Nanoparticles loaded with IL-2 and TGF-β promote transplantation tolerance to alloantigen.

We have previously reported that nanoparticles (NPs) loaded with IL-2 and TGF-β and targeted to T cells induced polyclonal T regulatory cells (Tregs) that protected mice from graft-versus-host disease (GvHD). Here, we evaluated whether administration of these NPs during alloantigen immunization could prevent allograft rejection by converting immunogenic responses to tolerogenic ones. Using C57BL/6 mice and BALB/c mice as either donors or recipients of allogeneic splenocytes, we found that treatment with the tolerogenic NPs in both strains of mice resulted in a marked inhibition of mixed lymphocyte reaction (MLR) to donor cell alloantigen but not to third-party control mouse cells after transfer of the allogeneic cells.

Nanoparticle-mediated Delivery of IL-2 To T Follicular Helper Cells Protects BDF1 Mice from Lupus-like Disease.

We recently reported that poly lactic-co-glycolic acid (PLGA) nanoparticles (NPs) loaded with interleukin (IL)-2 and targeted to T cells inhibited the development of lupus-like disease in BDF1 mice by inducing functional T regulatory cells (Tregs). Here we show that the protection from disease and the extended survival of BDF1 mice provided by IL-2-loaded NPs targeted to T cells is not only due to an induction of Tregs but also contributed by an inhibition of T follicular helper (T) cells. These results identify a dual protective activity of IL-2 in the control of lupus autoimmunity, namely the inhibition of effector T cells, in addition to the previously known induction of Tregs.

Metabolic and immunomodulatory control of type 1 diabetes via orally delivered bile-acid-polymer nanocarriers of insulin or rapamycin.

Oral formulations of insulin are typically designed to improve its intestinal absorption and increase its blood bioavailability. Here we show that polymerized ursodeoxycholic acid, selected from a panel of bile-acid polymers and formulated into nanoparticles for the oral delivery of insulin, restored blood-glucose levels in mice and pigs with established type 1 diabetes. The nanoparticles functioned as a protective insulin carrier and as a high-avidity bile-acid-receptor agonist, increased the intestinal absorption of insulin, polarized intestinal macrophages towards the M2 phenotype, and preferentially accumulated in the pancreas of the mice, binding to the islet-cell bile-acid membrane receptor TGR5 with high avidity and activating the secretion of glucagon-like peptide and of endogenous insulin.

Strategies to Use Nanoparticles to Generate CD4 and CD8 Regulatory T Cells for the Treatment of SLE and Other Autoimmune Diseases.

Autoimmune diseases are disorders of immune regulation where the mechanisms responsible for self-tolerance break down and pathologic T cells overcome the protective effects of T regulatory cells (Tregs) that normally control them. The result can be the initiation of chronic inflammatory diseases. Systemic lupus erythematosus (SLE) and other autoimmune diseases are generally treated with pharmacologic or biological agents that have broad suppressive effects.

Nanoparticles Engineered as Artificial Antigen-Presenting Cells Induce Human CD4 and CD8 Tregs That Are Functional in Humanized Mice.

Artificial antigen-presenting cells (aAPCs) are synthetic versions of naturally occurring antigen-presenting cells (APCs) that, similar to natural APCs, promote efficient T effector cell responses . This report describes a method to produce acellular tolerogenic aAPCs made of biodegradable poly lactic-co-glycolic acid (PLGA) nanoparticles (NPs) and encapsulating IL-2 and TGF-β for a paracrine release to T cells. We document that these aAPCs can induce both human CD4 and CD8 T cells to become FoxP3 T regulatory cells (Tregs).

Anti-CD2 Antibody-Coated Nanoparticles Containing IL-2 Induce NK Cells That Protect Lupus Mice a TGF-β-Dependent Mechanism.

We recently reported that the treatment with nanoparticles (NPs) loaded with tolerogenic cytokines suppressed the manifestations of lupus-like disease induced by the transfer of donor CD4 T cells from DBA/2 mice into (C57BL/6 × DBA/2)F (BDF1) mice. Although the protective effects were ascribed to the induction of adaptive CD4 and CD8 T regulatory cells, the results suggested that another population of immune cells could be involved. Here we report that NK cells critically contribute to the protection from lupus-like disease conferred by NPs to BDF1 mice, and that this effect is TGF-β-dependent.

Rebalancing Immune Homeostasis to Treat Autoimmune Diseases.

During homeostasis, interactions between tolerogenic dendritic cells (DCs), self-reactive T cells, and T regulatory cells (Tregs) contribute to maintaining mammalian immune tolerance. In response to infection, immunogenic DCs promote the generation of proinflammatory effector T cell subsets. When complex homeostatic mechanisms maintaining the balance between regulatory and effector functions become impaired, autoimmune diseases can develop.

Suppression of Murine Lupus by CD4+ and CD8+ Treg Cells Induced by T Cell-Targeted Nanoparticles Loaded With Interleukin-2 and Transforming Growth Factor β.

Objective: To develop a nanoparticle (NP) platform that can expand both CD4+ and CD8+ Treg cells in vivo for the suppression of autoimmune responses in systemic lupus erythematosus (SLE).

Methods: Poly(lactic-co-glycolic acid) (PLGA) NPs encapsulating interleukin-2 (IL-2) and transforming growth factor β (TGFβ) were coated with anti-CD2/CD4 antibodies and administered to mice with lupus-like disease induced by the transfer of DBA/2 T cells into (C57BL/6 × DBA/2)F (BDF1) mice. The peripheral frequency of Treg cells was monitored ex vivo by flow cytometry.

Paracrine co-delivery of TGF-β and IL-2 using CD4-targeted nanoparticles for induction and maintenance of regulatory T cells.

The cytokine milieu is critical for orchestration of lineage development towards effector T cell (Teff) or regulatory T cell (Treg) subsets implicated in the progression of cancer and autoimmune disease. Importantly, the fitness and survival of the Treg subset is dependent on the cytokines Interleukin-2 (IL-2) and transforming growth factor beta (TGF-β). The production of these cytokines is impaired in autoimmunity increasing the probability of Treg conversion to aggressive effector cells in a proinflammatory microenvironment.

Lack of coagulopathy after copperhead snakebites.

Study Objective: We determine the frequency and severity of abnormal laboratory measures of coagulation after suspected or known copperhead (Agkistrodon contortrix) envenomation.

Methods: We identified the charts of venomous snakebites in children presenting to St. Louis Children's Hospital over a period of time greater than 14 years and of all venomous snakebites in adults presenting to Barnes-Jewish Hospital over a period of time greater than 11 years.

Critical role of all-trans retinoic acid in stabilizing human natural regulatory T cells under inflammatory conditions.

Recent studies have demonstrated that thymus-derived naturally occurring CD4(+)Foxp3(+) regulatory T cells (Tregs) in human and mouse may be unstable and dysfunctional in the presence of proinflammatory cytokines. All-trans RA (atRA), the active derivative of vitamin A, has been shown to regulate Treg and T effector cell differentiation. We hypothesize atRA stabilizes human natural Tregs (nTregs) under inflammatory conditions.

Therapeutic polyclonal human CD8+ CD25+ Fox3+ TNFR2+ PD-L1+ regulatory cells induced ex-vivo.

We report that polyclonal CD8regs generated in one week ex-vivo with anti-CD3/28 beads and cytokines rapidly developed suppressive activity in vitro sustained by TGF-β. In immunodeficient mice, these CD8regs demonstrated a markedly protective, IL-10 dependent activity against a xeno-GVHD. They expressed IL-2Rα/β, Foxp3, TNFR2, and the negative co-stimulatory receptors CTLA-4, PD-1, PD-L1 and Tim-3.

Phenotypic and functional characteristic of a newly identified CD8+ Foxp3- CD103+ regulatory T cells.

TGF-β and Foxp3 expressions are crucial for the induction and functional activity of CD4(+)Foxp3(+) regulatory T (iTreg) cells. Here, we demonstrate that although TGF-β-primed CD8(+) cells display much lower Foxp3 expression, their suppressive capacity is equivalent to that of CD4(+) iTreg cells, and both Foxp3(-) and Foxp3(+) CD8+ subsets have suppressive activities in vitro and in vivo. CD8(+)Foxp3(-) iTreg cells produce little IFN-γ but almost no IL-2, and display a typical anergic phenotype.

Polyclonal CD4+Foxp3+ Treg cells induce TGFβ-dependent tolerogenic dendritic cells that suppress the murine lupus-like syndrome.

Interplay between Foxp3(+) regulatory T cells (Treg) and dendritic cells (DCs) maintains immunologic tolerance, but the effects of each cell on the other are not well understood. We report that polyclonal CD4(+)Foxp3(+) Treg cells induced ex vivo with transforming growth factor beta (TGFβ) (iTreg) suppress a lupus-like chronic graft-versus-host disease by preventing the expansion of immunogenic DCs and inducing protective DCs that generate additional recipient CD4(+)Foxp3(+) cells. The protective effects of the transferred iTreg cells required both interleukin (IL)-10 and TGFβ, but the tolerogenic effects of the iTreg on DCs, and the immunosuppressive effects of these DCs were exclusively TGFβ-dependent.

Emergency department patients with diabetes have better glycemic control when they have identifiable primary care providers.

Objectives: The objective was to determine if emergency department (ED) patients with diabetes mellitus (DM) who have primary care providers (PCPs) have better control of their DM than patients with no PCPs.

Methods: This was a prospective, cross-sectional, observation study at a large, adult, urban, academic ED with 85,000 annual visits. ED patients with a history of DM were eligible.

Antigen-specific transforming growth factor β-induced Treg cells, but not natural Treg cells, ameliorate autoimmune arthritis in mice by shifting the Th17/Treg cell balance from Th17 predominance to Treg cell predominance.

Objective: Transferred CD4+CD25+FoxP3+ Treg cells can prevent autoimmune disease, but generally fail to ameliorate established disease. This study was undertaken to compare the effects of antigen-specific Treg cells induced with interleukin-2 (IL-2) and transforming growth factor β (TGFβ) ex vivo (induced Treg [iTreg] cells) to the effects of equivalent expanded thymus-derived natural Treg (nTreg) cells on established collagen-induced arthritis (CIA).

Methods: CIA was induced in DBA/1 mice by immunization with type II collagen (CII), and before or shortly after immunization, mice were treated with iTreg or nTreg cells that were generated or expanded in vitro.

Characterization of protective human CD4CD25 FOXP3 regulatory T cells generated with IL-2, TGF-β and retinoic acid.

Background: Protective CD4+CD25+ regulatory T cells bearing the Forkhead Foxp3 transcription factor can now be divided into three subsets: Endogenous thymus-derived cells, those induced in the periphery, and another subset induced ex-vivo with pharmacological amounts of IL-2 and TGF-β. Unfortunately, endogenous CD4+CD25+ regulatory T cells are unstable and can be converted to effector cells by pro-inflammatory cytokines. Although protective Foxp3+CD4+CD25+ cells resistant to proinflammatory cytokines have been generated in mice, in humans this result has been elusive.

Role of SMAD and non-SMAD signals in the development of Th17 and regulatory T cells.

Whereas TGF-beta is essential for the development of peripherally induced Foxp3(+) regulatory T cells (iTreg cells) and Th17 cells, the intracellular signaling mechanism by which TGF-beta regulates development of both cell subsets is less understood. In this study, we report that neither Smad2 nor Smad3 gene deficiency abrogates TGF-beta-dependent iTreg induction by a deacetylase inhibitor trichostatin A in vivo, although the loss of the Smad2 or Smad3 gene partially reduces iTreg induction in vitro. Similarly, SMAD2 and SMAD3 have a redundant role in development of Th17 in vitro and in experimental autoimmune encephalomyelitis.

Identity of mysterious CD4+CD25-Foxp3+ cells in SLE.

Various abnormalities in CD4+CD25+ regulatory T cells (Tregs) in systemic lupus erythematosus (SLE) include increased Foxp3+ cells that are CD25 negative. Barring methodological technical factors, these cells could be atypical Tregs or activated non-Treg CD4+ cells that express Foxp3. Two groups have reached opposite conclusions that could possibly reflect the subjects studied.

Synergistic effect of TGF-beta superfamily members on the induction of Foxp3+ Treg.

TGF-beta plays an important role in the induction of Treg and maintenance of immunologic tolerance, but whether other members of TGF-beta superfamily act together or independently to achieve this effect is poorly understood. Although others have reported that the bone morphogenetic proteins (BMP) and TGF-beta have similar effects on the development of thymocytes and T cells, in this study, we report that members of the BMP family, BMP-2 and -4, are unable to induce non-regulatory T cells to become Foxp3+ Treg. Neutralization studies with Noggin have revealed that BMP-2/4 and the BMP receptor signaling pathway is not required for TGF-beta to induce naïve CD4+CD25- cells to express Foxp3; however, BMP-2/4 and TGF-beta have a synergistic effect on the induction of Foxp3+ Treg.

Regulatory T cells in systemic lupus erythematosus: past, present and future.

Regulatory/suppressor T cells (Tregs) maintain immunologic homeo-stasis and prevent autoimmunity. In this article, past studies and recent studies of Tregs in mouse models for lupus and of human systemic lupus erythematosus are reviewed concentrating on CD4+CD25+Foxp3+ Tregs. These cells consist of thymus-derived, natural Tregs and peripherally induced Tregs that are similar phenotypically and functionally.

Natural and TGF-beta-induced Foxp3(+)CD4(+) CD25(+) regulatory T cells are not mirror images of each other.

Foxp3(+) CD4(+) CD25(+) regulatory cell (Treg) subsets that maintain immunologic homeostasis have been considered to be a homogeneous population of naturally occurring, thymus-derived CD4(+)CD25(+) cells (nTregs). However, similar Foxp3+ Tregs can be induced from CD25(-) precursors in vivo, and ex vivo with interleukin 2 (IL-2) and transforming growth factor beta (TGF-beta) (iTregs). These two subsets differ in their principal antigen specificities and in the T-cell receptor signal strength and co-stimulatory requirements needed for their generation.