all-D proline-rich cell-penetrating peptides: a preliminary in vivo internalization study

2007 ◽  
Vol 35 (4) ◽  
pp. 794-796 ◽  
Author(s):  
S. Pujals ◽  
E. Sabidó ◽  
T. Tarragó ◽  
E. Giralt
Keyword(s):  
Self Assembly ◽  
Laser Scanning ◽  
White Blood Cells ◽  
Cell Penetrating Peptides ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Cell Penetrating ◽  
Peptide Derivatives ◽  
Degradation Problem

Proline-rich cell-penetrating peptides, particularly the SAP (sweet arrow peptide), (VRLPPP)3, have been proposed to be useful intracellular delivery vectors, as a result of their lack of cytotoxicity combined with their capacity to be internalized by cells. A common limitation of the therapeutic use of peptides is metabolic instability. In general, peptides are quickly degraded by proteases upon entry into the bloodstream. The use of all-D-peptide derivatives is emerging as a fruitful strategy to circumvent this degradation problem. In this context, we report on the internalization behaviour, protease-resistance enhancement and self-assembly properties of an all-D version of SAP [(vrlppp)3]. The cellular uptake of (vrlppp)3 was evaluated in an in vivo assay in mice. Both flow cytometry and confocal laser-scanning microscopy experiments showed that a carboxyfluoresceinated version of the molecule, carboxyfluorescein–(vrlppp)3, is internalized rapidly in white blood cells and kidney cells. Significant fluorescence was also detected in other organs such as the spleen and the liver. Finally, the toxicity of (vrlppp)3 was examined, and no significant differences in the main biochemical parameters nor in weight were detected compared with controls.

scholarly journals Cell-Penetrating Peptide and Transferrin Co-Modified Liposomes for Targeted Therapy of Glioma

Molecules ◽  
2019 ◽  
Vol 24 (19) ◽  
pp. 3540 ◽  
Author(s):  
Xi Wang ◽  
Yarong Zhao ◽  
Shiyan Dong ◽  
Robert J. Lee ◽  
Dongsheng Yang ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Therapeutic Agent ◽  
Spherical Shape ◽  
Cell Penetrating Peptides ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Great Promise ◽  
Uniform Size ◽  
Cell Penetrating ◽  
Low Toxicity

Glioma is one of the most aggressive and common malignant brain tumors. Due to the presence of the blood-brain barrier (BBB), the effectiveness of therapeutics is greatly affected. In this work, to develop an efficient anti-glioma drug with targeting and which was able to cross the BBB, cell-penetrating peptides (R8) and transferrin co-modified doxorubicin (DOX)-loaded liposomes (Tf-LPs) were prepared. Tf-LPs possessed a spherical shape and uniform size with 128.64 nm and their ξ-potential was 6.81 mV. Tf-LPs were found to be stable in serum within 48 h. Uptake of Tf-LPs in both U87 and GL261 cells was analyzed by confocal laser scanning microscopy and by flow cytometry. Tf-LPs were efficiently taken up by both U87 and GL261 cells. Moreover, Tf-LPs exhibited sustained-release. The cumulative release of DOX from Tf-LPs reached ~50.0% and showed excellent anti-glioma efficacy. Histology of major organs, including brain, heart, liver, spleen, lungs and kidney, and the bodyweight of mice, all indicated low toxicity of Tf-LPs. In conclusion, Tf-LPs showed great promise as an anti-glioma therapeutic agent.

scholarly journals Design of new acid-activated cell-penetrating peptides for tumor drug delivery

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3429 ◽  
Author(s):  
Jia Yao ◽  
Yinyun Ma ◽  
Wei Zhang ◽  
Li Li ◽  
Yun Zhang ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Targeted Delivery ◽  
Laser Scanning ◽  
Critical Role ◽  
Cell Penetrating Peptides ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Ph Response ◽  
Cell Penetrating ◽  
Ph Dependent

TH(AGYLLGHINLHHLAHL(Aib)HHIL-NH2), a histidine-rich, cell-penetrating peptide with acid-activated pH response, designed and synthesized by our group, can effectively target tumor tissues with an acidic extracellular environment. Since the protonating effect of histidine plays a critical role in the acid-activated, cell-penetrating ability of TH, we designed a series of new histidine substituents by introducing electron donating groups (Ethyl, Isopropyl, Butyl) to the C-2 position of histidine. This resulted in an enhanced pH-response and improved the application of TH in tumor-targeted delivery systems. The substituents were further utilized to form the corresponding TH analogs (Ethyl-TH, Isopropyl-TH and Butyl-TH), making them easier to protonate for positive charge in acidic tumor microenvironments. The pH-dependent cellular uptake efficiencies of new TH analogs were further evaluated using flow cytometry and confocal laser scanning microscopy, demonstrating that ethyl-TH and butyl-TH had an optimal pH-response in an acidic environment. Importantly, the new TH analogs exhibited relatively lower toxicity than TH. In addition, these new TH analogs were linked to the antitumor drug camptothecin (CPT), while butyl-TH modified conjugate presented a remarkably stronger pH-dependent cytotoxicity to cancer cells than TH and the other conjugates. In short, our work opens a new avenue for the development of improved acid-activated, cell-penetrating peptides as efficient anticancer drug delivery vectors.

scholarly journals Synthesis of Water-Soluble Iridium (III)-Containing Nanoparticles for Biological Applications

2015 ◽  
Vol 2015 ◽  
pp. 1-7
Author(s):  
Huanzhi Hou ◽  
Pengfei Sun ◽  
Quli Fan ◽  
Xiaomei Lu ◽  
Cheng Xue ◽  
...  
Keyword(s):  
Self Assembly ◽  
Laser Scanning ◽  
Morphological Characterization ◽  
Fluorescent Imaging ◽  
Water Soluble ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Biological Applications

Water-soluble nanoparticles (Ir/PGlc-NP, Ir/β-1,3-glucan-NP) based on water-soluble glycopolymers (PGlc),β-1,3-glucan polysaccharide, and conjugated phosphorescent Ir (III) complexes were successfully synthesized by self-assembly. The obtained nanoparticles have good spherical morphological characterization with a mean diameter of 50 nm measured by TEM. Ir/PGlc-NP and Ir/β-1,3-glucan-NP showed the same emission maxima at 565 nm in aqueous solution and both caused effective apoptosis and death of HepG2 and Hela cells after being irradiated at 445 nm for 30 min in vitro. Fluorescence cellular imaging was conducted by confocal laser scanning microscopy (CLSM) using HepG2 cells as the model cell in which the nanoparticles had successfully entered into the cytoplasm with high brightness. Furthermore, after injecting the nanoparticles into live mice in vivo, the real-time fluorescence imaging as well as the nanoparticles distribution in organs at 24 hours after administration indicated that these nanoparticles can serve as fluorescent imaging contrast for further biological applications.

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.

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