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PubMed1997

An in vitro study on thalassemic erythroid precursors in liquid culture.

Thalassemia is an inherited hematological disorder which can generally be classified according to the affected globin imbalance (alpha- or beta-globin) into two main types, i.e. alpha-thalassemia and beta-thalassemia, respectively. There is a wide range of cellular abnormalities associated with thalassemic erythrocytes such as hypochromia, microcytosis, reduced cellular deformability and membrane oxidative damage. The red cell abnormalities lead to premature destruction with marrow erythroid hyperplasia and ineffective erythropoiesis. The abnormalities in thalassemic red blood cells have been found along the erythroid differentiation pathway other than the mature stage as previously shown in bone marrow erythroid precursors and in reticulocytes, the penultimate stage of erythroid differentiation. However, there is a lag in our understanding of the more primitive erythroid stages due to the difficult and hazardous marrow aspiration and heterogeneity of cells derived. We have utilized a novel method of Two-Phase Liquid Culture (TPLC) of beta-thalassemia/HbE erythroid precursors instead of conventional semisolid culture. This type of liquid culture can given higher cell yield with quite synchronous cell differentiation stages and easily be applied for other cellular analytical techniques. The peripheral blood mononuclear cells (PBMC) obtained from non-splenectomized and splenectomized beta-thalassemia/HbE patients were first cultured in medium supplemented with 5637 conditioned medium for a 6-day period (phase I) and then transferred to medium supplemented with recombinant human erythropoietin to allow the terminal differentiation of erythroid precursors (phase II). During the phase I or II, the cultured cells were periodically sampled to determine the cell number, cytocentrifuged on glass slides and stained with Wright stain for morphological assessment of their differentiation stages and analyzed flow cytometrically by staining with fluoresceinated anti-transferrin receptor (anti-CD71) and R-phycoerythrin-conjugated anti-glycophorin A. After assessment by flow cytometry, the remaining stained cells were cytocentrifuged on glass slides and photographed by a fluorescent microscope and a laser scanning confocal microscope. The results of morphological assessment, flow cytometric analysis and microscopic pictures will be presented.

Cells, Cultured↗
PubMed2003

The cleaved peptide of PAR1 is a more potent stimulant of platelet-endothelial cell adhesion than is thrombin.

PURPOSE: Platelet-endothelial cell adhesion is an important pathologic response to vessel injury or inflammation. On binding to its endothelial or platelet G protein-linked seven-transmembrane domain receptor, protease-activated receptor-1 (PAR1), thrombin releases a 41-amino acid peptide (TR(1-41)). We examined the effect of TR(1-41) on platelet activation and on platelet-endothelial cell adhesion. METHODS: A monolayer of confluent human saphenous vein endothelial cells was incubated with washed human platelets. Platelets were stimulated with either TR(1-41), TR(21-41), scrambled TR(1-41), adenosine diphosphate (ADP)-epinephrine (EPI), thrombin, or thrombin receptor activating peptide (TRAP). Platelet activation was identified with flow cytometry. The magnitude of platelet-endothelial cell adhesion was determined with a laser scanning cytometer that scanned the monolayer of endothelial cells and identified fluorescently bound platelets. RESULTS: Maximal thrombin stimulation (0.1 U/mL) induced a threefold increase in platelets bound to endothelial cells compared with buffer alone. Stimulation with TR(1-41) (20 mmol/L) tripled the number of platelets bound to endothelial cells compared with thrombin. Scrambled sequence of TR(1-41) (20 mmol/L) and TR(21-41) (20 mmol/L), neither of which induces platelet activation, had minimal effect on platelet adhesion. Both TRAP (20 mmol/L) and ADP-EPI (20 mmol/L) induced less platelet-endothelial cell adhesion than did thrombin. TR(1-41)-induced platelet-endothelial cell adhesion was partially blocked by glycoprotein (GP)IIb-IIIa-specific monoclonal antibody, 10E5 (10 mg/mL). CONCLUSIONS: TR(1-41), the cleaved peptide of PAR1, is a more potent stimulant of platelet-endothelial cell adhesion than is thrombin, TRAP, or ADP-EPI, and this adhesion is at least in part mediated by the platelet GPIIb-IIIa receptor.

Cell Adhesion↗
PubMed2001

Expression and colocalization of cytokeratin 1 and urokinase plasminogen activator receptor on endothelial cells.

The cellular localization of human cytokeratin 1 (CK1), urokinase plasminogen activator receptor (uPAR), and gC1qR, high-molecular-weight kininogen (HK)-binding proteins on endothelial cells, was determined. CK1 was found on the external membrane of nonpermeabilized endothelial cells by immunoperoxidase staining, immunofluorescence, and transmission electron microscopy using immunogold. Human umbilical vein endothelial cells (HUVECs) had 7.2 +/- 0.2 x 10(4) specific CK1 membrane sites/cell by (125)I-F(ab')(2) anti-CK1 antibody binding. Flow cytometry studies confirmed the presence of CK1, uPAR, and gC1qR on HUVECs. On laser scanning confocal microscopy and transmission electron microscopy, CK1 and uPAR, but not gC1qR, colocalized on the cell surface of HUVECs. The HUVEC surface distribution of these proteins was distinctly different from that for von Willebrand factor. In competitive inhibition experiments, anti-CK1, anti-uPAR, or anti-gC1qR blocked both biotin-HK binding and prekallikrein (PK) activation on HUVECs with an inhibitory concentration of 50% (IC(50)) of 300 to 350 nM, 50 to 60 nM, or 35 to 100 nM, respectively. Also, antibodies to uPAR and gC1qR each inhibited 86% of kallikrein-mediated, 2-chain urokinase plasminogen activation, whereas antibodies to CK1 only inhibited 24% of plasminogen activation. On HUVECs, CK1 and uPAR, but not gC1qR, colocalized to be a multiprotein receptor complex for HK binding, PK activation, and 2-chain urokinase plasminogen activation.

Amino Acid Sequence↗
PubMed2005

Human gingival fibroblast integrin subunit expression on titanium implant surfaces.

BACKGROUND: Implant surface characteristics have been shown to modify cell behavior and regulate integrin expression. Integrin expression and resultant integrin-mediated cellular activity are essential components of tissue healing and homeostasis. Although both osseous and soft tissue healing around dental implants are critical to clinical success, there is limited information available on the effect of implant surfaces on integrin expression in soft tissues. Therefore, the aim of this study was to examine integrin expression for gingival fibroblasts on titanium surfaces and the influence of titanium surface roughness on integrin expression and cell morphology. METHODS: Human gingival fibroblasts were cultured on smooth (polished) and rough (sand-blasted acid-etched) titanium surfaces and a cell culture plastic (control) surface. To analyze integrin expression, total RNA was isolated from experimental and control cells, and levels of integrin subunit mRNA were assessed by reverse transcription-polymerase chain reaction (RT-PCR) using primers specific for the alpha2, alpha4, alpha5, alpha(v), and beta1 integrin subunits and aldolase (internal control). PCR products were analyzed by polyacrylamide gel electrophoresis (PAGE), confirmed via DNA sequencing, and quantified using computer-assisted densitometry. The expression of the integrin subunits was analyzed at the protein level using flow cytometry, as well as fluorescence and confocal laser microscopy. Cell morphology was evaluated using scanning electron microscopy (SEM). RESULTS: Our experiments demonstrated cellular expression of the alpha2, alpha4, alpha5, alpha(v), and beta1 integrin subunits at both mRNA and protein levels on all surfaces. In addition, the alpha4 and beta1 mRNA levels were significantly increased on smooth titanium relative to plastic surfaces (P <.05) with intermediate mRNA levels found on the rough titanium surfaces. The smooth titanium surfaces exhibited a flat monolayer of cells, while rough titanium surfaces showed cells orienting themselves along surface irregularities. CONCLUSIONS: These results demonstrate the presence of multiple integrin subunits in human gingival fibroblasts grown in contact with titanium implant surfaces and that titanium surface roughness alters cellular morphology but appears to have limited effects on integrin expression. This study provides insight into the complicated cellular and molecular events occurring at the implant surface that may be critical to optimizing the soft tissue interactions with the soft tissue-implant interface.

Cell Adhesion↗
PubMed1999

[Changes of F-actin in neutrophils under fluid shear stress].

Neutrophils are the main defender in the body, and their physiological characters are associated with functions. Because the environments of neutrophils maturation and storage are different from their function environment, they have to adapt thess changes. Filamentous actin (F-actin), as one of the important cytoskeleton components of neutrophils, has different quantity and distribution under different physiological conditions. Previous researches are all focused on static activation of neutrophils using various stimulants such as formyl-methionyl-leucyl-phenylalanine (f-MLP) and tumor necrosis factor(TNF). In our study, Low-Shear 30 and NXE-1 rheometers were used to provide steady or sinusoidaloscillatory fluid shear stress on separated neutrophils. Fluorescent agent Tritc-Phalloidin was added to label F-actin and the mean fluorescent intensity tested by flow cytometry was used as the indication of F-actin quantity and confocal laser scan microscope was used to detect the distribution of F-actin in neutrophils. We found that F-actin polymerization was significantly decreased under both steady and sinusoidal-oscillatory shear stress when the shear stress was below certain level. The density of the cortex F-actin near cell membrane in cells sheared became thinner than in that without shear stress. f-MLP and TNF both could increase actin polymerization in neutophils due to activation. When neutorphils were activated with f-MLP or TNF under shear stress, the quantity of F-actin in the cells was also decreased, but it was still higher than that in the cells absent of f-MLP or TNF. We concluded that neutrophils had different mechanical properties in different fluid environments. F-actin content and distribution would change according to the shear stress values to play their determinant role in neutrophils mechanical adaptation. The mechanism of actin polymerization in neutrophils for mechanical action is probably different from that for chemoattractants and cytokine activation.

Actins↗
PubMed2004

Cytomics--new technologies: towards a human cytome project.

BACKGROUND: Molecular cell systems research (cytomics) aims at the understanding of the molecular architecture and functionality of cell systems (cytomes) by single-cell analysis in combination with exhaustive bioinformatic knowledge extraction. In this way, loss of information as a consequence of molecular averaging by cell or tissue homogenisation is avoided. PROGRESS: The cytomics concept has been significantly advanced by a multitude of current developments. Amongst them are confocal and laser scanning microscopy, multiphoton fluorescence excitation, spectral imaging, fluorescence resonance energy transfer (FRET), fast imaging in flow, optical stretching in flow, and miniaturised flow and image cytometry within laboratories on a chip or laser microdissection, as well as the use of bead arrays. In addition, biomolecular analysis techniques like tyramide signal amplification, single-cell polymerase chain reaction (PCR), and the labelling of biomolecules by quantum dots, magnetic nanobeads, or aptamers open new horizons of sensitivity and molecular specificity at the single-cell level. Data sieving or data mining of the vast amounts of collected multiparameter data for exhaustive multilevel bioinformatic knowledge extraction avoids the inadvertent loss of information from unknown molecular relations being inaccessible to an a priori hypothesis. CHALLENGE: It seems important to address the challenge of a human cytome project using hypothesis-driven molecular information collection from disease associated cell systems, supplemented by systematic and exhaustive knowledge extraction. This will allow the description of the molecular setup of normal and abnormal cell systems within a relational knowledge system, permitting the standardised discrimination of abnormal cell states in disease. As one of the consequences, individualised predictions of further disease course in patients (predictive medicine by cytomics) by characteristic discriminatory data patterns will permit individualised therapies, identification of new pharmaceutical targets, and establishment of a standardised framework of relevant molecular alterations in disease. This special issue of Cytometry, on new technologies in cytomics, focuses on prominent examples of this presently fast-moving scientific field, and represents one of the preconditions for the formulation of a human cytome project.

Cell Biology↗
PubMed1993

Beam-shaping optics for a slit-scan flow cytometer.

Two sets of beam-shaping optical systems for slit-scan flow cytometry were made for 360 nm and 488 nm wavelength argon-ion lasers. A thin, ribbonlike laser beam was formed by compressing the zero-order component of Fraunhofer diffraction patterns caused by a slit vertically long and horizontally narrow by means of an objective with a numerical aperture of 0.5. The objectives were designed to include a jet stream that performed as a cylindrical lens. The characteristic values of the optical systems and lens alignments of the systems are described in this work. The beam size at focus was 0.95 micron in thinness and 85.0 microns in width on design values. The actual beam thinness for the UV laser was estimated to be 1.3 microns, based on shapes of pulses generated when fluorescence microspheres of diameter 1.0 microns passed across the beam. The usefulness of the optical systems was also proved by its ability to measure HeLa chromosomes.

Analog-Digital Conversion↗
PubMed2005

Effect of crocin on experimental atherosclerosis in quails and its mechanisms.

In the present study, we examined the prophylaxis effect of crocin on experimental atherosclerosis and its possible mechanisms. The atherosclerosis formation was induced by hyperlipidamic diet in quails. At the 9th week, serum lipid, MDA and NO were measured, and HE staining was used to investigate the histopathological changes of aorta. Bovine aortic endothelial cells (EC) were obtained from the thoracic aorta of newborn calves. After incubation of the cells with Ox-LDL (50 mg x L(-1)) for 24 h, the activities of LDH, NO in culture media and activity of NOS in endothelial cells were measured, flow cytometer was used to determine the rate of endothelial cells apoptosis. Peritoneal macrophages were obtained from thioglycolate-injected mice. Cholesterol and free cholesterol in cells were assayed after incubation of the cells with Ox-LDL. Bovine aortic smooth muscle cells (SMC) were obtained from the thoracic aorta of newborn calf. Proliferation was induced by 100 microg x L(-1) Ox-LDL and antiproliferative effect of crocin on SMCs were observed. SMCs cycle phases were measured by flow cytometry. SMCs were loaded with Fluo-3/AM and [Ca2+]i was measured by Laser Scanning Confocal Microscope (LSCM). Crocin could reduce the level of serum TC, TG, LDL-C and inhibit the formation of aortic plaque. Crocin could reduce MDA and inhibit the descending of NO in serum. Compared with control, Ox-LDL group could increase the activity of LDH and decrease activity of NO in culture media and activity of NOS in endothelial cells, preincubated with crocin, the effects of Ox-LDL were inhibited. Crocin could decrease the EC apoptosis induced by Ox-LDL. Crocin concentration-dependently inhibited the TC and CE elevation induced by Ox-LDL in macrophages. Crocin could inhibit the proliferation of SMCs induced by Ox-LDL. In the presence or absence of extracellular Ca2+, crocin concentration-dependently inhibited the [Ca2+]i elevation induced by 120 mg x L(-1)Ox-LDL, In the absence of extracellular Ca2+, crocin could inhibit the [Ca2+]i elevation induced by CHCl3 in a concentration-dependent manner. The results indicated that crocin could inhibit the formation of atherosclerosis in quails. Crocin had protective effects on endothelial cells. Crocin could decrease CE in macrophages and uptake of Ox-LDL, inhibiting the formation of foam cell, which would promote the initiation and progression of atherosclerosis. Crocin could inhibit the [Ca2+]i elevation in smooth muscle cell, Ca2+ is an important second messenger that regulates a variety of cellular processes, including smooth muscle cell proliferation and gene expression . Crocin exerted antiatherosclerotic effects through decreasing the level of Ox-LDL that plays an important role in the initiation and progression of atherosclerosis.

Animals↗
PubMed2002

Analysis of poly(D,L-lactic-co-glycolic acid) nanosphere uptake by human dendritic cells and macrophages in vitro.

PURPOSE: The purpose of this study was to demonstrate and characterize phagocytosis of poly(D,L-lactic-co-glycolic acid) (PLGA) nanospheres by human dendritic cells (DCs). METHODS: Parallel cultures of DCs and macrophages (Mphi) were established from peripheral blood leukocytes using media supplemented with granulocyte-macrophage colony stimulator factor and interleukin-4 (for DC) or granulocyte-macrophage colony stimulator factor alone (for Mphi). PLGA nanospheres containing tetramethylrhodamine-labeled dextran with or without an adjuvant, monophosphoryl lipid A, were prepared using a water/oil/water solvent evaporation technique. Cells were incubated with the nanospheres for 24 h. Confocal laser scanning microscopy was used to determine the intracellular location of the nanospheres and flow cytometry to measure the fraction of phagocytic cells in the culture and the amount of uptake per cell. After phagocytosis, cells were stained for MHC class II molecules, CD14, CD80, and CD86 to identify the phagocytic population. RESULTS: DCs phagocytosed PLGA nanospheres as efficiently as Mphi. Cell-surface marker expression conclusively established that the phagocytic cells were DC. CONCLUSIONS: DCs can take up PLGA nanospheres. Because DCs are the key professional antigen-presenting cells capable of stimulating naive T cells, our data suggest that PLGA nanospheres can be used as an efficient delivery system for vaccines designed to activate T cell-mediated immune responses.

Cells, Cultured↗
PubMed2005

Epstein-Barr virus-encoded latent membrane protein 1 modulates cyclin D1 by c-Jun/Jun B heterodimers.

In our recent studies, we found that LMP1 encoded by Epstein-Barr virus could accelerate the formation of active c-Jun/Jun B heterodimer. We studied the regulation of cyclinD1 by c-Jun/Jun B heterodimers by laser scanning confocal influorescence microscopy, Western blot, luciferase activity assay, super-EMSA and flow cytometry in the Tet-on-LMP1 HNE2 cell line, in which LMP1 expression was regulated by Tet-on system. c-Jun/Jun B heterodimers induced by LMP1 could up regulate cyclin D1 promoter activity and expression. Overexpression of cyclin D1 accelerated the progression of cell cycle.

Cell Line, Tumor↗
PubMed2004

Effects of puerarin on number and activity of endothelial progenitor cells from peripheral blood.

AIM: To investigate whether puerarin can augment endothelial progenitor cells (EPCs) numbers, promote EPC proliferative, migratory, adhesive, and in vitro vasculogenesis capacity. METHODS: EPCs were characterized as adherent cells by double staining of DiLDL-uptake and lectin binding under a laser scanning confocal microscope. Expression of KDR, VEGFR-2, and AC133 was detected by flow cytometry. EPCs proliferation, migration and in vitro vasculogenesis were determined with MTT assay, modified Boyden chamber assay, and in vitro vasculogenesis kit, respectively. EPCs adhesive assay was performed by replating those on fibronectin-coated dishes, then adherent cells were counted. RESULTS: Incubation of isolated human MNCs with puerarin 0.1-3 mmol/L increased the number of EPCs, EPC proliferative, migratory, adhesive, and in vitro vasculogenesis capacity in a concentration- and time-dependent manner, which reached maximum at 3 mmol/L, 24 h (approximately 1-fold increase, P<0.01). CONCLUSION: Puerarin enhanced EPCs functional activity.

AC133 Antigen↗
PubMed2001

Selective activation of cervical microvascular endothelial cells by human papillomavirus 16-e7 oncoprotein.

BACKGROUND: Human papillomavirus type 16 (HPV16) is strongly implicated in the etiology of cervical cancer, with the expression of HPV16-encoded E7 oncoprotein in infected epithelial cells contributing to their malignant transformation. Although nuclear E7 interacts with several nuclear targets, we have previously shown that extracellular E7 can cause suppression of immune cell function. Moreover, cervical microvascular endothelial (CrMVEn) cells treated with E7 increase their expression of adhesion molecules. High levels of some cytokines in serum and in cervicovaginal secretions are associated with the progression of cervical cancer. In this study, we investigated the effects of extracellular E7 on cytokine production and on cytoskeleton structure of CrMVEn cells and vascular endothelial cells from different organs. METHODS: Immunocytochemical staining and flow cytometry techniques were used to detect E7 in endothelial cells incubated with purified E7 protein. Laser scanning confocal microscopy was used to study the E7-induced modification of the endothelial cytoskeleton. An enzyme-linked immunosorbent assay was performed to measure the production of two cytokines, interleukin 6 (IL-6) and interleukin 8 (IL-8), by E7-treated endothelial cells. All statistical tests were two-sided. RESULTS: Extracellular E7 was taken up by CrMVEn cells and localized to the cytoplasm. CrMVEn cells showed a statistically significant (P<.02) increase in the production of IL-6 and IL-8 after treatment with E7 compared with the controls. CrMVEn cells also produced higher levels of these cytokines than did the other endothelial cells (P<.01). E7 also induced marked alterations in the endothelial cytoskeleton of CrMVEn cells as a result of actin fiber polymerization. CONCLUSION: These findings suggest a novel mechanism by which E7, as an extracellular factor, can play a role in the progression and dissemination of cervical cancer via its selective effects on endothelial cells.

Cell Adhesion↗
PubMed2005

Reversal of MDR1 gene-dependent multidrug resistance using short hairpin RNA expression vectors.

BACKGROUND: RNA interference using short hairpin RNA (shRNA) can mediate sequence-specific inhibition of gene expression in mammalian cells. A vector-based approach for synthesizing shRNA has been developed recently. Overexpression of P-glycoprotein (P-gp), the MDR1 gene product, confers multidrug resistance (MDR) to cancer cells. In this study, we reversed MDR using shRNA expression vectors in a multidrug-resistant human breast cancer cell line (MCF-7/AdrR). METHODS: The two shRNA expression vectors were constructed and introduced into MCF-7/AdrR cells. Expression of MDR1 mRNA was assessed by RT-PCR, and P-gp expression was determined by Western Blot and immunocytochemistry. Apoptosis and sensitization of the breast cancer cells to doxorubicin were quantified by flow cytometry and methyl thiazolyl tetrazolium (MTT) assays, respectively. Cellular daunorubicin accumulation was assayed by laser confocal scanning microscopy (LCSM). Statistical significance of differences in mean values was evaluated by Student's t tests. P < 0.05 was considered statistically significant. RESULTS: In MCF-7/AdrA cells transfected with MDR1-A and MDR1-B shRNA expression vectors, RT-PCR showed that MDR1 mRNA expression was reduced by 40.9% (P < 0.05), 30.1% (P < 0.01) (transient transfection) and 37.6% (P < 0.05), 28.0% (P < 0.01) (stable transfection), respectively. Western Blot and immunocytochemistry showed that P-gp expression was significantly and specifically inhibited. Resistance against doxorubicin was decreased from 162-fold to 109-fold (P < 0.05), 54-fold (P < 0.01) (transient transfection) and to 108-fold (P < 0.05), 50-fold (P < 0.01) (stable transfection). Furthermore, shRNA vectors significantly enhanced the cellular daunorubicin accumulation. The combination of shRNA vectors and doxorubicin significantly induced apoptosis in MCF-7/AdrR cells. CONCLUSIONS: shRNA expression vectors effectively reduce MDR expression in a sustained fashion and can restore the sensitivity of drug-resistant cancer cells to conventional chemotherapeutic agents.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2006

Leukemic lymphoblasts, a novel expression site of coagulation factor XIII subunit A.

Blood coagulation factor XIII (FXIII) is a protransglutaminase circulating as a tetramer formed by two types of subunits (A2B2). The intracellular dimeric form of FXIII (A2) is present in platelets, megakaryocytes, monocytes and macrophages and has been detected in mono- and megakaryocytic leukemias. The aim of our study was to investigate FXIII-A expression in newly diagnosed B cell acute lymphoblastic leukemia (ALL) samples. We examined 47 de novo ALL cases of B cell origin by triple color labeling with flow cytometry. FXIII-A was detected by a FITC conjugated monoclonal antibody combined with CD34 and CD45 staining. In selected cases FXIII-A was investigated on slides prepared from blasts and visualized with a fluorescent microscope. In addition, blasts were studied by Western blot analysis and FXIII-A was measured by a highly sensitive ELISA method. By flow cytometry 19 samples of the 47 cases were found to be FXIII-A positive. Antigen concentration was 3.11 +/- 1.19 fg/blast, while normal lymphoid precursors and mature lymphocytes from B-CLL did not contain FXIII-A. In the lysate of lymphoblasts that were positive by flow cytometry, a single band (82 kDa) corresponding to FXIII-A was detected on Western blots. Confocal laser scanning microscopic examination revealed the presence of FXIII-A in the cytoplasm of these lymphoblasts. This novel expression site of FXIII-A in leukemic lymphoblasts can be utilized as a diagnostic tool and may also gain functional significance in B-lineage ALL.

Adolescent↗
PubMed2005

Functional co-localization of monocytic aminopeptidase N/CD13 with the Fc gamma receptors CD32 and CD64.

Information about the function of aminopeptidase N/CD13 on monocytes is limited. In order to gain more insight into its interaction with other proteins, we have identified molecules that co-localize with the membrane ectoenzyme at the cell surface of monocytes. Using laser scanning and electron microscopy as well as fluorescence resonance energy transfer (FRET) measured by flow cytometry we show that monocytic CD13 co-localized with the Fc gamma receptor II/CD32 after Fc receptor ligation by a CD32-specific antibody. FRET was also observed between CD13 and the Fc gamma receptor I/CD64, but not with the myeloid marker CD33 representing a member of the sialoadhesin family. Our results imply a novel functional role of CD13 and Fc gamma receptors as members of a multimeric receptor complex. Further studies have to be done to elucidate common signaling pathways of these molecules.

CD13 Antigens↗
PubMed2006

[Protective effect of adeno-associated viral vector-mediated expression of human brain-derived neurotrophic factor in rat neurons against beta-amyloid-induced Alzheimer's disease in vitro].

OBJECTIVE: To achieve expression of human brain-derived neurotrophic factor (hBDNF) mediated by recombinant adeno-associated virus (rAAV) and explore the mechanism of its neuroprotective effects in rat neurons against beta-amyloid-induced Alzheimer's disease. METHODS: Using molecular cloning technique, rAAV vector containing hBDNF gene (AAV-hBDNF) was constructed to transfect SD rat hippocampal neurons exposed to beta-amyloid treatment. The changes in cell apoptosis were observed by MTT assay and flow cytometry, and the expression of hBDNF and Bcl-2 protein were determined by immunocytochemical staining. Laser scanning confocal microscopy (LSCM) was used to observe the changes of [Ca(2+)](i). RESULTS: The cultured rat hippocampal neurons were effectively transfected with AAV-hBDNF and expression of BDNF protein was obviously increased. hBNDF expression showed significant protective effects against beta-amyloid-induced neuronal damage, and the expression of Bcl-2 protein was increased significantly and the balance of [Ca(2+)](i) was maintained in BDNF-treated cells with beta-amyloid exposure. CONCLUSION: hBDNF expression can effectively protect cultured rat hippocampal cells from beta-amyloid-induced apoptosis through inhibiting the intracellular calcium overload and increasing the expression of Bcl-2 protein.

Alzheimer Disease↗
PubMed2005

[An experimental study on destruction of K562 and HL60 induced by 5-aminolaevulinic acid-based photodynamic therapy].

This experiment was designed to explore the pattern of K562 and HL60 leukemia cells death, the effects on their cell cycle and the cytoplasmic free calcium concentration ([Ca2+]i) induced by 5-aminolaevulinic acid-based photodynamic therapy (ALA-PDT). Under the transmission electron microscope (TEM), two kinds of leukemia cells' ultrastructure were observed. Flow cytometry combined with Annexin V-FITC/PI labeling was used to detect the pattern of K562 and HL60 cells' death induced by ALA-PDT. Flow cytometry combined with PI labeling was used to analyze the change in the cell cycle induced by ALA-PDT, and confocal laser scanning microscopy (CLSM) combining with calcium fluorescence probe was used to detect the change in the cytoplasmic free calcium concentration ([Ca2+]i). Immediately after irradiation, many typical apoptotic bodies were seen in the cells treated. Most of the cells treated were necrotic at 24 hours following irradiation. Flow cytometry analysis suggested that the main patterns of the cells' death were apoptosis immediately after irradiation and necrosis post-apoptosis at 24 hours post irradiation. Immediately and 24 hours after irradiation, the proportion of S phase of K562 was 57. 67% +/- 1.13% and 84.77% +/- 6.20% respectively, and the proportion of S phase of HL60 was 74.60% +/- 7.27% and 84.60% 1.74% respectively. Both [Ca+]i of the treated K562 and HL60 were increased obviously. In the best experiment condition, the initial pattern of the K562 and HL60 leukemia cells' death induced by PDT was apoptosis and the main pattern was necrosis post apoptosis. The two kinds of cells were arrested at S phase by ALA-PDT. During the death of the leukemia cells, the increase in intracellular free calcium concentration could be responsible for the ALA photodynamically induced damage to K562 and HL60 cells.

Aminolevulinic Acid↗