Background Metformin is originally introduced as a biguanide antidiabetic medication, has an anti-inflammatory effect via activating AMP-activated protein kinase (AMPK). Human adipose-derived stem cells (Ad-MSCs) are stromal cells derived from adipose tissue and known to have immunoregulatory activity. The study was undertaken to examine whether metformin-treated Ad-MSCs show more potent therapeutic effect in animal model of lupus and to clarify the underlying mechanism of potent immunoregulatory impact of metformin-treated Ad-MSCs.
Methods To examine the effects of metformin, Ad-MSCs were incubated for 72 hour in the presence of metformin. Cellular phenotype of Ad-MSCs was analyzed by flow cytometry. Indoleamine 2,3-dioxygenase, IL-10 and TGF-1 expression was analyzed by real-time PCR and ELISA. AMPK-mTOR pathway was analyzed by Western blotting in Ad-MSCs with or without metformin. MRL/lpr mice weekly injected 1×10^6 metformin-treated Ad-MSCs in 0.1 ml PBS to lateral tail vein for 7 weeks. All mice were sacrificed at the age of 16 weeks. Urinary albumin-to-creatinine ratios were then calculated. The amount of anti-dsDNA IgG antibody in mice sera was measured by ELISA. PAS stained kidney sections were used for assessment of histology. Deposition of IgG and C3 was detected by confocal microscope. Population of cellular subset in spleen, kidney and blood was analyzed by Flow cytometry.
Results In vitro, metformin-treated Ad-MSCs increased mRNA level of IDO, IL-10 and TGF-1 compared with untreated Ad-MSCs. Also, the concentrations of IDO, IL-10 and TGF- increased in culture supernatants. Metformin upregulated expression of p-AMPK and inhibited the expression of p-STAT3, p-mTOR, and p-Raptor in Ad-MSCs. Intravenously injected metformin-treated Ad-MSCs significantly reduced the splenomegaly and lymphadenopathy compared with untreated Ad-MSCs in MRL/lpr mice. In addition, Metformin-treated Ad-MSCs decreased anti-dsDNA antibodies in serum and proteinuria compared with untreated Ad-MSCs. Metformin-treated Ad-MSCs alleviated lupus nephritis, as judged by changes in the histopathology scores and immune complex deposition. Metformin-treated Ad-MSCs reduced CD90.2+T cells, CD90.2+CD4 CD8- (double negative) T cells, whereas CD4 +CD25+Foxp3+Treg cells were increased in splenocytes, kidney tissue and blood cells of MRL/lpr mice.
Conclusions Metformin can optimize the immunomodulatory potential of Ad-MSCs, suggesting a promising strategy of MSC use in lupus treatment.
Funding Source(s): None
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