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Arthritis on the ASK

Overview
Products for imaging arthritis
- Choosing the right agent
Mouse model for arthritis
Application notes and posters
Citations

Overview

Arthritis is inflammation in or around a joint, where two bones meet. A healthy joint is cushioned by cartilage and lined with synovium tissue which produces fluid to reduce friction during movement. Osteoarthritis occurs when wear and tear of the joint causes breakdown of the cartilage and inflammation of the synovial fluid. Osteoarthritis results from long term use of the joint and is associated with old age. Rheumatoid arthritis also results in breakdown of the joint, but as a result of an auto-inflammatory response.

Current methods to assess arthritis in animal models typically involve external measurements of limbs such as paw volume, paw thickness, and redness. While these methods do determine the overall progression of disease, they do not examine the underlying biology associated with the visible effects of the disease. Since arthritis involves not only inflammation pathways but also changes in vasculature and bone morphology, using fluorescent agents which can measure each of these changes individually allow for a more complete assessment of the overall clinical pathology. Preclinical imaging in the far-infrared region allows for non-invasive assessment of the biological response of the mouse over realistic timeframes of disease progression and for monitoring therapeutic efficacy.

Products for imaging arthritis

Our preclinical imaging agents fall into three main categories: Activatable, meaning they are optically silent until cleaved by a protease of interest; Targeted, meaning they bind to a receptor or other cellular membrane constituent which must be present in the model system of interest; or Vascular, meaning they localize to vasculature and are used to assess changes in vascular permeability associated with disease.

The following agents can be used to image arthritis in vivo. Many of these agents can also be used to study arthritis in vitro and ex vivo. Please select an agent below to learn more about its technical properties and validated protocols. All of our agents can be used on FMT and IVIS systems, provided you have the capability to measure the wavelengths of the probes.

Agent	Agent Type	Agent Mechanism	Available wavelengths and optimal in vivo imaging time (post injection)	Route of metabolism/ background tissue(s)	Validated imaging methods*
Cat B FAST™	Activatable, fluorescent	Detects cathepsin B activity; Cat B accumulates in synovial fluid in rheumatoid arthritis	680 FAST: 6-24 h 750 FAST: 6-24 h	Salivary glands, liver, kidneys	In vivo, flow cytometry, in vitro cell microscopy, frozen tissue labeling
Cat K FAST™	Activatable, fluorescent	Detects cathepsin K activity, which occurs in osteoclasts, chondrocytes and synovial fibroblasts during bone turnover	680: 6-24 h	Kidneys, liver	In vivo, flow cytometry, in vitro cell microscopy, frozen tissue labeling
MMPSense™	Activatable, fluorescent	Detects the activity of matrix metallo- proteases 2, 3, 7, 9, 12 and 13 (MMP2, MMP3, MMP7, MMP9, MMP12, MMP13); MMP 13 is responsible for collagen breakdown in chondrocytes, while MMP’s 2, 3, and 9 degrade the non-collagen matrix components of joints	645 FAST: 24 h (6-24 h) 680: 24 h (24-36 h) 750 FAST: 24 h (12-24 h)	Liver, kidneys (645 and 750 FAST) Liver only (680)	In vivo
ProSense^®	Activatable, fluorescent	Detects cathepsin B, L, S and plasmin: enzymes involved in degradation of the extracellular matrix	680: 24 h (24-48 h) 750: 24 h 750 FAST*: 6-24 h	Liver (680) Low liver, intestines (750) Low liver, bladder (750 FAST)	In vivo, flow cytometry, in vitro cell microscopy
RediJect COX-2 probe	Targeted, fluorescent	Efficiently target cyclooxygenase-2 (COX-2), which is normally absent from cells, but is found at high levels in inflammatory lesions	600 nm: 3 h	Kidney, liver	In vivo, in vitro cell microscopy
HypoxiSense™	Targeted, fluorescent	Binds to carbonic anhydrase IX which is expressed in hypoxic tissues; in Rheumatoid arthritis, dysfunctional vasculature leads to high amounts of hypoxia	680: 24 h	Kidney	In vivo, flow cytometry, in vitro cell microscopy, frozen tissue labeling
RediJect Inflammation Probe	Targeted, chemiluminescent	Detects myelo- peroxidase activity, which is released by activated phagocytes in both chronic and acute inflammation	Emits at 425 nm, 10 min post-injection, 5 min BLI acquisition time, internal fluorescent control imaged at 745 nm excitation and 800 nm emission 1-5 sec acquistion	Kidney	In vivo
OsteoSense^®	Targeted, fluorescent	Binds to hydroxyapatite, an indicator of bone remodeling	680: 3-24 h 750: 3-24 h 800: 3-24 h	Bladder	In vivo
AngioSense^®	Vascular, fluorescent	Will accumulate in areas of vascular leakage associated with tumorigenesis and inflammation; can be used to visualize the neovasculature	680: 24 h 750: 24 h	Low liver, lung	In vivo
AngioSPARK^®	Vascular, fluorescent	Nanoparticle for imaging vascularity, long PK profile, 20 hour half-life in plasma; can also be used to assess vascular leak associated with arthritis	680: 24 h 750: 24 h	Long term tissue accumulation	In vivo

Choosing the right agent

When choosing an agent for arthritis studies, several factors should be taken into consideration:

Fluorescent vs. Chemiluminscent agents: Be sure to choose the proper agent for your instrumentation. Some imagers can measure both chemiluminescence and fluorescence, while others can only measure one or the other.
Dye excitation/emission wavelength: Some fluorescent agents are available with excitation wavelengths that range from 645 nm to 800 nm. Be sure to pick a wavelength that is appropriate for both your instrument and application. Most microscopes filters are not suitable for wavelengths above 680 nm. However deep tissue imaging typically has less background fluorescence and works better with longer wavelengths (750 nm- 800 nm).
Agent clearancetime is typically faster for activatable agents (particularly the FAST™ platform) which allows for re-injection after shorter time periods.
Agent specificity: For targeted and activatable agents, it is important to remember that the protein or enzyme that the agent is targeting needs to be present and well expressed in your animal model.
In vivo distribution: Some agents might accumulate in organs at timepoints which could interfere with your study. Be sure to check the biodistribution profile of each agent of interest.
Type of imaging required for your study: Vascular agents for example, while useful in assessing vascularity changes in the mouse, do not make good agents for in vitro cell imaging or tissue analysis. If these types of analyses are desired, targeted or activatable agents might be a better choice for you.

View more information on agents for other bone biology applications.

Mouse model for arthritis

Antibody-induced arthritis

Monoclonal antibodies to collagen II can trigger an immune response in mice which provides a realistic model for rheumatoid arthritis. Eight days after treatment with the antibodies, the mouse exhibits pathology such as inflammation of the joint and synovium in addition to breakdown of the cartilage and bone. This model can be used in numerous mouse strains, including BALB/c which is typically resistant to other arthritis induction models (i.e. collagen-induced arthritis).

In Figure 2 (below), arthritis was induced in BALB/c mice and after 8 days the imaging agent CAT K 680 FAST™ was injected intravenously and the mouse was imaged 24 hours later. Fluorescent imaging results were compared to more traditional assessments of disease progression, such as clinical. As shown in Figure 3, 3D tomographic and 2D images were obtained from both arthritic and controlled mice and signal from fluorescence was compared to both paw thickness measurements and clinical score assessments.

Figure 1: Mouse antibody-induced arthritis. A) 5-week-old BALB/c mice are injected with 4 mg of ArthritoMab antibody cocktail followed 3 days later with LPS. After 8 days, CAT K 680 FAST™ is injected and the mouse is imaged 24 hour later, where fluorescent quantification is compared to paw thickness. B) Clinical score is typically assessed by visual assessment of paw swelling and redness.

Figure 2: Assessment of arthritic mice. A) 3D tomographic images were obtained by FMT 2500 B) Paw thickness (vernier caliper), clinical observation score and quantification of CAT K FAST fluorescence from tomographic datasets C) 2D epifluorescence images acquired using FMT 2500 D) Ex vivo tissue analysis and 2D Cat K FAST epifluorescent imaging.

In Figure 3 below, the activatable agents ProSense® and MMPSense™ are used assess protease activity in vivo, while the vascular imaging agent AngioSense™ detects changes in vascular leak associated with arthritis.

Figure 3: Activatable and Vascular agents used to image arthritis. A) The activatable agents ProSense and MMPSense are used to image protease activity associated with inflammation. B) The vascular agent AngioSense is used to image vascular leak using both planar imaging and intravital microscopy measurements in both healthy and arthritic tissue.

In addition to in vivo and ex vivo measurement as shown above, agents can also be used to label cells and imaged using fluorescence microscopy. In Figure 4A below, human synovial fibroblasts were isolated from rheumatoid arthritis patients.and labeled with CAT K 680 FAST. In Figure 4B rabbit synoviocytes (HIG-62 cell lines) were also labeled with the agent.

Figure 4: Uptake and activation of CAT K 680. Synovial fibroblasts isolated from human rheumatoid patient (A) or rabbit cell line (B). Cells were cultured with 1 mM CAT K FAST for 6 hours. Red: CAT K 680 FAST, Blue: DAPI nuclear stain; final magnification 40x.

Application notes and posters

Poster: Imaging of Cathepsin K activity in rodent models of bone turnover and soft tissue calcification
Poster: Development of a Fast Activating New Near Infrared-Labeled Agent for Detecting Cathepsin K Activity

Citations

Please visit our Citations library to search or browse references for arthritis studies using our preclinical imaging agents.