In vivo pre-activation of monocytes in patients with axial spondyloarthritis.

INTRODUCTION: Innate immune responses, including monocyte functions, seem to play an important role in the pathogenesis of axial spondyloarthritis (axSpA). Therefore, we characterized the phenotype and functional state of monocytes of patients with axSpA. METHODS: Fifty-seven patients with axSpA, 11 patients with rheumatoid arthritis (RA), and 29 healthy controls were included in the study. We determined the percentage of classic, intermediate, and non-classic monocytes according to CD14 and CD16 expression and the expression of Toll-like receptor (TLR) 1, 2, and 4 in whole blood by flow cytometry. The percentage of monocytes producing interleukin (IL)-1beta, IL-6, tumor necrosis factor alpha (TNFα), IL-12/23p40, and IL-1 receptor antagonist (IL-1ra) was detected by flow cytometry after stimulation of whole blood without and with different TLR and nucleotide-binding oligomerization domain ligands-i.e., lipopolysaccharide (LPS), fibroblast-stimulating lipopeptid-1, PAM3CSK4, and muramyl dipeptide (MDP)-for 5 h. IL-10 production was measured after 18 h of stimulation in supernatants by enzyme-linked immunosorbent assay. RESULTS: In patients with axSpA but not patients with RA, we found higher frequencies of classic monocytes than in controls (median of 90.4% versus 80.4%, P < 0.05), higher frequencies of monocytes spontaneously producing IL-1beta and IL-1ra (P < 0.05), and a higher percentage of monocytes producing IL-1beta after MDP stimulation (P < 0.05). Elevated cytokine production was confined to axSpA patients under conventional therapy (non-steroidal anti-inflammatory drugs) and not found in patients under TNFα inhibitor treatment. The LPS-induced production of IL-6 and IL-10 was lower in axSpA patients compared with controls (P < 0.05). Monocytic TLR expression was unaffected in patients with axSpA. CONCLUSION: Enhanced spontaneous and MDP-induced cytokine secretion by monocytes suggests in vivo pre-activation of monocytes in axSpA patients under conventional therapy which is reverted under TNF inhibitor treatment.

Serum adipokine levels in patients with ankylosing spondylitis and their relationship to clinical parameters and radiographic spinal progression.

OBJECTIVE: Adipokines have metabolic and inflammatory functions but can also affect bone metabolism. The purpose of this study was to determine the relationship between serum levels of adiponectin, resistin, and visfatin and markers of inflammation, disease activity, and radiographic spinal progression in patients with ankylosing spondylitis (AS). METHODS: Levels of adiponectin, resistin, and visfatin in the serum of 86 AS patients and 25 healthy controls were measured by enzyme-linked immunosorbent assay at baseline. Radiographic spinal progression was determined by the scoring of radiographs of the spine obtained at baseline and after 2 years. RESULTS: Mean (±SD) baseline levels of resistin and visfatin were significantly higher in AS patients than in healthy controls (11.6 ± 10.6 ng/ml versus 6.6 ± 3.2 ng/ml [P = 0.01] for resistin, and 20.9 ± 48.3 ng/ml versus 3.4 ± 2.6 ng/ml [P = 0.001] for visfatin). Adipokine serum levels did not correlate with disease activity or functional indices. Only resistin serum levels correlated with markers of inflammation. Baseline levels of visfatin, but not resistin or adiponectin, were significantly higher in patients with worsening of the modified Stoke Ankylosing Spondylitis Spine Score (mSASSS) by ≥2 units after 2 years (n = 19) as compared to patients without mSASSS worsening (37.7 ± 57.8 ng/ml versus 16.1 ± 44.6 ng/ml; P = 0.029) and in patients with syndesmophyte formation/progression (n = 22) as compared to patients without such progression (37.1 ± 55.3 ng/ml versus 15.3 ± 44.8 ng/ml; P = 0.023). Visfatin levels of >8 ng/ml at baseline were predictive of subsequent radiographic spinal progression (adjusted odds ratio 3.6 for mSASSS progression and 5.4 for syndesmophyte formation/progression). CONCLUSION: Serum levels of resistin and visfatin are elevated in AS patients. Elevated visfatin levels at baseline are predictive of subsequent progression of radiographic damage in AS patients.

Role of dendritic cells in the pathogenesis of Whipple's disease.

Accumulation of Tropheryma whipplei-stuffed macrophages in the duodenum, impaired T. whipplei-specific Th1 responses, and weak secretion of interleukin-12 (IL-12) are hallmarks of classical Whipple's disease (CWD). This study addresses dendritic cell (DC) functionality during CWD. We documented composition, distribution, and functionality of DC ex vivo or after in vitro maturation by fluorescence-activated cell sorting (FACS) and by immunohistochemistry in situ. A decrease in peripheral DC of untreated CWD patients compared to healthy donors was due to reduced CD11c(high) myeloid DC (M-DC). Decreased maturation markers CD83, CD86, and CCR7, as well as low IL-12 production in response to stimulation, disclosed an immature M-DC phenotype. In vitro-generated monocyte-derived DC from CWD patients showed normal maturation and T cell-stimulatory capacity under proinflammatory conditions but produced less IL-12 and failed to activate T. whipplei-specific Th1 cells. In duodenal and lymphoid tissues, T. whipplei was found within immature DC-SIGN(+) DC. DC and proliferating lymphocytes were reduced in lymph nodes of CWD patients compared to levels in controls. Our results indicate that dysfunctional IL-12 production by DC provides suboptimal conditions for priming of T. whipplei-specific T cells during CWD and that immature DC carrying T. whipplei contribute to the dissemination of the bacterium.

Macrophages accumulate in the gut mucosa of untreated HIV-infected patients.

BACKGROUND: Mucosal macrophages are involved in the maintenance of epithelial barrier integrity and the elimination of invading pathogens. Although an intestinal barrier defect and microbial translocation are hallmarks of human immunodeficiency virus (HIV) infection, recent data on gut mucosal macrophages in HIV infection are sparse. METHODS: Treatment-naive and treated HIV-infected patients and healthy controls were studied for frequencies and functional parameters of blood monocytes and macrophages in duodenal mucosa. RESULTS: We found mucosal enrichment of macrophages in untreated HIV infection associated with reduced monocyte counts in blood and increased monocyte expression of the gut-homing molecule integrin ß7. Increased CCR2 density on integrin ß7-expressing monocytes and mucosal secretion of CCL2 suggest that CCR2/CCL2-chemotaxis is involved in enhanced trafficking of blood monocytes to the gut. Secretion of macrophage-related proinflammatory molecules interleukin 1ß, CCL5, CXCL9, and CXCL10 was increased in the gut mucosa of untreated patients. Moreover, mucosal macrophages of untreated patients showed reduced phagocytic activity. CONCLUSIONS: These data suggest a role for gut mucosal macrophages in HIV immune pathogenesis: infiltrated macrophages in the intestinal mucosa may promote local inflammation and tissue injury, whereas their low phagocytic activity prevents the efficient elimination of luminal antigens that cross the damaged intestinal barrier.

Elevated serum level of the vascular endothelial growth factor predicts radiographic spinal progression in patients with axial spondyloarthritis.

OBJECTIVE: To investigate the role of serum vascular endothelial growth factor (VEGF) as a predictor of radiographic spinal progression in patients with axial spondyloarthritis (axSpA). METHODS: Altogether, 172 patients with definite axSpA (95 with ankylosing spondylitis and 77 with non-radiographic axSpA) were included in this study. Spinal radiographs obtained at baseline and after 2 years of follow-up were scored independently by two trained readers in a concealed and randomly selected order according to the modified Stoke Ankylosing Spondylitis Spine Score (mSASSS) scoring system and for the presence of syndesmophytes. Radiographic spinal progression after 2 years was defined as (1) mSASSS worsening by ≥2 units, and (2) new syndesmophyte formation or formation of a bridging syndesmophyte from two single syndesmophytes. Serum VEGF levels were detected at baseline. RESULTS: Mean baseline VEGF values were significantly higher in patients with mSASSS worsening by ≥2 units after 2 years (n=22) than in those without progression (562±357 vs 402±309 pg/mL, respectively, p=0.027) and in patients with syndesmophyte formation (n=18) again as compared with those without new bone formation (579±386 vs 404±307 pg/mL, respectively, p=0.041). VEGF as a predictor of radiographic spinal progression performed especially well in patients who were already at high risk for such a progression due to the presence of syndesmophytes at baseline (n=48). In these patients, a VEGF serum level of >600 pg/mL had a sensitivity of 53%, a specificity of 97% and an OR=36.6 (95% CI 3.9 to 341.5) as a predictor of mSASSS worsening by ≥2 units. For syndesmophyte formation, elevated VEGF demonstrated a sensitivity of 47%, a specificity of 94% and an OR=13.6 (95% CI 2.4 to 78.3). CONCLUSIONS: An elevated serum level of VEGF (>600 pg/mL) is highly specific as a predictor of radiographic spinal progression in patients with axSpA, especially in patients who are at high risk for further progression due to the presence of syndesmophytes.

Evaluation of arginine metabolism for the analysis of M1/M2 macrophage activation in human clinical specimens.

BACKGROUND: Macrophage heterogeneity reflects their plasticity in response to environmental stimuli. Usually human macrophages are characterized by analysis of surface molecules or cytokine expression while functional assays are established in the mouse system but lacking for various human specimens. METHODS: To evaluate the value of analysis of arginine metabolism for characterization of human macrophage differentiation, we analyzed nitrite production and arginase activity in plasma, duodenal biopsies, and in vitro differentiated macrophages of patients with classical Whipple's disease. RESULTS: We demonstrate that it is feasible to determine the content of urea in supernatants of stimulated duodenal biopsies, arginase activity in fresh duodenal biopsies and plasma samples, and arginase activity and nitrite production in lysates and supernatants of in vitro differentiated macrophages. However, only selected tests are appropriate to define macrophage polarization in human specimens. CONCLUSION: Analysis of arginine metabolism is not suitable for the characterization of in vitro differentiated human macrophages. Besides the measurement of nitrite in duodenal biopsy supernatants, the determination of arginase activity in human plasma seems to be a reasonable functional test to detect enhanced M2 macrophage activation and, thus, is of great value for the analysis of macrophage activity with a minimum of material and costs.

Regulatory T cells in patients with Whipple's disease.

Classical Whipple's disease (CWD) is caused by chronic infection with Tropheryma whipplei that seems to be associated with an underlying immune defect. The pathognomonic hallmark of CWD is a massive infiltration of the duodenal mucosa with T. whipplei-infected macrophages that disperse systemically to many other organ systems. An alleviated inflammatory reaction and the absence of T. whipplei-specific Th1 reactivity support persistence and systemic spread of the pathogen. In this article, we hypothesized that regulatory T cells (T(reg)) are involved in immunomodulation in CWD, and we asked for the distribution, activation, and regulatory capacity of T(reg) in CWD patients. Whereas in the lamina propria of CWD patients before treatment numbers of T(reg) were increased, percentages in the peripheral blood were similar in CWD patients and healthy controls. However, peripheral T(reg) of CWD patients were more activated than those of controls. Elevated secretion of IL-10 and TGF-ß in the duodenal mucosa of CWD patients indicated locally enhanced T(reg) activity. Enhanced CD95 expression on peripheral memory CD4(+) T cells combined with reduced expression of IFN-γ and IL-17A upon polyclonal stimulation by CD4(+) cells from untreated CWD patients further hinted to T(reg) activity-related exhaustion of effector CD4(+) T cells. In conclusion, increased numbers of T(reg) can be detected within the duodenal mucosa in untreated CWD, where huge numbers of T. whipplei-infected macrophages are present. Thus, T(reg) might contribute to the chronic infection and systemic spread of T. whipplei in CWD but in contrast prevent mucosal barrier defect by reducing local inflammation.