Emilio Jirillo

Enteric bacteria, lipopolysaccharides and related cytokines in inflammatory bowel disease: biological and clinical significance.

Ulcerative colitis (UC) and Crohns disease (CD) [inflammatory bowel disease (IBD)] are both characterized by an exaggerated immune response at the gut associated lymphoreticular tissue level. Such an abnormal and dysregulated immune response may be directed against luminal and/or enteric bacterial antigens, as also supported by murine models of inflammatory bowel disease (IBD) caused by organisms such as Citrobacter rodentium and Helicobacter hepaticus. Bacterial endotoxins or lipopolysaccharides (LPS) have been detected in the plasma of IBD patients and an abnormal microflora and/or an increased permeability of the intestinal mucosa have been invoked as cofactors responsible for endotoxemia. At the same time, the evidence that phagocytosis and killing exerted by polymorphonuclear cells and monocytes and the T-cell dependent antibacterial activity are decreased in IBD patients may also explain the origin of LPS in these diseases. In IBD, pro-inflammatory cytokines and chemokines have been detected in elevated amounts in mucosal tissue and/or in peripheral blood, thus suggesting a monocyte/macrophage stimulation by enteric bacteria and/or their constituents (e.g. LPS). On these grounds, in experimental models and in human IBD, anti-cytokine monoclonal antibodies and interleukin receptor antagonists are under investigation for their capacity to neutralize the noxious effects of immune mediators. Finally, the administration of lactobacilli is beneficial in human IBD and, in murine colitis, this treatment leads to a normalization of intestinal flora, reducing the number of colonic mucosal adherent and translocated bacteria.

Leptin as an immunomodulator

Leptin is an adipocyte-derived hormone/cytokine that links nutritional status with neuroendocrine and immune functions. In humans, leptin influences energy homeostasis and regulates neuroendocrine function primarily in states of energy deficiency. Initially described as an antiobesity hormone, leptin has subsequently been shown also to influence basal metabolism, hematopoiesis, thermogenesis, reproduction, and angiogenesis. As a cytokine, leptin can affect thymic homeostasis and the secretion of acute-phase reactants such as interleukin-1 (IL-1) and tumor-necrosis factor-alpha (TNF-α). Leptin links nutritional status and proinflammatory T helper 1 (Th1) immune responses and the decrease in leptin plasma concentration during food deprivation leads to impaired immune function. Similar to other pro-inflammatory cytokines, leptin promotes Th1-cell differentiation and can modulate the onset and progression of autoimmune responses in several animal models of disease. Here, we review the advances and controversy for a role of leptin in the pathophysiology of immune responses and discuss novel possible therapeutic implications for leptin modulators.

The role of the liver in the response to LPS: experimental and clinical findings

The liver plays an important physiological role in lipopolysaccharide (LPS) detoxification and, in particular, hepatocytes are involved in the clearance of endotoxin of intestinal derivation. In experimental shock models, tumor necrosis factor (TNF)-α induces hepatocyte apoptosis and lethal effects are due to secreted TNF-α and not to cell-associated TNF-α. An exaggerated production of TNF-α has been reported in murine viral infections, in which mice become sensitized to low amounts of LPS and both interferon (IFN)-γ and IFN-α/β are involved in the macrophage-induced release of TNF-α. The prominent role of LPS and TNF-α in liver injury is also supported by studies of ethanol-induced hepatic damage. In humans, evidence of LPS-induced hepatic injury has been reported in cirrhosis, autoimmune hepatitis, and primary biliary cirrhosis and a decreased phagocytic activity of the reticulo-endothelial system has been found in these diseases. The origin of endotoxemia in hepatitis C virus (HCV) infected patients seems to be multifactorial and LPS may be of exogenous or endogenous derivation. In endotoxemic HCV-positive patients responsive to a combined treatment with IFN-α/ribavirin (RIB), endotoxemia was no longer detected at the end of the therapeutic regimen. By contrast, 48% of the non-responders to this treatment were still endotoxemic and their monocytes displayed higher intracellular TNF-α and interleukin (IL)-1β levels than responders. Moreover, in responders, an equilibrium between IFN-γ and IL-10 serum levels was attained. In the non-responders, serum levels of IL-10 did not increase following treatment. This may imply that an imbalance between T helper (Th)1 and Th2 derived cytokines could be envisaged in the non-responders.

Salmonid T cells assemble in the thymus, spleen and in novel interbranchial lymphoid tissue

In modern bony fishes, or teleost fish, the general lack of leucocyte markers has greatly hampered investigations of the anatomy of the immune system and its reactions involved in inflammatory responses. We have previously reported the cloning and sequencing of the salmon CD3 complex, molecules that are specifically expressed in T cells. Here, we generate and validate sera recognizing a peptide sequence of the CD3ε chain. Flow cytometry analysis revealed high numbers of CD3ε+ or T cells in the thymus, gill and intestine, whereas lower numbers were detected in the head kidney, spleen and peripheral blood leucocytes. Subsequent morphological analysis showed accumulations of T cells in the thymus and spleen and in the newly discovered gill‐located interbranchial lymphoid tissue. In the latter, the T cells are embedded in a meshwork of epithelial cells and in the spleen, they cluster in the white pulp surrounding ellipsoids. The anatomical organization of the salmonid thymic cortex and medulla seems to be composed of three layers consisting of a sub‐epithelial medulla‐like zone, an intermediate cortex‐like zone and finally another cortex‐like basal zone. Our study in the salmonid thymus reports a previously non‐described tissue organization. In the intestinal tract, abundant T cells were found embedded in the epithelium. In non‐lymphoid organs, the presence of T cells was limited. The results show that the interbranchial lymphoid tissue is quantitatively a very important site of T cell aggregation, strategically located to facilitate antigen encounter. The interbranchial lymphoid tissue has no resemblance to previously described lymphoid tissues.

Antimicrobial and immunoregulatory functions of lactoferrin and its potential therapeutic application.

Lactoferrin is an iron-binding glycoprotein present in various secretions (e.g. milk, tears, saliva, pancreatic juice, etc.). It is also stored in specific granules of polymorphonuclear granulocytes from which it is released following activation. Lactoferrin exerts a bactericidal activity by damagingthe outer membrane of Gram-negative bacteria, as well as immunoregulatory functions by decreasing the release of interleukin-1 (IL-1), IL-2 and tumor necrosis factor- (TNF-) and enhancing monocyte and natural killer cell cytotoxicity. Lactoferrin binds with high affinity to lipid A, the toxic moiety of the lipopolysaccharide, or endotoxin from Gram-negative bacteria. Lipopolysaccharide interaction with monocytes/macrophages results in the production and release of TNF- , that plays an important role in inducing septic shock. In this respect, it has recently been demonstrated that lactoferrin inhibits the lipopolysaccharide interaction with CD14 on monocytes/macrophages by competition with the lipopolysaccharide binding protein. Therefore, besides its bactericidal activity, lactoferrin may also act by neutralizing the toxic effects of lipopolysaccharide and this protective role against endotoxin lethal shock has been demonstrated in animal models. Moreover, in vitro and in vivo neutralization of endotoxin by a human lactoferrin-derived peptide was also reported and lactoferrin or lactoferrin-derived peptides could represent useful tools for the treatment of endotoxin-induced septic shock. The recent production and characterization of monoclonal antibodies against different epitopes of human lactoferrin, including monoclonal antibodies selectively neutralizinglactoferrin binding to lipid A, may allow a better elucidation of the consequence of lactoferrin-lipopolysaccharideinteraction.

Vaccination-Induced Systemic Autoimmunity in Farmed Atlantic Salmon

Over half of the salmon consumed globally are farm-raised. The introduction of oil-adjuvanted vaccines into salmon aquaculture made large-scale production feasible by preventing infections. The vaccines that are given i.p. contain oil adjuvant such as mineral oil. However, in rodents, a single i.p. injection of adjuvant hydrocarbon oil induces lupus-like systemic autoimmune syndrome, characterized by autoantibodies, immune complex glomerulonephritis, and arthritis. In the present study, whether the farmed salmon that received oil-adjuvanted vaccine have autoimmune syndrome similar to adjuvant oil-injected rodents was examined. Sera and tissues were collected from vaccinated or unvaccinated Atlantic salmon (experimental, seven farms) and wild salmon. Autoantibodies (immunofluorescence, ELISA, and immunoprecipitation) and IgM levels (ELISA) in sera were measured. Kidneys and livers were examined for pathology. Autoantibodies were common in vaccinated fish vs unvaccinated controls and they reacted with salmon cells/Ags in addition to their reactivity with mammalian Ags. Diffuse nuclear/cytoplasmic staining was common in immunofluorescence but some had more specific patterns. Serum total IgM levels were also increased in vaccinated fish; however, the fold increase of autoantibodies was much more than that of total IgM. Sera from vaccinated fish immunoprecipitated ferritin and ∼50% also reacted with other unique proteins. Thrombosis and granulomatous inflammation in liver, and immune-complex glomerulonephritis were common in vaccinated fish. Autoimmunity similar to the mouse model of adjuvant oil-induced lupus is common in vaccinated farmed Atlantic salmon. This may have a significant impact on production loss, disease of previously unknown etiology, and future strategies of vaccines and salmon farming.

Non-specific immunity in aging: Deficiency of monocyte and polymorphonuclear cell-mediated functions

Peripheral blood monocytes obtained from 55 aged donors were evaluated for their chemotactic and phagocytic capacity. In the same subjects, polymorphonuclear cell-mediated functions were studied by chemotaxis, phagocytosis, nylon fiber adherence and nitroblue-tetrazolium reduction assay. Monocytes showed a normal chemotactic responsiveness to zymosan-activated serum, while the chemotactic activity induced by leukocyte-derived chemotactic factor and phagocytosis were rather depressed. A dramatic impairment of polymorphonuclear cell-mediated immune response was also observed. In fact, in spite of a normal nylon fiber adherence, chemotaxis, phagocytosis and nitroblue-tetrazolium reduction capacity were significantly depressed by the aging process. These data suggest that the deficiency of non-specific immunity may play an important role in the increased susceptibility to infections in aged donors.

Effect of Bifidobacterium bifidum and Lactobacillus acidophilus on gut mucosa and peripheral blood B lymphocytes

In 15 elderly individuals lyophilized Bifidobacterium bifidum (BB) and Lactobacillus acidophilus (LA) (Infloran) were administered in capsules (two capsules 4 times per day) for 28 days, while in 10 elderly controls placebo were given the same posology and for an equal period of time. The effects of this treatment on the immune system both at the periphery or the intestinal level were investigated. Results show that BB and LA significantly reduced the colonic inflammatory infiltration, without altering T, B and Leu7 + cell percentage. At the same time, a significant increase of B cell frequency in the peripheral blood was noted, in comparison to controls. The overall results suggest that the regular administration of BB and LA leads to a modulation of the immunological and inflammatory response in elderly subjects.

Low Grade Inflammation as a Common Pathogenetic Denominator in Age-Related Diseases: Novel Drug Targets for Anti-Ageing Strategies and Successful Ageing Achievement

Nowadays, people are living much longer than they used to do, however they are not free from ageing. Ageing, an inexorable intrinsic process that affects all cells, tissues, organs and individuals, is a post-maturational process that, due to a diminished homeostasis and increased organism frailty, causes a reduction of the response to environmental stimuli and, in general, is associated to an increased predisposition to illness and death. However, the high incidence of death due to infectious, cardiovascular and cancer diseases underlies a common feature in these pathologies that is represented by dysregulation of both instructive and innate immunity. Several studies show that a low-grade systemic inflammation characterizes ageing and that inflammatory markers are significant predictors of mortality in old humans. This pro-inflammatory status of the elderly underlies biological mechanisms responsible for physical function decline and age-related diseases such as Alzheimers disease and atherosclerosis are initiated or worsened by systemic inflammation. Understanding of the ageing process should have a prominent role in new strategies for extending the health old population. Accordingly, as extensively discussed in the review and in the accompanying related papers, investigating ageing pathophysiology, particularly disentangling age-related low grade inflammation, is likely to provide important clues about how to develop drugs that can slow or delay ageing.

Phagocytosis, killing, lymphocyte-mediated antibacterial activity, serum autoantibodies, and plasma endotoxins in inflammatory bowel Disease

OBJECTIVE:Alteration of mucosal and systemic immune responses may play an important role in the pathogenesis of inflammatory bowel disease (IBD). The aim of this study was to evaluate natural immune responses (i.e., phagocytosis, killing, and antibacterial activity), serum autoantibodies (antineutrophil cytoplasmic antibodies [ANCA] and antilactoferrin [LF] antibodies), and plasma endotoxins in patients affected by ulcerative colitis (UC) and Crohns disease (CD).METHODS:Blood samples were obtained from 71 patients with UC, 32 patients with CD, and 32 control subjects. Disease activity was scored using Trueloves criteria in patients with UC and the Crohns Disease Activity Index (CDAI) in patients with CD. Candida albicans served as a target for evaluation of phagocytosis and killing exerted by polymorphonuclear cells (PMN) and monocytes (MØ), whereas Salmonella typhi was used for assessing lymphocyte-mediated antibacterial activity. ANCA were detected by indirect immunofluorescence, whereas anti-LF antibodies were assayed by means of enzyme-linked immunosorbent assay. Plasma endotoxins were measured by Limulus amoebocyte lysate assay.RESULTS:Phagocytosis and killing exerted by PMN and MØ, as well as lymphocyte-mediated antibacterial activity, were significantly reduced (p < 0.0001) in patients affected by UC and CD in comparison with controls, irrespective of either disease activity or treatment. Plasma endotoxins were detected in 12/71 (17%) patients with UC, and in 10/32 (31%) patients with CD. ANCA were present in 42/71 (59%) patients with UC and in 3/32 (9%) patients with CD, whereas anti-LF antibodies were detected in 31 (44%) UC patients and in six (19%) CD patients. No significant differences in phagocytosis and killing exerted by PMN were found between ANCA-positive and ANCA-negative UC patients.CONCLUSIONS:Our data demonstrate an impairment of natural immunity exerted by peripheral blood phagocytes and lymphocytes in patients with UC and CD. ANCA and anti-LF antibodies were present mainly in UC patients but their presence did not affect PMN-mediated phagocytosis and killing. Finally, plasma endotoxins may contribute to the chronic inflammatory status, likely by inducing release of proinflammatory mediators.