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Recently, human platelet antigens (HPAs) polymorphisms are found to play a role in susceptibility to hepatitis C virus (HCV) infection and fibrosis progression. The aim of the current study was to evaluate the possible association between the HPAs polymorphisms with liver fibrosis progression in HCV patients. HPAs polymorphisms genotyping was performed in HCV patients (n = 71) by Sequence-specific primers-polymerase chain reaction. Fibrosis progression was evaluated using the Metavir scoring system and liver biopsy, and the patients were assigned to two groups, namely, G1 (n = 35) that included patients with F1 (portal fibrosis without septa) or F2 (few septa) and G2 (n = 36) that comprised patients with F3 (numerous septa) or F4 (cirrhosis). The data analyses were performed using Pearson's χ2 test. The genotype frequency of HPA-3ab was significantly higher in G1 patients than in G2 patients (P = 0.015). No statistically significant differences were found between the patient groups (G1 and G2) regarding the distributions of the allelic and genotypic frequencies of the HPA-1, -2, -4, -5, and -15 systems. Multivariate logistic regression showed an independent association between the genotype HPA-3aa/BB and severe fibrosis (F3-F4), when compared with genotype HPA-3ab, independent of the viral genotype, high alanine transaminase, sex, age, time of infection, diabetes, and high cholesterol as risk factors. The present study suggested that the HPA-3ab genotype could be noticed as a potential protecting factor against hepatic fibrosis. Therefore, the antigenic variation of integrins might be considered as a part of the coordinated inflammatory process involved in the progression of liver fibrosis.
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Antígenos de Plaquetas Humanas/genética , Hepatite C Crônica/genética , Cirrose Hepática/virologia , Polimorfismo Genético , Adulto , Idoso , Alelos , Progressão da Doença , Feminino , Estudos de Associação Genética , Genótipo , Hepatite C Crônica/complicações , Humanos , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
INTRODUCTION: Mt DNA (DAMPs) plays a key modulatory role in immune cells and may also mediate a variety of adverse transfusion reactions. MATERIAL AND METHOD: The cross-sectional study was performed on 22 (PRP) and 14 (SDP) between February 2019 and 2020. mtDNA DAMPs by quantitative real-time PCR was assessed on days 1, 3, and 5 of platelets storage. The data was entered in REST 2009 software, and the amount of fold change was calculated. Multiple t tests were also used. RESULTS: The mtDNA DAMPs fold change in SDP on days 1-3, 1.3 times, on days 3-5, 1.5 times, and on days 1-5, 2.1 times increased. The fold change on days 1-3, 0.8 times, on days 3-5, 0.6 times, and on days 1-5, 0.49 times decreased in PRP products. CONCLUSION: The method of preparation and processing can affect mtDNA DAMPs fold changes.
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Remoção de Componentes Sanguíneos , Plasma Rico em Plaquetas , Humanos , Plaquetas , DNA Mitocondrial/genética , Estudos TransversaisRESUMO
OBJECTIVE: Human platelet antigens (HPAs) are antigenic determinants on platelet membrane glycoproteins that stimulate the host's immune system and cause platelet destruction. In this study, we share our experience with implementing sequence-specific primer-polymerase chain reaction (PCR-SSP), real-time PCR, and PCR-RFLP (restriction fragment length polymorphism) and the validation process used to evaluate the results. METHODS: At the Ardabil Blood Transfusion Center, 10 samples were obtained from blood donors. Validation using PCR-SSP, real-time PCR, and PCR-RFLP methods for genotyping HPAs was done by sequencing. A commercial DNA sample and a commercial kit were also used for validation. RESULTS: The results of PCR-SSP, TaqMan Real-Time PCR, melting curve analysis (HPA-15), and PCR-RFLP (HPA-3) were 100% consistent with sequencing (gold standard) and commercial kit results. CONCLUSIONS: There was a 100% correlation between repeating the methods and the expected results for repeatability, and no false positives and negatives were observed.
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Antígenos de Plaquetas Humanas , Humanos , Antígenos de Plaquetas Humanas/genética , Antígenos de Plaquetas Humanas/análise , Polimorfismo de Fragmento de Restrição , Reação em Cadeia da Polimerase em Tempo Real , Doadores de Sangue , GenótipoRESUMO
INTRODUCTION: The aim of this study was to evaluate and compare two different apheresis and changes in some immunological factors in donors. MATERIAL AND METHODS: The cross-sectional study was performed from January 2017 to September 2018. Fifty six male blood donors were randomly divided into two groups. CD4, CD8, and CD25 markers by flow cytometry, and TGFBeta by real-time polymerase chain reaction (RT-PCR) method were done before and 7 days after the apheresis procedure. Independent Sample t-test, Mann-Whitney U Test, Wilcoxon signed ranked test, and Fisher exact test were used. RESULTS: WBC in MCS+ group after donation is significantly higher than before donation (P < 0.05) but no significant difference was seen between MCS+ and Trima groups in these two indicators. But in CD4, CD25, and TGFBeta, there was no significant difference between the two groups. CONCLUSION: There was no significant difference on CD4, CD25, and TGFBeta gene 7 days after donation.
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Doadores de Sangue , Plaquetoferese , Estudos Transversais , Humanos , Fatores Imunológicos , Masculino , Plaquetoferese/métodos , Fator de Crescimento Transformador betaRESUMO
Background: An analysis of red blood cell alloimmunization in patients with thalassemia can help to devise specific strategies to decrease the alloimmunization rate. This study explored the frequency and specificity of alloantibodies and autoantibodies against red blood cell (RBC) antigens in patients with thalassemia referring to the Iranian Blood Transfusion Organization (IBTO) Immunohematology Reference Laboratory (IRL) in Tehran. Materials and Methods: This study first examined the laboratory records of 23,113 patients suffering from different diseases referring to IBTO's IRL for pretransfusion testing in the 2008-2015 period. ABO and Rh(D) typing and antibody screening tests were performed for all 23,113 patient records and 685 (2.97%) beta-thalassemia patients with positive pre-transfusion test results (antibody screening and/or DAT) were selected for further investigation. Results: The antibody screening test was positive in 640 out of 685 thalassemic patients (93.4%). DAT was performed for 529 patients, 226 (33%) of which showed positive results. Meanwhile, 161 out of 685 beta-thalassemia patients (23.5%) had positive auto control test results, reflecting the possible presence of allo- and/or autoantibodies. The most common antigen-specific alloantibodies were directed against K and E RBC antigens with a frequency of 25% (Anti-K) and 11.91% (Anti-E), respectively. The development of two antibodies (double antibodies) in one patient was observed in 80 individuals (11.46%). Conclusion: Age, gender, history of pregnancy, and splenectomy were not contributing factors to the antibody presence in the patient population under study. Extended red blood cell phenotyping should be considered as an essential procedure for expected multi-transfused thalassemia patients before blood transfusion. Considering the high frequency of anti-K and anti-E observed in this study, it is recommended that thalassemia patients in Iran are tested through phenotyping of RBC units for K and E antigens before transfusion.
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Human platelet antigens (HPAs) are glycoproteins on the platelet surface that a single nucleotide mutation in the coding region gene could lead to the variation of different HPA polymorphisms. These antigens have shown variation among different races and may trigger immune responses during blood transfusion and pregnancy. Genotyping of HPAs is useful for managing these reactions and establishing a platelet registry to decrease platelet transfusion reactions. This study aimed to compare allelic and genotype frequencies of human platelet antigens in the Azeri ethnicity by TaqMan Real-time and polymerase chain reaction with sequence-specific primers (PCR-SSP) methods. DNA was extracted from the whole blood of 100 Azeri blood donors in the Ardabil Blood Transfusion Center. Genotyping of HPA-1 to -5 and -15 was performed by TaqMan Real-time PCR, and PCR-SSP and consistency of results were evaluated. The results of PCR-SSP and TaqMan Real-time PCR showed complete consistency. The allele frequencies were 91.5% and 8.5% for HPA-1a and -1b; 88% and 12% for HPA-2a and -2b; 58% and 42 % for HPA-3a and -3b; 100% for HPA-4a; 91% and 9% for HPA-5a and -5b; 56.5% and 43.5% for HPA-15a and -15b alleles. Not incompatibility was detected in HPAs genotyping by PCR-SSP and TaqMan Real-time PCR so that real-time PCR can be used as a robust and quick method for HPA genotyping. We found differences between Azeri blood donors and previously reported HPAs alleles' frequency in other ethnicities in the country. This fact highlights the need for a platelet registry to recruit platelet donors from different ethnicities and increase the number of donors by using faster methods.
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Antígenos de Plaquetas Humanas/genética , Doadores de Sangue , Tipagem e Reações Cruzadas Sanguíneas , Plaquetas/imunologia , Primers do DNA , Reação em Cadeia da Polimerase em Tempo Real , Transfusão de Sangue , Frequência do Gene , Genótipo , Histocompatibilidade , Humanos , Irã (Geográfico) , Valor Preditivo dos TestesRESUMO
BACKGROUND: Thalassemia is hereditary anemia with lifelong transfusion as treatment and hepatitis B virus (HBV) infection is one of the transfusion transmitted infections (TTI). HBV vaccination is obligatory for these patients by 3 double-dose injections. The authors studied the HBV status and immune response to vaccination by hepatitis B surface antibody (HBsAb) titration in their thalassemic patients. They also compared these results with their previous study to find out the effectiveness of a booster dose in the immunity of patients against HBV. MATERIALS AND METHODS: Hepatitis B surface antigen (HBsAg), HBsAb, and hepatitis B core antibody (HBcAb) were detected in sera of 416 patients at the Tehran Adult Thalassemia Clinic. The immune status was classified into 4 categories: (1) immune to HBV via the vaccination (positive vaccinal)--if HBs Ag: negative, HBsAb: positive, HBcAb: negative; (2) immune to HBV via the natural disease (past infection)--if HBs: negative, HBsAb and HBcAb: both positive; (3) nonimmune to HBV (negative)--if all three parameters were negative; (4) carrier of HBV (carrier state)--if HBs Ag was positive and HBsAb and HBc Ab: both negative. Also grading of immunity done by HBsAb titration as positive if HBsAb titer was more than 100 IU/mL, negative if HBsAb titer was less than 10 IU/mL, and weakly positive if antibody level was 10-100 IU/mL. RESULTS: There were 416 patients: 302 (72.5%) with thalassemia major (TM), 104 (25%) thalassemia intermedia (TI), 7 (1.6%) sickle thalassemia (ST), and 3(0.7%) alpha-thalassemia (HbH disease). The mean age was 25.6 +/- 8.3 yr and median age was 24 yr; there were 247 (59.4%) males and 169 (40.6%) females. A total of 257 patients (61.7%) were splenectomized. According to our classification 289 (69.4%) were immunized by vaccination; 80 (19.2%) were immunized by past infection; 44 (10.5%) were negative, and 3 (0.7%) were in carrier state of HBV. In grading of immunity to HBV vaccination, 319 (76.6%) patients had HBsAb > 100 IU/mL (positive), 77 (18.5%) between 10 and 100 IU/mL (weakly positive), and 20 (4.8%) less than 10 IU/mL (negative). There was no significant correlation between the level of HBsAb and splenectomy or type of thalassemia. CONCLUSION: Response rate to vaccination is more than 95% after complete course (3 doses) in healthy individuals but failure to fulfill vaccination seems a problem in chronic transfused patients. These results reflect advantages of a booster dose of vaccine, which increased the protection level among these high-risk patients from 46.9% (in the authors' previous data) up to 69.4% in this study.
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Anticorpos Anti-Hepatite B/sangue , Antígenos de Superfície da Hepatite B/imunologia , Vacinas contra Hepatite B/imunologia , Hepatite B/prevenção & controle , Talassemia beta/imunologia , Adolescente , Adulto , Criança , Estudos Transversais , Feminino , Hepatite B/sangue , Hepatite B/transmissão , Vacinas contra Hepatite B/administração & dosagem , Humanos , Imunização Secundária , Irã (Geográfico) , Masculino , Pessoa de Meia-Idade , Vacinação , Adulto Jovem , Talassemia beta/complicações , Talassemia beta/terapiaRESUMO
BACKGROUND: Gamma-irradiation of platelet concentrates may inactive contaminated lymphocytes and subsequently inhibit the synthesis of cytokine in platelet during storage. The aim of this study was to determine the effect of gamma-irradiation on the production of IL-8 and lymphocyte proliferation isolated from random donor platelet concentrates during 3-day storage. METHODS: In this study, we evaluated the effect of gamma-irradiation on both lymphocyte proliferative response and IL-8 production for 29 random donor platelet concentrates (RD-PCs). All PCs were prepared from single RD by platelet-rich plasma (PRP) method were divided in two groups: 1) non-irradiated RD-PCs (n = 13) and 2) irradiated RD-PCs (n = 16). The RD-PCs were treated by gamma-irradiation (30 Gy) on days 0 and 3. RESULTS: IL-8 increased in both groups after 3 days. It showed increase in concentration of IL-8 in irradiated and non-irradiated PCs during storage. Results of lymphocyte transformation test (LTT) indicated that the proliferation activity of lymphocyte was inhibited by gamma radiation. CONCLUSIONS: These data indicate that gamma irradiation inhibits proliferation of lymphocyte but does not inhibit production and accumulation of IL-8 during the storage of PCs.
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Plaquetas/citologia , Proliferação de Células/efeitos da radiação , Raios gama , Interleucina-8/biossíntese , Linfócitos/efeitos da radiação , Humanos , Linfócitos/citologia , Linfócitos/metabolismoRESUMO
OBJECTIVES: To determine if parthenolide (PTL) is cytotoxic for leukemia-like KG1a cells and if it involves in certain molecular-mediated resistance, especially osteopontin (OPN). METHODS: PTL/daunorubicin (DNR)-treated KG1a cells were examined for viability using MTT and colony-formation assay, and stained for apoptosis using AV/PI. The gene and protein expression were evaluated by qReal-time PCR and Western blotting analysis, respectively. OPN gene was inhibited by OPN siRNA. The cells were stained for various fractions using PE anti-CD34, FITC anti-CD38 and PerCP anti-CD123. RESULTS: Cell viability and proliferation assay exhibited KG1a cells are relatively refractory to used concentrations of PTL. OPN mRNA and protein levels increased in response to PTL. Suppression of OPN with siRNA increased the cytotoxic effects of PTL on KG1a cells. PTL treatment and OPN siRNA suppression in KG1a cells resulted in a decrease of mRNA expression of AKT, mTOR, ß-catenin, and Phosphatase and tensin homolog (PTEN). The sub-population cells of CD34+ and CD123+ from KG1a cells are enriched by PTL treatment. CONCLUSION: Parthenolide in spite of the reduction in gene expression of AKT, mTOR or beta-catenin, stimulates the OPN expression in KG1a cells. The OPN expression pattern in KG1a cells could be compatible with CD34+/CD123+ subtype enrichment by PTL which in turn implies OPN's unique role in resistance of cell populations characterized by CD34+/CD123+ phenotype.
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Antineoplásicos Fitogênicos/farmacologia , Leucemia Mieloide Aguda/tratamento farmacológico , Células-Tronco Neoplásicas/metabolismo , Osteopontina/metabolismo , Sesquiterpenos/farmacologia , Antígenos CD34/metabolismo , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Subunidade alfa de Receptor de Interleucina-3/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Osteopontina/genética , RNA Interferente Pequeno/administração & dosagem , Reação em Cadeia da Polimerase em Tempo RealRESUMO
BACKGROUND: Graft-versus-host disease is one of the major complications after allogeneic bone marrow transplantation, but it is not easy to anticipate the onset. Cytokines released by type 1 T helper cells are thought to play a pivotal role in acute graft-versus-host disease aGVHD. The ability to predict the likely occurrence of graft-versus-host-disease (GVHD) after Hematopoietic Stem cell Transplantation (HSCT) would be extremely valuable. By serially measuring serum levels of soluble IL-2 receptor (sIL-2R), IL-18 and following allogeneic HSCT we tried to define their effect on aGVHD as a complication of transplantation and determine useful markers for aGVHD predictors. SAMPLES AND METHODS: Serum sIL-2R, IL-18, levels were measured by sandwich ELISA in 219 sera samples from 39 patients (with hematological disorders before and after allogeneic HSCT) and 28 controls. All patients received transplants from HLA-identical siblings. RESULTS: 23 (58.9%) patients developed aGVHD (I-IV) and serum levels of sIL-2R and IL-18, in sera drawn before transplantation, in patients with acute graft-versus-host disease (aGVHD(+)), were increased in comparison to patients without acute graft-versus-host disease (aGVHD(-)) and to a control group and there were no significant differences in serum levels of sIL-2R and IL-18 in aGVHD(-) patients and controls. Serum level of IL-18, in aGVHD(+) patients, was increased during days 3-24 after HSCT, and there was a significant difference according to GVHD severity. In majority of patients with acute GVHD (60%), the peak levels of IL-18 and sIL-2R were achieved on day 10 after HSCT and the rise in sIL-2R and IL-18 preceded the clinical signs of GVHD (mean day 15 after BMT). The level of IL-18 in patients with aGVHD strongly correlated with the severity of aGVHD on Day 10 after HSCT. IL-18 level (before HSCT), in patients who received Busulfan and Fludarabin which were used to treat aGVHD, was lower than in patients who received Busulfan and Cyclophosphamide. CONCLUSION: Our data concluded that IL-18 plays an important role in the development of aGVHD and the IL-18 level might be an indicator of aGVHD, reflecting the severity of the disease. These findings suggest that IL-18 may play an important role in the pathogenesis of aGVHD and that measurement of serum IL-18 levels can be a useful indicator of aGVHD.
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Doença Enxerto-Hospedeiro/sangue , Transplante de Células-Tronco Hematopoéticas , Interleucina-18/sangue , Receptores de Interleucina-2/sangue , Doença Aguda , Adolescente , Adulto , Biomarcadores/sangue , Bussulfano/administração & dosagem , Criança , Pré-Escolar , Ciclofosfamida/administração & dosagem , Feminino , Doença Enxerto-Hospedeiro/tratamento farmacológico , Doença Enxerto-Hospedeiro/etiologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Imunossupressores/administração & dosagem , Irã (Geográfico) , Masculino , Pessoa de Meia-Idade , Fatores de TempoRESUMO
BACKGROUND: Platelet transfusion is accompanied by febrile nonhemolytic transfusion reactions. The generation of cytokines (like IL-1 beta, IL-6, IL-8, and TNF-alpha) in platelet concentrates by white cells is suggested to be responsible for febrile nonhemolytic transfusion reactions. The number of white cells in the platelet concentrates is crucial to cytokine generation. METHODS: This study was performed to determine whether WBC reduction in platelet concentrates by prestorage leukodepletion filters or inactivation by gamma radiation reduced the levels of these cytokines during storage for 3 days. Each of the platelet concentrates (n = 54) was prepared from a single random donor by platelet-rich plasma. This was then divided into four groups: 1) unfiltered, nonirradiated random-donner platelet concentrates (n = 13); 2) unfiltered, gamma-irradiated random-donner platelet concentrates (n = 16); 3) filtered, nonirradiated random-donner platelet concentrates (n = 14); and 4) filtered, gamma-irradiated random-donner platelet concentrates (n = 11). Cytokine levels in platelet concentrates supernatants were measured by ELISA kits according to the manufacturer's recommendations. RESULTS: Our results showed that IL-8 was detected in unfiltered, nonirradiated, and gamma-irradiated random-donner platelet concentrates but not in the filtered random-donner platelet concentrates. TNF-alpha was only detected in unfiltered, nonirradiated units. Compared with unfiltered platelet concentrates, prestorage filtration prevented a rise in the IL-8 and TNF-alpha on day 3 of storage. The concentration of IL-1 beta was lower than the minimum concentration value of the kit used for this purpose. IL-6 was detected only in 7 units of all filtered platelet concentrates on day 3. CONCLUSION: These data indicate that gamma irradiation can not prevent generation of IL-8 in platelet concentrates during storage, but prestorage leukoreduction of platelet concentrates can prevent accumulation of IL-6, IL-8, and TNF-alpha during storage.
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Plaquetas/metabolismo , Plaquetas/efeitos da radiação , Preservação de Sangue , Citocinas/biossíntese , Raios gama , Interleucina-8/análise , Fator de Necrose Tumoral alfa/análise , Plaquetas/química , Citocinas/efeitos adversos , Citocinas/análise , Ensaio de Imunoadsorção Enzimática , Filtração/métodos , Humanos , Interleucina-1/análise , Interleucina-6/análise , Contagem de Leucócitos , Transfusão de Plaquetas/efeitos adversos , Fatores de Tempo , Fator de Necrose Tumoral alfa/efeitos adversosRESUMO
The aim of this study was to evaluate mixed red cells population and red blood cell chimerism after hematopoietic stem cell transplantation. Red blood cell chimerism after hematopoietic stem cell transplantation was analyzed using a series of fluorescein isothiocyanate-conjugated monoclonal antibodies (BioAtlantic, France) directed against ABH, Rh (D, C, E, c, e), Kell, Duffy, Kidd, and Ss antigens on blood samples of 14 patients with hematologic disorders undergoing hematopoietic stem cell transplantation, by flow cytometric method on days 15, 30, and 60 after transplantation. All patients showed expression of donor red cell antigens within days 15 - 30 after hematopoietic stem cell transplantation. Graft versus host disease and ABO incompatibility did not affect the expression of chimerism. Flow cytometric analysis is a simple, accurate, and valuable test which is of significant help in monitoring chimerism in allogeneic hematopoietic stem cell transplantation.
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Transplante de Medula Óssea/métodos , Eritrócitos/citologia , Citometria de Fluxo/métodos , Sistema ABO de Grupos Sanguíneos , Adolescente , Adulto , Anticorpos Monoclonais/química , Antineoplásicos/farmacologia , Criança , Feminino , Fluoresceína-5-Isotiocianato/farmacologia , Doença Enxerto-Hospedeiro/terapia , Transplante de Células-Tronco Hematopoéticas , Humanos , Irã (Geográfico) , Masculino , Vidarabina/análogos & derivados , Vidarabina/farmacologiaRESUMO
BACKGROUND: This study was designed to evaluate platelet transfusion outcome via flow cytometric monocyte phagocytic assay (FMPA). METHODS: Fifteen patients with a history of multiple platelet transfusions and fifteen controls were enrolled in this study. CMFDA-labeled platelets were incubated with patients' sera and were finally incubated with monocytes in a tube and analyzed by flow cytometry. Monocytes that phagocytosed platelets were detected as a CMFDA-positive platelet population via monocyte gate. The FMPA results were compared with CCI results for the patients. RESULTS: The FMPA result correlated with 1-hour (r = -0.885, P = 0.001) and 24-hour (r = -0.884, P = 0.001) CCI. There is a significant difference in means of FMPA results between the patients with immune platelet refractoriness (68.46 ± 10.4%), non-refractory group (37.73 ± 15.21%) and the control group (18.27 ± 2.86%). CONCLUSION: Our data showed that FMPA has good results in evaluation of platelet transfusion outcome and may be useful as an indicator of platelet transfusion response.
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Plaquetas/citologia , Citometria de Fluxo/métodos , Leucemia Mieloide Aguda/complicações , Monócitos/citologia , Transfusão de Plaquetas , Trombocitopenia/terapia , Adulto , Idoso , Estudos de Casos e Controles , Feminino , Fluoresceínas , Corantes Fluorescentes , Humanos , Irã (Geográfico) , Masculino , Pessoa de Meia-Idade , Resultado do TratamentoRESUMO
BACKGROUND: Interfering with cell proliferation and survival is a critical role for antineoplastic drugs leading to cell death through induction of apoptosis. Alternative treatments with herbal extracts offer insights into acute myeloid leukemia (AML) therapy. Parthenolide (PTL), an extract from feverfew, induces apoptosis in primary human leukemia stem cells (LSCs) and bulk leukemic cell populations. Osteopontin (OPN) preserves cell viability in response to anticancer agents and its receptors could be utilized for therapeutic targeting of cancer cells. METHODS: U937 cells were cultured in RPMI 1640 with concentrations of 2, 4, 6, 8, and 10 µM PTL for 20-24 hours for MTT assays. Apoptosis assays were performed with Annexin V-Alexa Fluor-488/PI as Annexin V+/PI- and Annexin V+/PI+ to measure early and late apoptosis, respectively. Quantitative real-time PCR was used to measure OPN gene expression using the 2(-ΔΔCt) method. The PTL-treated cells were stained with FITC-CD38 antibody for flow cytometry analyses. Data were compared using one-way analysis of variance (ANOVA) by SPSS 19. RESULTS: Parthenolide inhibited growth of U937 cells with IC25 and IC50 values of 4 and 5.8 µM, respectively. Death induction with PTL was apoptotic. Flow cytometry showed a significant decrease in the percentage of CD38+ U937 cells in response to PTL. Osteopontin gene expression decreased in response to PTL. CONCLUSION: PTL induced apoptosis and reduced OPN gene expression in U937 cells.
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BACKGROUND: Acute myeloid leukemia (AML) is an immunophenotypically heterogeneous malignant disease, in which CD34 positivity is associated with poor prognosis. Osteopontin (OPN) plays different roles in physiologic and pathologic conditions like: survival, metastasis and cell protection from cytotoxic and apoptotic stimuli. Due to anti-apoptotic effect of OPN in normal and malignant cells, silencing of OPN leads to elevation of sensitivity towards chemotherapeutic agents and attenuates cancer cells migration and invasion. Therefore, the aim of this study was to evaluate OPN roles in modulating curcumin-mediated growth inhibitory on leukemic stem cells (LSCs) colony forming potential and survival in AML cell lines and primary CD34+/CD38- bone marrow-derived AML cells. MATERIALS AND METHODS: Primary human CD34+/CD38- cells were isolated from bone marrow mononuclear cells of 10 AML patients at initial state of diagnosis, using a CD34 Multi sort kit. The growth inhibitory effects of curcumin (CUR) were evaluated by MTT and colony-formation assays. Apoptosis was analyzed by 7AAD assay in CD34+ KG-1, U937 cell lines and primary isolated cells. Short interfering RNA (siRNA) against OPN was used for OPN silencing in both cell lines and primary AML cells. Then, transfected cells were incubated with/without curcumin. The change in OPN gene expression was examined by Real-time PCR. RESULTS: CUR inhibited proliferation and induced apoptosis in both KG-1 and U937 cells and also primary isolated AML cells. OPN silencing by siRNA increased the susceptibility of KG-1, U937 and primary CD34+/CD38- AML cells to apoptosis. Moreover, soft agar colony assays revealed that silencing of OPN with siRNA significantly decreased colony numbers in LSCs compared with the non-targeting group. Furthermore, CD34+/CD38- populations as a main LSCs compartment through OPN overexpression towards CUR treatment might be nullified the inhibitory effects of OPN siRNA on their survival and colony forming potential. CONCLUSION: Taken together, our results suggested that knockdown of OPN using OPN specific siRNA significantly decreased colony numbers in LSCs and this effect might be vetoed by LSCs via induction of OPN overexpressionin combination of CUR and siRNA.
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AIMS: Acute myeloid leukemia (AML) initiation and progression have been attributed to subpopulations of self-renewing leukemia stem cells (LSCs), which contribute to progression, recurrence and therapeutic resistance in leukemia. Osteopontin (OPN) plays an important role in promoting survival and drug resistance in LSCs. The aim of this study was to explore OPN roles in modulating curcumin-mediated LSC enrichment and survival in AML cell lines and primary CD34+/CD38- bone-marrow-derived AML cells. MATERIALS AND METHODS: The growth inhibitory effects of curcumin (CUR) were evaluated by MTT assay in U937 and CD34+ KG-1 AML cell lines as well as primary CD34+/CD38- bone-marrow derived AML cells isolated by MACS technique. The proportion of LSC markers (CD34, CD38 and CD123) were evaluated by flow cytometry. The expression levels of OPN, AKT, mTOR, PTEN, ß-catenin and NF-κB were investigated by qRT-PCR. Short interfering RNA (siRNA) against OPN was used in AML cells incubated with or without CUR. KEY FINDINGS: Proportions of CD34+/CD38-/CD123+ and CD34+/CD38+/CD123+ LSCs compartment co-expressing an increased level of OPN could be enriched in AML cell lines and in patient's primary cells by CUR treatment. The expression levels of AKT, mTOR, PTEN, and ß-catenin and NF-κB1, were also significantly up-regulated concurrently with OPN in the enriched CD34+ AML cells. SIGNIFICANCE: The increased in CUR-mediated OPN level is involved in a complex interplay of various signaling pathways resulting in cytoprotection and enrichment of CD34+ LSC compartment in CUR-treated AML cells. AKT/mTOR/PTEN/ß-catenin/NF-kB signaling pathways may play roles in modulating OPN-mediated LSC cell survival and enrichment.
Assuntos
Leucemia Mieloide Aguda/genética , Células-Tronco Neoplásicas/metabolismo , Proteína Oncogênica v-akt/metabolismo , Osteopontina/biossíntese , PTEN Fosfo-Hidrolase/biossíntese , Serina-Treonina Quinases TOR/biossíntese , beta Catenina/biossíntese , ADP-Ribosil Ciclase 1/metabolismo , Antígenos CD34/metabolismo , Antineoplásicos/farmacologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Linhagem Celular Tumoral , Curcumina/farmacologia , Humanos , Subunidade alfa de Receptor de Interleucina-3/metabolismo , NF-kappa B/metabolismo , Proteína Oncogênica v-akt/genética , Osteopontina/genética , PTEN Fosfo-Hidrolase/genética , RNA Interferente Pequeno/farmacologia , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/genética , beta Catenina/genéticaRESUMO
OBJECTIVES: Human cytomegalovirus (CMV), a double-strand DNA herpesvirus, can be transmitted via blood transfusion which is especially important for immunocompromised recipients and can cause a fatal infection. CMV seroprevalence in Iran was studied on blood donors, healthy subjects, and some patients. Highly variable rates were detected. The purpose of this study was to review CMV seroprevalence in blood donors and apparently healthy individuals, in Iran. MATERIALS AND METHODS: One hundred and fifty-eight electronic and paper-based resources and databases including published articles in internal and external journals, seminars, dissertations, and theses available in the database and different websites were used to be systematically reviewed as a meta-analysis. Less related articles to the issue, papers of specific high risk population, and articles with not enough information, were excluded. Eventually 22 articles that satisfied our selection criteria were systematically reviewed and analyzed. To explore heterogeneity between studies the I square (I(2)) index was used. Data were analyzed using the statistical software package (STATA) 11. RESULTS: The heterogeneity between selected studies was 97% with an I(2) statistic. In this study a random effects model was used for meta-analysis. The prevalence of CMV IgG and CMV IgM antibodies in the country were estimated to be 92% (95% CI: 90-94) and 2.6% (95% CI: 1.7-3.6), respectively. CONCLUSION: Given high rate of CMV seropositivity in Iran, it seems that CMVAbs screening would not be a reasonable and affordable approach to prevent CMV infection via transfusion especially for immune compromised recipients, so alternative strategies should be considered.
RESUMO
PURPOSE: Alloimmunization is a common consequence of chronic blood transfusion. Double alloantibody production may complicate the condition of such patients especially for finding matched blood. In this study, we evaluated the frequency of alloantibodies in thalassemic patients with previous history of transfusion reactions. SAMPLES AND METHODS: This study was performed on 441 multiply transfused thalassemia patients Antibody screening test was carried out using three cell-panel by gel method. Positive patients were followed up for antibody identification using 11-cell panel. Direct combs' test was performed to detect auto antibodies. RESULTS: In a total of 441 cases (362 thalassemia major and 79 intermedia), 234 were males (53.1%) and 207 females (46.9%); mean age 22 years, range 3-61 years. Alloimmunization was detected in 50(11.3%) patients, including 37(74%) patients with one alloantibody, 8(16%) with two antibodies, 4(8%) patients with unknown antibodies and one patient (2%) with autoantibody. The most common alloantibodies were anti-Rh antibodies (-E/e/C/c/Cw) (26%), anti-K (28%), anti-D (16%), and anti-Colton (4%). Double antibodies were detected in eight out of 50 patients, including: Anti-D+anti-C (8%), anti-D+anti-E (2%), anti-Kell+anti-D (2%), and anti-Kell+KPa (2%). A significant association was observed between the transfusion reaction history and the alloantibody detection results (p < 0.05). CONCLUSION: Antibody production against RBC antigens makes hard condition in regular blood transfusion. Double antibodies production may more complicate this situation. Thus, it is advisable to phenotype patients and matches the red cells in multiply transfused thalassemia patients.