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Molecules (Basel, Switzerland) Apr 2019We developed a tumor-targeted contrast agent based on linear polylysine (PLL) by conjugating a small molecular imaging agent, fluorescent molecule and targeting agent...
We developed a tumor-targeted contrast agent based on linear polylysine (PLL) by conjugating a small molecular imaging agent, fluorescent molecule and targeting agent amino phenylboronic acid onto the amino groups of polylysine, which can specifically target monosaccharide sialic acid residues overexpressing on the surface of tumor cell membranes. Further, 3,4,5,6-Tetrahydrophthalic anhydride (DCA) was attached to the free amino groups of the polylysine to change to a negative charge at physiology pH to lower the cytotoxicity, but it soon regenerated to a positive charge again once reaching the acidic intratumoral environment and therefore increased cell uptake. Laser confocal microscopy images showed that most of the polymeric contrast agents were bound to the cancer cell membrane. Moreover, the tumor targeting contrast agent showed the same magnetic resonance imaging (MRI) contrasting performance in vitro as the small molecule contrast agent used in clinic, which made it a promising tumor-targeting polymeric contrast agent for cancer diagnosis.
Topics: Animals; Contrast Media; Dogs; Hep G2 Cells; Humans; Madin Darby Canine Kidney Cells; Magnetic Resonance Imaging; Microscopy, Confocal; Neoplasms; Polylysine
PubMed: 30991689
DOI: 10.3390/molecules24081477 -
The Journal of Experimental Medicine Nov 2011This issue of the Journal of Experimental Medicine celebrates and honors the life of Ralph Steinman (1943-2011), winner of the 2011 Nobel Prize in Physiology or...
This issue of the Journal of Experimental Medicine celebrates and honors the life of Ralph Steinman (1943-2011), winner of the 2011 Nobel Prize in Physiology or Medicine. Ralph's science was rooted in fundamental discovery with the goal of translating these findings into clinical medicine. He recognized the power of immunology in treating human disease and passionately championed studies on vaccine design, immune therapy, and human immunology. One particular collaborative effort between the Steinman and Sekaly laboratories resulted in a paper published in this issue of the journal.
Topics: Adjuvants, Immunologic; Carboxymethylcellulose Sodium; Gene Expression Regulation; Humans; Immunity, Innate; Poly I-C; Polylysine; Signal Transduction
PubMed: 22110181
DOI: 10.1084/jem.20112321 -
ACS Applied Materials & Interfaces Apr 2017Temporary single-cell coating is a useful tool for cell processing, allowing manipulation of cells to prevent cell attachment and agglomeration, before re-establishing...
Temporary single-cell coating is a useful tool for cell processing, allowing manipulation of cells to prevent cell attachment and agglomeration, before re-establishing normal cell function. In this work, a speckled coating method using a known polycation [poly(l-lysine), PLL] is described to induce cell surface electrostatic charges on three different cell types, namely, two bone cancer cell lines and fibroblasts. The morphology of the PLL speckled coating on the cell surface, internalization and metabolization of the polymer, and prevention of cellular aggregations are reported. Polymer concentration was found to be the key parameter controlling both capsule morphology and cell health. This approach allows a temporary cell coating over the course of 1-2 h, with cells exhibiting phenotypically normal behavior after ingesting and metabolizing the polymer. The process offers a fast and efficient alternative to aid single-cell manipulation for bioprocessing applications. Preliminary work on the application of PLL speckled cell coating in enabling reliable bioprinting is also presented.
Topics: Capsules; Cations; Cell Line; Polylysine; Polymers
PubMed: 28323412
DOI: 10.1021/acsami.6b16434 -
Journal of Thrombosis and Haemostasis :... Jun 2023Hemorrhage, in particular noncompressible hemorrhage, is the leading cause of casualties in combat trauma and civilian trauma. Although systemic agents can stop bleeding...
An anticoagulant/procoagulant self-converting and bleeding site-targeting systemic nanotherapy for rapidly controlling noncompressible bleeding without risk of thrombosis.
BACKGROUND
Hemorrhage, in particular noncompressible hemorrhage, is the leading cause of casualties in combat trauma and civilian trauma. Although systemic agents can stop bleeding at both inaccessible and accessible injury sites, the application of systemic hemostats in clinics is strictly limited by the nontargeting ability of hemostats and their subsequent potential for thromboembolic complications.
OBJECTIVES
To engineer an anticoagulant/procoagulant self-converting and bleeding site-targeting systemic nanohemostat to rapidly control noncompressible bleeding without thrombosis risk.
METHODS
A multiscale computer simulation was taken to guide the self-assembly of sulindac (SUL, a prodrug of the antiplatelets agent) and poly-L-lysine (a cation polymer with platelets activation ability) for forming poly-L-lysine/SUL nanoparticles (PSNs). In vitro platelet-adhering ability, platelet activation effect, and hemostasis activity of PSNs were evaluated. Then, the biosafety, level of thrombosis, targeting ability, and hemostasis effect of systemic applied PSNs were carefully evaluated in various hemorrhage models.
RESULTS
PSNs were successfully prepared and showed good platelet adhesion and activation in vitro. The bleeding site-targeting ability and hemostatic efficiency in different bleeding models were leveled up by PSNs markedly compared with vitamin K and etamsylate in vivo. SUL in PSNs could be metabolized into sulindac sulfide at clot sites in 4 hours for antiplatelet aggregation, thus reducing thrombotic risk compared with other hemostatic agents, via the ingenious utilization of prodrug metabolism in terms of time intervals and the adhesion on platelets.
CONCLUSION
PSNs are expected to be a low-cost, safe, efficient, clinically translatable first-aid hemostat for first-aid scenarios.
Topics: Humans; Anticoagulants; Computer Simulation; Polylysine; Prodrugs; Hemorrhage; Hemostasis; Hemostatics; Thrombosis
PubMed: 36871669
DOI: 10.1016/j.jtha.2023.02.020 -
Frontiers in Immunology 2022Both Gram-negative and Gram-positive bacteria can release vesicle-like structures referred to as bacterial extracellular vesicles (BEVs), which contain various bioactive...
Both Gram-negative and Gram-positive bacteria can release vesicle-like structures referred to as bacterial extracellular vesicles (BEVs), which contain various bioactive compounds. BEVs play important roles in the microbial community interactions and host-microbe interactions. Markedly, BEVs can be delivered to host cells, thus modulating the development and function of the innate immune system. To clarify the compositions and biological functions of BEVs, we need to collect these vesicles with high purity and bioactivity. Here we propose an isolation strategy based on a broad-spectrum antimicrobial epsilon-poly-L-lysine (ϵ-PL) to precipitate BEVs at a relatively low centrifugal speed (10,000 × g). Compared to the standard ultracentrifugation strategy, our method can enrich BEVs from large volumes of media inexpensively and rapidly. The precipitated BEVs can be recovered by adjusting the pH and ionic strength of the media, followed by an ultrafiltration step to remove ϵ-PL and achieve buffer exchange. The morphology, size, and protein composition of the ϵ-PL-precipitated BEVs are comparable to those purified by ultracentrifugation. Moreover, ϵ-PL-precipitated BEVs retained the biological activity as observed by confocal microscopy studies. And THP-1 cells stimulated with these BEVs undergo marked reprogramming of their transcriptome. KEGG analysis of the differentially expressed genes showed that the signal pathways of cellular inflammatory response were significantly activated. Taken together, we provide a new method to rapidly enrich BEVs with high purity and bioactivity, which has the potential to be applied to BEVs-related immune response studies.
Topics: Anti-Bacterial Agents; Culture Media; Extracellular Vesicles; Host Microbial Interactions; Polylysine
PubMed: 36032173
DOI: 10.3389/fimmu.2022.930510 -
SLAS Discovery : Advancing Life... Jan 2017Although reverse transfection cell microarray (RTCM) is a powerful tool for mammalian cell studies, the technique is not appropriate for cells that are difficult to...
Although reverse transfection cell microarray (RTCM) is a powerful tool for mammalian cell studies, the technique is not appropriate for cells that are difficult to transfect. The lentivirus-infected cell microarray (LICM) technique was designed to improve overall efficiency. However, LICM presents new challenges because individual lentiviral particles can spread through the cell population, leading to cross-contamination. Therefore, we designed a cell-defined lentivirus microarray (CDLM) technique using cell-friendly biomaterials that are controlled by cell attachment timing. We selected poly-l-lysine (PLL) with Matrigel as the best combination of biomaterials for cell-defined culture. We used 2 µL PLL to determine by titration the optimum concentration required (0.04% stock, 0.005% final concentration). We also determined the optimum concentration of 10 µL of lentivirus particles for maximum reverse infection efficiency (1 × 10 infectious units [IFU]/mL stock, 62.5% final concentration) and established the best combination of components for the lentivirus mixture (10 µL of lentivirus particles and 2 µL each of siGLO Red dye, Matrigel, and 0.04% PLL). Finally, we validated both the effect of reverse infection in various cell lines and lentivirus spot activity in CDLM by storage period. This method provides an effective lentivirus-infected cell microarray for large-scale gene function studies.
Topics: Animals; Biocompatible Materials; Cell Line, Tumor; Green Fluorescent Proteins; Humans; Lentivirus; Mammals; Microarray Analysis; Polylysine; Reproducibility of Results
PubMed: 27703081
DOI: 10.1177/1087057116672417 -
Nanomedicine (London, England) Jan 2011We posit that cell-mediated drug delivery can improve transport of therapeutic enzymes to the brain and decrease inflammation and neurodegeneration seen during...
BACKGROUND
We posit that cell-mediated drug delivery can improve transport of therapeutic enzymes to the brain and decrease inflammation and neurodegeneration seen during Parkinson's disease. Our prior works demonstrated that macrophages loaded with nanoformulated catalase ('nanozyme') then parenterally injected protect the nigrostriatum in a murine model of Parkinson's disease. Packaging of catalase into block ionomer complex with a synthetic polyelectrolyte block copolymer precludes enzyme degradation in macrophages.
METHODS
We examined relationships between the composition and structure of block ionomer complexes with a range of block copolymers, their physicochemical characteristics, and loading, release and catalase enzymatic activity in bone marrow-derived macrophages.
RESULTS
Formation of block ionomer complexes resulted in improved aggregation stability. Block ionomer complexes with ε-polylysine and poly(L-glutamic acid)-poly(ethylene glycol) demonstrated the least cytotoxicity and high loading and release rates. However, these formulations did not efficiently protect catalase inside macrophages.
CONCLUSION
Nanozymes with polyethyleneimine- and poly(L-lysine)(10)-poly(ethylene glycol) provided the best protection of enzymatic activity for cell-mediated drug delivery.
Topics: Animals; Catalase; Cattle; Cell Line; Cell Survival; Cells, Cultured; Drug Delivery Systems; Macrophages; Male; Mice; Mice, Inbred C57BL; Nanoparticles; Neurons; Polyethylene Glycols; Polyethyleneimine; Polyglutamic Acid; Polylysine
PubMed: 21182416
DOI: 10.2217/nnm.10.129 -
The Journal of Biological Chemistry Dec 2000After synthesis in the cytosol, Ras proteins must be targeted to the inner leaflet of the plasma membrane for biological activity. This targeting requires a series of...
After synthesis in the cytosol, Ras proteins must be targeted to the inner leaflet of the plasma membrane for biological activity. This targeting requires a series of C-terminal posttranslational modifications initiated by the addition of an isoprenoid lipid in a process termed prenylation. A search for factors involved in the intracellular trafficking of Ras has identified a specific and prenylation-dependent interaction between tubulin/microtubules and K-Ras. In this study, we examined the structural requirements for this interaction between K-Ras and microtubules. By using a series of chimeras in which regions of the C terminus of K-Ras were replaced with those of Ha-Ras and vice versa, we found that the polylysine region of K-Ras located immediately upstream of the prenylation site is required for binding of K-Ras to microtubules. Studies in intact cells confirmed the importance of the K-Ras polylysine region for microtubule binding, as deletion or replacement of this region resulted in loss of paclitaxel-induced mislocalization of a fluorescent K-Ras fusion protein. The additional modifications in the prenyl protein processing pathway also affected the interaction of K-Ras with microtubules. Removal of the three C-terminal amino acids of farnesylated K-Ras with the specific endoprotease Rce1p abolished its binding to microtubules. Interestingly, however, methylation of the C-terminal prenylcysteine restored binding. Consistent with these results, localization of the fluorescent K-Ras fusion protein remained paclitaxel-sensitive in cells lacking Rce1, whereas no paclitaxel effect was observed in cells lacking the methyltransferase. These studies show that the polylysine region of K-Ras is critical for its interaction with microtubules and provide the first evidence for a functional consequence of Ras C-terminal proteolysis and methylation.
Topics: Amino Acid Sequence; Cells, Cultured; Methylation; Microtubules; Molecular Sequence Data; Paclitaxel; Polylysine; ras Proteins
PubMed: 11007785
DOI: 10.1074/jbc.M006687200 -
Theranostics 2024Intravesical chemotherapy is highly recommended after transurethral resection of bladder tumor for patients with bladder cancer (BCa). However, this localized adjuvant...
Intravesical chemotherapy is highly recommended after transurethral resection of bladder tumor for patients with bladder cancer (BCa). However, this localized adjuvant therapy has drawbacks of causing indiscriminate damage and inability to penetrate bladder mucosal. Fluorinated polylysine micelles (PLLF) were synthesized by reacting polylysine (PLL) with heptafluorobutyrate anhydride. Anti-apoptotic gene defender against cell death 1 (DAD1) was selected by different gene expression analysis between BCa patients and healthy individuals and identified by several biological function assays. The gene transfection ability of PLLF was verified by multiple and assays. The therapeutic efficiency of PLLF nanoparticles (NPs) targeting DAD1 were confirmed by intravesical administration using an orthotopic BCa mouse model. Decorated with fluorinated chains, PLL can self-assemble to form NPs and condense plasmids with excellent gene transfection efficiency . Loading with the CRISPR-Cas9 system designed to target DAD1 (Cas9-sgDAD1), PLLF/Cas9-sgDAD1 NPs strongly inhibited the expression of DAD1 in BCa cells and induced BCa cell apoptosis through the MAPK signaling pathway. Furthermore, intravesical administration of PLLF/Cas9-sgDAD1 NPs resulted in significant therapeutic outcomes without systemic toxicity . The synthetized PLLF can transmucosally deliver the CRISPR-Cas9 system into orthotopic BCa tissues to improve intravesical instillation therapy for BCa. This work presents a new strategy for targeting DAD1 gene in the intravesical therapy for BCa with high potential for clinical applications.
Topics: Mice; Animals; Humans; Urinary Bladder; Polylysine; CRISPR-Cas Systems; Urinary Bladder Neoplasms; Nanoparticles; Genetic Therapy
PubMed: 38164146
DOI: 10.7150/thno.88550 -
Scientific Reports Nov 2022Designing the architecture of L-lysine-based polymeric structures is a highly challenging task that requires careful control of the amino acid reactive groups....
Designing the architecture of L-lysine-based polymeric structures is a highly challenging task that requires careful control of the amino acid reactive groups. Conventional processes to obtain branched polylysine need several steps and the addition of specific catalysts. In the present work, to gain a better understanding and control of the formation of L-lysine-based polymers, we have investigated the correlation between the protonation state of L-lysine and the corresponding hydrothermally grown structures. The samples have been characterized by combining optical spectroscopies, such as UV-Vis, fluorescence, and synchrotron radiation circular dichroism with structural analysis by Nuclear Magnetic Resonance, Fourier Transform Infrared spectroscopy, and dynamic light scattering. We have observed that aqueous precursor solutions with alkaline pHs promote the formation of branched structures. In contrast, high pHs favour the reactivity of the ε-amino groups leading to linear structures, as shown by circular dichroism analyses. On the other hand, acidic conditions trigger the branching of the amino acid. Interestingly, the polymeric forms of L-lysine emit in the blue because the increasing number of intermolecular hydrogen bonds promote the intermolecular charge transfer responsible for the emission. Understanding the correlation between the L-lysine charged states and the polymeric structures that could form controlling the protonation-deprotonation states of the amino acid opens the route to a refined design of polypeptide systems based on L-lysine.
Topics: Polylysine; Circular Dichroism; Peptides; Poly A; Amino Acids; Polymers
PubMed: 36385123
DOI: 10.1038/s41598-022-24109-5