Рязанский государственный медицинский университет имени академика И.П. Павлова, Рязань, Российская Федерация _____________________________________________________________________________ Эндотелиальные клетки являются функционально-ведущим типом клеток внутренней оболочки сосудов и выполняют множество важных функций, включая поддержание гемостаза, регуляцию сосудистого тонуса, роста сосудов и процессов воспаления. Дисфункция эндотелия ассоциирована с широким спектром заболеваний, включая атеросклероз, артериальную гипертензию, сахарный диабет, аутоиммунные, инфекционные, онкологические заболевания и другие. В обзоре рассмотрены основные аспекты эмбрионального развития и морфологические особенности эндотелиальных клеток, описаны процессы васкуло-, ангио-и артериогенеза, представлены ключевые биологически активные вещества эндотелиального происхождения, а также иммуноцитохимические маркеры, позволяющие идентифицировать принадлежность к эндотелиоцитам. Информация, изложенная в статье, поможет читателю получить знания об эндотелиоцитах, что в эру активного развития клеточной биологии и молекулярной медицины важно для понимания патофизиологии и современных методов лечения пациентов с заболеваниями, ассоциированными с дисфункцией эндотелия. Ключевые слова: эндотелий; ангиогенез; васкулогенез; эндотелиальные маркеры; эндотелиальная дисфункция.
Aim. To study and compare cytotoxicity of the main types of synthetic prostheses used in arterial reconstructive surgery, including polytetrafluoroethylene (PTFE) and polyethylene-terephthalate (Dacron). Materials and Methods. On the culture of human umbilical vein endothelial cells (HUVEC) of the 3rd passage, MTS test was conducted that is used in laboratory examinations with attraction of cellular technologies to study cytotoxicity of medical drugs and medical products. The test implies use of MTS reagent that is 3-(4,5-dimethylthiazol-2-il)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium; additionally phenazine methosulfate (PMS) was used that plays the role of electron-binding reagent. In the experiment, cells were incubated with PTFE and Dacron within 24 hours at 37ᵒC with 5% CO2. For control, HUVEC cultured in the standard growth medium, were used. In the presence of PMS, MTS was reduced by mitochondrial dehydrogenases of endothelial cells to formazan staining blue. Supernatant of cell cultures was evaluated by photocolorimetric method on Stat Fax 3200 analyzer (microplate reader) of Awareness technology Inc. Palm City Fl. (USA). Results. The lowest mean values were noted in Dacron group 0.21 (0.20-0.22) optical density units, the highest values were noted in the control group 0.36 (0.35-0.38); parameters in PTFE group were 0.35 (0.33-0.36). In comparison of the groups statistically significant differences were found between the control group and Dacron group (р0.001), control and PTFE group (р=0.037), Dacron and PTFE (р0.001). Incubation with Dacron led to suppression of metabolic activity of cells by 41.7% as compared to the control group (р0.001). Metabolic activity of cells exposed to PTFE, approached that of the control group, that is, it corresponded to the optimal conditions of culturing of endothelial cells in vitro. Conclusion. In comparison with polyethylene-terephthalate (Dacron), polytetrafluoro-ethylene (PTFE) showed the least suppression of metabolic activity of endothelial cells in vitro.
The second part of the article aims at delivering stateof-the art information of the basics of laboratory work with endothelial cells, including main endothelial cell culture lines for in vitro experiments, 2D and 3D cultivation of endothelial cells. The authors also discuss current issues and perspectives of laboratory research work involving endothelial cells, derived from embryonic stem cells and induced pluripotent stem cells. In vivo study of endothelial cells and assessment of their morphology and function are associated with substantial technical difficulties, which necessitates the use of in vitro techniques aimed at evaluating pathogenesis and potential treatment methods of a number of diseases associated with endothelial dysfunction. Laboratory work involving cell cultures allows studying both physiological and pathological conditions using miniature and often automated systems, which has several considerable benefits over clinical trials and experiments carried out on animals. In vitro techniques have certain limitations, among which are the difficulties in interpreting the results of a work performed on a cell line and comparing those to in vivo conditions. Constant improvement of laboratory methods involving different endothelial cell lines, co-cultures, 3D cultivation systems, and induced pluripotent stem cells, is crucial. Understanding the principles of in vitro work with endothelial cell cultures and optimization of the current techniques will lead to the development and implementation of the accurate laboratory methods, which will help translate research into practice.
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