The role of leukotriene inhibition using a 5-lipoxygenase (5-LO) inhibitor in a joint contracture model

Abstract

Purpose: Arthrofibrosis is a common inflammatory complication of joint trauma and surgery. 5lipoxygenase (5-LO) is a key enzyme involved in inflammation. Inhibition of 5-LO has been shown to reduce inflammation in heart and lung models but has not been examined in a joint contracture model.

Methods: Twenty-six rats underwent joint contracture. Six rats served as non-surgical controls. A 5-LO inhibitor, caffeic acid (CA), suspended in 10% ethanol was orally administered to 14 rats and ethanol without CA to the remaining 12 rats daily for 21 days. Leukotriene B4 (LTB4) levels were measured, both systemically and locally. 5-LO levels in the posterior capsule were quantified by measuring the ratio of the length of the posterior capsule demonstrating 5-LO immunostaining to the total length of the capsule.

Results: Joint contracture was successfully achieved in all rats who underwent manipulation. Levels of 5- LO measured in the posterior capsule were significantly increased in the animals who underwent surgery (56%/44-64) compared to the non-surgical control animals (7%/4-9). LTB4 levels were found to be significantly lower in the non-surgical control animals (107.79 ± 34.08 pg/ml) compared to all surgical animals (157.6 ± 55.3 pg/ml).

Conclusion: Surgical intervention resulted in increased 5-LO activity of the synovial surface of the posterior capsule and increased LTB4 levels in the patellar tendon-fat pad. Oral administration of the 5LO inhibitor, CA, was ineffective at reducing systemic and local LTB4 levels and preventing knee joint contracture. Inhibiting 5-LO activity may still be effective in preventing arthrofibrosis and warrants further investigation.

Keywords: And 5-Lipoxygenase; Arthrofibrosis; Caffeic Acid; Contracture; Leukotriene.

Fig. 1

The eicosanoid pathway including relative metabolites and effects. Highlighted in red are the various treatment modalities that inhibit parts of this pathway. Metabolites include prostaglandin I₂ (PGI2), thromboxane A₂ (TXA2), and leukotrienes B₄, C₄, D₄, and E₄ (LTB4, C4, D4 and E4)

Fig. 2

Male rats were obtained from the commercial vendor Charles River. Group 1: Surgical with CA in 10% ethanol (EtOH). Group 2: Surgical without CA in 10% EtOH. Group 3: Non-surgical controls. Group 2 received 10% EtOH on their food. Animals were assigned to groups based on weight. Average weight across all three groups was 488.53 g. Of the 32 animals, seven were eliminated for the above reasons

Fig. 3

Radiograph of operative leg of a rat in the surgical without CA group with the immobilization suture intact

Fig. 4

Apparatus used for FTA measurement in all operative and nonoperative limbs. Landmarks were designated with india ink at the medial malleolus, the origin of the MCL and the center of the femoral head

Fig. 5

4 × 5 μm sagittal section of operative rat knee stained with Abcam anti-rabbit 5-LO antibody. Femoral and tibial bone marrow served as positive control for staining. Posterior capsule outlined in blue and posterior capsule demonstrating staining outlined in black

Fig. 6

Daily food consumption was recorded and converted into a percent consumption. Pre-surgery data indicated a mean 80% of total daily consumption

Fig. 7

Food consumption in the caffeic acid treatment group during postoperative days 1–3, when buprenorphine was dosed, compared to days 4–21 when no buprenorphine was used

Fig. 8

Immunostaining for 5-LO done on 5 μm sagittal sections of rat knee joint with the posterior capsule intact. Sections are from the medial mid-condylar region. a 1 × image of medial femoral condyle, medial tibial plateau and associated posterior capsule of an immobilized animal. Positive 5-LO activity is represented by uptake of brown stain. Bone marrow in femur and tiba represent a good internal control. b, c and d are 4 × images of (a) highlighting the staining in the posterior capsule of an immobilized animal