Targeting of intragraft reactive oxygen species by APP-103, a novel polymer product, mitigates ischemia/reperfusion injury and promotes the survival of renal transplants.

Inflammatory responses associated with ischemia/reperfusion injury (IRI) play a central role in alloimmunity and transplant outcomes. A key event driving these inflammatory responses is the burst of reactive oxygen species (ROS), with hydrogen peroxide (H2 O2 ) as the most abundant form that occurs as a result of surgical implantation of the donor organ. Here, we used a syngeneic rat renal transplant and IRI model to evaluate the therapeutic properties of APP-103, a polyoxalate-based copolymer molecule containing vanillyl alcohol (VA) that exhibits high sensitivity and specificity toward the production of H2 O2 . We show that APP-103 is safe, and that it effectively promotes kidney function following IRI and survival of renal transplants. APP-103 reduces tissue injury and IRI-associated inflammatory responses in models of both warm ischemia (kidney clamping) and prolonged cold ischemia (syngeneic renal transplant). Mechanistically, we demonstrate that APP-103 exerts protective effects by specifically targeting the production of ROS. Our data introduce APP-103 as a novel, nontoxic, and site-activating therapeutic approach that effectively ameliorates the consequences of IRI in solid organ transplantation.

Two unique human decidual macrophage populations.

Several important events occur at the maternal-fetal interface, including generation of maternal-fetal tolerance, remodeling of the uterine smooth muscle and its spiral arteries and glands, and placental construction. Fetal-derived extravillous trophoblasts come in direct contact with maternal decidual leukocytes. Macrophages represent ∼20% of the leukocytes at this interface. In this study, two distinct subsets of CD14(+) decidual macrophages (dMs) are found to be present in first-trimester decidual tissue, CD11c(HI) and CD11c(LO). Gene expression analysis by RNA microarray revealed that 379 probes were differentially expressed between these two populations. Analysis of the two subsets revealed several clusters of coregulated genes that suggest distinct functions for these subsets in tissue remodeling, growth, and development. CD11c(HI) dMs express genes associated with lipid metabolism and inflammation, whereas CD11c(LO) dMs express genes associated with extracellular matrix formation, muscle regulation, and tissue growth. The CD11c(HI) dMs also differ from CD11c(LO) dMs in their ability to process protein Ag and are likely to be the major APCs in the decidua. Moreover, these populations each secrete both proinflammatory and anti-inflammatory cytokines that may contribute to the balance that establishes fetal-maternal tolerance. Thus, they do not fit the conventional M1/M2 categorization.

HLA-G homodimer-induced cytokine secretion through HLA-G receptors on human decidual macrophages and natural killer cells.

Human decidual CD14(+) macrophages and CD56(+) NK cells were isolated from material obtained after first-trimester pregnancy terminations. Each cell type expressed a specific surface receptor for histocompatibility leukocyte antigen (HLA)-G (an MHC class Ib protein that is expressed on extravillous trophoblasts), LILRB1 on CD14(+) macrophages and KIR2DL4 on CD56(+) NK cells. Cross-linking with anti-LILRB1 or anti-KIR2DL4 resulted in up-regulation of a small subset of mRNAs including those for IL-6, IL-8, and TNFalpha detected using a microarray representing 114 cytokines. Incubation with transfectants expressing the HLA-G homodimer (but not with transfectants expressing the HLA-G monomer) resulted in secretion of the same cytokine proteins from both leukocyte sets. Moreover, cytokine secretion from both leukocyte sets was blocked by both the appropriate anti-receptor mAb and by anti-HLA-G. The amount of these cytokines secreted by decidual macrophages was substantially greater than that secreted by decidual NK cells. VEGF was constitutively secreted by both cell types. LILRB1, which contains an immunoreceptor tyrosine-based switch motif, functions here as an activating receptor, although it has been known as an inhibitory receptor. KIR2DL4 also functions as an activating receptor, although it also has the potential to function as an inhibitory receptor. Secretion of proinflammatory and proangiogenic proteins supports a role for these leukocytes in important processes that are essential for successful pregnancy, but they may represent only a portion of the proteins that are secreted.

Decidual macrophages and their roles at the maternal-fetal interface.

The semi-allogeneic fetus, whose genome consists of maternally and paternally inherited alleles, must coexist with an active maternal immune system during its 9 months in utero. Macrophages are the second most abundant immune cell at the maternal-fetal interface, although populations and functions for these populations remain ill defined. We have previously reported two distinct subsets of CD14(+) decidual macrophages found to be present in first trimester decidual tissue, 20 percent CD11c(HI) and 68 percent CD11c(LO). Interestingly, CD11c(HI) decidual macrophages express genes associated with lipid metabolism, inflammation, and antigen presentation function and specifically upregulate CD1 molecules. Conversely, CD11c(LO) decidual macrophages express genes associated with extracellular matrix formation, muscle regulation, and tissue growth. The large abundance of CD11c(HI) decidual macrophages and their ability to process antigens more efficiently than CD11c(LO) macrophages suggests that CD11c(HI) macrophages may be important antigen processing and presenting cells at the maternal-fetal interface, while CD11c(LO) macrophages may perform necessary homeostatic functions during placental construction. Thus, macrophage heterogeneity may be an important and necessary division of labor that leads to both an induction of maternal immune cell tolerance to fetal antigens as well as basic homeostatic functions in human pregnancy.

Human Islet Amyloid Polypeptide (hIAPP) Protofibril-Specific Antibodies for Detection and Treatment of Type 2 Diabetes.

Type 2 diabetes mellitus (T2D) is a major public health concern and is characterized by sustained hyperglycemia due to insulin resistance and destruction of insulin-producing ß cells. One pathological hallmark of T2D is the toxic accumulation of human islet amyloid polypeptide (hIAPP) aggregates. Monomeric hIAPP is a hormone normally co-secreted with insulin. However, increased levels of hIAPP in prediabetic and diabetic patients can lead to the formation of hIAPP protofibrils, which are toxic to ß cells. Current therapies fail to address hIAPP aggregation and current screening modalities do not detect it. Using a stabilizing capping protein, monoclonal antibodies (mAbs) can be developed against a previously nonisolatable form of hIAPP protofibrils, which are protofibril specific and do not engage monomeric hIAPP. Shown here are two candidate mAbs that can detect hIAPP protofibrils in serum and hIAPP deposits in pancreatic islets in a mouse model of rapidly progressing T2D. Treatment of diabetic mice with the mAbs delays disease progression and dramatically increases overall survival. These results demonstrate the potential for using novel hIAPP protofibril-specific mAbs as a diagnostic screening tool for early detection of T2D, as well as therapeutically to preserve ß cell function and target one of the underlying pathological mechanisms of T2D.

CD1 Antigen Presentation and Autoreactivity in the Pregnant Human Uterus.

PROBLEM: CD11c(HI) human decidual macrophages express several isoforms of CD1 molecules. Their expression pattern and function required investigation. METHOD OF STUDY: CD11c(HI) macrophages were isolated from decidua. Expression of CD1 isoforms and their ability to present lipid antigens to T cells was studied. RESULTS: CD1a, CD1c, and CD1d were all expressed on CD11c(HI) dMϕ, a pattern differing from those previously observed. Exposure of peripheral monocytes and dendritic cells to lipid isolates from decidua led to increased surface CD1a levels only. The CD1a and CD1c on dMϕ were able to present the appropriate lipid antigens to lipid antigen-specific T cells. Finally, autoreactivity of decidual T cells to CD1a was observed. CONCLUSION: The unique pattern of expression of CD1 isoforms on CD11c(HI) dMϕ is consistent with organ-specific roles of CD1 in human T-cell responses. dMϕ are able to present lipid antigens to both peripheral and decidual T cells and are major antigen-presenting cells in human decidua.

The HCMV membrane glycoprotein US10 selectively targets HLA-G for degradation.

Human cytomegalovirus (HCMV) encodes an endoplasmic reticulum (ER)-resident transmembrane glycoprotein, US10, expressed early in the replicative cycle of HCMV as part of the same cluster that encodes the known immunoevasins US2, US3, US6, and US11. We show that US10 down-regulates cell surface expression of HLA-G, but not that of classical class I MHC molecules. The unique and short cytoplasmic tail of HLA-G (RKKSSD) is essential in its role as a US10 substrate, and a tri-leucine motif in the cytoplasmic tail of US10 is responsible for down-regulation of HLA-G. Both the kinetics of HLA-G degradation and the mechanisms responsible appear to be distinct from those used by the US2 and US11 pathways, suggesting the existence of a third route of protein dislocation from the ER. We show that US10-mediated degradation of HLA-G interferes with HLA-G-mediated NK cell inhibition. Given the role of HLA-G in protecting the fetus from attack by the maternal immune system and in directing the differentiation of human dendritic cells to promote the evolution of regulatory T cells, HCMV likely targets the HLA-G-dependent axis of immune recognition no less efficiently than it interferes with classical class I MHC-restricted antigen presentation.