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Parkin, an E3 ubiquitin ligase, plays a role in maintaining mitochondrial homeostasis through targeting damaged mitochondria for mitophagy. Accumulating evidence suggests that the acetylation modification of the key mitophagy machinery influences mitophagy level, but the underlying mechanism is poorly understood. Here, our study demonstrated that inhibition of histone deacetylase (HDAC) by treatment of HDACis activates mitophagy through mediating Parkin acetylation, leading to inhibition of cervical cancer cell proliferation. Bioinformatics analysis shows that Parkin expression is inversely correlated with HDAC2 expression in human cervical cancer, indicating the low acetylation level of Parkin. Using mass spectrometry, Parkin is identified to interact with two upstream molecules, acetylase acetyl-CoA acetyltransferase 1 (ACAT1) and deacetylase HDAC2. Under treatment of suberoylanilide hydroxamic acid (SAHA), Parkin is acetylated at lysine residues 129, 220 and 349, located in different domains of Parkin protein. In in vitro experiments, combined mutation of Parkin largely attenuate the interaction of Parkin with PTEN induced putative kinase 1 (PINK1) and the function of Parkin in mitophagy induction and tumor suppression. In tumor xenografts, the expression of mutant Parkin impairs the tumor suppressive effect of Parkin and decreases the anticancer activity of SAHA. Our results reveal an acetylation-dependent regulatory mechanism governing Parkin in mitophagy and cervical carcinogenesis, which offers a new mitophagy modulation strategy for cancer therapy.
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Objective To explore the clinical efficacy and safety of panaxadiol saponins for the treatment of non-small cell lung cancer(NSCLC) with chemotherapy-induced leukopenia.Methods 92 NSCLC patients with leukopenia after chemotherapy were selected and divided into the observation group (46 cases) and the control group (46 cases) by random number table method.On the second day after the chemotherapy,the observation group was given panaxadiol saponins capsules,3 tablets/time,2 times/day.The control group was orally given placebo or reserpine,4 weeks for one course of treatment,the two groups were continuously treated for two courses.The clinical efficacy,number of leukocytes,improvement of TCM symptoms and adverse reactions were evaluated.Results After treatment for 4 weeks and 8 weeks,the WBC counts of the observation group were (4.48 ±0.77) × 109/L and (4.92 ± 0.89) × 109/L,respectively,which were significantly higher than those of the control group[(4.02 ± O.93) × 109/L and (4.57 ± 0.86) × 109/L],the differences were statistically significant(t =8.24,8.41,all P ＜ 0.05).After treatment for 4 weeks and 8 weeks,the TCM syndrome scores of the observation group were (24.02 ± 5.91)points and (21.73 ± 4.14) points,respectively,which were lower than those of the control group [(26.33 ± 5.08) points and (23.14 ± 3.90) points],the differences were statistically significant (t =9.68,9.63,all P ＜ 0.05).After treatment for 4 weeks and 8 weeks,the total effective rates of TCM were 76.09％ (35/46) and 82.61％ (38/46),respectively,which were significantly higher than those of the control group [63.04％ (29/46) and 63.04％ (29/46)],the differences were statistically significant(x2 =10.32,8.61,all P ＜ 0.05).The effective rates of leukopenia improvement of the observation group after treatment for 4 weeks and 8 weeks were 69.57％ (32/46) and 78.26％ (36/46),respectively,which were higher than those of the control group [56.52％ (26/46) and 65.22％ (30/46)],the differences were statistically significant(t =9.38,9.51,all P ＜ 0.05).There were no significant differences in adverse reactions between the two groups (P ＞ 0.05).Conclusion Panaxadiol saponins in the treatment of NSCLC chemotherapy-induced leukopenia can significantly improve the number of white blood cells,improve the clinical symptoms,and it has good safety.
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AIM:To investigate the effect of decitabine (Dacogen, DAC) on the proliferation and differentia-tion of K562 cells.METHODS:The K562 cells were treated with different concentrations of DAC .The colony formation ability of the cells was detected by the colony formation assay with semi-solid culture .The cell viability was detected with MTT assay.The morphologic features were observed under inverted microscope with Wright ’s staining.The changes of the cell cycle distribution and the expression of CD 11b and CD42b were analyzed with flow cytometry .The protein expression of CDK2, cyclin E1, P27, GATA-1 and PU.1 in the K562 cells was determined by Western blot .RESULTS:DAC signifi-cantly decreased the colony number of the cells and cell viability in a dose-dependent manner .The morphological changes of the cells displayed partial differentiation .After treated the K562 cells with DAC for 72 h, the cell proportion in S phase was obviously decreased , while the cell proportion in G 2/M phase was obviously increased in a dose-dependent manner . After treated the K562 cells with DAC for 7 d, the percentage of CD11b and CD14 positive cells was further elevated , and the protein expression of P27, GATA-1 and PU.1 was increased.However, the protein expression of CDK2 and cyclin E1 was decreased .CONCLUSION:DAC inhibits the proliferation and induces differentiation of the K 562 cells via regulation of cell cycle .
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Aim To investigate the effect of bufalin on proliferation and expression of WT1 in K562 cells. Methods The colony number of K562 cell was detec-ted with semi-solid culture assay. The cell cycle was measured by flowcytometry, and the expression of WT1 was observed with immunocytochemistry. Subcutaneous tumor models established by K562 cells in BALB/C nu/nu mice were divided into three groups, including model group, bufalin group and positive control group. After 21 days, the subcutaneous tumors were removed for calculating the inhibitory rate of tumor growth. HE staining and immunohistochemistry were used to ob-serve the morphological changes and the expression of WT1 . Results ① Bufalin could significantly decrease the colony number of K562 cell, arrest it at G0/G1 phase and down-regulate its expression of WT1 in a dose-dependent manner. ② Compared with the model group, the tumor inhibitory rate was much higher, while the volume and the weight were obviously lower in the other two groups. ③Bufalin could induce apop-tosis, necrosis, hemorrhage and fibrosis with HE stai-ning, and down-regulate the expression of WT1. Con-clusion Bufalin could inhibit the proliferation, arrest the cell cycle at G0/G1 phase and down-regulate the expression of WT1 in vitro. Bufalin could inhibit the tumor inhibitory rate, the volume and the weight of the subcutaneous tumors, induce apoptosis, necrosis, hemorrhage and fibrosis with HE staining and down-regulate the expression of WT1 .
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Aim To observe the effects of panaxdiol saponins component ( PDS-C) extracted and isolated from Chinese ginseng herb as new Chinese patent med-icine on the promotion of hematopoiesis and the regula-tion of the immune system in treating mice models with aplastic anemia ( AA ) . Methods For preparation of immune mediated AA models, BALB/c mice were ex-posed to sublethal doses of 5. 0 Gy γ radiation, fol-lowed by transplanted lymphocytes from DBA/2 donor mice. The mice models were divided into six groups in-cluding normal control, AA model, PDS-C treated groups with lower, medium and higher dosages, cy-closporine ( CsA) as positive drug control. Both PDS-C and CsA were administered by gastrogavage for 15 days. The peripheral blood cells counts and bone mar-row pathological examination were tested, the percenta-ges of Th1/Th2/Treg cells from spleen were measured, the protein expression levels of T-bet, GATA-3 and FOXP3 transcription factors in spleen cells were detec-ted. Results Curative effect of PDS-C on treating AA mice was satisfactory. The peripheral hemoglobin, white blood cells and platelet counts in PDS-C groups with medium and higher doses were significantly higher than those in model control. Meanwhile, PDS-C ele-vated the percentages of Th2 cells and Treg cells, but decreased the percentage of Th1 cells, as well as up-regulated the GATA-3 , FOXP3 and down-regulated the T-bet protein levels. Conclusion PDS-C possesses the activities of promoting hematopoiesis obviously. It can improve marrow myelosuppression, enhance the re-covery of hematopoiestic function, and elevate the pe-ripheral blood cells counts. PDS-C also pays its immu-noregulatory efficacy though recovering from unbal-anced Th1/Th2/Treg cells in treating immune media-ted AA mice.
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[ ABSTRACT] AIM:To observe the effects of total saponins of Panax ginseng ( TSPG) on the promotion of hema-topoiesis and to explore the underlying mechanism in treating aplastic anemia ( AA) in a mouse model.METHODS:For preparation of AA model, BALB/c mice were exposed to sublethal dose of 5.0 Gyγradiation, followed by transplantation of lymphocytes from DBA/2 donor mice.The experiment was divided into 6 groups, including normal control group, AA model group, TSPG treatment groups with low, medium and high doses, and positive control group with cyclosporine A ( CsA) .Both TSPG and CsA were administered by gastrogavage.After 15 d treatment, the peripheral hemogram was test-ed, the cytokine contents in serum were measured, and bone marrow semisolid culture of colony-forming assay was conduc-ted for determining hematopoietic progenitor cells.The protein levels of extracellular signal-regulated kinase 1/2 ( ERK1/2) and its phosphorylation status in the bone marrow cells were determined.RESULTS:Curative effect of TSPG in trea-ting AA mice was satisfactory.Peripheral counts of white blood cells and platelet, and concentration of hemoglobin in TSPG groups with medium and high doses were significantly higher than those in model control.TSPG increased the colony num-bers of CFU-E, BFU-E, CFU-GM and CFU-MK derived from hematopoietic progenitor cells.Meanwhile, up-regulation of the phosphorylated ERK1/2, decreased contents of Th1 cytokines and increased contents of Th2 cytokines in the serum were observed.CONCLUSION:TSPG possess the dual efficacies on the promotion of hematopoiesis and immunoregulation in AA mice by regulating the expression of Th1/Th2/Th17 cytokines and increasing the phosphorylation of ERK1/2.
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Aim To observe mesenchymal stem cell differentiation into osteoblast induced by TSPG, and explore how TSPG enhances the promotion of hemato-poiesis of osteoblast differentiation from mesenchymal stem cell. Methods MSCs were cultured by TSPG combined with osteoinductive medium. The cellular vi-ability of proliferation was detected with MTT assay. The content of alkaline phosphatase in the cultural su-pernatant was tested with pNPP assay. The ability of MSCs to form calcium nodes was also observed after a-lizarin red stain. The protein expression of RUNX2 was analyzed with Western blot. The content of cytokines associated with hematopoiesis was tested with Elisa as-say. The ability of promoting hematopoiesis was detec-ted with hematopoietic colony forming assay. Results Both MTT and pNPP assay showed that optical den-sity ( OD) values were increased in response to TSPG treatment in a dose-dependent manner. The mineraliza-tion formation ability was enhanced with TSPG-treat-ment. Meanwhile, the expression of RUNX2 protein was up-regulated in TSPG-treated cell. Moreover, the content of cytokines associated with hematopoiesis and the number of hematopoietic progenitor colony were in-creased by TSPG-treatment compared with the control group. Conclusion TSPG could induce MSCs differ-entiation in to osteoblast and enhance the effect of oste-oblast differentiation from MSCs on promoting hemato-poiesis.
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Objective To observe the effects of whole body γ-ray radiation on hematopoiesis and cytokines related to T cell subsets in mouse,to detect the expression of transcription factors of splenic T cell subsets,and to investigate the correlation between hematopoiesis injury and abnormal immune function.Methods Totally 50 BALB/c mice were divided into radiation group and blank control group with the random number table method.The former group were given 5.5 Gy 60Co γ-ray radiation on whole body and another received sham radiation.The numbers of white blood cells and platelets of radiation group were counted at 4,8,12 and 20 d after radiation,and these numbers of blank control mice were counted only at 20 d.Hematopoietic tissue proliferation was evaluated by biopsy sections of mice femur.The contents of Th1,Th2,and Th17 in peripheral blood were detected with cytometric bead array (CBA).The expressions of T-bet/GATA-3 and RORγt/Foxp3 proteins related to the differentiation of T cell subsets in spleen tissue were measured by Western blot.Results The numbers of white blood cells and platelets of radiation group mice were reduced obviously (t WBC =18.48,15.72,9.79,3.30; t PLT =22.52,19.74,11.78,4.70,P ＜ 0.05) compared with blank control group.Biopsy sections showed that bone marrow hematopoietic cells of the radiated mice were less than those of blank group,and adipocytes became more.At 8 d,the marrow suppressions were more obvious than those at 20 d.Serum contents of Th1 cytokines IFN-γ,TNF-α and Th2 cytokines IL-4,IL-6 in the radiation group were higher than those in the blank control group at 8 and 12 d(t IFN-γ =2.93,3.36,t TNF-α =6.09,8.11,6.43,4.49,tIL-4 =4.49,3.18,t IL-6=5.11,8.67,6.67,8.55,P＜0.05).IL-17A secreted mainly by Th17 cells was also higher than the blank (t =3.68,6.24,5.32,4.06,P ＜ 0.05).Compared with the blank control group,the expression of T-bet protein increased significantly (t =5.64,2.75,3.56,4.65,P ＜ 0.05),and the expressions of GATA-3,RORγt,and Foxp3 proteins decreased at 4,8 and 12 d except the RORγt at 20 d (tRORγt =6.79,4.31,4.47,tGATA-3 =3.88,8.06,2.84,3.23,tFoxp3 =10.00,8.06,2.89,5.93,P＜ 0.05).Conclusions 5.5 Gy whole body γ-ray radiation inhibits bone marrow hematopoiesis of BALB/c mice and makes the differentiation and function of T cells to be abnormal,which may be associated with bone marrow hematopoiesis obstacle.
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Aim To investigate the effect of resveratrol on proliferation and differentiation in K562 cells. Methods K562 cells were treated with different con-centrations of resveratrol for 6d. The colony number of K562 cells was detected with semi-solid culture assay. Expression of GATA-1 and PU. 1 in K562 cells was re-spectively measured with immunocytochemistry and Western blot. Expression of differentiation related anti-gen, CD11b, CD14 and CD42b, was measured with flowcytometry on K562 cells. Results Resveratrol could significantly decrease the colony number of K562 cells in a dose-dependent manner, and enhance the ex-pression of GATA-1,PU. 1,CD11b, CD14 and CD42b in K562 cells. Conclusion Resveratrol could inhibit the proliferation and induce differentiation of K562 cells via up-regulating the expression of GATA-1 and PU. 1 protein.
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The binding complex of chemokine receptor CXCR4 and its specific ligand CXCL12 triggers the downstream signaling pathway to form biological axis of CXCR4/CXCL12. The axis plays an important role in homing, repopulation of hematopoietic stem cells, as well as maintenance of normal hematopoiesis and homeostasis of hematopoietic microenvironment. Meanwhile, CXCR4/CXCL12 axis is also closely related with relapsed leukemia, since it may promote survival, proliferation, metastasis and drug resistance in leukemic cells.The expression level of CXCR4 could be treated as an important marker for predicting prognosis in patients wizh leukemia, therefore, the combination of CXCR4 inhibitors with routine chemotherapy may represent a powerful approach for the treatment of leukemia.
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[Objective]To explore the potential alteration of their osteogenesis and adipocyte differentiation in bone marrow mesenchymal stems cells from mice with immune-mediated aplastic anemia.[Methods]Balb/c mice model of immune-mediated aplastic anemia was established by radiation with sublethal dose of 60Co following the intravenously infusion lymphocytes of DAB/2 mice.The culture of the MSC and CFU-F and pathological examination of bone marrow were carried out 15 days later.The amount of calcium node and the frequency of adipocyte differentiation were evaluated respectively by alizarin red and oil red O.[Result]The number of CFU-F and the number of calcium node in model mice decreased more greatly than normal mice,but the ferenqucy of adipocyte differentiation was more increased greatly in model mice than normal mice;the pathological examination showed in the model mice,the hematopoietic structure was destroyed and filled with abundant adipocyte.[Conclusion]The potention of osteogenesis and adipocyte differentiation was altered in the mice with immune-mediated aplastic anemia.
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<p><b>OBJECTIVE</b>To observe whether the Ginsenosides (GS) could induce HL-60 cell line apoptosis from promyelocytic leukemia.</p><p><b>METHODS</b>HL-60 cells were treated with GS of various concentration to observe the effect of GS on cell morphological change, the DNA content change by flow cytometry, DNA ladder by electrophoresis, and apoptosis rate by Annexin V-FITC test of the cells.</p><p><b>RESULTS</b>GS could inhibit the growth of HL-60 cells and induce cell apoptosis in a certain scope of dose and reacting time.</p><p><b>CONCLUSION</b>GS could specifically induce apoptosis in HL-60 cells, which provides an experimental basis for treatment of leukemia with GS as an supplemntary agent of chemotherapy.</p>