Study of the influence of the extract of pipsissewa on cell cultures

Abstract

The development of new diuretics of plant origin is an actual direction. Chimaphila umbellata (L.) is a perennial herb with diuretic, astringent, analgesic and other effects; and it can treat various conditions such as edema, dropsy, etc. Pipsissewa herb helps the removal of nitrogenous and chloride salts from the body due to the content of arbutin glycoside, tannins (up to 5 %). The aim. Evaluation of the effect of pipsissewa extract on L929 cell culture. Materials and methods. Cell line L929 (fibroblasts of mouse adipose tissue) was obtained in the low-temperature bank of the Institute of Problems of Cryobiology and Cryomedicine of the National Academy of Sciences of Ukraine. Cells were cultured in DMEM medium (Bio West, France) enriched with 10 % FBS (Lonza, Germany) with 1 % antibiotic-antimycotic (Bio West, France), in a CO2 incubator (Thermo Fisher Scientific, USA) at 37 ºС in an atmosphere with 5 % CO2. Determination of the minimum toxic concentration at which the cells remained alive was evaluated by morphological features (shape, monolayer integrity, adhesion to plastic). The study of the effect of pipsissewa extract on various cell functions was determined by the following methods: the ability to preserve morphological integrity - by the phase-contrast microscopy method, energy exchange - by the MTT test method, pinocytotic function - by the neutral red absorption method, migratory function - by the scratch test method, proliferative activity - by the doubling calculation method population Results. It is proposed to use concentrations of 0.05, 0.02, 0.01, 0.005 % of pipsissewa extract for further research. After carrying out the MTT reaction, the transition of MTT to formazan was confirmed microscopically in the negative control (native cells), at PE concentrations of 0.01 % and below, and the absence of a reaction in the positive control (cells killed by ethanol) at PE concentrations above 0.02 %. When recording the parameters of the NP absorption reaction, it was determined that PE at a concentration of 0.02 % and higher sharply suppresses pinocytotic activity, despite the partial preservation of cell adhesion, reducing the concentration by two times no longer affects mitochondria. A concentration of 0.01 % reduces proliferative activity, and at a concentration of 0.005 %, no difference with the control values ​​was found. Conclusions. When studying the assessment of the effect of pipsisewa extract on L929 cell culture, a toxic effect on these cells was established when added to the culture medium at a concentration above 0.01 %. The toxic effect had a threshold effect. Migratory and proliferative functions were the most sensitive, energy, pinocytosis and preservation of morphological integrity of cells were less sensitive