In current in vitro study we have shown the impact of deuterium content in growth medium on proliferation rate of human cultured adipose-derived stem cells (ADSC). ADSCs have also demonstrated morphological changes when cultured in deuterated growth medium: the cell cultures did not reach confluence but acquired polygonal morphology with pronounced stress fibers. At high deuterium concentrations the ADSCs population doubling time increased which indicated the cell cycle retardation and decrease of cell proliferation rate. The deuterated and deuterium-depleted growth media demonstrated acute and chronic cytotoxicity, respectively. The minimal migration ability was observed in deuterated medium whereas the highest migration activity was observed in the medium with the deuterium content close to natural. The cells in deuterated growth medium demonstrated decrease in metabolic activity after three days in culture. In contrast, in deuterium-depleted medium there was an increase in ADSC metabolic activity.
To investigate the effects of the rhLIF recombinant human leukemia inhibitory factor on the structural and functional changes in the central nervous system in the cuprizone-induced experimental model of multiple sclerosis in mice and to evaluate the involvement of the brain macrophages, T-lymphocytes and antioxidant enzymes in the observed rhLIF effects. Methods. Young 129/Sv mice were fed with the cuprizone neurotoxin daily for three weeks. rhLIF was injected daily (50 μg/kg) after seven days of the cuprizone treatment. We used histology, flow cytometry, spectrophotometric and functional methods. Results. The cuprizone treatment resulted in an increase in the number of the brain and spinal cord neurons with destructive changes, the brain macrophages, CD3 +-cells and the content of malondialdehyde; it changes the behavior of animals, decreases the brain superoxide dismutase and glutathione peroxidase activities. The rhLIF injection partially or completely restored the cuprizone-changed parameters. The effect of rhLIF was preserved during two months after the experiment completion. Conclusions. The neuroprotective effect of rhLIF is connected with a decrease in the cuprizone-induced changes in the number of the brain T-cells and macrophages and the activity of antioxidant enzymes.
Introduction. The adult neural crest-derived stem cells (NCSCs) have significant perspectives for use in regenerative medicine. The most attractive sources for adult NCSC isolation are the hair follicles (HF) and skin dermis (SD) because of easy access and minimally invasive biopsy. The aim of this study was to compare the biological properties of HF- and SD-derived NCSCs after their large-scale expansion. Methods. The conventional explant method was used to obtain HF NCSCs. For the isolation of SD NCSCs, a new combined technique consisting of preplating and subsequent culturing in 3D blood plasma-derived fibrin hydrogel was applied. The studied cells were characterized by flow cytometry, ICC, qPCR, Bio-Plex multiplex assay, and directed multilineage differentiation assays. Results. We have obtained both adult SD and HF NCSCs from each skin sample (n=5). Adult SD and HF NCSCs were positive for key neural crest markers: SOX10, P75 (CD271), NESTIN, SOX2, and CD349. SD NCSCs showed a higher growth rate during the large-scale expansion compared to HF NCSCs (p<0.01). Final population of SD NCSCs also contained more clonogenic cells (p<0.01) and SOX10+, CD271+, CD105+, CD140a+, CD146+, CD349+ cells (p<0.01). Both HF and SD NCSCs had similar gene expression profiling and produced growth factors, but some quantitative differences were detected. Adult HF and SD NCSCs were able to undergo directed differentiation into neurons, Schwann cells, adipocytes, and osteoblasts. Conclusion. The HF and SD are suitable sources for large-scale manufacturing of adult NCSCs with similar biological properties. We demonstrated that the NCSC population from SD was homogenous and displayed significantly higher growth rate than HF NCSCs. Moreover, SD NCSC isolation is cheaper, easier, and minimally time-consuming method.
Cytokines and growth factors exhibit neurotropic, anti-inflammatory and immunomodulatory properties, and therefore can affect the functioning of the nervous system at demyelinating disorders.Purpose. To identify changes in T-lymphocytes, macrophages, oxidative stress and antioxidant defence factors, endocrine thymus function in the brain and behaviour of mice receiving neurotoxin cuprizone and recombinant human proteins: interleukin-10 (rhIL-10) and fibroblast growth factor (rhFGF-2).Materials and methods. 4-6-month-old 129/Sv mice received cuprizone with food every day for 3 weeks. From the 7th day of cuprizone diet, they received different doses of rhIL-10 and rhFGF-2. The content of СD3+ Т-cells, macrophages, malondialdehyde, activity of antioxidant enzymes in the brain and the level of thymulin in the blood were determined. Behavioural reactions were assessed in the “open field” test.Results. In the brain of mice receiving cuprizone and rhIL-10, there was a decrease in the number of СD3+ Т-cells and the activity of macrophages, which significantly increased under the influence of the toxin. After cytokine injection, the activity of superoxide dismutase, catalase and glutathione peroxidase increased significantly in the brain, as well as the level of thymulin in the blood. Motor, emotional and exploratory activity of mice, significantly suppressed by the cuprizone, was increased after the introduction of rhIL-10. The effect of rhIL-10 on the test parameters is more pronounced at a dose of 5 μg/kg than 50 μg/kg. After injection of rhFGF-2 in the mice with cuprizone diet, there is a decrease in the activity of brain macrophages and an increase of the level of thymulin in the blood depending on the dose of this factor; the motor activity of these animals increased regardless of the rhFGF-2 dose.Conclusion. The injections of rhIL-10 and rhFGF-2 provide dose-dependent positive effects on the pathogenetic factors of experimental demyelinating pathology, as well as the functional state of the nervous system. Whereas, the injections of rhIL-10 have more pronounced effects.
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