Monocyte Polarization and Kidney Tubule Inflammation

Human monocytes were cultured for 18 h in RPMI 1640 media supplemented with M-CSF, LPS, and IF-γ to induce M1 polarization [27 (link)]. M1-polarized cells were cultured alone or co-cultured with Lean- and MetS-MSC-derived EVs (50 μg of EV protein). Expression of inducible nitric oxide synthase (iNOS) and arginase-1 (1:200, Santa Cruz, CA) was evaluated using Western blotting [17 (link)]. In addition, pig proximal kidney tubular epithelial cells (LLC-PK1, ATCC, Manassas) were cultured in Medium-199 (Gibco BRL, USA) containing 3% FBS [28 (link)] alone or co-cultured with Lean- and MetS-MSC-derived EVs (5 μg of EV protein). Tubular epithelial cell inflammation was evaluated by immunofluorescent staining with antibodies against tumor necrosis factor (TNF)-α (Santa Cruz, 1:200) and monocyte chemoattractant protein (MCP)-1 (MyBioSource, San Diego, CA, http://www.mybiosource.com 1:7500).

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Meng Y., Eirin A., Zhu X.Y., O’Brien D.R., Lerman A., van Wijnen A.J, & Lerman L.O. (2018). The metabolic syndrome modifies the mRNA expression profile of extracellular vesicles derived from porcine mesenchymal stem cells. Diabetology & Metabolic Syndrome, 10, 58.

Publication 2018

arginase-1 LLC-PK1 Medium-199

Antibodies Arginase 1 Cells Epithelial cell Human Immunofluorescent Inducible nitric oxide synthase Inflammation Kidney M csf Monocyte chemoattractant protein 1 Monocytes Protein Tumor necrosis factor α

Corresponding Organization :

Other organizations : Mayo Clinic, Mayo Clinic in Florida, WinnMed, Mayo Clinic in Arizona

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Variable analysis

independent variables

Treatment with Lean-MSC-derived EVs
Treatment with MetS-MSC-derived EVs

dependent variables

Expression of inducible nitric oxide synthase (iNOS)
Expression of arginase-1
Tubular epithelial cell inflammation (evaluated by immunofluorescent staining with antibodies against tumor necrosis factor (TNF)-α and monocyte chemoattractant protein (MCP)-1)

control variables

Human monocytes cultured alone (without any EV treatment)
Pig proximal kidney tubular epithelial cells (LLC-PK1) cultured alone (without any EV treatment)

controls

Positive control: Human monocytes cultured with M-CSF, LPS, and IFN-γ to induce M1 polarization
Negative control: Human monocytes cultured alone, without any EV treatment
Negative control: Pig proximal kidney tubular epithelial cells (LLC-PK1) cultured alone, without any EV treatment

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