Characterisation of the protective immune response following subcutaneous vaccination of susceptible mice against Trichuris muris

Trichuris muris is a laboratory model for the human whipworm Trichuris trichiura which infects approximately 1 billion people in tropical and sub-tropical countries. The development of a vaccine would control trichuriasis by promoting the acquisition of immunity during childhood, thereby reducing faecal egg output by the community into their environment. Resistance to T. muris, defined as expulsion of the parasite prior to patency, requires the development of a T helper 2 (Th2) response during a primary infection. To our knowledge this is the first study to describe the protective immune response in the peripheral lymph nodes (PLN), mesenteric lymph nodes (MLN) and colonic mucosa following s.c. vaccination against T. muris. Susceptible AKR mice were either vaccinated with T. muris excretory-secretory product (ES) in incomplete Freund’s adjuvant (IFA) (ES/IFA) or injected with PBS in IFA (PBS/IFA) and for protection experiments were infected with embryonated infective T. muris eggs 10 days later. The ES/IFA vaccine induced the proliferation of PLN cells and their production of Th2 cytokines and the Th1-associated cytokine IFN-γ. Following a challenge infection, the ES/IFA vaccination offered susceptible mice complete protection. While MLN-derived IFN-γ was produced by infected mice following either ES/IFA vaccination or PBS/IFA, the protection of susceptible mice by ES/IFA was characterised by the production of MLN-derived Th2 cytokines. Goblet cell hyperplasia and the influx and alternative activation of macrophages were observed locally in the gut post-challenge infection. The rate of epithelial turnover did not appear to be increased by vaccination, suggesting that there are differences in the mechanisms of expulsion between ‘natural resistance’ and ‘vaccinated resistance’. High levels of serum IgG1 and cell-bound IgG1 in the colon of mice protected by the ES/IFA vaccine suggest that antibody may be involved in vaccination-induced worm expulsion.     

CBR2激活与小胶质细胞的活化和损伤的关系

目的：探讨大麻素CBR2受体激动剂AM1241预处理对脂多糖（LPS）和γ-干扰素（IFN-γ）所致炎症反应对小胶质细胞活化和损伤的影响。方法：联用LPS和IFN-γ作为小胶质细胞损伤模型,将细胞分为Control组、AM1241组、LPS/IFN-γ组和AM1241＋LPS/IFN-γ组;AM1241组和AM1241＋LPS/IFN-γ组经AM1241预处理2h,LPS/IFN-γ组和AM1241＋LPS/IFN-γ组用含LPS和IFN-γ的培养基培养24h。采用MTT法检测细胞代谢率,硝酸还原酶法检测细胞培养液中一氧化氮（NO）释放量,酶联免疫吸附剂测定细胞培养基中炎症因子释放量,倒置相差显微镜观察细胞形态。结果：与LPS/IFN-γ组相比,AM1241＋LPS/IFN-γ组细胞代谢率明显升高（P〈0.05）,NO、TNF-α、IL-1β和IL-10释放量明显减少（P〈0.05）,活化和损伤程度明显减轻。结论：大麻素CBR2受体激动剂AM1241预处理可减轻LPS和IFN-γ对小胶质细胞的活化和损伤。     

Activation, dysfunction and retention of T cells in vaccine sites after injection of incomplete Freund’s adjuvant, with or without peptide

We conducted a randomized clinical trial in 45 patients with resected AJCC stage IIB-IV melanoma to characterize cellular and molecular events at sites of immunization with incomplete Freund’s adjuvant (IFA) alone, or a melanoma vaccine in IFA. At a primary vaccine site, all patients received a multi-peptide melanoma vaccine in IFA. At a replicate vaccine site, which was biopsied, group 1 received IFA only; group 2 received vaccine in IFA. Lymphocytes isolated from replicate vaccine site microenvironments (VSME) were compared to time-matched peripheral blood mononuclear cells (PBMC) in ELISpot and flow cytometry assays. Compared to PBMC, the VSME had fewer naïve and greater proportions of effector memory CD8⁺ T cells (T_CD8). The vast majority of T_CD8 within the VSME were activated (CD69⁺), with a concentration of antigen-specific (tetramer^pos) cells in the VSME, particularly in vaccine sites with peptide (group 2). CXCR3⁺ lymphocytes were concentrated in the VSME of all patients, suggesting IFA-induced chemokine recruitment. T_CD8 expression of retention integrins αEβ7 and α1β1 was elevated in VSME, with the highest levels observed in antigen-specific cells in VSME containing peptide (group 2). T_CD8 retained in the VSME of both groups were strikingly dysfunctional, with minimal IFN-γ production in response to peptide stimulation and few tetramer^pos cells producing IFN-γ. These data suggest that vaccine-induced selective retention and dysfunction of antigen-specific T_CD8 within VSME may represent a significant mechanism underlying transient immune responses and low clinical response rates to peptide vaccines administered in IFA.     

Uncarinic acid C plus IFN-γ generates monocyte-derived dendritic cells and induces a potent Th1 polarization with capacity to migrate

Uncarinic acid C (URC) is triterpene isolated from Uncaria rhynchophylla and modulates human DC function in a fashion that favors Th1 cell polarization depending on TLR4 signaling. The induction of dendritic cells (DC) is critical for the induction of Ag-specific T lymphocyte responses and may be essential for the development of human vaccines relying on T cell immunity. Monocyte-derived DC used as adjuvant cells in cancer immunotherapy and have shown promising results. We studied the effect of interferon’s (IFN-α and IFN-γ) and TNF-α on phenotypic and functional maturation, and cytokine production of URC-primed DC in vitro. Human monocytes were exposed to either URC alone, or in combination with TNF-α, IFN-α or IFN-γ, and thereafter co-cultured with naïve T cells. We found that the expression levels of CD1a, CD83 and HLA-DR on URC-primed DC were influenced by IFN-γ and IFN-γ augmented the T cell stimulatory capacity in allo MLR to URC-primed DC. Moreover, the production of IL-12p70 by URC-primed DC was enhanced by IFN-γ. IL-12p70 production by URC-primed DC alone was influenced following treatment with anti-TLR4 mAb, but not DC differentiated with URC plus IFN-γ. URC plus IFN-γ-primed DC induced a substantial increase in the secretion of IFN-γ by T cells, which is dependent on IL-12 secretion. DC maturated with URC plus IFN-γ had an intermediate migratory capacity towards CCL19 and CCL21. In addition, the expression levels of CCR7 on URC-primed DC were enhanced by IFN-γ. In contrast, surface molecule up-regulation and function of URC-primed DC were slightly enhanced by TNF-α, and IFN-α. These results suggest that the enhancement of Th1 cells polarization to URC-primed DC induced by IFN-γ depends on the activation of IL-12p70 and independent on TLR4. DC differentiated with URC in combination with IFN-γ might be used on DC-based vaccine for cancer immunotherapy.     

Suppression of thymus- and activation-regulated chemokine (TARC/CCL17) production by 1,2,3,4,6-penta-O-galloyl-β-d-glucose via blockade of NF-κB and STAT1 activation in the HaCaT cells

Keratinocytes, one of major cell types in the skin, can be induced by TNF-α and IFN-γ to express thymus- and activation-regulated chemokine (TARC/CCL17), which is considered to be a pivotal mediator in the inflammatory responses during the development of inflammatory skin diseases, such as atopic dermatitis (AD). In this study, we examined the effect of 1,2,3,4,6-penta-O-galloyl-β-d-glucose (PGG), isolated from the barks of Juglans mandshurica, on TNF-α/IFN-γ induced CCL17 expression in the human keratinocyte cell line HaCaT. Pretreatment of HaCaT cells with PGG suppressed TNF-α/IFN-γ-induced protein and mRNA expression of CCL17. PGG significantly inhibited TNF-α/IFN-γ-induced NF-κB activation as well as STAT1 activation. Furthermore, pretreatment with PGG resulted in significant reduction in expression of CXCL9, 10, and 11 in the HaCaT cells treated with IFN-γ. These results suggest that PGG may exert anti-inflammatory responses by suppressing TNF-α and/or IFN-γ-induced activation of NF-κB and STAT1 in the keratinocytes and might be a useful tool in therapy of skin inflammatory diseases.     

LPS/IFN-γ-Induced RAW 264.7 Apoptosis is Regulated by Both Nitric Oxide–Dependent and –Independent Pathways Involving JNK and the Bcl-2 Family

Lipopolysaccharide (LPS) and interferon-gamma (IFN-γ) stimulate macrophages to produce nitric oxide (NO) via inducible nitric oxide synthase (iNOS) and activate stress signaling cascades including the c-jun-N-terminal kinase (JNK) pathway. These events trigger an apoptotic cascade that ultimately results in death. Since JNK regulates pro-apoptotic and anti-apoptotic Bcl-2 family members, the role of NO in LPS/IFN-γ-induced activation of JNK and its effects on the Bcl-2 family was examined in RAW 264.7 macrophage-like cells. Inhibition of JNK by siRNA verified a role for JNK in LPS/IFN-γ-induced apoptosis. Suppression of NO production by a pharmacologic agent, i.e. iNOS inhibitor L-NIL, altered the kinetics of JNK activation by LPS/IFN-γ. Examination of mitochondrial and nuclear compartments of RAW 264.7 cells demonstrated NO-dependent activation of mitochondrial JNK by LPS/IFN-γ, but NO-independent, cytokine-induced phosphorylation of Bim. NO did not affect phosphorylation, but did inhibit Bax phosphorylation. These results suggest a novel mechanism of LPS/IFN-γ-induced apoptosis in macrophages involving NO-independent phosphorylation of Bim and NO-dependent dephosphorylation of Bax.     

Tryptophan Depletion and the Kinase GCN2 Mediate IFN-γ-Induced Autophagy

IFN-γ is a master regulator of the immune responses that occur in the transplanted kidney, acting both on the immune system and on the graft itself. The cellular responses to IFN-γ are complex, and emerging evidence suggests that IFN-γ may regulate autophagic functions. Conversely, autophagy modulates innate and adaptive immune functions in various contexts. In this study, we identify a novel mechanism by which IFN-γ activates autophagy in human kidney epithelial cells and provide new insights into how autophagy regulates immune functions in response to IFN-γ. Our results indicate that IFN-γ promotes tryptophan depletion, activates the eIF2α kinase general control nonderepressible-2 (GCN2), and leads to an increase in the autophagic flux. Further, tryptophan supplementation and RNA interference directed against GCN2 inhibited IFN-γ-induced autophagy. This process is of functional relevance because autophagy regulates the secretion of inflammatory cytokines and growth factors by human kidney epithelial cells in response to IFN-γ. These findings assign to IFN-γ a novel function in the regulation of autophagy, which, in turn, modulates IFN-γ-induced secretion of inflammatory cytokines.     

Toll-like receptor homolog RP105 modulates the antigen-presenting cell function and regulates the development of collagen-induced arthritis

Introduction

RP105 is a Toll-like receptor homolog expressed on B cells, dendritic cells (DCs), and macrophages. We investigated the role of RP105 in the development of collagen-induced arthritis (CIA).

Methods

CIA was induced in RP105-deficient DBA/1 mice and the incidence and arthritis index were analyzed. The cytokine production by spleen cells was determined. The functions of the DCs and regulatory T cells (Tregs) from RP105-deficient or control mice were determined by adding these cells to the lymph node cell culture. Arthritis was also induced by incomplete Freund's adjuvant (IFA) plus collagen or by injecting anti-collagen antibody and lipopolysaccharide.

Results

RP105-deficient mice showed accelerated onset of arthritis and increased severity. Interferon-gamma (IFN-γ) and tumor necrosis factor-alpha production by spleen cells from RP105-deficient mice was increased in comparison with that from wild-type mice. The DCs from RP105-deficient mice induced more IFN-γ production, whereas Tregs from those mice showed less inhibitory effect against IFN-γ production. RP105-deficient mice also showed more severe arthritis induced by collagen with IFA.

Conclusions

These results indicate that RP105 regulates the antigen-presenting cell function and Treg development, which induced the attenuation of the cell-mediated immune responses and, as a result, suppressed the development of CIA.     

Changes in cells's secretory organelles and extracellular matrix during endochondral ossification in the mandibular condyle of the growing rat

The mandibular condyle from 20-day-old rats was examined in the electron microscope with particular attention to intracellular secretory granules and extracellular matrix. Moreover, type II collagen was localized by an immunoperoxidase method. The condyle has been divided into five layers: (1) the most superficial, articular layer, (2) polymorphic cell layer, (3) flattened cell layer, (4) upper hypertrophic, and (5) lower hypertrophic cell layers. In the articular layer, the cells seldom divide, but in the polymorphic layer and upper part of the flattened cell layer, mitosis gives rise to new cells. In these layers, cells produce two types of secretory granules, usually in distinct stacks of the Golgi apparatus; type a, cylindrical granules, in which 300-nm-long threads are packed in bundles which appear "lucent" after formaldehyde fixation; and type b, spherical granules loaded with short, dotted filaments. The matrix is composed of thick banded "lucent" fibrils in a loose feltwork of short, dotted filaments. The cells arising from mitosis undergo endochondral differentiation, which begins in the lower part of the flattened cell layer and is completed in the upper hypertrophic cell layer; it is followed by gradual cell degeneration in the lower hypertrophic cell layer. The cells produce two main types of secretory granules: type b as above; and type c, ovoid granules containing 300-nm-long threads associated with short, dotted filaments. A possibly different secretory granule, type d, dense and cigar-shaped, is also produced. The matrix is composed of thin banded fibrils in a dense feltwork. In the matrix of the superficial layers, the "lucency" of the fibrils indicated that they were composed of collagen I, whereas the "lucency" of the cylindrical secretory granules suggested that they transported collagen I precursors to the matrix. Moreover, the use of ruthenium red indicated that the feltwork was composed of proteoglycan; the dotted filaments packed in spherical granules were similar to, and presumably the source of, the matrix feltwork. The superficial layers did not contain collagen II and were collectively referred to as perichondrium. In the deep layers, the ovoid secretory granules displayed collagen II antigenicity and were likely to transport precursors of this collagen to the matrix, where it appeared in the thin banded fibrils. That these granules also carried proteoglycan to the matrix was suggested by their content of short dotted filaments. Thus the deep layers contained collagen II and proteoglycan as in cartilage; they were collectively referred to as the hyaline cartilage region.     

(+)-Nootkatone inhibits tumor necrosis factor α/interferon γ-induced production of chemokines in HaCaT cells

Chemokines are important mediators of cell migration, and thymus and activation-regulated chemokine (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22) are well-known typical inflammatory chemokines involved in atopic dermatitis (AD). (+)-Nootkatone is the major component of Cyperus rotundus. (+)-Nootkatone has antiallergic, anti-inflammatory, and antiplatelet activities. The purpose of this study was to investigate the effect of (+)-nootkatone on tumor necrosis factor α (TNF-α)/interferon γ (IFN-γ)-induced expression of Th2 chemokines in HaCaT cells. We found that (+)-nootkatone inhibited the TNF-α/IFN-γ-induced expression of TARC/CCL17 and MDC/CCL22 mRNA in HaCaT cells. It also significantly inhibited TNF-α/IFN-γ-induced activation of nuclear factor kappa B (NF-κB), p38 mitogen-activated protein kinase (MAPK), and protein kinase Cζ (PKCζ). Furthermore, we showed that PKCζ and p38 MAPK contributed to the inhibition of TNF-α/IFN-γ-induced TARC/CCL17 and MDC/CCL22 expression by blocking IκBα degradation in HaCaT cells. Taken together, these results suggest that (+)-nootkatone may suppress TNF-α/IFN-γ-induced TARC/CCL17 and MDC/CCL22 expression in HaCaT cells by inhibiting of PKCζ and p38 MAPK signaling pathways that lead to activation of NF-κB. We propose that (+)-nootkatone may be a useful therapeutic candidate for inflammatory skin diseases such as AD.     

Induction of type I IFN is a physiological immune reaction to apoptotic cell-derived membrane microparticles

Membrane microparticles (MMP) released from apoptotic cells deliver signals that secure the anti-inflammatory response beyond the nearest proximity of the apoptotic cell. Plasmacytoid dendritic cells (pDC) are sentinels prepared to detect cellular processes that endanger the organism. They play a key role in the regulation of both pro- and anti-inflammatory immune responses. Based on the assumption that pDC could participate in the initiation of the anti-inflammatory response to apoptotic cells, we investigated the effects of apoptotic cell-derived MMP on human pDC. The results obtained in our experiments confirmed that MMP released from apoptotic cells trigger IFN-α secretion from human pDC. They further suggest that pDC activation results from sensing of DNA contained in MMP. MMP-DNA displays a particularly strong stimulatory activity compared with MMP-RNA and other sources of DNA. Inhibition of MMP-induced IFN-α secretion by cytochalasin D, chloroquine, and an inhibitory G-rich oligodeoxynucleotide identify TLR9 as the receptor for MMP-DNA. In marked contrast to the pDC response in autoimmune patients, in healthy subjects MMP-mediated stimulation of pDC-derived IFN-α was found to be independent of FcγRIIA (CD32A). Based on our findings, we conclude that induction of pDC-derived IFN-α by MMP is a physiological event; future investigations are necessary to elucidate whether pDC activation promotes inflammation or propagates tolerance in the context of apoptotic cell clearance.     

Gamma interferon modulates myogenesis through the major histocompatibility complex class II transactivator, CIITA

Gamma interferon (IFN-γ) is an inflammatory cytokine that has complex effects on myogenesis. Here, we show that the IFN-γ-induced inhibition of myogenesis is mediated by the major histocompatibility complex (MHC) class II transactivator, CIITA, which binds to myogenin and inhibits its activity. In IFN-γ-treated myoblasts, the inhibition of muscle-specific genes includes the expression of myogenin itself, while in myotubes, myogenin expression is unaffected. Thus, CIITA appears to act by both repressing the expression and inhibiting the activity of myogenin at different stages of myogenesis. Stimulation by IFN-γ in skeletal muscle cells induces CIITA expression as well as MHC class II gene expression. The IFN-γ-mediated repression is reversible, with myogenesis proceeding normally upon removal of IFN-γ. Through overexpression studies, we confirm that the expression of CIITA, independent of IFN-γ, is sufficient to inhibit myogenesis. Through knockdown studies, we also demonstrate that CIITA is necessary for the IFN-γ-mediated inhibition of myogenesis. Finally, we show that CIITA, which lacks DNA binding activity, is recruited to muscle-specific promoters coincident with reductions in RNA polymerase II recruitment. Thus, this work reveals how IFN-γ modulates myogenesis and demonstrates a key role for CIITA in this process.     

Th17 cells in multiple sclerosis express higher levels of JAK2, which increases their surface expression of IFN-γR2

IFN-β inhibits the expansion of Th17 cells in active multiple sclerosis (AMS), and this might contribute to improve the clinical symptoms. The effectiveness of this inhibition, however, requires intact IFN-γ signaling in T cells. In this study, we report that both mRNA and cell surface expression of the signaling chain of the IFN-γ receptor (IFN-γR2) and its cognate tyrosine kinase JAK2 are enhanced in peripheral blood Th17 cells and clones from patients with AMS compared with those with inactive multiple sclerosis (IMS) or healthy subjects (HS). IFN-γ decreased the frequency of Th17 peripheral cells and proliferation of Th17 clones from AMS patients. Stimulation of PBMCs from HS in Th17-polarizing conditions resulted in the enhancement of JAK2 expression and accumulation of cell surface IFN-γR2. The role of JAK2 in the modulation of IFN-γR2 was demonstrated as its transduction prevented rapid internalization and degradation of IFN-γR2 in JAK2-deficient γ2A cells. In conclusion, these data identify JAK2 as a critical factor that stabilizes IFN-γR2 surface expression in Th17 cells from AMS patients, making them sensitive to IFN-γ. These data may have clinical implications for a better use of IFNs in multiple sclerosis and possibly other inflammatory diseases.     

Interferon-γ stimulates p11-dependent surface expression of annexin A2 in lung epithelial cells to enhance phagocytosis

Annexin A2 (p36) is usually present together with its natural ligand p11 as a heterotetramer complex, which has multiple biological functions depending on its cellular localization. However, the detailed mechanism of annexin A2 translocation and its physiological role in inflammation remain unclear. Here, we show that IFN-γ stimulation enhances surface translocation of annexin A2 on lung epithelial cells. While total annexin A2 protein remains unchanged, the expression of p11 is upregulated via the IFN-γ-activated JAK2/STAT1 signal pathway. Notably, IFN-γ-induced p11 expression is required for annexin A2 translocation to the cell surface. Since annexin A2 lacks a signal peptide for surface translocation by the classical endoplasmic reticulum-Golgi route, its mode of trafficking remains unclear. We observed that p11-dependent surface translocation of annexin A2 is associated with the exosomal secretion pathway. The IFN-γ-induced increase of annexin A2 in the exosomes is blocked in p11-silenced cells. Furthermore, IFN-γ-induced surface expression of annexin A2 mediates phagocytosis of apoptotic cells by lung epithelial cells. These findings provide insights into the surface translocation mechanism of annexin A2 and illustrate a pivotal function of surface annexin A2 in the phagocytic response to IFN-γ.     

MiR-375-3p alleviates the severity of inflammation through targeting YAP1/LEKTI pathway in HaCaT cells

ABSTRACT

Atopic dermatitis (AD) is a relapsing inflammatory skin disease with a complicated pathogenesis. This study aimed to investigate whether miR-375-3p could regulate AD through the Yes-associated protein 1 (YAP1) pathway. In this study, inflammatory response was induced by TNF-α and IFN-γ administration in HaCaT cells. We found that viability and inflammatory factor release, including interleukin-1β (IL-1β) and IL-6, were negatively related to miR-375-3p expression in HaCaT cells. We also found that YAP1 overexpression down-regulated lympho-epithelial Kazal type inhibitor (LEKTI) levels and aggravated viability and inflammation in TNF-α and IFN-γ-treated HaCaT cells. Dual-luciferase reporter assay proved the targeted binding of miR-375-3p and YAP1 3?-UTR. Additionally, the protective effect of miR-375-3p on inflammatory response in TNF-α and IFN-γ-treated HaCaT cells could be impeded by YAP1 overexpression. Collectively, our results suggested that miR-375-3p could modulate HaCaT cell viability and inflammation through the YAP1/LEKTI pathway.     

平喘固本合剂对昆系小鼠急性哮喘模型气道炎症作用影响

目的:探讨平喘固本合剂对昆系小鼠急性哮喘模型气道炎症作用影响及相关机制。方法:昆明系小鼠40只,随机分为对照组(A),哮喘模型组(B),喘固本合剂治疗组(C),布地奈德雾化治疗组(D)及平喘固本合剂联合布地奈德雾化治疗组(E)。用鸡卵蛋白(OVA)致敏建立昆系小鼠哮喘急性气道炎症模型,并给与药物治疗10天。对各组支气管肺泡灌洗液(BALF)中各种细胞进行分类并计数,观察肺组织的病理变化,同时应用ELISA法测定灌洗液中炎症相关因子的水平变化。结果:与B组比较,C组、D组、E组BALF中细胞总数、嗜酸性粒细胞数、巨噬细胞数及IL-4、IL-4/IFN-γ明显降低(P<0.05),IFN-γ明显升高(P<0.05)。对于IL-4、IL-4/IFN-γ水平的比较C组与D组无明显统计学差异,E组与两者具有明显统计学意义(P<0.05)。HE染色显示C组、D组、E组较单纯模型组炎症细胞浸润,平滑肌肥厚及黏膜肺组织水肿等炎症表现明显减轻。结论:平喘固本合剂对哮喘昆系小鼠气道炎症有明显的抑制作用,其作用机制可能与抑制IL-4的表达、炎性细胞聚积及促进IFN-γ的表达有关,并且可能与糖皮质激素有一定协同作用。     

Dissociation of human interferon-gamma-like activity from migration-inhibition factor

Supernatants harvested from concanavalin A-stimulated human peripheral mononuclear cells after 24 hr of incubation contain one interferon species similar to human interferon-gamma (IFN-γ) with a pI of 4.6–5.3 (first day pH 5 IFN-γ). In contrast, during the subsequent 24 hr of incubation two species with properties of IFN-γ are produced with pI of 3.6–4.0 (second day pH 4 IFN-γ) and 4.6–5.6 (second day pH 5 IFN-γ), respectively. First day pH 5 IFN-γ and second day pH 5 IFN-γ have been found to differ on the basis of trypsin sensitivity. This pattern of polymorphism is similar to the pattern previously described for human migration-inhibitory factor (MIF) which can be separated into first day pH 5 MIF, second day pH 3 MIF, and second day pH 5 MIF. However, IFN-γ-like species can be differentiated from MIF biochemically and antigenically. Fractions with second day pH 4 IFN-γ have no MIF activity and fractions with second day pH 3 MIF contain no IFN activity. In addition, first and second day pH 5 MIF, which also contain IFN-γ activity, can be separated from the latter by precipitation as well as neutralization with polyclonal and monoclonal anti-human MIF antibodies.     

Atorvastatin ameliorates experimental autoimmune neuritis by decreased Th1/Th17 cytokines and up-regulated T regulatory cells

Statins have anti-inflammatory and immune-regulating properties. To investigate the effects of atorvastatin on experimental autoimmune neuritis (EAN), an animal model of Guillain–Barré syndrome (GBS), atorvastatin was administered to Lewis rats immunized with bovine peripheral myelin in complete Freund’s adjuvant. We found that atorvastatin ameliorated the clinical symptoms of EAN, decreased the numbers of inflammatory cells as well as IFN-γ⁺ and IL-17⁺ cells in sciatic nerves, decreased the CD80 expression and increased the number of CD25⁺Foxp3⁺ cells in mononuclear cells (MNC), and decreased the levels of IFN-γ in MNC culture supernatants. These data provide strong evidence that atorvastatin can act as an inhibitor in EAN by inhibiting the immune response of Th1 and Th17, decreasing the expression of co-stimulatory molecule, and up-regulating the number of T regulatory cells. These data demonstrated that statins could be used as a therapeutic strategy in human GBS in future.     

Apoptotic neutrophils and nitric oxide regulate cytokine production by IFN-γ-stimulated macrophages

Early apoptotic neutrophils but not secondary necrotic ones down-regulate LPS-induced proinflammatory cytokine production of macrophages, thereby contributing to the resolution of inflammation. IFN-γ is also a well-known stimulant of macrophages, but how the apoptotic neutrophils affect IFN-γ-stimulated macrophages remains largely unexplored. Since IFN-γ induces the expression of inducible nitric oxide (NO) synthase, we examined the production of NO and various cytokines, including MIP-2, TNF-α, IL-12p40, IL-6, IL-10, and TGF-β, by IFN-γ-stimulated murine macrophages, the effect of coculturing the macrophages with early apoptotic or secondary necrotic neutrophils, and the regulatory role of NO in such cocultures. IFN-γ induced significant production of NO, IL-12p40, and IL-6 by macrophages, but not other cytokines. Early apoptotic neutrophils but not secondary necrotic ones promoted NO production, whereas secondary necrotic ones and their supernatants promoted TNF-α production. In contrast, both early apoptotic and secondary necrotic neutrophils suppressed IL-12p40 and IL-6 production. Furthermore, macrophages from inducible NO synthase-deficient mice produced significantly higher levels of MIP-2 than those from wild-type mice. Consistent with this, treatment of macrophages with l-NAME, an NO synthase inhibitor, also induced the production of a large amount of MIP-2. In conclusion, this study suggests that early apoptotic neutrophils are critical in the resolution of inflammation, but that secondary necrotic neutrophils may not cause an inflammatory response. Apoptotic neutrophils, however, appear not to modulate cytokine production via NO.     

IFN-α production by plasmacytoid dendritic cells stimulated with RNA-containing immune complexes is promoted by NK cells via MIP-1β and LFA-1

Several systemic autoimmune diseases display a prominent IFN signature. This is caused by a continuous IFN-α production by plasmacytoid dendritic cells (pDCs), which are activated by immune complexes (ICs) containing nucleic acid. The IFN-α production by pDCs stimulated with RNA-containing IC (RNA-IC) consisting of anti-RNP autoantibodies and U1 small nuclear ribonucleoprotein particles was recently shown to be inhibited by monocytes, but enhanced by NK cells. The inhibitory effect of monocytes was mediated by TNF-α, PGE(2), and reactive oxygen species, but the mechanisms for the NK cell-mediated increase in IFN-α production remained unclear. In this study, we investigated the mechanisms whereby NK cells increase the RNA-IC-induced IFN-α production by pDCs. Furthermore, NK cells from patients with systemic lupus erythematosus (SLE) were evaluated for their capacity to promote IFN-α production. We found that CD56(dim) NK cells could increase IFN-α production >1000-fold after RNA-IC activation, whereas CD56(bright) NK cells required costimulation by IL-12 and IL-18 to promote IFN-α production. NK cells produced MIP-1α, MIP-1β, RANTES, IFN-γ, and TNF-α via RNA-IC-mediated FcγRIIIA activation. The IFN-α production in pDCs was promoted by NK cells via MIP-1β secretion and LFA-mediated cell-cell contact. Moreover, NK cells from SLE patients displayed a reduced capacity to promote the RNA-IC-induced IFN-α production, which could be restored by exogenous IL-12 and IL-18. Thus, different molecular mechanisms can mediate the NK cell-dependent increase in IFN-α production by RNA-IC-stimulated pDCs, and our study suggests that the possibility to therapeutically target the NK-pDC axis in IFN-α-driven autoimmune diseases such as SLE should be investigated.