OBJECTIVETo study the platelet morphology and function of an inherited macrothrombocytopenia disorder with abnormal large granules.

METHODSPlatelet 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.

RESULTSBoth 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.

CONCLUSIONBoth 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.

Adult ; Blood Platelets ; metabolism ; ultrastructure ; Female ; Humans ; Integrin beta3 ; biosynthesis ; Microscopy, Immunoelectron ; Platelet Glycoprotein GPIb-IX Complex ; biosynthesis ; Platelet Membrane Glycoprotein IIb ; biosynthesis ; Thrombocytopenia ; genetics ; pathology

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.
Cell-Derived Microparticles/chemistry/*metabolism/radiation effects ; Erythrocytes/*cytology/radiation effects ; Flow Cytometry ; Gamma Rays ; Humans ; Membrane Glycoproteins/metabolism ; Metalloendopeptidases/metabolism ; Platelet Membrane Glycoprotein IIb/metabolism

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.
Dependovirus ; genetics ; metabolism ; Genetic Vectors ; genetics ; Humans ; Platelet Glycoprotein GPIIb-IIIa Complex ; biosynthesis ; genetics ; Platelet Membrane Glycoprotein IIb ; genetics ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Thrombasthenia ; metabolism ; therapy

OBJECTIVETo explore the changes and their clinical significance of D-dimer and platelet glycoprotein (GP) in patients with coronary heart disease.

METHODSD-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.

RESULTSThere 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.

CONCLUSIONSD-dimer and GPIIb, GPIIIa may be regarded as the indexes of coronary thrombosis and used for predicting the severity of coronary events.

Aged ; Angina, Unstable ; diagnosis ; metabolism ; Case-Control Studies ; Coronary Disease ; diagnosis ; metabolism ; Female ; Fibrin Fibrinogen Degradation Products ; metabolism ; Humans ; Male ; Middle Aged ; Myocardial Infarction ; diagnosis ; metabolism ; Platelet Glycoprotein GPIIb-IIIa Complex ; metabolism ; Platelet Membrane Glycoprotein IIb ; metabolism ; Platelet Membrane Glycoproteins ; metabolism

OBJECTIVETo 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).

METHODSCD41⁺ 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.

RESULTSThe 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.

CONCLUSIONThere 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.

Animals ; Aorta ; cytology ; Bone Morphogenetic Protein 4 ; pharmacology ; Cell Differentiation ; Gonads ; cytology ; Interleukin-3 ; pharmacology ; Mesonephros ; cytology ; Mice ; Platelet Membrane Glycoprotein IIb ; metabolism ; Proto-Oncogene Proteins c-kit ; metabolism ; Yolk Sac ; cytology

OBJECTIVEThe 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.

METHODSSeven-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.

RESULTSFITC-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.

CONCLUSIONSThe increase of tPA in the acute phase after HIBD may be helpful to clot dissolving, but it induces neuronal apoptosis and aggravates brain injury.

Animals ; Animals, Newborn ; Apoptosis ; Fibrin ; analysis ; Hypoxia-Ischemia, Brain ; metabolism ; pathology ; Immunohistochemistry ; In Situ Nick-End Labeling ; Platelet Membrane Glycoprotein IIb ; analysis ; Rats ; Rats, Wistar ; Reverse Transcriptase Polymerase Chain Reaction ; Tissue Plasminogen Activator ; analysis ; physiology

OBJECTIVETo study the effects of IL-13 on the differentiation and expression of transcription factor c-fos of human erythroleukemia cell line (HEL) cells.

METHODSReverse 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.

RESULTSIL-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).

CONCLUSIONIL-13 prompts the differentiation of HEL cells and up-regulates the expression of c-fos.

Cell Differentiation ; drug effects ; Cell Line, Tumor ; Flow Cytometry ; Humans ; Interleukin-13 ; pharmacology ; Leukemia, Erythroblastic, Acute ; metabolism ; Platelet Membrane Glycoprotein IIb ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; Receptors, Interleukin-13 ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Up-Regulation ; von Willebrand Factor ; biosynthesis ; genetics