MD Biosciences Blog

Continuing the discussion of imaging technologies, this week we will cover biofluorescence and bioluminescence as readouts for RA models.

Traditional extrinsic fluorescent dyes fluoresce within the range of 300-500nm. Unfortunately, at these wavelengths, biological samples autofluoresce. Recent advancement in technology and development of dyes that fluoresce close to the near infrared (NIR) range (700-900nm) has produced dyes better suited for in vivo studies. A charged-coupled detector (CCD) is used to detect the photons emitted from these dyes after appropriate excitation. Initially these studies were limited due to interference from thermal energy. However, cooling of the chip markedly increased the quality of the image (1). Imaging using these techniques, coined fluorescence molecular tomography (FMT) or optical fluorescence tomography (OFT) in some studies, has been used to follow therapeutic agents (2), assess cell surface molecule expression (3) and to determine disease state by quantifying enzymatic activity (4, 5). Enzymatic activity is assessed using activity based probes (APBs). ABPs consist of a dye and quencher attached to opposite ends of a peptide linker. When the peptide is cleaved by a protease the signal is released. Commercially available APBs Prosense750 (which detects Cathepsin activity) and ProSense680 (which detects MMP activity) were used by Peterson et al (5) in their study assessing the performance of different therapeutic agents. There are also non-specific dyes which accumulate due to increased vascular leakage (6).

Bioluminescence

Like fluorescence imaging, bioluminescence imaging offers a non-toxic, non-invasive means of following specific events longitudinally in live animals. However, unlike fluorescence imaging, bioluminescence imaging requires no exogenous excitation. The most commonly used bioluminescence assay involves the luciferase gene. The luciferase gene, found naturally in glow worms and fireflies, oxidises Luciferin. This reaction results in the release of a photon, which can be quantified and localised with imaging systems. Luciferase-expressing cells can be transferred into hosts or Tg mice expressing luciferase under specific promoters of interest can be used in bioluminescence studies. After injection of the substrate luciferin the cells of interest can be imaged. Nakajima et al (7) used luciferase bioluminescence to show that adoptively transferred CII-specific T cells home to the joints of CIA arthritic mice. In other studies mice expressing luciferase under the Nf-kB (8) or human IL-1b (9) reporter were used to investigate arthritis models and the animal’s response to therapy. One group used bioluminescence in the CIA model to directly visualise their novel therapy (a cytolytic adenovirus) (10). Luminol is also a bioluminescent agent. When exposed to an appropriate oxidising species, luminol emits a blue luminescence. MPO activity at sites of inflammation can generate the necessary oxidative species to catalyse this reaction. Investigators have exploited this using a model of LPS-induced arthritis where luminol bioluminescence was shown to co-localise with sites of inflammation (11).

View pre-clinical models of Rheumatoid Arthritis

References

1. Spibey CA, Jackson P, Herick K.  Electrophoresis. 2001 22(5):829-36.
2. Paiframan R, Airey M, Moore A, Vulger A, Nesbitt A. J Immunol Methods. 31;348(1-2):36-41.
3. Hansch A, Frey O, Sauner D, Hilger I, Haas M, Malich A, Bräuer R, Kaiser WA.  Arthritis Rheum. 2004 50(3):961-7.
4. Wunder A, Tung CH, Muller-Ladner U, Weissleder R, Mahmood U.  Arthritis Rheum. 2004 50(8):2459-65.
5. Peterson JD, Labranche T, Vasquez KO, Kossodo S, Melton M, Rader R, Listello JT, Abrams MA, Misko TP.  Arthritis Res Ther. 2010 12(3):R105.
6. Hansch A, Frey O, Hilger I, Sauner D, Haas M, Schmidt D, Kurrat C, Gajda M, Malich A, Brauer R, Kaiser WA.  Invest Radiol. 2004 39(10):626-32.
7. Nakajima A, Seroogy CM, Sandora MR, Tarner IH, Costa GL, Taylor-Edwards C, Bachmann MH, Contag CH, Fathman CG.  J Clin Invest. 2001 107(10):1293-301.
8. Carlsen H, Moskaug JO, Fromm SH, Blomhoff R. J Immunol 2002 168(3):1441.
9. Li L, Fei Z, Ren J, Sun R, Liu Z, Sheng Z, Wang L, Sun X, Yu J, Wang Z, Fei J. BMC Immunol. 2008 9:49.
10. Chen SY, Shiau AL, Shieh GS, Su CH, Lee CH, Lee HL, Wang CR, Wu CL. Arthritis Rheum. 2009 60(11):3290-302.
11. Gross S, Gammon ST, Moss BL, Rauch D, Harding J, Heinecke JW, Ratner L, D. Nat Med. 2009 15(4):455-61.