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Background-Pulmonary vascular injury by toxins can induce neointimal formation, pulmonary arterial hypertension (PAH), right ventricular failure, and death. We showed previously that simvastatin attenuates smooth muscle neointimal proliferation and pulmonary hypertension in pneumonectomized rats injected with the alkaloid toxin monocrotaline. The present study was undertaken to investigate the efficacy of simvastatin and its mechanism of reversing established neointimal vascular occlusion and pulmonary hypertension. Methods and Results-Pneumonectomized rats injected with monocrotaline at 4 weeks demonstrated severe PAH at 11 weeks (mean pulmonary artery pressure [mPAP]ϭ42 versus 17 mm Hg in normal rats) and death by 15 weeks. When rats with severe PAH received simvastatin (2 mg · kg Ϫ1 · d Ϫ1 by gavage) from week 11, there was 100% survival and reversal of PAH after 2 weeks (mPAPϭ36 mm Hg) and 6 weeks (mPAPϭ24 mm Hg) of therapy. Simvastatin treatment reduced right ventricular hypertrophy and reduced proliferation and increased apoptosis of pathological smooth muscle cells in the neointima and medial walls of pulmonary arteries. Longitudinal transcriptional profiling revealed that simvastatin downregulated the inflammatory genes fos, jun, and tumor necrosis factor-␣ and upregulated the cell cycle inhibitor p27Kip1, endothelial nitric oxide synthase, and bone morphogenetic protein receptor type 1a. Conclusions-Simvastatin reverses pulmonary arterial neointimal formation and PAH after toxic injury. (Circulation.

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Activation of T cells induces the production of T cell growth and survival factor interleukin (IL) 2. Regulatory T cells intrinsically fail to induce IL-2 expression upon activation and can suppress IL-2 production in conventional T cells. Thus, the control of IL-2 expression is critically important to T cell immune responses, yet the mechanisms remain incompletely understood. Nuclear factor (NF) 90 is a zinc-finger DNA- and double-stranded RNA-binding protein subunit that binds specifically to the antigen receptor response element (ARRE)/NF of activated T cells target sequence in the IL-2 proximal promoter. Inducible binding of NF90 to the IL-2 promoter in vivo is shown by chromatin immunoprecipitation. NF90 gene-targeted mice exhibit perinatal lethality. Compared with newborn NF90+/+ mice, newborn NF90−/− mice demonstrate severe impairment of IL-2 expression. Compared with wild-type cells, T cells deficient in NF90 are impaired in ARRE and IL-2 transcriptional activation and IL-2 mRNA stabilization. Fetal liver cells from NF90 gene-targeted mice were transplanted into irradiated adult recombination activating gene (RAG)–2−/− and IL-2Rγ−/− mice deficient in T cells, B cells, and natural killer cells. NF90+/+- and NF90−/−-RAG chimeric mice showed grossly normal repopulation of the thymus and spleen, but only NF90−/− T cells were severely impaired in IL-2 gene expression. Compared with littermates, NF90−/− RAG chimeric mice exhibited profound T cell lymphocytopenia in the peripheral circulation. Thus, NF90 regulates inducible IL-2 transcription, mRNA stability, and gene expression in T cells and represents a novel therapeutic target for the modulation of T cell immune responses.

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Experimental models of pulmonary hypertension that have used combinations of chronic hypoxia, pneumonectomy, and endothelial injury by monocrotaline or the vascular endothelial growth factor receptor antagonist SUGEN 5416 have been reviewed. 9 Whereas models based on hypoxia demonstrateBackground-Pulmonary endothelial injury triggers a reparative program, which in susceptible individuals is characterized by neointima formation, vascular narrowing, and the development of pulmonary arterial hypertension. The neointimal cells in human pathological plexiform lesions frequently coexpress smooth muscle α-actin and the endothelial von Willebrand antigen, creating a question about their cellular lineage of origin. Methods and Results-Experimental pulmonary hypertension with neointima formation develops in C57Bl/6 mice subjected to left pneumonectomy followed 1 week later by jugular vein injection of monocrotaline pyrrole (20 μg/μL and 1 μL/g; group P/MCTP).

IL-2 gene expression in activated T-cells is initiated by chromatin remodeling at the IL-2 proximal promoter and conversion of a transcriptional repressor into a potent transcriptional activator. A purine-box regulator complex was purified from activated Jurkat T-cell nuclei based on sequence-specific DNA binding to the antigen receptor response element (ARRE)/nuclear factor of activated T-cells (NF-AT) target DNA sequence in the proximal IL-2 promoter. ARRE DNA-binding subunits were identified as NF90, NF45 and systemic lupus erythematosis autoantigens, Ku80 and Ku70. Monoclonal antibodies to Ku80, Ku70 and NF90 specifically inhibit constitutive and inducible ARRE DNA-binding activity in Jurkat T-cells. Ku80, Ku70 and NF90 bind specifically to the IL-2 gene promoter in vivo, as demonstrated by chromatin immunoprecipitation. Activation of Jurkat T-cells and mouse primary spleen cells induces binding of Ku80 and NF90 to the IL-2 promoter in vivo, and decreases binding of Ku70 to the IL-2 promoter in vivo, and these dynamic changes are inhibited by immunosuppressants cyclosporin A and triptolide. Dynamic changes in binding of Ku80, Ku70 and NF90 to the IL-2 proximal promoter in vivo correlate with chromatin remodeling and transcriptional initiation in activated T-cells.

Triptolide (PG490, 97% pure) is a diterpenoid triepoxide with potent anti-inflammatory and immunosuppressive effects in transformed human bronchial epithelial cells and T cells (Qiu D, Zhao G, Aoki Y, Shi L, Uyei A, Nazarian S, Ng JC-H, and Kao PN. J Biol Chem 274: 13443-13450, 1999). Triptolide, with an IC(50) of approximately 20-50 ng/ml, inhibits normal and transformed human bronchial epithelial cell expression of interleukin (IL)-6 and IL-8 stimulated by phorbol 12-myristate 13-acetate (PMA), tumor necrosis factor-alpha, or IL-1 beta. Nuclear runoff and luciferase reporter gene assays demonstrate that triptolide inhibits IL-8 transcription. Triptolide also inhibits the transcriptional activation, but not the DNA binding, of nuclear factor-kappa B. A cDNA array and clustering algorithm analysis reveals that triptolide inhibits expression of the PMA-induced genes tumor necrosis factor-alpha, IL-8, macrophage inflammatory protein-2 alpha, intercellular adhesion molecule-1, integrin beta(6), vascular endothelial growth factor, granulocyte-macrophage colony-stimulating factor, GATA-3, fra-1, and NF45. Triptolide also inhibits constitutively expressed cell cycle regulators and survival genes cyclins D1, B1, and A1, cdc-25, bcl-x, and c-jun. Thus anti-inflammatory, antiproliferative, and proapoptotic properties of triptolide are associated with inhibition of nuclear factor-kappa B signaling and inhibition of genes known to regulate cell cycle progression and survival.