Endogenous heme oxygenase-1 down-regulates articular inflammation

R Brines1, ML Ferrándiz1, N Maicas1, A Loboda2, A Jozkowicz2, J Dulak2, MJ Alcaraz1. 1University of Valencia, Department of Pharmacology and IDM 46100, Spain, 2Jagiellonian University, Krakow, Department of Medical Biotechnology 30-387, Poland

Background: The enzyme heme oxygenase-1 (HO-1) is induced in many cell types as a defense mechanism against oxidative stress. Results of previous studies indicate that HO-1 induction plays a role in protecting tissues against injury through the modulation of immune responses or effector cells. Rheumatoid arthritis (RA) is characterized by a progressive degradation of joint cartilage and bone.Transfer of serum from K/BxN transgenic induces an autoimmune and inflammatory response with similarities to human RA.

Objective: We have investigated whether endogenous HO-1 would play a role in inflammatory arthritis by inducing K/BxN serum transfer arthritis in HO-1 wild type (HO-1+/+), heterozygous (HO-1+/-) and homozygous knock out (HO-1-/-) mice.

Methods: Arthritis was induced in 20-27-week-old C57/Black-6 xFVB(HO-1+/+, HO-1+/- and HO-1-/-) male mice (25±5 g) by intraperitoneal injection of 150 µl serum from K/BxN arthritic mice on days 0 and 2 (n=10 in arthritic groups and n=4 in naïve groups). All mice were maintained with a 12-hour light/dark cycle, at 22ºC and free access to sterilized food and water. Blood was collected from the retroorbital venous plexus at days 4, 7 and 10, as well as from tail at day 10, and animals were sacrificed by cervical dislocation at day 10. Histological analysis was performed on ankle sections. Levels of mediators were measured in serum and in ankle homogenates by ELISA or Multiplex. Results were expressed as mean±SEM and analysed by one way ANOVA and Dunnett\'s t-test.

Results: Naïve HO-1-/- mice showed increased counts of lymphocytes, granulocytes and monocytes with respect to HO-1+/+ or HO-1+/- animals which were also observed after arthritis induction. In addition, arthritis resulted in the production of anemia accompanied by increased platelet counts. Arthritis severity was increased in HO-1+/- (3.11±0.48, pË‚0.05) and in HO-1-/- (1.83±0.68) mice with respect to the HO-1+/+ group (1.35±0.26). Histological analysis showed the highest inflammatory cells migration and cartilage proteoglycan depletion, in the HO-1+/- group. The induction of arthritis resulted in a significant enhancement in myeloperoxidase activity with respect to the corresponding naïve animals with values of 0.650±0.039 units/g (p<0.05) and 0.823±0.053 units/g (p<0.01) in arthritic HO-1+/+ and HO-1+/- mice, respectively. An increase in serum levels of monocyte chemotactic protein-1 (46.2±8.3 pg/ml, p<0.01), plasminogen activator inhibitor-1 (28.3±5.5 pg/ml, p<0.01), E-selectin (428.2±45.7 ng/ml, p<0.05) and intercellular adhesion molecule-1 (63.5±8.5 ng/ml, p<0.01) was observed in arthritic HO-1-/- mice with respect to wild type animals (9.8±1.2 pg/ml, 7.4±0.4 pg/ml, 232.6±22.9 ng/ml and 63.5±8.5 ng/ml, respectively). In addition, higher levels of IL-6 (342.1±67.2 pg/g, p<0.05) and CXCL-1 (122.8±19.4 pg/g, p<0.05) were measured in ankle homogenates from arthritic HO-1+/- mice with respect to HO-1+/+ animals (195.7±27.9 and 93.8±9.7 pg/g, respectively) .

Conclusion: Our data show that endogenous HO-1 plays a protective role in articular inflammation, supporting an anti-inflammatory role of HO-1.

Maicas N, Ferrándiz ML, Brines R, Ibáñez L, Cuadrado A, Koenders MI, van den Berg WB, Alcaraz MJ. Antioxid. Redox Signal. (2011) 15:889-901.