Advantages of nanofibrous membranes for culturing of primary RPE cells compared to commercial scaffolds

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F67985904%3A_____%2F22%3A00560631" target="_blank" >RIV/67985904:_____/22:00560631 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/61389013:_____/22:00560631 RIV/68378041:_____/22:00560631 RIV/00064173:_____/22:43922457 RIV/00216208:11120/22:43922457 a 2 dalších

Výsledek na webu

<a href="https://asep.lib.cas.cz/arl-cav/cs/csg/?repo=crepo1&key=70763693065" target="_blank" >https://asep.lib.cas.cz/arl-cav/cs/csg/?repo=crepo1&key=70763693065</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1111/aos.15034" target="_blank" >10.1111/aos.15034</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Advantages of nanofibrous membranes for culturing of primary RPE cells compared to commercial scaffolds

Popis výsledku v původním jazyce

Purpose Dysfunction of the retinal pigment epithelium (RPE) causes numerous forms of retinal degeneration. RPE replacement is a modern option to save vision. We aimed to test the results of transplanting cultured RPEs on biocompatible membranes. Methods We cultivated porcine primary RPE cells isolated from cadaver eyes from the slaughterhouse on two types of membranes: commercial polyester scaffolds Transwell (Corning Inc., Kenneburg, ME, USA) with 0.4 mu m pore size and prepared Poly (L-lactide-co-DL-lactide) (PDLLA) nanofibrous membranes with an average pore size of 0.4 mu m. Results Five types of assays were used for the analysis: immunocytochemistry (ICC), phagocytosis assay, Western blotting, real-time qPCR (RT-qPCR) and electron microscopy. RT-qPCR demonstrated that RPEs cultured on nanofibrous membranes have higher expressions of BEST1 (bestrophin 1), RLBP1 (retinaldehyde-binding protein 1), RPE65 (retinal pigment epithelium-specific 65 kDa protein), PAX6 (transcription factor PAX6), SOX9 (transcription factor SOX9), DCT (dopachrome tautomerase) and MITF (microphthalmia-associated transcription factor). ICC of the RPEs cultured on nanofibrous membranes showed more intensive staining of markers such as BEST1, MCT1 (monocarboxylate transporter 1), Na+/K(+)ATPase, RPE65 and acetylated tubulin in comparison with commercial ones. Additionally, the absence of alpha-SMA proved the stability of the RPE polarization state and the absence of epithelial-to-mesenchymal transition. RPE possessed high phagocytic activity. Electron microscopy of both membranes confirmed a confluent layer of RPE cells and their genuine morphological structure, which was comparable to native RPEs. Conclusions Retinal pigment epitheliums cultured on polylactide nanofibrous membranes improved the final quality of the cell product by having better maturation and long-term survival of the RPE monolayer compared to those cultured on commercial polyester scaffolds. PDLLA-cultured RPEs are a plausible source for the replacement of non-functioning RPEs during cell therapy.

Název v anglickém jazyce

Advantages of nanofibrous membranes for culturing of primary RPE cells compared to commercial scaffolds

Popis výsledku anglicky

Purpose Dysfunction of the retinal pigment epithelium (RPE) causes numerous forms of retinal degeneration. RPE replacement is a modern option to save vision. We aimed to test the results of transplanting cultured RPEs on biocompatible membranes. Methods We cultivated porcine primary RPE cells isolated from cadaver eyes from the slaughterhouse on two types of membranes: commercial polyester scaffolds Transwell (Corning Inc., Kenneburg, ME, USA) with 0.4 mu m pore size and prepared Poly (L-lactide-co-DL-lactide) (PDLLA) nanofibrous membranes with an average pore size of 0.4 mu m. Results Five types of assays were used for the analysis: immunocytochemistry (ICC), phagocytosis assay, Western blotting, real-time qPCR (RT-qPCR) and electron microscopy. RT-qPCR demonstrated that RPEs cultured on nanofibrous membranes have higher expressions of BEST1 (bestrophin 1), RLBP1 (retinaldehyde-binding protein 1), RPE65 (retinal pigment epithelium-specific 65 kDa protein), PAX6 (transcription factor PAX6), SOX9 (transcription factor SOX9), DCT (dopachrome tautomerase) and MITF (microphthalmia-associated transcription factor). ICC of the RPEs cultured on nanofibrous membranes showed more intensive staining of markers such as BEST1, MCT1 (monocarboxylate transporter 1), Na+/K(+)ATPase, RPE65 and acetylated tubulin in comparison with commercial ones. Additionally, the absence of alpha-SMA proved the stability of the RPE polarization state and the absence of epithelial-to-mesenchymal transition. RPE possessed high phagocytic activity. Electron microscopy of both membranes confirmed a confluent layer of RPE cells and their genuine morphological structure, which was comparable to native RPEs. Conclusions Retinal pigment epitheliums cultured on polylactide nanofibrous membranes improved the final quality of the cell product by having better maturation and long-term survival of the RPE monolayer compared to those cultured on commercial polyester scaffolds. PDLLA-cultured RPEs are a plausible source for the replacement of non-functioning RPEs during cell therapy.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

30401 - Health-related biotechnology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2022

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Acta Ophthalmologica

ISSN

1755-375X

e-ISSN

1755-3768

Svazek periodika

100

Číslo periodika v rámci svazku

5

Stát vydavatele periodika

DK - Dánské království

Počet stran výsledku

14

Strana od-do

"E1172"-"E1185"

Kód UT WoS článku

000710006200001

EID výsledku v databázi Scopus

2-s2.0-85117446785