Hydrophilic Chlorin e6-Poly(amidoamine) Dendrimer Nanoconjugates for Enhanced Photodynamic Therapy
"> Figure 1
<p>Schematic diagram of synthesis of hydrophilic Ce6-PAMAM dendrimer nanoconjugate (DC).</p> "> Figure 2
<p>(<b>a</b>) TEM images and (<b>b</b>) particle size distribution of DC.</p> "> Figure 3
<p>(<b>a</b>) Photographs of free chlorin e6 (Ce6) and DC dissolved in DMSO or DW (0.2 mg/mL Ce6). The arrow shows the precipitate of Ce6. (<b>b</b>) UV-visible absorbance spectra and (<b>c</b>) fluorescence emission spectra (λ<sub>ex</sub> = 360 nm) of free Ce6 and DC in DMSO or DW (10 μg/mL Ce6).</p> "> Figure 4
<p>9,10-Dimethylnathracene (DMA) fluorescence change (F<sub>f</sub> − F<sub>s</sub>) due to singlet oxygen generation by free Ce6 and DC after laser irradiation in Dulbecco’s phosphate-buffered saline (DPBS) (pH 7.4), where F<sub>f</sub> and F<sub>s</sub> represent the fluorescence intensity of full DMA and each sample.</p> "> Figure 5
<p>Change in fluorescence intensity due to intracellular uptake of free Ce6 and DC (4 μg/mL Ce6) in HeLa cells as a function of incubation time (<span class="html-italic">n</span> = 5).</p> "> Figure 6
<p>Confocal laser scanning microscopy image of the intracellular distribution of (<b>a</b>) free Ce6 and (<b>b</b>) DC in HeLa cells after incubation for 2 h in the dark.</p> "> Figure 7
<p>In vitro phototoxicity and dark-toxicity of various concentrations of Ce6 and DC on HeLa cells with and without laser irradiation (<span class="html-italic">n</span> = 6).</p> "> Figure 8
<p>Fluorescence microscopy images of live HeLa cells and dead HeLa cells stained with calcein-AM (green) and ethidium homodimer-1 (EthD-1) (red) with (<b>a</b>) no drug, (<b>b</b>) free Ce6, and (<b>c</b>) DC (4 μg/mL Ce6) after irradiation by 2.5 J/cm<sup>2</sup>, 671 nm diode laser.</p> "> Figure 9
<p>Flow cytometric analysis of ROS generation in HeLa cells treated with free Ce6 and DC (4 μg/mL Ce6). (<b>a</b>) Shift in fluorescence peak due to the ROS generation in the presence of free Ce6 and DC before irradiation (Ce6(−) and DC(−)) and after irradiation (Ce6(+) and DC(+)) with a 2.5-J/cm<sup>2</sup>, 671-nm diode laser. (<b>b</b>) DCF fluorescence intensity measured with the flow cytometer for investigating the ROS level with free Ce6 and DC before and after irradiation (<span class="html-italic">n</span> = 5).</p> "> Figure 10
<p>Morphology of HeLa cells stained with neutral red and H-33258 (<b>a</b>) before photodynamic treatment (no drug and no light) and after treatment with (<b>b</b>) free Ce6 and (<b>c</b>) DC (4 μg/mL Ce6), followed by irradiation with a 2.5-J/cm<sup>2</sup>, 671-nm diode laser.</p> ">
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Synthesis of Ce6-PAMAM Dendrimer Nanoconjugate
2.3. Characterization of Ce6-PAMAM Dendrimer Nanoconjugate
2.4. Singlet Oxygen Detection
2.5. Cell Culture and Incubation Conditions
2.6. Intracellular Uptake and Distribution Tests
2.7. Cell Phototoxicity Assay
2.8. Generation of Reactive Oxygen Species
2.9. Apoptotic Analysis
3. Results and Discussion
3.1. Synthesis and Characterization of DC Nanoconjugate
3.2. Detection of Singlet Oxygen Production
3.3. Intracellular Uptake and Distribution of DC Nanoconjugate
3.4. In Vitro Phototoxicity of DC Nanoconjugate
3.5. Induction of Cell Apoptosis by DC Nanoconjugate
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Conflicts of Interest
References
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Lee, S.-R.; Kim, Y.-J. Hydrophilic Chlorin e6-Poly(amidoamine) Dendrimer Nanoconjugates for Enhanced Photodynamic Therapy. Nanomaterials 2018, 8, 445. https://doi.org/10.3390/nano8060445
Lee S-R, Kim Y-J. Hydrophilic Chlorin e6-Poly(amidoamine) Dendrimer Nanoconjugates for Enhanced Photodynamic Therapy. Nanomaterials. 2018; 8(6):445. https://doi.org/10.3390/nano8060445
Chicago/Turabian StyleLee, So-Ri, and Young-Jin Kim. 2018. "Hydrophilic Chlorin e6-Poly(amidoamine) Dendrimer Nanoconjugates for Enhanced Photodynamic Therapy" Nanomaterials 8, no. 6: 445. https://doi.org/10.3390/nano8060445