Lipofection

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The lipofection describes the transfection with liposomes , vesicles or micelles .

properties

By lipofection, nucleic acids (e.g. a vector ) are usually introduced into cells by binding to or inclusion in liposomes . After endocytosis , the liposomes fuse with the endosome membrane .

In the case of liposomes, vesicles or micelles with cationic lipids , complexation with the anionic nucleic acids can take place (nucleic acid adsorption ); in the case of uncharged lipids and membrane lipids , the nucleic acids must be enclosed in liposomes or vesicles by immobilization . Inclusion immobilization in liposomes or their production requires a higher expenditure of time than adsorption, but it can also smuggle liquids and thus also molecules with a less negatively charged surface (see zeta potential ) into cells.

Inclusion immobilization

Inclusion immobilization uses e.g. B. the ether- lipid infusion, a syringe with a membrane filter or ultrasound to generate the liposomes.

Less common methods are e.g. B. the preparation of erythrocytes - ghosts (English for 'ghosts'). Erythrocytes are lysed in a hypotonic solution. After the cell contents escape, the remaining emptied cell membranes are spontaneously resealed , including the surrounding medium to which the nucleic acids were previously added. The transfection is carried out by fusing the cell membrane analogous to the cell fusion . The method is relatively complex, but can take up relatively large volumes and release them into the target cell.

adsorption

The adsorption of nucleic acids is achieved by adding nucleic acids to a suspension with cationic lipids. Here, cationic lipids such as. B. DOTMA (N- [1- (2,3-Dioleyloxy) propyl] -N, N, N-trimethylammonium chloride), DOTAP (1,2-dioleoyl-3-trimethylammonium propane), DDA (dimethyldioctadecylammonium), DC- Chol (3b-N- (dimethylaminoethyl) carbamate-cholesterol) or DOSPER (1,3-di-oleoyloxy-2- (6-carboxy-spermyl) -propylamide) are used in conjunction with membrane lipids such as phosphatidylethanolamine , phosphatidylcholine and cholesterol . Due to the toxicity of the cationic lipids, the optimal ratios and amounts of lipids and nucleic acids for a cell type must be tested. The cationic lipids can trigger pro- apoptotic and pro-inflammatory effects. After an intravenous injection, a large part of the DNA-lipid complexes accumulates in the lungs . At least in the case of DNA (which are used by the same methods as RNA and are taken up by cells via the same mechanisms) there is only a weak correlation between uptake in cell culture and in vivo and no correlation between uptake in cell culture and the vaccination effect.

literature

Individual evidence

  1. ↑ a b S. E. McNeil, A. Vangala, VW Bramwell, PJ Hanson, Y. Perrie: Lipoplexes formulation and optimization: in vitro transfection studies reveal no correlation with in vivo vaccination studies. In: Curr Drug Deliv . (2010), Volume 7, No. 2, pp. 175-187. PMID 20158478 .
  2. ↑ C. Lonez, M. Vandenbranden, JM Ruysschaert: Cationic lipids activate intracellular signaling pathways. In: Adv Drug Deliv Rev . (2012), Volume 64, No. 15, pp. 1749-1758. doi : 10.1016 / j.addr.2012.05.009 . PMID 22634161 .
  3. ↑ W. Yeeprae, S. Kawakami, S. Suzuki, F. Yamashita, M. Hashida: Physicochemical and pharmacokinetic characteristics of cationic liposomes. In: Pharmazie (2006), Volume 61, pp. 2102-2105. PMID 16526555 .
  4. ↑ K. Paunovska, CD Sago, CM Monaco, WH Hudson, MG Castro, TG Rudoltz, S. Kalathoor, DA Vanover, PJ Santangelo, R. Ahmed, AV Bryksin, JE Dahlman: A Direct Comparison of in Vitro and in Vivo Nucleic Acid Delivery Mediated by Hundreds of Nanoparticles Reveals a Weak Correlation. In: Nano letters. Volume 18, number 3, 03 2018, pp. 2148-2157, doi : 10.1021 / acs.nanolett.8b00432 , PMID 29489381 , PMC 6054134 (free full text).