Rheumatoid arthritis (RA) is traditionally considered a chronic, inflammatory autoimmune disorder that causes the immune system to attack the tissues of joints. Once triggered, the immune response causes inflammation of the synovium, leading to edema, vasodilation and infiltration by activated T-cells (mainly CD4 in nodular aggregates and CD8 in diffuse infiltrates). Early and intermediate molecular mediators of inflammation include tumor necrosis factor alpha (TNF-α), interleukins IL-1, IL-6, IL-8 and IL-15, transforming growth factor beta, fibroblast growth factor and platelet-derived growth factor. Biomarkers of the disease also includes the activation/upregulation of NK-kappaB (NFkB), Inducible Nitric Oxide Synthase (iNOS), Serum Amyloid A-1 (SAA-1), Eosinophil Peroxidase (Epx), Cyclooxygenase-2 (COX2) and others.

There is no known cure for rheumatoid arthritis and current treatments focus on alleviating symptoms. One of the possible reasons for the lack of treatments may be due to the limitations of the preclinical animal models that are available. There are a number of different preclinical inflammation models of rheumatoid arthritis, each with strengths and weaknesses when compared with the human aspects of the disease. The following is a summary of the preclinical inflammation models of rheumatoid arthritis MIR has to offer:

Collagen-induced arthritis (CIA) is a preclinical animal inflammation model of rheumatoid arthritis (RA) that is widely used to address questions of disease pathogenesis and to validate therapeutic targets. Collagen-induced arthritis models can be performed in both in mice or rats by immunization with heterologous type II collagen in adjuvant. Susceptibility to collagen-induced rheumatoid arthritis is strongly associated with major histocompatibility complex class II genes, and the development of rheumatoid arthritis is accompanied by a robust T-cell and B-cell inflammation response to type II collagen.

Pathological Features:

The chief pathological features of collagen-induced arthritis include a proliferative synovitis with infiltration of polymorphonuclear and mononuclear cells, pannus formation, cartilage degradation, erosion of bone, and fibrosis. As in human rheumatoid arthritis, pro-inflammatory cytokines, such as tumor necrosis factor alpha (TNF-a), interleukin-1b (IL-1b) and IL-6 are increased in collagen-induced arthritis. Biological therapies designed to interfere with these mediators are active in these models.

Measurements:

Disease activity is assessed by measuring inflammation swelling in the affected joints (paw volume or thickness) over time. Treatments can be assessed in either prophylactic or therapeutic testing paradigms. Additional measure of disease activity include evaluation of serum IL-1b, IL-6, C-reactive protein (CRP) or serum amyloid A (SAA), and erythrocyte sedimentation rate. Bone lesion scoring conducted by preclinical Positron Emission Tomography (preclinical PET). MIR typically measures all four paws for thickness and gives an average paw thickness for each animal at each timepoint. Animal body weights are typically taken 2 times per week.

General Outline of Study:

Collagen-induced arthritis can be induced in either rats or mice using bovine type II collagen. Prophylactic studies in rats are carried out over 28 days (50 days for mice), while therapeutic studies require a 14 day protocol (26 for mice).

Histology and Analytical Services :

MIR Preclinical Services has established strategic partners for histology, immunohistochemistry, Phamacokinetics, blood chemistries and other pertainent preclinical services that clients may request. Each strategic partner is a specialist in the area of interest and MIR Preclinical Services can offer these services as a part of its offerings.

Reports:

Each week clients receive an interim report that contains all the data for a current study up to that day. This contains data on body weights, changes in body weights over time, paw measurements, changes in paw measurements over time, clinical based scoring, daily census, comments, necropsy findings and summary graphics. Clients will also receive an email detailing each significant study milestone including when animals are ordered, when animals are injected with inflammatory agents, when treatment begins, when treatment ends and when the study ends. Final reports contain detail materials and methods, statistical analysis, publication quality tables and figures, popular enpoint calculations (paw thickness, peak swelling, time to peak swelling, clinical grade scoring, overall severity, AUC, etc.)and all raw data for the study. Custom endpoints and analyses can also be generated upon request. MIR Preclinical Services strives to have final reports to clients within 3 weeks of the euthanasia of the last animal on study for preclinical pharmacology experiments and between 5 to 7 weeks for extensive preclinical imaging studies (depending on size and imaging modality).

Imaging in Collagen Induced Arthritis :

MIR Preclinical Services is the leading contract research organization in the application of preclinical imaging to preclinical models of cancer and arthritis. MIR has preclinical computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET) and biophotonic imaging capabilities. MIR Preclinical Services owns all of their own imaging equipment and operates this equipment on site. All of the company's imaging equipment is state-of-the-art and designed exclusively for small animals.

Control Rats
Methotrexate at 0.3mg/kg
Dexamethasone at 1.0mg/kg

Lipopolysaccharide (LPS) is an endotoxin that binds CD14/TLR4/MD2 receptor complex and induces a strong response from the immune system. This dramatically increases the secretion of pro-inflammatory cytokines in many cell types, but especially in macrophages. This model enhances CIA and produces a 100% incidence of arthritic symptoms in mice within 24 to 48 hours after LPS is injected. Unlike the ollagen-induced arthritis model, arthritis progresses rapidly, within a few hours compared to weeks.

Pathological Features:

Simailr to collagen-induced arthritis (CIA) described above.

Measurements:

Simailr to collagen-induced arthritis (CIA) described above.

General Outline of Study:

LPS synchronized c ollagen-induced arthritis can be induced in mice using bovine type II collagen. Prophylactic studies in rats are carried out over 50 days, while therapeutic studies require a 26 day protocol. LPS synchronizes the development of the disease so the study tends to have less overall variability than standard CIA mice models.

Collagen antibody induced arthritis is a model of human rheumatoid arthritis. The model has the advantage of inducing disease in many strains of mice that are resistant to the traditional collagen induced arthritis models. Collagen antibody induced arthritis relies on the injection of a cocktail of monoclonal antibodies directed against type II collagen (C-II), followed by a single injection of LPS. In animals, a significant part of the inflammatory attack on the joints is mediated by pathogenic antibodies directed against C-II. An advantage of this model is that compound assessment can be completed in a relatively short period of time in the Collagen antibody induced arthritis. In standard Collagen induced arthritis protocols, disease doesn’t begin to appear until 3 – 4 weeks and a typical study might last 6 – 8 weeks. In Collagen antibody induced arthritis, the disease appears within 7- 8 days and studies can be completed within 18 days. An additional 7 days can be added to assess the effects of agents on the resolution phase of the disease. Collagen antibody induced arthritis is dependent on IL-1β, and TNF-α, but is independent of the effects of IL-6. The model is quite robust and produces disease in virtually 100% of the injected animals.

Pathological Features:

Injected animals develop rapid and severe arthritis within 2 – 3 days following LPS injection. Swelling develops in all limbs (hind and fore paws). Published literature suggests that histologic changes can be observed in the joints after 18 days on study.

Measurements:

Footpad thickness, measured in mm via digital vernier calipers and clinical score. Clinical scoring is accomplished by awarding a score of 1 for each swollen digit, a score of 5 for a swollen footpad, and a score of 5 for a swollen wrist or ankle. These are added together to give a maximal score of 60 for each animal.

General Outline of Study:

Female mice are given an IV injection of 2mg of ArthritoMab™ antibody cocktail (MDBioscience). Six days later each animal receives an intraperitoneal injection of 100μg of LPS. Measurements of footpad thickness by caliper, and clinical scores are recorded every 2 days for 21 days. At study termination, limbs may be imaged or harvested for cytokine analyses.

Histology and Analytical Services :

MIR Preclinical Services has established strategic partners for histology, immunohistochemistry, Phamacokinetics, blood chemistries and other pertainent preclinical services that clients may request. Each strategic partner is a specialist in the area of interest and MIR Preclinical Services can offer these services as a part of its offerings. Standard joint histology, and Luminex analysis of cytokine levels in diseased paws.

Reports:

Each week clients receive an interim report that contains all the data for a current study up to that day. This contains data on body weights, changes in body weights over time, paw measurements, changes in paw measurements over time, clinical based scoring, daily census, comments, necropsy findings and summary graphics. Clients will also receive an email detailing each significant study milestone including when animals are ordered, when animals are injected with inflammatory agents, when treatment begins, when treatment ends and when the study ends. Final reports contain detail materials and methods, statistical analysis, publication quality tables and figures, popular enpoint calculations (paw thickness, peak swelling, time to peak swelling, clinical grade scoring, overall severity, AUC, etc.)and all raw data for the study. Custom endpoints and analyses can also be generated upon request. MIR Preclinical Services strives to have final reports to clients within 3 weeks of the euthanasia of the last animal on study for preclinical pharmacology experiments and between 5 to 7 weeks for extensive preclinical imaging studies (depending on size and imaging modality).

Imaging in Mouse Collagen Antibody Induced Arthritis :

We are currently assessing structural changes in the joint via CT.

This model is currently under development. Please call to inquire on availability.