scholarly journals Phospholipid of the rat glomerular basement membrane in experimental nephrosis

1972 ◽  
Vol 129 (3) ◽  
pp. 733-741 ◽  
Author(s):  
Kevin K. Fung ◽  
Norman Kalant
Keyword(s):  
Basement Membrane ◽  
Glomerular Basement Membrane ◽  
Plasma Membranes ◽  
Phospholipid Composition ◽  
Renal Tissue ◽  
Relative Increase ◽  
Phospholipid Content ◽  
The Individual

Phospholipids were found to be a constant component of rat glomerular basement-membrane preparations. The concentration fell during preparation of basement membrane by sonication of whole glomeruli, but then remained constant despite continued sonication. The proportions of the individual phospholipids were different from those of whole renal tissue or of isolated glomeruli. The basement-membrane preparations had no (Na++K+)-activated adenosine triphosphatase activity, an enzyme that is bound to plasma membranes. The concentration of lipid P was decreased on exposure in vivo or in vitro to antiserum against basement membrane; 7 days after injection of antiserum there was a change in the phospholipid composition, with a relative increase in phosphatidylcholine and a decrease in sphingomyelin content. The metabolic turnover rate of the lipid P remaining in the membrane was normal, as determined by 32P incorporation. The loss of phospholipid was associated with decreases in the relative concentrations of hydroxyproline, hydroxylysine and glycine, and relative increases in proline, lysine, serine, threonine and valine. Administration of aminonucleoside and daunomycin produced proteinuria but did not cause a decrease in lipid P. Anticollagen and anti-lymphocyte sera that attached to the basement membrane but failed to produce proteinuria, also failed to affect the phospholipid content.

scholarly journals Quantification of Na+,K+ pumps and their transport rate in skeletal muscle: Functional significance

2013 ◽  
Vol 142 (4) ◽  
pp. 327-345 ◽  
Author(s):  
Torben Clausen
Keyword(s):  
Skeletal Muscle ◽  
Plasma Membranes ◽  
Cell Damage ◽  
Total Content ◽  
Relative Increase ◽  
Transport Rate ◽  
Α Subunit ◽  
Muscle Inhibition

During excitation, muscle cells gain Na+ and lose K+, leading to a rise in extracellular K+ ([K+]o), depolarization, and loss of excitability. Recent studies support the idea that these events are important causes of muscle fatigue and that full use of the Na+,K+-ATPase (also known as the Na+,K+ pump) is often essential for adequate clearance of extracellular K+. As a result of their electrogenic action, Na+,K+ pumps also help reverse depolarization arising during excitation, hyperkalemia, and anoxia, or from cell damage resulting from exercise, rhabdomyolysis, or muscle diseases. The ability to evaluate Na+,K+-pump function and the capacity of the Na+,K+ pumps to fill these needs require quantification of the total content of Na+,K+ pumps in skeletal muscle. Inhibition of Na+,K+-pump activity, or a decrease in their content, reduces muscle contractility. Conversely, stimulation of the Na+,K+-pump transport rate or increasing the content of Na+,K+ pumps enhances muscle excitability and contractility. Measurements of [3H]ouabain binding to skeletal muscle in vivo or in vitro have enabled the reproducible quantification of the total content of Na+,K+ pumps in molar units in various animal species, and in both healthy people and individuals with various diseases. In contrast, measurements of 3-O-methylfluorescein phosphatase activity associated with the Na+,K+-ATPase may show inconsistent results. Measurements of Na+ and K+ fluxes in intact isolated muscles show that, after Na+ loading or intense excitation, all the Na+,K+ pumps are functional, allowing calculation of the maximum Na+,K+-pumping capacity, expressed in molar units/g muscle/min. The activity and content of Na+,K+ pumps are regulated by exercise, inactivity, K+ deficiency, fasting, age, and several hormones and pharmaceuticals. Studies on the α-subunit isoforms of the Na+,K+-ATPase have detected a relative increase in their number in response to exercise and the glucocorticoid dexamethasone but have not involved their quantification in molar units. Determination of ATPase activity in homogenates and plasma membranes obtained from muscle has shown ouabain-suppressible stimulatory effects of Na+ and K+.

Effects of plasma proteins on sieving of tracer macromolecules in glomerular basement membrane

2001 ◽  
Vol 281 (5) ◽  
pp. F860-F868 ◽  
Author(s):  
Matthew J. Lazzara ◽  
William M. Deen
Keyword(s):  
Basement Membrane ◽  
Glomerular Basement Membrane ◽  
Plasma Proteins ◽  
Prolate Spheroid ◽  
Osmotic Effect ◽  
Sieving Coefficient ◽  
Total Plasma Protein ◽  
Macromolecular Tracers

It was found previously that the sieving coefficients of Ficoll and Ficoll sulfate across isolated glomerular basement membrane (GBM) were greatly elevated when BSA was present at physiological levels, and it was suggested that most of this increase might have been the result of steric interactions between BSA and the tracers (5). To test this hypothesis, we extended the theory for the sieving of macromolecular tracers to account for the presence of a second, abundant solute. Increasing the concentration of an abundant solute is predicted to increase the equilibrium partition coefficient of a tracer in a porous or fibrous membrane, thereby increasing the sieving coefficient. The magnitude of this partitioning effect depends on solute size and membrane structure. The osmotic reduction in filtrate velocity caused by an abundant, mostly retained solute will also tend to elevate the tracer sieving coefficient. The osmotic effect alone explained only about one-third of the observed increase in the sieving coefficients of Ficoll and Ficoll sulfate, whereas the effect of BSA on tracer partitioning was sufficient to account for the remainder. At physiological concentrations, predictions for tracer sieving in the presence of BSA were found to be insensitive to the assumed shape of the protein (sphere or prolate spheroid). For protein mixtures, the theoretical effect of 6 g/dl BSA on the partitioning of spherical tracers was indistinguishable from that of 3 g/dl BSA and 3 g/dl IgG. This suggests that for partitioning and sieving studies in vitro, a good experimental model for plasma is a BSA solution with a mass concentration matching that of total plasma protein. The effect of plasma proteins on tracer partitioning is expected to influence sieving not only in isolated GBM but also in intact glomerular capillaries in vivo.

scholarly journals Renal glomerular proteoglycans. An investigation of their synthesis in vivo using a technique for fixation in situ

1988 ◽  
Vol 251 (2) ◽  
pp. 411-418 ◽  
Author(s):  
L A Beavan ◽  
M Davies ◽  
R M Mason
Keyword(s):  
Basement Membrane ◽  
Glomerular Basement Membrane ◽  
Membrane Fraction ◽  
Chondroitin Sulphate ◽  
Heparan Sulphate ◽  
Basement Membranes ◽  
Chondroitin Sulphate Proteoglycans

Newly synthesized rat glomerular [35S]proteoglycans were labelled in vivo after injecting Na2[35S]SO4 intraperitoneally. At the end of the labelling period (7 h) the kidneys were perfused in situ with 0.01% (w/v) cetylpyridinium chloride. This fixed proteoglycans in the tissue and increased their recovery 2-3-fold during subsequent isolation of glomeruli from the renal cortex. The glomeruli were fractionated by a modified osmotic lysis and detergent extraction procedure [Meezan, Brendel, Hjelle & Carlson (1978) in The Biology and Chemistry of Basement Membranes (Kefalides, N.A., ed.), Academic Press, New York; Kanwar & Farquhar (1979) Proc. Natl. Acad. Sci. U.S.A. 76, 4493-4497] to obtain a basement membrane preparation. The proteoglycans released at each stage of the procedure were characterized using DEAE-Sephacel ion-exchange chromatography, chondroitinase ABC and HNO2 digestion and Sepharose CL-4B gel-permeation chromatography. About 85% of the [35S]proteoglycans synthesized were of the heparan sulphate variety, the remainder being chondroitin sulphate proteoglycans. Three sizes of heparan sulphate proteoglycans were identified. The largest (HS1, Kav. 0.47) accounts for 44% of the total extractable heparan sulphates. About one third of HS1 were extracted from the glomerular basement-membrane fraction with 8 M-urea and 4 M-guanidine hydrochloride but the remainder were released from the glomerulus during preparation of the fraction. The two smaller molecules (HS2, Kav. 0.56 and HS3, Kav. 0.68) accounted for 27% and 28% of the extractable heparan sulphate respectively and were not associated with the basement membrane fraction. HS1, HS2 and HS3 were also isolated from non-fixed glomeruli labelled in vivo but with much lower recovery. In glomeruli labelled in vitro, heparan sulphate accounted for only 35% of the proteoglycans, the remainder being of the chondroitin sulphate type. Proteoglycans similar to HS1, HS2 and HS3 were present in glomeruli labelled in vitro but, in addition, a large, highly charged heparan sulphate (HS1a) was extracted from the glomerular basement-membrane fraction of these glomeruli. It accounted for 6% of the total heparan sulphate.

Synthesis of heparan sulfate proteoglycans by the isolated glomerulus

1988 ◽  
Vol 66 (10) ◽  
pp. 1078-1085 ◽  
Author(s):  
Douglas M. Templeton ◽  
Marina Khatchatourian
Keyword(s):  
Basement Membrane ◽  
Heparan Sulfate ◽  
Glomerular Basement Membrane ◽  
Time Course ◽  
In Vitro Model ◽  
Proteoglycan Synthesis ◽  
Proteoglycan Aggregates ◽  
Vitro Model

Incorporation of [35S]sulfate into newly synthesized macromolecules was studied in the isolated rat glomerulus and found to be linear between 6 and 24 h. When whole glomeruli were treated under conditions that dissociate proteoglycan aggregates, >90% of incorporated label was extracted. Of this, 80–90% was found to be the heparan sulfate proteoglycan. Similarly, a linear incorporation of [35S]sulfate into a glomerular basement membrane-enriched fraction was due almost entirely to proteoheparan sulfate. This predominance of heparan sulfate among the newly sulfated glycosaminoglycans has previously been observed in vivo and in the perfused kidney, but different patterns have hitherto been described in vitro. The present results suggest that under certain conditions, the isolated glomerulus is a suitable in vitro model for the study of proteoglycan synthesis. The pattern of incorporation of proteoglycans into the glomerular basement membrane reflects the time course and distribution of their synthesis by the whole glomerulus.

scholarly journals Anionic sites in the glomerular basement membrane. In vivo and in vitro localization to the laminae rarae by cationic probes.

1979 ◽  
Vol 81 (1) ◽  
pp. 137-153 ◽  
Author(s):  
Y S Kanwar ◽  
M G Farquhar
Keyword(s):  
Ionic Strength ◽  
Basement Membrane ◽  
Glomerular Basement Membrane ◽  
Left Kidney ◽  
High Ionic Strength ◽  
Basement Membranes ◽  
Mesangial Matrix ◽  
Anionic Sites

Cationized ferritin (CF) of narrow pI range (7.3-7.5) and the basic dye ruthenium red (RR) have been used as cationic probes to partially characterize anionic sites previously demonstrated in the glomerular basement membrane (GBM). When CF was given i.v. to normal rats and the left kidney was fixed by perfusion 15 min thereafter, clusters of CF molecules were found throughout the lamina rara interna (LRI), lamina rara externa (LRE), and mesangial matrix distributed at regular (approximately 60 nm) intervals. When kidneys were perfused with aldehyde fixative containing RR, small (20 nm) RR-stained particles were seen in the same locations distributed with the same 60 nm repeating pattern, forming a quasiregular, lattice-like arrangement. Fine (approximately 3 nm) filaments connected the sites and extended between them and the membranes of adjoining endothelial and epithelial cells. When CF was given i.v. followed by perfusion with RR in situ, both probes localized to the same sites. CF remained firmly bound after prolonged perfusion with 0.1-0.2 M KCl or NaCl. It was displaced by perfusion with buffers of high ionic strength (0.4-0.5 M KCl) or pH (less than 3.0 or greater than 10.0). CF also bound (clustered at approximately 60 nm intervals) to isolated GBM's, and binding was lost when such isolated GBM's were treated with buffers of high ionic strength or pH. These experiments demonstrate the existence of a quasi-regular, lattice-like network of anionic sites in the LRI and LRE and the mesangial matrix. The sites are demonstrable in vivo (by CF binding), in fixed kidneys (by RR staining), and in isolated GBM's (by CF binding). The results obtained with CF show that the binding of CF (and probably also RR) to the laminae rarae is electrostatic in nature since it is displaced by treatment with buffers of high ionic strength or pH. With RR the sites resemble in morphology and staining properties the proteoglycan particles found in connective tissue matrices and in association with basement membranes in several other locations.

scholarly journals Non-invasive skin sampling of tryptophan/kynurenine ratio in vitro towards a skin cancer biomarker

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Skaidre Jankovskaja ◽  
Johan Engblom ◽  
Melinda Rezeli ◽  
György Marko-Varga ◽  
Tautgirdas Ruzgas ◽  
...  
Keyword(s):  
Skin Cancer ◽  
Relative Increase ◽  
Cancer Biomarker ◽  
Cancer Diagnostics ◽  
Sampling Time ◽  
Skin Permeability ◽  
Experimental Conditions ◽  
Non Invasive

AbstractThe tryptophan to kynurenine ratio (Trp/Kyn) has been proposed as a cancer biomarker. Non-invasive topical sampling of Trp/Kyn can therefore serve as a promising concept for skin cancer diagnostics. By performing in vitro pig skin permeability studies, we conclude that non-invasive topical sampling of Trp and Kyn is feasible. We explore the influence of different experimental conditions, which are relevant for the clinical in vivo setting, such as pH variations, sampling time, and microbial degradation of Trp and Kyn. The permeabilities of Trp and Kyn are overall similar. However, the permeated Trp/Kyn ratio is generally higher than unity due to endogenous Trp, which should be taken into account to obtain a non-biased Trp/Kyn ratio accurately reflecting systemic concentrations. Additionally, prolonged sampling time is associated with bacterial Trp and Kyn degradation and should be considered in a clinical setting. Finally, the experimental results are supported by the four permeation pathways model, predicting that the hydrophilic Trp and Kyn molecules mainly permeate through lipid defects (i.e., the porous pathway). However, the hydrophobic indole ring of Trp is suggested to result in a small but noticeable relative increase of Trp diffusion via pathways across the SC lipid lamellae, while the shunt pathway is proposed to slightly favor permeation of Kyn relative to Trp.

Three-dimensional Growth of Renal Epithelial Cells in Vitro: A Tool in Toxicity Testing

1993 ◽  
Vol 21 (2) ◽  
pp. 191-195 ◽  
Author(s):  
Knut-Jan Andersen ◽  
Erik Ilsø Christensen ◽  
Hogne Vik
Keyword(s):  
Epithelial Cells ◽  
Three Dimensional ◽  
Toxicity Testing ◽  
Renal Tissue ◽  
Epithelial Cell Line ◽  
Multicellular Spheroids ◽  
Renal Epithelial Cell ◽  
Renal Epithelial Cells

The tissue culture of multicellular spheroids from the renal epithelial cell line LLC-PK1 (proximal tubule) is described. This represents a biological system of intermediate complexity between renal tissue in vivo and simple monolayer cultures. The multicellular structures, which show many similarities to kidney tubules in vivo, including a vectorial water transport, should prove useful for studying the potential nephrotoxicity of drugs and chemicals in vitro. In addition, the propagation of renal epithelial cells as multicellular spheroids in serum-free culture may provide information on the release of specific biological parameters, which may be suppressed or masked in serum-supplemented media.

scholarly journals Preclinical Testing of Radiopharmaceuticals for the Detection and Characterization of Osteomyelitis: Experiences from a Porcine Model

Molecules ◽  
2021 ◽  
Vol 26 (14) ◽  
pp. 4221
Author(s):  
Aage Kristian Olsen Alstrup ◽  
Svend Borup Jensen ◽  
Ole Lerberg Nielsen ◽  
Lars Jødal ◽  
Pia Afzelius
Keyword(s):  
Animal Model ◽  
Animal Models ◽  
Porcine Model ◽  
Animal Experiments ◽  
Radioactive Tracers ◽  
The Individual ◽  
Selection Of

The development of new and better radioactive tracers capable of detecting and characterizing osteomyelitis is an ongoing process, mainly because available tracers lack selectivity towards osteomyelitis. An integrated part of developing new tracers is the performance of in vivo tests using appropriate animal models. The available animal models for osteomyelitis are also far from ideal. Therefore, developing improved animal osteomyelitis models is as important as developing new radioactive tracers. We recently published a review on radioactive tracers. In this review, we only present and discuss osteomyelitis models. Three ethical aspects (3R) are essential when exposing experimental animals to infections. Thus, we should perform experiments in vitro rather than in vivo (Replacement), use as few animals as possible (Reduction), and impose as little pain on the animal as possible (Refinement). The gain for humans should by far exceed the disadvantages for the individual experimental animal. To this end, the translational value of animal experiments is crucial. We therefore need a robust and well-characterized animal model to evaluate new osteomyelitis tracers to be sure that unpredicted variation in the animal model does not lead to a misinterpretation of the tracer behavior. In this review, we focus on how the development of radioactive tracers relies heavily on the selection of a reliable animal model, and we base the discussions on our own experience with a porcine model.

scholarly journals Graphene and Reproduction: A Love-Hate Relationship

Nanomaterials ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 547
Author(s):  
Marina Ramal-Sanchez ◽  
Antonella Fontana ◽  
Luca Valbonetti ◽  
Alessandra Ordinelli ◽  
Nicola Bernabò ◽  
...  
Keyword(s):  
Graphene Oxide ◽  
Reproductive Function ◽  
Scientometric Analysis ◽  
Potential Toxicity ◽  
Good Tool ◽  
Vitro Fertilization ◽  
Number Of Publications ◽  
The Individual

Since its discovery, graphene and its multiple derivatives have been extensively used in many fields and with different applications, even in biomedicine. Numerous efforts have been made to elucidate the potential toxicity derived from their use, giving rise to an adequate number of publications with varied results. On this basis, the study of the reproductive function constitutes a good tool to evaluate not only the toxic effects derived from the use of these materials directly on the individual, but also the potential toxicity passed on to the offspring. By providing a detailed scientometric analysis, the present review provides an updated overview gathering all the research studies focused on the use of graphene and graphene-based materials in the reproductive field, highlighting the consequences and effects reported to date from experiments performed in vivo and in vitro and in different animal species (from Archea to mammals). Special attention is given to the oxidized form of graphene, graphene oxide, which has been recently investigated for its ability to increase the in vitro fertilization outcomes. Thus, the potential use of graphene oxide against infertility is hypothesized here, probably by engineering the spermatozoa and thus manipulating them in a safer and more efficient way.

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