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At least 415 records · Page 23Linked to original sources
PubMed1999

Chondrocyte-biocompatibility of DegraPol-foam: in vitro evaluations.

Histological and biochemical investigations were carried out in order to evaluate the chondrocyte compatibility of a recently developed biodegradable polyesterurethane-foam (DegraPol-foam). Therefore, cell adhesion, cell growth, and the preservation of chondrocyte phenotype was measured in rat xyphoid chondrocytes seeded on DegraPol-foam. Chondrocytes, isolated from xyphoids of adult male rats, exhibited relatively high cell adhesion on DegraPol-foam (about 60% of that found on TCPS). Scanning electron microscopy (SEM) showed that chondrocytes grew on the surface and into the open cell pores of the foam. Morphologically, cells found on the surface of the foam exhibited a flat cell appearance and built a confluent cell multilayer. In contrast, the interior of the foam cells showed rounded morphology in cell aggregates and cell islets. In addition, chondrocytes proliferated on the DegraPol-foam (doubling-time of about 12.5 days) and preserved their phenotype for up to 14 days. Compared to freshly isolated chondrocytes, cells seeded on the foam produced high concentrations of collagen type II for up to 2 weeks: the ratio of type II/I collagen was 1.2-1.4 fold higher than the ratio found in freshly isolated cells. No significant difference was observed in chondroitin sulfate levels produced by freshly isolated cells and cells cultured on DegraPol-foam for up to 14 days. To sum up, our results indicate that DegraPol-foam is a compatible substrate for chondrocytes.

3T3 Cells↗
PubMed2000

[Tissue engineering of vascularized bone and soft tissue transplants].

The utilization of in vitro angiogenesis in tissue engineering might be useful in order to establish an artificial vascular network. However, it remains unclear how far the in vitro preformation of vascular structures may contribute to the perfusion of larger artificial tissue aggregates regarding the improvement of oxygenation and nutrition. In an in vitro study, we developed a model of a vascularized tissue. Stromal cells of a target tissue, e.g., adipose tissue or bone tissue, were expanded in vitro and seeded onto microcarriers or microparticles. Densely covered microcarriers were brought into a fibrin matrix together with endothelial cells. In order to demonstrate the formation and stabilization of capillary-like structures, UEA-I labeled specimens were evaluated using laser scanning microscopy and digital image analysis. The stabilization of capillary-like structures was better with stromal cells from bone marrow than from adipose tissue. In one of the culture aggregates, the total length of capillary-like structures increased after 6 weeks of cultivation to up to 140 mm/mm3. Additional tests were performed utilizing hyperbaric oxygenation. In the oxygenation group, a significant increase in the length of capillary-like structures was found. The method implies the option of coculturing different tissue elements and of an in vitro preformation of vascularized tissues.

Bone Transplantation↗
PubMed2003

[PHI+], a novel Sup35-prion variant propagated with non-Gln/Asn oligopeptide repeats in the absence of the chaperone protein Hsp104.

BACKGROUND: The [PSI+] element of the budding yeast is an aggregated form of the translation release factor Sup35 that is propagated and transmitted cytoplasmically in a manner analogous to that of mammalian prions. The N-terminal of Sup35, necessary for [PSI+], contains oligopeptide repeats and multiple Gln/Asn residues. RESULTS: We replaced the Gln/Asn-rich prion repeats of Sup35 with non-Gln/Asn repeats from heterologous yeast strains. These non-Gln/Asn repeat Sup35s propagated a novel [PSI+] variant, [PHI+], that appeared de novo 103 times more frequent than [PSI+]. [PHI+] was stably inherited in a non-Mendelian fashion, but not eliminated upon the inactivation of Hsp104, unlike known [PSI+] elements. In vitro, non-Gln/Asn repeat domains formed amyloid fibres that were shorter and grew more slowly than did Gln/Asn-rich prion domains, while [PHI+] aggregates were smaller than [PSI+] aggregates in vivo. CONCLUSIONS: These findings suggest the existence of an alternative, Hsp104-independent pathway to replicate non-Gln/Asn variant Sup35 prion seeds.

Amino Acid Sequence↗
PubMed1991

Tissue and Cellular Localization of Proteinase Inhibitors I and II in the Fruit of the Wild Tomato, Lycopersicon peruvianum (L.) Mill.

The cellular and subcellular localization of proteinase inhibitor I and inhibitor II proteins in the fruit of the wild tomato species Lycopersicon peruvianum (L.) Mill., LA 107 was determined by immunoanalysis of tissue blots and protein-A gold immunocytochemistry. Tissue blot analysis showed that the proteinase inhibitor I proteins were located throughout the fruit tissue, with the exception of the seeds. Light microscopy, using immunocytochemical labeling, indicated that all the parenchyma cells of the pericarp contained inhibitor I and II proteins in dense vacuolar protein aggregates that were not membrane bound. The size, number, and morphology of the aggregates within individual cells varied greatly. The funiculus, ovule, and early embryonic tissues were devoid of inhibitor I and II. Immunocytochemical analysis using transmission electron microscopy confirmed that the proteinase inhibitor I proteins were principally located and stored in protein aggregates within the vacuole of the fruit parenchyma cells. Some cytoplasmic protein-A gold immunolabeling of inhibitor I proteins was evident, which may be related to the synthesis and intermediate transport steps preceding storage of the inhibitor I proteins in the vacuoles.

Journal Article↗
PubMed1994

Hepatocyte culture between woven capillary networks: a microscopy study.

A multi-compartment capillary membrane culture model with independently perfused three-dimensionally woven capillaries was developed for immobilization of hepatocytes in bioreactors. This enables spatial restructuring of cells and enhanced mass transfer performance with more efficient oxygenation and metabolite exchange. Seeding density defines cell behaviour in this model. With low densities cells attach to the membranes and flatten. Increasing density leads to spontaneous formation of aggregates which are immobilized between the capillaries.

Animals↗
PubMed2001

Optimization of hepatocyte spheroid formation for hepatic tissue engineering on three-dimensional biodegradable polymer within a flow bioreactor prior to implantation.

We hypothesize that in vitro conditioning of hepatocytes within biodegradable poly-L-lactic acid (PLLA) polymer matrices prior to implantation may increase hepatocyte survival and function after transplantation. The purpose of this study was to optimize the culture conditions of hepatocytes in a pulsatile flow bioreactor. PLLA discs were seeded with rat hepatocytes in a concentration of 2.5, 5, 10, 20 and 40 x 10(6) cells/ml. Seeded discs were exposed to recirculated perpendicular flow of 0, 7, 15, 24, 32, 52 ml/min of supplemented Williams' Medium E and harvested after 6 days in flow culture. Only under flow conditions the hepatocytes formed spheroidal aggregates (SphA) of 50-260 microm in diameter with a liver-like morphology and active metabolic function. The number of SphA was examined by phase contrast microscopy and the reductive enzyme function of the hepatocytes was tested using MTT. Hematoxylin and eosin histology showed vital hepatocytes within the SphA less than 200 microm in diameter but central necrosis in the SphA exceeding this size. Immunohistochemical staining confirmed albumin production of hepatocytes within the SphA. The optimal cell seeding concentration was 10 x 10(6) cells/ml with a flow speed of 24 ml/min. SphA of hepatocytes cultured with this flow bioreactor method may prove useful as a functional unit for tissue engineering of an in vivo liver substitute.

Albumins↗
PubMed1987

An assay for leukoagglutinating lectins using suspension cultured mouse lymphoma cells (BW5147) stained with neutral red.

A sensitive and rapid assay for leukoagglutinating lectins has been developed. This assay utilizes neutral red-stained mouse lymphoma cells from the suspension cultured cell line BW5147. The agglutination of the stained cells can be monitored visually in a manner similar to that for conventional assays for erythroagglutinating lectins using erythrocytes. The activity of lekoagglutinating lectins that are not capable of agglutinating erythrocytes can be quantified by this assay. The utility of the assay was demonstrated using leukoagglutinating and erythroagglutinating lectins from the seeds of Phaseolus vulgaris and Maackia amurensis.

Animals↗
PubMed1979

Effect of sodium dodecyl sulphate on the 12S fraction of mustard proteins (B. juncea).

The effect of SDS on the 12S protein fraction of mustard seed (B. juncea) has been followed by the techniques of ultracentrifugation, gel filtration, gel electrophoresis, viscosity, ultraviolet difference spectra and fluorescence spectra. At low concentrations of SDS, up to 0.1%, both aggregation and dissociation of the protein occurs. Only dissociation occurs at higher SDS concentrations and is complete at 0.5% SDS. Viscosity increases sharply up to 0.15% SDS, remains constant between 0.15 and 0.30% and then increases markedly again. SDS induces also difference spectra with minima at 280, 288 and 295 nm. Fluorescence emission intensity increases at SDS concentrations less than 0.05% and quenching occurs at higher SDS concentrations. The results suggest that SDS causes association, dissociation and denaturation of the protein molecule.

Brassica↗
PubMed2003

[Alpha-linolenic acid and cardiovascular diseases].

IMPORTANCE AND METABOLISM OF ALPHA-LINOLENIC ACID: Alpha-linolenic acid is an essential fatty acid which cannot be produced in the body and must be taken by food. Both in animals and humans, alpha-linolenic acid is desaturated and elongated into eicosapentaenoic and docosahexaenoic acid. It is also incorporated into plasma and tissue lipids and its conversion is affected by levels of linoleic acid. POTENTIAL ROLE IN PATHOGENESIS OF CARDIOVASCULAR DISEASES: Diet enriched in n-3 fatty acids, especially alpha-linolenic acid, reduces the incidence of cardiac death. Studies have shown that alpha linolenic acid prevents ventricular fibrillation which is the main cause of cardiac death. Studies in rats suggest that alpha-linolenic acid may be more effective in preventing ventricular fibrillations than eicosapentaenoic and docosahexaenoic acid. Furthermore, alpha-linolenic acid is the main fatty acid decreasing platalet aggregation which is an important step in thrombosis i.e. non-fatal myocardial infarction and stroke. DIETARY SOURCES AND NUTRITION RECOMMENDATIONS: Dietary sources include flaxseed and flaxseed oil, canola oil, soybean and soybean oil, pumpkin seed and pumpkin oil, walnuts and walnut oil. Strong evidence supports beneficial effects of alpha-linolenic acid and its dietary sources should be incorporated into balanced diet for prevention of cardiovascular diseases. The recommended daily intake is 2 g with a ratio of 5/1 for linoleic/alpha-linolenic acid.

Animals↗
PubMed2006

Effect of extract of Phyllanthus niruri on crystal deposition in experimental urolithiasis.

Phyllanthus niruri (Pn) is a plant that has been shown to interfere in the growth and aggregation of calcium oxalate (CaOx) crystals. In the present study we evaluated the effect of Pn on the preformed calculus induced by introduction of a CaOx seed into the bladder of male Wistar rats. Pn treatment (5 mg/ rat/day) was initiated immediately or 30 days after CaOx seeding and thus in the presence of a preformed calculus. Animals were sacrificed 50 or 70 days after surgery. The resulting calculi were weighed and analyzed by X-ray diffraction, stereomicroscopy and scanning electronic microscopy. Precocious Pn treatment reduced the number (75%, P < 0.05) and the weight (65%, P < 0.05) of calculi that frequently exhibited a matrix-like material on its surface, compared to the untreated CaOx group. In contrast, Pn treatment in the presence of a preformed calculus did not prevent further calculus growth; rather, it caused an impressive modification in its appearance and texture. Calculi from Pn-treated animals had a smoother, homogeneous surface compared to the spicule shape of calculi found in the untreated CaOx group. XRD analysis revealed the precipitation of struvite crystals over the CaOx seed and Pn did not change the crystalline composition of the calculi. This suggests that Pn interfered with the arrangement of the precipitating crystals, probably by modifying the crystal-crystal and/or crystal-matrix interactions. Results suggest that Pn may have a therapeutic potential, since it was able to modify the shape and texture of calculi to a smoother and probably more fragile form, which could contribute to elimination and/or dissolution of calculi.

Animals↗
PubMed1984

Some observations on the pharmacology of 'deep-heat', a topical rubifacient.

A topically applied rubifacient delivered by aerosol (Deep-Heat) was studied. After spray application to the forearms of volunteers, without massage, the erythema produced was measured by thermography and correlated with the concentration of 2 salicylate components of the mixture found in local and systemic venous blood. Maximum erythema occurred at about 30 minutes, while blood salicylate levels were maximal between 20 and 30 minutes after application. Methyl salicylate was absorbed before ethyl salicylate. Over the time period of the erythematous response oxygen levels in local venous blood were raised. Finally, platelets collected from venous blood draining from the sprayed site, when induced to clump by the addition of arachidonic acid in an aggregometer, showed increased resistance to clumping when compared with control cells. The mechanism of these observed phenomena and the mode of action of the constituents of Deep-Heat are discussed.

Administration, Topical↗
PubMed2001

The roles of turn formation and cross-strand interactions in fibrillization of peptides derived from the OspA single-layer beta-sheet.

We previously demonstrated that a beta-hairpin peptide, termed BH(9-10), derived from a single-layer beta-sheet of Borrelia OspA protein, formed a native-like beta-turn in trifluoroethanol (TFE) solution, and it assembled into amyloid-like fibrils at higher TFE concentrations. This peptide is highly charged, and fibrillization of such a hydrophilic peptide is quite unusual. In this study, we designed a circularly permutated peptide of BH(9-10), termed BH(10-9). When folded into their respective beta-hairpin structures found in OspA, these peptides would have identical cross-strand interactions but different turns connecting the strands. NMR study revealed that BH(10-9) had little propensity to form a turn structure both in aqueous and TFE solutions. At higher TFE concentration, BH(10-9) precipitated with a concomitant alpha-to-beta conformational conversion, in a similar manner to the BH(9-10) fibrillization. However, the BH(10-9) precipitates were nonfibrillar aggregation. The precipitation kinetics of BH(10-9) was exponential, consistent with a first-order molecular assembly reaction, while the fibrillization of BH(9-10) showed sigmoidal kinetics, indicative of a two-step reaction consisting of nucleation and molecular assembly. The correlation between native-like turn formation and fibrillization of our peptide system strongly suggests that BH(9-10) adopts a native-like beta-hairpin conformation in the fibrils. Remarkably, seeding with the preformed BH(10-9) precipitates changed the two-step BH(9-10) fibrillization to a one-step molecular assembly reaction, and disrupted the BH(9-10) fibril structure, indicating interactions between the BH(10-9) aggregates and the BH(9-10) peptide. Our results suggest that, in these peptides, cross-strand interactions are the driving force for molecular assembly, and turn formation limits modes of peptide assembly.

Antigens, Surface↗
PubMed1998

Insulin-like growth factor-I-mediated survival from anoikis: role of cell aggregation and focal adhesion kinase.

Anoikis is a form of cell death that occurs when cells are denied attachment to the extra-cellular matrix. Using p6 cells, that are 3T3 cells overexpressing the type 1 insulin-like growth factor receptor (IGF-IR), we show that these cells undergo apoptosis when seeded on polyHEMA plates in serum-free medium (SFM). IGF-I protects p6 cells from anoikis, without inducing mitogenesis or DNA synthesis. In the surviving p6 cells in suspension cultures, the focal adhesion kinase (FAK) is tyrosyl phosphorylated by IGF-I, although this phosphorylation occurs only after several hours. The importance of FAK in protection from anoikis is confirmed by v-src-transformed R-cells, in which FAK is constitutively phosphorylated, that survive even in SFM. Surviving cells, whether p6 or v-src transformed, tend to form large cell aggregates, whose appearance precedes the phosphorylation of FAK. These and other findings suggest that FAK phosphorylation in the case of IGF-I is a mediated effect rather than a direct one. When p6 cells are plated on polyHEMA dishes, IGF-I induces cell aggregation and this aggregation correlates with survival and the eventual phosphorylation of FAK.

3T3 Cells↗
PubMed1999

Potentiation of beta-folding of beta-amyloid peptide 25-35 by aluminum salts.

The formation of the beta pleated configuration of the amyloid peptide fragment 25-35 in aqueous solution, has been studied using thioflavin-T fluorescence as an indicator of such folding. Both phosphate and adenosine triphosphate (ATP) enhance the formation of aggregated beta-sheets. This phosphate-induced aggregation is greater in the presence of aluminum sulfate in a dose dependent manner. In the absence of ATP or phosphate, aluminum salts do not promote aggregation. It is proposed that a particulate aluminum phosphate complex may form critical nuclei upon whose surface the amyloid peptide can change its configuration. This capacity for seeding may be a relevant factor in the formation of insoluble proteinaceous materials such as amyloid plaques and neurofibrillary tangles found in Alzheimer's disease.

Alum Compounds↗
PubMed2002

Influence of urinary sialic acid on calcium oxalate crystal formation.

Using seed crystal method, whole-urine method, and scanning electron microscopy, the inhibitory effects of sialic acid and osteopontin (OPN) on aggregation/growth of CaOx crystals were investigated. Using the seed crystal method, sialic acid showed an inhibitory effect on CaOx crystal aggregation/growth in a concentration-dependent manner, but almost no effect was observed using the whole-urine method. OPN showed an inhibitory effect on aggregation/growth in both experimental systems. The inhibitory effect of asialo-OPN on aggregation/growth was approximately 20% lower than that of OPN in the experiment using the seed crystal method and approximately 15% lower in the experiment using the whole-urine method. Scanning electron microscopy showed that OPN and sialic acid inhibit the aggregation of CaOx crystals. The above findings show that sialic acid accounts for about 15-20% of the involvement of OPN in CaOx crystallization.

Calcium Oxalate↗
PubMed1992

Characterization of the charged components and their topology on the surface of plant seed oil bodies.

Oil bodies of plant seeds contain a triacylglycerol matrix surrounded by a monolayer of phospholipids embedded with alkaline proteins called oleosins. Oil bodies isolated from maize (Zea mays L.) in a medium of pH 7.2 maintained their entities but aggregated when the pH was lowered to 6.8 and 6.2. Aggregation did not lead to coalescence and was reversible with an elevation of the pH. Further decrease of the pH from 6.2 to 5.0 retarded the aggregation. Aggregation at pH 7.2 was induced with 2 mM CaCl2 or MgCl2 but not with NaCl. Aggregation at pH 6.8 was prevented by 10 microM sodium dodecyl sulfate but not with NaCl. We conclude that oil bodies have a negatively charged surface at pH 7.2 and an isoelectric point of about 6.0. This conclusion is supported by isoelectrofocusing results and by theoretical calculation of the positive charges in the oleosins and the negative charges in phosphatidylserine, phosphatidylinositol, and free fatty acids. Apparently, lowering of the pH from 7.2 to 6.2 protonates the histidine residues in the oleosins, and neutralizes the oil bodies. Further decrease of the pH to 5.0 likely protonates the free fatty acids and produces positively charged organelles. Similar charge properties were observed in the oil bodies isolated from rape, flax, and sesame seeds. An analysis of the oleosin secondary structures reveals an N-terminal amphipathic domain, a central hydrophobic anti-parallel beta-strand domain (not found in any other known protein), and a C-terminal amphipathic alpha-helical domain. In the two amphipathic domains, the positively charged residues are orientated toward the interior facing the negative charged lipids, whereas the negatively charged residues are exposed to the exterior. The negatively charged surface is a major factor in maintaining the oil bodies as stable individual entities.

Amino Acid Sequence↗