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Overexpression of Bcl-2 by activated human hepatic stellate cells: resistance to apoptosis as a mechanism of progressive hepatic fibrogenesis in humans
  1. E Novo1,
  2. F Marra2,
  3. E Zamara1,
  4. L Valfrè di Bonzo1,
  5. L Monitillo1,
  6. S Cannito1,
  7. I Petrai2,
  8. A Mazzocca2,
  9. A Bonacchi2,
  10. R S M De Franco2,
  11. S Colombatto1,
  12. R Autelli1,
  13. M Pinzani2,
  14. M Parola1
  1. 1Dip Medicina e Oncologia Sperimentale, University of Torino, Italy
  2. 2Dip Medicina Interna-Centro di Ricerca, Trasferimento e Alta Formazione “DENOTHE”, University of Florence, Italy
  1. Correspondence to:
    Professor M Parola
    Università degli Studi di Torino, Dip Medicina e Oncologia Sperimentale, C so Raffaello 30, 10125 Torino, Italy; maurizio.parola{at}unito.it

Abstract

Background and aims: Myofibroblast-like cells, originating from activation of hepatic stellate cells (HSC/MFs), play a key role in liver fibrosis, a potentially reversible process that may rely on induction of HSC/MFs apoptosis. While this possibility has been shown in cultured rat HSC, very limited data are currently available for human HSC/MFs.

Methods: Cultured human HSC/MFs were exposed to several proapoptotic stimuli, including those known to induce apoptosis in rat HSC/MFs, and induction of cell death and related mechanisms were investigated using morphology, molecular biology, and biochemical techniques.

Results: In this study we report that fully activated human HSC/MFs did not undergo spontaneous apoptosis and survived to prolonged serum deprivation, Fas activation, or exposure to nerve growth factor, tumour necrosis factor α (TNF-α), oxidative stress mediators, doxorubicin, and etoposide. Induction of caspase dependent, mitochondria driven apoptosis in HSC/MFs was observed only when protein synthesis or transcription were inhibited. Importantly, the process of HSC activation was accompanied by changes in expression of a set of genes involved in apoptosis control. In particular, activated human HSC/MFs in culture overexpressed Bcl-2. The role of Bcl-2 was crucial as Bcl-2 silenced cells became susceptible to TNF-α induced apoptosis. Finally, Bcl-2 was markedly expressed in HSC/MFs present in liver tissue obtained from patients with hepatitis C virus related cirrhosis.

Conclusions: Human activated HSC/MFs are resistant to most proapoptotic stimuli due to Bcl-2 overexpression and this feature may play a key role in the progression of fibrosis in chronic liver diseases.

  • CLDs, chronic liver diseases
  • DAPI, 4,6-diamidine-2-phenylindole di-hydrochloride
  • Δψm, mitochondrial membrane potential
  • ECL, enhanced chemiluminescence
  • ERK, extracellular regulated kinase
  • FasL, Fas ligand
  • HCV, hepatitis C virus
  • HSC, hepatic stellate cells
  • HSC/MFs, activated hepatic stellate cells in myofibroblast-like phenotype
  • JC-1, J-aggregate forming lipophilic cation
  • LDH, lactate dehydrogenase
  • mAb, monoclonal antibody
  • NGF, nerve growth factor
  • NFκB, nuclear factor κB
  • PARP, poly (ADP ribose) polymerase
  • SFI medium, serum free Iscove’s medium
  • α-SMA, α smooth muscle actin
  • siRNA, small interfering RNA
  • TBS, Tris buffered saline
  • h-TERT, human telomerase catalytic subunit
  • TIMP, tissue inhibitor of metalloproteinases
  • TNF-α, tumour necrosis factor α
  • TrkA, tyrosine kinase A
  • activated human hepatic stellate cells
  • liver fibrosis
  • apoptosis
  • Bcl-2
  • induction of cell death

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Footnotes

  • Published online first 19 January 2006

  • Conflict of interest: None declared.

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