Jane S. A. Voerman

Imiquimod-Induced Psoriasis-Like Skin Inflammation in Mice Is Mediated via the IL-23/IL-17 Axis

Topical application of imiquimod (IMQ), a TLR7/8 ligand and potent immune activator, can induce and exacerbate psoriasis, a chronic inflammatory skin disorder. Recently, a crucial role was proposed for the IL-23/IL-17 axis in psoriasis. We hypothesized that IMQ-induced dermatitis in mice can serve as a model for the analysis of pathogenic mechanisms in psoriasis-like dermatitis and assessed its IL-23/IL-17 axis dependency. Daily application of IMQ on mouse back skin induced inflamed scaly skin lesions resembling plaque type psoriasis. These lesions showed increased epidermal proliferation, abnormal differentiation, epidermal accumulation of neutrophils in microabcesses, neoangiogenesis, and infiltrates consisting of CD4+ T cells, CD11c+ dendritic cells, and plasmacytoid dendritic cells. IMQ induced epidermal expression of IL-23, IL-17A, and IL-17F, as well as an increase in splenic Th17 cells. IMQ-induced dermatitis was partially dependent on the presence of T cells, whereas disease development was almost completely blocked in mice deficient for IL-23 or the IL-17 receptor, demonstrating a pivotal role of the IL-23/IL-17 axis. In conclusion, the sole application of the innate TLR7/8 ligand IMQ rapidly induces a dermatitis closely resembling human psoriasis, critically dependent on the IL-23/IL-17 axis. This rapid and convenient model allows further elucidation of pathogenic mechanisms and evaluation of new therapies in psoriasis.

Markers of mouse macrophage development detected by monoclonal antibodies

In this review, we present and discuss a selected panel of antibody-defined markers expressed during different stages of mouse macrophage development. We distinguish four categories of markers, which are characteristic of: (1) macrophage precursors and immature macrophages (ER-MP12, ER-MP20, ER-MP54, ER-MP58); (2) mature macrophages in general (F4/80, BM8, Mac-1, Mac-2, ER-BMDM1); (3) macrophage subsets (ER-HR3, ER-MP23, ER-TR9, Forssman antigen, MOMA-1, MOMA-2, Monts-4, SER-4), and (4) IFN-gamma-stimulated macrophages (H-2Ia, LFA-1, ICAM-1, 158.2, MBR-2, TM-2, TM-4, and TM-5). It should be noted that many of the markers in this last category are inducible by other stimuli as well. The rigid classification of markers into four separate groups should be regarded as a digitalization of a continuum, thus inevitably implicating a simplification of the complex phenotypic changes that occur during mononuclear phagocyte development. Nevertheless, the current selection of antibodies against markers for different developmental stages of macrophages constitutes an important tool for characterization of mouse macrophages which participate in various biological processes.

Brain antigens in functionally distinct antigen-presenting cell populations in cervical lymph nodes in MS and EAE

Drainage of central nervous system (CNS) antigens to the brain-draining cervical lymph nodes (CLN) is likely crucial in the initiation and control of autoimmune responses during multiple sclerosis (MS). We demonstrate neuronal antigens within CLN of MS patients. In monkeys and mice with experimental autoimmune encephalomyelitis (EAE) and in mouse models with non-inflammatory CNS damage, the type and extent of CNS damage was associated with the frequencies of CNS antigens within the cervical lymph nodes. In addition, CNS antigens drained to the spinal-cord-draining lumbar lymph nodes. In human MS CLN, neuronal antigens were present in pro-inflammatory antigen-presenting cells (APC), whereas the majority of myelin-containing cells were anti-inflammatory. This may reflect a different origin of the cells or different drainage mechanisms. Indeed, neuronal antigen-containing cells in human CLN did not express the lymph node homing receptor CCR7, whereas myelin antigen-containing cells in situ and in vitro did. Nevertheless, CLN from EAE-affected CCR7-deficient mice contained equal amounts of myelin and neuronal antigens as wild-type mice. We conclude that the type and frequencies of CNS antigens within the CLN are determined by the type and extent of CNS damage. Furthermore, the presence of myelin and neuronal antigens in functionally distinct APC populations within MS CLN suggests that differential immune responses can be evoked.

Subsets of Macrophages and Dendritic Cells in Nonobese Diabetic Mouse Pancreatic Inflammatory Infiltrates: Correlation with the Development of Diabetes

Islet-specific T cells are essential in the development of type I diabetes. The role of non-lymphoid cells is relatively unclear, although infiltration of dendritic cells and macrophages is the first sign of islet autoimmunity in diabetes-prone nonobese diabetic (NOD) mice. BDC2.5 is one of the autoreactive T cell clones isolated from NOD mice. Transfer of BDC2.5 T cells into young NOD mice accelerates diabetes development, whereas transgenic expression of the BDC2.5 T cell receptor on NOD T cells (BDC2.5 TCR-Tg NOD) markedly reduces diabetes development. We show that, although the same antigen-specificity is involved, both models differ significantly in insulitis. BDC2.5 TCR-Tg NOD mice develop an extensive, but non-aggressive, peri-insulitis by 3 weeks of age. In these large peri-islet infiltrates, resembling secondary lymphoid tissue, BM8+ macrophages (Mφ) are virtually absent. In contrast, BDC2.5 T cell clone transfer results in an aggressive insulitis with small infiltrates, but relatively large numbers of BM8+ Mφ. Infiltration of BM8+ Mφ therefore correlates with islet destruction. This is, however, not observed for all Mφ; Monts-4+ Mφ follow a reverse pattern and are present in higher numbers in BDC2.5 TCR-Tg than in transferred mice. ER-MP23+ Mφ are reduced in both transferred and transgenic mice compared with wild-type NOD. Thus, this study underlines and extends previous data suggesting that Mφ are implicated in both early and late phases in diabetes development. Furthermore, our data imply that subsets of non-lymphoid cells have different roles in diabetes development. It is, therefore, important to recognize this heterogeneity when interpreting both in vivo and in vitro studies concerning non-lymphoid cells in diabetes.

Severe B cell deficiency and disrupted splenic architecture in transgenic mice expressing the E41K mutated form of Bruton's tyrosine kinase

To identify B‐cell signaling pathways activated by Brutons tyrosine kinase (Btk) in vivo, we generated transgenic mice in which Btk expression is driven by the MHC class II Ea gene locus control region. Btk overexpression did not have significant adverse effects on B cell function, and essentially corrected the X‐linked immunodeficiency (xid) phenotype in Btk− mice. In contrast, expression of a constitutively activated form of Btk carrying the E41K gain‐of‐function mutation resulted in a B cell defect that was more severe than xid. The mice showed a marked reduction of the B cell compartment in spleen, lymph nodes, peripheral blood and peritoneal cavity. The levels in the serum of most immunoglobulin subclasses decreased with age, and B cell responses to both T cell‐independent type II and T cell‐dependent antigens were essentially absent. Expression of the E41K Btk mutant enhanced blast formation of splenic B cells in vitro in response to anti‐IgM stimulation. Furthermore, the mice manifested a disorganization of B cell areas and marginal zones in the spleen. Our findings demonstrate that expression of constitutively activated Btk blocks the development of follicular recirculating B cells.

The novel calicheamicin-conjugated CD22 antibody inotuzumab ozogamicin (CMC-544) effectively kills primary pediatric acute lymphoblastic leukemia cells

We investigated whether the newly developed antibody (Ab) -targeted therapy inotuzumab ozogamicin (CMC-544), consisting of a humanized CD22 Ab linked to calicheamicin, is effective in pediatric primary B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cells in vitro, and analyzed which parameters determine its efficacy. CMC-544 induced dose-dependent cell kill in the majority of BCP-ALL cells, although IC50 values varied substantially (median 4.8 ng/ml, range 0.1–1000 ng/ml at 48 h). The efficacy of CMC-544 was highly dependent on calicheamicin sensitivity and CD22/CMC-544 internalization capacity of BCP-ALL cells, but hardly on basal and renewed CD22 expression. Although CD22 expression was essential for uptake of CMC-544, a repetitive loop of CD22 saturation, CD22/CMC-544 internalization and renewed CD22 expression was not required to achieve intracellular threshold levels of calicheamicin sufficient for efficient CMC-544-induced apoptosis in BCP-ALL cells. This is in contrast to studies with the comparable CD33 immunotoxin gemtuzumab ozogamicin (Mylotarg) in acute myeloid leukemia (AML) patients, in which complete and prolonged CD33 saturation was required for apoptosis induction. These data suggest that CMC-544 treatment may result in higher response rates in ALL compared with response rates obtained in AML with Mylotarg, and that therefore clinical studies in ALL, preferably with multiple low CMC-544 dosages, are warranted.

The dermal microenvironment induces the expression of the alternative activation marker CD301/mMGL in mononuclear phagocytes, independent of IL-4/IL-13 signaling

Recently, we have shown that mononuclear phagocytes comprise the majority of interstitial cells in the mouse dermis, as indicated by their phenotypic and functional characteristics. In particular, these cells express the mouse macrophage galactose‐/N‐acetylgalactosamine‐specificlectin (mMGL)/CD301, identified by the monoclonal antibody ER‐MP23, as well as other macrophage markers. As expression of mMGL is induced by IL‐4 or IL‐13 and is therefore a marker of alternatively activated macrophages, we asked whether dermal mononuclear phagocytes are genuinely alternatively activated. We observed that these cells expressed, next to mMGL, two other alternative activation markers, namely, the mannose receptor/CD206 and Dectin‐1. Yet, as this expression profile was similar in IL‐4 receptor α knockout mice, neither IL‐4 nor IL‐13 signaling appeared to be required for this phenotype. We also found that Langerhans cells (LC), which showed only a low level of mMGL in the epidermis, up‐regulated mMGL expression upon migration through the dermis, allowing these cells to internalize limited amounts of mMGL ligands. LC isolated from epidermal preparations did not show this up‐regulation when cultured in standard medium, but whole skin‐conditioned medium did stimulate mMGL expression by LC. The vast majority of mMGL molecules was present in the cytoplasm, however. LC, which arrived in skin‐draining lymph nodes, quickly down‐regulated mMGL expression, and dermally derived cells retained significant mMGL levels. Taken together, these data suggest that the dermal microenvironment induces mononuclear phagocyte subpopulations to express mMGL and possibly other markers of alternatively activated macrophages, independent of IL‐4/IL‐13 signaling.

First trimester interleukin 8 levels are associated with postpartum relapse in multiple sclerosis

Pregnancy has an ameliorating effect on multiple sclerosis (MS), but directly after delivery the risk of a relapse is increased. The pro-inflammatory chemokine interleukin 8 is associated with disease activity. We aimed to investigate whether pregnancy-induced fluctuations of interleukin 8 correlate with periods of enhanced and diminished disease activity. Thirty-six women with MS were prospectively studied before, during and after pregnancy. Serum levels of interleukin 8 were significantly decreased during the third trimester (p = 0.03). High first trimester serum levels of interleukin 8 were associated with a high risk of postpartum relapse (p = 0.007). These results help us to further understand the altered disease course during pregnancy.

Dendritic cells in the autoimmune insulitis in NOD mouse models of diabetes

Dendritic cells (DC) are thought to play important roles as antigen presenting cells in the autoimmune insulitis preceding insulin dependent diabetes mellitus (IDDM). Not only are DC the first cells appearing in the pancreatic inflammatory process in animal models of diabetes (NOD mouse1.2 and BB rat3.4), DC are also abundantly present in progressed human insulitis5. Although insulitis in the NOD mouse starts around 3 weeks, overt diabetes will not develop until an age of 20 weeks. The lag between occult insulitis and diabetes, which can also be observed in human IDDM, was always considered to reflect the time needed for accumulation of a sufficient number of destructive T cells in the islets.

A monoclonal antibody (ER-HR3) against murine macrophages. I. Ontogeny, distribution and enzyme histochemical characterization of ER-HR3-positive cells

We describe ER-HR3, a monoclonal antibody directed against bone marrow-derived haemopoietic reticulum cells. ER-HR3-positive cells have the electronmicroscopic and enzyme-histochemical characteristics of macrophages. Additionally, they are able to phagocytoze. The ER-HR3 antigen is expressed by a majority of blood monocytes and is present on a subpopulation of resident macrophages in multiple organs. ER-HR3-positive cells are abundant in the bone marrow, the splenic red pulp, the mesenteric lymphoid paracortex and the interfollicular areas of the Peyers patch. Few ER-HR3-positive cells have been observed in the thymic cortex and the connective tissues of the gastro-intestinal tract, the dermis and the renal medulla. Moreover, epidermal Langerhans cells express the antigen. No cross-reactivity with other cell types has been found. It is concluded that ER-HR3 has a unique distribution pattern distinct from other macrophage-specific antibodies.