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Cytotechnology

, Volume 68, Issue 4, pp 1337–1348 | Cite as

PPARγ ameliorated LPS induced inflammation of HEK cell line expressing both human Toll-like receptor 4 (TLR4) and MD2

  • Reyhaneh Darehgazani
  • Maryam Peymani
  • Motahare-Sadat Hashemi
  • Mir Davood Omrani
  • Abolfazl Movafagh
  • Kamran Ghaedi
  • Mohammad Hossein Nasr-Esfahani
Original Article

Abstract

TLR4 is transmembrane pattern-recognition receptor that initiates signals in response to diverse pathogen-associated molecular patterns especially LPS. Recently, there have been an increasing number of studies about the role of TLRs in the pathogenesis of several disorders as well as the therapeutic potential of TLR intervention in such diseases. Peroxisome proliferator-activated receptor-gamma (PPARγ) is a ligand-activated transcription factor with numerous biological effects. PPARγ has been shown to exert a potential anti-inflammatory effect through suppression of TLR4-mediated inflammation. Therefore, PPARγ agonists may have a potential to combat inflammatory conditions in pathologic states. The current study aims to show the decrease of inflammation by overexpression of PPARγ in a cell reporter model. To reach this goal, recombinant pBudCE4.1 (+) containing encoding sequences of human TLR4 and MD2 was constructed and used to transfect HEK cells. Subsequently, inflammation was induced by LPS treatment as control group. In the treatment group, overexpression of PPARγ prior to inflammation was performed and the expression of inflammatory markers was assessed in this condition. The expression of inflammatory markers (TNFα and iNOS) was defined by quantitative real time PCR and the amount of phosphorylated NF-κB was measured by western blot. Data indicated expression of TNFα and iNOS increased in LPS induced inflammation of stably transformed HEK cells with MD2 and TLR4. In this cell reporter model overexpression of PPARγ dramatically prevented LPS-induced inflammation through the blocking of TLR4/NF-κB signaling. PPARγ was shown to negatively regulate TLR4 activity and therefore exerts its anti-inflammatory action against LPS induced inflammation.

Keywords

HEK Inflammation MD2 PPARγ TLR4 

Abbreviations

BSA

Bovine serum albumin

CDS

Coding sequence

DAPI

4,6-Diamidino-2-phenylindole

DMEM

Dulbecco’s modified Eagle’s medium

EF-1α

Elongation factor 1α-subunit

FBS

Fetal bovine serum

GAPDH

Glyceraldehyde 3-phosphate dehydrogenase

HEK

Human embryonic kidney

IL

Interleukin

LPS

Lipopolysaccharide

LRR

Leucine-rich repeat

MD2

Myeloid differentiation protein 2

MTS/PMS

3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium)/phenazine methosulfate

OPC

Oligodendrocyte precursor cell

PAMP

Pathogen-associated molecular patterns

PBS

Phosphate buffered saline

PRR

Pattern-recognition receptor

PVDF

Polyvinylidenedifluoride

TLR

Toll-like receptor

Notes

Conflict of interest

None of the authors has any conflicts of interest to disclose and all authors support submission to this journal.

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Copyright information

© Springer Science+Business Media Dordrecht 2015

Authors and Affiliations

  • Reyhaneh Darehgazani
    • 1
    • 2
  • Maryam Peymani
    • 2
    • 3
  • Motahare-Sadat Hashemi
    • 2
  • Mir Davood Omrani
    • 1
  • Abolfazl Movafagh
    • 1
  • Kamran Ghaedi
    • 2
    • 4
  • Mohammad Hossein Nasr-Esfahani
    • 2
  1. 1.Department of Medical Genetics, Faculty of MedicineShahid Beheshti University of Medical SciencesTehranIran
  2. 2.Department of Cellular Biotechnology, Cell Science Research CenterRoyan Institute for BiotechnologyIsfahanIran
  3. 3.Department of Biology, Faculty of Basic Sciences, Shahrekord BranchIslamic Azad UniversitySahrekordIran
  4. 4.Biology Department, School of SciencesUniversity of IsfahanIsfahanIran

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