scholarly journals Folate (pteroylglutamate) uptake in human red blood cells, erythroid precursors and KB cells at high extracellular folate concentrations. Evidence against a role for specific folate-binding and transport proteins

1989 ◽  
Vol 260 (2) ◽  
pp. 401-411 ◽  
Author(s):  
A C Antony ◽  
M A Kane ◽  
S R Krishnan ◽  
R S Kincade ◽  
R S Verma
Keyword(s):  
Folate Receptor ◽  
Transport Proteins ◽  
Human Cells ◽  
Kb Cells ◽  
Uptake Mechanism ◽  
Human Red Blood Cells ◽  
Physiological Relevance ◽  
Cell Accumulation ◽  
Normal Human

Membrane-associated folate (pteroylglutamate, PteGlu)-binding proteins (FBPs) play an important role as PteGlu-transport proteins in malignant and normal human cells. Since high extracellular folate (PteGlu) concentrations (EFC) profoundly influenced uptake and toxicity of the anti-PteGlu methotrexate in malignant KB cells, we studied human cells to determine additional mechanisms for PteGlu uptake when the EFC was varied. At low EFC (less than 10 nM), the predominant mechanism for folate uptake in mature erythrocytes was through binding to externally oriented FBPs which were quantitatively insignificant (4-6 orders of magnitude lower) and of no apparent physiological relevance when compared with KB cells. However, the predominant mechanism of PteGlu accumulation at high EFC [10-250 nM] in intact erythrocytes and sealed right-side-out (RSO) ghosts was not FBP-mediated and non-specific. This conclusion was based on the findings that radiolabelled PteGlu uptake: (i) continued even in the presence of a 1000-fold excess of unlabelled PteGlu and was linear and not saturable up to 250 nM; (ii) was two-fold higher at pH 4.5 than 7.5; (iii) was less than 2-fold increased at 37 degrees C compared with 4 degrees C; and (iv) was unaffected after trypsin-mediated proteolysis of greater than 75% FBPs. The [3H]PteGlu and 125I-PteGlu (histamine derivative) accumulated intracellularly through the non-specific PteGlu-uptake mechanism was unaltered biochemically and in a soluble compartment. Raising the EFC 500-fold higher than controls during erythropoiesis in vitro resulted in reversal of the expected anti-(placental folate-receptor)-antiserum-induced megaloblastic changes in orthochromatic normoblasts derived from burst-forming unit-erythroid colonies. Furthermore, at EFC greater than 0.1 microM, KB-cell accumulation of [3H]PteGlu was also predominantly through a mechanism that did not involve specific FBPs. Thus, at high EFC, a major component of PteGlu transport in human cells is not mediated through FBPs and is likely to be a passive diffusion process.

Increase of Enzyme Activities following the in vitro Peroxidation of Normal Human Red Blood Cells

Enzyme ◽  
1988 ◽  
Vol 39 (1) ◽  
pp. 1-7 ◽  
Author(s):  
J. L. Vives Carrons ◽  
M. A. Pujades ◽  
D. Colomer
Keyword(s):  
Red Blood Cells ◽  
Enzyme Activities ◽  
Blood Cells ◽  
Human Red Blood Cells ◽  
Normal Human

scholarly journals Supra-physiological folic acid concentrations induce aberrant DNA methylation in normal human cells in vitro

Epigenetics ◽  
2012 ◽  
Vol 7 (7) ◽  
pp. 689-694 ◽  
Author(s):  
Michelle A. Charles ◽  
Ian T. Johnson ◽  
Nigel J. Belshaw
Keyword(s):  
Dna Methylation ◽  
Folic Acid ◽  
Human Cells ◽  
Normal Human ◽  
Normal Human Cells ◽  
Aberrant Dna Methylation

scholarly journals Effect of Disulfide Cyclization of Ultrashort Cationic Lipopeptides on Antimicrobial Activity and Cytotoxicity

2020 ◽  
Vol 21 (19) ◽  
pp. 7208
Author(s):  
Damian Neubauer ◽  
Maciej Jaśkiewicz ◽  
Emilia Sikorska ◽  
Sylwia Bartoszewska ◽  
Marta Bauer ◽  
...  
Keyword(s):  
Cyclic Peptide ◽  
Human Cells ◽  
Coarse Grained ◽  
Polar Head ◽  
Candida Sp ◽  
Transmission Electron ◽  
Significant Toxicity ◽  
Normal Human ◽  
Dynamics Simulations

Ultrashort cationic lipopeptides (USCLs) are considered to be a promising class of antimicrobials with high activity against a broad-spectrum of microorganisms. However, the majority of these compounds are characterized by significant toxicity toward human cells, which hinders their potential application. To overcome those limitations, several approaches have been advanced. One of these is disulfide cyclization that has been shown to improve drug-like characteristics of peptides. In this article the effect of disulfide cyclization of the polar head of N-palmitoylated USCLs on in vitro biological activity has been studied. Lipopeptides used in this study consisted of three or four basic amino acids (lysine and arginine) and cystine in a cyclic peptide. In general, disulfide cyclization of the lipopeptides resulted in peptides with reduced cytotoxicity. Disulfide-cyclized USCLs exhibited improved selectivity between Candida sp., Gram-positive strains and normal cells in contrast to their linear counterparts. Interactions between selected USCLs and membranes were studied by molecular dynamics simulations using a coarse-grained force field. Moreover, membrane permeabilization properties and kinetics were examined. Fluorescence and transmission electron microscopy revealed damage to Candida cell membrane and organelles. Concluding, USCLs are strong membrane disruptors and disulfide cyclization of polar head can have a beneficial effect on its in vitro selectivity between Candida sp. and normal human cells.

scholarly journals Development of Folate-Functionalized PEGylated Zein Nanoparticles for Ligand-Directed Delivery of Paclitaxel

Pharmaceutics ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 562 ◽  
Author(s):  
Zar Chi Soe ◽  
Wenquan Ou ◽  
Milan Gautam ◽  
Kishwor Poudel ◽  
Bo Kyun Kim ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Targeted Delivery ◽  
Folate Receptor ◽  
A549 Cells ◽  
Kb Cells ◽  
Cell Cycle Profile ◽  
Folate Receptors ◽  
Targeted Nanoparticles ◽  
Nanoparticle System

In this study, we investigated the active targeted delivery of a hydrophobic drug, paclitaxel (PTX), via receptor-mediated endocytosis by folate receptors expressed on cancer cells using a protein-based nanoparticle system. PTX was loaded on zein nanoparticles and conjugated with folate (PTX/Zein-FA) to estimate its chemotherapeutic efficacy in folate receptor-expressing KB cancer cells. PTX/Zein-FA nanoparticles were successfully developed, with a nanoparticle size of ~180 nm and narrow polydispersity index (~0.22). Accelerated release of PTX in an acidic environment was observed for PTX/Zein-FA. An in vitro cellular study of PTX/Zein-FAs in KB cells suggested that PTX/Zein-FA improved the cytotoxic activity of PTX on folate receptors overexpressed in cancer cells by inducing proapoptotic proteins and inhibiting anti-apoptotic proteins. In addition, PTX/Zein-FA exhibited anti-migratory properties and could alter the cell cycle profile of KB cells. A549 cells, which are folate receptor-negative cancer cells, showed no significant enhancement in the in vitro cellular activities of PTX/Zein-FA. We describe the antitumor efficacy of PTX/Zein-FA in KB tumor-bearing mice with minimum toxicity in healthy organs, and the results were confirmed in comparison with free drug and non-targeted nanoparticles.

scholarly journals EML4-ALK induces cellular senescence in mortal normal human cells and promotes anchorage-independent growth in immortalized normal human cells

2020 ◽  
Author(s):  
Akihiko Miyanaga ◽  
Masaru Matsumoto ◽  
Jessica A. Beck ◽  
Izumi Horikawa ◽  
Takahiro Oike ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cellular Senescence ◽  
Human Fibroblasts ◽  
Human Cells ◽  
Functional Study ◽  
Sequencing Analysis ◽  
Anchorage Independent Growth ◽  
Normal Human ◽  
Normal Human Cells

Abstract Background: Chromosomal inversions involving anaplastic lymphoma kinase (ALK) and echinoderm microtubule associated protein like 4 (EML4) generate a fusion protein EML4-ALK in non-small cell lung cancer (NSCLC). The understanding of EML4-ALK function can be improved by a functional study using normal human cells.Methods: Here we for the first time conduct such study to examine the effects of EML4-ALK on cell proliferation, cellular senescence, DNA damage, gene expression profiles and transformed phenotypes.Results: The lentiviral expression of EML4-ALK in mortal, normal human fibroblasts caused, through its constitutive ALK kinase activity, an early induction of cellular senescence with senescence-associated b-galactosidase activity, upregulation of p16INK4A and p21WAF1, and accumulated DNA damage. In contrast, when EML4-ALK was expressed in normal human fibroblasts immortalized by telomerase reverse transcriptase (hTERT), which is activated in the vast majority of NSCLC, the cells showed accelerated proliferation and acquired anchorage-independent growth ability in soft-agar medium, without accumulated DNA damage, chromosome aberration, nor p53 mutation. EML4-ALK induced the phosphorylation of STAT3 in both mortal and immortalized cells, but RNA sequencing analysis suggested that the different STAT3-regulated signaling pathways contributed to the different phenotypic outcomes in these cells. While EML4-ALK also induced anchorage-independent growth in hTERT-immortalized human bronchial epithelial cells in vitro, the expression of EML4-ALK alone did not cause detectable in vivo tumorigenicity in immunodeficient mice.Conclusions: Our data indicate that the immortalized state is critical for EML4-ALK to manifest its in vitro transforming activity in human cells. This study provides the isogenic pairs of human cells with and without EML4-ALK expression.

scholarly journals EML4-ALK induces cellular senescence in mortal normal human cells and promotes anchorage-independent growth in hTERT-transduced normal human cells

2020 ◽  
Author(s):  
Akihiko Miyanaga ◽  
Masaru Matsumoto ◽  
Jessica A. Beck ◽  
Izumi Horikawa ◽  
Takahiro Oike ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cellular Senescence ◽  
Human Fibroblasts ◽  
Human Cells ◽  
Functional Study ◽  
Sequencing Analysis ◽  
Anchorage Independent Growth ◽  
Normal Human ◽  
Normal Human Cells

Abstract Background: Chromosomal inversions involving anaplastic lymphoma kinase (ALK) and echinoderm microtubule associated protein like 4 (EML4) generate a fusion protein EML4-ALK in non-small cell lung cancer (NSCLC). The understanding of EML4-ALK function can be improved by a functional study using normal human cells.Methods: Here we for the first time conduct such study to examine the effects of EML4-ALK on cell proliferation, cellular senescence, DNA damage, gene expression profiles and transformed phenotypes.Results: The lentiviral expression of EML4-ALK in mortal, normal human fibroblasts caused, through its constitutive ALK kinase activity, an early induction of cellular senescence with accumulated DNA damage, upregulation of p16INK4A and p21WAF1, and senescence-associated b-galactosidase (SA-β-gal) activity. In contrast, when EML4-ALK was expressed in normal human fibroblasts transduced with telomerase reverse transcriptase (hTERT), which is activated in the vast majority of NSCLC, the cells showed accelerated proliferation and acquired anchorage-independent growth ability in soft-agar medium, without accumulated DNA damage, chromosome aberration, nor p53 mutation. EML4-ALK induced the phosphorylation of STAT3 in both mortal and hTERT-transduced cells, but RNA sequencing analysis suggested that the different signaling pathways contributed to the different phenotypic outcomes in these cells. While EML4-ALK also induced anchorage-independent growth in hTERT-immortalized human bronchial epithelial cells in vitro, the expression of EML4-ALK alone did not cause detectable in vivo tumorigenicity in immunodeficient mice.Conclusions: Our data indicate that the expression of hTERT is critical for EML4-ALK to manifest its in vitro transforming activity in human cells. This study provides the isogenic pairs of human cells with and without EML4-ALK expression.

scholarly journals Effect of Cationic Lipid Type in Folate-PEG-Modified Cationic Liposomes on Folate Receptor-Mediated siRNA Transfection in Tumor Cells

Pharmaceutics ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 181 ◽  
Author(s):  
Yoshiyuki Hattori ◽  
Satono Shimizu ◽  
Kei-ichi Ozaki ◽  
Hiraku Onishi
Keyword(s):  
Gene Silencing ◽  
Tumor Cells ◽  
Folate Receptor ◽  
Cationic Lipid ◽  
Intratumoral Injection ◽  
Cationic Liposomes ◽  
Kb Cells ◽  
In Vitro Transfection

In this study, we examined the effect of cationic lipid type in folate (FA)-polyethylene glycol (PEG)-modified cationic liposomes on gene-silencing effects in tumor cells using cationic liposomes/siRNA complexes (siRNA lipoplexes). We used three types of cationic cholesterol derivatives, cholesteryl (3-((2-hydroxyethyl)amino)propyl)carbamate hydroiodide (HAPC-Chol), N-(2-(2-hydroxyethylamino)ethyl)cholesteryl-3-carboxamide (OH-Chol), and cholesteryl (2-((2-hydroxyethyl)amino)ethyl)carbamate (OH-C-Chol), and we prepared three types of FA-PEG-modified siRNA lipoplexes. The modification of cationic liposomes with 1–2 mol % PEG-lipid abolished the gene-silencing effect in human nasopharyngeal tumor KB cells, which overexpress the FA receptor (FR). In contrast, FA-PEG-modification of cationic liposomes restored gene-silencing activity regardless of the cationic lipid type in cationic liposomes. However, the optimal amount of PEG-lipid and FA-PEG-lipid in cationic liposomes for selective gene silencing and cellular uptake were different among the three types of cationic liposomes. Furthermore, in vitro transfection of polo-like kinase 1 (PLK1) siRNA by FA-PEG-modified liposomes exhibited strong cytotoxicity in KB cells, compared with PEG-modified liposomes; however, in in vivo therapy, intratumoral injection of PEG-modified PLK1 siRNA lipoplexes inhibited tumor growth of KB xenografts, as well as that of FA-PEG-modified PLK1 siRNA lipoplexes. From these results, the optimal formulation of PEG- and FA-PEG-modified liposomes for FR-selective gene silencing might be different between in vitro and in vivo transfection.

scholarly journals Folate-Targeted mRNA Delivery Using Chitosan-Functionalized Selenium Nanoparticles: Potential in Cancer Immunotherapy

2019 ◽  
Vol 12 (4) ◽  
pp. 164 ◽  
Author(s):  
Maiyo ◽  
Singh
Keyword(s):  
Messenger Rna ◽  
Folate Receptor ◽  
Selenium Nanoparticles ◽  
Breast Adenocarcinoma ◽  
Kb Cells ◽  
Selenium Uptake ◽  
Ethidium Bromide Staining ◽  
Low Cytotoxicity ◽  
Uptake Studies

Systemic messenger RNA (mRNA) delivery, although still in its infancy, holds immense potential for application in cancer vaccination and immunotherapy. Its advantages over DNA transfection make it attractive in applications where transient expression is desired. However, this has proved challenging due to mRNA’s instability and susceptibility to degradation. Selenium is important for immune function and modulation, with selenium nanoparticles (SeNPs) finding a niche in biomedicine as drug delivery vehicles, owing to their biocompatibility, low toxicity, and biodegradability. In this investigation, we synthesized chitosan-coated SeNPs with a folic acid targeting moiety for Fluc mRNA delivery to cancer cells in vitro. Synthesized SeNPs were stable and well dispersed, and ranged from 59 to 102 nm in size. Nanoparticles bound and protected mRNA from RNase degradation, while exhibiting low cytotoxicity in the human embryonic kidney (HEK293), breast adenocarcinoma (MCF-7), and nasopharyngeal (KB) cells in culture. Moderate cytotoxicity evidenced in the colorectal carcinoma (Caco-2) and colon carcinoma (HT-29) cells was attributed to apoptosis induction by selenium, as confirmed by acridine orange/ethidium bromide staining. Selenium uptake studies corroborated the transfection results, where significant transgene expression was evident for the overexpressed folate receptor-positive KB cells when compared to the other cells with less or no folate receptors.

scholarly journals EML4-ALK induces cellular senescence in mortal normal human cells and promotes anchorage-independent growth in hTERT-transduced normal human cells

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Akihiko Miyanaga ◽  
Masaru Matsumoto ◽  
Jessica A. Beck ◽  
Izumi Horikawa ◽  
Takahiro Oike ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cellular Senescence ◽  
Human Fibroblasts ◽  
Human Cells ◽  
Functional Study ◽  
Sequencing Analysis ◽  
Anchorage Independent Growth ◽  
Normal Human ◽  
Normal Human Cells

Abstract Background Chromosomal inversions involving anaplastic lymphoma kinase (ALK) and echinoderm microtubule associated protein like 4 (EML4) generate a fusion protein EML4-ALK in non-small cell lung cancer (NSCLC). The understanding of EML4-ALK function can be improved by a functional study using normal human cells. Methods Here we for the first time conduct such study to examine the effects of EML4-ALK on cell proliferation, cellular senescence, DNA damage, gene expression profiles and transformed phenotypes. Results The lentiviral expression of EML4-ALK in mortal, normal human fibroblasts caused, through its constitutive ALK kinase activity, an early induction of cellular senescence with accumulated DNA damage, upregulation of p16INK4A and p21WAF1, and senescence-associated β-galactosidase (SA-β-gal) activity. In contrast, when EML4-ALK was expressed in normal human fibroblasts transduced with telomerase reverse transcriptase (hTERT), which is activated in the vast majority of NSCLC, the cells showed accelerated proliferation and acquired anchorage-independent growth ability in soft-agar medium, without accumulated DNA damage, chromosome aberration, nor p53 mutation. EML4-ALK induced the phosphorylation of STAT3 in both mortal and hTERT-transduced cells, but RNA sequencing analysis suggested that the different signaling pathways contributed to the different phenotypic outcomes in these cells. While EML4-ALK also induced anchorage-independent growth in hTERT-immortalized human bronchial epithelial cells in vitro, the expression of EML4-ALK alone did not cause detectable in vivo tumorigenicity in immunodeficient mice. Conclusions Our data indicate that the expression of hTERT is critical for EML4-ALK to manifest its in vitro transforming activity in human cells. This study provides the isogenic pairs of human cells with and without EML4-ALK expression.

Sign in / Sign up

Export Citation Format

Share Document