Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells

Background: Pharmacological factors used to induce insulin resistance (IR) in in vitro models may not mimic the full in vivo features of type 2 diabetes mellitus (T2DM). This study aimed to examine the ability of diabetic serum (DS) to induce IR and investigate whether adipose-derived mesenchymal st...

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Autores principales: Avinash Sanap, Ramesh Bhonde, Kalpana Joshi
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Publicado: KeAi Communications Co., Ltd. 2021
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spelling oai:doaj.org-article:a006f290cd43478e947fc74928b186ad2021-12-02T18:21:35ZMesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells2095-882X10.1016/j.cdtm.2020.09.001https://doaj.org/article/a006f290cd43478e947fc74928b186ad2021-03-01T00:00:00Zhttp://www.sciencedirect.com/science/article/pii/S2095882X20300797https://doaj.org/toc/2095-882XBackground: Pharmacological factors used to induce insulin resistance (IR) in in vitro models may not mimic the full in vivo features of type 2 diabetes mellitus (T2DM). This study aimed to examine the ability of diabetic serum (DS) to induce IR and investigate whether adipose-derived mesenchymal stem cell conditioned medium (ADMSC-CM) reverses DS-induced IR. Methods: DS was obtained from newly diagnosed T2DM patients. IR was induced in differentiated 3T3-L1 cells by employing dexamethasone, tumor necrosis factor alpha (TNF-α), palmitate and DS. Glucose uptake (2-[N-[7-nitrobenz-2-oxa-1,3-diazol-4-yl] amino]-2-deoxyglucose(2-NBDG) uptake assay), intracellular levels of reactive oxygen species (ROS), and superoxide radicals (O2−) (fluorescence microscopy and fluorometry) were analyzed in control and experimental samples. mRNA expression of key genes involved in glucose transport and inflammation were analyzed by using reverse transcription polymerase chain reaction (RT-PCR). Pro-inflammatory cytokines and phospho-insulin receptor substrate (IRS) (Ser-307) protein expression were analyzed by fluorescence activated cell sorter analysis. Statistical significance was determined by using one-way ANOVA followed by Tukey's multiple comparison tests. Results: ADMSC-CM significantly increased the DS-mediated decrease in 2-NBDG uptake (11.01 ± 0.50 vs. 7.20 ± 0.30, P < 0.01) and reduced DS-driven ROS (fluorescence count, 6.35 ± 0.46 vs. 9.80 ± 0.10, P < 0.01) and O2− (fluorescence count, 3.00 ± 0.10 vs. 4.60 ± 0.09, P < 0.01) production. Further, the ADMSC-CM restored DS-induced down regulation GLUT4 (1.52-fold, P < 0.05) as well as the up-regulation of PPARγ (0.35-fold, P < 0.01), and IKKβ (0.37-fold, P < 0.01) mRNA, and phospho-IRS (Ser-307) protein expression compared to the baseline (median fluorescence intensity, 88,192 ± 2720 vs. 65,450 ± 3111, P < 0.01). DS induced IR, similar to the traditionally used pharmacological factors, namely dexamethasone, TNF-α, and palmitate, which can be attributed to the significantly higher pro-inflammatory cytokines levels (TNF-α (2.28 ± 0.03 pg/mL vs. 2.38 ± 0.03 pg/mL, P < 0.01), interleukin 6 (IL)-6 (1.94 ± 0.02 pg/mL vs. 2.17 ± 0.04 pg/mL, P < 0.01), IL-17 (2.16 ± 0.02 pg/mL vs. 2.22 ± 0.002 pg/mL, P < 0.05), and interferon gamma (IFN-γ) (2.07 ± 0.02 pg/mL vs. 2.15 ± 0.04 pg/mL, P < 0.05)) in DS. Conclusions: DS can be explored as a novel inducer of IR in in vitro studies with further standardization, substituting the conventionally used pharmacological factors. Our findings also affirm the validity of ADMSC-CM as a prospective insulin sensitizer for T2DM therapy.Avinash SanapRamesh BhondeKalpana JoshiKeAi Communications Co., Ltd.articleInsulin resistanceInflammationMesenchymal stem cellsOxidative stressType 2 diabetes mellitusMedicine (General)R5-920ENChronic Diseases and Translational Medicine, Vol 7, Iss 1, Pp 47-56 (2021)
institution DOAJ
collection DOAJ
language EN
topic Insulin resistance
Inflammation
Mesenchymal stem cells
Oxidative stress
Type 2 diabetes mellitus
Medicine (General)
R5-920
spellingShingle Insulin resistance
Inflammation
Mesenchymal stem cells
Oxidative stress
Type 2 diabetes mellitus
Medicine (General)
R5-920
Avinash Sanap
Ramesh Bhonde
Kalpana Joshi
Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
description Background: Pharmacological factors used to induce insulin resistance (IR) in in vitro models may not mimic the full in vivo features of type 2 diabetes mellitus (T2DM). This study aimed to examine the ability of diabetic serum (DS) to induce IR and investigate whether adipose-derived mesenchymal stem cell conditioned medium (ADMSC-CM) reverses DS-induced IR. Methods: DS was obtained from newly diagnosed T2DM patients. IR was induced in differentiated 3T3-L1 cells by employing dexamethasone, tumor necrosis factor alpha (TNF-α), palmitate and DS. Glucose uptake (2-[N-[7-nitrobenz-2-oxa-1,3-diazol-4-yl] amino]-2-deoxyglucose(2-NBDG) uptake assay), intracellular levels of reactive oxygen species (ROS), and superoxide radicals (O2−) (fluorescence microscopy and fluorometry) were analyzed in control and experimental samples. mRNA expression of key genes involved in glucose transport and inflammation were analyzed by using reverse transcription polymerase chain reaction (RT-PCR). Pro-inflammatory cytokines and phospho-insulin receptor substrate (IRS) (Ser-307) protein expression were analyzed by fluorescence activated cell sorter analysis. Statistical significance was determined by using one-way ANOVA followed by Tukey's multiple comparison tests. Results: ADMSC-CM significantly increased the DS-mediated decrease in 2-NBDG uptake (11.01 ± 0.50 vs. 7.20 ± 0.30, P < 0.01) and reduced DS-driven ROS (fluorescence count, 6.35 ± 0.46 vs. 9.80 ± 0.10, P < 0.01) and O2− (fluorescence count, 3.00 ± 0.10 vs. 4.60 ± 0.09, P < 0.01) production. Further, the ADMSC-CM restored DS-induced down regulation GLUT4 (1.52-fold, P < 0.05) as well as the up-regulation of PPARγ (0.35-fold, P < 0.01), and IKKβ (0.37-fold, P < 0.01) mRNA, and phospho-IRS (Ser-307) protein expression compared to the baseline (median fluorescence intensity, 88,192 ± 2720 vs. 65,450 ± 3111, P < 0.01). DS induced IR, similar to the traditionally used pharmacological factors, namely dexamethasone, TNF-α, and palmitate, which can be attributed to the significantly higher pro-inflammatory cytokines levels (TNF-α (2.28 ± 0.03 pg/mL vs. 2.38 ± 0.03 pg/mL, P < 0.01), interleukin 6 (IL)-6 (1.94 ± 0.02 pg/mL vs. 2.17 ± 0.04 pg/mL, P < 0.01), IL-17 (2.16 ± 0.02 pg/mL vs. 2.22 ± 0.002 pg/mL, P < 0.05), and interferon gamma (IFN-γ) (2.07 ± 0.02 pg/mL vs. 2.15 ± 0.04 pg/mL, P < 0.05)) in DS. Conclusions: DS can be explored as a novel inducer of IR in in vitro studies with further standardization, substituting the conventionally used pharmacological factors. Our findings also affirm the validity of ADMSC-CM as a prospective insulin sensitizer for T2DM therapy.
format article
author Avinash Sanap
Ramesh Bhonde
Kalpana Joshi
author_facet Avinash Sanap
Ramesh Bhonde
Kalpana Joshi
author_sort Avinash Sanap
title Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
title_short Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
title_full Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
title_fullStr Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
title_full_unstemmed Mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3T3-L1 cells
title_sort mesenchymal stem cell conditioned medium ameliorates diabetic serum-induced insulin resistance in 3t3-l1 cells
publisher KeAi Communications Co., Ltd.
publishDate 2021
url https://doaj.org/article/a006f290cd43478e947fc74928b186ad
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AT rameshbhonde mesenchymalstemcellconditionedmediumamelioratesdiabeticseruminducedinsulinresistancein3t3l1cells
AT kalpanajoshi mesenchymalstemcellconditionedmediumamelioratesdiabeticseruminducedinsulinresistancein3t3l1cells
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