Expression and Clinical Significance of Exosome Derived MiR-181b-5p in Children with Acute Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To explore the expression level of exosome derived miR-181b-5p in different disease stages of children with acute lymphoblastic leukemia and its relationship with clinical characteristics.@*METHODS@#Bone marrow plasma samples of 86 children with ALL were collected. Exosomes were extracted by exosome extraction kit, and RNA in exosomes was extracted by TRIzol method. The levels of miR-181b-5p in the blood plasma exosomes of the patients in the newly diagnosed group, relapse group, remission group and control group were detected by qRT- PCR. The difference of miR-181b-5p expression level in each group was compared and analyzed, and the relationship between miR-181b-5p expression level and clinical characteristics was analyzed.@*RESULTS@#The expression level of exosomal miR-181b-5p in the newly diagnosed group and the relapsed group was significantly lower than that in the remission group and the control group (P< 0.05). The expression level of exosomal miR-181b-5p in T-ALL children was higher than that in B-ALL children (P<0.05). The expression level of plasma exosomal miR-181b-5p in male children was higher than that in female children (P<0.01).@*CONCLUSION@#Exosome derived miR-181b-5p changes dynamically in the course of ALL children, and can be used as a marker miRNA to monitor disease status. Exosomes can transmit information in the tumor microenvironment and serve as a potential carrier for biomolecular targeted therapy.

Study on the Mechanism of Multi-Drug Resistance of Agaricus Blazei Extract FA-2-b-β Mediated Wnt Signaling Pathway to Reverse Acute T Lymphoblastic Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate the mechanism of drug reversing resistance of Agaricus blazei extract FA-2-b-β on T cell acute lymphoblastic leukemia (T-ALL) cell lines.@*METHODS@#Cell proliferation was detected by CCK-8 assay; the apoptosis, cell cycle mitochondrial membrane potential, and intracellular rhodamine accumulation were detected by flow cytometry, and apoptosis-related gene and protein expression were detected by qPCR and Western blot; the membrane surface protein MDR1 was observed by immunofluorescence microscopy.@*RESULTS@#Different concentrations of FA-2-b-β significantly inhibited proliferation and induced apoptosis of CCRF-CEM and CEM/C1 (P<0.05), and CCRF-CEM cell cycle were arrested at S phase, and CEM/C1 cells were arrested at G0/G1 phase. Western blot and qPCR results show that FA-2-b-β inhibited ABCB1、ABCG2、CTNNB、MYC and BCL-2 expression, but upregulated Bax expression. In addition, FA-2-b-β reversed the resistance characteristics of CEM/C1 drug-resistance cells, which decreased mitochondrial membrane potential, and significantly increased the intracellular rhodamine accumulation, and weakening of the expression of the membrane surface protein MDR1. With the Wnt/β-catenin inhibitor (ICG001), the process was further intensified.@*CONCLUSION@#Agaricus Blazei Extract FA-2-b-β inhibits cell proliferation, promotes apoptosis, regulates the cell cycle, reduces mitochondrial energy supply, and down-regulate MDR1 expression to reverse the resistance of CEM/C1, which all suggest it is through regulating the Wnt signaling pathway in T-ALL.

Effect of MXRA7 on the Biological Functions of Acute B Lymphoblastic Leukemia Cell Line REH / 中国实验血液学杂志

OBJECTIVE@#To discover the relationship between matrix remodeling associated 7 (MXRA7) and acute B lymphoblastic leukemia (B-ALL), and explore the effect of MXRA7 on the biological functions of B-ALL cell line REH.@*METHODS@#The expression of MXRA7 in blood diseases was searched and analyzed through BloodSpot database. Real-time qPCR was used to detect the expression level of MXRA7 in B-ALL cell line 697 and REH cells. Lentivirus-mediated shRNA interference technology was utilized to knock down the expression of MXRA7 in REH cells. The effects of MXRA7 on the biological functions of REH cells were studied by in vitro experiments. Cell proliferation was detected by CCK-8 assay, cell cycle was detected by PI staining, cell apoptosis was detected by Annexin V and 7-AAD staining, and the expression of apoptosis pathway related proteins was detected by Western blot.@*RESULTS@#Database analysis showed that MXRA7 was highly expressed in B-ALL patients, and real-time qPCR results showed that MXRA7 was also highly expressed in cell lines 697 and REH cells. Knockdown of MXRA7 in REH cells inhibited the cell proliferation and increased the percentage of G0/G1 phase cells. After treatment with cytarabine, the apoptotic ratio was increased in MXRA7-impaired REH cells, and the activation of caspase-3 and caspase-9 were also increased.@*CONCLUSION@#Knockdown of MXRA7 can reduce the malignancy of REH cells by inhibiting the cell proliferation and increasing the sensitivity of REH cells to cytarabine. These results indicate MXRA7 may be as a novel target for the treatment of B-ALL, and the potential usefulness of MXRA7 in B-ALL deserves further investigation.

Characteristics of fusion gene expression in acute lymphoblastic leukemia / 中华病理学杂志

Objective: To analyze the genetic landscape of 52 fusion genes in patients with de novo acute lymphoblastic leukemia (ALL) and to investigate the characteristics of other laboratory results. Methods: The fusion gene expression was retrospectively analyzed in the 1 994 patients with de novo ALL diagnosed from September 2016 to December 2020. In addition, their mutational, immunophenotypical and karyotypical profiles were investigated. Results: In the 1 994 patients with ALL, the median age was 12 years (from 15 days to 89 years). In the panel of targeted genes, 15 different types of fusion genes were detected in 884 patients (44.33%) and demonstrated a Power law distribution. The frequency of detectable fusion genes in B-cell ALL was significantly higher than that in T-cell ALL (48.48% vs 18.71%), and fusion genes were almost exclusively expressed in B-cell ALL or T-cell ALL. The number of fusion genes showed peaks at<1 year, 3-5 years and 35-44 years, respectively. More fusion genes were identified in children than in adults. MLL-FG was most frequently seen in infants and TEL-AML1 was most commonly seen in children, while BCR-ABL1 was dominant in adults. The majority of fusion gene mutations involved signaling pathway and the most frequent mutations were observed in NRAS and KRAS genes. The expression of early-stage B-cell antigens varied in B-cell ALL patients. The complex karyotypes were more common in BCR-ABL1 positive patients than others. Conclusion: The distribution of fusion genes in ALL patients differs by ages and cell lineages. It also corresponds to various gene mutations, immunophenotypes, and karyotypes.

Yin-Yang-1 decreases Fas-induced apoptosis in acute lymphoblastic leukemia under hypoxic conditions: its implications in immune evasion / YY1 induce la disminución de la apoptosis a través de Fas en la leucemia linfoblástica aguda en condiciones hipóxicas: implicaciones en la evasión de la respuesta inmunitaria

Abstract Background: Acute lymphoblastic leukemia (ALL) is an aggressive malignant disease with high prevalence in pediatric patients. It has been shown that the downregulation of Fas expression is correlated with an inadequate response in ALL, although these mechanisms are still not well understood. Several reports demonstrated that hypoxia is involved in dysfunctional apoptosis. Yin-Yang-1 (YY1) transcription factor is involved in resistance to apoptosis, tumor progression, and it is increased in different types of cancer, including leukemia. The regulatory mechanism underlying YY1 expression in leukemia is still not understood, but it is known that YY1 negatively regulates Fas expression. The study aimed to evaluate the effect of YY1 on Fas expression under hypoxic conditions in ALL. Methods: Leukemia cell line RS4; 11 was cultured under normoxic and hypoxic conditions. YY1, Fas receptor, and hypoxia-inducible factor (HIF-1α) expression were analyzed. After treatment with a Fas agonist (DX2), apoptosis was analyzed through the detection of active caspase 3. Data were analyzed using Pearson’s correlation. Results: Leukemia cells co-expressed both HIF-1α and YY1 under hypoxia, which correlated with a downregulation of Fas expression. During hypoxia, the levels of apoptosis diminished after DX2 treatment. The analysis revealed that patients with high levels of HIF-1α also express high levels of YY1 and low levels of Fas. Conclusions: These results suggest that YY1 negatively regulates the expression of the Fas receptor, which could be involved in the escape of leukemic cells from the immune response contributing to the ALL pathogenesis.

Resumen Introducción: La leucemia linfoblástica aguda (LLA) es una enfermedad con alta prevalencia en la población pediátrica. El mecanismo por el cual el receptor de Fas participa en la regulación inmunitaria en los tumores es desconocido, pero se sabe que está subexpresado en LLA. El factor de transcripción Ying-Yang-1 (YY1) está involucrado en la resistencia a la apoptosis y la progresión tumoral; se encuentra aumentado en diferentes tumores, incluida la LLA. Aunque los mecanismos que regulan la expresión de YY1 en LLA son desconocidos, se sabe que YY1 regula la expresión del receptor de Fas. El objetivo de este trabajo fue evaluar el efecto de YY1 en la expresión de Fas en condiciones de hipoxia en la LLA. Métodos: Se cultivaron células RS4;11 en condiciones de hipoxia y se analizó la expresión de YY1, receptor de Fas y HIF-1α. La apoptosis fue inducida usando un agonista de Fas (DX2) y se analizó con la detección de caspasa 3 activa. Los datos se analizaron mediante correlación de Pearson. Resultados: Las células RS4;11 coexpresaron HIF-1αy YY1 en hipoxia, lo cual correlaciona con una baja expresión de Fas. La apoptosis se encontró disminuida durante condiciones de hipoxia, después del tratamiento con DX2. El análisis bioinformático mostró que los pacientes con altos niveles de HIF-1αpresentan YY1 elevado y bajos niveles del receptor de Fas. Conclusiones: Estos resultados sugieren que YY1 regula negativamente la expresión del receptor de Fas, lo cual podría estar involucrado en el escape de las células leucémicas a la respuesta inmunitaria, contribuyendo a la patogénesis de la LLA.

Estimation of salivary amylase and total proteins in leukemia patients and its correlation with clinical feature and radiographic finding.

Background: Leukemia is a fatal disease. The oral manifestations of the leukemias occur early in the course of the disease and these oral features can at times act as a diagnostic indicator. Saliva has been used as a diagnostic aid in a number of systemic diseases. Materials and Methods: In our study, samples of unstimulated saliva of 30 leukemia patients who were not on chemotherapy were collected and analyzed for salivary amylase and total protein. The oral manifestations and radiographic changes (OPG) were recorded. The correlation between the oral manifestations and the salivary components (salivary amylase and total protein) was assessed for prognostic significance. Results: In the present study when the mean values of salivary amylase (1280±754 U/ml) and total protein (647.2±320.7 mg%) were compared with that in control subjects. There was a statistically significant difference for amylase levels (P<.05). On intraoral examination the study subjects showed pallor, gingivitis, gingival enlargement, petechiae, and ecchymosis. On the OPG, the radiographic features included generalized rarefaction of bone (20%), thinning of lamina dura (3.4%), generalized alveolar crest bone resorption (30%), thinning of walls of alveolar crypts (6.7%), besides others, e.g., periapical abscess (10%). Conclusions: The saliva of leukemic patients demonstrated obvious changes in composition. A rise in salivary amylase and total protein levels was evident, with the increase in amylase levels being statistically significant.

Aspectos mamográficos e ultra-sonográficos de leucemia linfóide aguda na mama masculina / Mammography and ultrasound in a case of acute lymphoblastic leukemia in the male breast

A neoplasia mamária no homem é um evento pouco freqüente, quanto mais em paciente jovem.Quando essa neoplasia é do tipo leucemia linfóide aguda torna-se uma situação rara. Dessa forma, odiagnóstico apropriado com métodos de imagem, como mamografia e ultra-sonografia, além da biópsiacom agulha e do estudo imunoistoquímico, auxiliam na indicação da terapêutica sistêmica.

The male breast localization of an acute lymphoblastic leukemia is a rare event. The use of mammography,ultrasound and fine needle or core biopsy could help in this diagnosis. The imunohistochemistrycomplete an orientation for the systemic treatment.

Comparative analysis of differential expression of sialic acids and adhesion molecules on mononuclear cells of bone marrow and peripheral blood in childhood acute lymphoblastic leukaemia at diagnosis and clinical remission.

Childhood acute lymphoblastic leukaemia (ALL) is characterized by the neoplasm of immature haematopoietic precursor cells (HPCs). We report significant differences between the expression of sialoglycoproteins and adhesion molecules on mononuclear cells (MNCs) of bone marrow (BM) and peripheral blood (PB) from individual children at diagnosis of the disease. Lymphoblasts in PB predominantly expressed 9-O-acetylated sialoglycoproteins (Neu5,9Ac2-GPs), sialic acid, alpha2-3 linked sialic acid, L- and P-selectins and vascular cell adhesion molecule -1 (VCAM-1) on their surface compared to BM, as determined with selective lectins and monoclonal antibodies (mAbs) by flow cytometric analysis. CD34+CD38+ cells present either in diagnostic PB or BM always showed enhanced expression of both alpha2-3 and alpha2-6 linked sialic acids, Neu5,9Ac2-GPs, L- and P-selectins and VCAM-1, compared to CD34+CD38- population, as confirmed by higher mean fluorescence intensity (MFI). Expression of ICAM-1 was reverse. However, MFI of Neu5,9Ac2-GPs was always higher both in CD34+CD38+ and CD34+CD38- population in PB compared to BM. Diverse trend of these cell surface macromolecules was observed during clinical remission. This is the first comparative study between PB and BM, where significant differential distribution of sialylated macromolecules and adhesion molecules was observed. Hence, supervising these cell surface macromolecules at various stages of treatment might help in minimal residual disease detection, identifying mobilization factor(s) and in isolation of normal HPCs for autologous BM transplantation.

O-acetyl sialic acid binding lectin as a probe for detection of subtle change on cell surface induced during acute lymphoblastic leukemia (ALL) and its clinical application.

A novel probe, a 9-O-acetylated sialic acid binding lectin, namely achatininH (ATNH) has been used for the detection of changes on the cell surface during acute lymphoblastic leukemia (ALL). ATNH does not agglutinate normal human erythrocytes, however it is capable of agglutinating erythrocytes and peripheral blood mononuclear cells (PBMC) of patients suffering from ALL. The differential expression of a key receptor, 9-O-acetylated sialo glyco conjugate (9-O-AcSG), on PBMC was observed using a simple lymphoproliferative assay (LA). The extent of expression of 9-O-AcSG was used as an index to distinguish ALL patients of different clinical stages and assess the probability of relapse. The amount of ATNH needed for maximum stimulation served as a tool to indirectly measure the extent of expression of 9-O-AcSG on PBMC surface. The acetylated sialo glycoconjugate was expressed at a very high concentration during acute phase of the disease. Subsequently it decreased during treatment persisted during maintenance therapy and reappeared with relapse. PBMC of normal human donors required 80 times more ATNH in comparison to the untreated acute phase ALL patients. No cross reactivity was found in non Hodgkin's lymphoma, chronic myelogenous leukemia and thalassaemia patients.

Alteraciones bioquímicas y citoquímicas de hidrolasas ácidas lisosomales en leucemias / Biochemical and citochemical alterations of lisosomal acid hydrolases in leukemias

La actividad enzimática total de ß-galactosidasa (ß-gal), hexosaminidasa (hex) y fosfatasa ácida (Fac) fue determinada bioquímicamente, tanto en suero como en sobrenadantes de homogenatos celulares de sujetos normales y pacientes portadores de leucemias, empleándose sustratos paranitrofenilados específicos. La actividad de ß-gal en leucemias mieloides, tanto agudas (LMA-M1) como crónicas (LMC), sólo mostró un incremento significativo en sobrenadantes de homogenato celulares, respecto a los valores de neutrófilos normales. En leucemias linfoidales agudas (LLA), como crónicas (LLC), su comportamiento no ofreció variaciones. Tanto los sueros como los sobrenadantes de homogenatos celulares de LMA-M1 y LMC mostraron un franco incremento en la actividad de hex, mientras en LLA y LLC esta actividad no mostró variaciones. La actividad sérica de fosfatasa ácida estuvo incrementada en el 86% de las LMC. En los sobrenadantes de homogenatos celulares de LLA y LLC, esta actividad enzimática se mostró significativamente disminuida respecto de los valores de linfocitos normales. En los tres casos de LMA-M4 analizados, fue observado en el contenido celular niveles elevados de ß-gal, hex y Fac (lo que estaría correlacionado con la presencia de monocitos y/o monoblastos normales, con alto contenido de hidrolasas ácidas). Citoquímicamente fue demostrado en médula ósea y sangre periférica de pacientes con LMA-M1 una ligera o nula actividad de hex, en tanto que en LLA la reacción fue localizada asimétricamente en un polo celular o en gránulos citoplasmáticos. Los resultados encontrados demuestran una gran heterogeneidad en el contenido lisosomal de los diferentes tipos de leucemias

Simplified method for the analysis of cellular karyotype and phenotype in leukemias

The objective of this study was to develop a simplified method for the simultaneous analysis of cellular karyotype and phenotype which would permit the identification of cell origin. We studied 6 patients with AML, 3 with CML (one of which was in blastic transformation) and one ALL. We used a method in which the suspension of bone marrow cells was incubated in TC 199 medium with colchicine and with hypotonic solution formed from glycerol, NaCl, KCl, CaCl2, MgCl2 and sucrose. The slides were prepared from this cell suspension by cytospin and stained for peroxidase, PAS, esterases and iron. The karyotype was studied by direct method and culture. It was possible to relate the cytogenetic marker with cytochemistry characteristics in the same cell in 3 cases, showing the feasibility of cytochemistry techniques in cytogenetical preparations. The best preparations were found through peroxidase. The presence of iron granules allowed identification of erythroblastic lineage in the combined staining. Mitosis with a marker chromosome of leukemic clone in an AML cell with negative peroxidase probably showed a proliferation of more primitive precursor not sufficiently differentiated to show markers