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1.
The effects of synthetic polycation polyallylamine (PAA) on adhesion of CHL V-79 RJK fibroblasts and CHL V-79 RJK40 cells resistant to 40 degrees C, and attachment to these cells to polycation immobilized on polystyrene surface were studied. We have also investigated the cytotoxicity of PAA. It was shown that cell adherence to polystyrene plastic coated with PAA was enhanced or decreased in dependence of the PAA concentration used for surface coating and did not depend on heat resistance of investigated cell lines. The effect of PAA on cell adhesion to uncoated polystyrene surface after cell exposure with PAA depended not only on the polycation concentration, but also on the extent of heat resistance of investigated cell lines. Pretreatment of CHL V-79 RJK cells with PAA at the nontoxic concentrations led to inhibition of cell adhesion, and no change in adhesive properties of thermoresistant cells was found under the same conditions. PAA was toxic for CHL V-79 RJK and CHL V-79 RJK40 cells only at concentrations of 100 microg/ml (MTT assay). PAA-induced acute toxicity was accompanied by necrotic-like cell damage. Possible mechanisms of the PAA effect on the behaviour of cells with different structural and metabolic characteristics that are due to the temperature of cell cultivation are discussed.  相似文献   

2.
Karyotypic destabilization in cells of Chinese hamster fibroblasts CHL V-79 RJK with ectopically overexpressed antiapoptotical human bcl-2 gene from pSFFV-bcl-2 vector has been analysed. Analysis of G-banded metaphase chromosomes from 4 clones with different levels of bcl-2 expression revealed an increased level of chromosomal instability in bcl-2-transfected cells. Besides, an increased percentage of aneu- and polyploid cells and high level of cells with different chromosomal aberrations was observed. The degree of karyotypic instability positively correlated with the level of bcl-2 expression in bcl-2-transfected cells. Cells of a clone with the highest bcl-2 expression at the 13th passage of cultivation displayed an almost 100% polyploidization and the presence of specific aberrations and a tricentric marker chromosome. Selection of cells with non-random specific chromosome changes was observed in pSFFV-bcl-2-transfected CHL V-79 RJK cells in the process of their long-term cultivation. By contrast, cells of the parental cell line, as well as the control pSFFV-neo transfectants, displayed a stable karyotype throughout the long period of cultivation. It is important that the presence of morphological markers of gene amplifications--DOO, DM, MH--was observed in bcl-2-transfected cells. These findings suggest that the overexpression of antiapoptotic human bcl-2 gene may result in destabilization of the karyotype structure in cells of Chinese hamster fibroblasts CHL V-79 RJK. The character and level of destabilization correlate with the level of ectopic overexpression of this gene.  相似文献   

3.
Chinese hamster fibroblasts CHL V-79 RJK were subjected to multistep selection in the presence of etoposide, known as an inhibitor of topoisomerase II and inductor of apoptosis. The karyotype of cells stably resistant to etoposide was analysed at progressive stages of selection using G-type staining of metaphase chromosomes. Multiple changes in the karyotype of resistant cells were observed at an early stage of selection (0.2 mg/ml of etoposide) and included: random chromosome breaks leading to formation of new chromosome markers, high frequency of aneuploid and polyploid cells, morphological instability and extracopy of q-shoulder of chromosome 1. On advanced stages of selection we observed an increased frequency of specific minute chromosome and the appearance of chromosomes with homogeneously or differentially stained regions (HSR and DSR). These data suggest that different mechanisms may be involved in developing cellular resistance to etoposide at progressive stages of selection.  相似文献   

4.
In general, anchorage-dependent animal cells cultivated on a solid culture substrate, such as polystyrene, are collected by trypsin treatment. This treatment may have detrimental effects such as the proteolysis of the cell membrane proteins. To avoid these effects, cell cultivation using a liquid/liquid interface system has been investigated. In this cultivation method, the cells grow at the interface between a culture medium and a hydrophobic liquid. In this study, various fluorocarbons (FC-40, FC-70, KPF-91, KPF-102, and KPF-142) were used as substrates for the interface, and the cultivation of fibroblast cells (L-929; the mouse-derived cell line) at the interfaces was investigated. Early in the cultivation period, the growth of L-929 cells depended on the substrate type. Although cell cultivation at the interfaces was possible, it was slower than that at the polystyrene surface. Cell spreading at the interfaces was relatively small, which indicates that cell adhesion at the interfaces may be weak. In particular, the cells at the MEM/FC-70 interface anchored with one another and formed multicellular hemispherical aggregations shaped like spheroids. The difference in the adhesions to the interfaces appears to be dependent on the contaminants contained in the fluorocarbons because the physical properties of the fluorocarbon did not affect the cell growth and adhesion. Moreover, subcultivation from the interfaces to the same interface was possible without trypsin treatment. In this case, the delay of the growth at the interfaces did not occur because the cells were not affected by trypsin treatment.  相似文献   

5.
Cell-cell adhesion occurs when human erythrocytes and yeast cells are suspended together in suprathreshold concentrations of polylysine in saline. The threshold polycation concentration for adhesion depends on cell concentration and decreases with increasing polycation molecular weight. The threshold concentration was similar for erythrocyte-erythrocyte adhesion and for yeast-erythrocyte adhesion. Transmission electron micrographs show that the erythrocytes adhere to yeast as if to engulf the cell. The regions of close contact between the erythrocyte membrane and the yeast cell walls are spatially discrete. The contact separation distance for the asymmetric erythrocyte-yeast adhesion is very similar to that (0.83 μm) observed when polylysine-induced adhesion occurs in the symmetrical erythrocyte-erythrocyte system. The spacing is attributed to the growth of a squeezing wave as an interfacial instability, on the intercellular aqueous layer. Freeze-fracture electron microscopy of cells that were not fixed during preparation for microscopy confirms the discrete nature of contacts between polylysine treated erythrocytes.  相似文献   

6.
Cell-cell adhesion occurs when human erythrocytes and yeast cells are suspended together in suprathreshold concentrations of polylysine in saline. The threshold polycation concentration for adhesion depends on cell concentration and decreases with increasing polycation molecular weight. The threshold concentration was similar for erythrocyte-erythrocyte adhesion and for yeast-erythrocyte adhesion. Transmission electron micrographs show that the erythrocytes adhere to yeast as if to engulf the cell. The regions of close contact between the erythrocyte membrane and the yeast cell walls are spatially discrete. The contact separation distance for the asymmetric erythrocyte-yeast adhesion is very similar to that (0.83 μm) observed when polylysine-induced adhesion occurs in the symmetrical erythrocyte-erythrocyte system. The spacing is attributed to the growth of a squeezing wave as an interfacial instability, on the intercellular aqueous layer. Freeze-fracture electron microscopy of cells that were not fixed during preparation for microscopy confirms the discrete nature of contacts between polylysine treated erythrocytes.  相似文献   

7.
Cell-cell adhesion occurs when human erythrocytes and yeast cells are suspended together in suprathreshold concentrations of polylysine in saline. The threshold polycation concentration for adhesion depends on cell concentration and decreases with increasing polycation molecular weight. The threshold concentration was similar for erythrocyte-erythrocyte adhesion and for yeast-erythrocyte adhesion. Transmission electron micrographs show that the erythrocytes adhere to yeast as if to engulf the cell. The regions of close contact between the erythrocyte membrane and the yeast cell walls are spatially discrete. The contact separation distance for the asymmetric erythrocyte-yeast adhesion is very similar to that (0.83 micron) observed when polylysine-induced adhesion occurs in the symmetrical erythrocyte-erythrocyte system. The spacing is attributed to the growth of a squeezing wave as an interfacial instability, on the intercellular aqueous layer. Freeze-fracture electron microscopy of cells that were not fixed during preparation for microscopy confirms the discrete nature of contacts between polylysine treated erythrocytes.  相似文献   

8.
F Laval 《Nucleic acids research》1994,22(23):4943-4946
The E. coli fpg gene encodes the formamido-pyrimidine-DNA-glycosylase (FPG protein) which specifically removes the formamido-pyrimidine and C8-oxoGuanine residues from gamma-irradiated DNA. The fpg gene was ligated in the psV2 vector and transfected into the Chinese hamster CHO and V-79 cells. The transfected cells expressed a formamido-pyrimidine-DNA-glycosylase activity 30 to 40-fold over the constitutive level. The resistance of CHO and V-79 cells to the lethal effect of gamma-rays was similar in control and transfected cells. Furthermore CHO cells expressing the fpg gene had the same resistance to the lethal effect of hydrogen peroxide as control cells. However, the sensitivity to the mutagenic effect of gamma-rays, measured as 6-thioguanine resistance, decreased both in CHO and V-79 transfected cells. Since the lethal effect of gamma-rays was not modified in cells overproducing the FPG protein, the results suggest that this protein protects the cells against the mutagenic lesions formed by ionizing radiations, and among them C8-oxoguanine.  相似文献   

9.
Adhesion of cells to polystyrene surfaces   总被引:7,自引:2,他引:5       下载免费PDF全文
《The Journal of cell biology》1983,97(5):1500-1506
The surface treatment of polystyrene, which is required to make polystyrene suitable for cell adhesion and spreading, was investigated. Examination of surfaces treated with sulfuric acid or various oxidizing agents using (a) x-ray photoelectron and attenuated total reflection spectroscopy and (b) measurement of surface carboxyl-, hydroxyl-, and sulfur-containing groups by various radiochemical methods showed that sulfuric acid produces an insignificant number of sulfonic acid groups on polystyrene. This technique together with various oxidation techniques that render surfaces suitable for cell culture generated high surface densities of hydroxyl groups. The importance of surface hydroxyl groups for the adhesion of baby hamster kidney cells or leukocytes was demonstrated by the inhibition of adhesion when these groups were blocked: blocking of carboxyl groups did not inhibit adhesion and may raise the adhesion of a surface. These results applied to cell adhesion in the presence and absence of serum. The relative unimportance of fibronectin for the adhesion and spreading of baby hamster kidney cells to hydroxyl-rich surfaces was concluded when cells spread on such surfaces after protein synthesis was inhibited with cycloheximide, fibronectin was removed by trypsinization, and trypsin activity was stopped with leupeptin.  相似文献   

10.
Previous studies suggested that Chinese hamster V79 cells possess two mechanisms for their mutual adhesion, Ca2+-dependent and Ca2+-independent ones. We could prepare cells with only the Ca2+-dependent mechanism intact by dispersing cell monolayers with trypsin (0.01%) containing Ca2+. In the present study, we found that cells dispersed with a very low concentration of trypsin (0.0001%) in the absence of Ca2+ retain only the Ca2+-independent mechanism intact. Fab fragments of antibodies directed against surface antigens of V79 cells inhibited the aggregation of V79 cells by the Ca2+-independent mechanism, but did not inhibit the aggregation of these cells by the Ca2+-dependent mechanism. These results suggest that the two mechanisms of cell adhesion are based on different cellular components. Molecules responsible for the Ca2+-independent adhesion mechanism are probably cell surface components, because they were released from cells by the treatment with 0.01% trypsin without losing their specific antigenicity. The presence of adhesion mechanisms similar to those in V79 cells was shown in neural retinal cells of chick embryos. It was assumed, therefore, that these mechanisms of cell adhesion are generally present among a variety of cell types.  相似文献   

11.
Results of karyological analysis of cells CHL V-79 RJK selected for resistance to ethidium bromide (EB) causing multidrug resistance (MDR) (line Vebr-5) were compared with the data of microfluorimetric determination of DNA content in individual chromosomes of the karyotype. The analysis was performed at the 11th and 88th passages. Karyotyping of Vebr-5 has shown the presence of an additional genetic material (ADM) in the form of homogenously or differentially stained regions (HSRs and DSRs, respectively) in two chromosomes (Z1 and Z6, loci 1 p29-31 and 1q26, respectively). HSRs in Z6, in the region of localization of the wild type of gene mdr, had unstable length and structure characteristic of morphological markers of amplification of genes of the family mdr. During long cultivation of Vebr-5 in the presence of EB (88 passages), the instability of HSRs in Z6 increased. Results of microfluorimetric analysis of Vebr-5 at the 11th passage have shown an increase in the DNA content not only in chromosomes Z1 and Z6 marked by HSRs, but also in three chromosomes (Z5, Z12 and Z13) that have no visual morphological changes. The corresponding analysis at the 88th passage has also revealed non-random changes in the DNA content in four more chromosomes: an increase in Z14, while a decrease in chromosomes 8, Z7, and Z9. A decrease of the DNA content in chromosomes is considered to be a result of a partial loss of genetic material, while its increase is a result of its translocation and (or) amplification. Coefficient of variation of the DNA content changes for large chromosomes amounted to about 9%. while for small chromosomes it is about 26%, which indicates that small chromosomes have greater potential for instability than the large ones. The data obtained not only confirm, but also enlarge the concept of directions and character of destabilization of the cell genetic apparatus in the process of neoplastic transformation due to the MDR acquisition by cells.  相似文献   

12.
Integration of living cells with novel microdevices requires the development of innovative technologies for manipulating cells. Chemical surface patterning has been proven as an effective method to control the attachment and growth of diverse cell populations. Patterning polyelectrolyte multilayers through the combination of layer‐by‐layer self‐assembly technique and photolithography offer a simple, versatile, and silicon compatible approach that overcomes chemical surface patterning limitations, such as short‐term stability and low‐protein adsorption resistance. In this study, direct photolithographic patterning of two types of multilayers, PAA (poly acrylic acid)/PAAm (poly acryl amide) and PAA/PAH (poly allyl amine hydrochloride), were developed to pattern mammalian neuronal, skeletal, and cardiac muscle cells. For all studied cell types, PAA/PAAm multilayers behaved as a cytophobic surface, completely preventing cell attachment. In contrast, PAA/PAH multilayers have shown a cell‐selective behavior, promoting the attachment and growth of neuronal cells (embryonic rat hippocampal and NG108‐15 cells) to a greater extent, while providing little attachment for neonatal rat cardiac and skeletal muscle cells (C2C12 cell line). PAA/PAAm multilayer cellular patterns have also shown a remarkable protein adsorption resistance. Protein adsorption protocols commonly used for surface treatment in cell culture did not compromise the cell attachment inhibiting feature of the PAA/PAAm multilayer patterns. The combination of polyelectrolyte multilayer patterns with different adsorbed proteins could expand the applicability of this technology to cell types that require specific proteins either on the surface or in the medium for attachment or differentiation, and could not be patterned using the traditional methods. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009  相似文献   

13.
Results of a karyological analysis of cells CHL V-79 RJK selected for their resistance to ethidium bromide (EB) causing multidrug resistance (MDR) (subline Vebr-5) were compared with data from the microfluorimetric determination of the DNA content in individual chromosomes of the karyotype. The analysis was performed at the 11th and 88th passages. Karyotyping of Vebr-5 has shown the presence of an additional genetic material (AGM) in the form of homogenously or differentially stained regions (HSR or DSR, respectively) in two chromosomes (Z1 and Z6, loci 1p29-31 and 1q26, respectively). HSR in Z6, in the site of localization of the mdr gene of the wild type had unstable length and structure, which is characteristic of the morphological markers of the amplification of genes of the mdr family. During the long cultivation of Vebr-5 in the presence of EB (88 passages), the instability of HSR (DSR) in Z6 increased. Results of a microfluorimetric analysis of Vebr-5 at the 11th passage have shown a statistically significant increase of the DNA content not only in chromosomes Z1 and Z6 marked by HSR (DSR), but also in three chromosomes (Z5, Z12, and Z13) that have no visual morphological changes. The corresponding analysis at the 88th passage has also revealed nonrandom changes in the DNA content in four more chromosomes, including an increase in Z14 and a decrease in chromosomes 8, Z7, and Z9. A decrease in the DNA content in chromosomes is considered to be the result of a partial loss of genetic material, while its increase is considered to be the result of its translocation and/or amplification. While the coefficient of the variation of the DNA content changes about 9% for large chromosomes, it amounted to about 26% for the small ones, indicating that small chromosomes to have a greater potential for instability than large chromosomes. The obtained data not only confirm, but also enlarge, the concept of the directions and character of the destabilization of the cell genetic apparatus in the process of neoplastic transformation due to MDR acquisition by these cells.  相似文献   

14.
The form of contact seam (whether a continuous parallel seam or membranes in spatially periodic contact) has been characterized for normal and for neuraminidase pretreated human erythrocytes following adhesion in solutions of polylysine in the molecular mass range 10–225 kDa at concentrations from 0.5 to 1.0 mg/mL. The adhesion contact seam was spatially periodic for all normal control cells in polylysine. The lateral separation of contacts decreased from 1.6 to 0.8 μm as the concentration of 225 kDa polylysine was increased threefold from the adhesion threshold value. The separation distance did not change further even at high polymer concentrations that increased the electrophoretic velocity to positive values over twice the modulus of the velocity of control cells. The probability of cell adhesion decreased at these high polymer concentrations. The lateral contact separation increased and cell adhesion decreased for cells pretreated with neuraminidase. Cell adhesion did not occur when neuraminidase reduced the cell electrophoretic velocity modulus by 30%. Following neuraminidase pretreatments that allowed a small amount of adhesion, the cell contact seam was continuous rather than spatially peridic. The results show that a procedure that increases (e.g., polymer concentration increase) or decreases (e.g., enzyme removal of polycation crosslinking site) attraction leads to shorter (to a limiting value) or longer lateral contact separation, respectively.  相似文献   

15.
The physicochemical forces that mediate attachment of yeasts to the phylloplane are unknown. Cell surface charge and hydrophobicity and adhesion to polystyrene, glass, and barley were assessed for wild-type Rhodosporidium toruloides and attachment-minus (Att) mutants. Cells were grown under conditions promoting (excess carbon) or not promoting (excess nitrogen) capsule production. Hydrophobicity was measured by adhesion to xylenes, and surface charge characteristics were assessed by attachment to either DEAE (positive)- or carboxymethyl (CM) (negative)-Sephadex ion-exchange beads. Hydrophobicity and adhesiveness of nonencapsulated, wild-type R. toruloides decreased from mid-log to late stationary phase. Encapsulated wild-type R. toruloides cells were more hydrophobic and more adhesive than nonencapsulated cells. However, two encapsulated Att mutants were more hydrophobic than the wild type and levels of adhesion of R. toruloides were similar on polystyrene and less hydrophobic glass surfaces. Adhesion of wild-type yeast to barley and polystyrene was correlated with attachment to CM-Sephadex beads, indicating a positive cell surface charge. Sixteen Att mutants did not exhibit a positive cell surface charge, and wild-type yeast cells that did not attach to CM-Sephadex did not adhere to either polystyrene or barley. Wild-type R. toruloides attached to CM-Sephadex beads by the poles of the cells, indicating a localization of positive charge which was also visualized with India ink. We conclude that localized, positive charge, and not hydrophobic interactions, mediates attachment of R. toruloides to barley leaves.  相似文献   

16.
The effect of collagen tripeptide fragment GER on the adhesion and spreading of mouse embryonic fibroblasts STO to different substrates (polystyrene plastic, poly-L-lysine, fibronectin, gelatin) has been studied. It was found that tripeptide GER was involved in fibroblast adhesion and spreading. The cell response depended both on the mode of tripeptide addition to culture medium and the substrate type. Coincubation of fibroblasts with tripeptide stimulated the cell attachment and spreading to untreated plastic and plastic coated with fibronectin or gelatin but did not change cell adhesion to immobilized poly-L-lysine. Preincubation of cells with tripeptide resulted in partial inhibition of fibroblast adhesion and spreading on fibronectin- and gelatin-coated substrata. It was shown that activation and inhibition of adhesive processes after tripeptide treating was higher on fibronectin than gelatin. The data obtained support the assumption about concerted action of tripeptide GER (activity was dependent both on the used concentration of the tripeptide and the mode of tripeptide addition to culture medium) and chemical characteristics of substrate (polymers of styrene and L-lysine, ECM proteins in native (fibronectin) or partly denatured (gelatin) form) on the cell adhesion and spreading. The main targets that GER peptide may affect during the formation of cell-substrate interactions are discussed.  相似文献   

17.
The form of contact seam (whether a continuous parallel seam or membranes in spatially periodic contact) has been characterized for normal and for neuraminidase pretreated human erythrocytes following adhesion in solutions of polylysine in the molecular mass range 10-225 kDa at concentrations from 0.5 to 1.0 mg/mL. The adhesion contact seam was spatially periodic for all normal control cells in polylysine. The lateral separation of contacts decreased from 1.6 to 0.8 microns as the concentration of 225 kDa polylysine was increased threefold from the adhesion threshold value. The separation distance did not change further even at high polymer concentrations that increased the electrophoretic velocity to positive values over twice the modulus of the velocity of control cells. The probability of cell adhesion decreased at these high polymer concentrations. The lateral contact separation increased and cell adhesion decreased for cells pretreated with neuraminidase. Cell adhesion did not occur when neuraminidase reduced the cell electrophoretic velocity modulus by 30%. Following neuraminidase pretreatments that allowed a small amount of adhesion, the cell contact seam was continuous rather than spatially periodic. The results show that a procedure that increases (e.g., polymer concentration increase) or decreases (e.g., enzyme removal of polycation crosslinking site) attraction leads to shorter (to a limiting value) or longer lateral contact separation, respectively.  相似文献   

18.
Effects of antineoplastic prostaglandins (PG), PGD2 and 9-deoxy-delta 9-PGD2, on colony formation of cloned Dunn osteosarcoma (TA 102), normal Swiss 3T3 and V-79 cell lines were evaluated. PGD2 significantly inhibited the colony formation of TA 102 cells in a dose-dependent manner at concentrations between 0.5 and 5 micrograms/ml. The IC50 value was calculated to be 0.72 microgram/ml. A dose-dependent inhibition of TA 102 colony formation was also observed with 9-deoxy-delta 9-PGD2 between 0.01 to 1 microgram/ml, the IC50 value being 0.22 microgram/ml. These prostaglandins did not exert cytocidal effects in vitro on Swiss 3T3 cells at concentrations between 0.01 to 1 microgram/ml. The two agents had no significant cytocidal effects on V-79 cells except for 9-deoxy-delta 9-PGD2 at a concentration of 5 ug/ml. These results suggest that PGD2 and 9-deoxy-delta 9-PGD2 are considered to have cytocidal activity on Dunn osteosarcoma cells in dosages which do not affect non-malignant cells.  相似文献   

19.
The adhesion of embryonic chicken retinal cells and mouse N2A neuroblastoma cells to purified embryonic chicken retinal NCAM adsorbed on a solid substratum was examined using a quantitative centrifugal adhesion assay. Both cell types adhered to NCAM and the adhesion was specifically inhibited by monovalent anti-NCAM antibody fragments. N2A cell adhesion depended on the amount of NCAM applied to the substratum, was cation independent, and was insensitive to treatment with the cytoskeletal perturbing drugs colchicine and cytochalasin D. These results indicated that the tubulin and actin cytoskeletons were not critically required for adhesion to NCAM and make it unlikely that the cell surface ligand for NCAM is an integrin. Adhesion was however temperature dependent, strengthening greatly after a brief incubation at 37 degrees C. CHO cells transfected with NCAM cDNAs did not adhere specifically to substratum-bound NCAM and pretreatment of N2A cells and retinal cells with anti-NCAM antibodies did not inhibit adhesion to substratum-bound NCAM. These results suggest that a heterophilic interaction between substratum-adsorbed NCAM and a non-NCAM ligand on the surface of the probe cells affects adhesion in this system and support the possibility that heterophilic adhesion may be a function of NCAM in vivo.  相似文献   

20.
A decreased sensitivity of the Chinese hamster cells (line V-79) to gamma-radiation under the influence of nitric oxide induction was shown elsewhere. This effect is connected hypothetically with post-radiation reparation of DNA. The investigation of the nitric oxide donor effect on sensitivity of these to UV-radiation is of interest, because this radiation is an important ecological factor of the environment. The question of retention of nitric oxide positive effect on UV and gamma-radiation sensitivity in malignant HeLa cells is no less actual, because these cells significantly differ from normal cells of line V-79. We demonstrated that the donor of nitric oxide enhances stability of the Chinese hamster cells (line V-79) to UV-radiation, as well as to gamma-radiation independently of the time of cell incubation with sodium oxide donor before or after irradiation. The inefficiency of nitric oxide as a factor increasing UV-stability of cells was shown for malignant HeLa cells. A 1 h long incubation of these cells with NO-donor before gamma-irradiation decreased the number of chromosome aberrations, and conversely, the addition of this agent to the HeLa cell culture after gamma-irradiation did not change the radiostability. It may be inferred that distinctions in behaviour of nitric oxide in cultures of V-79 and HeLa cells using UV-radiation may be explained by transformation of the latter special features of their damage, and by the following reparation.  相似文献   

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