Fludarabine, busulfan, and melphalan conditioning regimen in allogeneic hematopoietic stem cell transplantation for adult patients with myeloid malignancies: A multicenter retrospective study.

Relapse remains the main cause of treatment failure in patients with myeloid malignancies even after allogeneic hematopoietic stem cell transplantation (allo-HSCT). We observed a particularly low incidence of relapse in patients prepared with fludarabine, busulfan and melphalan in our previous study and this multicenter retrospective analysis aimed to confirm the feasibility of the regimen and to identify the potential prognostic factors. This study was performed using registry data from adults patients with myeloid malignancies who underwent their first allo-HSCT following fludarabine(≥100 mg/m2), busulfan (≥3.2 mg/kg) and melphalan (≥100 mg/m2) based conditioning at nine transplantation centers in China between Jan. 2020 and Mar. 2022. A total of 221 consecutive patients (AML n = 171, MDS-IB-1 or 2 n = 44, CMML n = 6) with median age of 46 were enrolled in this study. The median follow-up was 507 days for survivors. The 2-year NRM, CIR, OS and DFS were 10.6% ± 2.2%, 14.8% ± 3.3%, 79.4% ± 3.7% and 74.6% ± 3.7%, respectively. In multivariate analyses, high HCT-CI (≥3) was the only independent factor for higher NRM [hazard ratio (HR), 2.96; 95% confidence interval (CI), 1.11 to 7.90; p = 0.030] and ECOG score ≥2 was the only independent factor for inferior OS (HR, 2.43; 95%CI, 1.15 to 5.16; p = 0.020) and DFS (HR, 2.12; 95%CI, 1.13 to 4.02; p = 0.020). AML diagnosis and positive measurable residual disease (MRD) at transplantation were predictors for higher CIR (HR = 7.92, 95%CI 1.05-60.03, p = 0.045; HR = 3.64, 95%CI 1.40-9.44, p = 0.008; respectively), while post-transplantation cyclophosphamide based graft-versus-host disease prophylaxis was associated with lower CIR (HR = 0.24 95%CI 0.11-0.54, p = 0.001). The intensity of conditioning regimen did not impact CIR, NRM, DFS and OS. These results supported that double alkylating agents of busulfan and melphalan based conditioning regimens were associated with low relapse rate and acceptable NRM in adult patients with myeloid malignancies. The optimal dose remained to be confirmed by further prospective studies.

[Chidamide Promotes Osteogenic Differentiation of Bone Marrow Mesenchymal Stromal Cells from Patients with Myelodysplastic Syndromes].

OBJECTIVE: To explore the effects and mechanisms of chidamide on the osteogenic differentiation of bone marrow mesenchymal stromal cells (MSC) from myelodysplastic syndromes (MDS). METHODS: MSC were isolated and cultured from bone marrow of MDS patients and healthy donors. CCK-8 assay was used to detect the effects of chidamide on the proliferation of MSC. The effects of chidamide on the activity of histone deacetylase (HDAC) in MSC was measured by a fluorescence assay kit and Western blot. Alkaline phosphatase (ALP) activity was detected on day 3 and calcium nodule formation was observed by Alizarin Red staining on day 21 after osteogenic differentiation. The expression of early and late osteogenic genes was detected on day 7 and day 21, respectively. RT-PCR and Western blot were used to detect the effects of chidamide on mRNA and protein expression of RUNX2 which is the key transcription factor during osteogenesis. RESULTS: As the concentration of chidamide increased, the proliferation of MSC was inhibited. However, at a low concentration (1 µmol/L), chidamide had no significant inhibitory effect on MSC proliferation but significantly inhibited HDAC activity. In MSC from both MDS patients and healthy donors, chidamide (1 µmol/L) significantly increased ALP activity, calcium nodule formation, thereby mRNA expression of osteogenic genes, and restored the reduced osteogenic differentiation ability of MDS-MSC compared to normal MSC. Mechanistic studies showed that the osteogenic-promoting effect of chidamide may be related to the upregulation of RUNX2 . CONCLUSION: Chidamide can inhibit HDAC activity in MSC, upregulate the expression of the osteogenic transcription factor RUNX2, and promote the osteogenic differentiation of MDS-MSC.

Incorporating mutations and bone marrow fibrosis into the revised international prognostic scoring system in myelodysplastic syndromes.

The independent prognostic significance of bone marrow fibrosis (BMF) in myelodysplastic syndromes (MDS) is challenged under currently molecular prognostic models. In this study, the clinical and genetic data from 438 MDS patients were analyzed retrospectively. The patients were randomly divided into training (n = 306) and validation (n = 132) cohorts. The independent significant prognostic factors included age, IPSS-R, BMF, TP53 and U2AF1. Using their weighted coefficients, we developed a simplified prognostic system. Four risk groups were produced: low, intermediate, high and very high. The new model yielded more clearly separated survival curves than the IPSS-R. In addition, our model achieved higher C-indexes (0.61 in the training cohort and 0.63 in the validation cohort) than the IPSS-RM model (0.59 and 0.58) and IPSS-R (0.57 and 0.56). In conclusion, BMF was an independent significant prognostic factor for MDS, and adding BMF into the IPSS-R improved its predictive capability.

S100a9 deficiency accelerates MDS-associated tumor escape via PD-1/PD-L1 overexpression.

In recent studies, the tolerable safety profile and positive bone marrow (BM) response suggest a beneficial use of anti-PD-1 agents in the treatment of Myelodysplastic Syndromes (MDS), but the underlying mechanism is still unknown. MDS is mainly characterized by ineffective hematopoiesis, which may contribute to inflammatory signaling or immune dysfunction. Our previous studies focused on inflammatory signaling, and the results showed that S100a9 expression was higher in low-risk MDS and lower in high-risk MDS. In this study, we combine the inflammatory signaling and immune dysfunction. SKM-1 cells and K562 cells co-cultured with S100a9 acquire apoptotic features. Moreover, we confirm the inhibitory effect of S100a9 on PD-1/PD-L1. Importantly, PD-1/PD-L1 blockade and S100a9 can both activate the PI3K/AKT/mTOR signaling pathway. The cytotoxicity is higher in lower-risk MDS-lymphocytes than in high-risk MDS-lymphocytes, and S100a9 partially rescues the exhausted cytotoxicity in lymphocytes. Our study demonstrates that S100a9 may inhibit MDS-associated tumor escape via PD-1/PD-L1 blockade through PI3K/AKT/mTOR signaling pathway activation. Our findings indicate the possible mechanisms by which anti-PD-1 agents may contribute to the treatment of MDS. These insights may provide mutation-specific treatment as a supplementary therapy for MDS patients with high-risk mutations, such as TP53, N-RAS or other complex mutations.

Decreased FOXO1 Expression Is Correlated with Poor Prognosis in Myelodysplastic Syndromes.

Myelodysplastic syndrome is one of the main hematological malignancies that threaten the health of the elderly. However, biomarkers which predict the progression and prognosis of MDS are still controversial and puzzling. FOXO1 gene plays an important role in a variety of intracellular functions, including tumor suppression and cellular immune regulation. However, there is no research report on the correlation between FOXO1 and the clinical features of MDS including immune environment. In this study, we observed that FOXO1 expression is associated with neutrophil count, blasts, chromosome and different MDS scoring systems. FOXO1 expression is closely related to MDS cell immune polarization, and the increase expression of FOXO1 is significantly related to the amplification of immune cell polarization ratio. In addition, FOXO1 expression is associated with progression-free survival and overall survival in MDS patients. Moreover, in a multivariate model FOXO1 low-expression was an independent predictor of poor survival in MDS. In summary, FOXO1 may play a candidate tumor suppressor in MDS, and FOXO1 is a useful independent prognostic predictor in MDS, and it may provide a candidate target therapy in future.

Bone Marrow Fibrosis at Diagnosis and during the Course of Disease Is Associated with TP53 Mutations and Adverse Prognosis in Primary Myelodysplastic Syndrome.

The prognostic significance of bone marrow fibrosis (MF) grade in patients with myelodysplastic syndrome (MDS) is still debated and the molecular changes remain unclear. In our large cohort, a normal reticulum was found in 211 (25.9%) patients, whereas MF1, MF2 and MF3 were detected in 478 (58.7%), 90 (11.1%) and 35 (4.3%) patients at initial diagnosis, respectively. Patients with MF often correlated with some poor prognostic characteristics, including older age, anemia, unfavorable karyotype, higher BM blast and a higher IPSS-R category. For the entire cohort, the median OS was not reached, 30, 16 and 15 months for patients with MF 0, 1, 2 and 3, respectively. After adjusting for IPSS-R, the hazard ratio for mortality was 1.56 (95% CI, 1.18-2.06) for patients with MF1, 2.29 (95% CI, 1.61-3.27) for patients with MF2 and 2.75 (95% CI, 1.69-4.49) for patients with MF3 compared with those with MF0. The mutational landscape of 370 patients showed that TP53, U2AF1 and KMT2D mutations were more frequent in patients with MF2-3. In addition, of the 408 patients with MF0-1, 62 patients (15.1%) progressed to MF2-3 during the follow-up interval. The clinical features, mutation landscape and prognosis of patients with progressed fibrosis were similar to those of patients with MF2-3 at diagnosis. We concluded that BM fibrosis (MF1, 2 and 3) was an adverse prognosis feature in primary MDS and correlated with TP53 mutations both at the time of initial diagnosis and during the course of the disease. Therefore, BM fibrosis should be included in the revised prognostic scoring system and carefully considered in treatment selection.

Aryl hydrocarbon receptor signaling pathway plays important roles in the proliferative and metabolic properties of bone marrow mesenchymal stromal cells.

Bone marrow mesenchymal stromal cells (BMMSCs) are widely sourced and easily amplified in vitro; thus, they have a great potential in the treatment of hemopathies. Recent findings suggested that BMMSCs express the aryl hydrocarbon receptor (AHR). However, few studies have reported on the regulation of proliferative behaviors and metabolism by AHR in BMMSCs. In the present study, we found that activating AHR reduced the proliferation of BMMSCs and enhanced their mitochondrial function, whereas inhibiting AHR exerted the opposite effects. This study may provide the basis for further unveiling the molecular mechanisms and therapeutic potential of AHR in BMMSCs.

Withdrawal Notice: Roles of Aryl-hydrocarbon Receptor in Metabolism and Proliferative Behaviors Of Bone Marrow Mscs: Implications For Targeted Therapy

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Comparative Study on Iron Content Detection by Energy Spectral CT and MRI in MDS Patients.

Objective: The purpose of this study was to identify the difference between dual energy spectral computed tomography (DECT) and magnetic resonance imaging (MRI) used to detect liver/cardiac iron content in Myelodysplastic syndrome (MDS) patients with differently adjusted serum ferritin (ASF) levels. Method: Liver and cardiac iron content were detected by DECT and MRI. Patients were divided into different subgroups according to the level of ASF. The receiver operating characteristic curve (ROC) analysis was applied in each subgroup. The correlation between iron content detected by DECT/MRI and ASF was analyzed in each subgroup. Result: ROC curves showed that liver virtual iron content (LVIC) Az was significantly less than liver iron concentration (LIC) Az in the subgroup with ASF < 1,000 ng/ml. There was no significant difference between LVIC Az and LIC Az in the subgroup with 1,000 ≤ ASF < 2,500 ng/ml and 2,500 ≤ ASF < 5,000 ng/ml. LVIC Az was significantly higher than LIC Az in the subgroup with ASF <5,000 and 5,000 ≤ ASF ng/ml. In patients undergoing DECT and MRI examination on the same day, ASF was significantly correlated with LVIC, whereas no significant correlation was observed between ASF and LIC. After removing the data of ASF > 5,000 mg/L in LIC, LIC became correlated with ASF. There was no significant difference between the subgroup with 2,500 ≤ ASF < 5,000 ng/ml and 5,000 ng/ml ≤ ASF in LIC expression. Furthermore, both LIC and liver VIC had significant correlations with ASF in patients with ASF < 2,500 ng/ml, while LVIC was still correlated with ASF, LIC was not correlated with ASF in patients with 2,500 ng/ml ≤ ASF. Moreover, neither cardiac VIC nor myocardial iron content (MIC) were correlated with ASF in these subgroups. Conclusion: MRI and DECT were complementary to each other in liver iron detection. In MDS patients with high iron content, such as ASF ≥ 5,000 ng/ml, DECT was more reliable than the MRI in the assessment of iron content. But in patients with low iron content, such as ASF < 1,000 ng/ml, MRI is more reliable than DECT. Therefore, for the sake of more accurately evaluating the iron content, the appropriate detection method can be selected according to ASF.

AHR signaling pathway reshapes the metabolism of AML/MDS cells and potentially leads to cytarabine resistance.

Emerging evidence suggests that aryl hydrocarbon receptor (AHR) promotes the initiation, invasion, progression, and metastasis of cancer cells. However, its effects in patients with myelodysplastic syndrome/acute myeloid leukemia (MDS/AML) remain undefined. In this study, we aimed to investigate the effects of AHR activation on malignant cells in patients with MDS/AML. We found that AHR was expressed aberrantly in patients with MDS/AML. Further studies demonstrated that inhibiting AHR decreased the mitochondrial dehydrogenase content and the mitochondrial membrane potential (MMP) in MDS/AML cells. Activating AHR with L-kynurenine (Kyn) increased AHR expression, which was accompanied by an increase in mitochondrial dehydrogenase content and MMP in MDS/AML cells. Moreover, the expression level of mitochondria-associated mitochondrial transcription factor A was increased after activating AHR with L-Kyn when compared with that in the control group but decreased after inhibiting the AHR signal. Activating AHR in MDS/AML cells enhanced the resistance to cytarabine. These findings indicated that activating the AHR signaling pathway reshaped the metabolism in MDS/AML cells, thus contributing to the resistance to cytarabine.

Analysis of the influencing factors related to liver and cardiac iron overload in MDS patients detected by MRI in the real world.

OBJECTIVES: We aim to explore and analyze the related influencing factors of liver and cardiac iron overload in MDS patients detected by magnetic resonance imaging (MRI). METHODS: We have detected cardiac T2* and liver T2* by MRI in 105 MDS patients. Among them, 20 patients accepted MRI examination before and after iron chelation therapy (ICT). Results: We found that adjusted ferritin (ASF) was significantly correlated with liver T2* and cardiac T2*. RBC transfusion volume, brain natriuretic peptide (BNP) and age were the related factors of cardiac T2*, while RBC transfusion volume and erythropoietin (EPO) were related factors of liver T2*. After ICT, the changes of ASF and liver T2* were earlier than cardiac T2*. Chronic hepatitis but virus copy normal's has no significant effect on liver iron deposition. CONCLUSION: These results showed special attention should be paid to these related influencing factors of liver and cardiac T2* expression when we evaluated iron overload and detected the efficacy of ICT in MDS patients.

A genetic development route analysis on MDS subset carrying initial epigenetic gene mutations.

MDS development is a dynamic process during which the accumulation of somatic mutations leads to specific malignant evolution. To elucidate the differential roles of gene mutations in typical MDS, we used targeted sequencing to investigate clonal patterns from 563 patients and focused on cases (199/563 cases) with initial mutations (ASXL1, DNMT3A and TET2) at MDS diagnosis. The consistency of frequency and distribution in patients with or without aberrant chromosomes suggested early events of these initial mutations. Some additional driver mutations (SF3B1, U2AF1 or RUNX1) played roles to keep the basic disease features, or give rise to different phenotypes (BCOR, EZH2 or TP53) in individual patients. Notably, analysis in paired samples before and after MDS progression showed that the mutations identified as last events (involving active signaling, myeloid transcription or tumor suppressor) seemed necessary for MDS development to be AML. Last mutations can exist at MDS diagnosis, or emerge at AML transformation, and involve a small group of genes. Single-allele CEBPA mutations and diverse TP53 mutations were checked as the most common last event mutations. Considering the necessity of last event mutations and limited gene involvement in AML transformations, it is possible to validate a small group of last events involved mutations to develop some new strategies to block MDS progression.

Integration Analysis of JAK2 or RUNX1 Mutation With Bone Marrow Blast Can Improve Risk Stratification in the Patients With Lower Risk Myelodysplastic Syndrome.

Despite the improvements in prognostication of the revised International Prognostic Scoring System (IPSS-R) in myelodysplastic syndrome (MDS), there remain a portion of patients with lower risk (low/intermediate risk, LR) but poor prognostics. This study aimed to evaluate the relative contribution of mutational status when added to the IPSS-R, for estimating overall survival (OS) and progression-free survival (PFS) in patients with LR-MDS. We retrospectively analyzed clinical and laboratory variables of 328 patients diagnosed with MDS according to the FAB criteria. Twenty-nine-gene NGS assay was applied to bone marrow samples obtained at diagnosis. 233 (71.04%) patients were classified as LR-MDS. Univariate analysis showed association between inferior outcome (OS and PFS) and presence of JAK2 (p = 0.0177, p = 0.0002), RUNX1 (p = 0.0250, p = 0.0387), and U2AF1 (p = 0.0227, p = 0.7995) mutations. Multivariable survival analysis revealed JAK2 (p < 0.0001) and RUNX1 (p = 0.0215) mutations were independently prognostic for PFS in LR-MDS. Interestingly, bone marrow blast >1.5% could further predict disease progression of patients with LR-MDS (HR 8.06, 95%CI 2.95-22.04, p < 0.0001). Incorporation of JAK2, RUNX1 mutation and bone marrow blast in the IPSS-R can improve risk stratification in patients with LR-MDS. In summary, our result provided new risk factors for LR-MDS prognostics to identify candidates for early therapeutic intervention.

Cellular senescence induced by S100A9 in mesenchymal stromal cells through NLRP3 inflammasome activation.

Bone marrow stromal cells from patients with myelodysplastic syndrome (MDS) display a senescence phenotype, but the underlying mechanism has not been elucidated. Pro-inflammatory signaling within the malignant clone and the bone marrow microenvironment has been identified as a key pathogenetic driver of MDS. Our study revealed that S100A9 is highly-expressed in lower-risk MDS. Moreover, normal primary mesenchymal stromal cells (MSCs) and the human stromal cell line HS-27a co-cultured with lower-risk MDS bone marrow mononuclear cells acquired a senescence phenotype. Exogenous supplemented S100A9 also induced cellular senescence in MSCs and HS-27a cells. Importantly, Toll-like receptor 4 (TLR4) inhibition or knockdown attenuated the cellular senescence induced by S100A9. Furthermore, we showed that S100A9 induces NLRP3 inflammasome formation, and IL-1ß secretion; findings in samples from MDS patients further confirmed these thoughts. Moreover, ROS and IL-1ß inhibition suppressed the cellular senescence induced by S100A9, whereas NLRP3 overexpression and exogenous IL-1ß supplementation induces cellular senescence. Our study demonstrated that S100A9 promotes cellular senescence of bone marrow stromal cells via TLR4, NLRP3 inflammasome formation, and IL-1ß secretion for its effects. Our findings deepen the understanding of the molecular mechanisms involved in MDS reprogramming of MSCs and indicated the essential role of S100A9 in tumor-environment interactions in bone marrow.

The efficacy and toxicity of the CHG priming regimen (low-dose cytarabine, homoharringtonine, and G-CSF) in higher risk MDS patients relapsed or refractory to decitabine.

PURPOSE: Myelodysplastic syndromes (MDSs) refractory or relapsed after hypomethylating agents (HMAs) remain a therapeutic challenge. The CHG regimen has been demonstrated to be effective in initially treating higher risk MDS. The current study evaluated the efficacy and toxicity of the CHG regimen in patients who were resistant to decitabine. METHODS: Patients with higher risk MDS relapsed or refractory to decitabine were enrolled in this study. Each patient received the CHG regimen (cytarabine (25 mg/day, days 1-14) and homoharringtonine (1 mg/day, days 1-14) intravenously with G-CSF (300 µg/day) subcutaneously from day 0 until neutrophil count recovery to 2.0 × 109 cells/L). Next gene sequencing with a 31-gene panel was carried out in patients. RESULTS: Thirty-three patients were enrolled, including 12 relapsed and 21 refractory cases. The overall response rate (ORR) was 39.4% (13 of 33), with 9 (27.3%) achieving complete remission (CR), 2 having marrow CR (mCR), and 2 achieving partial remission (PR). The CR rate was higher in patients harboring fewer gene mutations (0-1) (55.6%) than in those with more gene mutations (> 1) (12.5%) (p = 0.021). The median overall survival (OS) of the 33 patients was 7.0 months. Patients who achieved a response had significantly longer survival times than were found in those without a response (21.0 M vs. 4.0 M, p < 0.0001). The regimen was endurable for most of the patients. CONCLUSIONS: The CHG priming regimen provided a safe and effective salvage regimen for higher risk MDS patients who were resistant to decitabine. Further studies involving larger samples will be needed. Clinical trial No. ChiCTR-ONC-11001501.

Clinical significance of hyaluronan levels and its pro-osteogenic effect on mesenchymal stromal cells in myelodysplastic syndromes.

BACKGROUND: Hyaluronan (HA), a major component of the extracellular matrix, has been proven to play a crucial role in tumor progression. However, it remains unknown whether HA exerts any effects in myelodysplastic syndromes (MDS). METHODS: A total of 82 patients with MDS and 28 healthy donors were investigated in this study. We firstly examined the bone marrow (BM) serum levels of HA in MDS by radioimmunoassay. Then we determined HA production and hyaluronan synthase (HAS) gene expression in BM mesenchymal stromal cells (MSC) and mononuclear cells derived from MDS patients. Finally, we investigated the effects of HA on osteogenic differentiation of MSC. RESULTS: The BM serum levels of HA was increased in higher-risk MDS patients compared to normal controls. Meanwhile, patients with high BM serum HA levels had significantly shorter median survival than those with low HA levels. Moreover, the HA levels secreted by MSC was elevated in MDS, especially in higher-risk MDS. In addition, HAS-2 mRNA expression was also up-regulated in higher-risk MDS-MSC. Furthermore, we found that MSC derived from MDS patients with high BM serum HA levels had better osteogenic differentiation potential. Moreover, MSC cultured in HA-coated surface presented enhanced osteogenic differentiation ability. CONCLUSIONS: Our results show that elevated levels of BM serum HA are related to adverse clinical outcome in MDS. Better osteogenic differentiation of MSC induced by HA may be implicated in the pathogenesis of MDS.

Dicer1 downregulation by multiple myeloma cells promotes the senescence and tumor-supporting capacity and decreases the differentiation potential of mesenchymal stem cells.

Bone marrow mesenchymal stem cells (BMMSCs) facilitate the growth of multiple myeloma (MM) cells, but the underlying mechanisms remain unclear. This study demonstrates that the senescence of MM-MSCs significantly increased, as evidenced by a decrease in proliferation and increase in the number of cells positive for senescence-associated ß-galactosidase activity. Senescent MM-MSCs displayed decreased differentiation potential and increased tumor-supporting capacity. Dicer1 knockdown in the MSCs of healthy controls promoted cellular senescence and tumor-supporting capacity, while decreasing the differentiation capacity. Dicer1 overexpression in MM-MSCs reversed the effects on differentiation and reduced cellular senescence. In addition, decreased expression of the microRNA-17 family was identified as a favorable element responsible for increasing senescence, with the expression of p21 increased in Dicer1 knockdown cells. Furthermore, we observed decreased expression of miR-93 and miR-20a in MM-MSCs, while upregulation of miR-93/miR-20a decreased cellular senescence, as evidenced by the increased p21 expression. Importantly, we found that myeloma cells could induce the senescence of MSCs from healthy controls, as observed from the decreased expression of Dicer1 and miR-93/miR-20a and increased expression of p21. Overall, MM cells downregulate Dicer1 in MSCs, which leads to senescence; in turn, senescent MSCs promote MM cell growth, which most likely contributes to disease progression.

Iron overload promotes mitochondrial fragmentation in mesenchymal stromal cells from myelodysplastic syndrome patients through activation of the AMPK/MFF/Drp1 pathway.

Iron overload (IO) has been reported to contribute to mesenchymal stromal cell (MSC) damage, but the precise mechanism has yet to be clearly elucidated. In this study, we found that IO increased cell apoptosis and lowered cell viability in MSCs, accompanied by extensive mitochondrial fragmentation and autophagy enhancement. All these effects were reactive oxygen species (ROS) dependent. In MSCs with IO, the ATP concentrations were significantly reduced due to high ROS levels and low electron respiratory chain complex (ETC) II/III activity. Reduced ATP phosphorylated AMP-activated protein kinase (AMPK). Activation of AMPK kinase complexes triggered mitochondrial fission. Moreover, gene knockout of AMPK via CRISPR/Cas9 reduced cell apoptosis, enhanced cell viability and attenuated mitochondrial fragmentation and autophagy caused by IO in MSCs. Further, AMPK-induced mitochondrial fragmentation of MSCs with IO was mediated via phosphorylation of mitochondrial fission factor (MFF), a mitochondrial outer-membrane receptor for the GTPase dynamin-related protein 1 (Drp1). Gene knockdown of MFF reversed AMPK-induced mitochondrial fragmentation in MSCs with IO. In addition, MSCs from IO patients with myelodysplastic syndrome (MDS) showed increased cell apoptosis, decreased cell viability, higher ROS levels, lower ATP concentrations and increased mitochondrial fragmentation compared with MSCs from non-IO patients. In addition, iron chelation or antioxidant weakened the activity of the AMPK/MFF/Drp1 pathway in MDS-MSCs with IO from several patients, accompanied by attenuation of mitochondrial fragmentation and autophagy. Taken together, the AMPK/MFF/Drp1 pathway has an important role in the damage to MDS-MSCs caused by IO.

[Inhibitory Effect of Decitabine on Proliferation of MDS-L Cells and Its Mechanism].

OBJECTIVE: To investigate the inhibitory effect of decitabine (DAC) in various dosages on the proliferention of MDS-RAEB cell line MDS-L and its mechanism. METHODS: LC-MS/MS method was used to test the blood DAC concentration of 2 groups of MDS patients being treated with DAC 20 and 15 mg/m2×5 d. In according to the various blood DAC concentration levels, the MDS-L cells were treated with different DAC dosages for 24, 48, 72 and 96 h, respectively. The CCK-8 method was applied to determine the cell proliferation, the flow cytometry was used to analyze the cell cycle and cell apoptosis changes, the P15INK4B DNA methylation status was measured by methylation specific PCR using EZ DNA Methylation-Gold Kit. RESULTS: The blood DAC concentration of MDS patients treated with DAC 20 mg/m2×5 d was 174.08±80.15(84.7-311) ng/ml, which was significantly higher than 89.87±32.94(43.2-165)ng/ml for the group treated with 15 mg/m2×5 d (P=0.014). DAC could notably inhibit the proliferation of MDS-L cells, and the effect was in dose- and- time-dependent manner(r=0.786). However, when DAC concentration was ≥0.1 µg/ml, the proliferation inhibition rates were not significantly different between various dosages. After DAC treatment, MDS-L cells in G1 phase increased notably, while cells in S phase decreased significantly. Also, the P15INK4B DNA methylation status of MDS-L cells decreased after being treated with DAC for 96 h, but the difference was not significant between various dosages. CONCLUSION: DAC can significantly suppress MDS-L cell proliferation, block MDS-L cells in G1 phase and induce the apoptosis at low concentration (0.1-0.2 µg/ml).