD34, CD38, CD123, CD96 and TIM-3 to analyze the relationship between leukemia stem cells associated antigens expressions and relapse. Results: Of 47 t (8;21) AML patients tested, the median percentages of CD34(+)CD38(-), CD34(+) CD38(-)CD123(+), CD34(+)CD38(-) CD96(+) and CD34(+) CD38(-) TIM-3(+) cells among nucleated cells were 2.37%, 0.24%, 0.27% and 0.06%, respectively. All the frequencies of CD34(+)CD38(-), CD34(+)CD38(-)CD123(+), CD34(+)CD38(-)CD96(+) and CD34(+) CD38(-)TIM-3(+) cells had no impact on the achievement of CR after the first course of induction. All higher frequencies of CD34(+)CD38(-), CD34(+)CD38(-)CD123(+), CD34(+)CD38(-)CD96(+) cells were related to higher 2-year CIR rate. Whereas, the frequency of CD34(+) CD38(-) TIM-3(+) cells had no impact on CIR rate. Both high frequency of CD34(+) CD38(-) cells and the high level of minimal residual diseases (patients with <3-log reduction in the RUNX1-RUNX1T1 transcript level after the second consolidation therapy) were independent poor prognostic factors of CIR[P=0.025, HR=6.9 (95%CI 1.3-37.4) ; P=0.031, HR=11.1 (95%CI 1.2-99.2) ]. Conclusion: Different leukemia stem cells associated antigens had distinct prognostic significance in t (8;21) AML. High frequencies of CD34(+) CD38(-), CD34(+) CD38(-) CD123(+) and CD34(+)CD38(-)CD96(+) cells at diagnosis predicted relapse in patients with t (8;21) AML.
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Humanos , ADP-Ribosil Ciclasa 1 , Antígenos CD , Citometría de Flujo , Subunidad alfa del Receptor de Interleucina-3 , Leucemia Mieloide Aguda , Células Madre Neoplásicas , Pronóstico , Células MadreRESUMEN
Objective To study the mechanism of the Hedgehog signaling transduction intervened by polypeptide extract from scorpion venom (PESV) on K562/BALB/c-nu leukemia mice. Trying to analyze the molecular mechanisms and targets of the inhibited effect of PESV on chronic myeloid leukemia (CML) in vivo. Methods After establishing the K562/BALB/c-nu leukemia mice successfully, the model mice were divided into six groups which were the blank group, the PESV high, medium, and low doses (0.3, 0.6, 1.2 mg/kg) group, the Imatinib (50 mg/kg) group, and the model group. After 14 d drug intervention, the levels of gene and protein expression of Hedgehog signaling pathway upstream factors Shh, Ptch, and Smo were detected by qRT-PCR and Western blotting, and the protein expression of downstream factor Gli1 was determined by ELISA test. Results Compared to the model group, the genetic and protein expression of Shh which was an upstream factor were increased in the PESV groups. The mRNA and protein expression of Ptch and Smo in PESV low-dose and medium-dose groups were decreased. There were no significant differences of upstream factors between Imatinib group and model group. The concentration of downstream Gli1 protein significantly decreased within low-dose and medium-dose PESV groups, while there was no significant difference between high-dose PESV group and Imatinib group. Conclusion PESV can inhibit the expression of Hedgehog signaling pathway upstream factor Ptch, Smo and downstream factor Gli1 on the mRNA and protein level, while Imatinib has no obvious inhibitory effect on the Hedgehog signaling pathway.
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Objective@#To investigate the characteristic and prognostic significance of leukemia stem cells associated antigens expressions including CD34, CD38, CD123, CD96 and TIM-3 in t (8;21) AML.@*Methods@#Bone marrow samples of 47 t (8;21) AML patients were collected at diagnosis from October 2015 to April 2018 in Peking University Peoples’ Hospital, then flow cytometry method was performed to detect the expression frequencies of CD34, CD38, CD123, CD96 and TIM-3 to analyze the relationship between leukemia stem cells associated antigens expressions and relapse.@*Results@#Of 47 t (8;21) AML patients tested, the median percentages of CD34+CD38-, CD34+ CD38-CD123+, CD34+CD38- CD96+ and CD34+ CD38- TIM-3+ cells among nucleated cells were 2.37%, 0.24%, 0.27% and 0.06%, respectively. All the frequencies of CD34+CD38-, CD34+CD38-CD123+, CD34+CD38-CD96+ and CD34+ CD38-TIM-3+ cells had no impact on the achievement of CR after the first course of induction. All higher frequencies of CD34+CD38-, CD34+CD38-CD123+, CD34+CD38-CD96+ cells were related to higher 2-year CIR rate. Whereas, the frequency of CD34+ CD38- TIM-3+ cells had no impact on CIR rate. Both high frequency of CD34+ CD38- cells and the high level of minimal residual diseases (patients with <3-log reduction in the RUNX1-RUNX1T1 transcript level after the second consolidation therapy) were independent poor prognostic factors of CIR[P=0.025, HR=6.9 (95%CI 1.3-37.4) ; P=0.031, HR=11.1 (95%CI 1.2-99.2) ].@*Conclusion@#Different leukemia stem cells associated antigens had distinct prognostic significance in t (8;21) AML. High frequencies of CD34+ CD38-, CD34+ CD38- CD123+ and CD34+CD38-CD96+ cells at diagnosis predicted relapse in patients with t (8;21) AML.
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Objective:To observe the influence of the CD96 and CD71 expressions in the surface of leukemia stem cells (LSC) in the therapeutic effects and prognosis of the children with acute leukemia,and to clarify the relationships between the molecular biological characteristics of LSC in the children with leukemia and their therapeutic effects and prognosis.Methods:Eighty children with acute leukemia were selected as the subjects.Among them, 39 cases were acute lymphoblastic leukemia (ALL) and 41 cases were acute myeloid leukemia (AML).The CD96 and CD71 expressions on the surface of LSC were detected with flow cytometry;the therapeutic effects of the first cycle chemotherapy, the survival rate of 5-year, the incidence of infection after chemotherapy, the recurrence rate after chemotherapy, and the incidence of extramedullary infiltration of the children were observed and compared.Results: The CD96 expression on the surface of LSC was positive in 38 cases (47.5%) and the CD71 expression on the surface of LSC was positive in 45 cases (56.3%);the difference of positive expression rates of CD96 and CD71 was not significant (χ2=1.227, P=0.268).The positive expression rates of CD96 and CD71 on the surface of LSC of the children with AML were significantly higher than those in the children with ALL, and the differences were statistically significant (χ2=22.225, χ2=34.028, P<0.01).The distribution of therapeutic effects and the clinical efficiency of the first cycle chemotherap, in the children with negative CD96 expression on the surface of LSC were superior to those with positive CD96 expression on the surface of LSC;the distribution of therapeutic effects and the clinical efficiency of the first cycle chemotherapy in the children with negative CD71 expression on the surface of LSC were superior than those with positive CD71 expression on the surface of LSC;the differences between two groups were statistically significant (χ2=11.323, χ2=16.589, P<0.05;U=2.939, U=2.291, P<0.05).The survival rate of 5-year in the children with negative CD96 expression on the surface of LSC was higher than those with positive CD96 expression;the incidence of infection after chemotherapy,the recurrence rate after chemotherapy and the incidence of extramedullary infiltration were lower than those with positive CD96 expression.The incidence of infection after chemotherapy and the recurrence rate after chemotherapy in the children with negative CD71 expression on the surface of LSC were lower than those with positive CD71 expression,and the differences between two groups were statistically significant (χ2=5.051,χ2=13.288, P<0.05).Conclusion: The expressions of CD96 and CD71 on the surface of LSC in the children with acute leukemia has relationship with the subtypes of the disease and the therapeutic effects of the first cycle chemotherapy, which can be used as markers for the diagnosis and evaluation of therapeutic effects.The expression level of CD96 is related to the prognosis of the patients, which can be used as an indicator for predicting the prognosis of patients.
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Leukemia stem cells (LSC) that were found in chronic myeloid leukemia (CML) responsible for the abnormal proliferation with the potential of self-renewal and multi-directional differentiation are involved in the pathophysiological process for drug resistance and relapse of CML. Autophagy, a conservative lysosomal degradation process that mediates cell degradation and recycling process, plays crucial roles in maintaining the homeostasis and function of intracellular environment. Recent studies suggested that autophagy is involved in the regulation of LSC differentiation and also closely related to the chemo-sensitivity of CML. In this review, we focused on the role of autophagy on chemotherapy sensitivity of CML as well as the leukemia stem cell function for the development of new anti-leukemia drugs.
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[ ABSTRACT] Tyrosine kinase inhibitors ( TKIs) are now advocated as the first-line treatment for chronic myeloid leukemia ( CML) , but facing resistance and relapse .Leukemia stem cells ( LSCs ) are leukemia-initiating cells as the source of resistance and relapse .It is therefore important to discover the molecular biomarker of LSCs for developing anti -LSC strategies in leukemic therapy .15-Lipoxygenase (15-LO) is a key enzyme in the pathway of arachidonic acid and plays an important role in the occurrence and development of CML , which is specifically required for chronic myeloid LSCs . This review summarizes the influence of 15-LO on the chronic myeloid LSC characteristics of marked survival , self-renewal, proliferation , differentiation and apoptosis .
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Objective To investigate the effects of curcumin on proliferation and apoptosis of CD34+CD38-KG1a cells and its synergetic effect with busulfan on CD34+CD38-KG1a cells.Methods The expressions of CD34 and CD38 on the surface of KG1a cells and the effect of curcumin on the cell cycle and apoptosis in CD34+CD38-KG1a cells were detected by flow cytometry.MTT assay was used to analyze curcumin’s inhibitory effects on proliferation and synergistic effect with busulfan on CD34+CD38-KG1a.Clone formation rate was measured by methylcellulose colony-formation assay.Morphological changes of apoptotic cells were observed with the inverted optical microscope.The expression of Bcl-2 at the protein level was detected by Western blot.Results The percentage of CD34+CD38-KG1a was (98.2±3.2)% in KG1a cells.Curcumin could inhibit the proliferation in time-and dose-dependent manners and reduce the colony-formation ability of CD34+CD38-KG1a.The coefficient of drug interaction between curcumin and busulfan was less than 1.CD34+CD38-KG1a cells were arrested in the G0/G1 phase by decreasing S phase cells.Meanwhile,curcumin induced the apoptosis of CD34+CD38-KG1a cells. Apoptotic cells became bigger than normal ones,with unclear cell structure and rough edge of cell membrane.The expression of Bcl-2 at the protein level was down-regulated by curcumin.Conclusion Curcumin inhibited the proliferation of CD34+CD38-KG1a cells by reducing colony-formation ability,arresting cells in the G0/G1 phase and inducing apoptosis.Besides,there was a synergistic effect between curcumin and busulfan in CD34+CD38-KG1a cells.The down-regulated expression of Bcl-2 at the protein level may be associated with curcumin-induced apoptosis of CD34+CD38-KG1a cells.
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Objective: To investigate the mechanisms on the regulation of telomere and telomerase in the process of senescence induction of human-derived CD34+CD38- leukemia stem cells (LSC) subpopulation by Angelica sinensis polysaccharide (ASP). Methods: The human-derived CD34+CD38- LSC subpopulation was isolated from acute myelogenous leukemia bone marrow mononuclear cells by magnetic activated cell sorting. The inhibition of ASP on CD34+CD38- LSC subpopulation proliferation was detected by CCK-8 assay. The percentage of senescent cells was detected by SA-β-Gal staining. The colony-formed ability was detected by Colony-forming Assay. The levels of telomerase activities and telomerase reverse transcriptase (TERT) gene expression were performed by TRAP-PCR and quantitative RT-PCR, respectively. The changes of telomere length were tested by Southern blotting assay. Results: The CD34+CD38- LSC subpopulation could be effectively isolated by MACS. The purity of CD34+CD38- LSC population is up to (91.15 ± 2.41)%; The cells showed the features of well-stacked morphology, high transparency, well refraction under inverted phase contrast microscope. ASP had a remarkable dose-dependent inhibition on CD34+CD38- LSC proliferation in vitro culture (P < 0.05). The number of SA-β-Gal staining positive cells had been increased compared to the cells in control group (P < 0.01), a decrease in colony-forming abilities (P < 0.01), a decrease level on TERT gene and telomerase activities (P < 0.05), and a shorter length on telomere of CD34+CD38- LSC after 40 μg/mL ASP co-culture for 48 h (P < 0.05). Conclusion: ASP could induce the senescence of human derived leukemia bone marrow CD34+CD38- LSC via regulating the cell telomere system in vitro co-culture.
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Objective To assay and determine whether the human acute monocytic leukemia cell line THP-1 contains side populations (SP) cells,and to increase the proportion of SP cells using cytarabine (Ara-C).Methods Fluorescent microscope and flow cytometry (FCM) were employed for detecting the percentage of SP cells in THP-1 cells.Then,SP and non-SP (NSP) subpopulations were collected and identified.Finally,THP-1 cells were incubated with different concentrations of Ara-C for 24 hours and detected the proportion of SP cells,respectively.Results The results demonstrated that the percentage of SP cells was (1.81 ± 0.99) ％ in THP-1 cells.A majority of the SP cells remained in the G0/G1 phase,and the expressions of CD34 + and CD34 + CD38-and the proliferative ability of the SP cells were higher than those of NSP cells (P ＜ 0.05).The mRNA expression of multidrug resistance genes (ABCG2,ABCB1),apoptosis regulation genes (Bcl-2) and the Bcl-2/Bax ratio of SP cells were higher than those of NSP cells.SP cells have been shown to be more tumorigenic than NSP cells.After co-culture with Ara-C,the proportion of SP cells increased significantly and presented in a concentration-dependent manor.Conclusions All of these findings suggest that the THP-1 cell line contains SP cells and the SP cells possess some intrinsic stem cell properties.The proportion of SP cells can be increased when co-cultured with Ara C,and this technique is a useful and important application for the study of LSCs.