Saratov JOURNAL of Medical and Scientific Research

St. Petersburg Polytechnic University

Experimental study of synthetic polymeric materials as a basis in the creation of the advaced carrier matrix for the cultivation of limbal stem cells

Year: 2019, volume 15 Issue: №2 Pages: 495-501
Heading: Ophtalmology Article type: Original article
Authors: Karpovich V.V., Kulikov A.N., Churashov S.V., Chernysh V.F., Grigoriev S.G., Blinova M.I., Nashchekina Yu.A., Aleksandrova O.I., Khorolskaya Yu.I., Nikonov P.O., Tsobkallo E.S., Moskalyuk O.A., Melnikov A.S., Serdobintsev P.Yu., Mashel' T.V., Pisugina G.A., Perepletchikova D.A., Khoroshikh D.A.
Organization: Institute of Cytology, Russian Academy of Sciences, St. Petersburg Polytechnic University, St. Petersburg State University, St. Petersburg State University of Industrial Technology and Design
Summary:

Purpose: to study in the experiment the properties of three diferent types of synthetic polyester matrices, to conduct their comparative assessment and determine the optimum as a carrier for the cultivation and transplantation of limbal stem cells while eliminating limbal insufciency. Material and Methods. Transparency, mechanical properties (strength, elongation at break, modulus of rigidity), biocompatibility with cell cultures of the cornea, as well as the study of the timing of matrix biodegradation in vivo were carried out. Results. The study examined the optical and mechanical properties of matrices made from polylactide-glycolide (PLG), polylactide-caprolactone (PLC) and poly-e-caprolactone (PCL). It was experimentally shown that human and rabbit limbal stem cells, as well as human corneal epithelium cells, adhered on the surface of all types of matrices under investigation, and during cultivation they retained the typical structure of actin cytoskeleton, the ability to proliferate and migrate. Diferences in the interaction of diferent cell cultures with diferent types of carriers were revealed. The terms of biodegradation of PLC matrices with a thickness of 5 μm was about 30 days. Conclusion. The results indicate that it is possible to use 5 μm thick PLC matrices as a carrier of cultured limbal stem cells.

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On preparation of am-niotic membrane as a scafold for cultivated cells to create corneal bioengineering constructs

Year: 2019, volume 15 Issue: №2 Pages: 409-413
Heading: Ophtalmology Article type: Original article
Authors: Aleksandrova OI, Gavrilyuk IO, Mashel TV, Chernysh VF, Churashov SV, Kulikov AN, Blinova MI.
Organization: Institute of Cytology, Russian Academy of Sciences, Military Medical Academy n.a. S. M. Kirov, St. Petersburg Polytechnic University
Summary:

Aim: to determine the preparation of amniotic membrane (AM) necessary for its use as a scafold for cultured cells to create bioengineered constructions (BEC). Material and Methods. Native AM was placed in a special clamping device and subjected to additional mechanical, thermal and enzymatic treatment: removal of mucus residues from its surface and cryoconservation of AM scafolds was performed at –80°C, — 20°C with subsequent decellularization with a 0.25 % Tripsin-EDTA mixture. The lifetime assessment of the morphology of cells cultivated on AM scafolds was performed using a Nikon Eclipse TS100 inverted microscope equipped with camera. The viability and metabolic activity of AM cells was determined by means of an MTT test using a UNIFLAN AIFR-01 tablet spectrophotometer (Picon, Russia) at a wavelength of 570 nm and a reference wavelength of 620 nm. Results. It has been established that the presence of mucous residueson the surface of native AM, which are not removed during standard mechanical processing, negatively afect the survival of the cell test system. The efects of cryoconservation of AM scafolds revealed the positive efects of this process together with enzymatic decellularization for improving the viability of cells cultured on scafolds. Conclusion. Standard mechanical processing of native AM does not guarantee complete cleaning of its surface from mucus residues that interfere with adhesion and even distribution of cultured cells. It is necessary to reliably control the removal of AM surface mucus before immobilization and further manipulations. Cryopreservation and subsequent decellularization of AM scafolds contributes to the increased viability of the cell test system. AM scafolds, purifed from amniotic mucus, cryopreserved at –80 ° C in a mixture of DMEM-F12 and DMSO (1:1) and enzymatic decellularization after thawing, turned out to be the best of the studied substrates for cell cultivation.

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