scholarly journals Heparin Inhibits Leukocyte Rolling in Pial Vessels and Attenuates Inflammatory Changes in a Rat Model of Experimental Bacterial Meningitis

1997 ◽  
Vol 17 (11) ◽  
pp. 1221-1229 ◽  
Author(s):  
Joerg R. Weber ◽  
Klemens Angstwurm ◽  
Thomas Rosenkranz ◽  
Ute Lindauer ◽  
Dorette Freyer ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Bacterial Meningitis ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Leukocyte Rolling ◽  
Central Nervous System Inflammation ◽  
Inflammatory Processes

Heparin is a natural proteoglycan that was first described in 1916. In addition to its well characterized effect on blood coagulation, it is becoming clear that heparin also modulates inflammatory processes on several levels, including the interference with leukocyte–endothelium interaction. Anecdotal observations suggest a better clinical outcome of heparin-treated patients with bacterial meningitis. The authors demonstrate that heparin, a glycosaminoglycan, inhibits significantly in the early phase of experimental pneumococcal meningitis the increase of 1) regional cerebral blood flow (125 ± 18 versus 247 ± 42%), 2) intracranial pressure (4.5 ± 2.0 versus 12.1 ± 2.2 mm Hg), 3) brain edema (brain water content: 78.23 ± 0.33 versus 79.49 ± 0.46%), and 4) influx of leukocytes (571 ± 397 versus 2400 ± 875 cells/μL) to the cerebrospinal fluid compared with untreated rats. To elucidate the possible mechanism of this observation, the authors investigated for the first time leukocyte rolling in an inflammatory model in brain venules by confocal laser scanning microscopy in vivo. Heparin significantly attenuates leukocyte rolling at 2, 3, and 4 hours (2.8 ± 1.3 versus 7.9 ± 3.2/100 μm/min), as well as leukocyte sticking at 4 hours (2.1 ± 0.4 versus 3.5 ± 1.0/100 μm/min) after meningitis induction compared with untreated animals. The authors conclude that heparin can modulate acute central nervous system inflammation and, in particular, leukocyte–endothelium interaction, a key process in the cascade of injury in bacterial meningitis. They propose to evaluate further the potential of heparin in central nervous system inflammation in basic and clinical studies.

Confocal Laser Microscopy of Physiologically Characterized Fluorescently Labeled Central Nervous System Neurons

1988 ◽  
Vol 46 ◽  
pp. 274-275
Author(s):  
J.N. Turner ◽  
J. Swann ◽  
K. Smith ◽  
M. Siemens ◽  
D. Szarowski ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Laser Scanning ◽  
Tissue Sample ◽  
Lucifer Yellow ◽  
Single Cells ◽  
Brain Slices ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Efficient Detection

Confocal laser scanning microscopy (CLSM) is capable of three-dimensional imaging of fluorescently labeled single cells. Efficient detection via a photomultiplier and optical sectioning with high rejection of light from other specimen levels make it possible to image cells surrounded by either labeled or unlabeled tissue. It is no longer necessary to restrict high resolution light microscopy to cultured cells or those near the surface of a tissue sample. Cells can be observed üin situ” in a physiologically characterized environment. Central nervous system neurons can be electrophysiologically characterized and then injected with a fluorescent dye such as lucifer yellow. The CLSM can excite the dye and image the fluorescent emission in thick tissue preparations (hundreds of micrometers) making possible a new approach to the correlation of physiology and anatomy.Brain slices 350 μm thick were obtained from hippocampus and inferior colliculus of immature rats and incubated in oxygenated artificial cerebrospinal fluid. Cells were penetrated with micropipets, characterized electrophysiologically and ionophoretically injected with 5% lucifer yellow in LiAc.

Vacuoles Induced in Mammalian Cells by Helicobacter Cytotoxin are Acidic Organelles

1990 ◽  
Vol 48 (3) ◽  
pp. 168-169
Author(s):  
M. H. Chestnut ◽  
C. E. Catrenich
Keyword(s):  
Acridine Orange ◽  
Mammalian Cells ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Ulcer Disease ◽  
Gastroduodenal Disease ◽  
H Pylori

Helicobacter pylori is a non-invasive, Gram-negative spiral bacterium first identified in 1983, and subsequently implicated in the pathogenesis of gastroduodenal disease including gastritis and peptic ulcer disease. Cytotoxic activity, manifested by intracytoplasmic vacuolation of mammalian cells in vitro, was identified in 55% of H. pylori strains examined. The vacuoles increase in number and size during extended incubation, resulting in vacuolar and cellular degeneration after 24 h to 48 h. Vacuolation of gastric epithelial cells is also observed in vivo during infection by H. pylori. A high molecular weight, heat labile protein is believed to be responsible for vacuolation and to significantly contribute to the development of gastroduodenal disease in humans. The mechanism by which the cytotoxin exerts its effect is unknown, as is the intracellular origin of the vacuolar membrane and contents. Acridine orange is a membrane-permeant weak base that initially accumulates in low-pH compartments. We have used acridine orange accumulation in conjunction with confocal laser scanning microscopy of toxin-treated cells to begin probing the nature and origin of these vacuoles.

scholarly journals Targeted Liposomal Chemotherapies to Treat Triple-Negative Breast Cancer

Cancers ◽  
2021 ◽  
Vol 13 (15) ◽  
pp. 3749
Author(s):  
Yingnan Si ◽  
Ya Zhang ◽  
Hanh Giai Ngo ◽  
Jia-Shiung Guan ◽  
Kai Chen ◽  
...  
Keyword(s):  
Targeted Delivery ◽  
Laser Scanning ◽  
Triple Negative ◽  
Imaging System ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Surface Binding ◽  
Synthesis Inhibitor ◽  
Tnbc Cell

Triple-negative breast cancers (TNBCs) are highly aggressive and recurrent. Standard cytotoxic chemotherapies are currently the main treatment options, but their clinical efficacies are limited and patients usually suffer from severe side effects. The goal of this study was to develop and evaluate targeted liposomes-delivered combined chemotherapies to treat TNBCs. Specifically, the IC50 values of the microtubule polymerization inhibitor mertansine (DM1), mitotic spindle assembly defecting taxane (paclitaxel, PTX), DNA synthesis inhibitor gemcitabine (GC), and DNA damage inducer doxorubicin (AC) were tested in both TNBC MDA-MB-231 and MDA-MB-468 cells. Then we constructed the anti-epidermal growth factor receptor (EGFR) monoclonal antibody (mAb) tagged liposomes and confirmed its TNBC cell surface binding using flow cytometry, internalization with confocal laser scanning microscopy, and TNBC xenograft targeting in NSG female mice using In Vivo Imaging System. The safe dosage of anti-EGFR liposomal chemotherapies, i.e., <20% body weight change, was identified. Finally, the in vivo anti-tumor efficacy studies in TNBC cell line-derived xenograft and patient-derived xenograft models revealed that the targeted delivery of chemotherapies (mertansine and gemcitabine) can effectively inhibit tumor growth. This study demonstrated that the targeted liposomes enable the new formulations of combined therapies that improve anti-TNBC efficacy.

scholarly journals Degradation of Drug Delivery Nanocarriers and Payload Release: A Review of Physical Methods for Tracing Nanocarrier Biological Fate

Pharmaceutics ◽  
2021 ◽  
Vol 13 (6) ◽  
pp. 770
Author(s):  
Patrick M. Perrigue ◽  
Richard A. Murray ◽  
Angelika Mielcarek ◽  
Agata Henschke ◽  
Sergio E. Moya
Keyword(s):  
Drug Delivery ◽  
Laser Scanning ◽  
Single Photon ◽  
Photon Emission ◽  
Correlation Spectroscopy ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Emission Computed Tomography ◽  
Systemic Delivery

Nanoformulations offer multiple advantages over conventional drug delivery, enhancing solubility, biocompatibility, and bioavailability of drugs. Nanocarriers can be engineered with targeting ligands for reaching specific tissue or cells, thus reducing the side effects of payloads. Following systemic delivery, nanocarriers must deliver encapsulated drugs, usually through nanocarrier degradation. A premature degradation, or the loss of the nanocarrier coating, may prevent the drug’s delivery to the targeted tissue. Despite their importance, stability and degradation of nanocarriers in biological environments are largely not studied in the literature. Here we review techniques for tracing the fate of nanocarriers, focusing on nanocarrier degradation and drug release both intracellularly and in vivo. Intracellularly, we will discuss different fluorescence techniques: confocal laser scanning microscopy, fluorescence correlation spectroscopy, lifetime imaging, flow cytometry, etc. We also consider confocal Raman microscopy as a label-free technique to trace colocalization of nanocarriers and drugs. In vivo we will consider fluorescence and nuclear imaging for tracing nanocarriers. Positron emission tomography and single-photon emission computed tomography are used for a quantitative assessment of nanocarrier and payload biodistribution. Strategies for dual radiolabelling of the nanocarriers and the payload for tracing carrier degradation, as well as the efficacy of the payload delivery in vivo, are also discussed.

Probing Enhanced Cytochrome P450 2B1/2 Activity in Rat Hepatocyte Spheroids through Confocal Laser Scanning Microscopy

2001 ◽  
Vol 10 (3) ◽  
pp. 329-342 ◽  
Author(s):  
Emmanouhl S. Tzanakakis ◽  
Chang-Chun Hsiao ◽  
Taku Matsushita ◽  
Rory P. Remmel ◽  
Wei-Shou Hu
Keyword(s):  
Cytochrome P450 ◽  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Constitutive Expression ◽  
Rat Hepatocytes ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser

Cytochrome P450 (CYP450) enzymes are essential for xenobiotic metabolism. Although CYP450s are found in many tissues, CYP2B1/2 are primarily expressed in the rat liver. The constitutive expression in vivo of CYP2B1/2 is low but it is induced in the presence of various drugs such as phenobarbital (PB). In this study, CYP2B1/2 activity in cultured hepatocytes was assessed in situ with the introduction of a fluorogenic sub-strate, pentoxyresorufin. The product of 7-pentoxyresorufin-O-dealkylation (PROD), which is catalyzed specifically by CYP2B1/2, was detected using confocal laser scanning microscopy (CLSM). Primary hepatocytes cultured as monolayers on collagen-coated surfaces exhibited background PROD activity and minimal PB inducibility after 4 days in culture. In contrast, rat hepatocytes organized in compacted aggregates, or spheroids, exhibited higher levels of PROD activity and retained their ability for PB induction. The results from the CLSM analysis were verified by RT-PCR and Western immunoblotting analysis. Furthermore, CLSM in conjunction with image processing techniques and three-dimensional reconstruction revealed the localization of enhanced PROD activity in the center of spheroids. The results support the use of CLSM as a powerful tool for investigating CYP2B1/2 activity in cultured rat hepatocytes.

scholarly journals The Interaction of N-Acetylcysteine and Serum Transferrin Promotes Bacterial Biofilm Formation

2018 ◽  
Vol 45 (4) ◽  
pp. 1399-1409 ◽  
Author(s):  
Supeng Yin ◽  
Bei Jiang ◽  
Guangtao Huang ◽  
Yulong Zhang ◽  
Bo You ◽  
...  
Keyword(s):  
Human Serum ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Serum Proteins ◽  
Venous Catheter ◽  
Bacterial Biofilm ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Bacterial Strains

Background/Aims: N-acetylcysteine (NAC) is a novel and promising agent with activity against bacterial biofilms. Human serum also inhibits biofilm formation by some bacteria. We tested whether the combination of NAC and human serum offers greater anti-biofilm activity than either agent alone. Methods: Microtiter plate assays and confocal laser scanning microscopy were used to evaluate bacterial biofilm formation in the presence of NAC and human serum. qPCR was used to examine expression of selected biofilm-associated genes. Extracellular matrix (ECM) was observed by transmission electron microscopy. The antioxidants GSH or ascorbic acid were used to replace NAC, and human transferrin, lactoferrin, or bovine serum albumin were used to replace serum proteins in biofilm formation assays. A rat central venous catheter model was developed to evaluate the effect of NAC on biofilm formation in vivo. Results: NAC and serum together increased biofilm formation by seven different bacterial strains. In Staphylococcus aureus, expression of genes for some global regulators and for genes in the ica-dependent pathway increased markedly. In Pseudomonas aeruginosa, transcription of las, the PQS quorum sensing (QS) systems, and the two-component system GacS/GacA increased significantly. ECM production by S. aureus and P. aeruginosa was also enhanced. The potentiation of biofilm formation is due mainly to interaction between NAC and transferrin. Intravenous administration of NAC increased colonization by S. aureus and P. aeruginosa on implanted catheters. Conclusions: NAC used intravenously or in the presence of blood increases bacterial biofilm formation rather than inhibits it.

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