Maintenance of mesenchymal stem cells culture due to the cells with reduced attachment rate

Shuvalova, N. S.; Maslova, O.O.; Sukhorada, O. M.; Deryabina, О. G.; Кордюм, В. А.

doi:10.7124/bc.000809

Cited by 5 publications

(4 citation statements)

References 15 publications

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“…In the flacks containing the conditioned media from MSC cultures, the attached cells, able to form clones, can be viewed already in 2-5 days. The morphological and surface markers of the cells obtained by this method resembled those of the original culture, confirming that "self-replated" cells descend from the cells, the adhesion of which to a substrate reduced before or during the division [11].…”

Section: Growth Analysis Of Obtained "Self-replating" Culturessupporting

confidence: 53%

“…In our previous work we have suggested the method of optimization of human WJ-MSC cultivation. It allowed preservation and multiplication of the cells, that spontaneously detached during cultivation (presumably, in the phase prior to mitosis), thus obtaining the populations of MSC, that avoided the damaging influence of a passing procedure on the surface molecules [11]. Besides, we have shown a positive effect of the nitrogen-and argon-based cultivation gas mixtures containing 3 % O 2 [12].…”

Section: Molecular and Cell Biotechnologiesmentioning

confidence: 83%

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Proliferation of Wharton jelly mesenchymal stem cells, derived by preserving the cells with reduced attachment rate, under various gas conditions

Shuvalova

Кордюм

2015

Biopolym. Cell

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To estimate the proliferation rates in cultures of the human Wharton jelly mesenchymal stem cells (WJ-MSCs), obtained by the method of preservation of spontaneously detached cells, in various gas mixtures, containing physiological oxygen concentrations. Methods. Starting from the first passage, WJ-MSC were cultured for 4 subsequent passages,7 days between replating ("main" line"). At 0, 1 and 2 passage, day 3 of cultivation, conditioned media was collected and transferred to another flack with complete growth media. The attached cells from conditioned media were cultivated until the clones size and confluence (70-80 %) became sufficient for replating, and after that were subsequently passed with trypsin-EDTA solution ("side" lines). Besides the cultivation in standard condition of CO 2-incubator, the cultivation was conducted in the nitrogen-based gas mixture (3 % oxygen, 4 % carbon dioxide, 93 % nitrogen) and argonbased gas mixture (3 % oxygen, 4 % carbon dioxide, 93 % argon). At each passage, the number of cells was counted. Results. The proliferation level in "side" lines, obtained from 0 and 1 passages, had a lot of similarities with that of "main" line. We observed the trend of multiplication rate reduction in "side" lines during in vitro maintenance, similar to that in "main" line. For cultures, obtained at the passage 2, the level of proliferation was significantly lower. The cultivation in both gas mixtures with 3% O 2 concentration had beneficial effect on the level of cell multiplication: the number of cells was significantly higher. The effect of argon-based mixture was more pronounced. Conclusion. The physiologic oxygen tension allows optimizing the cultivation of WJ-MSC, obtained by the suggested method of preserving the cells with reduced attachment ability.

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Section: Growth Analysis Of Obtained "Self-replating" Culturessupporting

confidence: 53%

Section: Molecular and Cell Biotechnologiesmentioning

confidence: 83%

Proliferation of Wharton jelly mesenchymal stem cells, derived by preserving the cells with reduced attachment rate, under various gas conditions

Shuvalova

Кордюм

2015

Biopolym. Cell

Self Cite

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“…The first attached cells were visible on the 7–10th day. After 14 days the clones reached the size and confluence (70–80%) sufficient for passing, which was performed by standard method with the use of trypsin-EDTA mixture (Shuvalova et al., 2013). The surface marker proteins CD34, CD45, CD90, CD73, CD105 expression was determined at the second passage by flow cytometry (BD FACS Aria) with FITC- and PE- conjugated antibodies (UsBiological, USA) according to minimal criteria for defining multipotent mesenchymal stromal cells (Dominici et al., 2006).…”

Section: Methodsmentioning

confidence: 99%

Intravenously Injected Mesenchymal Stem Cells Penetrate the Brain and Treat Inflammation-Induced Brain Damage and Memory Impairment in Mice

et al. 2019

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Neuroinflammation is regarded as one of the pathogenic factors of Alzheimer disease (AD). Previously, we showed that mice regularly injected with bacterial lipopolysaccharide (LPS) possessed the AD-like symptoms like episodic memory decline, elevated amounts of amyloid beta (Aβ) peptide (1–42), and decreased levels of nicotinic acetylcholine receptors (nAChRs) in the brain. The use of mesenchymal stem cells (MSCs), which can differentiate into multiple cell types, including neurons, is an attractive idea of regenerative medicine, in particular, for neurodegenerative disorders like AD. In the present study, we aimed to investigate whether pathogenic effect of LPS on the brain and behavior of mice can be prevented or treated by injection of MSCs or MSC-produced soluble factors. Fluorescently-labeled MSCs, injected intravenously, were found in the brain blood vessels of LPS-treated mice. Mice co-injected with LPS and MSCs did not demonstrate episodic memory impairment, Aβ (1–42) accumulation, and nAChR decrease in the brain and brain mitochondria. Their mitochondria released less cytochrome c under the effect of Ca 2+ compared to mitochondria of LPS-only-treated mice. Moreover, MSCs could reverse the pathogenic symptoms developed 3 weeks after LPS injection. Cultured MSCs produced IL-6 in response to LPS and MSCs effect in vivo was accompanied by additional stimulation of both micro- and macroglia. Xenogeneic (human) MSCs were almost as efficient as allogeneic (mouse) ones and regular injections of human MSC-conditioned medium also produced positive effect. These data allow suggesting MSCs as a potential therapeutic tool to cure neuroinflammation-related cognitive pathology.

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“…Most of reagents were purchased from Sigma-Aldrich (Saint Louis, USA). Antibodies against α7(179-190) or α7(1-208) nAChR fragments were generated and biotinylated by us previously as described [20,21]. The kits for IL-1β and IL-6 determination (Invitrogen, Waltham, USA) and Neutravidin-peroxidase conjugate (Molecular Probes) were purchased from Thermo Fisher Scientific.…”

Section: Methodsmentioning

confidence: 99%

Extracellular vesicles derived from mesenchymal stem cells ameliorate cognitive impairment caused by neuroinflammation in young but not aged mice

Lykhmus,

Kalashnyk,

Skok

et al. 2024

Explor Neurosci

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Aim: The aim of this work was to study the effects of extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) on inflammation-impaired cognitive functions and the brain of mice. Methods: Young mice (~3-month-old) and aged mice (~18-month-old) were injected with bacterial lipopolysaccharide (LPS) and obtained intravenously donor 106 human umbilical cord MSCs, EVs isolated from a similar amount of MSCs or conditioned medium (CM) of MSCs. Subsequently, the mice were examined in behavioral tests and the mouse brains were analyzed for the levels of pro-inflammatory cytokines, α7 nicotinic acetylcholine receptors (α7 nAChRs) and amyloid beta 1-42 (Aβ1-42). Results: EVs prevented LPS-induced memory impairment in mice, whereas CM provided a weaker and temporal effect. Both EVs and MSCs injected once after regular injections of LPS stably improved memory of young mice. In contrast, both cells and EVs provided only transient effect in aged mice injected with LPS. The brains of aged LPS-treated mice contained elevated amounts of IL-1β and IL-6; both MSCs and EVs decreased them significantly. The brains of non-treated aged mice contained decreased levels of α7 nAChRs and increased levels of Aβ1-42 and α7-bound Aβ1-42 compared to the brains of young mice. LPS treatment decreased α7 nAChRs in both young and aged mice, while both MSCs and EVs restored them up to the control level. In young mice, LPS treatment increased the level of Aβ1-42 and α7-bound Aβ1-42, whereas MSCs and EVs decreased it. In contrast, neither LPS nor MSCs/EVs influenced the elevated level of Aβ1-42 but increased α7-bound Aβ1-42 in the brains of aged mice. Conclusions: Regenerative potential of MSCs and MSC-derived EVs is sufficient to support cognitive functions of LPS-treated young mice but is quite poor for aged animals, possibly, due to decreased levels of α7 nAChRs and accumulated Aβ1-42 in their brains.

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Maintenance of mesenchymal stem cells culture due to the cells with reduced attachment rate

Cited by 5 publications

References 15 publications

Proliferation of Wharton jelly mesenchymal stem cells, derived by preserving the cells with reduced attachment rate, under various gas conditions

Proliferation of Wharton jelly mesenchymal stem cells, derived by preserving the cells with reduced attachment rate, under various gas conditions

Intravenously Injected Mesenchymal Stem Cells Penetrate the Brain and Treat Inflammation-Induced Brain Damage and Memory Impairment in Mice

Extracellular vesicles derived from mesenchymal stem cells ameliorate cognitive impairment caused by neuroinflammation in young but not aged mice

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