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MiR-185/AKT and miR-29a/collagen 1a pathways are activated in IPF BAL cells

Eliza Tsitoura1,2, Athol U Wells3, Kostantinos Karagiannis1

  • 1Laboratory of Molecular and Cellular Pneumonology, Medical School, University of Crete, Heraklion, Crete, Greece.

Oncotarget
|November 5, 2016
PubMed

Insights

Idiopathic pulmonary fibrosis (IPF) shows altered microRNA levels in lung cells. Specifically, miR-29a and miR-185 are downregulated, impacting collagen and AKT pathways, crucial for IPF progression.

Area of Science:

  • Pulmonary Medicine
  • Molecular Biology
  • Biochemistry

Background:

  • Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with limited treatment options.
  • MicroRNAs (miRNAs) play critical roles in cellular processes, but their specific roles in IPF pathogenesis are not fully understood.
  • Bronchoalveolar lavage (BAL) cells offer a direct source for investigating molecular changes in IPF.

Purpose of the Study:

  • To investigate the expression profiles of fibrosis-related and IPF progression-associated microRNAs in BAL cells from IPF patients.
  • To explore the functional relationship between identified miRNAs and key fibrotic markers, such as collagen.
  • To elucidate the role of miR-185 in macrophage response to profibrotic stimuli and its connection to the AKT signaling pathway.

Main Methods:

  • Quantitative real-time PCR was used to measure the expression of selected miRNAs (miR-29a, miR-29b, miR-29c, let-7d, miR-21, miR-185, miR-210, miR-302c-3p, miR-376c, miR-423-5p) in BAL cells.
  • COL1A1 mRNA levels were quantified in BAL cells.
  • Immunofluorescence staining was performed on BAL cells to detect Collagen 1 alpha expression.
  • THP-1 macrophage cell line was used to study miR-185 regulation by TGF-beta and pro-inflammatory stimuli.
  • Western blotting was employed to assess total AKT and phosphorylated AKT (AKTser473) levels in IPF BAL cells and THP-1 macrophages.
  • miR-185 inhibition experiments were conducted in THP-1 macrophages.

Main Results:

  • miR-29a and miR-185 were significantly downregulated in BAL cells of IPF patients.
  • miR-302c-3p and miR-376c were not expressed in BAL cells.
  • Downregulation of miR-29a correlated inversely with increased COL1A1 mRNA levels, with Collagen 1 alpha predominantly found in alveolar macrophages.
  • miR-185 was downregulated in THP-1 macrophages stimulated with TGF-beta but upregulated by pro-inflammatory stimuli.
  • While mRNA levels of known miR-185 targets (AKT1, DNMT1, HMGA2) did not correlate, increased total AKT and AKTser473 phosphorylation were observed in IPF BAL cells.
  • Inhibition of miR-185 in THP-1 macrophages led to increased AKTser473 phosphorylation.

Conclusions:

  • BAL cell microRNA signatures are altered in IPF, with specific miRNAs like miR-29a and miR-185 showing significant changes.
  • The miR-29a/collagen axis and the miR-185/AKT axis represent important molecular pathways dysregulated in IPF BAL cells.
  • These findings highlight the potential of targeting specific miRNAs or their downstream pathways for therapeutic interventions in IPF.
Keywords:
AKTIPFImmune responseImmunityImmunology and Microbiology Sectionbronchoalveolar lavage fluidcollagenmicroRNA

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