Low dose irinotecan improves advanced lupus nephritis in mice potentially by changing DNA relaxation and anti–double-stranded DNA binding

Objective Albeit clear advances in the treatment of SLE, many patients still present with refractory lupus nephritis requiring new treatment strategies for this disease. Here we determined whether reduced doses of the topoisomerase I inhibitor irinotecan, which is known as chemotherapeutic agent, were able to suppress SLE in NZB/W F1 mice. We further evaluated the potential mechanism how irinotecan influenced the course of SLE. Methods NZB/W F1 mice were treated with low dose irinotecan either from week 24 of age or from established glomerulonephritis defined by a proteinuria ≥grade 3+. Binding of anti-dsDNA antibodies was measured by ELISA; and DNA relaxation was visualized by gel electrophoresis. Results Significantly reduced irinotecan dosages improved lupus nephritis and prolonged survival in NZB/W F1 mice. The lowest dose successfully used for the treatment of established murine lupus nephritis was more than 50 times lower than the dose usually applied for chemotherapy in humans. As a mechanism, low dose irinotecan reduced B cell activity; however, the levels of B cell activity in irinotecan-treated mice were similar to those in Balb/c mice of the same age suggesting that irinotecan did not induce a clear immunosuppression. In addition, incubation of double-stranded (ds) DNA with topoisomerase I increased binding of murine and human anti-dsDNA antibodies showing for the first time that relaxed DNA is more susceptible to anti-dsDNA antibody binding. This effect was reversed by addition of the topoisomerase I inhibitor camptothecin. Conclusion Our results propose topoisomerase I inhibitors as a novel and targeted therapy for SLE. © 2014 American College of Rheumatology.

Inhibition of matrix metalloproteinases attenuates brain damage in experimental meningococcal meningitis.

BackgroundApproximately 7% of survivors from meningococcal meningitis (MM) suffer from neurological sequelae due to brain damage in the course of meningitis. The present study focuses on the role of matrix metalloproteinases (MMPs) in a novel mouse model of MM-induced brain damage.MethodsThe model is based on intracisternal infection of BALB/c mice with a serogroup C Neisseria meningitidis strain. Mice were infected with meningococci and randomised for treatment with the MMP inhibitor batimastat (BB-94) or vehicle. Animal survival, brain injury and host-response biomarkers were assessed 48 h after meningococcal challenge.ResultsMice that received BB-94 presented significantly diminished MMP-9 levels (p¿<¿0.01), intracerebral bleeding (p¿<¿0.01), and blood-brain barrier (BBB) breakdown (p¿<¿0.05) in comparison with untreated animals. In mice suffering from MM, the amount of MMP-9 measured by zymography significantly correlated with both intracerebral haemorrhage (p¿<¿0.01) and BBB disruption (p¿<¿0.05).ConclusionsMMPs significantly contribute to brain damage associated with experimental MM. Inhibition of MMPs reduces intracranial complications in mice suffering from MM, representing a potential adjuvant strategy in MM post-infection sequelae.

Persistent and compartmentalised disruption of dendritic cell subpopulations in the lung following influenza A virus infection.

Immunological homeostasis in the respiratory tract is thought to require balanced interactions between networks of dendritic cell (DC) subsets in lung microenvironments in order to regulate tolerance or immunity to inhaled antigens and pathogens. Influenza A virus (IAV) poses a serious threat of long-term disruption to this balance through its potent pro-inflammatory activities. In this study, we have used a BALB/c mouse model of A/PR8/34 H1N1 Influenza Type A Virus infection to examine the effects of IAV on respiratory tissue DC subsets during the recovery phase following clearance of the virus. In adult mice, we found differences in the kinetics and activation states of DC residing in the airway mucosa (AMDC) compared to those in the parenchymal lung (PLDC) compartments. A significant depletion in the percentage of AMDC was observed at day 4 post-infection that was associated with a change in steady-state CD11b+ and CD11b- AMDC subset frequencies and significantly elevated CD40 and CD80 expression and that returned to baseline by day 14 post-infection. In contrast, percentages and total numbers of PLDC were significantly elevated at day 14 and remained so until day 21 post-infection. Accompanying this was a change in CD11b+and CD11b- PLDC subset frequencies and significant increase in CD40 and CD80 expression at these time points. Furthermore, mice infected with IAV at 4 weeks of age showed a significant increase in total numbers of PLDC, and increased CD40 expression on both AMDC and PLDC, when analysed as adults 35 days later. These data suggest that the rate of recovery of DC populations following IAV infection differs in the mucosal and parenchymal compartments of the lung and that DC populations can remain disrupted and activated for a prolonged period following viral clearance, into adulthood if infection occurred early in life.

Fluorescence lifetime imaging of the ocular fundus in mice

PURPOSE Fundus autofluorescence (AF) is characterized not only by its intensity or excitation and emission spectra but also by the lifetimes of the fluorophores. Fluorescence lifetime is influenced by the fluorophore's microenvironment and may provide information about the metabolic tissue state. We report quantitative and qualitative autofluorescence lifetime imaging of the ocular fundus in mice. METHODS A fluorescence lifetime imaging ophthalmoscope (FLIO) was used to measure fluorescence lifetimes of endogenous fluorophores in the murine retina. FLIO imaging was performed in 1-month-old C57BL/6, BALB/c, and C3A.Cg-Pde6b(+)Prph2(Rd2)/J mice. Measurements were repeated at monthly intervals over the course of 6 months. For correlation with structural changes, an optical coherence tomogram was acquired. RESULTS Fundus autofluorescence lifetime images were readily obtained in all mice. In the short spectral channel (498-560 nm), mean ± SEM AF lifetimes were 956 ± 15 picoseconds (ps) in C57BL/6; 801 ± 35 ps in BALB/c mice; and 882 ± 37 ps in C3A.Cg-Pde6b(+)Prph2(Rd2)/J mice. In the long spectral channel (560-720 nm), mean ± SEM AF lifetimes were 298 ± 14 ps in C57BL/6 mice, 241 ± 10 ps in BALB/c mice, and 288 ± 8 ps in C3A.Cg-Pde6b(+)Prph2(Rd2)/J mice. There was a general decrease in mean AF lifetimes with age. CONCLUSIONS Although fluorescence lifetime values differ among mouse strains, we found little variance within the groups. Fundus autofluorescence lifetime imaging in mice may provide additional information for understanding retinal disease processes and may facilitate monitoring of therapeutic effects in preclinical studies.

A Lentiviral Vector Expressing Japanese Encephalitis Virus-like Particles Elicits Broad Neutralizing Antibody Response in Pigs.

BACKGROUND Japanese encephalitis virus (JEV) is the major cause of viral encephalitis in Southeast Asia. Vaccination of domestic pigs has been suggested as a "one health" strategy to reduce viral disease transmission to humans. The efficiency of two lentiviral TRIP/JEV vectors expressing the JEV envelope prM and E glycoproteins at eliciting protective humoral response was assessed in a mouse model and piglets. METHODOLOGY/PRINCIPAL FINDINGS A gene encoding the envelope proteins prM and E from a genotype 3 JEV strain was inserted into a lentiviral TRIP vector. Two lentiviral vectors TRIP/JEV were generated, each expressing the prM signal peptide followed by the prM protein and the E glycoprotein, the latter being expressed either in its native form or lacking its two C-terminal transmembrane domains. In vitro transduction of cells with the TRIP/JEV vector expressing the native prM and E resulted in the efficient secretion of virus-like particles of Japanese encephalitis virus. Immunization of BALB/c mice with TRIP/JEV vectors resulted in the production of IgGs against Japanese encephalitis virus, and the injection of a second dose one month after the prime injection greatly boosted antibody titers. The TRIP/JEV vectors elicited neutralizing antibodies against JEV strains belonging to genotypes 1, 3, and 5. Immunization of piglets with two doses of the lentiviral vector expressing JEV virus-like particles led to high titers of anti-JEV antibodies, that had efficient neutralizing activity regardless of the JEV genotype tested. CONCLUSIONS/SIGNIFICANCE Immunization of pigs with the lentiviral vector expressing JEV virus-like particles is particularly efficient to prime antigen-specific humoral immunity and trigger neutralizing antibody responses against JEV genotypes 1, 3, and 5. The titers of neutralizing antibodies elicited by the TRIP/JEV vector are sufficient to confer protection in domestic pigs against different genotypes of JEV and this could be of a great utility in endemic regions where more than one genotype is circulating.

In Vitro and In Vivo Effects of the Bumped Kinase Inhibitor 1294 in the Related Cyst-Forming Apicomplexans Toxoplasma gondii and Neospora caninum.

We report on the in vitro effects of the bumped kinase inhibitor 1294 (BKI-1294) in cultures of virulent Neospora caninum isolates Nc-Liverpool (Nc-Liv) and Nc-Spain7 and in two strains of Toxoplasma gondii (RH and ME49), all grown in human foreskin fibroblasts. In these parasites, BKI-1294 acted with 50% inhibitory concentrations (IC50s) ranging from 20 nM (T. gondii RH) to 360 nM (N. caninum Nc-Liv), and exposure of intracellular stages to 1294 led to the nondisjunction of newly formed tachyzoites, resulting in the formation of multinucleated complexes similar to complexes previously observed in BKI-1294-treated N. caninum beta-galactosidase-expressing parasites. However, such complexes were not seen in a transgenic T. gondii strain that expressed CDPK1 harboring a mutation (G to M) in the gatekeeper residue. In T. gondii ME49 and N. caninum Nc-Liv, exposure of cultures to BKI-1294 resulted in the elevated expression of mRNA coding for the bradyzoite marker BAG1. Unlike in bradyzoites, SAG1 expression was not repressed. Immunofluorescence also showed that these multinucleated complexes expressed SAG1 and BAG1 and the monoclonal antibody CC2, which binds to a yet unidentified bradyzoite antigen, also exhibited increased labeling. In a pregnant mouse model, BKI-1294 efficiently inhibited vertical transmission in BALB/c mice experimentally infected with one of the two virulent isolates Nc-Liv or Nc-Spain7, demonstrating proof of concept that this compound protected offspring from vertical transmission and disease. The observed deregulated antigen expression effect may enhance the immune response during BKI-1294 therapy and will be the subject of future studies.

Dose-dependent effects of experimental infection with the virulent Neospora caninum Nc-Spain7 isolate in a pregnant mouse model.

Pregnant BALB/c mice have been widely used as an in vivo model to study Neospora caninum infection biology and to provide proof-of-concept for assessments of drugs and vaccines against neosporosis. The fact that this model has been used with different isolates of variable virulence, varying infection routes and differing methods to prepare the parasites for infection, has rendered the comparison of results from different laboratories impossible. In most studies, mice were infected with similar number of parasites (2 × 10(6)) as employed in ruminant models (10(7) for cows and 10(6) for sheep), which seems inappropriate considering the enormous differences in the weight of these species. Thus, for achieving meaningful results in vaccination and drug efficacy experiments, a refinement and standardization of this experimental model is necessary. Thus, 2 × 10(6), 10(5), 10(4), 10(3) and 10(2) tachyzoites of the highly virulent and well-characterised Nc-Spain7 isolate were subcutaneously inoculated into mice at day 7 of pregnancy, and clinical outcome, vertical transmission, parasite burden and antibody responses were compared. Dams from all infected groups presented nervous signs and the percentage of surviving pups at day 30 postpartum was surprisingly low (24%) in mice infected with only 10(2) tachyzoites. Importantly, infection with 10(5) tachyzoites resulted in antibody levels, cerebral parasite burden in dams and 100% mortality rate in pups, which was identical to infection with 2 × 10(6) tachyzoites. Considering these results, it is reasonable to lower the challenge dose to 10(5) tachyzoites in further experiments when assessing drugs or vaccine candidates.

IN VITRO INDUCTION OF XENOIMMUNE RESPONSES TO LIPOSOMES CONTAINING HUMAN COLON TUMOR ANTIGENS

Liposomes prepared with human LS174T colon tumor cell membranes induce specific primary and secondary xenogeneic immune responses in BALB/c splenocytes in vitro. The multilamellar vesicular liposomes were prepared by adding sonicated membrane fragments in 8 mM CaCl(,2) to a dried lipid film. Cytoxic splenocytes generated in vivo exhibited specificity for the LS174T cell; liposomes elicited higher levels of cytotoxicity than did membranes (P < 0.01). Secondary blastogenic responses elicited in in vivo-primed spleen cells by liposomes also produced a significantly greater (P < 0.005) response than membranes. Subsequently, in vitro induction of primary blastogenic and cytotoxic responses by liposomes were accomplished and revealed similar kinetics to that of whole LS174T cell immunogens. Specificity of the in vitro-primed spleen cells was clearly demonstrated (P < 0.01) on a variety of human tumor cells using both the primed lymphocyte and cell-mediated cytotoxicity assays. The results of competitive inhibition tests with autologous lymphoblasts demonstrated that 30% of the cytotoxic activity was directed against lymphocyte antigens.^ The adjuvant effect of liposomes was shown to be mediated primarily by tumor antigens exposed on the outer surface of liposomes. Trypsinization of the liposomes which eliminated 96% of the surface protein reduced the ability of liposomes to induce cytotoxic splenocytes. The generation of cytolytic activity with liposomes of increasing protein concentration showed that while 10 (mu)g protein was threshold, 100 (mu)g protein induced maximal responses. In addition, membrane fluidity studies revealed that rigid liposomes were significantly (P < 0.05) more efficacious than fluid liposomes in inducing cytotoxicity.^ The induction of the primary response required the presence of nonadherent splenocytes bearing the Thy-1, Lyt-1, and Lyt-2 surface markers. The role of a Lyt-123 subpopulation was suggested by the inability of both the Lyt-1 and Lyt-2 depleted populations to completely restore the cytolytic levels to normal. In addition, the interaction of I-A('+) spleen adherent cells with liposomes for at least 8 hours was required to generate maximal cytotoxic activity. The phenotype of the cytotoxic effector was Thy-1('+), Lyt-2('+), and I-A('d-).^ Incorporation of tumor antigens into liposomes has thus enabled primary immunization in vitro to human colon cancer antigens and may afford an adaptable means to evaluate and to select specific immune responses, as well as to identify colon tumor-specific determinants.^

Characterization of recombinant class I MHC alloantigen action upon the survival of disparate heart allografts

Previous studies have led to the development of allochimeric class I MHC proteins as agents that effectively induce donor-specific transplantation tolerance in a rat system with or without additional immunosuppression. Within the α1-helical region of RT1.Au, an epitope that conferred immunologic tolerance was discovered. Studies presented herein were designed to test our central hypothesis that allochimeric proteins onfer tolerance in a mouse model. To test this hypothesis, portal vein (PV) injection of wild-type H2Kd and H2Dd proteins were produced in a bacterial expression system and found to specifically prolong the survival of BALB/c (H2d) heart allografts in C57BL/10 (H2b) recipients. Although a single PV injection of 50 μg α1–α 3 H2Kd alone was ineffective, 50 μg α1 –α3 alone slightly prolonged BALB/c heart allograft survivals. In contrast, the combination of 25 μg α1–α 3 H2Kd and 25 μg α1–α 3 H2Dd proteins prolonged BALB/c graft survivals to 20.2 ± 6.4 days (p < 0.004). The effect was donor-specific, since a combination of 25 μg α1–α3 H2Kd and 25 μg α1–α3 H2Dd proteins failed to affect survivals of third-party C3H (H2k k) heart allografts, namely 9.0 ± 0.0 days in treated versus 7.8 ± 0.5 days in untreated hosts. Thus, the combination of two H2K d and H2Dd proteins is more effective in prolonging allograft survival than a single protein produced in a bacterial expression system. A single PV injection (day 0) of 25 μg α1–α 2 H2Kd and 25 μg α1–α 2 H2Dd proteins to C57BL/10 mice prolonged the survival of BALB/c heart allografts to 22.4 ± 4.5 days. Within a WF to ACI rat heart allograft system, a single PV injection of 20 μg 70–77 u-RT1.Aa induced specific tolerance of allografts. This therapy could be combined with CsA to induce transplantation tolerance. However, combination of 70–77u-RT1.Aa with CTLA4Ig, rapamycin, or AG-490 effectively blocked the induction of transplantation tolerance. Tolerance generated by allochimeric protein could be adoptively transferred to naive recipients. Intragraft cytokine mRNA levels showed a bias towards a Th2-type phenotype. Additionally, studies of cytokine signaling and activation of transcription factors revealed a requirement that these pathways remain available for signaling in order for transplantation tolerance to occur. These studies suggest that the generation of regulatory cells are required for the induction of transplantation tolerance through the use of allochimeric proteins. ^

The role of interleukin-6 in the angiogenesis of ovarian carcinoma

The purpose of this study was to characterize the effects of IL-6 on endothelial cells and to investigate the role of IL-6 in the angiogenesis of ovarian carcinomas. We evaluated human ovarian carcinoma clinical specimens and determined that high expression of IL-6 was associated with increased tumor vascularization. Additionally, endothelial cells derived from the ovary and mesentery expressed the IL-6 receptor (IL-6R), and their stimulation with the exogenous ligand activated downstream signaling molecules and enhanced cell migration. Dual immunohistochemical staining for CD-31 and IL-6R revealed IL-6R expression on human endothelial cells within normal ovary and ovarian carcinomas. To further investigate the possible proangiogenic function of IL-6, Gelfoam sponges containing IL-6 or bFGF were implanted into the subcutis of BALB/c mice. IL-6 containing sponges were vascularized to the same extent as bFGF containing sponges. ^ Chronic stress can adversely affect disease progression. Stimulation of ovarian carcinoma cell lines with concentrations of catecholamines achieved in individuals experiencing chronic stress resulted in a substantial increase in IL-6 production. It was determined that stress mediators regulate IL-6 expression through the β-adrenergic receptor and Src. These data illustrate one mechanism by which chronic stress may influence tumor progression. ^ To investigate whether IL-6 contributes to the angiogenesis of ovarian carcinomas, we isolated low IL-6 expressing clones from the SKOV3.ip1 cell line and transfected them with a plasmid encoding the IL-6 gene. We observed no difference in tumor weight between high and low IL-6 expressing cells. However, while low IL-6 expressing tumors were highly vascularized, high IL-6 expressing tumors appeared hypervascularized. Immunohistochemical analysis revealed that all tumors exhibited robust expression of additional proangiogenic molecules. ^ Collectively, these studies indicate that IL-6 secreted by ovarian cancer cells is a highly proangiogenic cytokine. However, IL-6 is but one of several proangiogenic molecules produced by ovarian cancer, and its inhibition may not be sufficient to inhibit angiogenesis of ovarian carcinoma. The findings presented in this dissertation provide insight into the function of IL-6 as a regulator of angiogenesis. Understanding of the role of proangiogenic molecules such as IL-6 in ovarian carcinoma may have important implications for therapy directed at the vascular component of this disease. ^

CIRCADIAN RHYTHM IN THE BIOAVAILABILITY OF ORALLY ADMINISTERED THEOPHYLLINE

The effect of time (i.e., biologic time structure) of drug administration on the bioavailability of theophylline was investigated in man after both a single dosage as well as after repeated, or chronic, drug administrations. Preliminary laboratory investigations on Balb-C mice showed the toxic

Boosting with recombinant vaccinia increases immunogenicity and protective efficacy of malaria DNA vaccine

To enhance the efficacy of DNA malaria vaccines, we evaluated the effect on protection of immunizing with various combinations of DNA, recombinant vaccinia virus, and a synthetic peptide. Immunization of BALB/c mice with a plasmid expressing Plasmodium yoelii (Py) circumsporozoite protein (CSP) induces H-2Kd-restricted CD8+ cytotoxic T lymphocyte (CTL) responses and CD8+ T cell- and interferon (IFN)-γ-dependent protection of mice against challenge with Py sporozoites. Immunization with a multiple antigenic peptide, including the only reported H-2Kd-restricted CD8+ T cell epitope on the PyCSP (PyCSP CTL multiple antigenic peptide) and immunization with recombinant vaccinia expressing the PyCSP induced CTL but only modest to minimal protection. Mice were immunized with PyCSP DNA, PyCSP CTL multiple antigenic peptide, or recombinant vaccinia expressing PyCSP, were boosted 9 wk later with the same immunogen or one of the others, and were challenged. Only mice immunized with DNA and boosted with vaccinia PyCSP (D-V) (11/16: 69%) or DNA (D-D) (7/16: 44%) had greater protection (P < 0.0007) than controls. D-V mice had significantly higher individual levels of antibodies and class I-restricted CTL activity than did D-D mice; IFN-γ production by ELIspot also was higher in D-V than in D-D mice. In a second experiment, three different groups of D-V mice each had higher levels of protection than did D-D mice, and IFN-γ production was significantly greater in D-V than in D-D mice. The observation that priming with PyCSP DNA and boosting with vaccinia-PyCSP is more immunogenic and protective than immunizing with PyCSP DNA alone supports consideration of a similar sequential immunization approach in humans.

The crucial role of polymorphonuclear leukocytes in resistance to Salmonella dublin infections in genetically susceptible and resistant mice

Macrophages are considered to be the mediators of resistance to extra-intestinal Salmonella infections. Nevertheless, the initial cellular response to Salmonella infections consists primarily of polymorphonuclear leukocytes (PMN). To determine whether PMN serve an important function for the infected host, we made mice neutropenic with the rat mAb to RB6–8C5 and infected them i.v. with ≈103 Salmonella dublin or an isogenic derivative that lacks the virulence plasmid (LD842). We infected BALB/c mice, which have a point mutation in the macrophage-expressed gene Nramp1 that makes them susceptible to Salmonella, and BALB/c.D2 congenic mice, which have the wild-type Nramp1 gene that makes them resistant to Salmonella. Both mouse strains were resistant to LD842, and neutropenia made only the BALB/c strain susceptible to this infection. Neutropenic congenic mice, however, were susceptible only to wild-type S. dublin (plasmid+). These results show a complex interplay between plasmid-virulence genes in Salmonella, host macrophages, and PMN. Mice with normal macrophages need PMN to defend against nontyphoid Salmonella that carry a virulence plasmid but not against Salmonella without virulence plasmids. Mice with a mutant Nramp1 gene need PMN to defend against all Salmonella, even those that lack virulence plasmids. These results, plus the evidence that PMN kill Salmonella efficiently in vitro, suggest that Salmonella have adapted to grow inside macrophages where they are relatively sheltered from PMN. The adaptations that allow Salmonella to survive in macrophages do not protect them from PMN.

Collagen-binding growth factors: Production and characterization of functional fusion proteins having a collagen-binding domain

The autocrine/paracrine peptide signaling molecules such as growth factors have many promising biologic activities for clinical applications. However, one cannot expect specific therapeutic effects of the factors administered by ordinary drug delivery systems as they have limited target specificity and short half-lives in vivo. To overcome the difficulties in using growth factors as therapeutic agents, we have produced fusion proteins consisting of growth factor moieties and a collagen-binding domain (CBD) derived from Clostridium histolyticum collagenase. The fusion proteins carrying the epidermal growth factor (EGF) or basic fibroblast growth factor (bFGF) at the N terminal of CBD (CBEGF/CBFGF) tightly bound to insoluble collagen and stimulated the growth of BALB/c 3T3 fibroblasts as much as the unfused counterparts. CBEGF, when injected subcutaneously into nude mice, remained at the sites of injection for up to 10 days, whereas EGF was not detectable 24 h after injection. Although CBEGF did not exert a growth-promoting effect in vivo, CBFGF, but not bFGF, strongly stimulated the DNA synthesis in stromal cells at 5 days and 7 days after injection. These results indicate that CBD may be used as an anchoring unit to produce fusion proteins nondiffusible and long-lasting in vivo.

Gene therapy in allergic encephalomyelitis using myelin basic protein-specific T cells engineered to express latent transforming growth factor-β1

A myelin basic protein (MBP)-specific BALB/c T helper 1 (Th1) clone was transduced with cDNA for murine latent transforming growth factor-β1 (TGF-β1) by coculture with fibroblasts producing a genetically engineered retrovirus. When SJL x BALB/c F1 mice, immunized 12–15 days earlier with proteolipid protein in complete Freund’s adjuvant, were injected with 3 × 106 cells from MBP-activated untransduced cloned Th1 cells, the severity of experimental allergic encephalomyelitis (EAE) was slightly increased. In contrast, MBP-activated (but not resting) latent TGF-β1-transduced T cells significantly delayed and ameliorated EAE development. This protective effect was negated by simultaneously injected anti-TGF-β1. The transduced cells secreted 2–4 ng/ml of latent TGF-β1 into their culture medium, whereas control cells secreted barely detectable amounts. mRNA profiles for tumor necrosis factor, lymphotoxin, and interferon-γ were similar before and after transduction; interleukin-4 and -10 were absent. TGF-β1-transduced and antigen-activated BALB/c Th1 clones, specific for hemocyanin or ovalbumin, did not ameliorate EAE. Spinal cords from mice, taken 12 days after receiving TGF-β1-transduced, antigen-activated cells, contained detectable amounts of TGF-β1 cDNA. We conclude that latent TGF-β1-transduced, self-reactive T cell clones may be useful in the therapy of autoimmune diseases.