Down-regulation of Musashi-2 exerts antileukemic effects on acute lymphoblastic leukemia cells and increases sensitivity to dexamethasone.

Musashi-2 (MSI2), implicated in the oncogenesis and propagation of a broad array of malignancies, inclusive of certain leukemia, remains a nascent field of study within the context of acute lymphoblastic leukemia (ALL). Using lentiviral transfection, ALL cells with stable MSI2 knockdown were engineered. A suite of analytic techniques - a CCK-8 assay, flow cytometry, qRT-PCR, and western blotting - were employed to evaluate cellular proliferation, cell cycle arrest, and apoptosis and to confirm differential gene expression. The suppression of MSI2 expression yielded significant results: inhibition of cell proliferation, G0/G1 cell cycle arrest, and induced apoptosis in ALL cell lines. Furthermore, it was noted that MSI2 inhibition heightened the responsiveness of ALL cells to dexamethasone. Significantly, the depletion of MSI2 prompted the translocation of GR from the cytoplasm to the nucleus upon dexamethasone treatment, consequently leading to enhanced sensitivity. Additionally, the FOXO1/4 signaling pathway contributed to the biological effects of ALL cells evoked by MSI2 silencing. Our study offers novel insight into the inhibitory effects of MSI2 suppression on ALL cells, positing MSI2 as a promising therapeutic target in the treatment of ALL.

Decreased serum apolipoprotein A1 level predicts poor prognosis of patients with de novo myelodysplastic syndromes.

BACKGROUND: Myelodysplastic syndromes (MDS) is a group of heterogeneous myeloid clonal diseases originating from hematopoietic stem cells. It has been demonstrated that apolipoproteins A1(ApoA1) are associated with disease risk in many cancer types. However, there still lacks evidence regarding the link between ApoA1 and MDS. This study was designed to investigate the prognostic value of pretreatment ApoA1 levels in MDS patients. METHODS: We retrospectively analyzed a cohort of 228 MDS patients to explore the prognostic value of the serum ApoA1 levels at diagnosis. Patients were divided into the high ApoA1 group and the low ApoA1 group. The prognostic significance was determined by univariate and multivariate Cox hazard models. RESULTS: MDS patients with low ApoA1 levels had significantly shorter overall survival (OS, P < 0.0001) along with a higher frequency of TP53 mutation (P = 0.002). Based on univariate analysis, age (≥ 60 years), gender (male), lower levels of hemoglobin (< 10 g/dl), HDL (≤0.91 mmol/L), higher bone marrow blast percentage (> 5%), higher IPSS-R scores and poorer karyotype were significantly associated with decreased OS. However, low ApoA1 level did not influence leukemia-free survival (LFS, P = 0.367). Multivariate Cox proportional hazards regression analysis indicated that low ApoA1 level (≤ 1.02 g/L) was also an independent adverse prognostic factor for OS in MDS (P = 0.034). CONCLUSIONS: Decreased ApoA1 level predicts a poor prognosis of MDS patients and thus provides a novel evaluation factor for them that is independent of the IPSS-R system.

Ameliorative effect of melatonin against increased intestinal permeability in diabetic rats: possible involvement of MLCK-dependent MLC phosphorylation.

The increased intestinal permeability and functional impairment play an important role in type 2 diabetes (T2D), and melatonin may possess enteroprotection properties. Therefore, we used streptozotocin-induced diabetic rat model to investigate the regulation of intestinal permeability by melatonin. Rats were randomly divided into three groups, including control, diabetes mellitus (DM), and DM rats treated with melatonin. Melatonin was administered (10 mg/kg/day) by gavage for 24 weeks. The DM rats significantly increased the serum fasting blood glucose and lipid levels, which were alleviated by melatonin treatment. Importantly, the intestinal epithelial permeability was significantly increased in DM rats but was ameliorated following treatment with melatonin. These findings also indicated the expression of myosin light chain kinase (MLCK) and phosphorylation of MLC targeting subunit (MYPT) induced myosin light chain (MLC) phosphorylation level was markedly elevated in hyperglycemic and hyperlipidemic status. They were partly associated with down-regulated membrane type 1 and 2 (MT1 and MT2) expression, and up-regulated Rho-associated protein kinase (ROCK) expression and increased extracellular signal-regulated kinase (ERK) phosphorylation. However, the changes in target protein expression were reversed by melatonin. In conclusion, our results show melatonin beneficial effects on impaired intestinal epithelial permeability in T2D by suppressing ERK/MLCK- and ROCK/MCLP-dependent MLC phosphorylation.

Circ_0012152 Accelerates Acute Myeloid Leukemia Progression through the miR-652-3p/SOX4 Axis.

OBJECTIVE: Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by abnormal myeloid blast expansion. Recent studies have demonstrated that circular RNAs play a role in AML pathogenesis. In this study, we aimed to investigate the clinical significance of circ_0012152 in AML and elucidate its underlying molecular mechanism in the pathogenesis of this condition. METHODS: Circ_0012152 expression was detected by quantitative real-time polymerase chain reaction in samples obtained from 247 patients with AML and 40 healthy controls. A systematic analysis of clinical characteristics and prognostic factors was also conducted. Cell growth was assessed using the Cell Counting Kit-8 (CCK-8) assay, and apoptosis and cell cycle progression were evaluated by flow cytometry. Moreover, RNA pull-down was performed to identify target microRNAs, and transcriptome RNA sequencing and bioinformatics analyses were utilized to identify downstream mRNA targets. RESULTS: Circ_0012152 was significantly upregulated in samples from patients with AML and served as an independent adverse prognostic factor for overall survival (OS) (hazard ratio: 2.357; 95% confidence interval 1.258-4.415). The circ_0012152 knockdown reduced cell growth, increased apoptosis, and inhibited cell cycle progression in AML cell lines. RNA pull-down and sequencing identified miR-652-3p as a target microRNA of circ_0012152. Cell growth inhibition by circ_0012152 knockdown was significantly relieved by miR-652-3p inhibitors. We suggested that miR-652-3p targeted SOX4, as the decrease in SOX4 expression resulting from circ_0012152 knockdown was upregulated by miR-652-3p inhibitors in AML cells. CONCLUSION: Circ_0012152 is an independent poor prognostic factor for OS in AML, and it promotes AML cell growth by upregulating SOX4 through miR-652-3p.

Decreased transthyretin predicts a poor prognosis in primary myelodysplastic syndrome.

Background: This study aims to investigate the prognostic significance of transthyretin in newly diagnosed myelodysplastic syndromes (MDS). Methods: The clinical, laboratory, and follow-up data of 280 newly diagnosed patients with MDS were collected. The relationship between serum transthyretin levels and overall survival (OS) and leukemia-free survival (LFS) were analyzed by Kaplan-Meier analysis and Cox Regression Model. Result: In the MDS cohort, there were 121 cases in the low transthyretin group and 159 cases in the normal transthyretin group. MDS patients with decreased transthyretin had a higher risk score on the Revised International Prognostic Scoring System (IPSS-R) (p = 0.004) and on the molecular IPSS (IPSS-M) (p = 0.005), a higher frequency of TP53 mutation (p < 0.0001), a shorter OS (p < 0.0001) and LFS (p < 0.0001). Multivariate analyses showed that higher IPSS-R and IPSS-M score were adverse factors for OS (p = 0.008 and p = 0.015, respectively) and LFS (p = 0.024 and p = 0.005, respectively). Mutations of TP53 and NRAS were also poor factors for LFS (p = 0.034 and p = 0.018, respectively). Notably, decreased transthyretin was an independent adverse predictor for OS (p = 0.009, HR = 0.097, 95%CI, 0.017-0.561) but not for LFS (p = 0.167) when IPSS-R was included in the Cox regression model and an independent poor one for OS (p = 0.033, HR = 0.267, 95%CI, 0.080-0.898) and LFS (p = 0.024, HR = 0.290, 95%CI, 0.099-0.848) while IPSS-M involved. Conclusion: The results indicate that decreased transthyretin could be an independent adverse prognostic factor in patients with MDS and may provide a supplement to IPSS-R and IPSS-M.

MLL-SEPT5 Fusion Transcript in Myelodysplastic Syndrome Patient With t(11;22)(q23;q11).

Objectives: This study aimed to identify unknown mixed lineage leukemia (MLL) translocation partner genes in a de novo patient with myelodysplastic syndrome (MDS) with t(11;22)(q23;q11) and investigate the clinical and molecular features of this patient. Methods: Bone marrow cells were assessed by karyotype analysis to reveal chromosomal abnormalities. Fluorescence in situ hybridization (FISH) was performed to detect MLL gene rearrangement using an MLL-specific break-apart probe. LDI-PCR and RT-PCR were performed, and the PCR products were sequenced using an Illumina MiSeq sequencer (Illumina, San Diego, CA, USA). The sequence data of the PCR products were analyzed using bioinformatics tools. Meanwhile, clinical data were collected to evaluate the prognosis of the patient. Results: Chromosomal karyotype analysis showed that the karyotype of the patient was 46, XX, t(11;22)(q23;q11)[10]/46, XX[1]. Subsequently, FISH data confirmed MLL gene rearrangement in the patient. LDI-PCR precisely showed that SEPT5 was the MLL translocation partner gene. RT-PCR and sequencing analysis disclosed the presence of MLL-SEPT5 fusion transcript and confirmed the fusion between MLL exon 8 and SEPT5 exon 3. Moreover, the patient had a recurrence shortly after allogeneic hematopoietic stem cell transplantation. Conclusion: Although the MLL-SEPT5 fusion transcript was occasionally described in acute myeloid leukemia, it was first identified in MDS. Patients with MLL-SEPT5 fusion gene exhibited a poor prognosis even with an aggressive treatment.

Prognostic significance of U2AF1 mutations in myelodysplastic syndromes: a meta-analysis.

INTRODUCTION: Although the effects of U2 small nuclear RNA auxiliary factor 1 gene (U2AF1) mutations on the outcomes of patients with myelodysplastic syndromes (MDS) have previously been investigated, their prognostic significance remains controversial. We performed a meta-analysis to investigate the impact of U2AF1 mutations on MDS progression. METHODS: Two reviewers independently extracted information such as hazard ratios (HRs) and 95% confidential intervals (CIs) for overall survival (OS) and leukemia-free survival (LFS) as well as the number of surviving patients each year after diagnosis from the included studies. RESULTS: Thirteen studies with a total of 3038 patients were included. The summary odds ratio (OR) for U2AF1 mutations with an OS of 5 years was 0.37, the summary HR for U2AF1 mutations in OS was 1.60, and the summary OR for an OS of 5 years in patients with U2AF1S34 and U2AF1Q157 was 3.68. There were no significant differences in leukemia-free survival or hypomethylating therapy response between patients with and without U2AF1 mutations. CONCLUSION: U2AF1 mutations were associated with poor survival in MDS patients, and patients with U2AF1Q157 had a worse OS than those with U2AF1S34. Our findings suggest that MDS patients with U2AF1 mutations could benefit more from hypomethylation therapy.

A Novel External Quality Assessment of Chromosomal Karyotype Analysis Based on Chimera Image Assembly.

OBJECTIVE: This study aimed to establish a new external quality assessment (EQA) of chromosomal karyotype analysis. METHODS: Chimeric assembly A1 was established by collecting chimeric chromosome images prepared artificially from chromosomally abnormal amniocytes remaining after prenatal diagnosis. Chimeric assembly B1 and nonchimeric assembly C1 were constructed through the collection of chimeric and nonchimeric chromosome images from prenatal diagnosis, respectively. Then, chromosome images were selected randomly from assemblies A1, B1, or C1 to send to 20 technicians via email to verify the validity of a new EQA of chromosomal karyotype analysis. RESULTS: According to the EQA of 20 technicians, 47,XX,+mar from assembly A was easily misdiagnosed as 47,XX,+19 or 47,XXY, and 45,XX,t(13;22) (q10;q10) was misdiagnosed as 45,XX,13S+,-22. The total misdiagnosis rate was 3.8%. For assembly B, 46,X,+mar and 46,X,idic(Y) were easily misdiagnosed as 46,XY and 46,X,+mar, respectively. In addition, some testers missed 47,XXX in 47,XXX[2]/46,XX[48], as well as 47,XX,+18 in 46,XX [47]/47,XX,+18[3], and 45,X and 47,XXX in 46,XX[47]/45,X[2]/47,XXX[1]. The total misdiagnosis rate was 4.2%. All karyo-types from assembly C were correctly diagnosed, although incorrect descriptions used for 4% of cases. CONCLUSION: The quality of chromosome karyotype analysis can be comprehensively evaluated by a new EQA based on assembly A1 or B1.

Hsa_circ_0012152 and Hsa_circ_0001857 Accurately Discriminate Acute Lymphoblastic Leukemia From Acute Myeloid Leukemia.

Acute leukemia (AL) is a group of highly heterogeneous hematological malignancies. Circular RNAs (circRNAs) are covalently closed circRNA molecules implicated in the development of many diseases. However, the role of circRNAs in AL remains largely unknown. Therefore, this study aimed to identify new classification diagnostic biomarkers for subgroups of AL. The circRNA expression signatures discriminating acute lymphoblastic leukemia (ALL) from acute myeloid leukemia (AML) were identified by microarray, followed by reverse transcription quantitative polymerase chain reaction (RT-qPCR) validation. Receiver operating characteristic curve analysis was used to evaluate the diagnostic efficiencies of hsa_circ_0001857 and hsa_circ_0012152, and hsa_circ_0012152 was selected for Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis. The results showed that the circRNA expression profiles, hsa_circ_0001857, and hsa_circ_0012152 could clearly discriminate ALL from AML. The target genes of hsa_circ_0012152 might be involved in biological processes, such as myeloid cell differentiation, covalent chromatin modification, histone modification, and rat sarcoma (Ras) protein signal transduction, and participate in pathways such as mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3'-kinase (PI3K)-Akt signaling pathway. Hsa_circ_0012152 might be involved in the initiation and development of AML through miR-491-5p/epidermal growth factor receptor (EGFR)/MAPK1 or miR-512-3p/EGFR/MAPK1 axis. Our results showed that circRNA expression profiles and specifically expressed circRNAs were promising classification biomarkers to designate AL into ALL or AML.

Tip of the iceberg: roles of circRNAs in hematological malignancies.

Circular RNAs (circRNAs) are a new class of covalently closed RNA molecules whose 3'- and 5'-ends are linked by a back-splicing event. Emerging evidence has shown that circRNAs play a vital role in the occurrence and development of many diseases and are promising biomarkers and therapeutic targets. However, knowledge of circRNAs in hematological malignancies is limited. In this review, the biogenesis, categories, characteristics, and functions of circRNAs are summarized, especially the roles of circRNAs in hematopoiesis and hematological malignancies.

Melatonin inhibits colon cancer RKO cell migration by downregulating Rho­associated protein kinase expression via the p38/MAPK signaling pathway.

Melatonin is predominately produced and secreted by the pineal gland, and inhibits cell growth in various cancer cell lines such as colorectal cancer. However, the precise mechanisms involved have not been fully elucidated. In the present study, the potential molecular mechanism underlying the efficacy of melatonin on migration in RKO colon cancer cells was investigated. The effects of melatonin and H­1152, a selective inhibitor of Rho­associated protein kinase (ROCK), on the migration of RKO cells were analyzed by an in vitro wound healing assay. The localization of zonula occludens­1 (ZO­1) and occludin were observed by immunofluorescence. Reverse transcription­quantitative polymerase chain reaction (qPCR) was performed to analyze the relative mRNA levels of ROCK, ZO­1 and occludin. In addition, western blot analysis was implemented to examine the expression of ROCK, phospho (p)­myosin phosphatase targeting subunit 1 (MYPT1), p­myosin light chains (MLC) and p­p38. The results revealed that the expression levels of ROCK2, p­MYPT1 and p­MLC in RKO cells were decreased, and the membrane protein expression of ZO­1 and occludin increased when the cells were treated with melatonin. qPCR demonstrated that melatonin downregulated ROCK2 gene expression, and upregulated the expression of the ZO­1 and occludin genes. The levels of ZO­1 and occludin localized in the tight junctions were markedly increased in the immunofluorescence assay. In addition, the phosphorylation levels of p38 were reduced when the cells were treated with melatonin, and treatment with H­1152 downregulated p38 phosphorylation. The results indicated that melatonin may inhibit the migration of RKO colon cancer cells by downregulating ROCK expression via the p38/mitogen­activated protein kinase signaling pathway.

Melatonin inhibits the Migration of Colon Cancer RKO cells by Down-regulating Myosin Light Chain Kinase Expression through Cross-talk with p38 MAPK.

BACKGROUND: Melatonin, which is mainly produced by the pineal gland, has a good inhibitory effect on cell growth of multiple cancer types. However, the underlying molecular mechanisms of anti-tumor activity for colon cancer have not been fully elucidated. In this study, we investigated the effects of melatonin on migration in human colon cancer RKO cells and the potential molecular mechanisms. MATERIALS AND METHODS: The viability of RKO cells was investigated by MTT assay after treatment with melatonin, SB203580 (p38 inhibitor) and phorbol 12-myristate 13-acetate (PMA, MAPK activator) alone or in combination for 48h. The effects of melatonin, and ML-7, a selective inhibitor of myosin light chain kinase (MLCK), and SB203580, and PMA on the migration of RKO cells were analyzed by in vitro scratch-wound assay. The relative mRNA levels of MLCK was assessed by real-time quantitative RT-PCR. Western blotting analysis was performed to examine the expression of MLCK, phosphorylation of myosin light chain (pMLC) and p38 (pp38). RESULTS: The proliferation and migration of human colon cancer RKO cells were inhibited significantly after treatment with melatonin. The expression levels of MLCK and phosphorylation of MLC of RKO cells were reduced, and real-time quantitative RT-PCR showed that melatonin had significant effects on suppressing the expression of MLCK. Furthermore, the phosphorylation level of p38, which showed the same trend, was also reduced when cells were treated by melatonin. In addition, ML-7 (25umol/l) could down-regulate the phosphorylation of p38. CONCLUSIONS: Melatonin could inhibit the proliferation and migration of RKO cells, and further experiments confirmed that p38 MAPK plays an important role in regulating melatonin-induced migration inhibition through down-regulating the expression and activity of MLCK.