Amphiphilic Polypeptides for VEGF siRNA Delivery into Retinal Epithelial Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Methods
2.2.1. Synthesis and Characterization of Polypeptides
2.2.2. Preparation and Characterization of Polypeptide Particles
2.2.3. Encapsulation and Release of RNA, Duplex Oligo-dT-dA and siRNA
2.2.4. Cytotoxicity of Particles
2.2.5. Cellular Uptake
2.2.6. VEGF Gene Silencing
2.2.7. Total RNA Isolation, Reverse Transcription and Quantitative Real-Time PCR Analysis
2.2.8. Western Blotting
2.2.9. Statistical Analysis
3. Results and Discussion
3.1. Synthesis and Characterization of Polypeptides
3.2. Preparation and Characterization of Polypeptide Particles
3.3. Entrapment and Release of Duplex Oligo-dT-dA
3.4. Cytotoxicity
3.5. VEGF Gene Silencing
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Sample | Polymer Characteristics | |||
---|---|---|---|---|
SEC | SLS | |||
Mn | Mw | Ð | Mw | |
P(Lys(Z)n-co-Glu(OBzl)m-co-Phek) | ||||
KEF1 | 20,170 | 23,600 | 1.17 | 23,000 |
KEF2 | 9400 | 17,000 | 1.80 | 17,500 |
KEF3 | 10,000 | 13,200 | 1.32 | 17,400 |
P(Lys(Z)n-co-Glu(OBzl)m-co-Ilek) | ||||
KEI1 | 21,350 | 28,400 | 1.33 | 18,800 |
KEI2 | 16,090 | 21,240 | 1.32 | 17,100 |
KEI3 | 17,560 | 22,300 | 1.27 | - |
Sample | Determined Polymer Composition (mol%) | ||||
---|---|---|---|---|---|
HPLC | 1HNMR | ||||
Lys | Glu | Phe/Ile | Lys + Glu | Phe/Ile | |
P(Lysn-co-Glum-co-Phek) | |||||
KEF1 | 55 | 25 | 20 | 76 | 24 |
KEF2 | 75 | 9 | 16 | 75 | 25 |
KEF3 | 21 | 54 | 25 | 64 | 36 |
P(Lysn-co-Glum-co-Ilek) | |||||
KEI1 | 66 | 16 | 18 | 85 | 15 |
KEI2 | 57 | 31 | 12 | 88 | 12 |
KEI3 | 54 | 36 | 10 | - | - |
Sample | Particle Characteristics | ||
---|---|---|---|
DH, nm | PDI | ζ-Potential, mV | |
P(Lysn-co-Glum-co-Phek) | |||
KEF1 | 200 ± 8 | 0.22 | +12 ± 2 |
KEF2 | 550 ± 14 | 0.31 | +18 ± 5 |
KEF3 | 900 ± 38 | 0.39 | −6 ± 3 |
P(Lysn-co-Glum-co-Ilek) | |||
KEI1 | 232 ± 11 | 0.14 | +49 ± 2 |
KEI2 | 180 ± 19 | 0.19 | +45 ± 3 |
KEI3 | 440 ± 31 | 0.30 | +31 ± 5 |
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Osipova, O.; Sharoyko, V.; Zashikhina, N.; Zakharova, N.; Tennikova, T.; Urtti, A.; Korzhikova-Vlakh, E. Amphiphilic Polypeptides for VEGF siRNA Delivery into Retinal Epithelial Cells. Pharmaceutics 2020, 12, 39. https://doi.org/10.3390/pharmaceutics12010039
Osipova O, Sharoyko V, Zashikhina N, Zakharova N, Tennikova T, Urtti A, Korzhikova-Vlakh E. Amphiphilic Polypeptides for VEGF siRNA Delivery into Retinal Epithelial Cells. Pharmaceutics. 2020; 12(1):39. https://doi.org/10.3390/pharmaceutics12010039
Chicago/Turabian StyleOsipova, Olga, Vladimir Sharoyko, Natalia Zashikhina, Natalya Zakharova, Tatiana Tennikova, Arto Urtti, and Evgenia Korzhikova-Vlakh. 2020. "Amphiphilic Polypeptides for VEGF siRNA Delivery into Retinal Epithelial Cells" Pharmaceutics 12, no. 1: 39. https://doi.org/10.3390/pharmaceutics12010039