scholarly journals PERMEABILITY OF MUSCLE CAPILLARIES TO EXOGENOUS MYOGLOBIN

1973 ◽  
Vol 57 (2) ◽  
pp. 424-452 ◽  
Author(s):  
Nicolae Simionescu ◽  
Maia Simionescu ◽  
George E. Palade
Keyword(s):  
Gel Filtration ◽  
Intercellular Junctions ◽  
Cell Periphery ◽  
Pore System ◽  
Regional Heterogeneity ◽  
Plasmalemmal Vesicles ◽  
Small Pore ◽  
Perinuclear Cytoplasm ◽  
Muscle Capillaries

Whale skeletal muscle myoglobin (mol wt 17,800; molecular dimensions 25 x 34 x 42 Å) was used as a probe molecule for the pore systems of muscle capillaries. Diaphragms of Wistar-Furth rats were fixed in situ at intervals up to 4 h after the intravenous injection of the tracer, and myoglobin was localized in the tissue by a peroxidase reaction. Gel filtration of plasma samples proved that myoglobin molecules remained in circulation in native monomeric form. At 30–35 s postinjection, the tracer marked ∼75% of the plasmalemmal vesicles on the blood front of the endothelium, 15% of those located inside and none of those on the tissue front. At 45 s, the labeling of vesicles in the inner group reached 60% but remained nil for those on the tissue front. Marked vesicles appeared on the latter past 45 s and their frequency increased to ∼80% by 60–75 s, concomitantly with the appearance of myoglobin in the pericapillary spaces. Significant regional heterogeneity in initial labeling was found in the different segments of the endothelium (i.e., perinuclear cytoplasm, organelle region, cell periphery, and parajunctional zone). Up to 60 s, the intercellular junctions and spaces of the endothelium were free of myoglobin reaction product; thereafter, the latter was detected in the distal part of the intercellular spaces in concentration generally equal to or lower than that prevailing in the adjacent pericapillary space. The findings indicate that myoglobin molecules cross the endothelium of muscle capillaries primarily via plasmalemmal vesicles. Since a molecule of this size is supposed to exit through both pore systems, our results confirm the earlier conclusion that the plasmalemmal vesicles represent the large pore system; in addition, they suggest that the same structures are, at least in part, the structural equivalent of the small pore system of this type of capillaries.

Plasmalemmal vesicles function as transcytotic carriers for small proteins in the continuous endothelium

1997 ◽  
Vol 272 (2) ◽  
pp. H937-H949 ◽  
Author(s):  
S. A. Predescu ◽  
D. N. Predescu ◽  
G. E. Palade
Keyword(s):  
Capillary Permeability ◽  
Pore System ◽  
Plasmalemmal Vesicles ◽  
Small Proteins ◽  
Structural Identity ◽  
Heart Perfusion ◽  
Small Pore ◽  
Vesicular Carriers ◽  
Tissue Fractionation

We investigated the location and the structural identity of the small pore system, postulated by the pore theory of capillary permeability, using a murine heart perfusion system and small protein molecules as preferential probes for the small pores. Dinitrophenylated proteins were perfused in situ in the absence and in the presence of N-ethylmaleimide (NEM), a reagent known to interfere with membrane fusion of vesicular carriers with their target membranes. The exit pathways of the tracers from vascular lumina to the interstitia were followed by immunoelectron microscopy and by tissue fractionation biochemistry to quantitate their transport and to estimate the extent of transport inhibition by NEM. After 5 min of perfusion, all tracers used were found essentially restricted to plasmalemmal vesicles (PVs) within the endothelium and NEM inhibited their transport by 80-85%. The transport of [14C]inulin and [14C]sucrose, assumed to follow the paracellular pathway, was marginally affected by NEM. These findings indicate that PVs function as structural equivalents of the small pore system for molecules >2 nm in diameter.

scholarly journals Permeability of muscle capillaries to small heme-peptides. Evidence for the existence of patent transendothelial channels.

1975 ◽  
Vol 64 (3) ◽  
pp. 586-607 ◽  
Author(s):  
N Simionescu ◽  
M Siminoescu ◽  
G E Palade
Keyword(s):  
Plasma Proteins ◽  
Intercellular Junctions ◽  
Rat Diaphragm ◽  
Enzymic Hydrolysis ◽  
Graphic Analysis ◽  
Heme Peptides ◽  
Future Work ◽  
Hydrolysis Of ◽  
Muscle Capillaries

Two heme-peptides (HP) of about 20-A diameter (heme-undecapeptide [H11P], mol wt approximately 1900 and heme-octapeptide [H8P], mol wt approximately 1550), obtained by enzymic hydrolysis of cytochrome c, were sued as probe molecules in muscle capillaries (rat diaphragm). They were localized in situ by a perixidase reaction, enhanced by the addition of imidazole to the incubation medium. Chromatography of plasma samples showed that HPs circulate predominantly as monomers for the duration of the experiments and are bound by aldehyde fixatives to plasma proteins to the extent of approximately 50% (H8P) to approximately 95% (H11P). Both tracers cross the endothelium primarily via plasmalemmal vesicles which become progressively labeled (by reaction product) from the blood front to the tissue front of the endothelium, in three successive resolvable phases. By the end of each phase the extent of labeling reaches greater than 90% of the corresponding vesicle population. Labeled vesicles appear as either isolated units or chains which form patent channels across the endothelium. The patency of these channels was checked by specimen tilting and graphic analysis of their images. No evidence was found for early or preferential marking of the intercellular junctions and spaces by reaction product. It is concluded that the channels are the most likely candidate for structural equivalents of the small pores of the capillary wall since they are continuous, water-filled passages, and are provided with one or more strictures of less than 100 A. Their frequency remains to be established by future work.

scholarly journals Binding and transcytosis of glycoalbumin by the microvascular endothelium of the murine myocardium: evidence that glycoalbumin behaves as a bifunctional ligand.

1988 ◽  
Vol 107 (5) ◽  
pp. 1729-1738 ◽  
Author(s):  
D Predescu ◽  
M Simionescu ◽  
N Simionescu ◽  
G E Palade
Keyword(s):  
Intercellular Junctions ◽  
Concomitant Administration ◽  
Multivesicular Bodies ◽  
Microvascular Endothelium ◽  
Coated Pits ◽  
Gold Complexes ◽  
Plasmalemmal Vesicles ◽  
Bifunctional Ligand ◽  
Albumin Molecule

The binding and transport of glycoalbumin (gA) by the endothelium of murine myocardial microvessels were studied by perfusing in situ 125I-gA or gA-gold complexes (gA-Au) and examining the specimens by radioassays and EM, respectively. After a 3-min perfusion, the uptake of radioiodinated gA is 2.2-fold higher than that of native albumin; it is partially (approximately 55%) competed by either albumin or D-glucose, and almost completely abolished by the concomitant administration of both competitors or by gA. D-mannose and D-galactose are not effective competitors. Unlike albumin-gold complexes that bind restrictively to plasmalemmal vesicles, gA-Au labels the plasma-lemma proper, plasmalemmal vesicles open on the lumen, and most coated pits. Competing albumin prevents gA-Au binding to the membrane of plasmalemmal vesicles, while glucose significantly reduces the ligand binding to plasmalemma proper. Competition with albumin and glucose gives additive effects. Transcytosis of gA-Au, already detected at 3 min, becomes substantial by 30 min. No tracer exit via intercellular junctions was detected. gA-Au progressively accumulates in multivesicular bodies. The results of the binding and competition experiments indicate that the gA behaves as a bifunctional ligand which is recognized by two distinct binding sites: one, located on the plasma membrane, binds as a lectin the glucose residues of gA; whereas the other, confined to plasmalemmal vesicles, recognizes presumably specific domains of the albumin molecule.

scholarly journals INTESTINAL CAPILLARIES

1969 ◽  
Vol 41 (1) ◽  
pp. 33-58 ◽  
Author(s):  
F. Clementi ◽  
G. E. Palade
Keyword(s):  
Electron Microscopy ◽  
Horseradish Peroxidase ◽  
Intestinal Mucosa ◽  
Intercellular Junctions ◽  
Post Injection ◽  
Pore System ◽  
Blood Capillaries ◽  
Large Pore ◽  
Small Pore ◽  
Histochemical Procedure

Horseradish peroxidase (mol. diam. ≃50 A) and ferritin (mol. diam. ≃110 A) were used as probe molecules for the small and large pore system, respectively, in blood capillaries of the intestinal mucosa of the mouse. Peroxidase distribution was followed in time, after intravenous injection, by applying the Graham-Karnovsky histochemical procedure to aldehyde-fixed specimens. The tracer was found to leave the plasma rapidly and to reach the pericapillary spaces 1 min post injection. Between 1 min and 1 min 30 sec, gradients of peroxidase reaction product could be demonstrated regularly around the capillaries; their highs were located opposite the fenestrated parts of the endothelium. These gradients were replaced by even distribution past 1 min 30 sec. Ferritin, followed directly by electron microscopy, appeared in the pericapillary spaces 3–4 min after i.v. injection. Like peroxidase, it initially produced transient gradients with highs opposite the fenestrated parts of the endothelium. For both tracers, there was no evidence of movement through intercellular junctions, and transport by plasmalemmal vesicles appeared less efficient than outflow through fenestrae. It is concluded that, in the blood capillaries of the inintestinal mucosa, the diaphragms of the endothelial fenestrae contain the structural equivalents of the small pore system. The large pore system seems to be restricted to a fraction of the fenestral population which presumably consists of diaphragm-free or diaphragm-deficient units.

Plasmalemmal vesicles represent the large pore system of continuous microvascular endothelium

1993 ◽  
Vol 265 (2) ◽  
pp. H725-H733 ◽  
Author(s):  
D. Predescu ◽  
G. E. Palade
Keyword(s):  
Intercellular Junctions ◽  
Pore System ◽  
Microvascular Endothelium ◽  
Plasmalemmal Vesicles ◽  
Morphological Studies ◽  
Large Pore ◽  
In Transit

In the capillary physiology literature, molecules and particles larger than 10 nm are assumed to leave the plasma mostly through large pores located at the level of intercellular junctions in microvessels lined with a continuous endothelium. In morphological studies of similar microvessels, outgoing particles > 10 nm were detected in endothelial plasmalemmal vesicles not in intercellular junctions. Because the probes may not be found in transit through the junctions because they may be swept away by strong currents generated by Starling forces, we have examined a large number of junctions in arteriolar, capillary, and venular segments of bipolar vascular fields of mouse diaphragms collected after perfusion with large pore probes. The results presented in this study indicate that 1) the perfused probes accumulate in the luminal introits of the junctions as filtration residues that decrease in size and frequency from arterioles to venules, and 2) large pore probes move across the endothelium exclusively through plasmalemmal vesicles.

scholarly journals PERMEABILITY OF INTESTINAL CAPILLARIES

1972 ◽  
Vol 53 (2) ◽  
pp. 365-392 ◽  
Author(s):  
Nicolae Simionescu ◽  
Maia Simionescu ◽  
George E. Palade
Keyword(s):  
Intercellular Junctions ◽  
Liver Glycogen ◽  
Pore System ◽  
Concentration Gradients ◽  
Good Definition ◽  
Plasmalemmal Vesicles ◽  
Tracer Particles ◽  
One Step ◽  
Dynamic Structures ◽  
Capillary Walls

The pathway followed by macromolecules across the wall of visceral capillaries has been studied by using a set of tracers of graded sizes, ranging in diameter from 100 A (ferritin) to 300 A (glycogen). Polysaccharide particles, i.e. dextran 75 (mol wt ∼75,000; diam ∼125 A), dextran 250 (mol wt 250,000; diam ∼225 A), shellfish glycogen (diam ∼200 A) and rabbit liver glycogen (diam ∼300 A), are well tolerated by Wistar-Furth rats and give no vascular reactions ascribable to histamine release. Good definition and high contrast of the tracer particles were obtained in a one-step fixation—in block staining of the tissues by a mixture containing aldehydes, OsO4 and lead citrate in phosphate or arsenate buffer, pH 7.4, followed by lead staining of sections. The glycogens and dextrans used move out of the plasma through the fenestrae and channels of the endothelium relatively fast (3–7 min) and create in the pericapillary spaces transient (2–5 min) concentration gradients centered on the fenestrated sectors of the capillary walls. The tracers also gained access to the plasmalemmal vesicles, first on the blood front and subsequently on the tissue front of the endothelium. The particles are temporarily retained by the basement membrane. No probe moved through the intercellular junctions. It is concluded that, in visceral capillaries, the fenestrae, channels, and plasmalemmal vesicles, viewed as related parts in a system of dynamic structures, are the structural equivalent of the large pore system.

CTAB-assisted size controlled synthesis of SAPO-34 and its contribution toward MTO performance

2018 ◽  
Vol 47 (29) ◽  
pp. 9861-9870 ◽  
Author(s):  
Syed ul Hasnain Bakhtiar ◽  
Xiaotong Wang ◽  
Sher Ali ◽  
Fulong Yuan ◽  
Zhibin Li ◽  
...  
Keyword(s):  
Controlled Synthesis ◽  
Pore System ◽  
Light Olefin ◽  
Diffusion Limitations ◽  
Small Pore ◽  
Methanol To Olefin ◽  
Olefin Selectivity ◽  
Size Controlled

SAPO-34 shows higher light olefin selectivity in the reaction of methanol to olefin (MTO), but its small pore system implies diffusion limitations to bigger molecular products and results in coking too.

Direct visualization of a vast cortical calcium compartment in Paramecium by secondary ion mass spectrometry (SIMS) microscopy: possible involvement in exocytosis

1995 ◽  
Vol 108 (5) ◽  
pp. 1895-1909 ◽  
Author(s):  
N. Stelly ◽  
S. Halpern ◽  
G. Nicolas ◽  
P. Fragu ◽  
A. Adoutte
Keyword(s):  
Mass Spectrometry ◽  
Secondary Ion Mass Spectrometry ◽  
Membrane Vesicles ◽  
Cell Periphery ◽  
A Value ◽  
External Reference ◽  
Ion Mass Spectrometry ◽  
First Time ◽  
Secondary Ion

The plasma membrane of ciliates is underlaid by a vast continuous array of membrane vesicles known as cortical alveoli. Previous work had shown that a purified fraction of these vesicles actively pumps calcium, suggesting that alveoli may constitute a calcium-storage compartment. Here we provide direct confirmation of this hypothesis using in situ visualization of total cell calcium on sections of cryofixed and cryosubstituted cells analyzed by SIMS (secondary ion mass spectrometry) microscopy a method never previously applied to protists. A narrow, continuous, Ca-emitting zone located all along the cell periphery was observed on sections including the cortex. In contrast, Na and K were evenly distributed throughout the cell. Various controls confirmed that emission was from the alveoli, in particular, the emitting zone was still seen in mutants totally lacking trichocysts, the large exocytotic organelles docked at the cell surface, indicating that they make no major direct contribution to the emission. Calcium concentration within alveoli was quantified for the first time in SIMS microscopy using an external reference and was found to be in the range of 3 to 5 mM, a value similar to that for sarcoplasmic reticulum. After massive induction of trichocyst discharge, this concentration was found to decrease by about 50%, suggesting that the alveoli are the main source of the calcium involved in exocytosis.

scholarly journals Visualization of mRNA translation in living cells

2006 ◽  
Vol 175 (1) ◽  
pp. 67-76 ◽  
Author(s):  
Alexis J. Rodriguez ◽  
Shailesh M. Shenoy ◽  
Robert H. Singer ◽  
John Condeelis
Keyword(s):  
Mrna Translation ◽  
Leading Edge ◽  
Intercellular Junctions ◽  
Mrna Localization ◽  
Living Cells ◽  
C2c12 Cells ◽  
Cell Periphery ◽  
Actin Mrna ◽  
Cell Asymmetry ◽  
Cellular Compartments

The role of mRNA localization is presumably to effect cell asymmetry by synthesizing proteins in specific cellular compartments. However, protein synthesis has never been directly demonstrated at the sites of mRNA localization. To address this, we developed a live cell method for imaging translation of β-actin mRNA. Constructs coding for β-actin, containing tetracysteine motifs, were transfected into C2C12 cells, and sites of nascent polypeptide chains were detected using the biarsenial dyes FlAsH and ReAsH, a technique we call translation site imaging. These sites colocalized with β-actin mRNA at the leading edge of motile myoblasts, confirming that they were translating. β-Actin mRNA lacking the sequence (zipcode) that localizes the mRNA to the cell periphery, eliminated the translation there. A pulse-chase experiment on living cells showed that the recently synthesized protein correlated spatially with the sites of its translation. Additionally, localization of β-actin mRNA and translation activity was enhanced at cell contacts and facilitated the formation of intercellular junctions.

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