Technical improvements in the clotted plasma droplet MIF assay in vitro.

The migration inhibitory activity of culture supernatants of rat and human lymphocytes stimulated with PHA and Con A-Sepharose was tested on cell migration from clotted plasma droplets. This technique was improved by using homologous as well as heterologous plasma and fibrinogen solutions for suspending the migratory cells, automatic micropipettes for performing the technique and purified cell populations as target. The effects of calcium chloride and of the cell concentration in the plasma droplets on the migration indices obtained in the MIF assay were tested.     

Type 1 and type 2 tumor infiltrating effector cell subpopulations in progressive breast cancer

Effector T cells fall into two subpopulations based on cytokine-secretion. Type 1 cells secrete IFN-gamma, whereas type 2 cells secrete IL-4, IL-10, and GM-CSF. NKT cells represent a third subpopulation that secretes similar cytokines and have been associated with immunoregulation. Using the TS/A adenocarcinoma, we assessed the phenotype and kinetics of tumor-infiltrating lymphocytes (TIL) in mice challenged subcutaneously in the mammary region. Flow cytometric analysis shows that T cells do not infiltrate the primary tumor site until days 7-14 following tumor challenge. Both CD4 and CD8 TILs were predominantly CD44(High) and expressed CD25, CD69, and CD95 cell surface activation markers. Activated CD4/CD44(High) TIL numbers reached peak levels at day 21 that precipitously decreased by day 28 whereas corresponding CD8 cell numbers progressively increased, however, at lower levels and with later kinetics. Intracellular cytokine staining showed that greater numbers of IL-4-producing Th2 cells were elicited and with earlier kinetics than that of IFN-gamma-producing Th1 cells. T cells co-expressing DX5 (CD3(+)/DX5(+)) emerged (>21 days), suggesting a recruitment of NK-like T cells at later stages of tumor progression. Moreover, tumors selectively up-regulated TGF-beta, MIF, and IP-10 gene expression at times as early as day 4, with peak levels at day 7 in vivo. Such gene expression remained elevated and correlated with a continued progression in tumor growth suggesting that preferential effector cell recruitment and production of select factors during different stages of tumor maturation may aid in regulating effective endogenous antitumor responses in progressive breast cancer.     

Immunologic events in pigeon breeders'' disease

The immunologic and physiologic status of a group of symptomatic and asymptomatic pigeon breeders was studied in an attempt to define the immunologic events occurring in pigeon breeders' disease. Antibody activity to antigen(s) present in pigeon dropping extract (PDE) and pigeon serum (PS) was detected in the serum of both symptomatic and asymptomatic breeders. Antibody activity, however, tended to be greater in the symptomatic pigeon breeders. When subjects were challenged with PS via aerosol, serum complement activity became depressed only in asymptomatic patients. Cellular hypersensitivity to antigens present in PDE was detected in vitro in peripheral lymphocyte populations of 4 of 5 symptomatic breeders and in none of the asymptomatic breeders; cellular hypersensitivity to antigens in PS was not demonstrated in any of the individuals tested. These findings indicate that cell-mediated hypersensitivity, as well as humoral immunologic processes, may be involved in the pathogenesis of the hypersensitivity pneumonitis found in pigeon breeders.     

Chemical Characterization of Macrophage Cytotoxicity Factor,Macrophage Migration Inhibitory Factor,T-Helper Cell-Replacing Factor and Colony-Stimulating Factor from Culture Supernatants of Concanavalin A-Stimulated Murine Spleen Cells

Supernatants from Concanavalin A-stimulated murine spleen cells were subjected to hydrophobic interaction chromatography on phenyl-Sepharose. Macrophage cytotoxicity factor (MCF), macrophage migration inhibitory factor (MIF), T-helper cell-replacing factor (TRF) and colony-stimulating factor (CSF) were bound at high ionic strength and were released stepwise at low ionic strength. CSF thus could be separated from MCF, MIF and TRF and the bulk of other proteins. Chromatography of pools containing MCF, MIF and TRF on Sephadex did not lead to a separation of the three activities which were all found in a molecular weight range of 25.000-55.000. Isoelectric focusing of these pools in pH range from 4 to 9 gave two peaks for MCF at pH 8.2 and 7.2, whereas MIF activity focused from pH 4.5 to 5.5. TRF activity was found in a single sharp peak at pH 5.3. The results demonstrate that the four biological activities can be distinguished on a chemical basis and are accessible for purification and chemical characterization.     

Role for macrophage migration inhibitory factor in acute respiratory distress syndrome

The critical role of macrophage migration inhibitory factor (MIF) in mediating inflammatory lung injury in acute respiratory distress syndrome (ARDS) has been raised recently. The present study has identified enhanced MIF protein expression in alveolar capillary endothelium and infiltrating macrophages in lung tissues from ARDS patients. The possibility that MIF up-regulates its synthesis in an autocrine fashion in ARDS was tested using cultured endothelial cells stimulated with MIF and a murine model of lipopolysaccharide (LPS)-induced acute lung injury. MIF induced significant MIF and tumour necrosis factor (TNF)-alpha synthesis in cultured endothelial cells and the effect was blocked by neutralizing anti-MIF antibody. A similar blocking effect was observed when MIF-stimulated endothelial cells were pretreated with neutralizing anti-TNF-alpha antibody or glucocorticoid, supporting the notion that MIF induced TNF-alpha production via an amplifying pro-inflammatory loop. Treatment with anti-MIF or glucocorticoid effectively attenuated pulmonary pathology and the synthesis of MIF or TNF-alpha in mice with LPS-induced acute lung injury. Mildly augmented expression of aquaporin 1 (AQP1) was also detected in alveolar capillary endothelium in ARDS. In vitro studies revealed that both MIF and TNF-alpha induced a small increase of AQP1 synthesis in cultured endothelial cells. These findings suggest that MIF plays a crucial pathological role leading to alveolar inflammation in ARDS. Anti-MIF and early glucocorticoid therapy may represent a novel therapeutic approach for reducing alveolar inflammation in ARDS.     

Macrophage Migration Inhibitory Factor contributes to drive phenotypic and functional macrophages activation in response to Toxoplasma gondii infection

Cytokines are small molecules secreted by numerous cells. Macrophage Migration Inhibitory Factor (MIF) is a cytokine initially described due to its function of inhibiting random macrophage migration. Currently, new functions have been described for MIF, such as stimulating inflammatory functions in response to infections by microorganisms including, Toxoplasma gondii. However, the primordial MIF function related to macrophages has been little addressed. The main purpose of the study was to recapitulate MIF function on macrophages in response to T. gondii infection. To achieve this goal, peritoneal macrophages were collected from C57BL/6WT and Mif1^-/- mice after recruitment with thioglycolate. Macrophages were cultured, treated with 4-Iodo-6-phenylpyrimidine (4-IPP), and infected or not by T. gondii for 24 h. Following this, the culture supernatant was collected for cytokine, urea and nitrite analysis. In addition, macrophages were evaluated for phagocytic activity and T. gondii proliferation rates. Results demonstrated that T. gondii infection triggered an increase in MIF production in the WT group as well as an increase in the secretion of IL-10, TNF, IFN-γ, IL-6 and IL-17 in the WT and Mif1^-/- macrophages. Regarding the comparison between groups, it was detected that Mif1^-/- macrophages secreted more IL-10 compared to WT. On the other hand, the WT macrophages produced greater amounts of TNF, IFN-γ, IL-6 and IL-17. Urea production was more pronounced in Mif1^-/- macrophages while nitrite production was higher in WT macrophages. T. gondii showed a greater ability to proliferate in Mif1^-/- macrophages and these cells also presented enhanced phagocytic activity. In conclusion, T. gondii infection induces macrophage activation inciting cytokine production. In presence of MIF, T. gondii infected macrophages produce pro-inflammatory cytokines compatible with the M1 activation profile. MIF absence caused a dramatic reduction in pro-inflammatory cytokines that are balanced by increased levels of urea and anti-inflammatory cytokines. These macrophages presented increased phagocytic capacity and shared features activation with the M2 profile.     

Hormonal influences on the extinction of conditioned taste aversion

Conditioned taste aversion for a 5% glucose solution (sugar water) was induced in rats by an i.p. injection of LiCl 30 min after the first presentation of sugar water. Extinction of conditioned taste aversion was measured either in the forced-drinking test or in the preference-drinking test. In the forced-drinking test sugar water was the only fluid presented to the animals during extinction sessions. In the preference-drinking test the animals had the choice of tap water or sugar water. The rate of extinction was much slower in the preference test.The ACTH-analogues, ACTH_4–10 and ACTH_{4–10 7D Phe}, and -MSH delayed extinction in the preference test but not extinction in the forced-drinking test. ACTH_11–24 was without any effect. MSH-release inhibiting factor (MIF) facilitated extinction in the forced-drinking test but did not alter extinction in the preference test. The peptides did not affect intake of tap water or preference of sugar water over tap water by control rats.     

Bone formation in axial spondyloarthritis: Is disease modification possible?

Axial spondyloarthritis (SpA) is a chronic disease characterised by new bone formation (NBF) in the axial skeleton as well as at peripheral entheseal sites. NBF is thought to arise in areas of previous inflammation or osteitis visualised on MRI, with mechanical stress playing a role in disease pathogenesis. The interface between bone and immune cells is complex with the RANKL-OPG system being key to NBF. The IL-17/23 axis and other cytokines such as TNFα and MIF are thought to play a central role. The transition from inflammation to NBF is mediated via the Wnt, BMP and Hedgehog signalling pathways. An altered microbiome has been reported in SpA, which is a potential trigger of NBF in SpA. There is now data to show that treatment with TNF inhibitors prevents NBF and hence modifies disease progression. More research into identifying newer targets for disease modification is needed to alter the course of the disease.     

CD74: an emerging opportunity as a therapeutic target in cancer and autoimmune disease

Introduction: CD74, also known as the invariant chain, participates in several key processes of the immune system, including antigen presentation, B-cell differentiation and inflammatory signaling. Despite being described more than 3 decades ago, new functions and novel interactions for this evolutionarily conserved molecule are still being unraveled. As a participant in several immunological processes and an indicator of disease in some conditions, it has potential as a therapeutic target.

Areas covered: The relationship between the structure of CD74 variants and their physiological functions is detailed in this review. The function of CD74 in several cell lineages is examined with a focus on the interactions with cathepsins and, in an inflammatory milieu, the pro-inflammatory cytokine macrophage migratory inhibitory factor. The role of CD74 signaling in inflammatory and carcinogenic processes is outlined as is the use of CD74 as a therapeutic target (in cancer) and tool (as a vaccine).

Expert opinion: CD74 has several roles within the cell and throughout the immune system. Most prominent amongst these are the complex relationships with MIF and cathepsins. Modulation of CD74 function shows promise for the effective amelioration of disease.