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1.
Hematol., Transfus. Cell Ther. (Impr.) ; 43(4): 430-436, Oct.-Dec. 2021. graf, ilus
Article in English | LILACS | ID: biblio-1350823

ABSTRACT

ABSTRACT Background: In Philadelphia chromosome-negative myeloproliferative neoplasm (MPN) models, reactive oxygen species (ROS) are elevated and have been implicated in genomic instability, JAK2/STAT signaling amplification, and disease progression. Although the potential effects of ROS on the MPN phenotype, the effects of ruxolitinib treatment on ROS regulation have been poorly explored. Herein, we have reported the impact of ruxolitinib on redox signaling transcriptional network, and the effects of diphenyleneiodonium (DPI), a pan NOX inhibitor, in JAK2V617F-driven cellular models. Method: Redox signaling-related genes were investigated in SET2 cells upon ruxolitinib treatment by RNA-seq (GEO accession GSE69827). SET2 and HEL cells, which represent JAK2V617F-positive MPN cellular models with distinct sensitivity to apoptosis induced by ruxolitinib, were used. Cell viability was evaluated by MTT, apoptosis by annexin V/PI and flow cytometry, and cell signaling by quantitative PCR and Western blot. Main results: Ruxolitinib impacted on a network composed of redox signaling-related genes, and DUOX1 and DUOX2 were identified as potential modulators of ruxolitinib response. In SET2 and HEL cells, DPI reduced cell viability and, at low doses, it significantly potentiated ruxolitinib-induced apoptosis. In the molecular scenario, DPI inhibited STAT3, STAT5 and S6 ribosomal protein phosphorylation and induced PARP1 cleavage in JAK2V617F-positive cells. DPI combined with ruxolitinib increased PARP1 cleavage in SET2 cells and potentiated ruxolitinib-reduced STAT3, STAT5 and S6 ribosomal protein in HEL cells. Conclusion: Our study reveals a potential adaptation mechanism for resistance against ruxolitinib by transcriptionally reprogramming redox signaling in JAK2V617F cells and exposes redox vulnerabilities with therapeutic value in MPN cellular models.


Subject(s)
Janus Kinase 2 , Myelodysplastic-Myeloproliferative Diseases/drug therapy , Oxidation-Reduction , NADPH Oxidases , Dual Oxidases , Myeloproliferative Disorders
2.
Clinics ; 76: e2348, 2021. graf
Article in English | LILACS | ID: biblio-1153978

ABSTRACT

OBJECTIVES: TTP488, an antagonist of the receptor for advanced glycation end-products, was evaluated as a potential treatment for patients with mild-to-moderate Alzheimer's disease (AD). However, the mechanism underlying the protective action of TTP488 against AD has not yet been fully explored. METHODS: Healthy male rats were exposed to aberrant amyloid β (Aβ) 1-42. Lipopolysaccharide (LPS) and the NOD-like receptor family pyrin domain containing 1 (NLRP1) overexpression lentivirus were injected to activate the NLRP1 inflammasome and exacerbate AD. TTP488 was administered to reverse AD injury. Finally, tofacitinib and fludarabine were used to inhibit the activity of Janus tyrosine kinase (JAK) and signal transducer and activator of transcription (STAT) to prove the relationship between the JAK/STAT signaling pathway and TTP488. RESULTS: LPS and NLRP1 overexpression significantly increased the NLRP1 levels, reduced neurological function, and aggravated neuronal damage, as demonstrated by the impact latency time of, time spent by, and length of the platform covered by, the mice in the Morris water maze assay, Nissl staining, and immunofluorescence staining in rats with AD. CONCLUSIONS: TTP488 administration successfully reduced AD injury and reversed the aforementioned processes. Additionally, tofacitinib and fludarabine administration could further reverse AD injury after the TTP488 intervention. These results suggest a new potential mechanism underlying the TTP488-mediated alleviation of AD injury.


Subject(s)
Animals , Male , Mice , Rats , Janus Kinases/metabolism , Alzheimer Disease/drug therapy , Tyrosine , Transducers , Signal Transduction , Amyloid beta-Peptides , Janus Kinase 2 , Receptor for Advanced Glycation End Products , Imidazoles
3.
Journal of Experimental Hematology ; (6): 1540-1547, 2021.
Article in Chinese | WPRIM | ID: wpr-922292

ABSTRACT

OBJECTIVE@#To analyze the disease types, clinical manifestations, efficacy and outcome of JAK2 V617F and BCR-ABL double-mutant myeloproliferative neoplasms (MPN), and provide a reference for the diagnosis, treatment and prognosis of MPN.@*METHODS@#The clinical characteristics, diagnosis, therapeutic efficacy and outcome of JAK2 V617F and BCR-ABL double-mutant MPN were analyzed comprehensitively by combining a clinical case diagnosed and treated in our hospital with literature cases from CNKI and PubMed databases.@*RESULTS@#A total of 38 related literatures were retrieved from the two databases by searching "JAK2 V617F" and "BCR-ABL" as key words from 1990 to 2019, and 59 cases were involved. Among all the 60 cases, 41 were males (68.3%) with a median age of 61 (32-77) years old, while 19 were females (31.7%) with a median age of 58 (21-82) years old. The BCR-ABL fusion gene and JAK2 V617F mutation were found simultaneously in 21 cases (35%), 19 cases (31.7%) with JAK2 V617F mutation were found during the treatment of Philadelphia chromosome (Ph)-positive chronic myelogenous leukemia (CML). Ph@*CONCLUSION@#As cases of BCR-ABL and JAK2 V617F double-mutant MPN are reported, simultaneous detection of JAK2 V617F mutation and BCR-ABL fusion gene in MPN patients is necessary to avoid misdiagnosis and missed diagnosis.


Subject(s)
Adult , Aged , Aged, 80 and over , Female , Fusion Proteins, bcr-abl/genetics , Humans , Janus Kinase 2/genetics , Male , Middle Aged , Myeloproliferative Disorders/genetics , Polycythemia Vera , Thrombocythemia, Essential , Young Adult
4.
Journal of Experimental Hematology ; (6): 1533-1539, 2021.
Article in Chinese | WPRIM | ID: wpr-922291

ABSTRACT

OBJECTIVE@#To investigate the relationship between JAK2 gene mutation and clinical indicators in patients with myeloproliferative neoplasms (MPN).@*METHODS@#122 MPN patients in the Department of Hematology, Xiyuan Hospital, China Academy of Chinese Medical Sciences from September 2017 to January 2020 were retrospectively analyzed. The relationship between JAK2 gene mutation and sex, age, peripheral blood cell count, splenomegaly, and thrombosis and bleeding events were analyzed.@*RESULTS@#In 122 patients with MPN, the patients with polycythemia vera (PV) accounted for 36 (29.5%), the patients with essential thrombocythemia (ET) accounted for 56 (45.9%), the patients with myelofibrosis (MF) accounted for 30 (24.6%). The JAK2 gene mutation rate in MPN patients was 64.6% (79/122), and the JAK2 gene mutation rate in PV, ET and MF groups were 77.7% (28/36), 60.7% (34/56) and 56.7% (17/30), the JAK2 gene mutation rate of the patients in PV group was statistically significant as compared with those in the ET group (P<0.05). The hemoglobin (Hb) count of the patients in JAK2 gene mutation group was higher than those in wild-type group [(150.0±39.6)g/L vs (129.4±38.9)g/L, P<0.05]; the white blood cell (WBC) count of the patients in JAK2 gene mutation group was higher than those in the wild type group [(9.5±4.7)×10@*CONCLUSION@#The mutation rate of JAK2 gene in MPN patients is higher, and the mutation rate of JAK2 gene in PV patients is higher than that in ET and MF patients; JAK2 gene mutations in MPN patients are related to hemogram index; the incidence of splenomegaly is the highest in MF patients, and splenomegaly is related to the occurrence of JAK2 gene mutations in MF patients.


Subject(s)
Humans , Janus Kinase 2/genetics , Mutation Rate , Myeloproliferative Disorders/genetics , Polycythemia Vera , Retrospective Studies
5.
Journal of Experimental Hematology ; (6): 1236-1241, 2021.
Article in Chinese | WPRIM | ID: wpr-888544

ABSTRACT

OBJECTIVE@#To analyze the comprehensive laboratory test data of BCR-ABL1 fusion gene and JAK2 V617F mutation co-expressed in myeloproliferative neoplasm (MPN) patients, and investigate its relative clinical significance.@*METHODS@#Data of 1 332 MPN patients were comprehensively analyzed, BCR-ABL1 (P190/P210/P230) fusion gene and JAK2 V617F mutation were detected by real time-polymerase chain reaction (RT-PCR) technique, the CALR, MPL, JAK2 12 and 13 exon mutations were detected by the First Generation Sequencing, the bone marrow cell morphology and pathological characteristics were evaluated by bone marrow smear and biopsy technique, the immune phenotypes of bone marrow cells were evaluated by flow cytometry, the chromosome karyotypes of bone marrow cells were analyzed by chromosome G banding technique.@*RESULTS@#Four of the 1 332 patients were found to have the co-existence of BCR-ABL1 fusion gene and the JAK2 V617F mutation, with a 0.3% incidence and a median age of 70 years old, including 2 cases of polycythemia vera, 1 case of primary myelofibrosis, and 1 case of chronic myeloid leukemia-accelerated phase. The clues of double positive genes of such patients at the time of initial diagnose could not be cued only by age, physical signs and cell morphology, they should be analyzed by comprehensive test data.@*CONCLUSION@#The co-existence of BCR-ABL1 fusion gene and JAK2 V617F mutation in the same case is a kind of disease with special clinical significance. The application of multiple detection methods can improve the detection of this disease, which is conducive to early detection, reasonable diagnosis and treatment by clinicians.


Subject(s)
Aged , Fusion Proteins, bcr-abl/genetics , Humans , Janus Kinase 2/genetics , Laboratories , Mutation , Myeloproliferative Disorders/genetics , Polycythemia Vera
6.
Acta méd. colomb ; 45(4): 41-47, Oct.-Dec. 2020. tab, graf
Article in English | LILACS, COLNAL | ID: biblio-1278140

ABSTRACT

Abstract Background and objective: in the global scientific literature, the frequency of JAK2 is highly heterogenous in chronic myeloproliferative neoplasms. The objective of this study was to analyze the prevalence of the JAK2 mutation in primary myelofibrosis (PMF) and compare it according to the detection method used, from 2007-2018. Materials and methods: a systematic review with meta-analysis, using 21 searches in three multidisciplinary databases. The PRISMA guideline phases of identification, screening, selection and inclusion were applied. Reproducibility and evaluation of the methodological quality were ensured. The analyses were based on frequencies and meta-analysis for the prevalence of the mutation with its 95% confidence interval. Results: twenty-nine studies with 744 patients were included, mainly from Korea, Brazil and China. The most commonly used technique was AS-PCR, and the prevalence of JAK2 with this technique ranged from 33.3 to 71.4%; with real-time PCR ranging from 42.9 to 77.3%, sequencing from 14.3-57.4%, and ARMS from 36.4-83.3%. The prevalence of JAK2 showed no statistically significant differences according to the type of diagnostic test used. Conclusion: high frequencies of the JAK2V617F mutation are seen in PMF, which shows that this entity should not be diagnosed solely based on clinical and hematological characteristics, but also on the patients' genetic screening.


Resumen Antecedentes y objetivo: en la literatura científica mundial la frecuencia de JAK2 presenta una alta heterogeneidad en las neoplasias mieloproliferativas crónicas. El objetivo de esta investigación fue analizar la prevalencia de la mutación JAK2 en mielofibrosis Primaria (MFP) y compararla según la técnica de detección en los años 2007-2018. Material y métodos: revisión sistemática con metaanálisis, usando 21 búsquedas en tres bases de datos multidisciplinarias. Se aplicaron las fases de identificación, tamización, elección e inclusión de la guía PRISMA. Se garantizó reproducibilidad y evaluación de la calidad metodológica. Los análisis se basaron en frecuencias y metaanálisis para la prevalencia de la mutación con su intervalo de confianza de 95%. Resultados: se incluyeron 29 estudios con 744 pacientes, los cuales provienen principalmente de Corea, Brasil y China. La técnica más empleada fue AS-PCR, las prevalencias de JAK2 con esta técnica oscilaron entre 33.3 y 71.4%; con PCR en tiempo real entre 42.9 y 77.3%, con secuenciación de 14.3-57.4% y en ARMS de 36.4-83.3%. La prevalencia de JAK2 no presentó diferencias estadísticamente significativas según el tipo de prueba diagnóstica utilizada. Conclusión: se evidencian altas frecuencias de la mutación JAK2V617F en MFP, lo que evidencia que el diagnóstico de esta entidad no debe realizarse únicamente por características clínicas y hematológicas sino también por la tamización genética de los pacientes.


Subject(s)
Humans , Male , Female , Meta-Analysis , Patients , Genetic Testing , Prevalence , Janus Kinase 2 , Primary Myelofibrosis , Mutation
7.
Electron. j. biotechnol ; 45: 46-52, May 15, 2020. tab, graf, ilus
Article in English | LILACS | ID: biblio-1177424

ABSTRACT

BACKGROUND: The present study analyzed the synergistic protective effect of ß-alanine and taurine against myocardial ischemia/reperfusion. Myocardial infarct size, lipid peroxidation, and levels of glutathione peroxidase (Gpx), superoxide dismutase (SOD), reduced glutathione (GSH), catalase, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), reactive oxygen species (ROS), apoptosis, and the mRNA and protein expression of Janus kinase 2 (JAK2) and signal transducer and activator 3 of transcription (STAT3) were determined. The molecular docking was carried out by using AutoDock 4.2.1. RESULTS: Combined treatment with ß-alanine and taurine reduced myocardial infarct size, lipid peroxidation, inflammatory marker, ROS levels, and apoptosis and increased Gpx, SOD activity, GSH, and catalase activity. Furthermore, combined treatment significantly reduced JAK2 and STAT3 mRNA and protein expression compared with the control. The small molecule was docked over the SH2 domain of a STAT3, and binding mode was determined to investigate the inhibitory potential of ß-alanine and taurine. ß-Alanine bound to SH2 domain with ΔG of -7.34 kcal/mol and KI of 1.91 µM. Taurine bound to SH2 domain with ΔG of -7.38 kcal/mol and KI of 1.95 µM. CONCLUSION: Taken together, these results suggest that the combined supplementation of ß-alanine and taurine should be further investigated as an effective therapeutic approach in achieving cardioprotection in myocardial ischemia/reperfusion.


Subject(s)
Animals , Male , Rats , Taurine/therapeutic use , Cardiotonic Agents/therapeutic use , Reperfusion Injury/drug therapy , beta-Alanine/therapeutic use , Myocardial Ischemia/drug therapy , Superoxide Dismutase , Immunohistochemistry , Lipid Peroxidation , Reactive Oxygen Species , Rats, Wistar , Reverse Transcriptase Polymerase Chain Reaction , Disease Models, Animal , Janus Kinase 2 , Molecular Docking Simulation , Glutathione Peroxidase , Heart Diseases/drug therapy , Inflammation
8.
Rev. bras. anal. clin ; 52(1): 27-31, 20200330.
Article in Portuguese | LILACS | ID: biblio-1104127

ABSTRACT

Trombose essencial é uma das doenças mieloproliferativas crônicas, rara e de etiologia ainda desconhecida, mas que apresenta risco alto de eventos trombóticos e/ou hemorrágicos, uma vez que acomete as células megacariocíticas e, consequentemente, as plaquetas. O objetivo deste trabalho foi realizar uma revisão das publicações sobre o tema abordado. O estudo caracteriza-se como revisão bibliográfica de artigos das bases de dados da Literatura Latino-Americana e do Caribe em Ciências da Saúde (Lilacs), National Library of Medicine (PubMed), Scientific Electronic Library Online (SciELO) e Google acadêmico. Foram incluídos artigos disponíveis e em qualquer idioma de publicação, com a finalidade de aumentar o referencial teórico. Posteriormente à seleção e análise dos artigos, foram encontrados alguns pontos em comum, como a dificuldade em se diagnosticar a doença. De acordo com os estudos, a maioria dos doentes é assintomática, mas pode apresentar desde eventos trombóticos até mesmo embolia pulmonar. Atualmente, o tema vem crescendo, principalmente abordando técnicas moleculares mais específicas para a descoberta da doença em seu estágio inicial. A análise dos artigos demonstrou a dificuldade do diagnóstico da trombose essencial, sendo a sua identificação crucial nos estágios iniciais.


Essential thrombosis is one of the chronic myeloproliferative diseases, rare and of unknown etiology, but which presents a high risk of thrombotic and / or hemorrhagic events. Once it attacks the megakaryocytic cells and consequently the platelets. The objective of this work is to review the publications on the subject. The study is characterized as a bibliographical review of articles from the Latin American and Caribbean Literature in Health Sciences (LILACS), National Library of Medicine (PubMed), Scientific Electronic Library Online (SciELO) and Google Acadêmico. Articles were included and in any language of publication, in order to increase the theoretical reference. Subsequent to the selection and analysis of the articles, some common points were found such as the difficulty in diagnosing the disease. According to the studies, the majority of patients are asymptomatic but may present from thrombotic events to even pulmonary embolism. Currently, the topic has been growing, mainly addressing molecular techniques more specific to the discovery of the disease in its initial stage. The analysis of the articles demonstrated the difficulty of diagnosing essential thrombosis, which is crucial in the initial stages.


Subject(s)
Thrombocytosis , Janus Kinase 2 , Thrombocythemia, Essential
9.
Journal of Experimental Hematology ; (6): 1998-2003, 2020.
Article in Chinese | WPRIM | ID: wpr-880005

ABSTRACT

OBJECTIVE@#To evaluate the proformance of multiplex PCR and capillary electrophoresis(MPCE) in the detection of JAK2V617F and CALR mutation in myeloproliferative neoplasms(MPN).@*METHODS@#The specificity primers of JAK2617F gene mutation and the primers of CALR gene were designed at the same time. The JAK2V617F and CALR gene primers were labeled with Cy5 fluorescence, all the primers were mixed in one tube for multiplex PCR and the PCR prodcuts were analysised by capillary electrophoresis. Then detection limit and sensitivity of MPCE were evaluated, and compared with comercial diagnostic kit.@*RESULTS@#JAK2V617F and CALR gene mutations could be detect by MPCE in one PCR test. JAK2V617F mutation could be detected at 0.01 ng genomic DNA, double positive JAK2V617F and CLAR gene mutations could be detected at 0.1 ng genomic DNA, at least 0.1% JAK2V617F positive mutation could be detected. The consistency between MPCE and commercial diagnostic gene mutation kit was 100%.@*CONCLUSION@#It is developed that a new gene mutation detection method of JAK2 V617F and CLAR gene based on MPCE in our experiment and it can be used as a new reagent for molecular diagnosis of MPN patients.


Subject(s)
Calreticulin/genetics , Electrophoresis, Capillary , Humans , Janus Kinase 2/genetics , Mutation , Myeloproliferative Disorders/genetics , Neoplasms , Patients , Polymerase Chain Reaction
10.
Journal of Experimental Hematology ; (6): 1321-1325, 2020.
Article in Chinese | WPRIM | ID: wpr-827118

ABSTRACT

OBJECTIVE@#To investigate the clinical characteristics of essential thrombocytopenia (ET) patients with positive mutations including JAK2, CALR, MPL, or negative mutations.@*METHODS@#A total of 66 newly diagnosed ET cases from January 2016 to December 2018 in Department of Hematology, Huaian No.1 People's Hospital affiliated to Nanjing Medical University were analyzed. Statistical analysis data included the patient's sex, age, symptoms, thrombosis and embolism events, spleen omegaly, platelet count (Plt), leukocyte (WBC) count, hemoglobin (Hb), fibrinogen (FIB), thrombus elastic diagram (TEG), serum potassium, blood glucose (GLU), lactate dehydrogenase (LDH), JAK2, CALR and MPL mutations, treatment options, and efficacy.@*RESULTS@#All the patients were not MPL-positive, and divided in three groups: JAK2 mutation (46 cases, 69.7%), CALR mutation (9 cases, 13.6%) and gene negative mutation (11 cases, 16.7%) group. The average age of patients in the JAK2 mutation group was 63.2 years old, and significantly higher than that in the CALR mutation group (51.8 year) and gene negative group (50.2 year) (P<0.05). Compared with the JAK2 mutation group and gene negative group, the CALR mutation group had lower WBC count (6.3×10/L vs 13.79×10/L) (P=0.003) (6.3×10/L vs 9.70×10/L) (P=0.009). Also the Hb level of patients in CALR mutation group was lower than the JAK2 mutation group (121.22 g/L vs 136.2 g/L) (P=0.036). However, there was higher tumor burden in the CALR mutation group, compared with the gene negative mutation group (300.11 U/L vs 227.4 U/L) (P=0. 033). There was no significant difference among the three groups, such as the Plt counts, serum potassium level, GLU level and FIB level (P>0.05). In addition, thrombus and embolism appeared in 30.3% (20/66) cases. 18.2% (12/66) cases were complicated with hyperkalemia, which significantly correlated with Plt counts (r=0.518). TEG was performed in 34 patients, of which 41.2% (14/34) had abnormal TEG and 55.9% (19/34) were accompanied by Plt count > 1 000 ×10/L, but there was no significant correlation between them (r=0.134). After routine clinical treatment, all the 66 cases achieved partial or complete hematological remission, but the disease usually repeated. Until now 4.5% (3/66) cases had been converted to myelofibrosis (MF) all with JAK2 mutation, but without advancing to acute myeloid leukemia.@*CONCLUSION@#ET patients with JAK2 mutation have higher incidence, moreover were in older age. However, the patients with CALR mutations display lower WBC count and Hb level, but higher tumor burden. In short, the multiple gene mutations of ET showed different clinical features closely relates with the prognosis, thus providing guidance for the clinical diagnosis and treatment.


Subject(s)
Aged , Calreticulin , Genetics , Humans , Janus Kinase 2 , Genetics , Middle Aged , Mutation , Primary Myelofibrosis , Thrombocythemia, Essential , Thrombocytopenia
11.
Journal of Experimental Hematology ; (6): 1326-1331, 2020.
Article in Chinese | WPRIM | ID: wpr-827117

ABSTRACT

OBJECTIVE@#To investigate the clinical characteristics, laboratorial and bone marrow pathological features of primary thrombocytopenia (ET) patients with different mutations of CALR, JAK2 and MPL genes.@*METHODS@#The chinical data of 120 cases of ET in Jiangsu provincial people's hospital/ The First Affiliated Hospital of Nanjing Medical University from January 2015 to December 2017 were collected and analyzed, including 76 cases with JAK2 gene mutation, 40 cases with CALR gene mutation, 2 cases with MPL gene mutations, 2 cases without gene mutation.@*RESULTS@#Among the ET patients, compared with the JAK2 gene mutation, CALR gene mutation showed statistically significant deareament of white blood cells and hemoglobin (P=0.001, P=0.01) and the male platelets in CALR group showed significant increament (P=0.04). Fourthermore, the average number of megakaryocytes and its cluster numbers in each hight power field of vision showed statistically significant decreament in CALR group as compared with JAK2 group (P=0.001, P=0.001), and thrombotic events in CALR group were signicantly lower than those in JAK2 group (7.5% vs 18.4%) (P=0.03).@*CONCLUSION@#Mutations of CALR, JAK2 have different clinical characteristics and blood pathological changes of Chinese ET patients, and their clinical significance is worth to explore.


Subject(s)
Bone Marrow , Calreticulin , Genetics , China , Humans , Janus Kinase 2 , Genetics , Male , Mutation , Receptors, Thrombopoietin , Genetics , Thrombocythemia, Essential
12.
Rev. Assoc. Med. Bras. (1992) ; 65(6): 772-774, June 2019.
Article in English | LILACS | ID: biblio-1041042

ABSTRACT

SUMMARY The essential thrombocythemia is one of the seven described forms of myeloproliferative neoplasms. It is characterized by megakaryocytic hyperplasia with consequent thrombocytosis maintained in the peripheral blood, favoring the occurrence of thrombo-hemorrhagic phenomena. We present the case of an 81-year-old woman with a history of ischemic stroke in the context of a sustained thrombocytosis, which led to a spinal study and a search for the V617F mutation in the JAK2 gene, which was positive. The patient started cytoreductive therapy with hydroxyurea with favorable current evolution.


Subject(s)
Humans , Female , Aged , Stroke/etiology , Thrombocythemia, Essential/complications , Risk Factors , Stroke/genetics , Janus Kinase 2/genetics , Thrombocythemia, Essential/genetics , Mutation
13.
Rev. Hosp. Clin. Univ. Chile ; 30(3): 207-217, 2019. tab, graf, ilus
Article in Spanish | LILACS | ID: biblio-1051370

ABSTRACT

V617F mutation in exon 14 of Janus Kinase 2 gene (jak-2) is used as a molecular marker for the diagnosis of Philadelphia negative myeloproliferative neoplasms (Phi-) such as Polycythemia Vera (PV), Essential Thrombocythemia (ET) and Primary Myelofibrosis (MFP). To detect this mutation, we used conventional polymerase chain reaction technique (PCR), a simple and inexpensive technique, however, has some drawbacks that current technology allows to solve. During the last years, more sensitive molecular techniques have been incorporated in clinical practice to support the diagnosis, prognosis and follow-up of hematological patients. For its implementation in the clinical routine should be considered technical and economic aspects, so in this work, we evaluate the Real Time PCR technique as a diagnostic method for the detection of the Jak-2-V617F mutation, using in house primers design. Our result show that the technique implemented has a concordance index of 0.87 with the conventional PCR used in the molecular diagnosis of myeloproliferative neoplasms. In addition, it has the same specificity, greater sensitivity and, shorter execution time in relation to conventional PCR. The implementation of this diagnostic method in our Hospital is technically possible and commercially convenient. (AU)


Subject(s)
Humans , Janus Kinase 2/analysis , Real-Time Polymerase Chain Reaction/methods , Myeloproliferative Disorders/diagnosis , Real-Time Polymerase Chain Reaction/trends
14.
Biol. Res ; 52: 29, 2019. tab, graf
Article in English | LILACS | ID: biblio-1011431

ABSTRACT

BACKGROUND: Acute kidney injury (AKI), which is mainly caused by sepsis, has high morbidity and mortality rates. CXCL8(3-72) K11R/G31P (G31P) can exert therapeutic effect on inflammatory diseases and malignancies. We aimed to investigate the effect and mechanism of G31P on septic AKI. METHODS: An AKI mouse model was established, and kidney injury was assessed by histological analysis. The contents of serum creatinine (SCr) and blood urea nitrogen (BUN) were measured by commercial kits, whereas neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1) were detected by enzyme-linked immunosorbent assay (ELISA) kits. The expressions of CXCL8 in serum and kidney tissues were determined using ELISA and immunohistochemical analysis, respectively. Apoptosis rate of renal tissue was detected by terminal deoxynucleotidyl transfer-mediated dUTP nick end labeling (TUNEL) analysis. The expressions of inflammatory cytokines were measured by quantitative real-time PCR and Western blot, respectively. The apoptosis-related proteins, JAK2, STAT3, NF-κB and IκB were determined by Western blot. RESULTS: G31P could reduce the levels of SCr, BUN, HGAL and KIM-1 and inhibit the renal tissue injury in AKI mice. G31P was also found to suppress the serum and nephric CXCL8 expressions and attenuated the apoptosis rate. The levels of inflammatory cytokines, pro-apoptotic proteins were decreased, while the anti-apoptotic proteins were increased by G31P in AKI mice. G31P also inhibited the activation of JAK2, STAT3 and NF-κB in AKI mice. CONCLUSION: These results suggest that G31P could protect renal function and attenuate the septic AKI. Our findings provide a potential target for the treatment of AKI.


Subject(s)
Animals , Male , Mice , NF-kappa B/metabolism , Sepsis/complications , STAT3 Transcription Factor/metabolism , Janus Kinase 2/metabolism , Acute Kidney Injury/etiology , Signal Transduction , Apoptosis , Sepsis/pathology , Disease Models, Animal , Acute Kidney Injury/metabolism , Acute Kidney Injury/pathology , Mice, Inbred C57BL
15.
Journal of Breast Cancer ; : 362-374, 2019.
Article in English | WPRIM | ID: wpr-764284

ABSTRACT

PURPOSE: The chemical structure of tubulosine has been known since the mid-1960s. However, little is known about its biological and pharmacological functions. The aim of this study was to investigate the novel functions of tubulosine in cancer treatment, specifically in breast cancer. METHODS: An Unpaired (Upd)-induced Drosophila cell line and interleukin (IL)-6-stimulated human breast cancer cell lines were used to investigate the biological and pharmacological activities of tubulosine in vitro. To investigate the activities of tubulosine, we performed molecular and cellular experiments such as Western blot and reverse transcription polymerase chain reaction analyses, immunoprecipitation and terminal deoxynucleotidyl transferase dUTP nick end labeling assays, and immunofluorescence staining using breast cancer cell lines. RESULTS: Tubulosine exhibited anticancer activity in IL-6-stimulated human breast cancer cells. Moreover, tubulosine reduced the tyrosine phosphorylation level and transcriptional activity of signal transducer and activator of transcription (STAT) protein at 92E in Upd-induced Drosophila cells. Additionally, tubulosine suppressed IL-6-induced Janus kinase 2 (JAK2)/STAT3 signaling, resulting in decreased viability and induction of apoptotic cell death in breast cancer cells. Interestingly, inhibition of IL-6-induced JAK2/STAT3 signaling by tubulosine was associated with the blocking of IL-6 receptor (IL-6R) and glycoprotein 130 (gp130) binding. CONCLUSION: Tubulosine exhibits anticancer activity through functional inhibition of IL-6-induced JAK2/STAT3 signaling by targeting IL-6Rα/gp130 binding in breast cancer cells. These findings suggest that tubulosine may hold promise for the treatment of inflammation-associated cancers, including breast cancer.


Subject(s)
Blotting, Western , Breast Neoplasms , Cell Death , Cell Line , DNA Nucleotidylexotransferase , Drosophila , Fluorescent Antibody Technique , Glycoproteins , Humans , Immunoprecipitation , In Vitro Techniques , Interleukin-6 , Interleukins , Janus Kinase 2 , Phosphorylation , Phosphotransferases , Polymerase Chain Reaction , Receptors, Interleukin-6 , Reverse Transcription , STAT3 Transcription Factor , Transducers , Tyrosine
16.
Article in Chinese | WPRIM | ID: wpr-771926

ABSTRACT

OBJECTIVE@#To explore the symptomatic burden of patients with essential thrombocythemia (ET) and its relation with clinical characteristics including the mutation status, therapeutic protocols and sex.@*METHODS@#Total of 173 Chinese ET patients were selected and grouped on the basis of disease characteristics (mutation status, therapeutic pro to- cols, and sex).@*RESULTS@#All the groups showed low-to-high symptom burden, with the highest in the Hu (hydroxyurea)-group (total symptom score [TSS], 14.7; range, 7.6-14.7). In the JAK2V617F-positive, Hu-treated, and female groups TSS and independent symptom scores were higher than those in the control group. The CALR-positive and IFN-α-treated groups had lower overall and individual scores as compared with groups lacking the corresponding characteristics. As the number of characteristics (JAK2V617F-positive, Hu-treated, and female) increases, the severity of symptoms gradually increased.@*CONCLUSION@#The different characteristics have various effects on symptom burden in ET patients. The accumulation of certain characteristics will lead to more severe symptom burden, thus the patient's symptom burden should be considered comprehensively when making up the treatment schemes and prognosis.


Subject(s)
Asian Continental Ancestry Group , Calreticulin , Female , Humans , Hydroxyurea , Janus Kinase 2 , Mutation , Thrombocythemia, Essential
17.
Acta Physiologica Sinica ; (6): 698-704, 2019.
Article in Chinese | WPRIM | ID: wpr-777141

ABSTRACT

The aim of this study was to investigate the relationship between the effects of different doses of X-rays on DNA damage and JAK/STAT signaling pathway activation in A549 cells. The A549 cells were radiated with X-rays at doses of 2, 4, and 8 Gy. The proliferation of A549 cells was detected by CCK8 method. The content of interleukin 6 (IL-6) in culture medium at different time points after irradiation was detected by enzyme-linked immunoassay, and the expression levels of IL-6 receptor (IL-6R) and p53 binding protein 1 (53BP1) were detected by immunofluorescent staining. The expression levels of JAK2, p-JAK2, STAT3 and p-STAT3 were detected by Western blot. The results showed that, compared with the control group, X-ray irradiation reduced the cellular proliferation, up-regulated the expression of 53BP1, increased the IL-6 content in the medium supernatant, and up-regulated the protein expression levels of IL-6R, JAK2, p-JAK2, STAT3, and p-STAT3. The above effects of X-ray irradiation were dose-dependent. These results suggest that the mechanism by which X-rays cause DNA damage in A549 cells may involve activation of the JAK/STAT signaling pathway.


Subject(s)
A549 Cells , DNA Damage , Radiation Effects , Humans , Janus Kinase 2 , Metabolism , Receptors, Interleukin-6 , Metabolism , STAT3 Transcription Factor , Metabolism , Signal Transduction , Tumor Suppressor p53-Binding Protein 1 , Metabolism , X-Rays
18.
Article in English | WPRIM | ID: wpr-776901

ABSTRACT

Macrophages show significant heterogeneity in function and phenotype, which could shift into different populations of cells in response to exposure to various micro-environmental signals. These changes, also termed as macrophage polarization, of which play an important role in the pathogenesis of many diseases. Numerous studies have proved that Hesperidin (HDN), a traditional Chinese medicine, extracted from fruit peels of the genus citrus, play key roles in anti-inflammation, anti-tumor, anti-oxidant and so on. However, the role of HDN in macrophage polarization has never been reported. Additional, because of its poor water solubility and bioavailability. Our laboratory had synthesized many hesperidin derivatives. Among them, hesperidin derivatives-12 (HDND-12) has better water solubility and bioavailability. So, we evaluated the role of HDND-12 in macrophage polarization in the present study. The results showed that the expression of Arginase-1 (Arg-1), interleukin-10 (IL-10), transforming growth factor β (TGF-β) were up-regulated by HDND-12, whereas the expression of inducible Nitric Oxide Synthase (iNOS) was down-regulated in LPS- and IFN-γ-treated (M1) RAW264.7 cells. Moreover, the expression of p-JAK2 and p-STAT3 were significantly decreased after stimulation with HDND-12 in M1-like macrophages. More importantly, when we taken AG490 (inhibitor of JAK2/STAT3 signaling), the protein levels of iNOS were significantly reduced in AG490 stimulation group compare with control in LPS, IFN-γ and HDND-12 stimulation cells. Taken together, these findings indicated that HDND-12 could prevent polarization toward M1-like macrophages, at least in part, through modulating JAK2/STAT3 pathway.


Subject(s)
Animals , Cytokines , Genetics , Metabolism , Enzyme Inhibitors , Pharmacology , Gene Expression Regulation , Hesperidin , Chemistry , Pharmacology , Inflammation , Genetics , Metabolism , Janus Kinase 2 , Metabolism , Macrophages , Allergy and Immunology , Metabolism , Medicine, Chinese Traditional , Mice , Molecular Structure , Phosphorylation , STAT3 Transcription Factor , Metabolism , Signal Transduction
19.
Article in Chinese | WPRIM | ID: wpr-776763

ABSTRACT

OBJECTIVE@#To develop a system for rapid detection of JAK2 V617F mutation among patients with myeloproliferative diseases.@*METHODS@#Specific primers and TagMan probes were designed for the mutant and wild type alleles based on the principle of real-time PCR. A complete system including the method for detection and product for quality control were established through the evaluation of sensitivity and accuracy of the method, double-blind trial, and preparation of negative and positive controls through site-directed mutagenesis and molecular cloning.@*RESULTS@#A system for rapid detection of the JAK V617F mutation has been developed. Compared with Sanger sequencing, the sensitivity and specificity of the method have both reached 100%. Meanwhile, 1000 normal samples and 1 case with the JAK2 V617F mutation were detected, which gave a population rate of 1‰.@*CONCLUSION@#The system was fast, accurate, cheap, high throughput, and easy to use. It can be utilized as a routine test. Although the JAK2 V617F mutation is rare in the population, it should be screened among myeloproliferative neoplasm patients.


Subject(s)
Alleles , DNA Mutational Analysis , Double-Blind Method , Humans , Janus Kinase 2 , Genetics , Mutation , Myeloproliferative Disorders , Genetics , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity
20.
Journal of Experimental Hematology ; (6): 1574-1579, 2019.
Article in Chinese | WPRIM | ID: wpr-775683

ABSTRACT

OBJECTIVE@#To investigate the relationship between the polymorphism of TET2 gene SNP rs3733609 and JAK2V617F allele burden in patients with myeloproliferative neoplasms (MPN).@*METHODS@#The exon 9 of TET2 gene was amplified by RT-PCR, and the nucleotide sequence of SNP rs3733609 site was analyzed by gene sequencing. The MGB Taqman probe PCR method was used to detect the JAK2V617F allele burden. The correlation of TET2 gene SNP rs3733609 C/T with the JAK2V617F allele burden and clinical parameters was analyzed.@*RESULTS@#TET2 gene rs3733609 C/T heterozygosity (normal T/T) could be detected in 19 cases of 85 cases of JAK2V617F positive MPN (22.4%) patients, while the TET2 gene rs3733609 C/T heterozygosity could be detected only in 9 of the 106 healthy volunteers, and the incidence was only 8.5% (9/106). Compared with the negative group (TET2 rs3733609 T/T), there was no significant difference in the median age, hemoglobin level and platelet count in the patients with TET2 gene SNP rs3733609 (CT/TC) positive, but the WBC count of peripheral blood and JAK2V617F allele burden significantly increased. In JAK2V617F high allele burden group, TET2 gene SNP rs3733609 was positive in 7 cases (36.8%, 7/19), the ratio was higher than that in the low allele burden group(18.2%, 12/66).@*CONCLUSION@#TET2 SNP rs3733609 C/T may be a new susceptible allelee, which affects the clinical characteristics and clonal evolution of MPN patients.


Subject(s)
Alleles , DNA-Binding Proteins , Genetics , Exons , Humans , Janus Kinase 2 , Genetics , Mutation , Myeloproliferative Disorders , Genetics , Neoplasms , Proto-Oncogene Proteins , Genetics
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