Effects of High Glucose Concentration on Pericyte-Like Differentiated Human Adipose-Derived Mesenchymal Stem Cells
Abstract
:1. Introduction
2. Results
2.1. Cell Proliferation and Viability
2.2. Cell Migration
2.3. ROS Level Measurements
2.4. High Glucose Effects on ASC mRNA Levels of Anti-Inflammatory Cytokines, Pro-Inflammatory Cytokines, and Angiogenic Factors
2.5. Three-Dimensional Cultures in Matrigel
3. Discussion
4. Materials and Methods
4.1. Materials
4.2. Cell Cultures
4.2.1. HRPC Cultures
4.2.2. HREC Cultures
4.2.3. ASC Cultures
4.2.4. ASCs Undergoing Pericyte-Like Differentiation
4.3. Cell Proliferation Assay
4.4. Cell Viability Assay
4.5. Wound-Healing Assay
4.6. ROS Measurements
4.7. Extraction of Total RNA and Real-Time Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
4.8. Three-Dimensional Cultures in Matrigel
4.9. Statistical Analysis
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
ASCs | human adipose-derived mesenchymal stem cells |
DR | diabetic retinopathy |
ROS | reactive oxygen species |
PmASCs | ASCs pre-cultured in the pericyte medium |
TGF-β1 | transforming growth factor-β1 |
IL-10 | interleukin-10 |
IL-1β | interleukin-1β |
TNF-α | tumor necrosis factor-α |
BRB | blood retinal barrier |
VEGF | vascular endothelial growth factor |
PM | pericyte medium |
α-SMA | smooth muscle actin α |
NG2 | neural/glial antigen 2 |
HRECs | human retinal endothelial cells |
MMP-9 | matrix metallopeptidase-9 |
HRPCs | pericytes |
NG | normal glucose |
HG | high glucose |
ECM | endothelial cell medium |
DCFDA | 2′,7′-dichlorodihydrofluorescein diacetate |
MTT | 3-[4,5-dimethylthiazol-2-y l]-2,5-diphenyl tetrasodium bromide |
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Gene | Sequence (5′-3′) | Amplicon (bp) | Accession Number |
---|---|---|---|
IL-10 | Fw: GACTTTAAGGGTTACCTGGGTTG | 112 | NM_000572.3 |
Rv: TCACATGCGCCTTGATGTCTG | |||
TGF-β1 | Fw: CGTCTGCTGAGGCTCAAGT | 74 | NM_000660.7 |
Rv: CGCCAGGAATTGTTGCTGTA | |||
TNF-α | Fw: AGCCCATGTTGTAGCAAACC | 134 | NM_000594.4 |
Rv: TGAGGTACAGGCCCTCTGAT | |||
IL-1β | Fw: AGCTACGAATCTCCGACCAC | 186 | NM_000576.3 |
Rv: CGTTATCCCATGTGTCGAAGAA | |||
VEGF | Fw: ATCTTCAAGCCATCCTGTGTGC | 121 | NM_001025366.3 |
Rv: GAGGTTTGATCCGCATAATCTG | |||
Angiopoietin-2 | Fw: CTCGAATACGATGACTCGGTG | 76 | NM_001386337.1 |
Rv: TCATTAGCCACTGAGTGTTGTTT | |||
MMP9 | Fw: CACTGTCCACCCCTCAGAGC | 264 | NM_004994.3 |
Rv: GCCAACTTGTCGGCGATAAGG | |||
18S rRNA | Fw: TAAGTCCCTGCCCTTTGTACACA | 69 | NR_146119 |
Rv: GATCCGAGGGCCTCACTAAAC |
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Mannino, G.; Longo, A.; Gennuso, F.; Anfuso, C.D.; Lupo, G.; Giurdanella, G.; Giuffrida, R.; Lo Furno, D. Effects of High Glucose Concentration on Pericyte-Like Differentiated Human Adipose-Derived Mesenchymal Stem Cells. Int. J. Mol. Sci. 2021, 22, 4604. https://doi.org/10.3390/ijms22094604
Mannino G, Longo A, Gennuso F, Anfuso CD, Lupo G, Giurdanella G, Giuffrida R, Lo Furno D. Effects of High Glucose Concentration on Pericyte-Like Differentiated Human Adipose-Derived Mesenchymal Stem Cells. International Journal of Molecular Sciences. 2021; 22(9):4604. https://doi.org/10.3390/ijms22094604
Chicago/Turabian StyleMannino, Giuliana, Anna Longo, Florinda Gennuso, Carmelina Daniela Anfuso, Gabriella Lupo, Giovanni Giurdanella, Rosario Giuffrida, and Debora Lo Furno. 2021. "Effects of High Glucose Concentration on Pericyte-Like Differentiated Human Adipose-Derived Mesenchymal Stem Cells" International Journal of Molecular Sciences 22, no. 9: 4604. https://doi.org/10.3390/ijms22094604