Assessment of a novel nanoparticle hyperthermia therapy in a murine model of osteosarcoma.

OBJECTIVE: To evaluate the effects of nanoparticle hyperthermia therapy on monocyte function and tumor-derived factors associated with macrophage polarization in a murine osteosarcoma model. STUDY DESIGN: Experimental study. ANIMALS: Female C3H mice. METHODS: Peripheral blood monocyte cell surface phenotype, monocyte chemotaxis, tumor messenger RNA expression, and survival were compared among osteosarcoma (OS)-bearing mice treated with nanoparticle hyperthermia therapy, OS-bearing mice with osteomyelitis, OS-bearing mice, vehicle control mice, and normal control mice. RESULTS: OS-bearing mice with osteomyelitis had a higher proportion of "nonclassical" monocytes (Ly6Clo ) compared with all other experimental groups. There were alterations in monocyte expression of multiple chemokine receptors among experimental groups including CXCR2, CCR2, and CXCR4. Monocytes from OS-bearing mice treated with hyperthermia therapy exhibited greater chemotaxis compared with monocytes from OS-bearing mice with osteomyelitis. CONCLUSION: OS likely induced alterations in monocyte phenotype and function. Nanoparticle hyperthermia therapy increased in vitro monocyte chemotaxis. CLINICAL IMPACT: Enhancing monocyte/macrophage function in dogs with OS may enhance antitumor immunity.

CD4+CD25+ T regulatory cells activated during feline immunodeficiency virus infection convert T helper cells into functional suppressors through a membrane-bound TGFß / GARP-mediated mechanism.

BACKGROUND: We and others have previously reported that cell membrane-bound TGFß (mTGFß) on activated T regulatory (Treg) cells mediates suppressor function. Current findings suggest that a novel protein known as Glycoprotein A Repetitions Predominant (GARP) anchors mTGFß to the Treg cell surface and facilitates suppressor activity. Recently, we have described that GARP+TGFß+ Treg cells expand during the course of FIV infection. Because Treg cells are anergic and generally exhibit poor proliferative ability, we asked how Treg homeostasis is maintained during the course of feline immunodeficiency virus (FIV) infection. RESULTS: Here, we report that Treg cells from FIV+ cats express GARP and mTGFß and convert T helper (Th) cells into phenotypic and functional Treg cells. Th to Treg conversion was abrogated by anti-TGFß or anti-GARP treatment of Treg cells or by anti-TGFßRII treatment of Th cells, suggesting that Treg cell recruitment from the Th pool is mediated by TGFß/TGFßRII signaling and that cell-surface GARP plays a major role in this process. CONCLUSIONS: These findings suggest Th to Treg conversion may initiate a cascade of events that contributes to the maintenance of virus reservoirs, progressive Th cell immunosuppression, and the development of immunodeficiency, all of which are central to the pathogenesis of AIDS lentivirus infections.

Optimized adenovirus-antibody complexes stimulate strong cellular and humoral immune responses against an encoded antigen in naive mice and those with preexisting immunity.

The immune response to recombinant adenoviruses is the most significant impediment to their clinical use for immunization. We test the hypothesis that specific virus-antibody combinations dictate the type of immune response generated against the adenovirus and its transgene cassette under certain physiological conditions while minimizing vector-induced toxicity. In vitro and in vivo assays were used to characterize the transduction efficiency, the T and B cell responses to the encoded transgene, and the toxicity of 1 × 10(11) adenovirus particles mixed with different concentrations of neutralizing antibodies. Complexes formed at concentrations of 500 to 0.05 times the 50% neutralizing dose (ND(50)) elicited strong virus- and transgene-specific T cell responses. The 0.05-ND(50) formulation elicited measurable anti-transgene antibodies that were similar to those of virus alone (P = 0.07). This preparation also elicited very strong transgene-specific memory T cell responses (28.6 ± 5.2% proliferation versus 7.7 ± 1.4% for virus alone). Preexisting immunity significantly reduced all responses elicited by these formulations. Although lower concentrations (0.005 and 0.0005 ND(50)) of antibody did not improve cellular and humoral responses in naïve animals, they did promote strong cellular (0.005 ND(50)) and humoral (0.0005 ND(50)) responses in mice with preexisting immunity. Some virus-antibody complexes may improve the potency of adenovirus-based vaccines in naïve individuals, while others can sway the immune response in those with preexisting immunity. Additional studies with these and other virus-antibody ratios may be useful to predict and model the type of immune responses generated against a transgene in those with different levels of exposure to adenovirus.

Transforming growth factor-beta/transforming growth factor-betaRII signaling may regulate CD4+CD25+ T-regulatory cell homeostasis and suppressor function in feline AIDS lentivirus infection.

We have reported that CD4+CD25+ T-regulatory (Treg) cells are fully activated for suppressor function in feline immunodeficiency virus (FIV)-infected cats. Studies have suggested that surface transforming growth factor-beta (TGFbeta; membrane TGFbeta [mTGFbeta]) is a feature of activated CD4+CD25+ Treg cells and may play a role in Treg homeostasis and suppressor function. Herein, we explore the role of TGFbeta in feline Treg homeostasis and suppressor function and what effect FIV infection of cats might have on these processes. Stimulation of CD4+CD25- T helper (Th) cells with Concanavalin A (ConA) plus TGFbeta converts them to Treg-like cells capable of suppressor function. Reverse-transcription polymerase chain reaction and flow cytometry revealed that these ConA/TGFbeta-converted Treg cells upregulate Foxp3 and mTGFbeta. ConA stimulation of CD4+CD25- T cells upregulates TGFbeta receptor II (RII), and pretreatment of these cells with anti-TGFbeta-RII antibodies blocks the TGFbeta-induced conversion to Treg cells. Pretreatment of ConA/TGFbeta-converted Treg cells with anti-TGFbeta antibodies also abrogates their suppressor function, suggesting that Treg homeostasis and suppressor function may be mediated by mTGFbeta. Finally, we show that treatment of CD4+CD25+ mTGFbeta-positive Treg cells from FIV-infected cats with anti-TGFbeta antibodies or treatment of ConA-stimulated CD4+CD25- Th target cells with anti-TGFbeta-RII antibodies diminishes suppressor function. These data suggest that the recruitment of Treg cells from the Th pool and suppressor function of Treg cells are dependent on the TGFbeta/TGFbeta-RII signaling pathway and that this pathway is constitutively upregulated in asymptomatic chronically FIV-infected cats.