The generation of humanized BLT mice by the cotransplantation of human fetal thymus and liver tissues and CD34؉ fetal liver cells into nonobese diabetic/severe combined immunodeficiency mice allows for the long-term reconstitution of a functional human immune system, with human T cells, B cells, dendritic cells, and monocytes/ macrophages repopulating mouse tissues. Here, we show that humanized BLT mice sustained high-level disseminated human immunodeficiency virus (HIV) infection, resulting in CD4 ؉ T-cell depletion and generalized immune activation. Following infection, HIV-specific humoral responses were present in all mice by 3 months, and HIVspecific CD4؉ and CD8 ؉ T-cell responses were detected in the majority of mice tested after 9 weeks of infection. Despite robust HIV-specific responses, however, viral loads remained elevated in infected BLT mice, raising the possibility that these responses are dysfunctional. The increased T-cell expression of the negative costimulator PD-1 recently has been postulated to contribute to T-cell dysfunction in chronic HIV infection. As seen in human infection, both CD4 ؉ and CD8 ؉ T cells demonstrated increased PD-1 expression in HIV-infected BLT mice, and PD-1 levels in these cells correlated positively with viral load and inversely with CD4 ؉ cell levels. The ability of humanized BLT mice to generate both cellular and humoral immune responses to HIV will allow the further investigation of human HIV-specific immune responses in vivo and suggests that these mice are able to provide a platform to assess candidate HIV vaccines and other immunotherapeutic strategies.

Sphingosine 1-phosphate (S1P) is a key endogenous regulator of the response to lung injury, maintaining endothelial barrier integrity through interaction with one of its receptors, S1P 1 . The short-term administration of S1P or S1P 1 receptor agonists enhances endothelial monolayer barrier function in vitro, and attenuates injuryinduced vascular leak in the lung and other organ systems in vivo. Although S1P 1 agonists bind to and activate S1P 1 , several of these agents also induce receptor internalization and degradation, and may therefore act as functional antagonists of S1P 1 after extended exposure. Here we report on the effects of prolonged exposure to these agents in bleomycin-induced lung injury. We demonstrate that repeated administration of S1P 1 agonists dramatically worsened lung injury after bleomycin challenge, as manifested by increased vascular leak and mortality. Consistent with these results, prolonged exposure to S1P 1 agonists in vitro eliminated the ability of endothelial cell monolayers to respond appropriately to the barrier-protective effects of S1P, indicating a loss of normal S1P-S1P 1 signaling. As bleomycin-induced lung injury progressed, continued exposure to S1P 1 agonists also resulted in increased pulmonary fibrosis. These data indicate that S1P 1 agonists can act as functional antagonists of S1P 1 on endothelial cells in vivo, which should be considered in developing these agents as therapies for vascular leak syndromes. Our findings also support the hypothesis that vascular leak is an important component of the fibrogenic response to lung injury, and suggest that targeting the S1P-S1P 1 pathway may also be an effective therapeutic strategy for fibrotic lung diseases.

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