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At least 973 records · Page 54Linked to original sources
PubMed2005

Stable ciliary activity in human nasal epithelial cells grown in a perfusion system.

PURPOSE: Explore the usefulness of a perfusion system in order to establish human nasal epithelial cell cultures suitable for long-term in vitro ciliary beat frequency (CBF) and cilio-toxicity studies. METHODS: The cells were obtained by protease digestion of nasal biopsy material. The cells were plated at a density of 0.8-1 x 10(6)/cm2 on Vitrogen-coated polyethylene terephthalate membranes, and cultured under submerged conditions in a CO2 incubator or in a perfusion system (initiated on days 8-9 after plating). The CBF was determined at 24.1 +/- 0.8 degrees C by a computerized microscope photometry system. The morphology of the cultured cells was characterized by transmission electron microscopy (TEM). RESULTS: Under CO2 incubator culture conditions, stable ciliary activity was expressed and maintained from day 2 to day 24. Under perfusion system culture conditions, the CBF (mean+/-S.D., n = 4) amounted to 8.4 +/- 0.9 and 8.8 +/- 0.4 Hz on days 7 and 14, respectively. These values were lower as compared to the corresponding CBF obtained in the CO2 incubator cultures (9.5 +/- 0.6 and 9.9 +/- 1.0 Hz, respectively). Reference cilio-stimulatory (glycocholate) and cilio-inhibitory (chlorocresol) compounds were used to assess CBF reactivity. In the CO2 incubator and 7- and 14-days perfusion system cultures, glycocholate (0.5%) showed a reversible cilio-stimulatory effect of 23, 26 and 21%, respectively, while chlorocresol (0.005%) exerted a reversible cilio-inhibitory effect of 36, 40 and 36%, respectively. TEM revealed polarized cuboidal to columnar epithelial morphology, with well-differentiated ciliated cells under CO2 and perfusion system conditions (up to day 23). CONCLUSION: Culturing human nasal epithelial cells on Vitrogen-coated polyethylene terephthalate membranes in submerged conditions in a CO2 incubator and in a perfusion system offers the possibility for long-term preservation (up to 22-24 days) of stable and reactive CBF in vitro.

Animals↗
PubMed2005

Hyaluronic acid combined with mannitol to improve protection against free-radical endothelial damage: experimental model.

PURPOSE: To evaluate the protective properties of combined sodium hyaluronate 2% and mannitol 0.5% (Visiol) on the corneal endothelium in the presence of oxidative stress induced by hydrogen peroxide (H(2)O(2)). SETTING: Instituto Oftalmológico de Alicante, Universidad Miguel Hernández, Alicante, Spain. METHODS: This was an exploratory randomized controlled parallel-group, masked-assessor study of 3 sodium hyaluronate-based ophthalmic viscosurgical devices (OVDs): Visiol, Healon (sodium hyaluronate 1%), and Viscoat (sodium hyaluronate 3%-chondroitin sodium 4%). The OVDs were tested for protective effects on the endothelium following oxidative stress induced by H(2)O(2) at increased concentrations: control (lactated Ringer's solution), 1 mM, 10 mM, and 100 mM. Groups without OVD were used as controls at the same concentrations of peroxide. Each animal received the same treatment in both eyes (10 eyes per group). Endothelial cell lesion was assessed using the Janus green photometry absorbance technique. RESULTS: At 10 mM peroxide concentration, the value of endothelial cell lesion was significantly lower in the Visiol (16.8%, P=.0056), Healon (22.2%, P=.0302), and Viscoat (21.6%, P=.0336) groups than in the control group (29.4%, no OVD). There was a trend in favor of Visiol to more efficiently reduce cell lesions of the endothelium, than Healon (P=.055) and Viscoat (P=.1013). Values of endothelial cell lesion at peroxide concentrations of 1 mM and 100 mM showed the same trends than those observed at 10 mM. CONCLUSIONS: All of the OVDs tested efficiently reduced endothelial lesions against free radicals compared with the control group in which no OVD was used. The following sequence for the efficacy of endothelial cell protection was established: Visiol>Viscoat>Healon>no OVD.

Animals↗
PubMed2025

Biochemical and Structural Analyses of the Tardigrade DNA-Damage Suppressor Protein, Dsup.

Tardigrades are extremophiles that withstand harsh environments through unique molecular strategies. One such strategy involves Damage Suppressor (Dsup), a protein shown to protect cells from radiation-induced DNA damage. Little is known about the biochemical and structural characteristics of Dsup that lead to DNA protection. To gain insight into the mechanism of DNA protection by Dsup, we examined its fundamental biochemical and structural properties using mass photometry, biolayer interferometry, small-angle X-ray scattering, and microfluidic modulation spectroscopy. We found that Dsup is largely intrinsically disordered and binds DNA with high affinity via a multi-valent interface. This interaction induced conformational changes in both Dsup and the DNA, suggesting a potential structural mechanism of its DNA protection ability. We propose that Dsup alters DNA structure, possibly by partially unwinding it, to reduce its susceptibility to damage. These findings offer new insights into how a disordered protein such as Dsup functions as radioprotectants in extreme environments.

Tardigrada↗
PubMed2005

Alterations of ionized Mg2+ in human blood after exercise.

Magnesium (Mg) is the second most abundant intracellular cation with modulating properties in a number of metabolic processes, e.g. in glycolysis, and intracellular signalling processes, e.g. regulation of ion channels and transporters. There are conflicting data available about the regulation of Mg in blood cells during exercise. Moreover, there are no data available about changes of the metabolic important fraction of ionized Mg(2+) both in blood and in blood cells during exercise. The present study investigated the changes of ionized Mg(2+) and total Mg concentration in different compartments after a stepwise treadmill ergometer test. Intracellular ionized Mg(2+) of thrombocytes and erythrocytes was determined by the magnesium sensitive fluorescent dyes mag-fura-2 and Mag-Green using fluorescence spectroscopy and flow cytometry, respectively. Ionized Mg(2+) in blood/serum was measured by an ion-sensitive microelectrode. Total cellular and serum Mg concentration were investigated using atomic absorbance spectroscopy and photometry, respectively. The present results shown that at the end of the ergometer test, ionized Mg(2+) in both blood and serum and total serum Mg decreased. In contrast, intracellular concentration of ionized Mg increased in both thrombocytes and erythrocytes. Total intracellular Mg was unchanged making a Mg(2+) shift between the intra- and extracellular compartment unlikely. The present study therefore demonstrated opposite changes of the ratio [ionized Mg(2+)]/[total Mg] in the intracellular and the extracellular compartment after anaerobic exercise. In in vitro experiments, similar changes of ionized Mg(2+) in both compartments could be mimicked by application of weak acids like propionic and lactic acid. It is concluded changes in the fraction of ionized Mg(2+) should be high enough to influence intracellular signalling and metabolic processes.

Adult↗
PubMed2004

7Li MR measures of blood lithium--correlation with chemical analysis data.

Lithium is used in the treatment and prophylaxis of manic-depressive illness. A narrow therapeutic range of lithium (0.4-1.2 meq/l) requires constant monitoring of these levels to avoid neurotoxicity and overdose. It is general practice to measure plasma lithium levels as a guide to monitor the therapy. Efforts to predict the concentration of Li at its active sites in the brain have led to the use of red blood cells (RBCs) because they are similar to neuronal cells. Thus RBC lithium is a very relevant clinical parameter for monitoring therapy and to observe the changes at intracellular levels under varying treatment conditions. A measure of both plasma and RBC lithium may be of significant value to physicians and researchers as lithium profile for RBCs correlate more closely with the brain lithium than plasma lithium. Although methods to measure lithium in blood such as atomic absorption or flame photometry exist, a complete quantitation of both plasma and RBC lithium requires a tedious physical separation of the two components prior to chemical analysis. On the other hand, lithium MR technique, via the use of shift reagents, can provide both plasma and RBC lithium in a single study. Here we have performed a correlation study of lithium results obtained from MR with the vitros dry-slide method on blood samples from rats treated with lithium. The results show a high degree of correlation between the two methods. Additionally, the MR measurements made on dilute samples of blood indicate that small blood samples with lithium concentration in the neighborhood of 0.08 meq/l can be measured with high accuracy and reproducibility needed for clinical purposes.

Animals↗
PubMed2006

Clinical characteristics of Vogt-Koyanagi-Harada syndrome in Chinese patients.

PURPOSE: To characterize the clinical features of Vogt-Koyanagi-Harada syndrome (VKH) in Chinese patients. DESIGN: Retrospective noncomparative case series. PARTICIPANTS: A total of 410 consecutive VKH patients examined from August 1995 to April 2005. METHODS: Charts of these patients were reviewed according to international VKH criteria. Patients who consulted us within 2 weeks after a uveitis attack were classified into group 1; between 2 weeks and 2 months into group 2; and after 2 months into group 3. The history and clinical findings of all of the patients were reviewed. Laser flare-cell photometry, fundus fluorescein angiography, indocyanine green angiography, optical coherence tomography, B-scan ultrasonography, and ultrasound biomicroscopy were performed in certain cases. Corticosteroids were mainly used to treat patients in groups 1 and 2, whereas cyclosporine or chlorambucil in combination with corticosteroids were prescribed for patients in group 3. MAIN OUTCOME MEASURES: Demographics, clinical presentation, and ancillary examinations of the patients in all 3 groups. RESULTS: The patients were diagnosed as having either complete (n = 273), incomplete (n = 76), or probable (n = 61) VKH syndrome. Exudative retinal detachment and either choroiditis or chorioretinitis were the main findings in group 1. Posterior uveitis with mild to moderate anterior uveitis simulating a nongranulomatous inflammation as evidenced by dust keratic precipitates, anterior chamber cells, and flare was noted in group 2. Recurrent granulomatous anterior uveitis with a "sunset glow" fundus was observed in group 3. Complicated cataract was the most common complication in the patients in group 3. At the final visit, the uveitis was completely controlled in all patients and a best-corrected visual acuity of <20/200 was 1.9%/eye-year, 1.2%/eye-year, and 6%/eye-year in groups 1, 2, and 3, respectively. CONCLUSION: Vogt-Koyanagi-Harada syndrome in Chinese patients is characterized by early posterior uveitis, and if the syndrome is not controlled, subsequent recurrent granulomatous anterior uveitis. Good visual prognosis is possible if the patients are managed according to a tailored immunosuppressive treatment protocol.

Adolescent↗
PubMed2005

Sperm concentration determination between hemacytometric and CASA systems: why they can be different.

Determination of sperm concentration is a critical component of semen analysis. Traditionally, the hemacytometer has been the standard for calibrating other technologies used to estimate sperm concentration, including photometry, Coulter counters, flow cytometry, and computer-automated semen analysis (CASA). Disposable capillary-loaded slides are commonly used in conjunction with most CASA systems currently in use. Questions have been raised regarding differences in sperm concentration measurements between CASA systems (using 20 microm disposable slides) and hemacytometry. This review explains that these differences are largely due to the Segre-Silberberg (SS) effect, which occurs during Poiseuille flow in thin, capillary-loaded slides. The SS effect can lead to errors in estimation of particle concentration, as demonstrated with latex beads and suspensions of human or porcine spermatozoa. The SS effect does not appear to have time to develop in the hemacytometer, which at 100 microm is considerably deeper than most disposable slides. Thus, hemacytometry, when properly performed, remains the gold standard for estimation of sperm concentration. When using thin (20 microm) slides with CASA systems, recognition of the appropriate compensation factor to adjust for the SS effect is critical for accuracy.

Animals↗
PubMed1997

Digital angiography of experimental choroidal melanomas using benzoporphyrin derivative.

PURPOSE: To examine benzoporphyrin derivative angiography as a modality for studying photosensitizer biodistribution in experimental choroidal melanomas. METHODS: A liposomal preparation of benzoporphyrin derivative was used in this study. Digital benzoporphyrin derivative angiograms were performed in 10 rabbits (six for experimental choroidal melanomas, two for normal choroids, and two for irides) using a Topcon ImageNet H1024 digital imaging system, a Kodak Megaplus video camera, and a Topcon TRC-50-VT fundus camera. Only one eye from each rabbit was used. Filters specifically designed for benzoporphyrin derivative (peak absorption at 580 nm and peak emission at 695 nm) were used. Benzoporphyrin derivative (1 mg/kg) was injected into an ear vein while images of tumor, normal choroid, or iris were being obtained. Follow-up images were obtained during the first 3 hours and at 24 hours after injection. Fluorescence microscopy was performed in all 10 rabbits using 1 mg/kg of benzoporphyrin derivative. Tumor-bearing eyes were enucleated at the same time points that angiograms were performed, and the two sets of results were compared for maximum dye accumulation. RESULTS: Digital angiography demonstrated that maximal benzoporphyrin derivative fluorescence occurred in tumors 15 to 45 minutes after injection. Fluorescence photometry corroborated these results. CONCLUSION: Photosensitizer angiography is a valid modality for determining the optimum treatment time for photodynamic therapy.

Animals↗
PubMed1998

Characterization of a heat modifiable protein, Escherichia coli outer membrane protein OmpA in binary surfactant system of sodium dodecyl sulfate and octylglucoside.

A membrane protein, OmpA of Escherichia coli, in the process of refolding from its heat-modified form in the presence of sodium dodecyl sulfate (SDS) to its non-heated one by the addition of systematic amounts of octylglucoside (OG) was characterized by means of dynamic light scattering and the size exclusion chromatography combined with low angle laser light scattering photometry. Upon heating in the presence of SDS only, the amount of SDS bound to OmpA was increased from 1.8 to 2.3 g/g of protein and its hydrodynamic radius increased from 3.7 to 4.7 nm. On the addition of OG, the once denatured OmpA regained its original size above the weight fraction of OG in the total amount of surfactants, 0.8. During the process, the hydrodynamic radius was observed to decrease cooperatively at the weight fraction of 0.6, while no change took place in the molar mass of the protein. The refractive index increment of OmpA reflecting the amount of surfactant binding also regained the value before the heating in parallel with the change of size. Examination of the amount of surfactants bound to the membrane protein according to known properties of the binary surfactant micellar system of the surfactants showed that SDS was principally responsible for the denaturing phenomena of OmpA.

Bacterial Outer Membrane Proteins↗
PubMed1976

The significance of DNA flow-through fluorescence cytophotometry for the diagnosis of prostate carcinoma.

Flow-through fluorescence cytophotometric determination of nuclear DNA content was employed for the diagnosis of prostate carcinoma. Fine needle aspiration biopsy material from the prostate of 220 patients was used for study. A false negative rate of 11.4% and a false positive rate of 29.7% were obtained when the results of flow-through photometry were compared with those of traditional cytodiagnosis. It was found that 4.5% of the specimens were unsuitable for cytologic diagnosis and 10.9% for flow-through cytophotometry. False negative DNA histograms may be due to two factors: either the number of tumor cells is small or there are tumor cells whose nuclear DNA content does not differ from that of a normal cell population. False positive findings result from proliferating cells in inflammatory activation. Errors in preparation of the material and mechanical mistakes, such as cellular clumping and coincidences, are less likely causes. The greater percentage of specimens which were inadequate for cytophotometry was due to the large number of cells needed for a utilizable flow-through photometric histogram. The high rate of false negative and false positive results (11.4% and 29.7%, respectively) argues against using flow-through photometric nuclear DNA determination for the diagnosis of prostate carcinoma.

Biopsy, Needle↗
PubMed1985

Transient dichroism in photoreceptor membranes indicates that stable oligomers of rhodopsin do not form during excitation.

If a photoexcited rhodopsin molecule initiates the formation of rhodopsin oligomers during the process of visual excitation, the rate of rotational diffusion of the rhodopsin molecules involved should change markedly. Using microsecond-flash photometry, we have observed the rotational diffusion of rhodopsin throughout the time period of visual excitation and found that no detectable change occurs in its rotational diffusion rate. Partial chemical cross-linking of the retina yields oligomers of rhodopsin and causes a significant decrease in the rotational diffusion rate of rhodopsin even when as little as 20% of rhodopsin is dimeric. Moreover, the pattern of oligomers formed by cross-linking, taken together with the magnitude of decreases in rotational diffusion rate accompanying the cross-linking reaction, suggests that rhodopsin is a monomer in the dark-adapted state. The experiments reported here show that photoexcited rhodopsin molecules do not irreversibly associate with unbleached neighbors during the time course of the receptor response. Hence, it is not likely that stable oligomers of rhodopsin trigger the excitation of the photoreceptor cell.

Animals↗
PubMed1987

Measurement of tissue potassium in vivo using 39K nuclear magnetic resonance.

39K nuclear magnetic resonance (NMR) spectra were readily obtained, in vivo, from rat muscle, kidney, and brain in 5-10 min with signal-to-noise ratios of approximately 20:1. Quantitation of the K+ signal was achieved by reference to an external standard of KCl/dysprosium nitrate as well as by reference to the proton signal from tissue water. In vitro NMR studies of isolated tissue showed a K+ visibility (NMR K+/total tissue K+) of 96%, 62 +/- 8%, 47 +/- 1.9%, 45 +/- 3.5%, and 43 +/- 2.5% for blood, brain, muscle, kidney, and liver, respectively. Absolute tissue K+ was determined by flame photometry of acid-digested tissue. Changes in tissue K+ status by chronic K+ depletion or acute K+ loading produced changes of 39K NMR signal intensity that were equal to changes of absolute tissue K+. Acidosis, alkalosis, mannitol, or RbCl infusion did not significantly change the NMR K+ signal. These results indicate that the changes in K+ detected by NMR were specifically and accurately detected. To investigate the factors that affect the 39K NMR signal, the effects of liver homogenate on 39K NMR signal intensity were studied. Addition of homogenate produced a 60% loss of signal intensity, suggesting that a large portion of cell K+ may be only 40% visible. Addition of RbCl to undiluted homogenate increased the NMR K+ signal by 11 +/- 2 mumol/g. Addition of H2O or NaCl had no effect, suggesting that Rb+ was replacing K+ in sites of low (less than 40%) NMR visibility. These results demonstrate that 39K NMR experiments can be performed using intact organs. To explain the lack of detectable K+ and changes in K+ NMR visibility, a three compartment model is proposed.

Animals↗
PubMed1991

Formation of liquid crystalline phase of actin filament solutions and its dependence on filament length as studied by optical birefringence.

We studied the formation and structure of liquid crystalline phase of F-actin solutions by polarized light photometry, assuming that a small domain of the liquid crystalline phase works as a linear retardation plate. Transmittance of polarized light due to the birefringence of liquid crystalline phase appeared above a threshold concentration of F-actin. The threshold increased with a decrease in filament length, which was regulated by calcium-activated gelsolin. The intensity increased linearly with increasing concentrations until it reached a stationary value. The deviation of optical axis direction of the putative retardation plate was estimated 7-15 degrees. These results indicate that:(a) the liquid crystalline phase is formed above a threshold concentration of F-actin; (b) the threshold is proportional to the inverse of filament length; (c) the ordered phase coexists with the isotropic one, increasing the volume fraction with increasing concentrations until all filaments take the liquid crystalline structure; (d) the filaments in liquid crystalline phase take a highly ordered array. These results can be attributed to the excluded volume effect of rod-like molecules on the formation of liquid crystalline structure.

Actin Cytoskeleton↗
PubMed1994

Cation conductance and efflux induced by polyene antibiotics in the membrane of skeletal muscle fiber.

Cation conductance and efflux induced by polyene antibiotics amphotericin B (AMB), amphotericin B methyl ester (AME), nystatin, mycoheptin, and levorin on frog isolated skeletal muscle fibers and whole sartorius muscles were investigated. Conductance was measured under current-clamp conditions using a double sucrose-gap technique. Cation efflux was studied using flame emission photometry. Some new data were obtained concerning the effects of levorin and mycoheptin on biological membranes. The power dependence of polyene-induced cation transport on antibiotic concentration in muscle membrane was lower than that in bilayers. The decline in the equilibrium conductance caused by polyene removal (except for levorin) was very fast. There was reverse temperature dependence of AMB- and nystatin-induced conductances. Both induced conductance and efflux values demonstrated a correlation with the order of antifungal activities: levorin > AMB, mycoheptin > AME > nystatin, except for AME, which was more potent on yeastlike cells. These effects were interpreted in terms of possible differences in the kinetics of channel formation in biological and model membranes and in light of the role of nonconducting antibiotic forms in biological membranes.

Amphotericin B↗
PubMed1998

Mathematical modeling and fluorescence imaging to study the Ca2+ turnover in skinned muscle fibers.

A mathematical model was developed for the simulation of the spatial and temporal time course of Ca2+ ion movement in caffeine-induced calcium transients of chemically skinned muscle fiber preparations. Our model assumes cylindrical symmetry and quantifies the radial profile of Ca2+ ion concentration by solving the diffusion equations for Ca2+ ions and various mobile buffers, and the rate equations for Ca2+ buffering (mobile and immobile buffers) and for the release and reuptake of Ca2+ ions by the sarcoplasmic reticulum (SR), with a finite-difference algorithm. The results of the model are compared with caffeine-induced spatial Ca2+ transients obtained from saponin skinned murine fast-twitch fibers by fluorescence photometry and imaging measurements using the ratiometric dye Fura-2. The combination of mathematical modeling and digital image analysis provides a tool for the quantitative description of the total Ca2+ turnover and the different contributions of all interacting processes to the overall Ca2+ transient in skinned muscle fibers. It should thereby strongly improve the usage of skinned fibers as quantitative assay systems for many parameters of the SR and the contractile apparatus helping also to bridge the gap to the intact muscle fiber.

Animals↗
PubMed1999

The actin-based nanomachine at the leading edge of migrating cells.

Two fundamental parameters of the highly dynamic, ultrathin lamellipodia of migrating fibroblasts have been determined-its thickness in living cells (176 +/- 14 nm), by standing-wave fluorescence microscopy, and its F-actin density (1580 +/- 613 microm of F-actin/microm(3)), via image-based photometry. In combination with data from previous studies, we have computed the density of growing actin filament ends at the lamellipodium margin (241 +/- 100/microm) and the maximum force (1.86 +/- 0.83 nN/microm) and pressure (10.5 +/- 4.8 kPa) obtainable via actin assembly. We have used cell deformability measurements (. J. Cell Sci. 44:187-200;. Proc. Natl. Acad. Sci. USA. 79:5327-5331) and an estimate of the force required to stall the polymerization of a single filament (. Proc. Natl. Acad. Sci. USA. 78:5613-5617;. Biophys. J. 65:316-324) to argue that actin assembly alone could drive lamellipodial extension directly.

3T3 Cells↗
PubMed2000

Directed sampling for electrolyte analysis and water content of micro-punch samples shows large differences between normal and ischemic rat brain cortex.

Changes in sodium, potassium, and water content in brain tissue are important in the progression of pathology that follows ischemic stroke. Determining these parameters regionally in rodent models of experimental ischemia has been limited because typical tissue weights of more than 35 mg are too large. Identifying ischemic tissue to direct tissue sampling towards ischemic cortex is also represents a difficult generally unresolved area. We suggest that larger differences between normal and ischemic cortex of sodium, potassium, and water content than previously observed can be obtained from directed sampling of 2-mg brain tissue in a model of focal cerebral ischemia. In five rats, the middle cerebral artery and both common carotid arteries were occluded for 4.9+/-0.13 h (mean+/-SEM). Punch-sampling of 1-mm diameter tissue cores for water content (H(2)O%) by the wet-dry method, and [Na(+)] and [K(+)] by flame photometry, was guided by the observation of a subtle change in the surface reflectivity of ischemic cortex of quickly dried, 20-microm frozen brain sections, that was confirmed by MAP2 immunohistochemistry. The ratio of the lesion areas as determined by the reflective change and MAP2 immunoreactivity was 0.96+/-0.03 (n=5). In ischemic cortex H(2)O% was 79.9%+/-0.8%, [Na(+)] was 550+/-25 mEq/kg dry-weight, and [K(+)] 94.2+/-19.2 mEq/kg dry-weight (n=5), all significantly different from the values in border zone cortex, and in cortex contralateral to ischemic cortex and border zone (for all samples n=60, mean wet weight 2.037+/-0.046 mg). Differences between ischemic and normal cortex were 5.4+/-1.1%, 317+/-21 mEq/kg dry-weight, -304+/-27 mEq/kg dry-weight (n=5) for H(2)O%, [Na(+)], and [K(+)]. These differences between ischemic and normal cortex are 1.4-2.5, 1-3.11, and 1.4-3.5 times greater, respectively, than previous results obtained using samples weighing 35 mg or more. These results extend the association of sodium and potassium with ischemic brain edema in the rodent model, and show that these classical measurements can keep pace with the regionality of histochemical and morphological methods.

Animals↗
PubMed1991

Participation of D-galactose-specific receptors of liver macrophages in recognition of fibronectin-opsonized particles.

The interaction of immobilized human or rat plasma fibronectin with isolated rat liver macrophages was studied in a model system using colloidal gold of 17-nm diameter (Au-17) as test particles. Plasma fibronectin (pFn)-coated gold particles were rapidly bound and endocytosed via the coated pit-coated vesicle pathway as demonstrated by photometry, and light and electron microscopy. The isolated macrophages bind 2.5 +/- 2 particles/10 microns of plasma membrane (incubation at 4 degrees), equalling a binding capacity of approximately 3.5 x 10(4) pFn-Au-17 particles per cell. Binding and uptake (at 37 degrees) was specifically inhibited by D-galactose-related carbohydrates, but not by D-mannose, N-acetyl-D-glucosamine, nor by excess soluble pFn. Uptake was also inhibited by lactosylated bovine serum albumin at a concentration of 10(-6) M but not by bovine serum albumin. India ink uptake by the liver macrophages in the presence of fibronectin was also inhibited by D-galactose-related monosaccharides. The presence of terminal, nonreducing D-galactosyl groups on pFn could be demonstrated by agglutination experiments with the D-galactose-specific plant lectin, Ricinus communis agglutinin (RCA), which could also be used for isolation of pFn from rat plasma. The 29-kDa molecular mass D-galactose-specific receptor, known to be expressed on the liver macrophage membrane and recently shown to be a membrane-bound form of C-reactive protein, was found to bind the pFn-coated gold particles in dot blotting experiments. It was concluded that the D-galactose-specific macrophage receptor binds to terminal D-galactose-related units of immobilized pFn and participates in recognition of fibronectin-opsonized particles.

Animals↗