Skip to main content

Advertisement

SpringerLink
Log in

Inhibition of latent transforming growth factor-β1 activation by lentivirus-mediated short hairpin RNA targeting the CD36 gene in NR8383 cells

  • Published:
Molecular Biology Reports Aims and scope Submit manuscript

Abstract

CD36, a cell surface receptor for thrombospondin-1 (TSP-1), is believed to interact with latent transforming growth factor-β1 (L-TGF-β1) thereby activating its fibrogenic bioactivity. In this study, a lentiviral vector expressing a short hairpin RNA (shRNA) targeting the rat CD36 gene (Lv-shCD36) is developed and tested. To observe the inhibitory effect of Lv-shCD36 on the activation of L-TGF-β1, a rat alveolar macrophage cell line (NR8383), infected with either Lv-shCD36 or Lv-shCD36-NC (non-silenced control lentivirus), was treated with 0.1 μg/ml bleomycin, which is known to stimulate alveolar macrophages to release increasing amounts of TGF-β1. The results show that Lv-shCD36 can suppress expression of CD36 mRNA and protein in bleomycin-treated NR8383 cells. By quantifying active and total TGF-β1 in the supernatant, it was discovered that the quantity of total TGF-β1 is not significantly different between the three groups, while the quantity and percent of active TGF-β1 in the Lv-shCD36 group was significantly lower than in either the bleomycin-treated group or the Lv-shCD36-NC group, respectively (P < 0.05). These results suggest that Lv-shCD36 can inhibit activation of L-TGF-β1 secreted in bleomycin-treated NR8383 cells by decreasing the expression of CD36 on the cell membrane, thereby reducing binding of CD36 to TSP-1.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Khalil N, Whitman C, Zuo L, Danielpour D, Greenberg A (1993) Regulation of alveolar macrophage transforming growth factor-beta secretion by corticosteroids in bleomycin induced pulmonary inflammation in the rat. J Clin Invest 92:1812–1818. doi:10.1172/JCI116771

    Article  CAS  PubMed  Google Scholar 

  2. Cutroneo KR, White SL, Phan SH, Ehrlich HP (2007) Therapies for bleomycin induced lung fibrosis through regulation of TGF-beta1 induced collagen gene expression. J Cell Physiol 211:585–589. doi:10.1002/jcp.20972

    Article  CAS  PubMed  Google Scholar 

  3. Chen Y, Chen J, Dong J, Liu W (2005) Antifibrotic effect of interferon gamma in silicosis model of rat. Toxicol Lett 155:353–360. doi:10.1016/j.toxlet.2004.10.011

    Article  CAS  PubMed  Google Scholar 

  4. Daniels CE, Wilkes MC, Edens M, Kottom TJ, Murphy SJ, Limper AH, Leof EB (2004) Imatinib mesylate inhibits the profibrogenic activity of TGF-β and prevents bleomycin mediated lung fibrosis. J Clin Invest 114:1308–1316

    CAS  PubMed  Google Scholar 

  5. Yehualaeshet T, O’Connor R, Green-Johnson J, Mai S, Silverstein R, Murphy-Ullrich JE, Khalil N (1999) Activation of rat alveolar macrophage-derived latent transforming growth factor-β1 by plasmin requires interaction with thrombospondin-1 and its cell surface receptor, CD36. Am J Pathol 155:841–851

    CAS  PubMed  Google Scholar 

  6. Yang YL, Lin SH, Chuang LY, Guh JY, Liao TN, Lee TC, Chang WT, Chang FR, Hung MY, Chiang TA, Hung CY (2007) CD36 is a novel and potential anti-fibrogenic target in albumin-induced renal proximal tubule fibrosis. J Cell Biochem 101:735–744. doi:10.1002/jcb.21236

    Article  CAS  PubMed  Google Scholar 

  7. Yehualaeshet T, O’Connor R, Begleiter A, Murphy-Ullrich JE, Silverstein R, Khalil N (2000) A CD36 synthetic peptide inhibits bleomycin-induced pulmonary inflammation and connective tissue synthesis in the rat. Am J Respir Cell Mol Biol 23:204–212

    CAS  PubMed  Google Scholar 

  8. Gartel AL, Kandel ES (2006) RNA interference in cancer. Biomol Eng 23:17–34. doi:10.1016/j.bioeng.2006.01.002

    Article  CAS  PubMed  Google Scholar 

  9. Lu PY, Xie F, Woodle MC (2005) In vivo application of RNA interference: from functional genomics to therapeutics. Adv Genet 54:117–142

    CAS  PubMed  Google Scholar 

  10. Abbas-Terki T, Blanco-Bose W, Déglon N, Pralong W, Aebischer P (2002) Lentiviral-mediated RNA interference. Hum Gene Ther 13:2197–2201. doi:10.1089/104303402320987888

    Article  CAS  PubMed  Google Scholar 

  11. Li M, Rossi JJ (2008) Lentiviral vector delivery of siRNA and shRNA encoding genes into cultured and primary hematopoietic cells. Methods Mol Biol 433:287–299. doi:10.1007/978-1-59745-237-3_18

    Article  CAS  PubMed  Google Scholar 

  12. Zielske SP, Stevenson M (2005) Importin 7 may be dispensable for human immunodeficiency virus type 1 and simian immunodeficiency virus infection of primary macrophages. J Virol 79:11541–11546. doi:10.1128/JVI.79.17.11541-11546.2005

    Article  CAS  PubMed  Google Scholar 

  13. Pullmann R Jr, Juhaszova M, López de Silanes I, Kawai T, Mazan-Mamczarz K, Halushka MK, Gorospe M (2005) Enhanced proliferation of cultured human vascular smooth muscle cells linked to increased function of RNA-binding protein HuR. J Biol Chem 280:22819–22826. doi:10.1074/jbc.M501106200

    Article  CAS  PubMed  Google Scholar 

  14. Sakoda T, Kasahara N, Hamamori Y, Kedes L (1999) A high-titer lentiviral production system mediates efficient transduction of differentiated cells including beating cardiac myocytes. J Mol Cell Cardiol 31:2037–2047. doi:10.1006/jmcc.1999.1035

    Article  CAS  PubMed  Google Scholar 

  15. Soneoka Y, Cannon PM, Ramsdale EE, Griffiths JC, Romano G, Kingsman SM, Kingsman AJ (1995) A transient three-plasmid expression system for the production of high titer retroviral vectors. Nucleic Acids Res 23:628–633. doi:10.1093/nar/23.4.628

    Article  CAS  PubMed  Google Scholar 

  16. Denholm EM, Rollins SM (1993) Expression and secretion of transforming growth factor-beta by bleomycin-stimulated rat alveolar macrophages. Am J Physiol Lung Cell Mol Physiol 264:L36–L42

    CAS  Google Scholar 

  17. Xu Y, Mirmalek-Sani SH, Lin F, Zhang J, Oreffo RO (2007) Adipocyte differentiation induced using nonspecific siRNA controls in cultured human mesenchymal stem cells. RNA 13:1179–1183. doi:10.1261/rna.527207

    Article  CAS  PubMed  Google Scholar 

  18. Febbraio M, Hajjar DP, Silverstein RL (2001) CD36: a class B scavenger receptor involved in angiogenesis, atherosclerosis, inflammation, and lipid metabolism. J Clin Invest 108:785–791

    CAS  PubMed  Google Scholar 

  19. Pfeifer A, Eigenbrod S, Al-Khadra S, Hofmann A, Mitteregger G, Moser M, Bertsch U, Kretzschmar H (2006) Lentivector-mediated RNAi efficiently suppresses prion protein and prolongs survival of scrapie-infected mice. J Clin Invest 116:3204–3210. doi:10.1172/JCI29236

    Article  CAS  PubMed  Google Scholar 

  20. Liu L, Zhang Q, Zhang Y, Wang S, Ding Y (2006) Lentivirus-mediated silencing of Tiam1 gene influences multiple functions of a human colorectal cancer cell line. Neoplasia 8:917–924. doi:10.1593/neo.06364

    Article  CAS  PubMed  Google Scholar 

  21. Katayama K, Wada K, Miyoshi H, Ohashi K, Tachibana M, Furuki R, Mizuguchi H, Hayakawa T, Nakajima A, Kadowaki T, Tsutsumi Y, Nakagawa S, Kamisaki Y, Mayumi T (2004) RNA interfering approach for clarifying the PPARgamma pathway using lentiviral vector expressing short hairpin RNA. FEBS Lett 560:178–182. doi:10.1016/S0014-5793(04)00100-0

    Article  CAS  PubMed  Google Scholar 

  22. Kuchibhotla S, Vanegas D, Kennedy DJ, Guy E, Nimako G, Morton RE, Febbraio M (2008) Absence of CD36 protects against atherosclerosis in ApoE knock-out mice with no additional protection provided by absence of scavenger receptor A I/II. Cardiovasc Res 78:185–196. doi:10.1093/cvr/cvm093

    Article  CAS  PubMed  Google Scholar 

  23. Khalil N (2001) Post translational activation of latent transforming growth factor beta (L-TGF-beta): clinical implications. Histol Histopathol 16:541–551

    CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by grants from the National Natural Science Foundation Committee of China (No. 30500232), the Specialized Research Fund for the doctoral program of higher education (No. 20060159006), and the Liaoning Province Natural Science Foundation (No. 20072102).

Author information

Authors and Affiliations

  1. Division of Pneumoconiosis, School of Public Health, China Medical University, 92 North 2nd, 110001, Heping District, Shenyang, People’s Republic of China

    Xin Wang, Ying Chen, Lina Lv & Jie Chen

Authors
  1. Xin Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Ying Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Lina Lv
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Jie Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jie Chen.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wang, X., Chen, Y., Lv, L. et al. Inhibition of latent transforming growth factor-β1 activation by lentivirus-mediated short hairpin RNA targeting the CD36 gene in NR8383 cells. Mol Biol Rep 37, 1649–1655 (2010). https://doi.org/10.1007/s11033-009-9579-2

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11033-009-9579-2

Keywords

Search

Navigation