The IL-1 receptor antagonist (IL-1Ra) knockout mice spontaneously develop a Th17 driven arthritis. When cross bred with TLR4 knock out (TLR4-/-) these animals showed reduced inflammation, joint destruction, and diminished IL-17 levels.
An age-matched (4-5 weeks), sex-mismatched reciprocal bone marrow transplantation was performed with TLR4-/- and TLR4+/+ mice in the IL-1Ra-/- background by 7.5 Gy irradiation and intravenous injection of freshly harvested bone marrow cells. Bone marrow was stained for Y-chromosome. Peritoneal macrophages were stimulated with lipopolysaccharide. Clinical manifestation of disease was monitored macroscopically (0 to 2 per joint). Animals were sacrificed at the age of 25 weeks. Bone erosion was scored on Safranin-O stained histological slides (0-5). T cell analysis was performed on cells from spleen and draining lymph nodes using flow cytometry.
Reconstitution of bone marrow was successful as determined by Y-chromosome staining. Peritoneal macrophages from animals engrafted with TLR4+/+ bone marrow showed normal IL-6 mRNA up regulation after LPS challenge, whereas IL-6 mRNA up regulation was significantly reduced by 90% in animals that received TLR4-/- bone marrow. Lack of TLR4 on either the engrafted bone marrow cells or the radio-resistant cells did not affect disease incidence, ranging from 55 to 90% between all groups and comparable to previous experiments. However, animals that lacked TLR4 on the engrafted bone marrow derived cells, radio-resistant cells, or both showed a reduced macroscopic arthritis score (mean 1.0, 0.9, and 0.9, respectively) compared to animals expressing Tlr4 on all cells (mean 1.6). This was also reflected in bone erosion scores of the joints. T-cell analysis showed no differences in Th1 or Th17 in the spleen. However, the percentage of IL-17 positive cells was reduced in the draining lymph nodes of animals lacking TLR4 on the bone marrow derived cells, resident cells, or both, averaging 0.7%, 0.7%, and 0.57% respectively, whereas lymph nodes of complete TLR4+/+ animals contained 1.1% IL-17 cells.
These data suggest that TLR4 plays an equally important role on both the bone marrow derived and local resident cells in Th17 development and in aggravating experimental arthritis. TLR4 could play a local role by creating a cytokine environment favouring Th17. On the other hand, TLR4 activation on the bone marrow derived antigen presenting cells might promote a more aggressive Th17 phenotype.
Disclosure: B. T. van Den Brand, None; S. Abdollahi-Roodsaz, None; M. B. Bennink, None; O. J. Arntz, None; W. van den Berg, None; F. A. van de Loo, None.