Tumor Necrosis Factor Induces Obliterative Pulmonary Vascular Disease in a Novel Model of Connective Tissue Disease-Associated Pulmonary Arterial Hypertension.

OBJECTIVE: Connective tissue disease (CTD)-associated pulmonary arterial hypertension (PAH) is the second most common etiology of PAH and carries a poor prognosis. Recently, it has been shown that female human tumor necrosis factor (TNF)-transgenic (Tg) mice die of cardiopulmonary disease by 6 months of age. This study was undertaken to characterize this pathophysiology and assess its potential as a novel model of CTD-PAH. METHODS: Histologic analysis was performed on TNF-Tg and wild-type (WT) mice to characterize pulmonary vascular and right ventricular (RV) pathology (n = 40 [4-5 mice per group per time point]). Mice underwent right-sided heart catheterization (n = 29) and micro-computed tomographic angiography (n = 8) to assess vascular disease. Bone marrow chimeric mice (n = 12), and anti-TNF-treated mice versus placebo-treated mice (n = 12), were assessed. RNA sequencing was performed on mouse lung tissue (n = 6). RESULTS: TNF-Tg mice displayed a pulmonary vasculopathy marked by collagen deposition (P < 0.001) and vascular occlusion (P < 0.001) with associated RV hypertrophy (P < 0.001) and severely increased RV systolic pressure (mean ± SD 75.1 ± 19.3 mm Hg versus 26.7 ± 1.7 mm Hg in WT animals; P < 0.0001). TNF-Tg mice had increased α-smooth muscle actin (α-SMA) staining, which corresponded to proliferation and loss of von Willebrand factor (vWF)-positive endothelial cells (P < 0.01). There was an increase in α-SMA-positive, vWF-positive cells (P < 0.01), implicating endothelial-mesenchymal transition. Bone marrow chimera experiments revealed that mesenchymal but not bone marrow-derived cells are necessary to drive this process. Treatment with anti-TNF therapy halted the progression of disease. This pathology closely mimics human CTD-PAH, in which patient lungs demonstrate increased TNF signaling and significant similarities in genomic pathway dysregulation. CONCLUSION: The TNF-Tg mouse represents a novel model of CTD-PAH, recapitulates key disease features, and can serve as a valuable tool for discovery and assessment of therapeutics.

iNOS dependent and independent phases of lymph node expansion in mice with TNF-induced inflammatory-erosive arthritis.

INTRODUCTION: A pivotal effect of lymphatic vessel (LV) function in joint homeostasis was identified in the tumor necrosis factor-transgenic (TNF-Tg) mouse model of rheumatoid arthritis (RA). Specifically, loss of LV contractions is associated with progressive synovitis and erosions. Furthermore, draining lymph node expansion is a biomarker of arthritic progression, and both macrophages and lymphatic endothelial cells express inducible nitric oxide synthase (iNOS), which disrupts LV contraction and transport of immune cells to the draining lymph nodes. Therefore, to directly assess these relationships, we tested the hypothesis that TNF-Tg mice with global genetic ablation of iNOS (iNOS-/-) will show delayed draining lymph node expansion, maintained LV contractions, and decreased synovitis and erosions. METHOD: iNOS-/-× TNF-Tg female and male mice, and control littermates (iNOS-/-, TNF-Tg, and WT), were examined with (1) ultrasound to determine popliteal lymph node (PLN) volume and (2) near-infrared imaging (NIR) to assess popliteal LV contraction frequency, and differences between genotypes were assessed at 3, 4, 5, and 6 months of age. Knees and PLN were harvested at 4 months in females and 6 months in males, to assess synovitis, bone erosions, and cellular accumulation in PLN sinuses via histology. RESULTS: Initially, an increase in PLN volume was observed for both female and male iNOS-/-× TNF-Tg and TNF-Tg compared to their WT and iNOS-/- counterparts at 2 and 3 months, respectively. Subsequently, TNF-Tg PLNs continue to increase in volume, while iNOS-/-× TNF-Tg did not increase in volume from the initial timepoints. WT and iNOS-/- PLN volume was unchanged throughout the experiment. LV contraction frequency was increased at 4 months in females and 5 months in males, in the iNOS-/-× TNF-Tg mice compared to the TNF-Tg. Synovitis and erosions were moderately reduced in iNOS-/-× TNF-Tg versus TNF-Tg knees in females, while no differences in knee pathology were observed in males. CONCLUSIONS: Genetic iNOS ablation maintains draining lymph node volume and LV function during TNF-induced inflammatory arthritis and is associated with moderately decreased joint inflammation and damage.