Mapping the Distinctive Populations of Lymphatic Endothelial Cells in Different Zones of Human Lymph Nodes

The lymphatic sinuses in human lymph nodes (LNs) are crucial to LN function yet their structure remains poorly defined. Much of our current knowledge of lymphatic sinuses derives from rodent models, however human LNs differ substantially in their sinus structure, most notably due to the presence of trabeculae and trabecular lymphatic sinuses that rodent LNs lack. Lymphatic sinuses are bounded and traversed by lymphatic endothelial cells (LECs). A better understanding of LECs in human LNs is likely to improve our understanding of the regulation of cell trafficking within LNs, now an important therapeutic target, as well as disease processes that involve lymphatic sinuses. We therefore sought to map all the LECs within human LNs using multicolor immunofluorescence microscopy to visualize the distribution of a range of putative markers. PROX1 was the only marker that uniquely identified the LECs lining and traversing all the sinuses in human LNs. In contrast, LYVE1 and STAB2 were only expressed by LECs in the paracortical and medullary sinuses in the vast majority of LNs studied, whilst the subcapsular and trabecular sinuses lacked these molecules. These data highlight the existence of at least two distinctive populations of LECs within human LNs. Of the other LEC markers, we confirmed VEGFR3 was not specific for LECs, and CD144 and CD31 stained both LECs and blood vascular endothelial cells (BECs); in contrast, CD59 and CD105 stained BECs but not LECs. We also showed that antigen-presenting cells (APCs) in the sinuses could be clearly distinguished from LECs by their expression of CD169, and their lack of expression of PROX1 and STAB2, or endothelial markers such as CD144. However, both LECs and sinus APCs were stained with DCN46, an antibody commonly used to detect CD209.     

Role of solvent properties of aqueous media in macromolecular crowding effects

Analysis of the macromolecular crowding effects in polymer solutions show that the excluded volume effect is not the only factor affecting the behavior of biomolecules in a crowded environment. The observed inconsistencies are commonly explained by the so-called soft interactions, such as electrostatic, hydrophobic, and van der Waals interactions, between the crowding agent and the protein, in addition to the hard nonspecific steric interactions. We suggest that the changes in the solvent properties of aqueous media induced by the crowding agents may be the root of these “soft” interactions. To check this hypothesis, the solvatochromic comparison method was used to determine the solvent dipolarity/polarizability, hydrogen-bond donor acidity, and hydrogen-bond acceptor basicity of aqueous solutions of different polymers (dextran, poly(ethylene glycol), Ficoll, Ucon, and polyvinylpyrrolidone) with the polymer concentration up to 40% typically used as crowding agents. Polymer-induced changes in these features were found to be polymer type and concentration specific, and, in case of polyethylene glycol (PEG), molecular mass specific. Similarly sized polymers PEG and Ucon producing different changes in the solvent properties of water in their solutions induced morphologically different α-synuclein aggregates. It is shown that the crowding effects of some polymers on protein refolding and stability reported in the literature can be quantitatively described in terms of the established solvent features of the media in these polymers solutions. These results indicate that the crowding agents do induce changes in solvent properties of aqueous media in crowded environment. Therefore, these changes should be taken into account for crowding effect analysis.     

Switchable MRI contrast agents based on morphological changes of pH-responsive polymers

Magnetic resonance imaging (MRI) contrast agents are effective tools in both medical diagnosis and life science research. Various smart contrast agents have been developed for the visualization of biological phenomena. These contrast agents have molecular switches that increase or reduce MRI signal intensity in response to the target biological reaction. Therefore, novel approaches to the design of molecular switches for versatile in vivo studies using MRI are eagerly anticipated. Here, we report one such approach for the development of molecular switches based on morphological changes of pH-responsive polymers. We designed and synthesized three types of contrast agents based on a linear homopolymer or spherical copolymers with two different cross-linking degrees. The relaxivity measurements showed that these agents have molecular switches that respond to pH changes, and fluorescence studies indicated that these switches are based on the alteration of the molecular tumbling caused by pH-responsive morphological changes. As a result, the spherical polymers possess promising characteristics for the development of switchable MRI contrast agents.     

Probing the limits of molecular imprinting: strategies with a template of limited size and functionality

A series of polymers molecularly imprinted with the general anaesthetic propofol were synthesized using both semi- and non-covalent approaches. The polymers were evaluated with respect to template rebinding in both aqueous and organic media. In aqueous media, the observed propofol binding in these polymer systems was largely hydrophobic and non-specific in nature. In non-polar solvents such as hexane, electrostatic (hydrogen bonding) interactions dominate resulting in some selectivity. The implication of these results, in conjunction with those obtained using structures of similar size in other studies, is that propofol, a template possessing limited functionality and size, appears to define the lower limit for template size and degree of functionalization that can be used for the creation of ligand-selective recognition sites in molecularly imprinted polymers. Furthermore, studies with alternative ligands indicate that the steric crowding of a ligand's functionality to the polymer contributes to the extent of polymer-ligand recognition.     

Significance of zinc in nephrotoxicity of contrast media used in imaging diagnostics of the cardiovascular system

Nephrotoxicity is an undesirable reaction of contrast media used in X-ray or magnetic resonance diagnostics. In addition to a direct toxic effect on renal tubules, the hemodynamic factor is considered to be the main cause of kidney damage and malfunction. The factors that increase the probability of a nephrotoxic effect of contrast media include oldage, diabetes, arterial hypertension, circulatory system insufficiency, neoplastic diseases, and prior kidney damage. Decreased serum zinc is observed in all those conditions. In this article, the influence of contrast media on zinc homeostasis and the possibility of a nephrotoxic reaction caused by these agents is discussed.     

Synthesis and characterization of poly(L-glutamic acid) gadolinium chelate: a new biodegradable MRI contrast agent

Most currently evaluated macromolecular contrast agents for magnetic resonance imaging (MRI) are not biodegradable. The goal of this study is to synthesize and characterize poly(l-glutamic acid) (PG) gadolinium chelates as biodegradable blood-pool MRI contrast agents. Two PG chelates of gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA) were synthesized through the use of difunctional and monofunctional DTPA precursors. The conjugates were characterized with regard to molecular weight and molecular weight distribution, gadolinium content, relaxivity, and degradability. Distributions of the polymeric MRI contrast agents in various organs were determined by intravenous injection of (111)In-labeled polymers into mice bearing murine breast tumors. MRI scans were performed at 1.5 T in mice after bolus injection of the polymeric chelates. PG-Hex-DTPA-Gd, obtained from aminohexyl-substituted PG and DTPA-dianhydride, was partially cross-linked and was undegradable in the presence of cathepsin B. On the other hand, PG-Bz-DTPA-Gd synthesized directly from PG and monofunctional p-aminobenzyl-DTPA(acetic acid-tert-butyl ester) was a linear polymer and was degradable. The relaxivities of the polymers at 1.5 T were 3-8 times as great as that of Gd-DTPA. Both polymers had high blood concentrations and were primarily accumulated in the kidney. However, PG-Bz-DTPA-Gd was gradually cleared from the body and had significantly less retention in the blood, the spleen, and the kidney. MRI with PG-Bz-DTPA-Gd in mice showed enhanced vascular contrast at up to 2 h after the contrast agent injection. The ability of PG-Bz-DTPA-Gd to be degraded and cleared from the body makes it a favorable macromolecular MRI contrast agent.     

Histology of the caprine hemal node

Caprine hemal nodes were studied by light microscopy after glutaraldehyde fixation and epoxy resin embedding. A node consisted of a capsule, subcapsular and other sinuses, cortex, medulla and hilus. Elements of circulating blood filled the interstices of the reticular meshwork and associated macrophages which traversed the lumina of subcapsular and medullary sinuses. The latter were rare in 1-month-old goats, progressively increased in number and size in 2- to 4-month-old goats and coalesced with each other and the subcapsular sinus in adult animals. The cortical tissue appeared as lymphoid nodules. Circumferential lymphatic vessels abutted on outer margins of the nodules and gave origin to several radial lymphatics which branched and anastomosed between the medullary blood sinuses. Medullary cords were organized around the radial lymphatics. A single efferent lymphatic was formed at the hilum by confluence of the radial lymphatics. Our study, in contrast to earlier reports, shows that caprine hemal nodes possess efferent lymphatics. The present data suggest that the hemal nodes are involved, in addition to classical functions, in blood storage by hemoconcentration.     

Studies on the stimulation of cAMP metabolism in human lymphocytes by latex polymers

The purpose of this investigation was to further characterize the marked increase in intracellular cAMP which follows the interaction of human lymphocytes and latex polymers. Six distinct cell types, each of which either bind or ingest these latex particles, were studied; however, only lymphocytes responded with increases in intracellular cAMP. The initial attachment of the latex particles to the lymphocyte surface was independent of temperature, cyclic nucleotides and divalent cations in the external milieu. The subsequent cAMP response was maximal at physiologic temperatures and modulated by agents thought to alter microfilament and microtubule function. Four different types of polymers produced increases in intralymphocytic cAMP and the maximal increases were confined to particles having a mean diameter of 0.4–2.02 μm. Within this latter size range, there was a close correlation between the number of membrane-associated particles and the magnitude of the cAMP response. Similarities to the lymphocyte-lectin activation system included:

1. 1. A requirement for binding of the latex polymers to the external plasma membrane.

2. 2. A biphasic cAMP response characterized by an early rise followed by a later fall.

3. 3. Modulation of this response by pharmacologic agents which compromise microtubule and microfilament function.

In contrast to the lectin-induced activation, latex beads inhibited amino acid transport and phosphatidylinositol turnover and did not lead to later increases in DNA synthesis. These data suggest that latex polymers attach to receptors on the plasma membrane different from those responsible for lymphocyte activation, and through cAMP induce metabolic responses dissimilar to those associated with lectin activation.     

The effect of the presence of globular proteins and elongated polymers on enzyme activity

We have studied the effect of a crowded (macromolecular) solution on reaction rates of the decarboxylating enzymes urease, pyruvate decarboxylase and glutamate decarboxylase. A variety of crowding agents were used including haemoglobin, lysozyme, various dextrans and polyethylene glycol. Enzyme reaction rates of all three enzymes show two different types of effect that separate the globular proteins from the polysaccharides/polymers. Increasing concentration of globular proteins caused a dramatic rise in enzyme activity up to 30% crowding concentration then the activity decreased, whereas the polymers caused a concentration dependent decrease in activity. The viscosities of the globular proteins were low compared to the polymers. The increased activity with proteins may be due to the decreased amount of free water, which leads to higher effective concentration of substrates, or to an increased oligomeric state by self-association. The lower activities of all enzymes with all agents at high concentrations may be explained by a decrease in rates of diffusion. An increase in protein crowding (decrease in cell water content) may contribute to pathological conditions including cataract and Alzheimer's disease.     

Risk-group patients due to the administration of iodine and gadolinium preparations for diagnostic clinical study

The paper discusses the concept of risk-group patients who have an increased probability of immediate systemic adverse reactions to contrast agents used clinically in various radiodiagnostic techniques. It analyzes the pathogenesis of allergoid reactions to X-ray and magnetic resonance, contrast media. On this basis, the authors describe the abnormalities and nosological entities the patients had in their medical history before diagnostic studies will be indicative of the increased risk of these complications to the administration of a contrast agent.     

Effects of different gelling agents on the different stages of rice regeneration in two rice cultivars

Plant tissue culture technology offers a solution for meeting the increasing commercial demand on economically important plants such as rice, a widespread dietary staple. However, significant genotype-specific morphogenetic responses constitute a considerable on rice regeneration in plant biotechnology contexts. Aside from genotype dependency, the components of the nutrient media including gelling agents have an important impact on regeneration efficiency. The current study explores the effect of different gelling agents on various stages of rice regeneration in two Egyptian rice cultivars-Sakha104 and Giza178. Media solidified with varying concentrations of a variety of gelling agents (agar, bacto agar, gelrite and phytagel) were tested for their impact on the frequency of callus induction, shoot regeneration and rooting. The results indicated gellan gum (gelrite and phytagel) was superior to agar products (agar and bacto agar) for callus induction. By contrast, no significant differences were found between different gelling agents for shoot regeneration. Gellan gum and media solidified with bacto agar were found to lead to significantly higher root regeneration than agar. The Sakha104 cultivar showed better responses than Giza 178 for callus induction and similar performance to the Giza 178 cultivar for root regeneration irrespective of the gelling agent. This work provides insights into the impact of different gelling agents on the morphogenetic response of two rice cultivars and can be used to help maximize the frequency of rice regeneration.     

Three-dimensional reconstruction of the human capillary network and the intramyocardial micronecrosis

Three-dimensional reconstruction of the human heart was performed to define the structure of the intramyocardial microvasculature. A total of 200 consecutive serial sections of 6 μm each were prepared from the left ventricular tissue of an autopsied human heart with normal coronary arteries. The corresponding arteriole, venule, and all capillaries were reconstructed using three-dimensional software. The capillary network extended right and left along the cardiomyocyte with major and minor axes of about 130 and 120 μm, respectively. The capillary length from an arteriole to an adjacent venule was about 350 μm. Two types of sack-like structures, the precapillary sinus and the capillary sinus, were present in the capillary network, and many capillaries diverged from these sinuses. The cardiomyocytes were covered with reticular capillaries. In contrast, the precapillary and capillary sinuses were surrounded by many cardiomyocytes. The arterial and venous capillaries were positioned alternately, forming a lattice pattern. Intramyocardial microcirculatory units forming a capillary network from an arteriole to adjacent venules on both sides were present. The sizes of myocardial micronecroses corresponded to that of the intramyocardial microcirculatory unit. These results show that the capillary network is an ordered and anatomically regulated structure and that the microcirculatory unit and the precapillary and capillary sinuses may play an important role in maintaining the intramyocardial microcirculation during contraction and relaxation.     

Osmotic perturbations induce differential movements in the core and periphery of proteins, membranes and micelles

Polymeric structures, namely, micelles, membranes and globular proteins share the property of two distinct regions: a hydrophobic core and a hydrophilic exterior. The dynamics of these regions of the polymeric structures were probed using selective fluorophores 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-anilinonaphthalene-8-sulfonate (ANS), respectively. Perturbation of the polymers by external osmotic pressure, ionic strength and temperature was monitored in the two regions using steady state measurements of fluorescence intensity and anisotropy. While the fluorescence lifetime of DPH and ANS did not change significantly, parallel change in steady state anisotropy values and the rotational correlation time indicated mobility in the probe/probe-domain. Osmotic perturbation of the polymers in electrolyte media led to decreased DPH mobility. Enhanced ellipticity at 222 nm in bovine serum albumin was observed in 1.5 M NaCl and sucrose media. ANS exhibited a decreased anisotropy with progressive dehydration in proteins in NaCl media, in dimyristoylphosphatidylcholine (DMPC) vesicles in sucrose media, and in neutral laurylmaltoside micelles in both NaCl and sucrose media. Thus, ANS showed responses opposite to that of DPH in these systems. A comparison with several domain selective probes indicated that DPH reported findings common to depth probes while ANS reported data common to interfacial probes used for voltage monitoring.     

非水介质中辣根过氧化物酶（HRP）荧光活性测定法

建立了一种分析ＨＲＰ催化活力的新方法。该方法基于单体（底物）、聚合物（产物）的荧光发射光谱不重叠,使用荧光光谱仪,通过测量底物荧光淬灭来检测ＨＲＰ在非水介质中（二氧六环－水、乙醇－水、丙酮－水体系）催化酚类、芳香胺类物质聚合的活力。此方法迅速、简便,结果是定量并可重复的,并能定量地计算底物转化率。     

Differential uptake of MRI contrast agents indicates charge-selective blood-brain interface in the crayfish

This study provides a new perspective on the long-standing problem of the nature of the decapod crustacean blood-brain interface. Previous studies of crustacean blood-brain interface permeability have relied on invasive histological, immunohistochemical and electrophysiological techniques, indicating a leaky non-selective blood-brain barrier. The present investigation involves the use of magnetic resonance imaging (MRI), a method for non-invasive longitudinal tracking of tracers in real-time. Differential uptake rates of two molecularly distinct MRI contrast agents, namely manganese (Mn(II)) and Magnevist? (Gd-DTPA), were observed and quantified in the crayfish, Cherax destructor. Contrast agents were injected into the pericardium and uptake was observed with longitudinal MRI for approximately 14.5?h. Mn(II) was taken up quickly into neural tissue (within 6.5?min), whereas Gd-DTPA was not taken up into neural tissue and was instead restricted to the intracerebral vasculature or excreted into nearby sinuses. Our results provide evidence for a charge-selective intracerebral blood-brain interface in the crustacean nervous system, a structural characteristic once considered too complex for a lower-order arthropod.     

无胶筛分毛细管电泳分离DNA片段的现状与展望

毛细管电泳已DNA片段分离分析的重要手段。本简述了毛细管电泳中采用无胶筛分介质分离DNA片段的机理研究,介绍了筛分介质近年的研究发展状况,依据分离介质的化学组成,分单聚物、共聚物和混聚物等3个部分进行了评述,并对其发展前景进行了展望。     

Comparative Histochemical Analysis of Cell Wall Polysaccharides by Enzymatic and Chemical Extractions of Two Fruits

A protocol for extracting polysaccharides from cell walls has been modified and used to analyze histochemically two fruits with opposite characteristics. Grapes are nonclimacteric fruits and are harvested at full maturity. In contrast, kiwi fruits are climacteric and are harvested and consumed before they are physiologically mature. The two fruits were analyzed histochemically using two protocols. One method is defined as chemical, and is based on subsequential extractions of pectins by chemical agents. The other is defined as enzymatic because it removes pectins using pectinase followed by hot ammonium oxalate. In both protocols, two types of hemicellulosic polymers are removed by 1 M and 4 M/KOH leaving a cellulosic residue on the slide. Both protocols remove the same amount of pectins, thus confirming their precision. The sum of hemicellulose and the cellulosic insoluble residue are equivalent using the two methods, but the relative amounts of the cellulose and hemicellulosic polymers were dependent upon the method of extraction. When the enzyme was used to extract the pectins, there was less cellulose and more hemicellulose. The removal of polysaccharides by ammonium oxalate and by guanidinethio-cyanate in the enzymatic and the chemical protocols, respectively, yielded approximately the same amount of removed material.

Similar results were obtained from both fruits. Grape, being softer than kiwi fruit, was relatively richer in pectic substances and less rich in hemicellulose and cellulose polymers. No difference in cell wall material could be ascribed to the different ripening habits.     

Comparative Histochemical Analysis of Cell Wall Polysaccharides by Enzymatic and Chemical Extractions of Two Fruits

A protocol for extracting polysaccharides from cell walls has been modified and used to analyze histochemically two fruits with opposite characteristics. Grapes are nonclimacteric fruits and are harvested at full maturity. In contrast, kiwi fruits are climacteric and are harvested and consumed before they are physiologically mature. The two fruits were analyzed histochemically using two protocols. One method is defined as chemical, and is based on subsequential extractions of pectins by chemical agents. The other is defined as enzymatic because it removes pectins using pectinase followed by hot ammonium oxalate. In both protocols, two types of hemicellulosic polymers are removed by 1 M and 4 M/KOH leaving a cellulosic residue on the slide. Both protocols remove the same amount of pectins, thus confirming their precision. The sum of hemicellulose and the cellulosic insoluble residue are equivalent using the two methods, but the relative amounts of the cellulose and hemicellulosic polymers were dependent upon the method of extraction. When the enzyme was used to extract the pectins, there was less cellulose and more hemicellulose. The removal of polysaccharides by ammonium oxalate and by guanidinethio-cyanate in the enzymatic and the chemical protocols, respectively, yielded approximately the same amount of removed material.

Similar results were obtained from both fruits. Grape, being softer than kiwi fruit, was relatively richer in pectic substances and less rich in hemicellulose and cellulose polymers. No difference in cell wall material could be ascribed to the different ripening habits.     

Morphology, constraints, and scaling of frontal sinuses in the hartebeest, Alcelaphus buselaphus (Mammalia: Artiodactyla, Bovidae)

The frontal sinuses of bovid mammals display a great deal of diversity, which has been attributed to both phylogenetic and functional influences. In-depth study of the hartebeest (Alcelaphus buselaphus), a large African antelope, reveals a number of previously undescribed details of frontal sinus morphology. In A. buselaphus, the frontal sinuses conform closely to the shape of the frontal bone, filling nearly the entire element. However, the horncores are never extensively pneumatized, contrasting with the condition seen in many other bovids. This evidence is inconsistent with the hypothesis that sinuses are opportunistic pneumatizing agents, suggesting that phylogenetic factors also play a role in determining sinus size. Both cranial sutures and neurovasculature appear to constrain the growth of sinuses in part. In turn, the sinus also affects the growth of the parietal; apparently this element is not truly pneumatized by the sinus in most cases, but the bone's shape changes under the influence of the sinus. Furthermore, the sinuses present relatively few struts when compared with the sinuses of some other bovids, such as Ovis. By adapting methods previously developed for measuring structural parameters of trabecular bone, it is possible to quantify certain aspects of sinus morphology. These include the number of bony struts within the sinus, the spacing of these struts, and the size of individual cavities within the sinus. Some differences in the number of struts are evident between subspecies. Similarly, significant differences occur in the relative number of struts between male and female A. buselaphus, which may be related to behavior. The volume of the sinus is strongly correlated with the size of the frontal, but less so with overall cranial size. This finding illustrates the importance of choosing variables carefully when comparing sinus sizes and growth between species.     

Personalized application of three different concentrations of iodinated contrast media in coronary computed tomography angiography

No study has evaluated the impact of different iodinated contrast media on coronary contrast enhancement, using an injection protocol according to body surface area (BSA). Thus, the present study aimed to examine the usefulness and safety of personalized application of different iodine concentrations of contrast media in coronary computed tomographic (CT) angiography with a 2nd dual-source CT scanner in eliminating differences in coronary contrast enhancement based on a BSA-adapted injection protocol of contrast media. A total of 270 enrolled participants were randomly assigned to three groups: ioversol 320, ioversol 350 and iopromide 370 (n = 90 per group). The three groups were administered contrast media at a BSA-adjusted volume and flow rate with a fixed injection time of 15 seconds, and they subsequently received a 30-mL saline flush. All patients were scanned with a prospective electrocardiogram-gated protocol in a craniocaudal direction using a second-generation 128-slice dual-source CT system. The three iodinated contrast media used in coronary CT angiography exhibited similar diagnostic quality and safety. No significant differences were found in the contrast enhancement degrees, image quality scores, radiation doses and incidences of adverse effects among the three groups. The three contrast media used in coronary CT angiography with 320, 350 and 370 mg/mL iodine, respectively, have comparable diagnostic quality and safety. However, more large-scale, multinational, multi-centre and prospective trials are warranted.