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In recent years, the field of infectious diseases has been hit by the overwhelming amount of information generated while the human microbiome is being disentangled. Based on the interaction between the microbiota and the immune system, the implications regarding infectious diseases are probably major and remain a challenge. Aims: This review was conceived as a comprehensive tool to provide an overview of the available evidence regarding the influence of the microbiome on infectious diseases in children. Methods: We present the main findings aroused from microbiome research in prevention, diagnosis and treatment of infectious disease under a paediatric perspective, to inform clinicians of the potential relevance of microbiome‐related knowledge for translation to clinical practice. Results and Conclusion: The evidence shown in this review highlights the numerous research gaps ahead and supports the need to move forward to integrating the so‐called microbiome thinking into our routine clinical practice. [ABSTRACT FROM AUTHOR]

Objective CXCL12γ is an alternative splicing isoform of CXCL12 with enhanced affinity for heparan sulfate (HS) proteoglycans. This study was undertaken to investigate the distribution and potential function of CXCL12γ in rheumatoid arthritis (RA) synovium and normal lymphoid tissue, where its immobilization to HS may be relevant in pathologic or homeostatic immune cell migration and activation. Methods Expression of CXCL12 or CXCL12γ was immunodetected in RA and normal synovium, lymphoid tissue, and cultured cells with anti-pan-CXCL12 or anti-CXCL12γ-specific monoclonal antibodies. CXCL12α and CXCL12γ messenger RNA expression was analyzed by quantitative reverse transcription-polymerase chain reaction. Binding of wild-type CXCL12 isoforms or their HS binding-defective mutants to monocyte-derived dendritic cells (DCs) was analyzed by flow cytometry. The effect of DC-bound CXCL12α and CXCL12γ on T cell activation was analyzed in DC/T cell allogeneic cultures. Results CXCL12γ expression was increased in RA compared to normal synovium and preferentially located in endothelia and DC-SIGN-positive cells. This distribution was also observed in lymphoid organs. Surface-bound CXCL12γ was detected in a fraction of freshly isolated DCs. Monocyte-derived DCs, but not monocytes, showed a high capacity to bind CXCL12γ in an HS-dependent manner. Surface-bound CXCL12α and CXCL12γ on monocyte-derived DCs were potent inhibitors of allogeneic T cell activation, in contrast to the T cell-stimulatory effects of soluble CXCL12 proteins. Conclusion CXCL12γ shows a specific and similar distribution in RA synovium and lymphoid tissue, consistent with its higher HS binding affinity. Presentation of CXCL12 to T cells on membrane HS in DCs can play a distinct regulatory role in T cell activation. [ABSTRACT FROM AUTHOR]

Eur J Clin Invest 2010; 40 (6): 511–517 Background Iron is essential for mammalian metabolism and its cellular concentration is controlled by regulating its acquisition and storage. Haemochromatosis is a condition involving iron overload that is characterised by increased duodenal iron absorption and a progressive accumulation of iron in vital organs. Hepcidin is the main hormone that regulates iron homoestasis and it is secreted by the liver. Materials and methods We have studied how extended hepcidin administration affects the iron load status, plasma and tissue iron concentration, erythropoiesis and the expression of proteins involved on iron homeostasis in haemochromatotic ( Hfe −/− ) and wild-type mice. Results Hepcidin reverted the high plasma iron concentrations in Hfe −/− mice to normal values. The high concentration of hepatic iron was not altered in the liver of these Hfe −/− mice. Hepcidin administration did not disturb erythropoiesis in either Hfe −/− or wild-type mice and likewise, hepcidin did not modify the expression of any protein analysed in the liver, duodenum or spleen of Hfe −/− and wild-type mice. These data confirm that hepcidin administration diminishes plasma iron concentrations. Conclusion Treatment with sustained doses of hepcidin diminishes plasma iron concentrations in Hfe −/− mice. [ABSTRACT FROM AUTHOR]

Objective: Hyperplasia and phenotypic changes in fibroblasts are often observed in chronic inflammatory lesions, and yet the autonomous pathogenic contribution of these changes is uncertain. The purpose of this study was to analyze the intrinsic ability of fibroblasts from chronically inflamed synovial tissue to drive cell recruitment and angiogenesis., Methods: Fibroblasts from patients with rheumatoid arthritis (RA) or osteoarthritis (OA), as well as fibroblasts from healthy synovial tissue and healthy skin, were cultured and subcutaneously engrafted into immunodeficient mice. Cell infiltration and angiogenesis were analyzed in the grafts by immunohistochemical studies. The role of vascular endothelial growth factor (VEGF), CXCL12, and hypoxia-inducible transcription factor 1alpha (HIF-1alpha) in these processes was investigated using specific antagonists or small interfering RNA (siRNA)-mediated down-regulation of HIF-1alpha in fibroblasts., Results: Inflammatory (OA and RA) synovial fibroblasts, compared with healthy dermal or synovial tissue fibroblasts, induced a significant enhancement in myeloid cell infiltration and angiogenesis in immunodeficient mice. These activities were associated with increased constitutive and hypoxia-induced expression of VEGF, but not CXCL12, in inflammatory fibroblasts compared with healthy fibroblasts. VEGF and CXCL12 antagonists significantly reduced myeloid cell infiltration and angiogenesis. Furthermore, targeting of HIF-1alpha expression by siRNA or of HIF-1alpha transcriptional activity by the small molecule chetomin in RA fibroblasts significantly reduced both responses., Conclusion: These results demonstrate that chronic synovial inflammation is associated with stable fibroblast changes that, under hypoxic conditions, are sufficient to induce inflammatory cell recruitment and angiogenesis, both of which are processes relevant to the perpetuation of chronic inflammation.

Objective: Vasoactive intestinal peptide (VIP) has shown potent antiinflammatory effects in murine arthritis and ex vivo in human rheumatoid arthritis (RA) synovial cells. To investigate the potential endogenous participation of this system in the pathogenesis of RA, we analyzed the expression and regulation of VIP and its functional receptors in human fibroblast-like synoviocytes (FLS) from patients with osteoarthritis (OA) and patients with RA., Methods: The expression of VIP was studied by reverse transcription-polymerase chain reaction (RT-PCR), enzyme immunoassay, and immunofluorescence in cultured FLS, and by immunohistochemical analysis in synovial tissue. The expression and function of the potential VIP receptors in FLS were studied by RT-PCR, determination of intracellular cAMP production, cell membrane adenylate cyclase (AC) activity, and interleukin-6, CCL2, and CXCL8 production in response to VIP or specific agonists and antagonists., Results: VIP expression was detected in human FLS at the messenger RNA and protein levels, and it was significantly decreased in RA FLS compared with OA FLS. VIP receptor type 1 (VPAC1) was the dominant AC-coupled receptor in OA FLS, in contrast with RA FLS, in which VPAC2 was dominant. Tumor necrosis factor alpha-treated OA FLS reproduced the VIP and VPAC receptor expression pattern of RA FLS. The antagonistic effects of VIP on FLS proinflammatory factor production were reproduced by VPAC1- and VPAC2-specific agonists in OA FLS and RA FLS, respectively., Conclusion: VIP expression is down-regulated in RA and in tumor necrosis factor alpha-treated FLS, suggesting that down-regulation of this endogenous antiinflammatory factor may contribute to the pathogenesis of RA. In RA FLS, VPAC2 mediates the antiinflammatory effects of VIP, suggesting that VPAC2 agonists may be an alternative to VIP as antiinflammatory agents.

Objective: The expansion of an aggressive population of fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA) synovium occurs despite their expression of functional death receptors and exposure to death receptor ligands. FLS can survive Fas challenge because of the constitutive expression of FLIP apoptosis inhibitor. We investigated whether Fas signaling plays a pathogenetic role by activating a nonapoptotic proinflammatory program in RA FLS., Methods: Cultured RA FLS were stimulated with an agonistic anti-Fas antibody in the presence or absence of the caspase inhibitor Z-VAD-FMK or after RNA interference with a short hairpin RNA expression plasmid directed against FLIP. NF-kappaB and activator protein 1 (AP-1) activation was studied by electrophoretic mobility shift assays and p65 immunofluorescence analysis, and expression of messenger RNA (mRNA) for monocyte chemoattractant protein 1, interleukin-8, IkappaB alpha, and matrix metalloproteinases (MMPs) 1, 9, and 13 was examined by reverse transcription-polymerase chain reaction. Chemotactic activity of Fas-activated FLS-conditioned media was studied in Transwell migration assays., Results: Fas stimulation activated NF-kappaB and AP-1, and this response required caspase activity, since Z-VAD-FMK inhibitor precluded it. FLIP was processed to p43 protein after Fas stimulation in a caspase-dependent manner, and inhibition of FLIP expression resulted in reduced Fas-triggered NF-kappaB activation. Fas stimulation increased expression of mRNA for IkappaB alpha, MMPs, and chemokines, and Fas-activated RA FLS displayed increased chemotactic activity for monocytic cells., Conclusion: Fas triggering may contribute to the proinflammatory features of RA FLS by activating NF-kappaB and AP-1 and by expression of relevant target genes, such as MMPs and chemokines. Fas proinflammatory signaling is dependent upon caspase activity and FLIP expression. These data implicate FLIP as a potentially important molecular switch that turns the Fas signaling away from apoptosis and toward induction of a proinflammatory phenotype in RA FLS.

Objective: To assess whether an interleukin-1 receptor antagonist gene (IL1RN) polymorphism is associated with disease susceptibility and/or severity in a Spanish population of patients with rheumatoid arthritis (RA)., Methods: An 86-bp variable-number tandem repeat polymorphism within IL1RN intron 2 was analyzed by polymerase chain reaction in genomic DNA obtained from 247 unrelated patients with RA (group A) and 287 healthy control subjects. The polymorphism analysis was repeated in a second group of 194 patients with RA (group B). Clinical information from patients in group A was used to compare activity and severity data in patients stratified according to the different alleles or genotypes. Odds ratios (ORs) with 95% confidence intervals (95% CIs) were used to determine the strength of the association of the different alleles or genotypes with RA activity or severity., Results: In the control group, the allelic frequencies were 76% for IL1RN*1 (4 repeats), 21% for IL1RN*2 (2 repeats), 3% for IL1RN*3 (5 repeats), and 0.3% for IL1RN*4 (3 repeats). In group A patients with RA, both the frequency (OR 1.47, 95% CI 1.1-1.96, P = 0.007) and carriage rate (OR 1.6, 95% CI 1.1-2.2, P = 0.01) of allele IL1RN*2 were significantly increased. The increased frequency of IL1RN*2 was confirmed in group B patients with RA (OR 1.44, 95% CI 1.1-1.97, P = 0.01). In patients with RA, homozygosity for IL1RN*2 was associated with an increased number of affected articular areas during the first year of followup but not with other parameters of disease activity or severity., Conclusion: Our results suggest that IL1RN has a role in determining susceptibility to RA in the Spanish population.

Objective: Hyperplasia of fibroblast-like synoviocytes (FLS) contributes to chronic inflammation and joint destruction in rheumatoid arthritis (RA). FLICE-inhibitory protein (FLIP) is an antiapoptotic protein that might prevent apoptotic elimination of FLS in response to death ligands such as tumor necrosis factor alpha (TNFalpha) or Fas ligand, which are present in RA synovium. Previous studies on FLIP expression by osteoarthritis (OA) and RA FLS have shown variable results, and the specific role of FLIP as an apoptosis inhibitor in these cells remains unclear. We undertook this study to investigate the expression and antiapoptotic function of FLIP in FLS., Methods: We studied the expression of FLIP by immunohistochemistry and immunoblotting in synovial tissues or cultured FLS from RA and OA patients. FLS apoptosis was induced by an agonistic anti-Fas monoclonal antibody and FLS were then quantified. We studied the effects of cycloheximide (CHX), TNFalpha, and FLIP antisense oligonucleotide on FLIP expression and FLS apoptotic susceptibility., Results: FLIP(L) was the isoform mainly expressed in lining synoviocytes and cultured FLS. Synovial tissues and cultured FLS from OA and RA tissues displayed similar patterns and levels of expression of FLIP. Fas-induced apoptosis was variable in different FLS lines, but differences between OA and RA groups were not detected. TNFalpha induced increases in FLIP(L) and FLIP(S) expression and protected RA FLS from apoptosis, while CHX induced the opposite effects. Down-regulation of FLIP by antisense oligonucleotide strongly sensitized RA FLS to Fas-mediated apoptosis., Conclusion: Apoptosis susceptibility and FLIP expression are similar in OA and RA FLS. Down-regulation of FLIP sensitizes RA FLS to Fas-mediated apoptosis and may be a valuable tool for targeting RA FLS hyperplasia.