High Dickkopf-1 levels are associated with chronic inflammation in children with sickle cell disease.

OBJECTIVES: Sickle bone disease (SBD) is a chronic complication of sickle cell disease (SCD) whose pathogenesis is not completely understood. Chronic inflammation associated with SCD could alter bone remodeling. Our aim was to analyze the serum levels of bone remodeling markers in a group of SCD children to evaluate their involvement in the SBD. METHODS: We enrolled 26 SCD subjects and 26 age-matched controls, who lived in the same geographic area. DKK-1, sclerostin, RANKL, and OPG serum levels were evaluated. Neutrophil-lymphocyte ratio (NLR) was also evaluated as a marker of inflammation. RESULTS: The analysis of bone remodeling markers did not show any significant difference between the two groups except for DKK-1 levels that were significantly higher in the patients than controls (p < .05). A significant direct correlation between NLR and DKK-1 (p = .004) was found. An inverse correlation between NLR and osteocalcin (p = .01) has also been observed. CONCLUSIONS: The chronic inflammation, which represents a peculiar characteristic in SCD patients, would represent the primary causal agent of the activation of osteoblastogenesis inhibitors responsible of bone impairment in these subjects. Further studies will be needed to better explain the role of these inhibitors in SCD, to prevent or treat bone damage in this population.

Shedding "LIGHT" on the Link between Bone and Fat in Obese Children and Adolescents.

Obesity may affect bone health, but literature reports are contradictory about the correlation of body mass index (BMI) and bone markers. LIGHT, one of the immunostimulatory cytokines regulating the homeostasis of bone and adipose tissue, could be involved in obesity. The study involved 111 obese subjects (12.21 ± 3.71 years) and 45 controls. Patients underwent the evaluation of bone status by quantitative ultrasonography (QUS). LIGHT amounts were evaluated in sera by ELISA, whereas its expression on peripheral blood cells was evaluated by flow cytometry. Osteoclastogenesis was performed by culturing peripheral blood mononuclear cells (PBMCs) with or without anti-LIGHT antibodies. Obese patients showed significant high BMI-standard deviation score (SDS), weight-SDS, and Homeostatic model assessment for insulin resistance (HOMA-IR) that negatively correlated with the reduced Amplitude Dependent Speed of Sound (AD-SoS)-Z-score and Bone Transmission Time (BTT-Z)-score. They displayed significantly higher serum levels of LIGHT compared with controls (497.30 ± 363.45 pg/mL vs. 186.06 ± 101.41 pg/mL, p < 0.001). LIGHT expression on monocytes, CD3+-T-cells, and neutrophils was also higher in obese patients than in the controls. Finally, in PBMC cultures, the addition of anti-LIGHT antibodies induced a significant osteoclastogenesis inhibition. Our study highlighted the high serum levels of LIGHT in obese children and adolescents, and its relationship with both the grade of obesity and bone impairment.

Mechanisms of Enhanced Osteoclastogenesis in Alkaptonuria.

Alkaptonuria (AKU) is a rare disorder characterized by the deficiency of the enzyme homogentisate 1,2-dioxygenase and consequent homogentisate accumulation, which leads to progressive and severe osteoarthopathy starting from the second decade of life. Thus, in AKU patients, bone involvement represents an important clinical issue, which we investigated. Serum levels of receptor activator of NF-κB ligand (RANKL), osteoprotegerin, sclerostin, Dickkopf-1, and bone remodeling markers were measured in nine AKU patients (two children and seven adults) and 22 controls, together with lumbar spine bone mineral density (LS-BMD) and femoral-BMD. In the two AKU children, the average of LS-BMD and femoral-BMD Z-scores were within the normal range, but reduced with respect to the controls. Otherwise, in the adult AKU patients, LS-BMD T-score was inside the normal range, but femoral-BMD T-score reached osteopenic levels. Consistently, in AKU adults, higher RANKL and C-terminal telopeptide of collagen type 1 and lower osteoprotegerin levels were observed than in controls. Otherwise, spontaneous osteoclastogenesis was already evident in peripheral blood mononuclear cell cultures from AKU children, together with a high percentage of circulating osteoclast precursors. Osteoclastogenesis was sustained by the high levels of tumor necrosis factor-α, RANK, RANKL, and LIGHT. In conclusion, the altered osteoclastogenesis was observed already in AKU children, despite the absence of evident injury. Thus, a preventive approach in young patients, targeting osteoclast activity, may prevent the macroscopic bone disease that appears in adult AKU.

Irisin serum levels are positively correlated with bone mineral status in a population of healthy children.

BACKGROUND: Irisin is a myokine secreted by skeletal muscle during physical activity. Irisin treatment increased cortical bone mineral density (BMD) in young healthy mice and restored bone and muscle mass loss in a mouse model of disuse-induced osteoporosis and muscular atrophy. In humans, Irisin was positively correlated with BMD in young athletes. Considering that the bone mass reached during childhood is one of the most important determinants of lifelong skeletal health, we sought to determine if Irisin levels were correlated with bone mineral status in children. METHODS: Irisin and bone metabolic markers were quantified in sera and bone mineral status was evaluated by quantitative ultrasound in a population of 34 healthy children (9.82 ± 3.2 years). RESULTS: We found that Irisin levels were positively correlated with the amplitude-dependent speed of sound Z-score (r = 0.305; p < 0.001), bone transmission time Z-score (r = 0.375; p < 0.001) and osteocalcin (r = 0.370; p < 0.001), and negatively with Dickkopf WNT Signaling Pathway Inhibitor 1 (r = -0.274; p < 0.001). CONCLUSION: In a regression analysis model, Irisin was one of the determinants of bone mineral status to a greater extent than bone alkaline phosphatase and parathyroid hormone, indicating that Irisin might be considered as one of the bone formation markers during childhood.

Osteoclastogenic potential of peripheral blood mononuclear cells in cleidocranial dysplasia.

Cleidocranial dysplasia (CCD) is an autosomal dominant skeletal dysplasia characterized by hypoplastic or aplastic clavicles, dental abnormalities, and delayed closure of the cranial sutures. In addition, mid-face hypoplasia, short stature, skeletal anomalies and osteoporosis are common. We aimed to evaluate osteoclastogenesis in a child (4 years old), who presented with clinical signs of CCD and who have been diagnosed as affected by deletion of RUNX2, master gene in osteoblast differentiation, but also affecting T cell development and indirectly osteoclastogenesis. The results of this study may help to understand whether in this disease is present an alteration in the bone-resorptive cells, the osteoclasts (OCs). Unfractionated and T cell-depleted Peripheral Blood Mononuclear Cells (PBMCs) from patient were cultured in presence/absence of recombinant human M-CSF and RANKL. At the end of the culture period, OCs only developed following the addition of M-CSF and RANKL. Moreover, real-time PCR experiment showed that freshly isolated T cells expressed the osteoclastogenic cytokines (RANKL and TNFα) at very low level, as in controls. This is in accordance with results arising from flow cytometry experiments demonstrating an high percentage of circulating CD4(+)CD28(+) and CD4(+)CD27(+) T cells, not able to produce osteoclastogenic cytokines. Also RANKL, OPG and CTX serum levels in CCD patient are similar to controls, whereas QUS measurements showed an osteoporotic status (BTT-Z score -3.09) in the patient. In conclusions, our findings suggest that the heterozygous deletion of RUNX2 in this CCD patient did not alter the osteoclastogenic potential of PBMCs in vitro.

High dickkopf-1 levels in sera and leukocytes from children with 21-hydroxylase deficiency on chronic glucocorticoid treatment.

Children with 21-hydroxylase deficiency (21-OHD) need chronic glucocorticoid (cGC) therapy to replace congenital deficit of cortisol synthesis, and this therapy is the most frequent and severe form of drug-induced osteoporosis. In this study, we enrolled 18 patients (9 females) and 18 sex- and age-matched controls. We found in 21-OHD patients high serum and leukocyte levels of dickkopf-1 (DKK1), a secreted antagonist of the Wnt/ß-catenin signaling pathway known to be a key regulator of bone mass. In particular, we demonstrated by flow cytometry, confocal microscopy, and real-time PCR that monocytes, T lymphocytes, and neutrophils from patients expressed high levels of DKK1, which may be related to the cGC therapy. In fact, we showed that dexamethasone treatment markedly induced the expression of DKK1 in a dose- and time-dependent manner in leukocytes. The serum from patients containing elevated levels of DKK1 can directly inhibit in vitro osteoblast differentiation and receptor activator of NF-κB ligand (RANKL) expression. We also found a correlation between both DKK1 and RANKL or COOH-terminal telopeptides of type I collagen (CTX) serum levels in 21-OHD patients on cGC treatment. Our data indicated that DKK1, produced by leukocytes, may contribute to the alteration of bone remodeling in 21-OHD patients on cGC treatment.

Brain-Type Creatine Kinase Release from Cultured Osteoclasts Exposed to Neridronate in Children Affected by Osteogenesis Imperfecta Type 1.

Brain-type creatine kinase (CK-BB) increases during osteoclastogenesis, with high circulating amounts in type I osteogenesis imperfecta (OI) following treatment with neridronate, a bisphosphonate able to inhibit osteoclast activity and survival. The aim of this study was to demonstrate the correlation between osteoclastogenesis and CK-BB release from OI patients' osteoclasts treated with different concentrations of neridronate. Our patients showed reduced bone quality, increased levels of CTX I, a marker of bone resorption, and decreased levels of OPG, an inhibitor of osteoclastogenesis. In OI patients, the presence of MCSF and RANKL determined an increased secretion of CK-BB from osteoclasts (p = 0.04) compared with control conditions without these cytokines; interestingly, in the absence of these factors, the secretion of CK-BB is significantly elevated at 3 µmol/L compared with 0.03 and 1 µmol/L (p = 0.007). In healthy donors' cultures, the higher concentration of CK-BB can be detected following stimulation with 3 µmol/L neridronate compared with the untreated condition both with and without MCSF and RANKL (p = 0.03 and p = 0.006, respectively). Consistently, in osteoclast cultures, neridronate treatment is associated with a decrease in multinucleated TRAP+ cells, together with morphology changes typical of apoptosis. Consistently, in the media of the same osteoclast cultures, we demonstrated a significant increase in caspase-3 levels. In conclusion, our findings support the idea that CK-BB levels increase in the serum of OI-treated patients.

Bone Remodeling Markers in Children with Acute Lymphoblastic Leukemia after Intensive Chemotherapy: The Screenshot of a Biochemical Signature.

PURPOSE: to investigate the effects of intensive chemotherapy and glucocorticoid (GC) treatment on bone remodeling markers in children with acute lymphoblastic leukemia (ALL). METHODS: A cross-sectional study was carried out in 39 ALL children (aged 7.64 ± 4.47) and 49 controls (aged 8.7 ± 4.7 years). Osteoprotegerin (OPG), receptor activator of NF-κB ligand (RANKL), osteocalcin (OC), C-terminal telopeptide of type I collagen (CTX), bone alkaline phosphatase (bALP), tartrate-resistant acid phosphatase 5b (TRACP5b), procollagen type I N-terminal propeptide (P1NP), Dickkopf-1 (DKK-1), and sclerostin were assessed. Statistical analysis was conducted using the principal component analysis (PCA) to study patterns of associations in bone markers. RESULTS: ALL patients showed significantly higher OPG, RANKL, OC, CTX, and TRACP5b than the controls (p ≤ 0.02). Considering ALL group, we found a strong positive correlation among OC, TRACP5b, P1NP, CTX, and PTH (r = 0.43-0.69; p < 0.001); between CTX and P1NP (r = 0.5; p = 0.001); and between P1NP and TRAcP (r = 0.63; p < 0.001). The PCA revealed OC, CTX, and P1NP as the main markers explaining the variability of the ALL cohort. CONCLUSIONS: Children with ALL showed a signature of bone resorption. The assessment of bone biomarkers could help identify ALL individuals who are most at risk of developing bone damage and who need preventive interventions.

Bone Remodeling in an Mps-1h Girl after Hematopoietic Stem Cell Transplantation along with Enzymatic Replacement Therapy.

BACKGROUND: Mucopolysaccharidosis-1H (Hurler syndrome, MPS-1H) is the most severe form of a lysosomal storage disorder (LSD) caused by variants in IDUA, encoding alpha- L-iduronidase (IDUA). MPS-1H is also associated with various degrees of skeletal defects due to the accumulation of partially degraded glycosaminoglycans (GAGs) in the lysosomes of connective tissue cells. The efficacy of hematopoietic stem cell transplantation (HSCT) and enzymatic replacement therapy (ERT) on MPS-1H skeletal manifestations is still considered unsatisfactory. CASE PRESENTATION: We report the case of a young girl, who manifested significant changes in bone remodeling markers and osteoclastogenesis potential after HSCT combined with ERT. She received ERT and underwent two HSCTs. The skeletal alterations at the time of diagnosis showed a trend toward improvement of both mobility and radiological features after HSCT. We observed the highest levels of Receptor activator of nuclear factor-kappa-Β ligand (RANKL) and RANK/osteoprotegerin (OPG) ratio at diagnosis and during ERT, consistently with spontaneous osteoclastogenesis. Conversely, after the successful HSCT with ongoing ERT, the highest levels of osteocalcin were observed and all markers of bone formation and resorption improved. CONCLUSION: The combination therapy of ERT and HSCT was effective in reducing osteoclast activity and increasing osteoblast activity, and these changes were according to the child's bone phenotype, IDUA activity, and Glycosaminoglycan (GAG) trends. These results represent one of the few pieces of human evidence in this context.

Extra virgin olive oil extract rich in secoiridoids induces an anti-inflammatory profile in peripheral blood mononuclear cells from obese children.

Obesity represents an important public health challenge of the twenty first century reaching epidemic proportions worldwide; this is especially true for the pediatric population. In this context, bioactive compounds from foods are crucial to counteract chronic inflammation as a typical feature of obesity. In particular, extra virgin olive oil (EVOO) is one of the most important functional foods exerting, among others, an anti-inflammatory activity not only due to its major (monounsaturated fatty acids) but also to its minor (phenolics) components, as reported in the last years. However, only a limited number of studies were performed on pediatric population, and even fewer are those focusing on EVOO phenolics that investigate the correlation of the chemical characterization with the biological function. Thus, starting from our in vitro data identifying an EVOO chemical profile characterized by a high content of secoiridoids correlating with an anti-inflammatory effect, we studied the ability of an EVOO extract with the same chemical profile to retain this function ex vivo. Specifically, peripheral blood mononuclear cells (PBMCs) collected from obese children were treated with EVOO and olive oil extracts, characterized by a low polyphenol content, to study the ability of secoiridoids to dampen the inflammatory response. A reduction of pro-inflammatory CD14+CD16+ monocytes was detected by cytofluorimetric analysis when PBMCs were treated with EVOO as compared to olive oil extracts. According to this, a down modulation of CCL2 and CCL4 chemokines involved in the recruitment of inflammatory cells, was reported in the supernatants of EVOO relative to olive oil extracts treated PBMCs. Moreover, a high-throughput gene expression analysis revealed that PBMCs molecular profile from obese children is greatly modulated after the treatment with EVOO extract in terms of metabolic and inflammatory pathways. Importantly, some of the significantly modulated genes were involved in the pathways promoting the development of severe obesity. Overall, our ex vivo data demonstrated the ability of EVOO to reduce the inflammatory milieu of PBMCs from obese children both at protein and molecular levels. Of note, a good correlation between the EVOO chemical profile and the biological modulations in terms of anti-inflammatory activity was reported.

Effects of Sweet Cherry Polyphenols on Enhanced Osteoclastogenesis Associated With Childhood Obesity.

Childhood obesity is associated with the development of severe comorbidities, such as diabetes, cardiovascular diseases, and increased risk of osteopenia/osteoporosis and fractures. The status of low-grade inflammation associated to obesity can be reversed through an enhanced physical activity and by consumption of food enrich of anti-inflammatory compounds, such as omega-3 fatty acids and polyphenols. The aim of this study was to deepen the mechanisms of bone impairment in obese children and adolescents through the evaluation of the osteoclastogenic potential of peripheral blood mononuclear cells (PBMCs), and the assessment of the serum levels of RANKL and osteoprotegerin (OPG). Furthermore, we aimed to evaluate the in vitro effects of polyphenol cherry extracts on osteoclastogenesis, as possible dietary treatment to improve bone health in obese subjects. High RANKL levels were measured in obese with respect to controls (115.48 ± 35.20 pg/ml vs. 87.18 ± 17.82 pg/ml; p < 0.01), while OPG levels were significantly reduced in obese than controls (378.02 ± 61.15 pg/ml vs. 436.75 ± 95.53 pg/ml, respectively, p < 0.01). Lower Ad-SoS- and BTT Z-scores were measured in obese compared to controls (p < 0.05). A significant elevated number of multinucleated TRAP+ osteoclasts (OCs) were observed in the un-stimulated cultures of obese subjects compared to the controls. Interestingly, obese subjects displayed a higher percentage of CD14+/CD16+ than controls. Furthermore, in the mRNA extracts of obese subjects we detected a 2.5- and 2-fold increase of TNFα and RANKL transcripts compared to controls, respectively. Each extract of sweet cherries determined a dose-dependent reduction in the formation of multinucleated TRAP+ OCs. Consistently, 24 h treatment of obese PBMCs with sweet cherry extracts from the three cultivars resulted in a significant reduction of the expression of TNFα. In conclusion, the bone impairment in obese children and adolescents is sustained by a spontaneous osteoclastogenesis that can be inhibited in vitro by the polyphenol content of sweet cherries. Thus, our study opens future perspectives for the use of sweet cherry extracts, appropriately formulated as nutraceutical food, as preventive in healthy children and therapeutic in obese ones.

Reduced interleukin-5 production by peripheral CD4+ T cells in common variable immunodeficiency patients.

We evaluated the capacity of peripheral CD4+ T helper cells in four Common Variable Immunodeficiency (CVI) patients to secrete interleukin-4 (IL-4) and IL-5. While in control CD4+ T cells, stimulated via CD3 and cultured in presence of IL-2 or IL-15, a 10 fold increased production of IL-5 (146 +/- 30; 142 +/- 25 pg/ml) was found, a 4 fold increment of this cytokine was, instead, detected in 3 out of 4 CVI patients (34 +/- 13; 39 +/- 12 pg/ml) (p < 0.05). In conclusion, the reduction of IL-5, involved in the late regulation of B cell differentiation into Ig-secreting plasma cells, may contribute to the defective antibody production in CVI patients.

Cytokine pattern and endothelium damage markers in Henoch-Schönlein purpura.

In a longitudinal cohort study our aim was to evaluate the cytokine pattern of children affected by Henoch-Schonlein purpura (HSP) and to correlate this pattern to vascular endothelium damage and to nephropathy. The following parameters were monitored at the onset of the disease (T0) and after 6 months of follow-up (T1): clinical scores, serum levels of tumor necrosis factor alpha (TNF-alpha), interleukin 2 (IL-2), soluble IL-2 receptor (IL-2sRalpha), fibrinogen, von Willebrand factor antigen (vWf:Ag) and soluble thrombomodulin (TMD) levels. A total of 24 children (9 M, 15 F), affected by HSP, aged between 3-14 years (median 6 years), were enrolled into the study. IL-2 serum levels were significantly increased at the onset of the disease compared to control group and T1. The same pattern was observed for IL-2sRalpha and TNF-alpha. Fibrinogen and vWf:Ag concentrations were significantly higher at the onset of disease than t1 and in control group. TMD levels resulted constantly within the normal range. Concerning the analyzed parameters, no significant difference resulted to be in subjects with and without renal involvement (hematuria and/or proteinuria). Finally, raised serum TNF-alpha concentration, related to vascular endothelium damage as shown by increased vWf:Ag levels, occurred invariably in children affected by HSP both with and without renal involvement.

High irisin levels are associated with better glycemic control and bone health in children with Type 1 diabetes.

AIM: Irisin is a new peptide produced mainly by the skeletal muscle playing an important role both in glucose/energy homeostasis and bone metabolism. Childhood type 1 diabetes mellitus (T1DM) is associated with decreased bone mass. We aimed to evaluate irisin levels in TD1M children and their correlation with bone metabolism and glycaemic control. METHODS: Ninety-six T1DM subjects (12.2 ±â€¯4 years), 56 on multiple daily injections (MDI), 40 on continuous subcutaneous insulin infusion (CSII), and 34 controls were included in the study. Irisin and bone remodeling markers were quantified in sera from patients and controls. Bone mineral density (BMD) was evaluated by QUS. RESULTS: Increased irisin levels were found in T1DM patients respect to controls (p < 0.001). With adjustment for age, irisin levels significantly correlated negatively with HbA1c% (r = -0.105, p < 0.001), years of diabetes (r = -0.07, p < 0.04), 25(OH)-Vitamin D (r = -0.175, p < 0.0001), and positively with BTT-Z-score (r = 0.088, p = 0.016), and osteocalcin (r = 0.059, p < 0.04). We detected the highest levels of irisin in CSII patients compared to MDI and controls (p < 0.001 and p < 0.007 respectively). CONCLUSIONS: We demonstrated high irisin levels in T1DM children and the association of highest irisin amounts to a better glycaemic control and bone health in TDM1 subjects on CSII.

High Sclerostin and Dickkopf-1 (DKK-1) Serum Levels in Children and Adolescents With Type 1 Diabetes Mellitus.

Context: Childhood type 1 diabetes mellitus (T1DM) is associated with decreased bone mass. Sclerostin and dickkopf-1 (DKK-1) are Wnt inhibitors that regulate bone formation. Objective: To evaluate sclerostin and DKK-1 levels in T1DM children and to analyze the influence of glycemic control on bone health. Design and setting: Cross-sectional study conducted at a clinical research center. Participants: One hundred and six T1DM subjects (12.2 ± 4 years), 66 on multiple daily injections (MDIs) and 40 on continuous subcutaneous infusion of insulin (CSII), and 80 controls. Results: The average bone transmission time (BTT) and amplitude-dependent speed of sound (AD-SoS) z scores were lower in patients with diabetes than in controls. Significantly increased DKK-1 (3593 ± 1172 vs 2652 ± 689 pg/mL; P < 0.006) and sclerostin (29.45 ± 12.32 vs 22.53 ± 8.29; P < 0.001) levels were found in patients with diabetes with respect to controls, particularly in patients on MDI compared with ones on CSII. Glycemic control was improved in CSII patients compared with MDI ones (P < 0.001) and was also associated with significantly higher BMI-SDS (P < 0.002) and BTT z scores (P < 0.02). With adjustment for age, multiple linear regression analysis of DKK-1 and sclerostin as dependent variables showed that levels of glycated hemoglobin, glucose, 25(OH) vitamin D, osteocalcin, and parathyroid hormone; years of diabetes; and BMI-SDS and AD-SoS z score were the most important predictors (P < 0.0001). Conclusions: Our study highlighted (1) the high serum levels of DKK-1 and sclerostin in T1DM children and their relationship with altered glycemic control and (2) the effect of CSII on improvement of glycemic control and bone health in T1DM children.

Mechanisms of enhanced osteoclastogenesis in girls and young women with Turner's Syndrome.

Subjects with hypergonadotropic hypogonadism due to Turner's syndrome show low cortical mineral density, osteoporosis and risk of fractures. It is not clear if this bone fragility derives from chromosomal abnormalities or is the result of inadequate bone formation due to estrogen deficiency. The aim of this study was to investigate the cellular mechanisms underlying bone fragility in subjects with Turner's syndrome before induction of puberty and after hormonal replacement therapy (HRT). For this purpose, we have evaluated the osteoclastogenic potential of non-fractioned and T-cell depleted cultures of peripheral blood mononuclear cells (PBMCs) belonging to girls with Turner's syndrome who had not been treated with HRT yet, girls and young women who were on HRT and age-matched controls. Untreated subjects showed high FSH serum levels, whereas the other subjects displayed normal FSH serum levels. T-cell immunophenotype was analyzed through flow cytometry. Biochemical and DXA analyses were performed. Spontaneous osteoclastogenesis in non-fractioned and T-cell depleted cultures of PBMC belonging to girls with high FSH levels was more evident than in cultures of subjects with normal FSH levels. In the former, osteoclastogenesis was sustained by monocytes expressing high levels of c-fms, TNF-α and RANK, and T-cells producing high RANKL and TNF-α; in the latter it was supported by T-cells expressing high RANKL levels. CD4(+)CD25(high) T-cells were reduced in all subjects, whereas CD3(+)/CD16(+)/CD56(+) NKT-cells were increased in those with high FSH levels. High RANKL and CTX levels were detected in the sera. Bone impairment was already detectable by DXA in subjects aged under 10, although it became more evident with aging. In conclusion, our results demonstrated that bone fragility in subjects with Turner's syndrome is associated to enhanced osteoclastogenesis. This process seems to be due to high FSH serum levels before HRT, whereas it is caused by high RANKL during HRT.

Clinical significance of serum cytokine levels and thrombopoietic markers in childhood idiopathic thrombocytopenic purpura.

BACKGROUND: Biological markers useful for defining children with newly diagnosed immune thrombocytopenic purpura (ITP) who are likely to develop the chronic form of the disease are partially lacking. The purpose of this study was to assess the clinical role of both immunological and thrombopoietic markers in children with ITP and correlate their levels with different disease stages. MATERIALS AND METHODS: We enrolled 28 children with ITP at the onset of their disease, who were followed-up for a whole year and divided according to whether their disease resolved within the 12 months (n=13) or became chronic (n=15), 11 subjects with chronic ITP off therapy for at least 1 month at the time of enrolment, and 30 healthy matched controls. Serum levels of T helper type 1 and 2 and T regulatory-associated cytokines, such as interferon γ, tumour necrosis factor α, interleukin (IL) 2, IL6, IL10, and thrombopoietin were measured in all children using quantitative immunoenzymatic assays, while reticulated platelets were evaluated by flow cytometric analysis. RESULTS: Serum IL10 levels were significantly higher in patients with an acute evolution of ITP than in either healthy controls (p<0.001) or patients with chronic progression of ITP (p<0.05). Reticulated platelet count and thrombopoietin levels were significantly higher in ITP patients at the onset of their disease, whether with acute resolution or chronic progression, than in healthy subjects (p<0.01; p<0.001), but did not differ between the groups of patients. CONCLUSION: IL-10 seems to predict the clinical course of ITP, as it is significantly higher at the onset of disease in patients who obtain disease remission in less than 1 year.

Osteoclastogenesis in children with 21-hydroxylase deficiency on long-term glucocorticoid therapy: the role of receptor activator of nuclear factor-kappaB ligand/osteoprotegerin imbalance.

CONTEXT: Children with 21-hydroxylase deficiency (21-OHD) need chronic glucocorticoid (cGC) therapy to replace congenital deficit of cortisol synthesis. cGC therapy is the most frequent and severe form of drug-induced osteoporosis, and different mechanisms have been proposed to explain its pathogenesis. OBJECTIVE: We investigated the osteoclastogenic potential of peripheral blood mononuclear cells (PBMCs) from 18 children with 21-OHD on cGC therapy and 25 controls who never received GCs. We also evaluated the presence of circulating osteoclast precursors (OCPs) and the role of T cells in osteoclast formation. RESULTS: Spontaneous osteoclastogenesis, without adding macrophage-colony stimulating factor and receptor activator of nuclear factor-kappaB ligand (RANKL), and significantly higher osteoclasts resorption activity occurred in 21-OHD patients. Conversely, macrophage-colony stimulating factor and RANKL were essential to trigger and sustain osteoclastogenesis in controls. Furthermore, in 21-OHD patients, we identified a significant percentage of CD11b-CD51/CD61 and CD51/61-RANK-positive cells, which are OCPs strongly committed. Additionally, we demonstrated a T cell-dependent osteoclastogenesis from 21-OHD patients' PBMCs. T cells from patients expressed high levels of RANKL and low levels of osteoprotegerin (OPG) with respect to controls. Moreover, 21-OHD patients had higher soluble RANKL and lower OPG serum levels compared with controls; thus, soluble RANKL to OPG ratio was significantly higher in patients than controls. CONCLUSIONS: The present study showed for the first time a high osteoclastogenic potential of PBMCs from 21-OHD patients on cGC therapy. This spontaneous osteoclastogenesis seems to be supported by both the presence of circulating OCPs and factors released by T cells.

Effects of deferiprone on immune status and cytokine pattern in thalassaemia major.

OBJECTIVE: The present study was undertaken to evaluate the possible occurrence of immunological abnormalities in thalassaemia major patients treated with deferiprone (L1). METHODS: Longitudinal observational cohort study. RESULTS: The absolute number of CD8+ lymphocytes was high and the CD4/CD8 ratio low before L1 treatment; these parameters returned to normal after 3 months of L1 treatment. TNF-alpha, IL-2 and IL-2sRalpha were elevated before L1 treatment (11.83 +/- 1.75, 11.75 +/- 3.91, 1,409 +/- 621 pg/ml, respectively), while IL-6 was normal (2.58 +/- 0.79 pg/ml). After 12 months of treatment, IL-10 was higher than in previous periods, although always within the normal range. TNF-alpha, IL-2 and IL-2sRalpha returned to normal after 12, 6, and 3 months of L1 treatment, respectively.