Megacariopoyesis humana in vitro: determinación de la concentración óptima de trombopoyetina / In vitro human megacaryocytopoiesis: determination of thrombopoietin optimal concentration / Megacariópoiesis humana in vitro: determinação da concentração ótima da trombopoietina

El estudio de la megacariopoyesis humana se ha visto obstaculizado por la relativa escasez de megacariocitos en la médula ósea (0,05-0,2 % de las células medulares), lo que ha llevado a la optimización de protocolos de expansión in vitro a partir de precursores de diversos orígenes (cordón umbilical, médula ósea y sangre periférica con o sin movilización previa). Los cultivos celulares a partir de precursores han permitido la producción y el estudio tanto de megacariocitos así como de proplaquetas y plaquetas Sin embargo, la producción in vitro óptima de megacariocitos que culminen todos los estadios de diferenciación es un reto aún no resuelto. En este trabajo reportamos los hallazgos concernientes a la determinación de las condiciones y concentraciones de trombopoyetina para lograr una óptima relación entre la cantidad de trombopoyetina empleada y el porcentaje y grado de diferenciación megacariocítica en muestras obtenidas de cinco donantes alogénicos aceptados para trasplante de médula ósea.

The study of human megakaryocytopoiesis has been hampered by the relative scarcity of megakaryocytes in bone marrow (0.05-0.2 % of medullary cells), which has led to the optimization of protocols of in vitro expansion of precursors from diverse sources (umbilical cord, bone marrow and peripheral blood with or without previous mobilization). Cell cultures from different precursors have allowed the production and study of megakaryocytes as well as proplatelets and platelets. However, the in vitro production of megakaryocytes that culminate all stages of differentiation is a challenge that has not yet been resolved. In this work we report the findings related to the determination of thrombopoietin treatment conditions and concentrations to achieve an optimal relationship between the amount of thrombopoietin and the percentage and degree of megakaryocytic differentiation in five allogeneic donors that were accepted for bone marrow transplantation.

O estudo da megacariopoiese humana tem sido dificultado pela relativa escassez de megacariócitos na medula óssea (0,05-0,2 % das células medulares), o que levou à otimização dos protocolos de expansão in vitro a partir de precursores de diversas origens (cordão umbilical, medula óssea e sangue periférico com ou sem mobilização prévia). Culturas de células a partir de precursores permitiram a produção e o estudo tanto de megacariócitos e de proplaquetas e plaquetas. No entanto, a produção ótima in vitro de megacariócitos que culminam em todas as fases de diferenciação é um desafio ainda não resolvido. Neste trabalho, relatamos as descobertas relativas à determinação das condições e concentrações de trombopoietina para obter uma relação ótima entre a quantidade de trombopoietina usada e a taxa e o grau de diferenciação megacariocítica em amostras obtidas de cinco doadores alogênicos aceitos para transplante de medula óssea.

Origin, tempo, and mode of the spread of DENV-4 Genotype IIB across the state of São Paulo, Brazil during the 2012-2013 outbreak

BACKGROUND Dengue virus type 4 (DENV-4) was first reported in Brazil in 1982 and since then no more cases were detected again in Brazil until 2010, when the virus was reintroduced. Over the following years, the virus spread to several Brazilian states and resulted in about 1,400,000 dengue cases, in 2013. The largest number of cases were documented in the Southeast macro-region. OBJECTIVES To determine the phylogeography of DENV-4 Genotype IIB strains isolated during the epidemics in 2012-2013 in São Paulo, Brazil, we aimed to contextualise the contribution of viruses sampled in different localities across the overall movement of DENV-4 in Brazil. METHODS Based on the envelope gene sequences retrieved from GenBank, we employed a Bayesian phylogeographic approach to assess the spatiotemporal dynamics of DENV-4 Genotype IIB in São Paulo, Brazil. FINDINGS The dispersal dynamics of DENV-4 Genotype IIB in Brazil indicated Rio de Janeiro and Mato Grosso states as the most likely routes toward São Paulo before the 2012-2013 outbreak. Likewise, Guarujá and São José do Rio Preto facilitated viral spread and transmission to other localities in the South and Southeast macro-regions in Brazil. CONCLUSIONS The spread pattern of DENV-4 Genotype IIB strains across the country supports two independent introductions of the virus in São Paulo in a short period of time. Furthermore, São Paulo appears to have played a pivotal role in the dissemination of DENV-4 to other locations in Brazil.

Effect of Irradiation on Microparticles in Red Blood Cell Concentrates

Changes in microparticles (MP) from red blood cell (RBC) concentrates in the context of irradiation have not been investigated. The aim of this study was to evaluate how irradiation affects the number of MPs within transfusion components. Twenty RBC concentrates, within 14 days after donation, were exposed to gamma rays (dose rate: 25 cGy) from a cesium-137 irradiator. Flow cytometry was used to determine the numbers of MPs derived from RBC concentrates before and 24 hr after irradiation. The mean number of MPs (±standard deviation) in RBC concentrates was 21.9×10(9)/L (±22.7×10(9)/L), and the total number of MPs ranged from 2.6×10(9)/L to 96.9×10(9)/L. The mean number of MPs increased to 22.6×10(9)/L (±31.6×10(9)/L) after irradiation. Before irradiation, the CD41-positive and CD235a-positive MPs constituted 9.5% (1.0×10(9)/L) and 2.2% (263×10(6)/L) of total MPs, respectively. After irradiation, CD41-positive MPs increased to 12.1% (1.5×10(9)/L) (P=0.014), but the CD235a-positive MPs decreased to 2.0% (214×10(6)/L) of the total MPs (P=0.369). Irradiation increases the number of CD41-positive MPs within RBC concentrates, suggesting the irradiation of RBC concentrates could be associated with thrombotic risk of circulating blood through the numerical change.

Analysis of β -thalassemia mutations in Guizhou Province / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the spectrum of β -thalassemia mutations in Guizhou Province.</p><p><b>METHODS</b>For 542 individuals suspected to have β -thalassemia by decreased mean corpuscular volume (MCV) and corpuscle hemoglobin (MCH) by routine blood test and hemoglobin electrophoresis, reverse dot blot hybridization (RDB) was performed to detect 17 known β -thalassemia mutations, including 8 common and 9 rare mutations. For cases where no mutation was identified, the entire human β -globin gene was screened to find other rare mutations. The distribution and frequencies of detected β -thalassemia mutations were then analyzed.</p><p><b>RESULTS</b>A total of 460 individuals were diagnosed as β -thalassemia by DNA analysis, which included 352 heterozygotes, 67 compound heterozygotes and 41 mutant homozygotes. A total of 12 β -thalassemia mutations were detected in these individuals. The mutations have ranked from high to low frequency as: CD17 (40.74%), CD41-42 (33.69%), IVS-II-654 (13.76%), -28 (3.70%), β E (3.35%), CD71-72(1.94%), CD43 (1.06%), IVS-I-1 (0.71%), CD27-28 (0.35%), -29(0.35%), CAP (0.18%), and CD121 (0.18%). The former six mutations have accounted for 97.18% of all. CD121 (GAA> TAA) detected from a heterozygote, as a dominant mutation, has been firstly found in the Chinese population.</p><p><b>CONCLUSION</b>The spectrum of β -thalassemia in Guizhou Province showed certain distinct characteristics, with CD17 being the most common mutation. The newly discovered mutation of CD121 has expanded the spectrum of β -thalassemia in Chinese population. Our result may provide valuable information for the prevention and control of β -thalassemia in Guizhou.</p>

Differentiation potential of CD41⁺ cells derived from the mouse aorta-gonad-mesonephros region, yolk sac and embryonic circulating blood / 中华血液学杂志

<p><b>OBJECTIVE</b>To compare the differentiation ability difference of hematopoietic, mesenchymal and endothelial potential between CD41⁺ cells derived from the mouse aorta-gonadmesonephros (AGM) region, yolk sac (YS) and embryonic circulating blood (CB).</p><p><b>METHODS</b>CD41⁺ cells were sorted from AGM, YS and CB. The CD45 and c-kit expression were studied in CD41⁺ cells by flow cytometry. IL-3 and bone morphogenetic protein 4 (BMP-4) treatment together with semi solid culture were used to assess hematopoietic potential difference of CD41⁺ cells. Immunofluorescence staining of α-SMA was used to assess mesenchymal potential difference. The endothelial cell induction system was used to assess endothelial potential difference.</p><p><b>RESULTS</b>The proportions of CD45+ cells in CD41⁺ population were 51.9% (AGM), 45.8% (YS) and 22.2% (CB), respectively, while those of c-kit⁺ cells were 40.0% (AGM), 39.6% (YS) and 36.2% (CB), respectively. After stimulated by IL-3 factor, the number of total colonies increased in all three groups-derived CD41⁺ cells compared to that of unstimulated group[(14.1±1.9) vs (1.2±0.2), (32.4±1.1) vs (18.4±2.2) and (41.8±0.9) vs (10.4±1.8)], (P<0.01). After stimulated by BMP-4 factor, compared to unstimulated group, CFU-Mix colony number in CD41⁺ cells from AGM region and YS were significantly decreased[(0.5±0.6) vs (3.2±0.8), (1.3±0.7) vs (7.4±1.7)](P<0.01), but there was no difference in CB group[(2.5±0.5) vs (3.9±1.5)](P>0.01). The mesenchymal marker α-SMA was highly expressed in CD41⁺ cells from AGM region and YS, but lowly expressed in CD41⁺ cells from CB.</p><p><b>CONCLUSION</b>There are some differences between CD41⁺ cells in AGM region, YS and CB on hematopoietic cell surface marker expression, hematopoietic colony formation with IL-3 and BMP-4 stimulation.</p>

Analysis of clinical features and genotype in three Chinese pedigrees with Glanzmann thrombasthenia / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the clinical feature and alpha II b beta 3 gene mutations of three Glanzmann thrombasthenia (GT) pedigrees.</p><p><b>METHODS</b>Platelet counts (BPC), blood film, bleeding time, platelet aggregation and flow cytometry were used for phenotype diagnosis of all the patients. All the exons of alpha II b and beta 3 genes were amplified by polymerase chain reaction (PCR) and direct sequencing was performed for mutational screening. One hundred and three healthy blood donors were as normal controls.</p><p><b>RESULTS</b>Three probands showed normal BPC, defective platelets aggregation, prolonged bleeding time and significantly reduced platelet aggregation to ADP, epinephrine, and collagen, while relatively normal aggregation to ristocetin. Flow cytometry showed platelet surface expressed alpha II b beta 3 was strongly reduced in proband 1 and proband 3 and mildly reduced in the amount of surface expressed alpha II b beta 3 (63%) in proband 2. Sequencing results showed that proband 1 had a G10A homozygous mutation in alpha II b, and a G1412T homozygous mutation in beta3. Compound heterozygous mutations in beta3, G1199A and 1525delC were identified in proband 2. No mutations in alpha II b beta 3 gene were identified in proband 3.</p><p><b>CONCLUSIONS</b>Compound homozygous mutations, GI0A in alpha II b and G1412T in beta3, lead to GT in proband 1. Compound heterozygous mutations in beta3, G1199A and 1525delC, lead to GT in proband 2. The mutations of G10A, G1412T and 1525delC were reported for the first time in GT patients.</p>

Changes and significance of endogenous tissue plasminogen activators in cerebral hypoxia-ischemia in neonatal rats / 中国当代儿科杂志

<p><b>OBJECTIVE</b>The mechanism of neonatal hypoxic-ischemic brain damage (HIBD) remains unclear and effective treatment approach is limited for this disorder. Many studies have shown that tissue-type plasminogen activator (tPA) plays an important role in nervous system. This study investigated the effect of tPA in HIBD in neonatal rats.</p><p><b>METHODS</b>Seven-day-old Wistar rat pups were used for the Rice-Vannucci model of neonatal hypoxia-ischemia (HI). Brain samples were collected 1, 4, and 24 hrs after HI. FITC-Dextran was injected into the left ventricle of pups after HI to observe reperfusion defects of the neonatal brain. RT-PCR and tPA zymogram were used to detect the expression and activity of tPA. Double immunostaining, terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) and DAPI staining were used to detect the expression of integrin GPIIb and fibrin and neuronal apoptosis.</p><p><b>RESULTS</b>FITC-Dextran perfusion analysis indicated there was obvious infarct area in the neonatal brain and the expression of integrin GPIIb and fibrin increased significantly 1 hr after HI compared with the contralateral side. The infarct area decreased and the expression of integrin GPIIb and fibrin were reduced 4 hrs after HI. The expression and activity of tPA increased significantly in neonatal rats after HI, and peaked at 4 hrs after HI. The number of apoptotic neural cells increased progressively with the prolonged reperfusion time following HI.</p><p><b>CONCLUSIONS</b>The increase of tPA in the acute phase after HIBD may be helpful to clot dissolving, but it induces neuronal apoptosis and aggravates brain injury.</p>

Novel frame-shift mutation of 540 A deletion in GP IIb gene from a patient with Glanzmann thrombasthenia / 中南大学学报(医学版)

OBJECTIVE@#To explore the molecular mechanism of Glanzmann thrombasthenia (GT).@*METHODS@#All 45 exons of alphaIIb and beta3 subunit genes as well as their splicing sites were amplified by polymerase chain reaction(PCR) with 40 primer pairs, and then the PCR products were used to screen the gene mutation by single strand conformation polymorphism-polyacrylamide gel electrophoresis (SSCP-PAGE). The mutation was further confirmed by direct DNA sequencing.@*RESULTS@#A DNA band alterated migration was detected after SSCP-PAGE. DNA sequencing showed that a base deletion within the band at the site of 540 in GPIIb gene(540A) was found.@*CONCLUSION@#The frame-shift mutation caused by the deletion of 540A in GPIIb gene is a novel mutation which is a genetic defect in patients with GT.

3 polymorphisms of gene GP IIb and the function of GP IIb T13959 G in the platelet transfusion refractoriness / 中南大学学报(医学版)

OBJECTIVE@#To investigate the relationship among 3 polymorphisms of GP IIb and the function of GP IIb T13959 G in the platelet transfusion refractoriness(PTR).@*METHODS@#The 26th exon, the 30th exon and the 21st intron of gene GP IIb in 110 individuals were amplified by polymerase chain reaction (PCR), and the PCR products were analyzed with single-strand conformation polymorphism(SSCP) and sequenced to investigate whether there was linkage among the polymorphisms of the gene. Human platelet antigen-3 (HPA-3) gene frequency was detected by Fok I enzyme in 147 patients with hematologic diseases, and was compared with that in 110 normal individuals. Forty-four patients who received apheresis platelet transfusion repeatedly were randomly divided into the HPA-3 homotype group and the control group. The antibodies of the platelet were detected after 3 times of platelet transfusion.@*RESULTS@#There were polymorphisms of gene GP IIb in the 26th, 30th exon and the 21st intron, and the mutations were: T changed into G in 13,959 th of the 26th exon; C changed into T in 16,997 th of the 30th exon; the 9 bps deletion occurred in 11,996-12,004 th of the 21st intron. The 3 polymorphisms had synchronization in the individuals. The results of Fok I enzyme indicated that the frequency of HPA-3a was 83.6% (92/110)and 81.9%(119/147), and that of HPA-3b was 16.4%(18/110) and 19.1%(28/147) in the normal individuals and the patients respectively. There was no significant difference between the patients and normal individuals (P>0.05). After the platelet transfusion, the antibodies of all the cases of the homotype platelet transfusion were negative, but the antibodies in 2 cases of the control group were positive, and there was antibody to HPA-3a in one of the antibodies positive cases.@*CONCLUSION@#(1)There is close linkage among the polymorphisms of gene GP IIb, which is T->G in 13 959 th of the 26th exon, C->T in 16,997 th of the 30th exon, and the 9 bps deletion in 11,996-12,004 th in the 21st intron. (2)The gene frequency of HPA-3a/3b is similar in the normal individuals and patients with hematologic diseases. (3) HPA-3 system may be one of the reasons for PTR in Chinese.

The immunoreactivity of IgG and its fragments from ITP patients and their effects on platelet aggregation function / 中华血液学杂志

<p><b>OBJECTIVE</b>To prepare ITP plasma IgG and its F(ab')2 fragments and investigate their immunoreactivity to platelet GPIIb/IIIa and/or GPIb/IX and their effects on platelet aggregation function.</p><p><b>METHODS</b>The ITP patients having inhibitory autoantibody to the platelet aggregation were selected by modified MAIPA and platelet aggregation test with turbidimetry. Plasma IgG and its F(ab')2 fragments were prepared by streptococcal protein A affinity column and pepsin digestion. The immunoreactivity and the effects on platelet aggregation function of the whole antibody and its fragments were detected by modified MAIPA and platelet aggregation test, respectively.</p><p><b>RESULTS</b>(1) Anti-platelet GPIIb/IIIa and/or GPIb/IX autoantibodies were detected in 34 of 68 (53.6%) ITP patients' plasmas and that from 5 patients significantly inhibited the platelet aggregation induced by ADP or ristocetin. (2) By using protein A column combined with protease digestion, pure IgG and its F(ab')2 fragments were successfully obtained. (3) The purified IgG and its F(ab')2 fragments retained the ability to bind to their respective glycoproteins and inhibited the platelet aggregation function, whereas the IgG depleted plasma lost the ability of binding to the platelet GPs.</p><p><b>CONCLUSIONS</b>F(ab')2 fragment of the IgG antibody is a functional fragment, which not only has the binding ability to the platelet GPs but also inhibits the platelet aggregation function in a dose-dependent manner.</p>

Construction of rAAV2-GPIIb/IIIa vector and test of its expression and function in vitro / 中国实验血液学杂志

This study was aimed to explore the possibility of rAAV2 vector-mediating gene therapy for Glanzmann' s thrombasthenia. The rAAV2-GPIIb/IIIa vector was constructed. The GPIIb/IIIa gene expression in mammal cell were examined by different methods, such as: detection of mRNA expression in BHK-21 cells after 24 hours of infection (MOI = 1 x 10(5) v.g/cell) was performed by RT-PCR; the relation between MOI and quantity of GPII6/IIIa gene expression was detected by FACS after 48 hours of infection; GPIIb/IIIa protein expression in BHK-21 cells after 48 hours of infection (MOI = 10(5) v x g/cell) was assayed by Western blot, GPIIb/IIIa protein expression on cell surface was detected by immunofluorescence, and the biological function of expressing product was determined by PAC-1 conjunct experiments. The results showed that GPIIb/IIIa gene expression in mRNA level could be detected in BHK-21 cells after 24 hours of infection at MOI = 1 x 10(5) v x g/cell and the GPIIb/IIIa gene expression in protein level could be detected in BHK-21 cells after 48 hours of infection at MOI = 1 x 10(5) v x g/cell. In certain range, quantity of GPIIb/IIIa gene expression increased with MOI, but overdose of MOI decreased quantity of GPIIb/IIIa gene expression. Activated product of GPIIb/IIIa gene expression could combined with PAC-I, and possesed normal biological function. In conclusion, rAAV2 vactor can effectively mediate GPIIb and GPIIIa gene expressing in mammal cells, and the products of these genes exhibit biological function. This result may provide a basis for application of rAAV2 vector in Glanzmann's thrombasthenia gene therapy in furture.

Effects of interleukin 13 on the differentiation and expression of transcription factor c-fos of HEL cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the effects of IL-13 on the differentiation and expression of transcription factor c-fos of human erythroleukemia cell line (HEL) cells.</p><p><b>METHODS</b>Reverse transcription polymerase chain reaction (RT-PCR) was used to observe the mRNA expression of IL-13 receptor a 1, GP i b, vWF and c-fos, and Western blot and cytometry were used to analyse their protein expression.</p><p><b>RESULTS</b>IL-13 receptor a 1 was expressed on HEL cells. IL-13 (100 ng/ml ) up-regulated the mRNA expression of GP II b and vWF. The ratio of luminous absorption (LA) of GP I b to p-actin bands ( AB) was 1. 303 in control group, whereas was 2. 912 in experiment group; being 2. 23-fold higher than that in control group (P < 0. 05). The ratio of LA to AB for vWF was 0.217 in control group, and 0. 506 in experiment group; indicating a 2. 33-fold increase in experiment group (P <0. 05). The protein expression of GP I b and vWF was significantly increased in experiment group, compared with that in control group. IL-13 inducing the increased expression of c-fos mRNA and protein of HEL cells peaked at 30 min and 60 min, respectively. The ratio of LA to AB for c-fos was also increased at 30 min and 60 min (P <0. 05).</p><p><b>CONCLUSION</b>IL-13 prompts the differentiation of HEL cells and up-regulates the expression of c-fos.</p>

A preliminary study of an inherited macrothrombocytopenia disorder with abnormal large granules / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the platelet morphology and function of an inherited macrothrombocytopenia disorder with abnormal large granules.</p><p><b>METHODS</b>Platelet size and structure were investigated by both light microscopy and electron microscopy. The platelet membrane expression of GP I b, GP II b, GPIII a, P-selectin and CD63 were analyzed by using respective monoclonal antibodies. Platelet 5-hydroxy-tryptamine was measured with spectrophotofluorometer.</p><p><b>RESULTS</b>Both the patient and her father had large granules in their platelets, with exocytosis being easily observed. The expressions of GP I b, GP II b and GP II a on the platelets were in normal range, while P-selectin and CD63 were somewhat increased. The abnormal large granules were not the alpha granules, lysosomes or dense bodies.</p><p><b>CONCLUSION</b>Both morphological and functional abnormalities of the platelets from the patient are clearly distinguishable from other hereditary giant platelet disorders. It would probably represent a novel platelet disorder.</p>

Trombocitopenia em unidade coronariana: relato de caso e revisão de literatura / Thrombocytopenia in the coronary care unit: case report and review of the literature

Trombocitopenia é um achado frequente em unidades de terapia intensiva cardiológica; neste ambiente onde grande parte do arsenal terapêutico diminui a coagulabilidade sanguínea, a baixa contagem plaquetária representa um desafio ao médico intensivista. Este relato de caso ocorreu no Hospital Johns Hopkins, em Baltimore(EUA), em 2003, quando o autor esteve em visita a esta instituição. É apresentada uma revisão da literatura e uma breve discussão do caso

Study on the expansion of megakaryocyte progenitors in vitro from cord blood / 中国实验血液学杂志

This study was aimed to investigate the effect of various cytokines on megakeryocytes expansion in vitro from human cord blood CD34(+) cells in order to establish an optimal culture system for MK expansion. Mononuclear cells were obtained by Ficoll-Hapaque density gradient separation. CD34(+) cells were positively isolated using a CD34 progenitor cell isolation kit. CD34(+) cells were placed into 24 well plates at a concentration of 2 x 10(4) per well. Each well contained 1 ml of IMDM with the present of effective MK cells growth cytokines. Clonogenic potentials of MK progenitor were assayed using a methylcellulose cultures system. The results suggested that four cytokines (IL-3 + IL-6 + TPO + FLT3L) culture system could effectively induce and expand cord blood CD41(+) MK cells. The number of CD41(+) cells expanded 154.67 +/- 32.21-fold on day 7, and 193.23 +/- 25.24-fold on day 14. In conclusion, established expansion system in vitro for MK cells provides experimental foundation for recovery of platelets after cord blood transplantation.

The changes and their clinical significance of D-dimer and platelet glycoprotein in patients with coronary heart disease / 中华心血管病杂志

<p><b>OBJECTIVE</b>To explore the changes and their clinical significance of D-dimer and platelet glycoprotein (GP) in patients with coronary heart disease.</p><p><b>METHODS</b>D-dimer and GP in 20 patients with stable angina (SA group), 48 patients with unstable angina (UA group), and 20 control cases were measured. The changes of D-dimer and GP in patients with and without coronary events were compared. The sensitivity of those changes in the diagnosis of coronary events was evaluated.</p><p><b>RESULTS</b>There were significant differences of D-dimer and GP between UA group and SA group or control group (P < 0.01), while there was no significant difference between SA group and control group (P > 0.05). There were also significant differences of D-dimer and GP between patients with coronary events and patients without coronary events (P < 0.05). In the sensitivity test for detecting coronary events, D-dimer and GPIIb, GPIIIa were much more sensitive than other parameters.</p><p><b>CONCLUSIONS</b>D-dimer and GPIIb, GPIIIa may be regarded as the indexes of coronary thrombosis and used for predicting the severity of coronary events.</p>

Significance of detecting platelet associated antibody and platelet membrane glycoprotein for diagnosis of immune thrombocytopenia / 中国实验血液学杂志

The aim of this study was to explore application value of detecting platelet associated antibody and platelet membrane glycoprotein in the diagnosis and prognosis for immune thrombocytopenia. The platelet associated immunoglobulin (PAIg) and platelet membrane glycoprotein (CD41, CD61, GPIIb/IIIa) in 76 cases of immune thrombocytopenia and 30 healthy subjects were determined by FCM. The results showed that PAIg level in ITP patients included PAIgG (31.25 +/- 18.06)%, PAIgM (32.41 +/- 15.51)%, PAIgA (23.39 +/- 16.67)% which were remarkedly higher than in health control (10.48 +/- 5.05)%, (9.40 +/- 4.42)% and (7.23 +/- 3.61)% (P < 0.001). In patients with secondary immune thrombocytopenia (chronic aplastic anemia, SLE, Evans syndrome, liver cirrhosis hypersplenism, etc), PAIg level was higher than that in control group, while the platelet membrane glycoprotein in the blood of these patients was lower than that in control group. The level of PAIg decreased (P < 0.05) after treatment, but platelet membrane glycoprotein increased (P < 0.01). The result suggested that measurements for platelet membrane glycoprotein and platelet associated antibody by FCM were practical with high sensitivity, rapidity and simplicity used as a routine method in diagnosis and evaluation of the therapeutic effects in immune thrombocytopenia patients.

Study on bone marrow megakaryocytes in children patients with idiopathic thrombocytopenic purpura / 中国实验血液学杂志

To observe the morphological characteristics and hematopoietic function of bone marrow megakaryocyte (MK) in children patients with idiopathic thrombocytopenic purpura (ITP), and to preliminary analyse the cause and mechanism of thrombocytopenia. CD41 McAb immunohistochemical technique was used to detect micromegakaryocyte in bone marrow smear. Plasma clot culture and CD41 McAb immunohistochemical technique were used for the MK-colony forming assay. The results showed that there was no statistical difference of the positive rate of micromegakaryocyte between groups of ITP and control, but type I lymphocyte-like micromegakaryocyte was infrequent. The number of micromegakaryocyte and the formation rates of CFU-MK and BFU-MK in ITP group were significantly higher than those in control group. The normal MK releasing platelet could be easily found in the culture system. The MK colony formation rate was decreased in a patient with chronic ITP. In conclusion, the increment of type II, III, IV micromegakaryocytes is one of pathologic phenomenon of ITP. These small megakaryocytes can develop and mature to normal megakaryocytes in the condition of ex vivo culture. The developmental abnormity of MK is a possible reason for thrombocytopenia among partial patients with ITP, especially the chronic cases.