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Cryo-Temperature Pretreatment Increases the Pro-Angiogenic Capacity of Three-Dimensional Mesenchymal Stem Cells via the PI3K-AKT Pathway.

ABSTRACT: To increase the potential and effectiveness of three-dimensional (3D) mesenchymal stem cells (MSCs) for clinical applications, this study explored the effects of short cryo-temperature pretreatment on MSC function. Adipose-derived MSCs (A-MSCs) were cultured via the ordinary monolayer method and 3D hanging drop spheroid method. When the cells adhered to the wall or formed a spheroid, they were subjected to hypothermic stress at 4°C for 1 h and then divided into three recovery periods at 37°C, specifically 0, 12, and 24 h. The control group was not subjected to any treatment throughout the study. Monolayer and 3D spheroid A-MSCs were analyzed via RNA sequencing after hypothermic stress at 4°C for 1 h. Subsequently, each group of cells was collected and subjected to phenotype identification via flow cytometry, and mRNA expression was detected via reverse transcription-quantitative polymerase chain reaction analysis. Western blot analysis was performed to analyze the PI3K-AKT signaling pathway in A-MSCs. The effects of A-MSCs on angiogenesis in vivo were examined using a chick chorioallantoic membrane assay. Transwell assays were performed to determine whether the culture supernatant from each group could induce the chemotaxis of human umbilical vein endothelial cells (HUVECs). Three-dimensional spheroid culture did not change the phenotype of A-MSCs. The expression of fibroblast growth factors, hepatocyte growth factors, and other angiogenesis-related factors in A-MSCs was upregulated. A-MSCs subjected to hypothermic stress promoted angiogenesis under both monolayer and 3D spheroid cultures. Moreover, the chemotaxis of HUVECs to the 3D spheroid culture supernatant increased substantially. Short cryo-temperature pretreatment could stimulate 3D spheroid A-MSCs and activate the PI3K-AKT pathway. This approach has the advantages of promoting angiogenesis and maintaining cell viability.

SUBMITTER: Zhu H

PROVIDER: S-EPMC9218451 | biostudies-literature | 2022 Jan-Dec

REPOSITORIES: biostudies-literature

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Cryo-Temperature Pretreatment Increases the Pro-Angiogenic Capacity of Three-Dimensional Mesenchymal Stem Cells <i>via</i> the PI3K-AKT Pathway.

Zhu Huasu H Zhuang Yong Y Li Dong D Dong Na N Ma Huixian H Liu Linghong L Shi Qing Q Ju Xiuli X

Cell transplantation 20220101

To increase the potential and effectiveness of three-dimensional (3D) mesenchymal stem cells (MSCs) for clinical applications, this study explored the effects of short cryo-temperature pretreatment on MSC function. Adipose-derived MSCs (A-MSCs) were cultured via the ordinary monolayer method and 3D hanging drop spheroid method. When the cells adhered to the wall or formed a spheroid, they were subjected to hypothermic stress at 4°C for 1 h and then divided into three recovery periods at 37°C, sp ...[more]

PMID: 35727010

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Project description:BackgroundAngiogenesis strongly reflects poor breast cancer outcome and an important contributor to breast cancer (BC) metastasis; therefore, anti-angiogenic intervention is a potential tool for cancer treatment. However, currently used antibodies against vascular endothelial growth factor A (VEGFA) or inhibitors that target the VEGFA receptor are not effective due to weak penetration and low efficiency. Herein, we assessed the anti-BC angiogenic role of muscone, a natural bioactive musk constituent, and explored possible anti-cancer mechanisms of this compound.MethodsCCK-8, EdU, scratch and Transwell assessments were employed to detect the muscone-mediated regulation of breast cancer (BC) and human umbilical vein endothelial cells (HUVECs) proliferation and migration. Tube formation, matrigel plug assay and zebrafish assay were employed for assessment of regulation of tumor angiogenesis by muscone. In vivo xenograft mouse model was constructed to compare microvessel density (MVD), vascular leakage, vascular maturation and function in muscone-treated or untreated mice. RNA sequencing was performed for gene screening, and Western blot verified the effect of the VEGFA-VEGFR2 pathway on BC angiogenic inhibition by muscone.ResultsBased on our findings, muscone suppressed BC progression via tumor angiogenic inhibition in cellular and animal models. Functionally, muscone inhibited BC cell proliferation and migration as well as tumor cell-conditioned medium-based endothelial cell proliferation and migration. Muscone exhibited a strong suppressive influence on tumor vasculature in cellular and animal models. It abrogated tumor cell growth in a xenograft BC mouse model and minimized tumor microvessel density and hypoxia, and increased vascular wall cell coverage and perfusion. Regarding the mechanism of action, we found that muscone suppressed phosphorylation of members of the VEGF/PI3K/Akt/MAPK axis, and it worked synergistically with a VEGFR2 inhibitor, an Akt inhibitor, and a MAPK inhibitor to further inhibit tube formation.ConclusionOverall, our results demonstrate that muscone may proficiently suppress tumor angiogenesis via modulation of the VEGF/PI3K/Akt/MAPK axis, facilitating its candidacy as a natural small molecule drug for BC treatment.

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Dataset Information

Cryo-Temperature Pretreatment Increases the Pro-Angiogenic Capacity of Three-Dimensional Mesenchymal Stem Cells via the PI3K-AKT Pathway.

Publications

Cryo-Temperature Pretreatment Increases the Pro-Angiogenic Capacity of Three-Dimensional Mesenchymal Stem Cells <i>via</i> the PI3K-AKT Pathway.

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