Makarova M. Morphological changes of corneal tissues in experimental keratitis and silver nanoparticles influence

Українська версія

Thesis for the degree of Candidate of Sciences (CSc)

State registration number

0421U100840

Applicant for

Specialization

  • 14.03.09 - Гістологія, цитологія, ембріологія

07-04-2021

Specialized Academic Board

Д 20.601.02

Ivano-Frankivsk national medical university

Essay

The thesis is dedicated to the study of silver nanoparticles with size of 30 nm obtained by the citrated and electron beam synthesis methods and their impact on the anteriorepithelium and corneal stroma of the rabbits. The objective of the work was to study with the usage of general morphological, morphometrical and electron microscopic methods the morphological and the ultrastructural changes in the corneal tissues of the healthyrabbit's eyesand in experimental keratitis under conditions of exposure to silver nanoparticles in various forms (instillations and application of films). Using the histoimmunocytochemical method, we studied the impact of silver nanoparticles with size of 30 nm on the expression oflymphocyte activation molecular markers using the panel of monoclonal antibodies CD5+, СD7+, CD25+, CD38+, CD45+, CD54+, CD95+ and CD150+ before and after the cultivation of silver nanoparticles with the peripheral blood lymphocytes of the patients with viral keratitis. Simulation of bacterial keratitis was carried out by means of layer-by-layer cornea trepanation in the optic zone up to 2/3 of the stroma thickness with the subsequent stromectomy and instillation of Staphylococcus aureus culture into the conjunctival cavity and subconjunctival administration of betamethasone, as well as daily instillations of dexamethasone solution for 5 days. All the animals underwent clinical and microbiological testing, and they were excluded fromthe experiment on the 3rd and 6th days and their eye bulbs were taken for morphological studies. The material was studied under supermicroscope PEM-100 - 01. It has been established that the application of films containing silver nanoparticles with size of 30 nm, obtained by citrate method in the rabbits with bacterial keratitis, facilitates the decrease of exudative phase of inflammatory process, strengthening of proliferative response, which leads to regeneration of the anterior corneal epithelium and quick stroma regeneration, through the increase of the amount of fibroblasts, and also to the strengthening of protein synthetic ability and active formation of tender collagenic acontoid proper to the healthy corneal tissue. It was detected that microbicidal, immune modulating properties of nanoparticles and their presentation in the nidus in the living organism depend on the synthetic procedure of nanodimensional structures and form of administration. It has been developed and implemented into practice the determination method of receptor modifying influence of silver nanoparticles on the activation molecules of lymphoid cells, which can be used for evaluation ofbiological influenceof metals on blood cells functional activity. The conducted studiesshowed that the exposureof silver nanoparticles with size of 30 nmensure the increase of activity of the receptor apparatus of immunocompetent cells, the strengthening of specialized functions of lymphocytes, which take part in the specific immune response in the species with viral keratitis. Therefore,in healthy donors the level of expression of molecular markersof activation was significantly increased CD5+ by 38%; CD25+ by 98%; CD38+ by 71,4%; CD45+ by 50,5%; CD54+ by 74,3%; CD95+ by 81,1%; CD150+ by 51,5%; the highest activation in 4,1 times was noticed in the marker CD7+ (р<0,05). In species with viral keratitis only the level of expression of CD7+ increased by 29,9%, CD25+ by 39,5%, and CD45+ by 34,4% (p<0,05). The cultivation of neutrophils with silver nanoparticles increases the phagocytic activity of neutrophils by 35,7% in patients with viral keratitis, and in donors by 24,9% (p<0,05). It can therefore be concluded that the immunomodulatory effect of silver nanoparticles is noted both in relation to lymphoid cells and neutrophils. Key words: cornea, keratitis, silver nanoparticles, markers of activation of lymphocytes. Branch-Medicine.

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