Immunoscintigraphic localization of inflammatory lesions: concept, radiolabelling and in vitro testing of a granulocyte specific antibody.

Current nuclear medicine techniques for the localization of inflammatory processes are based on injection of 111In labelled autologous granulocytes which need to be isolated and radiolabelled in vitro before reinjection. A new technique is presented here that obviates the need for cell isolation by the direct intravenous injection of a granulocyte specific 123I labelled monoclonal antibody. In this publication the basic parameters of the antibody granulocyte interaction are described. Antibody binding does not inhibit vital functions of the granulocytes, such as chemotaxis and superoxide generation. Scatchard analysis of binding data reveals an apparent affinity of the antibody for granulocytes of 6.8 X 10(9) l/mol and approximately 7.1 X 10(4) binding sites per cell. Due to the high specificity of the antibody, the only expected interference is from CEA producing tumors.

Alterations of bone microarchitecture in young patients with inflammatory bowel diseases are associated with fracture risk during growth

Aims: Inflammatory bowel diseases (IBD) appearing during childhood and adolescence compromise peak bone mass acquisition and increase fracture risk. The structural determinants of bone fragility in IBD however remain unknown. Methods: We investigated volumetric bone mineral density (vBMD), trabecular and cortical bone microstructure at distal radius and tibia by high-resolution pQCT (XtremeCT, Scanco, Switzerland), aBMD at distal radius, hip and spine and vertebral fracture assessment (VFA) by DXA in 107 young patients (mean age 22.8 yrs, range 12.2-33.7 yrs; 62 females and 45 males) with Crohn's disease (n=75), ulcerative colitis (n=25), undetermined colitis (n=2), and no definitive diagnosis (n=5), and in 389 healthy young individuals. Results: Mean disease duration was 6.1 yrs, 89/107 IBD patients received corticosteroids, 83 other immunomodulators, and 59 vitamin D. Clinical fractures were reported by 38 patients (mean age at 1st fracture, 12.6 yrs), the vast majority of the forearm, arm or hand; 5 had vertebral crush fractures (Grade 1 or 2) and 11 had multiple fractures. As compared to healthy controls (matched 2:1 for age, sex, height and fracture history), the 102 patients with established IBD had similar weight but significantly lower aBMD at all sites, lower trabecular (Tb) BV/TV and number, and greater Tb separation and inhomogeneous Tb distribution (1/SD TbN) at both distal radius and tibia, lower tibia cortical thickness (CTh), but no differences in cortical vBMD nor bone perimeter. Among IBD's, aBMD was not associated with fractures (by logistic regression adjusted for age, age square, sex, height, weight and protein intake). However, radius and tibia Tb BV/TV, thickness and SD 1/TbN, as well as radius Tb separation and perimeter, were significantly associated with fracture risk (fully adjusted as above), whereas cortical vBMD and CTh were not. After adjustment for aBMD at radius, respectively at femur neck, radius SD 1/TbN and tibia BV/TV, TbTh and perimeter remained independently associated with fracture risk. Conclusions: Young subjects with IBD have low bone mass and poor bone microarchitecture compared to healthy controls. Alterations of bone microarchitecture, particularly in the trabecular bone compartment, are specifically associated with increased fracture risk during growth.

The role of chemokines and chemokine receptors in eosinophil activation during inflammatory allergic reactions

Chemokines are important chemotactic cytokines that play a fundamental role in the trafficking of leukocytes to sites of inflammation. They are also potent cell-activating factors, inducing cytokine and histamine release and free radical production, a fact that makes them particularly important in the pathogenesis of allergic inflammation. The action of chemokines is regulated at the level of agonist production and processing as well as at the level of receptor expression and coupling. Therefore, an analysis of the ligands must necessarily consider receptors. Eosinophils are target cells involved in the allergic inflammatory response since they are able to release a wide variety of mediators including CC and CXC chemokines and express their receptors. These mediators could damage the airway epithelial cells and might be important to stimulate other cells inducing an amplification of the allergic response. This review focuses on recently emerging data pertaining to the importance of chemokines and chemokine receptors in promoting eosinophil activation and migration during the allergic inflammatory process. The analysis of the function of eosinophils and their chemokine receptors during allergic inflammation might be a good approach to understanding the determinants of asthma severity and to developing novel therapies.

Rôle différentiel des cellules épithéliales intestinales et pulmonaires dans le recrutement des cellules Th17 vers les sites de réplication du virus de l'immunodéficience humaine de type 1

L’infection à VIH-1 est associée à une forte déplétion des lymphocytes T CD4+ à polarisation Th17 au niveau des tissus lymphoïdes associés aux muqueuses intestinales (GALT, gut-associated lymphoid tissues). Ceci conduit à la translocation microbienne, qui est une cause d’activation immunitaire chronique et de progression de la maladie. Les cellules épithéliales (CE) jouent un rôle critique dans le maintien de l’intégrité et de l’homéostasie au niveau des muqueuses intestinales via le recrutement des cellules de l’immunité innée (e.g., neutrophiles) et adaptative (e.g., cellules Th17). Les neutrophiles produisent des molécules antivirales (e.g., défensines-) et ont la capacité de limiter la réplication virale au niveau des muqueuses. Les cellules Th17 jouent un double rôle lors de l’infection à VIH. Elles contribuent d’une part à la défense contre différents pathogènes opportunistes en augmentant, via la production d’IL-17, la capacité des CE à attirer les cellules Th17 et les neutrophiles. D’autre part, les cellules Th17 jouent un rôle délétère en tant que cibles de réplication virale et sources de cytokines pro-inflammatoires. La fréquence des cellules Th17 est diminuée dans les GALT mais pas dans les poumons des patients infectés par le VIH, suggérant qu’il existe des mécanismes différents par lesquels les cellules Th17 sont recrutées vers ces sites anatomiques. Nous avons testé l’hypothèse selon laquelle le VIH interfère avec la capacité des CE intestinales et non pas pulmonaires à produire des chimiokines (CK) responsables de l’attraction des cellules Th17 et des neutrophiles. Nous avons démontré que les CE intestinales et pulmonaires produisent des CK spécifiques pour les cellules Th17 (CCL20) et les neutrophiles (CXCL8) en réponse à des stimuli pro-inflammatoires tels que l’IL-1 et le TNF-. Le TNF- agit en synergie avec l’IL-17, un « signal de danger » récemment identifié, et augmente la capacité des CE intestinales mais pas pulmonaires à produire la chimiokine CCL20. Cette synergie s’explique par l’augmentation préférentielle de l’expression du récepteur à l’IL-17 à la surface des CE intestinales suite à la stimulation par le TNF-. L’exposition au VIH n’affecte pas la production de CCL20 et de CXCL8 par les CE intestinales, mais altère la capacité des CE alvéolaires à produire ces chimiokines en accord avec la permissivité sélective de ces dernières à l’infection par le VIH. En conclusion, nos résultats démontrent que (i) le VIH n’interfère pas directement avec la capacité des CE intestinales à recruter des cellules Th17 et des neutrophils et que (ii) la production de CCL20 par ces cellules est dépendantes de la synergie entre le TNF- et l’IL-17. Ainsi, la déplétion des cellules Th17 et la pénurie en IL-17 dans les GALT des sujets infectés pourrait causer de façon préférentielle des altérations fonctionnelles au niveau des CE intestinales, se traduisant par l’altération du recrutement des cellules Th17 en réponse au CCL20.

Loss of 3-chlorotyrosine by inflammatory oxidants: Implications for the use of 3-chlorotyrosine as a bio-marker in vivo

Activated neutrophils generate the potent oxidant hypochlorous acid (HOCl) from the enzyme myeloperoxidase (MPO). A proposed bio-marker for MPO-derived HOCl in vivo is 3-chlorotyrosine, elevated levels of which have been measured in several human inflammatory pathologies. However, it is unlikely that HOCl is produced as the sole oxidant at sites of chronic inflammation as other reactive species are also produced during the inflammatory response. The work presented shows that free and protein bound 3-chlorotyrosine is lost upon addition of the pro-inflammatory oxidants, HOCl, peroxynitrite, and acidified nitrite. Furthermore, incubation of 3-chlorotyrosine with activated RAW264.7 macrophages or neutrophil-like HL-60 cells resulted in significant loss of 3-chlorotyrosine. Therefore, at sites of chronic inflammation where there is concomitant ONOO- and HOCl formation, it is possible measurement of 3-chlorotyrosine may represent an underestimate of the true extent of tyrosine chlorination. This finding could account for some of the discrepancies reported between 3-chlorotyrosine levels in tissues in the literature. (c) 2008 Elsevier Inc. All rights reserved.

The pro-inflammatory oxidant hypochlorous acid induces Bax-dependent mitochondrial permeabilisation and cell death through AIF-/EndoG-dependent pathways

At sites of chronic inflammation, such as in the inflamed rheumatoid joint, activated neutrophils release hydrogen peroxide (H2O2) and the enzyme myeloperoxidase to catalyse the formation of hypochlorous acid (HOCl). 3-chlorotyrosine, a marker of HOCl in vivo, has been observed in synovial fluid proteins from rheumatoid arthritis patients. However the mechanisms of HOCl-induced cytotxicity are unknown. We determined the molecular mechanisms by which HOCl induced cell death in human mesenchymal progenitor cells (MPCs) differentiated into a chondrocytic phenotype as a model of human cartilage cells and show that HOCl induced rapid Bax conformational change, mitochondrial permeability and release of intra-mitochondrial pro-apoptotic proteins which resulted in nuclear translocation of AIF and EndoG. siRNA-mediated knockdown of Bax substantially prevented mitochondrial permeability, release of intra-mitochondrial pro-apoptotic proteins. Cell death was inhibited by siRNA-mediated knockdown of Bax, AIF or EndoG. Although we observed several biochemical markers of apoptosis, caspase activation was not detected either by western blotting, fluorescence activity assays or by using caspase inhibitors to inhibit cell death. This was further supported by findings that (1) in vitro exposure of recombinant human caspases to HOCl caused significant inhibition of caspase activity and (2) the addition of HOCl to staurosporine-treated MPCs inhibited the activity of cellular caspases. Our results show for the first time that HOCl induced Bax-dependent mitochondrial permeability which led to cell death without caspase activity by processes involving AIF/EndoG-dependent pathways. Our study provides a novel insight into the potential mechanisms of cell death in the inflamed human joint. (c) 2006 Elsevier Inc. All rights reserved.

Oncostatin M is a novel glucocorticoid-dependent neuroinflammatory factor that enhances oligodendrocyte precursor cell activity in demyelinated sites

The innate immune reaction to tissue injury is a natural process, which can have detrimental effects in the absence of negative feedbacks by glucocorticoids (GCs). Although acute lipopolysaccharide (LPS) challenge is relatively harmless to the brain parenchyma of adult animals, the endotoxin is highly neurotoxic in animals that are treated with the GC receptor antagonist RU486. This study investigated the role of cytokines of the gp130-related family in these effects, because they are essential components of the inflammatory process that provide survival signals to neurons. Intracerebral LPS injection stimulated expression of several members of this family of cytokines, but oncostatin M (Osm) was the unique ligand to be completely inhibited by the RU486 treatment. OSM receptor (Osmr) is expressed mainly in astrocytes and endothelial cells following LPS administration and GCs are directly responsible for its transcriptional activation in the presence of the endotoxin. In a mouse model of demyelination, exogenous OSM significantly modulated the expression of genes involved in the mobilization of oligodendrocyte precursor cells (OPCs), differentiation of oligodendrocyte, and production of myelin. In conclusion, the activation of OSM signaling is a mechanism activated by TLR4 in the presence of negative feedback by GCs on the innate immune system of the brain. OSM absence is associated with detrimental effects of LPS, whereas exogenous OSM favors repair response to demyelinated regions. (C) 2010 Elsevier Inc. All rights reserved.

The role of chemokines and chemokine receptors in eosinophil activation during inflammatory allergic reactions

Chemokines are important chemotactic cytokines that play a fundamental role in the trafficking of leukocytes to sites of inflammation. They are also potent cell-activating factors, inducing cytokine and histamine release and free radical production, a fact that makes them particularly important in the pathogenesis of allergic inflammation. The action of chemokines is regulated at the level of agonist production and processing as well as at the level of receptor expression and coupling. Therefore, an analysis of the ligands must necessarily consider receptors. Eosinophils are target cells involved in the allergic inflammatory response since they are able to release a wide variety of mediators including CC and CXC chemokines and express their receptors. These mediators could damage the airway epithelial cells and might be important to stimulate other cells inducing an amplification of the allergic response. This review focuses on recently emerging data pertaining to the importance of chemokines and chemokine receptors in promoting eosinophil activation and migration during the allergic inflammatory process. The analysis of the function of eosinophils and their chemokine receptors during allergic inflammation might be a good approach to understanding the determinants of asthma severity and to developing novel therapies.

Evaluation of subcutaneous and alveolar implantation surgical sites in the study of the biological properties of root-end filling endodontic materials

Objective: The aim of this study was to compare two methodologies used in the evaluation of tissue response to root-end filling materials in rats. Material and Methods: Forty rats were divided into 4 groups: in Groups I and II (control groups), empty polyethylene tubes were implanted in the extraction site and in the subcutaneous tissue, respectively; in Groups III and IV, polyethylene tubes filled with ProRoot MTA were implanted in the extraction site and in the subcutaneous tissue, respectively. The animals were killed 7 and 30 days after tube implantation, and the hemi-maxillas and the capsular subcutaneous tissue, both with the tubes, were removed. Specimens were processed and evaluated histomorphologicaly under light microscopy. The scores obtained were analyzed statistically by the Kruskal-Wallis test (p<0.05). Results: There were no statistically significant differences between the implantation methods (p=0.78033, p=0.72039). It was observed that the 30-day groups presented a more mature healing process due to smaller number of inflammatory cells. Conclusions: The present study showed no differences in tissue responses as far as the implantation site and the studied period were concerned. Alveolar socket implantation methodology represents an interesting method in the study of the biological properties of root-end filling endodontic materials due to the opportunity to evaluate bone tissue response.

MAP Kinase Phosphatase-1 Protects against Inflammatory Bone Loss

The mitogen-activated protein (MAP) kinase phosphatase (MKP) family plays an important function in regulating the pro-inflammatory cytokines by deactivating MAP kinases. MKP-1 is essential for the dephosphorylation of p38 MAP kinase that regulates expression of IL-6, TNF-alpha, and IL-1 beta. We hypothesized that MKP-1 regulates inflammatory bone loss in experimental periodontitis. Wild-type and Mkp-1(-/-) mice received A. actinomycetemcomitans LPS injection in the palatal region or PBS control 3 times/wk for 30 days. Mice were killed, and maxillae were assessed by microcomputed tomography, histological analysis, and TRAP staining for measurement of bone loss, extent of inflammation, and degree of osteoclastogenesis. Results indicated that, in LPS-injected Mkp-1(-/-) mice, significantly greater bone loss occurred with more inflammatory infiltrate and a significant increase in osteoclastogenesis compared with Mkp-1(-/-) control sites or either wild-type group. Analysis of these data indicates that MKP-1 plays a key role in the regulation of inflammatory bone loss.

Non-inflammatory destructive periodontal disease: a clinical, microbiological, immunological and genetic investigation

Periodontitis comprises a group of multifactorial diseases in which periodontopathogens accumulate in dental plaque and trigger host chronic inflammatory and immune responses against periodontal structures, which are determinant to the disease outcome. Although unusual cases of non-inflammatory destructive periodontal disease (NIDPD) are described, their pathogenesis remains unknown. A unique NIDPD case was investigated by clinical, microbiological, immunological and genetic tools. The patient, a non-smoking dental surgeon with excessive oral hygiene practice, presented a generalized bone resorption and tooth mobility, but not gingival inflammation or occlusion problems. No hematological, immunological or endocrine alterations were found. No periodontopathogens (A. actinomycetemcomitans, P. gingivalis, F. nucleatum and T. denticola) or viruses (HCMV, EBV-1 and HSV-1) were detected, along with levels of IL-1 beta and TNF-alpha in GCF compatible with healthy tissues. Conversely ALP, ACP and RANKL GCF levels were similar to diseased periodontal sites. Genetic investigation demonstrated that the patient carried some SNPs, as well HLA-DR4 (*0404) and HLA-B27 alleles, considered risk factors for bone loss. Then, a less vigorous and diminished frequency of toothbrushing was recommended to the patient, resulting in the arrest of alveolar bone loss, associated with the return of ALP, ACP and RANKL in GCF to normality levels. In conclusion, the unusual case presented here is compatible with the previous description of NIDPD, and the results that a possible combination of excessive force and frequency of mechanical stimulation with a potentially bone loss prone genotype could result in the alveolar bone loss seen in NIDPD.

Acute abdomen caused by a perforated inflammatory myofibroblastic tumor of the jejunum

Inflammatory myofibroblastic tumors are rare pseudosarcomatous tumors found in virtually all anatomic sites. Our case report describes an elderly female patient with an inflammatory myofibroblastic tumor in the proximal jejunum, accidentally discovered at laparotomy for an acute abdomen. The localization in the jejunum is a very rare finding, and perforation has not been described before.

Targeting the sphingosine kinase/sphingosine 1-phosphate pathway to treat chronic inflammatory kidney diseases

Chronic kidney diseases including glomerulonephritis are often accompanied by acute or chronic inflammation that leads to an increase in extracellular matrix (ECM) production and subsequent glomerulosclerosis. Glomerulonephritis is one of the leading causes for end-stage renal failure with high morbidity and mortality, and there are still only a limited number of drugs for treatment available. In this MiniReview, we discuss the possibility of targeting sphingolipids, specifically the sphingosine kinase 1 (SphK1) and sphingosine 1-phosphate (S1P) pathway, as new therapeutic strategy for the treatment of glomerulonephritis, as this pathway was demonstrated to be dysregulated under disease conditions. Sphingosine 1-phosphate is a multifunctional signalling molecule, which was shown to influence several hallmarks of glomerulonephritis including mesangial cell proliferation, renal inflammation and fibrosis. Most importantly, the site of action of S1P determines the final effect on disease progression. Concerning renal fibrosis, extracellular S1P acts pro-fibrotic via activation of cell surface S1P receptors, whereas intracellular S1P was shown to attenuate the fibrotic response. Interference with S1P signalling by treatment with FTY720, an S1P receptor modulator, resulted in beneficial effects in various animal models of chronic kidney diseases. Also, sonepcizumab, a monoclonal anti-S1P antibody that neutralizes extracellular S1P, and a S1P-degrading recombinant S1P lyase are promising new strategies for the treatment of glomerulonephritis. In summary, especially due to the bifunctionality of S1P, the SphK1/S1P pathway provides multiple target sites for the treatment of chronic kidney diseases.

Subgingival microflora in inflammatory bowel disease patients with untreated periodontitis

OBJECTIVE To analyze the subgingival microflora composition of inflammatory bowel disease (IBD) patients with untreated chronic periodontitis and compare them with systemically healthy controls also having untreated chronic periodontitis. METHOD Thirty IBD patients [15 with Crohn's disease (CD) and 15 with ulcerative colitis (UC)] and 15 control individuals participated in the study. All patients had been diagnosed with untreated chronic periodontitis. From every patient, subgingival plaque was collected from four gingivitis and four periodontitis sites with paper points. Samples from the same category (gingivitis or periodontitis) in each patient were pooled together and stored at -70 °C until analysis using a checkerboard DNA-DNA hybridization technique for 74 bacterial species. RESULTS Multiple-comparison analysis showed that the groups differed in bacterial counts for Bacteroides ureolyticus, Campylobacter gracilis, Parvimonas micra, Prevotella melaninogenica, Peptostreptococcus anaerobius, Staphylococcus aureus, Streptococcus anginosus, Streptococcus intermedius, Streptococcus mitis, Streptococcus mutans, and Treponema denticola (P<0.001). CD patients had significantly higher levels of these bacteria than UC patients either in gingivitis or in periodontitis sites (P<0.05). CD patients harbored higher levels of P. melaninogenica, S. aureus, S. anginosus, and S. mutans compared with controls both at gingivitis and at periodontitis sites (P<0.05). UC patients harbored higher levels of S. aureus (P=0.01) and P. anaerobius (P=0.05) than controls only in gingivitis sites. CONCLUSION Our study showed that even with similar clinical periodontal parameters, IBD patients harbor higher levels of bacteria that are related to opportunistic infections in inflamed subgingival sites that might be harmful for the crucial microbe-host interaction.

Periodontitis and Gingivitis in Inflammatory Bowel Disease

BACKGROUND: The oral cavity is frequently affected in patients with inflammatory bowel disease (IBD), especially in patients with Crohn's disease (CD). Periodontitis is thought to influence systemic autoimmune or inflammatory diseases. We aimed to analyze the relationship of periodontitis and gingivitis markers with specific disease characteristics in patients with IBD and to compare these data with healthy controls. METHODS: In a prospective 8-month study, systematic oral examinations were performed in 113 patients with IBD, including 69 patients with CD and 44 patients with ulcerative colitis. For all patients, a structured personal history was taken. One hundred thirteen healthy volunteers served as a control group. Oral examination focussed on established oral health markers for periodontitis (bleeding on probing, loss of attachment, and periodontal pocket depth) and gingivitis (papilla bleeding index). Additionally, visible oral lesions were documented. RESULTS: Both gingivitis and periodontitis markers were higher in patients with IBD than in healthy control. In univariate analysis and logistic regression analysis, perianal disease was a risk factor for periodontitis. Nonsmoking decreased the risk of having periodontitis. No clear association was found between clinical activity and periodontitis in IBD. In only the CD subgroup, high clinical activity (Harvey-Bradshaw index > 10) was associated with 1 periodontitis marker, the loss of attachment at sites of maximal periodontal pocket depth. Oral lesions besides periodontitis and gingivitis were not common, but nevertheless observed in about 10% of patients with IBD. CONCLUSIONS: IBD, and especially perianal disease in CD, is associated with periodontitis. Optimal therapeutic strategies should probably focus on treating both local oral and systemic inflammation.