Catuxa Prado

Glucocorticoids enhance Th17/Th1 imbalance and signal transducer and activator of transcription 3 expression in systemic lupus erythematosus patients

OBJECTIVE To investigate the relative amounts of Th17 and Th1 cells present in SLE patients and the possible effects of treatments or disease features on these populations. METHODS Peripheral blood mononuclear cells were collected from 75 SLE patients and 19 healthy controls and the proportion of Th17 and Th1 populations were assessed by flow cytometry measuring the amount of IL-17 and IFN-γ-producing cells. Gene expression of signal transducers and activators of transcription 3 (STAT3), STAT4, IL-6R and IL-12R were determined in 30 patients and 8 healthy individuals by real-time RT-PCR. Data were related to clinical and immunological parameters and to the treatment followed during the past 3 months. RESULTS Th17 cells and the Th17/Th1 ratio were significantly increased in SLE patients treated with glucocorticoids compared with healthy individuals, untreated patients or those under other treatments. No association was detected with clinical parameters, but patients with anti-ENA antibodies also displayed increased Th17 responses. Disease activity (SLEDAI) is associated with the Th17/Th1 index only in glucocorticoid-treated patients. In line with these results, gene expression of STAT3 and IL-6R was up-regulated in patients taking these drugs. Accordingly, we found a positive correlation between the Th17/Th1 ratio and STAT3 levels. CONCLUSIONS The present work provides the first evidence that aberrant Th17/Th1 balance in SLE is linked to the use of glucocorticoids and suggests that the up-regulatory effect of these drugs on the Th17 population could be associated with their ability to increase STAT3 and IL-6R expression. Additionally, anti-ENA positivity could represent a potential biomarker for Th17 bias.

Cytokines and regulatory T cells in rheumatoid arthritis and their relationship with response to corticosteroids.

Objective. To analyze circulating cytokines and regulatory T cells (Treg) in patients with rheumatoid arthritis (RA) of different durations, and their association with functional interleukin 10 (IL-10) and tumor necrosis factor-α (TNF-α) genotypes in patients treated with corticosteroids. Methods. Serum levels of IL-6, IL-10, IL-17, IL-18, TNF-α, and transforming growth factor-ß (TGF-ß) were quantified in 196 patients and 61 healthy controls. Percentage of CD4+CD25high cells was determined by flow cytometry and Foxp3 expression by real-time reverse-transcription polymerase chain reaction. Data were related to clinical measurements and presence of the genotype −1082GG IL-10/−308GG TNF-α, previously associated with good response to corticosteroids. Results. Levels of TNF-α, IL-6, and IL-18 were significantly higher in patients compared to controls, while TGF-ß and IL-10 were lower. Serum samples of patients at disease onset (n = 32) had increased IL-6 and decreased TGF-ß, but there were no differences in other cytokines. These patients also presented a higher percentage of CD4+CD25high cells than those with established disease, although no significant differences were detected in Foxp3. Patients under corticosteroid treatment who were carriers of the good responder genotype had higher levels of TGF-ß, Foxp3, and Treg compared to patients with other genotypes, while relatively lower levels of TNF-α and IL-17 were observed. Conclusion. Patients at onset of RA present fewer alterations in cytokine levels and Treg than those with longer disease duration, supporting the role of disease progression in subsequent changes. The antiinflammatory balance observed in high IL-10/low TNF-α patients treated with prednisone supports the use of these genetic polymorphisms as predictors of response to corticosteroid therapy.

Interleukin 10 and Tumor Necrosis Factor-α Genotypes in Rheumatoid Arthritis — Association with Clinical Response to Glucocorticoids

Objective. There are dysregulated levels of interleukin 10 (IL-10) and tumor necrosis factor-α (TNF-α) in rheumatoid arthritis (RA), and their role in the disease is controversial. We analyzed the association of functional polymorphisms of IL-10 and TNF-α with susceptibility and disease characteristics at the time of diagnosis, and we also evaluated their possible use as predictors of clinical response to treatments. Methods. Patients with recent-onset RA (n = 162) and healthy controls (n = 373) were genotyped for −1082 IL-10 and −308 TNF-α polymorphisms and data were related to clinical and immunological measurements of patients at the time of diagnosis. Response to treatment after 6 months was determined in 125 patients by the absolute change in Disease Activity Score (DAS28) and the American College of Rheumatology criteria for improvement. Results. We found a reduced frequency of the low IL-10 producer genotype (−1082AA) in patients with RA compared to controls (26.5% vs 38.9%; p = 0.006), while it is a risk factor for anticyclic citrullinated peptide antibodies (anti-CCP) positivity (p = 0.028). Evaluation of clinical response to treatments indicated that carriage of the high IL-10 genotype was associated with a favorable outcome (p = 0.009), specifically to prednisone therapy (p = 0.0003). No significant effects were observed with TNF-α polymorphism alone; however, in combination with the IL-10 genotype, it increased the strength of these associations. Conclusion. Results show an association between the low IL-10 producer genotype and protection from RA; nevertheless, when other specific genetic and/or environmental factors trigger onset of RA, this genotype may predispose to development of anti-CCP+ RA disease with reduced response to prednisone treatment.

Effects of glucocorticoid treatment on CD25−FOXP3+ population and cytokine-producing cells in rheumatoid arthritis

OBJECTIVES To investigate CD25(-)FOXP3(+) cells in RA patients and their possible relationship with disease features and response to glucocorticoids (GCs). METHODS Peripheral blood mononuclear cells were collected from 147 RA patients, 29 healthy controls and 75 SLE patients as disease controls. The proportion of CD4(+)FOXP3(+) cells with negative, low or high CD25 expression and the levels of IL-10-, TNF-α-, IL-17- and IFNγ-producing cells were assessed by flow cytometry. The presence of the high IL-10 genotype (-1082GG), associated with good response to GC, was determined by PCR amplification and hybridization with allele-specific fluorescently labelled probes. Data were related to treatment and clinical parameters. RESULTS The CD25(-)FOXP3(+) population was significantly increased in RA patients and negatively correlated with DAS-28 and other disease parameters. The IL-10 genotype did not influence the frequency of these cells in controls or the entire RA group; however, GC-treated patient carriers of the high IL-10 genotype presented significantly higher levels of this population in addition to an increased percentage of IL-10-secreting cells and relatively low amounts of TNF-α-, IFN-γ- and IL-17-positive cells. Finally, a prospective study confirmed that genetically high IL-10 producers significantly increase CD25(-)FOXP3(+) cells after 6 months of GC treatment. CONCLUSION The present study provides the first evidence of increased CD25(-)FOXP3(+) cells in RA patients, which were associated with disease activity and with GC treatment in carriers of the high IL-10 genotype, suggesting that this population plays a role in the clinical response to prednisone in RA.

IFNα Serum Levels Are Associated with Endothelial Progenitor Cells Imbalance and Disease Features in Rheumatoid Arthritis Patients

Introduction IFNα has been largely implicated in the ethiopathogenesis of autoimmune diseases but only recently it has been linked to endothelial damage and accelerated atherosclerosis in autoimmunity. In addition, proinflammatory conditions are supposed to be implicated in the cardiovascular status of these patients. Since a role for IFNα in endothelial damage and impaired Endothelial Progenitor Cell (EPC) number and function has been reported in other diseases, we aimed to evaluate the potential associations of IFNα serum levels on EPC populations and cytokine profiles in Rheumatoid Arthritis (RA) patients. Methods pre-EPC, EPC and mature EPC (mEPC) populations were quantified by flow cytometry analyzing their differential CD34, CD133 and VEGFR2 expression in blood samples from 120 RA patients, 52 healthy controls (HC), and 83 systemic lupus erythematosus (SLE) patients as disease control. Cytokine serum levels were measured by immunoassays and clinical and immunological data, including cardiovascular (CV) events and CV risk factors, were retrospectively obtained by reviewing clinical records. Results Long-standing, but not recent onset RA patients displayed a significant depletion of all endothelial progenitor populations, unless high IFNα levels were present. In fact, the IFNhigh RA patient group (n = 40, 33%), showed increased EPC levels, comparable to SLE patients. In addition, high IFNα serum levels were associated with higher disease activity (DAS28), presence of autoantibodies, higher levels of IL-1β, IL-6, IL-10 and MIP-1α, lower amounts of TGF-β, and increased mEPC/EPC ratio, thus suggesting higher rates of endothelial damage and an endothelial repair failure. Finally, the relationship between high IFNα levels and occurrence of CV events observed in RA patients seems to support this hypothesis. Conclusions IFNα serum marker could be used to identify a group of RA patients with increased disease activity, EPC imbalance, enhanced proinflammatory profile and higher cardiovascular risk, probably due, at least in part, to an impaired endothelial repair.

Antibacterial and Antifungal Activity of ZnO Containing Glasses

A new family of non-toxic biocides based on low melting point (1250°C) transparent glasses with high content of ZnO (15–40wt%) belonging to the miscibility region of the B2O3-SiO2-Na2O-ZnO system has been developed. These glasses have shown an excellent biocide activity (logarithmic reduction >3) against Gram- (E. coli), Gram+ (S. aureus) and yeast (C. krusei); they are chemically stable in different media (distilled water, sea-like water, LB and DMEN media) as well as biocompatible. The cytotoxicity was evaluated by the Neutral Red Uptake using NIH-3T3 (mouse embryonic fibroblast cells) and the cell viability was >80%. These new glasses can be considered in several and important applications in the field of inorganic non-toxic biocide agents such as medical implants, surgical equipment, protective apparels in hospitals, water purifications systems, food packaging, food storages or textiles.

Mechanical performance of a biocompatible biocide soda–lime glass-ceramic

A biocompatible soda-lime glass-ceramic in the SiO2-Na2O-Al2O3-CaO-B2O3 system containing combeite and nepheline as crystalline phases, has been obtained at 750°C by two different routes: (i) pressureless sintering and (ii) Spark Plasma Sintering. The SPS glass-ceramic showed a bending strength, Weibull modulus, and toughness similar values to the cortical human bone. This material had a fatigue limit slightly superior to cortical bone and at least two times higher than commercial dental glass-ceramics and dentine. The in vitro studies indicate that soda-lime glass-ceramic is fully biocompatible. The in vivo studies in beagle jaws showed that implanted SPS rods presented no inflammatory changes in soft tissues surrounding implants in any of the 10 different cases after four months implantation. The radiological analysis indicates no signs of osseointegration lack around implants. Moreover, the biocide activity of SPS glass-ceramic versus Escherichia coli, was found to be >4log indicating that it prevents implant infections. Because of this, the SPS new glass-ceramic is particularly promising for dental applications (inlay, crowns, etc).

Calcium and Zinc Containing Bactericidal Glass Coatings for Biomedical Metallic Substrates

The present work presents new bactericidal coatings, based on two families of non-toxic, antimicrobial glasses belonging to B2O3–SiO2–Na2O–ZnO and SiO2–Na2O–Al2O3–CaO–B2O3 systems. Free of cracking, single layer direct coatings on different biomedical metallic substrates (titanium alloy, Nb, Ta, and stainless steel) have been developed. Thermal expansion mismatch was adjusted by changing glass composition of the glass type, as well as the firing atmosphere (air or Ar) according to the biomedical metallic substrates. Formation of bubbles in some of the glassy coatings has been rationalized considering the reactions that take place at the different metal/coating interfaces. All the obtained coatings were proven to be strongly antibacterial versus Escherichia coli (>4 log).

Relationship between FOXP3 positive populations and cytokine production in systemic lupus erythematosus

In this work we studied CD4+FOXP3+ populations in systemic lupus erythematosus (SLE) and the relationship with Th cytokine production. We found an increment in CD25-FOXP3+ population in SLE associated with CD4+ downregulation and disease progression. CD25low cells were also upregulated and showed increased percentages of FOXP3+ and CD127-/low cells, supporting the activated status of SLE lymphocytes. Despite the normal levels of CD25highFOXP3+ cells, the negative correlations observed in controls with the frequency of IFNγ, TNFα and IL-10 secreting cells were disrupted in patients, supporting a defective Treg function. Also, CD25high cells showed an altered balance in the production of these cytokines. In addition, CD25highFOXP3+ cells correlated directly with IL-17A and IL-8 but not with TGFβ in SLE. The increased proportion of IL-17+ cells among the CD25high subset and the positive correlation between IL-17 levels and Treg cells suggest a trans-differentiation of Treg into Th17 cells in SLE.

Performance of a New Al2O3/Ce–TZP Ceramic Nanocomposite Dental Implant: A Pilot Study in Dogs.

Although titanium remains as the prevalent material in dental implant manufacturing new zirconia-based materials that overcome the major drawbacks of the standard 3Y-yttria partially-stabilized zirconia (Y-TZP) are now emerging. In this study, a new ceramic nanocomposite made of alumina and ceria-stabilized TZP (ZCe-A) has been used to produce dental implants with the mechanic and topographic characteristics of a pilot implant design to evaluate bone and soft tissue integration in a dog model (n = 5). Histological cross-section analysis of the implanted ceramic fixations (n = 15) showed not only perfect biocompatibility, but also a high rate of osseous integration (defined as the percentage of bone to implant contact) and soft tissue attachment. This clinical success, in combination with the superior mechanical properties achieved by this Al2O3/Ce-TZP nanocomposite, may place this material as an improved alternative of traditional 3Y-TZP dental implants.