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Results for “Extracellular Traps”

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At least 703 records · Page 39Linked to original sources

PubMed2000

Spin-trapping detection of superoxides in polymorphonuclear leukocytes stimulated with serum-opsonized zymosan.

To clarify where oxygen radicals are generated in polymorphonuclear leukocytes (PMNs) during phagocytosis, superoxides (O2-) from opsonized symosan (OZ)--stimulated human PMNs were detected by the ESR and spin-trapping methods. PMNs were preactivated with OZ for the indicated periods of time at 37 degrees C. Then a spin-trapping agent, 5, 5-dimethyl-1-pyrroline N-oxide (DMPO), was added to them, and they were further incubated for 30 sec for ESR observations. The ESR spectra consisted of two components due to the DMPO-OOH and DMPO-OH spin adducts. To clarify where these spin-adducts were present, cells were separated from extracellular fluid by brief centrifugation and resuspended in Hanks' balanced salt solution. ESR examination of two fractions showed that the DMPO-OOH adducts was present in the cell fraction, whereas the DMPO-OH adducts were present in the extracellular fluid. When DMSO was used as a scavenger of hydroxyl radicals (.OH), DMPO-CH3 adducts were observed in the fluid fraction but not in the cell fraction. Both spin adducts were completely abolished by Cu, Zn-SOD but not catalase. These results indicated that O2- were produced inside phagosomes of OZ-stimulated PMNs and .OH were produced outside them by spontaneous decomposition of the DMPO-OOH adducts.

Electron Spin Resonance Spectroscopy↗

PubMed2002

[Regulative function of extracelluar regulated protein kinases and telomerase in apoptosis of hepatocarcinomatous cell SMMC-7721].

OBJECTIVE: To study the changes of telomerase activity and protein expression of phosphorylated (activated) extracellular regulated protein kinases (ERK1 and ERK2) in the course of inhibiting hepatocarcinomatous cell proliferation and inducing cell apoptosis by three kinds of chemotherapy drugs: Harringtonine (HRT), Vincristine (VCR), and Etoposide (Vp16). To discuss the regulative function to hepatocarcinomatous cell apoptosis and interrelation of telomerase and ERK. METHODS: Cytotoxicity assay, flow cytometry analysis, telomerase repeat amplification protocol assay (TRAP), bioluminescence analysis, and western blot were used in this experiment. RESULTS: HRT, VCR, and Vp16 could inhibit cell proliferation (0.28% 0.08%, 0.25% 0.16%, 0.24% 0.11%), induce apoptosis (21.12%, 28.83%, 12.30%), inhibit telomerase activity, and down-regulate the protein expression of phosphorylated ERK. CONCLUSIONS: It might be through ERK signal transduction pathways that chemotherapy drugs down-regulate telomerase activity and induce apoptosis.

Apoptosis↗

PubMed2002

RACK1, an insulin-like growth factor I (IGF-I) receptor-interacting protein, modulates IGF-I-dependent integrin signaling and promotes cell spreading and contact with extracellular matrix.

The insulin-like growth factor I (IGF-I) receptor (IGF-IR) is known to regulate a variety of cellular processes including cell proliferation, cell survival, cell differentiation, and cell transformation. IRS-1 and Shc, substrates of the IGF-IR, are known to mediate IGF-IR signaling pathways such as those of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K), which are believed to play important roles in some of the IGF-IR-dependent biological functions. We used the cytoplasmic domain of IGF-IR in a yeast two-hybrid interaction trap to identify IGF-IR-interacting molecules that may potentially mediate IGF-IR-regulated functions. We identified RACK1, a WD repeat family member and a Gbeta homologue, and demonstrated that RACK1 interacts with the IGF-IR but not with the closely related insulin receptor (IR). In several types of mammalian cells, RACK1 interacted with IGF-IR, protein kinase C, and beta1 integrin in response to IGF-I and phorbol 12-myristate 13-acetate stimulation. Whereas most of RACK1 resides in the cytoskeletal compartment of the cytoplasm, transformation of fibroblasts and epithelial cells by v-Src, oncogenic IR or oncogenic IGF-IR, but not by Ros or Ras, resulted in a significantly increased association of RACK1 with the membrane. We examined the role of RACK1 in IGF-IR-mediated functions by stably overexpressing RACK1 in NIH 3T3 cells that expressed an elevated level of IGF-IR. RACK1 overexpression resulted in reduced IGF-I-induced cell growth in both anchorage-dependent and anchorage-independent conditions. Overexpression of RACK1 also led to enhanced cell spreading, increased stress fibers, and increased focal adhesions, which were accompanied by increased tyrosine phosphorylation of focal adhesion kinase and paxillin. While IGF-I-induced activation of IRS-1, Shc, PI3K, and MAPK pathways was unaffected, IGF-I-inducible beta1 integrin-associated kinase activity and association of Crk with p130(CAS) were significantly inhibited by RACK1 overexpression. In RACK1-overexpressing cells, delayed cell cycle progression in G(1) or G(1)/S was correlated with retinoblastoma protein hypophophorylation, increased levels of p21(Cip1/WAF1) and p27(Kip1), and reduced IGF-I-inducible Cdk2 activity. Reduction of RACK1 protein expression by antisense oligonucleotides prevented cell spreading and suppressed IGF-I-dependent monolayer growth. Our data suggest that RACK1 is a novel IGF-IR signaling molecule that functions as a positive mediator of cell spreading and contact with extracellular matrix, possibly through a novel IGF-IR signaling pathway involving integrin and focal adhesion signaling molecules.

Animals↗

PubMed1997

Effects of calmodulin antagonists on calcium pump and cytosolic calcium level in human neutrophils.

The concentration of cytosolic free calcium was monitored in suspensions of intact human neutrophils in phosphate-buffered saline by means of the fluorescent indicator Indo 1 trapped in the cytosol. Trifluoperazine and n-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide markedly reduced the amplitude of the transient increase in cytosolic Ca2+ triggered by CaCl2 as well as by N-formyl-methionyl-leucyl-phenylalanine. The effect of the calmodulin antagonists on the calcium burst observed upon cell activation was much more pronounced in the presence of extracellular free calcium than in EGTA-containing media; it was not inhibited by wortmannin or thapsigargin. Nevertheless, trifluoperazine and n-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide inhibited the plasma-membrane Ca2+ ATPase if added to plasma membrane-enriched fractions of neutrophils. These results suggest that calmodulin antagonists affect calcium ion influx even if they inhibit plasma membrane Ca2+ ATPase.

Calcium↗

PubMed2000

Snapshots of usher-mediated protein secretion and ordered pilus assembly.

Type 1 pilus biogenesis was used as a paradigm to investigate ordered macromolecular assembly at the outer cell membrane. The ability of Gram-negative bacteria to secrete proteins across their outer membrane and to assemble adhesive macromolecular structures on their surface is a defining event in pathogenesis. We elucidated genetic, biochemical, and biophysical requirements for assembly of functional type 1 pili. We discovered that the minor pilus protein FimG plays a critical role in nucleating the formation of the adhesive tip fibrillum. Genetic methods were used to trap pilus subunits during their translocation through the outer membrane usher protein, providing data on the structural interactions that occur between subunit components during type 1 pilus formation. Electron microscopic and biochemical analyses of these stepwise assembly intermediates demonstrated that translocation of pilus subunits occurs linearly through the usher's central channel, with formation of the pilus helix occurring extracellularly. Specialized pilin subunits play unique roles both in this multimerization and in the final ultrastructure of the adhesive pilus.

Animals↗

PubMed2000

Aberrant trafficking of the B cell receptor Ig-alpha beta subunit in a B lymphoma cell line.

The B cell Ag receptor (BCR) has two important functions: first, it binds and takes up Ag for presentation to T lymphocytes; and second, it transmits signals that regulate B cell development. Normal expression of the BCR requires the association of the Ag binding subunit, membrane IgM (mIgM), with the signaling component, the Ig-alpha beta heterodimer. After assembly in the endoplasmic reticulum, the intact BCR travels through the secretory pathway to the cell surface. In this paper, we report two variants of the B lymphoma cell lines, WEHI 279 and WEHI 231, that have both lost the ability to express mu heavy chain and consequently do not express mIgM. However, these variants do express the Ig-alpha beta heterodimer. In one variant, WEHI 279*, the Ig-alpha beta remained trapped intracellularly in the absence of mIgM. The other variant, 303.1.5.LM, expressed an aberrantly glycosylated Ig-alpha beta on the cell surface that was capable of signaling after cross-linking with anti-Ig-beta Abs. Further characterization uncovered a point mutation in the 303.1.5.LM mb1 gene that would change a proline for a leucine in the extracellular domain of Ig-alpha. The 303.1.5.LM Ig-alpha beta could not associate with a wild-type mIgM after mu heavy chain was reconstituted by DNA transfection. Thus, this mutation could define a region of the Ig-alpha polypeptide that is important for recognition by the endoplasmic reticulum quality control system, for association with glycosylating enzymes, and for the association of Ig-alpha beta subunits with mIgM subunits to create a complete BCR complex.

Animals↗

PubMed1997

[Penetration of azithromycin into human neutrophils: effect of hydrogen peroxide production].

BACKGROUND: The aim of this study was to evaluate the uptake of azythromycin at therapeutic concentrations by human polymorphonuclear leukocytes (PMN), as well as the effect of environmental temperature, pH, and cell viability on this uptake. The effect of azythromycin and other macrolides on hydrogen peroxide production by PMN was also assessed. METHODS: Uptake of radiolabeled azythromycin by PMN was determined by a radiometric technique. Hydrogen peroxide production by PMN was measured by the phenol red method. RESULTS: The intracellular concentrations achieved by azythromycin in PMN were 20 to 60 fold the extracellular ones, even at extracellular concentration of 0.125 mg/l. The uptake was significantly affected by low temperature, acid pH, and cell viability. Hydrogen peroxide production was not affected by the macrolides studied. CONCLUSIONS: Azythromycin at therapeutical concentrations achieves high intracellular accumulation in human PMN. The effect of environmental temperature, pH and cell viability on uptake points out that a passive diffusion, with lysosome trapping, is the essential mechanism of azythromycin uptake by phagocytic cells. In spite of the high intracellular concentrations achieved by macrolides, hydrogen peroxide production by PMN is not affected by these compounds.

Anti-Bacterial Agents↗

PubMed2001

Lactate dehydrogenase release is facilitated by brief sonication of rat hippocampal slices and isolated retinas following acute neuronal damage.

Although useful for determining neuronal damage in cell cultures, the lactate dehydrogenase (LDH) assay is not suitable for acute brain preparations because LDH release is typically delayed relative to neuronal deterioration. The slow release suggests that LDH may remain trapped inside damaged cells until late in the degenerative process. To test this, we examined whether brief sonication facilitates LDH release from acutely damaged neurons. In rat isolated retinas and hippocampal slices, LDH release was minimal following acute administration of iodoacetate or kainate. However, these toxins promoted significant LDH release, when toxin exposure was followed by brief sonication. Increases in extracellular LDH correlated with changes in neuronal morphology. These findings suggest that sonication may facilitate the use of the LDH assay in acute brain preparations.

Acute Disease↗

PubMed1997

Induction of transcription from the long terminal repeat of the intracysternal particles type A (IAP) by X-irradiation.

Intracisternal A particles (IAPs) are retrovirus-like entities that are present in many embryonic and transformed cells of Mus musculus. They present long terminal repeats (LTRs) which control the promotion and regulation of their transcription. Using a construction expressing a reporter gene under the control of the entire long terminal repeat (LTR) of IAP in transfected murine fibroblast BALB/c 3T3 cells clone D152, we were able to show that the IAP-LTR is activated by X-irradiation in a time-dependent manner. The relative CAT activity increased with increasing X-irradiation doses, reaching a maximum at 75-150 cGy, followed by a drop in activation. In addition, X-induced D152 mouse cells produced extracellular factor(s), in response to X-irradiation, which activated the IAP-LTR in non-irradiated cells. This factor(s) was detected both when transfected cells were cocultured with inducing cells and when conditioned medium from irradiated cultures was added to the cell cultures. The use of suramin, a strong polyanonic molecule which has been reported to trap growth factors, induces a high reduction of the indirect activation.

3T3 Cells↗

PubMed1993

Extracellular production of singlet oxygen by stimulated macrophages quantified using 9,10-diphenylanthracene and perylene in a polystyrene film.

The extracellular production of singlet oxygen (O2(1 delta g)) by stimulated macrophages was measured using a modification of our quantitative method initially developed to measure the intracellular production of O2(1 delta g) by neutrophils (Steinbeck, M. J., Khan, A. U., and Karnovsky, M. J. (1992) J. Biol. Chem. 267, 13425-13433). Glass coverslips were coated with the specific chemical trap for O2(1 delta g), 9,10-diphenylanthracene (DPA) and perylene, which is an internal standard, in a methylene chloride solution containing 0.3 mg/ml polystyrene. On evaporation, the polystyrene formed an even coating of DPA and perylene over the surface of a glass coverslip (PDP film). Unstimulated macrophages or macrophages stimulated with 4 beta-phorbol 12-myristate 13-acetate (PMA) or formyl-methionyl-leucyl-phenylalanine (fMLP) were then added to the PDP film in a darkened room and incubated at 37 degrees C for 30 min in a humidified 5% CO2 atmosphere. Both unstimulated and stimulated cells adhered to the PDP film in approximately equivalent numbers. Only stimulated cells produced measurable amounts of O2(1 delta g) in a dose-dependent response to either PMA or fMLP. The production of O2(1 delta g) by macrophages stimulated with PMA was maximal in response to 25 ng, 17.8 +/- 1.3 nmol of O2(1 delta g)/approximately 1.00 x 10(6) cells. The maximal response for fMLP was at a concentration of 1 microM, 18.4 +/- 1.0 nmol of O2(1 delta g)/approximately 1.00 x 10(6) cells. The specific detection of O2(1 delta g) by this method was confirmed by thermally releasing O2(1 delta g) from the DPA-O2(1 delta g) reaction product, DPA-endoperoxide, regenerating the original DPA compound. Production of O2(1 delta g) by the stimulated cells was inhibited 80-89% by the addition of 60-120 micrograms of superoxide dismutase, an enzyme that converts superoxide to hydrogen peroxide and ground state molecular oxygen or 79-84% with the addition of 2 mM histidine, an avid quencher of O2(1 delta g). Neither of these additions interfered with adhesion of the cells to the PDP film. The ability of superoxide dismutase to inhibit the production of O2(1 delta g) suggested that O2(1 delta g) was produced via a superoxide-dependent route. The ability of an oxidase to produce O2(1 delta g) secondary to superoxide production was substantiated further using a xanthine oxidase-acetaldehyde system. Purified xanthine oxidase produced both superoxide and O2(1 delta g), and their production was inhibited by the addition of superoxide dismutase.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1998

The role of laminins in basement membrane function.

Laminins are a family of multifunctional macromolecules, ubiquitous in basement membranes, and represent the most abundant structural noncollagenous glycoproteins of these highly specialised extracellular matrices. Their discovery started with the difficult task of isolating molecules produced by cultivated cells or extracted from tissues. The development of molecular biology techniques has facilitated and accelerated the identification and the characterisation of new laminin variants making it feasible to identify full-length polypeptides which have not been purified. Further, genetically engineered laminin fragments can be generated for studies of their structure-function relationship, permitting the demonstration that laminins are involved in multiple interactions with themselves, with other components of the basal lamina, and with cells. It endows laminins with a central role in the formation, the architecture, and the stability of basement membranes. In addition, laminins may both separate and connect different tissues, i.e. the parenchymal and the interstitial connective tissues. Laminins also provide adjacent cells with a mechanical scaffold and biological information either directly by interacting with cell surface components, or indirectly by trapping growth factors. In doing so they trigger and control cellular functions. Recently, the structural and biological diversity of the laminins has started to be elucidated by gene targeting and by the identification of laminin defects in acquired or inherited human diseases. The consequent phenotypes highlight the pivotal role of laminins in determining heterogeneity in basement membrane functions.

Animals↗

PubMed1999

Keratocytes pull with similar forces on their dorsal and ventral surfaces.

As cells move forward, they pull rearward against extracellular matrices (ECMs), exerting traction forces. However, no rearward forces have been seen in the fish keratocyte. To address this discrepancy, we have measured the propulsive forces generated by the keratocyte lamella on both the ventral and the dorsal surfaces. On the ventral surface, a micromachined device revealed that traction forces were small and rearward directed under the lamella, changed direction in front of the nucleus, and became larger under the cell body. On the dorsal surface of the lamella, an optical gradient trap measured rearward forces generated against fibronectin-coated beads. The retrograde force exerted by the cell on the bead increased in the thickened region of the lamella where myosin condensation has been observed (Svitkina, T.M., A.B. Verkhovsky, K.M. McQuade, and G. G. Borisy. 1997. J. Cell Biol. 139:397-415). Similar forces were generated on both the ventral (0.2 nN/microm(2)) and the dorsal (0.4 nN/microm(2)) surfaces of the lamella, suggesting that dorsal matrix contacts are as effectively linked to the force-generating cytoskeleton as ventral contacts. The correlation between the level of traction force and the density of myosin suggests a model for keratocyte movement in which myosin condensation in the perinuclear region generates rearward forces in the lamella and forward forces in the cell rear.

Actins↗

PubMed2001

Nitric oxide enhances MPP(+)-induced hydroxyl radical generation via depolarization activated nitric oxide synthase in rat striatum.

We examined the effect of N(G)-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase (NOS) inhibitor, on extracellular potassium ion concentration ([K(+)](o))-enhanced hydroxyl radical (.OH) generation due to 1-methyl-4-phenylpyridinium ion (MPP(+)) was examined in the rat striatum. Rats were anesthetized, and sodium salicylate in Ringer's solution (0.5 nmol/microl per min) was infused through a microdialysis probe to detect the generation of.OH as reflected by the non-enzymatic formation of 2,3-dihydroxybenzoic acid (DHBA) in the striatum. Induction of KCl (20, 70 and 140 mM) increased MPP(+)-induced.OH formation trapped as 2,3-dihydroxybenzoic acid (DHBA) in a concentration dependent manner. However, the application of L-NAME (5 mg/kg i.v.) abolished the [K(+)](o) depolarization-induced.OH formation with MPP(+). Dopamine (DA; 10 microM) also increased the levels of DHBA due to MPP(+). However, the effect of DA after application of L-NAME did not change the levels of DHBA. On the other hand, the application of allopurinol (20 mg/kg i.v., 30 min prior to study), a xanthine oxidase (XO) inhibitor was abolished the both [K(+)](o)- and DA-induced.OH generation. Moreover, when iron(II) was administered to MPP(+) then [K(+)](o) (70 mM)-pretreated animals, a marked increase in the level of DHBA. However, when corresponding experiments were performed with L-NAME-pretreated animals, the same results were obtained. Therefore, NOS activation may be no relation to Fenton-type reaction via [K(+)](o) depolarization-induced.OH generation. The present results suggest that [K(+)](o)-induced depolarization augmented MPP(+)-induced.OH formation by enhancing NO synthesis.

1-Methyl-4-phenylpyridinium↗

PubMed2005

A proteomic approach to characterize protein shedding.

Shedding (i.e. proteolysis of ectodomains of membrane proteins) plays an important pathophysiological role. In order to study the feasibility of identifying shed proteins, we analyzed serum-free media of human mammary epithelial cells by mass spectrometry following induction of shedding by the phorbol ester, 4 beta-phorbol 12-myristate 13-acetate (PMA). Different means of sample preparation, including biotinylation of cell surface proteins, isolation of glycosylated proteins, and preparation of crude protein fractions, were carried out to develop the optimal method of sample processing. The collected proteins were digested with trypsin and analyzed by reversed-phase capillary liquid chromatography interfaced to an ion-trap mass spectrometer. The resulting peptide spectra were interpreted using the program SEQUEST. Analyzing the sample containing the crude protein mixture without chemical modification or separation resulted in the greatest number of identifications, including putatively shed proteins. Overall, 45 membrane-associated proteins were identified including 22 that contain at least one transmembrane domain and 23 that indirectly associate with the extracellular surface of the plasma membrane. Of the 22 transmembrane proteins, 18 were identified by extracellular peptides providing strong evidence they originate from regulated proteolysis or shedding processes. We combined results from the different experiments and used a peptide count method to estimate changes in protein abundance. Using this approach, we identified two proteins, syndecan-4 and hepatoma-derived growth factor, whose abundances increased in media of cells treated with PMA. We also detected proteins whose abundances decreased after PMA treatment such as 78 kDa glucose-regulated protein and lactate dehydrogenase A. Further analysis using immunoblotting validated the abundance changes for syndecan-4 and 78 kDa glucose-regulated protein as a result of PMA treatment. These results demonstrate that tandem mass spectrometry can be used to identify shed proteins and to estimate changes in protein abundance.

Biotinylation↗

PubMed1988

Characterization of Na+/H+ exchange in a rabbit corneal epithelial cell line (SIRC).

Continuous intracellular pH (pHi) measurements were performed in SIRC rabbit corneal epithelial cells using the pH-sensitive absorbance of intracellularly trapped 5(and 6)-carboxy-4',5'-dimethylfluorescein. Steady-state pHi in nominally bicarbonate free Ringer's solution averaged 6.87 +/- 0.02 (mean +/- S.E., n = 53). After intracellular acidification induced by the NH4Cl-prepulse technique, there was a sodium-dependent pHi recovery towards the normal steady-state pHi. The initial pHi recovery rate was a saturable function of extracellular sodium concentration with an apparent Km for external sodium of about 25 mM and a Vmax of about 0.28 pH units/min. Virtually no pHi recovery was observed in the absence of extracellular sodium. Sodium removal during steady state acidified the cells by 0.36 +/- 0.05 pH units (mean +/- S.E., n = 13) within 5 min. There was a dose-dependent inhibition of pHi recovery after NH4Cl prepulse by amiloride with an IC50 of about 15 microM. Amiloride in a concentration of 1 mM almost completely abolished pHi recovery. Amiloride (1 mM) applied during steady state induced an intracellular acidification of 0.2 +/- 0.03 pH units (mean +/- S.E., n = 7) within 5 min. These findings suggest that a Na+/H+ exchange is present in SIRC rabbit corneal epithelial cells. Na+/H+ exchange seems to be the major process involved in pHi recovery in SIRC cells after an intracellular acid load. Na+/H+ exchange also plays a role in the maintenance of steady-state pHi.

Amiloride↗

PubMed1989

Palmitic acid stimulates glucose incorporation in the adipocyte by a mechanism likely involving intracellular calcium.

The effect of palmitic acid on basal and insulin-stimulated incorporation of glucose into rat adipocytes was studied. Palmitic acid (2.40 mM) stimulated basal as well as insulin-stimulated glucose incorporation in rat adipocytes three and twofold, respectively. Similar degrees of stimulation of basal glucose oxidation by palmitate were also observed. The ability of palmitic acid to stimulate glucose uptake was additive with respect to the stimulation induced by insulin and was proportional to the palmitic acid concentration between 0.15 mM and 2.40 mM. Stimulation of glucose incorporation by palmitic acid was inhibited by preincubating the cells with quin2-AM, which accumulates intracellularly yielding the trapped chelator form. quin2, which binds intracellular Ca2+.The concentration of quin2-AM required for half-maximal inhibition of palmitic acid stimulated glucose incorporation was 3.8 +/- 1.2 microM (mean +/- SEM). The inhibition of palmitic acid-stimulated glucose incorporation by quin2-AM (10 microM) was overcome by incubating cells with the Ca2+ ionophore, A23187, in the presence of extracellular Ca2+ (2.6 mM). Chelation of extracellular Ca2+ with EGTA did not significantly affect the magnitude of palmitic acid-stimulated glucose incorporation. Dantrolene (12.5-100 microM) failed to affect basal or palmitic acid-stimulated glucose incorporation. These findings suggest that palmitic acid stimulates incorporation of glucose in the adipocyte by a mechanism dependent upon intracellular but not extracellular Ca2+.

Adipose Tissue↗

PubMed1995

Specific accumulation of exogenous fibronectin in experimental glomerulosclerosis.

The prognosis of patients showing glomerulosclerosis as a complication of an immunologically mediated kidney disease is poor. To improve the diagnosis and treatment of these patients, it is important to understand the processes involved in the development of glomerulosclerosis. In this study, we investigated the molecular composition of experimental end-stage glomerular sclerotic lesions and their pathogenesis in chronic graft-versus-host disease (GvHD) in the mouse and chronic serum sickness in the rat. Accumulation studies were performed to determine the degree of specific trapping of constituents from the circulation. Two different models were investigated to determine whether differences in disease initiation resulted in different compositions of the glomerulosclerotic lesions. In both models, glomerulosclerosis was preceded by expansion of the mesangial matrix and thickening of the glomerular basement membrane (GBM). The end-stage sclerotic lesions consisted mainly of fibronectin, which appeared to displace the other extracellular matrix (ECM) components peripherally in the mesangial matrix and GBM. The abundance of fibronectin in the lesions was not reflected in the mRNA levels for this component. Indeed, antibodies directed against the cellular form of fibronectin did not stain positive in the end-stage lesions. These findings, together with accumulation studies, suggest that specific accumulation rather than de novo synthesis of fibronectin plays a major role in the development of experimental glomerulosclerosis, which appears to be independent of the pathway of induction.

Animals↗

PubMed1989

Change in cytoplasmic free Ca2+ concentration in rabbit platelets contacting with albumin coated and uncoated polystyrene surfaces.

Direct measurement of cytoplasmic free Ca2+ concentration in rabbit platelets in contact with polystyrene latexes was examined by monitoring an intracellular trapped fluorescent indicator dye, Fura 2, as a quantitative evaluation of platelet activation on material surfaces. It was found that an increase in cytoplasmic free calcium levels in platelets in contacting latexes was strongly reduced by the adsorption of albumin on the latex surfaces. Further, such an elevation of cytoplasmic free calcium levels was found to require a calcium influx from an extracellular medium across the platelet-plasma membrane.

Animals↗