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Summary The discovery of serum biomarkers specific for paediatric lupus nephritis (pLN) will facilitate the non-invasive diagnosis, follow-up and more appropriate use of treatment. The aim of this study was to explore the role of serum high-mobility group box 1 (HMGB1) protein, antibodies against nucleosomes (anti-NCS), complement factor C1q (anti-C1q) and glomerular basement membrane (anti-GBM) in pLN. Serum samples of 42 patients with paediatric systemic lupus erythematosus (pSLE) (22 with pLN and 20 without renal involvement), 15 patients with other autoimmune nephritis (AN) and 26 healthy controls (HCs) were examined using enzyme-linked immunosorbent assay (ELISA). The activity of both pSLE and pLN was assessed by the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) tool. The levels of all four biomarkers were significantly higher in pLN compared to AN and to HCs. The anti-NCS, anti-GBM and HMGB1 serum levels were significantly higher in pLN than in pSLE without renal involvement. The anti-C1q and the HMGB1 serum levels were correlated positively with pSLE activity. The HMGB1 serum levels were also correlated positively with pLN activity. These findings suggest that serum anti-NCS, anti-GBM and HMGB1 may serve as biomarkers specific for the presence of nephritis in pSLE. HMGB1 emerged as a useful biomarker for the assessment of pLN and pSLE activity, whereas anti-C1q only of pSLE activity.

Sickle cell disease pathogenesis is a complex interplay of multiple factors associated with vascular endothelial activation, intense oxidative stress, and increased sickle cell adhesion. The aim of this study was to determine and compare three panels of plasma circulating biomarkers at 'steady state' and during veno-occlusive crises (VOC) in a cohort of children and adolescents with SCD and healthy controls. The following biomarkers were assessed: acute phase reactants, endothelial factors, and adhesion molecules. Forty-one SCD pediatric patients and 28 healthy children were enrolled. Patients at 'steady state' presented significantly elevated plasma levels of endothelin-1 (ET-1), soluble-VCAM-1 (sVCAM-1), soluble P-selectin (sP-selectin), and d-dimers compared to the control group. ET-1, sP-selectin, platelet-derived growth factor (PDGF), von Willebrand factor (vWf), d-dimers, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP) seems to represent additional, but not independent, prognostic markers of VOC crisis. Elevated plasma levels of sP-selectin, ET-1, and sVCAM-1 were associated with VOC frequency. The present study provides preliminary evidence of a possible association between these biomarkers and the endothelial activation at steady state and VOC in childhood SCD. Further prospective studies are required to confirm the potential independent prognostic value of these markers in different stages of pediatric SCD.

Our aim was to evaluate bone status in boys with haemophilia using dual energy X-ray absorptiometry (DXA) and quantitative ultraSonography (QUS), and in addition, to compare these two methods with the use of biochemical markers of bone turnover. Twenty-six boys with a mean decimal age of 12.08 ± 4.44 years were included in the study which included a DXA scan at lumbar spine and radial, as well as tibial QUS. Serum levels of soluble receptor activator of nuclear factor κB ligand (sRANK-L), osteoprotegerin (OPG) and osteocalcin (OC) were measured and joint evaluation was performed using the Hemophilia Joint Health Score (HJHS). With regard to the study results, only 2 of 26 patients (7.7%) had bone mineral density (BMD) Z-scores < -2, and 4 patients (15.4%) had BMD Z-scores between -1 and -2. Only one patient had radial and other two had tibial QUS Z-scores < -2. No agreement was recorded between QUS and DXA in identifying patients at risk for osteoporosis (k = 0.275, P = 0.063). Haemophiliacs had significantly higher serum levels of sRANK-L (21.04 ± 4.78 vs. 18.58 ± 2.28 ng mL(-1), P = 0.038) and of OC (5.35 ± 2.29 vs. 3.09 ± 0.61 ng mL(-1), P = 0.002) and significantly decreased levels of OPG (15.78 ± 2.53 vs. 23.79 ± 4.39 pg mL(-1), P < 0.001) compared with controls. QUS Z-scores at tibia significantly correlated with HJH Scores (r = -0.450, P = 0.040), whereas lumbar BMD Z-scores significantly correlated with body mass index Z-scores (r = 0.500, P = 0.009). More studies are warranted to identify the most accurate densitometric method for assessing bone status in haemophiliacs.

Children with steroid sensitive nephrotic syndrome (SSNS) is thought to have dysregulated type-1/type-2 cytokine network. Interleukin (IL)-18 is a cytokine, which may enhance both type-1 and type-2 responses, depending on the cytokines milieu. This prospective study aimed to assess type-1/type-2 cytokine synthesis and production profile in different stages of SSNS and define the potent involvement of IL-18. Twenty-three children with SSNS, aged 2.5-14 years, were studied; 23/23 both in active stage before treatment initiation and in remission still on steroids; 15/23 in remission off steroids as well. Data were compared with those obtained from 25 age-matched controls. The following parameters were assessed: Basic T cell populations, percentages of CD3+/CD69+/IFN-gamma+ and CD3+/CD69+/IL-4+ T cells as well as serum levels of IFN-gamma, IL-2, IL-4, IL-13 and IL-18. No difference in IL-2 levels was found between nephrotic children of all disease stages and controls (p0.05). Percentage of CD3+/CD69+/IL-4+ T cells and serum levels of IL-4, IL-13 and IL-18 were significantly higher in the active stage of SSNS compared with the remission stages and controls (p0.05). On the contrary, percentage of CD3+/CD69+/IFN-gamma+ T cells as well as serum IFN-gamma were significantly lower during active disease stage compared with remission stages and controls (p0.05). In children with SSNS, of all disease stages, serum levels of IL-18 were significantly correlated with both IL-4 and IL-13 (r=0.628 and p0.0001, r=0.71 and p0.0001, respectively). It seems that a type-2 cytokine synthesis and production pattern prevails in children with active SSNS and IL-18 expression is significantly correlated with this type-2 immune response.

Deferiprone (L1), has previously been reported to be associated with immunological abnormalities in iron loaded thalassemia patients. However, other factors may also have similar effects such as the level of iron overload, chronic immuno-stimulation due to transfusions, splenectomy and deferoxamine (DFO). During chelation therapy with DFO, several complications have been reported, which were due to pharmacological activity and high dose toxicity with regard to both acoustic and visual effects, as well as peripheral nerve disorders that were measured by nerve conduction velocities. The immune and neural status of 44 beta-thalassemic patients, aged 10-30 years (mean 19.4 +/- 4.9), receiving L1 as a monotherapy (n = 21), or in combination with DFO (n = 23), has been followed for 2 years by monitoring the level of immunoglobulins (IgG, IgM, IgA), the level of T and B lymphocytes (CD4/CD8), the auto antibodies: anti nuclear (ANA), anti-double-stranded (anti-ds DNA), anti reticulin (anti-R1), anti-extra nuclear (anti-ENA), anti histone (anti-AHA), anti liver-kidney-muscle (anti-LKM), anti-smooth muscle (anti-SMA), anti-thyroid (anti-ATA), anti-mitochondrial (anti-AMA) antibodies and the C-reactive protein. The percentage of patients with disorders of the immune and nervous system concerned very few cases. None of our patients with pathological findings in their immunological or neurophysiological examinations presented any signs or symptoms of involvement of the immune or nervous system. Further advantages have been identified for the oral use of L1 and its combination with DFO, including synergistic efficacy and lower dosing with limited toxicity.

The objective of this study was to investigate the effect of treatment with recombinant human granulocyte-colony stimulating factor (rhG-CSF) on the neutrophil count and function of preterm neonates with documented sepsis. For this purpose 62 preterm neonates with proven sepsis and 19 healthy preterm ones were studied. Of the 62 patients, 27 septic neonates had an absolute neutrophil count (ANC)5000/mm3 (group A) and were scheduled not to receive rhG-CSF and 35/62 had an ANC5000/mm3 (n=35) and were randomly assigned either to receive rhG-CSF (group B) or not to receive it (group C). rhG-CSF (10 microg/kg) was administered for 3 consecutive days (0, 1, 2). The ANC, plasma levels of G-CSF (ELISA), neutrophil respiratory burst activity (NRBA) and neutrophil expression of CD11a, CD11b and CD11c (flow cytometry) were measured in all septic neonates on days 0 (onset of sepsis), 1, 3 and 5 and in the healthy neonates once within the first 2 days of life. We found that on day 0, G-CSF levels of all groups of septic neonates were significantly higher than those of the healthy ones. The highest levels were observed in group A. NRBA was diminished only in groups B and C and the expression of CD11a and CD11c was reduced in all groups of septic neonates. Administration of rhG-CSF resulted in a rapid and significant increase in ANC, NRBA and CD11a, CD11b and CD11c expression that persisted throughout the follow up. CONCLUSION; The administration of granulocyte colony stimulating factor to septic neonates significantly increases the absolute granulocyte count and enhances the neutrophil respiratory burst and beta2 integrin expression.

We investigated the simultaneous changes in serum levels of HMGB1 and IFN-α as well as in LAIR-1 expression on plasmatoid dendritic cells (pDCs) of juvenile systemic lupus erythematosus (jSLE) patients in order to explore their involvement in the disease pathogenesis and their correlation with disease activity and other characteristics. In total, 62 blood samples were studied from 26 jSLE patients (18 girls), aged 8–16 years. Twenty healthy subjects (16 girls) of comparable age were included as healthy controls (HCs). Concentrations of serum HMGB1 and IFN-α were assessed by ELISA and LAIR-1 expression on pDCs by five-color flow cytometry. The disease activity index was assessed by SLEDAI and ECLAM scores. It was found that mean serum levels both of HMGB1 and IFN-α were significantly increased in jSLE patients compared to HCs and in jSLE patients with active disease with or without active nephritis compared to those with inactive disease. Mean serum levels of HMGB1 were positively correlated with levels of IFN-α and both were positively correlated with the SLEDAI and ECLAM scores. The expression of LAIR -1 on pDCs of jSLE patients was significantly lower than that of HCs. In conclusion, our findings indicate that serum HMGB1 not only represents a potential marker of disease activity but together with the lack of LAIR-1 inhibitory function may contribute to the sustained inflammatory action of IFN-α in jSLE. In this regard, blocking the action of HMGB1 and its receptors or enhancing the expression/inhibitory function of LAIR-1 on pDCs should be included in future immune interventions for controlling jSLE.

The aim of this study was to evaluate the impact of prematurity, sepsis and stress on the neutrophil respiratory burst activity (NRBA) of neonates. For this purpose 122 healthy neonates (89 term and 33 preterm), 33 preterm stressed neonates, 59 septic neonates (12 term and 47 preterm) and 26 healthy adults were studied. The NRBA was assessed after in vitro stimulation by PMA using a whole blood flow cytometric microassay with dihydrorhodamine 123 (DHR 123). It was found that the percentage of responding neutrophils in term neonates was comparable to that found in adults (medians 83.5 and 89.8%, respectively), whereas it was significantly lower in the healthy preterm neonates (median 70.6%, p < 0.05). The NRBA was further depressed in the stressed (median = 63%) and septic neonates, both term and preterm (medians 60.5 and 54.3%, respectively). No correlation with the levels of G-CSF, TNF-α and IL-1β, which were found to be higher in the stressed and septic neonates, was observed. These findings indicate that prematurity, sepsis and stress significantly depress the respiratory burst activity of neonatal neutrophils.

Our aim was to evaluate bone status in boys with haemophilia using dual energy X-ray absorptiometry (DXA) and quantitative ultraSonography (QUS), and in addition, to compare these two methods with the use of biochemical markers of bone turnover. Twenty-six boys with a mean decimal age of 12.08 ± 4.44 years were included in the study which included a DXA scan at lumbar spine and radial, as well as tibial QUS. Serum levels of soluble receptor activator of nuclear factor κB ligand (sRANK-L), osteoprotegerin (OPG) and osteocalcin (OC) were measured and joint evaluation was performed using the Hemophilia Joint Health Score (HJHS). With regard to the study results, only 2 of 26 patients (7.7%) had bone mineral density (BMD) Z-scores-2, and 4 patients (15.4%) had BMD Z-scores between -1 and -2. Only one patient had radial and other two had tibial QUS Z-scores-2. No agreement was recorded between QUS and DXA in identifying patients at risk for osteoporosis (k = 0.275, P = 0.063). Haemophiliacs had significantly higher serum levels of sRANK-L (21.04 ± 4.78 vs. 18.58 ± 2.28 ng mL(-1), P = 0.038) and of OC (5.35 ± 2.29 vs. 3.09 ± 0.61 ng mL(-1), P = 0.002) and significantly decreased levels of OPG (15.78 ± 2.53 vs. 23.79 ± 4.39 pg mL(-1), P0.001) compared with controls. QUS Z-scores at tibia significantly correlated with HJH Scores (r = -0.450, P = 0.040), whereas lumbar BMD Z-scores significantly correlated with body mass index Z-scores (r = 0.500, P = 0.009). More studies are warranted to identify the most accurate densitometric method for assessing bone status in haemophiliacs.

Bronchoalveolar lavage (BAL) is a useful bronchoscopic technique. Studies in "normal" children are limited.To provide data on BAL reference values from Greek children and compare BAL cellular and noncellular components in children with inflammatory and non-inflammatory lung diseases.Seventy two children, aged 2.5 months to 16 years, underwent diagnostic bronchoscopy and BAL. Patients were divided in two groups whether lung inflammation was absent or present. Differential cytology, flow cytometry for lymphocyte subsets and cytokine and chemokine measurements were performed on BAL fluid.Alveolar macrophages were the predominant cellular population in normal children. Patients with inflammatory pneumonopathies had significantly more neutrophils. There was no difference in lymphocyte subpopulations. Values of CD4+/CD8+ ratio in BAL was similar to that reported in adults. Levels of IL-8 and TNF- alpha were significantly higher in children with inflammatory lung diseases.This study provides the first data on BAL of "normal" Greek children. BAL from patients with pulmonary inflammation was characterised by neutrophilia. Finally, we propose that measurement of IL-8 and TNF-a levels in BAL could help in early identification of inflammation in the tracheobronchial tree.

There are limited studies on renal involvement in β-thalassemia, mainly involving patients on deferoxamine, reporting both glomerular and tubular dysfunction. The aim of the present study was to investigate renal involvement in young thalassemia patients, using both conventional and early markers of renal dysfunction, and to correlate findings to iron chelation therapy. Forty-two patients aged 4–23 years were studied and, for analysis purposes, were divided into two groups based on chelation therapy (group A receiving deferasirox and group B receiving deferoxamine and deferiprone combination therapy). In addition to conventional renal biochemistries, creatinine clearance, estimated glomerular filtration rate, serum cystatin C (Cys C), fractional excretion of sodium, tubular phosphorus reabsorption and urine calcium, protein, β2-microglobulin (β2-MG) and glucose levels were measured. A considerable number of patients demonstrated impaired renal function with elevated Cys C levels (36%), glomerular dysfunction with proteinuria (24%) and tubulopathy with hypercalciuria (35.5%) and elevated excretion of β2-MG (33.5%). Renal involvement seems to be present even in young patients with β-thalassemia, therefore, routine use of early markers of renal dysfunction is recommended. Further studies are needed in order to investigate the role of new chelators in tubular function parameters.

Background The course and outcome of oligo-JIA is more or less unpredictable. So far, no internationally acceptable prognostic biomarkers, regarding the progression of oligo- to the extended disease type have been defined. Objectives The aim of this study was to assess the immunophenotype plus cytokine profile in synovial fluid (SF) of patients (pts) with JIA and to evaluate the findings in respect to the severity of the disease course in order to define those pts who need early aggressive treatment. Methods 73 SF samples from the knees of 57 JIA patients (F: 48, median age 11.49 yrs) were studied. 20/57 pts had oligo-persistent (O-per), 27 oligo-extended (O-ext) and 10 polyarthritis (poly). 25 age-matched children (25 SF samples) with recent traumatic knee arthritis served as controls. The SF immunophenotype and cytokine levels were assessed by flow cytometry and ELISA respectively. Results The median CD4:CD8 T cell ratio was found to be significantly lower in 19/57 pts (24/73 samples) from all 3 JIA pt groups as compared to the controls (0.75 vs 1.55, p Conclusions A low ratio of CD4:CD8 T cells in combination with high levels of IL-17 and IL-23 in SF of JIA pts can predict a prolonged, polyarticular course of the disease and may be used as “the window of opportunity” for a more targeted treatment. References Hunter PJ, et al. Biologic predictors of extension of oligoarticular juvenile idiopathic arthritis as determined from synovial fluid cellular composition and gene expression. Arthritis Rheum 2010;62: 896-907. Disclosure of Interest None declared

SummaryThe aim of this study was to investigate the effect of prematurity, neonatal sepsis, respiratory distress syndrome (RDS) and perinatal asphyxia on monocyte HLA-DR expression of neonates using a flow cytometric method based on monocyte negative selection. The subjects were one hundred and thirty-one neonates (59 healthy, 44 septicaemic, 20 with RDS and eight with perinatal asphyxia) and 20 healthy adults. Monocyte HLA-DR expression was measured using one-colour HLA-DR labelling in a gate for monocytes obtained using the combination of CD3-CD19–PE/CD15–FITC MoAbs. In addition, the common dual staining method using MoAbs against two CD14 epitopes (TUK4, MO2) was evaluated. With the one-colour HLA-DR labelling higher purity and recovery values of monocytes were achieved than with the dual labelling method. Healthy neonates had significantly lower percentages of HLA-DR+ monocytes than adults (69 ± 13% versus 91·5 ± 2·5%) and comparable mean fluorescence intensity (MFI) (119 ± 25 versus 131 ± 26). Values did not differ significantly between healthy term and preterm neonates. Preterm neonates with RDS had a significantly lower percentage of HLA-DR+ monocytes than the healthy preterm neonates. In neonates with asphyxia both parameters were comparable to those of the healthy ones. Septicaemic neonates presented significantly lower values of both parameters than the healthy, RDS and asphyxiated neonates. Monocyte negative selection provides a reliable estimation of HLA-DR expression on monocytes. Expression of monocyte HLA-DR is lower in healthy neonates in comparison with adults and is further decreased in neonates with sepsis and RDS, but it is not influenced by prematurity and perinatal asphyxia.

Sir, Pathogenesis of steroid-sensitive nephrotic syndrome (SSNS) is thought to be related mainly to T-cell dysfunction [1]. However, the beneficial use of rituximab in cases of frequently relapsing SSNS provided evidence of B cell involvement [2–4]. Our aim was, thus, to investigate prospectively the levels of the circulating CD19+ B cells in children with a first episode of SSNS in sequential stages (presentation, remission on steroids and remission off steroids). Twenty-three children (M/F = 13/10, age = 2.5–14 years, median = 4.32 years) with a first episode of SSNS were studied; 23/23 both at presentation (before steroids initiation) and in remission on steroids (40 mg/m2 on alternate day); 15/23 were tested as well in remission off steroids for at least 6 months. Twenty-five age-matched children who had come to the outpatient haematology clinic in order to be tested for b-thalassaemia trait were found to be negative and served as healthy controls (controls 1). Considering that the presentation of SSNS may be associated with a recent infection, mainly a respiratory tract infection, twenty age-matched children with an upper respiratory tract infection acted as a second control group (controls 2). The percentages of CD3± T cells, as a pan T-cell marker, and the percentages of CD19+ and CD20± B cells were evaluated in all children. The above-mentioned parameters were determined in each sample by flow cytometry using the lysed whole blood method. The duochrome phycoerythrin-cyanin5 (PE-Cy5) conjugated CD3± monoclonal antibody (MoAb) and phycoerythrin (PE) conjugated CD19+ and CD20± MoAbs purchased from Beckman Coulter were used. The samples were analysed with a EPICS-XL flow cytometer. The results were expressed as percentages (%) of fluorescence-positive cells as well as actual numbers (cells/μL) of the circulating CD19+ and CD20+ B cells, based on the white blood cell count. Statistical analysis was performed using the Statistical Package for the Social Sciences for Windows (SPSS version 11.5). The paired t-test and independent-samples t-test were used to compare differences between study groups with and without paired data, respectively. Pearson's coefficient of correlation (r) was used to determine the correlations. A P ≤ 0.05 was considered to be statistically significant. In 5 of 23 children with a first episode of SSNS, there was a recent history of an upper respiratory tract infection. Remission was achieved in all children within 6–15 days after steroid initiation. During remission, all patients presented normoalbuminaemia and were free of proteinuria and albuminuria. Percentages of CD3± T cells were found to be within normal limits in all patients (at presentation of SSNS, in remission still on steroids and in remission off steroids) compared with the two control groups (P ≥ 0.05). As depicted in Figure ​Figure1,1, the circulating CD19+ B cells were significantly higher at presentation of SSNS (mean percentage = 18.13 ± 5.43, mean actual number = 695.34 ± 258.29) compared with remission on steroids (mean percentage = 13.57 ± 4.22 and P 0.05). Moreover, circulating CD19+ B cells were inversely correlated with disease activity (r = −0.465, P 0.05). Fig. 1 Percentage of CD19+ B cells in children with a first episode of SSNS in sequential stages (presentation, remission on steroids, remission off steroids) and in controls 1 and 2. The pathogenesis of SSNS has mainly been focused on T cells dysfunction. The success of rituximab, a chimeric anti-CD20 antibody, in the treatment of cases of frequently relapsing SSNS initiated interest in pathogenic pathways involving B cells [2–4]. There is an interaction between B cells and T cells, and B cells are involved in the presentation of antigens to T cells. However, the effect of B-cell depletion on T-cell function is unknown. Kemper et al. [5] suggested that in children with SSNS, there is a combined T- and B-cell activation. In agreement with our results, recent studies showed that CD19+ B cells may be elevated in relapsing nephrotic children [6,7]. Children with both steroid-sensitive and steroid-resistant nephrotic syndrome were included in this study, and there were no paired data of the circulating CD19+ B cells in remission off steroids. It is worth noting that in our study not all children with SSNS presented elevated CD19+ B cells. We intend to follow up this cohort of 23 nephrotic children in order to identify whether the long-term outcome of the nephrotic syndrome is associated with the up-regulation of CD19+ B cells in these patients taking into account steroid dependency and frequency of relapses. Our findings demonstrated an up-regulation of circulating CD19+ and CD20± B cells in children with a first episode of SSNS. This indicates that B cells may play a role in SSNS and that rituximab may be considered in certain cases. Conflict of interest statement. None declared.

Background Nephritis is the most common severe manifestation of pediatric Systemic Lupus Erythematosus (pSLE) and the major predictor of poor prognosis. Research for the detection of novel and accurate biomarkers, predictive of the nephritis outcome, is globally in progress. Ethnicity affects the disease outcome but data on purely Caucasian populations are still limited. Objectives The exploration of the role of serum biomarkers, namely anti-nucleosome antibodies (anti-NCS), anti-C1q antibodies (anti-C1q), anti-glomerular basement membrane antibodies (anti-GBM) and high mobility group box 1 (HMGB1) in pSLE nephritis and the association of their levels with pSLE nephritis and pSLE disease activity. Methods Twenty-four patients (16 female) with pSLE nephritis (44 serum samples, 22 in active nephritis) and 21 patients (18 female) with pSLE without nephritis (32 serum samples, 19 in active pSLE) were enrolled in the study. The disease control group included 17 patients with nephritis of other causality (Henoch-Schonlein purpura nephritis, IgA nephropathy, postinfectious glomerulonephritis, membranous glomerulonephritis), who provided equal serum samples. The SLICC renal activity score was applied for assessing pSLE nephritis disease activity and ECLAM for global pSLE disease activity. The biomarkers’ levels were determined by ELISA. Results The pSLE nephritis patients had significantly higher serum levels of anti-NCS [median: 48.89 (IQR: 31.48-80.81) U/ml versus 12.5 (11.5-27.8) U/ml, p Conclusions In this pure Caucasian Northern Greek pSLE population, high serum anti-GBM levels were found to be associated with the presence of nephritis, but not with the disease activity. Serum anti-GBM, anti-NCS, anti-C1q and HMGB1 may be used to differentiate patients with pSLE nephritis from patients with nephritis of other causality. Furthermore, serum anti-NCS and anti-C1q may be useful for the estimation of pSLE disease activity. References Mok CC. Biomarkers for lupus nephritis: a critical appraisal. J Biomed Biotechnol. 2010;2010:638413. Petri M et al. Systemic lupus international collaborating clinics renal activity/response exercise: development of a renal activity score and renal response index. Arthritis Rheum. 2008 Jun;58(6):1784-8. Disclosure of Interest None Declared

Aiming to define the evolution pattern of 10 acute-phase proteins in early infancy, we measured nephelometrically the serum concentrations of albumin, prealbumin, retinol-binding protein, transferrin, ceruloplasmin, hemopexin, haptoglobin, alpha 1-acid glycoprotein, alpha 2-macroglobulin, and alpha 1-antitrypsin in 395 term and preterm infants (gestational ages 26-41 weeks). Measurements were performed within 24 h after birth and then at the end of 1 (n = 171), 3 (n = 155), and 6 (n = 90) months afterwards. Data obtained from 250 healthy adults were used as adult reference values. All proteins increased progressively with postnatal age, except for alpha 1-antitrypsin, which remained stable from birth to the 6th month. Concentrations of almost all measured proteins were significantly lower in preterm than in term infants in the first 3 months. Compared with adult values, alpha 2-macroglobulin and alpha 1-antitrypsin were higher in infants throughout the 6 months. The other proteins were significantly lower at birth than adult values but after 6 months, only albumin, prealbumin, retinol-binding protein, and alpha 1-acid glycoprotein still remained lower in infants. Thus both gestational and postnatal age should be considered when interpreting concentrations of these proteins in early infancy.

INTRODUCTION: Nephrotic syndrome (NS) is thought to be associated with primary immune disturbances. OBJECTIVE: The aim of our study was to investigate prospectively the immune disturbances in steroid-sensitive (SS) and steroid-resistant (SR) NS and identify whether these immune disturbances may predict the response to steroid therapy. METHODS: Thirty children with SS NS and 7 children with SR NS (aged 2 to 14 years) were studied. To evaluate the possible relationship between immune disturbances and response to treatment, patients were evaluated during different disease stages. Data were compared with those obtained from 25 age-matched controls. The following parameters were assessed: basic B- and T-cell populations, percentages of CD23+, CD3+/CD69+/interferon γ+ (IFN-γ+) cells, and CD3+/CD69+/interleukin 4+ (IL-4+) T cells, and serum levels of IL-13 and IL-18. RESULTS: In patients with SS NS percentages of CD23+ and CD19+ B cells, CD3+/CD69+/IL-4+ T cells and serum levels of IL-13, IL-18 were significantly higher in the active stage compared with the remission stage on steroids, remission off steroids, and controls (P < .05). On the contrary, percentages of CD3+/CD69+/IFN-γ+ T cells were significantly decreased (P < .05). In patients with SR NS, percentages of CD23+B cells, CD3+/CD69+/IL-4+ T cells, and serum levels of IL-13 and IL-18 presented no significant difference between active stage and partial remission. Percentages of CD19+ B cells and CD3+/CD69+/IFN-γ+ T cells were elevated in active stage compared with remission stage of patients with SR NS and in controls (P < .05). CONCLUSIONS: These findings suggest that when a type-2 immune response is found in the active stage of NS, one could predict a good response to steroid therapy.

Background: Neutropenia is a common manifestation in neonates with intrauterine growth retardation (IUGR) related to preeclampsia and has been associated with decreased neutrophil production. In addition, the increased serum levels of soluble Fas Ligand (sFasL) found in neonates born to mothers with preeclampsia have been associated with increased apoptosis. Neutropenia is uncommon in neonates with isolated IUGR. AIM. To assess neutrophil apoptosis in neonates with IUGR born to normotensive mothers. Material and methods: Neutrophil apoptosis was studied in 19 neonates with IUGR born to mothers without hypertension and 19 gestational age matched, appropriate for gestational age (AGA) neonates during the first 12 hours after birth. Neutrophil apoptosis was evaluated by flow cytometric assessment of neutrophils stained with fluorescein isothiocynate (FICT)-annexin-V, propidium iodide (PI) or expressing the Fas molecule. Results: No significant difference in the percentage of neutrophills stained with FICT-annexin-V (mean7,1%, SD 3,9% vs mean 7,8% SD 2,5%, respectively) or with PI (mean 7,6%, SD 4,3% and mean 7,8%, SD 3,4%, respectively) was found between the IUGR and the AGA neonates. Also the percentage of neutrophills expressing the Fas did not differ significantly between the IUGR (mean 30,2%, SD 11,3%) and AGA neonates (mean 24,8%, SD 9,3%). Conclusion: Neutrophil apoptosis in IUGR neonates of normotensive mothers is not increased compared to AGA neonates. We speculate that neutropenia in IUGR is rather attributed to preeclampsia than intrauterine starvation.

Effect of Cetirizine on Circulating Indicators of Persistent Inflammation and Endothelial Cell Activation in Patients With Sickle Cell Disease

ELEVATED CIRCULATING INDICATORS OF PERSISTENT INFLAMMATION AND ENDOTHELIAL CELL ACTIVATION IN PATIENTS WITH SICKLE CELL DISEASE. 110

Increased proportions of CD10, CD19 and HLA-DR+ cells in bone marrow specimens of children with nonmalignant hematologic conditions have been recently identified. In the last 3 years we have been following up 11 children with hematologic disorders who revealed an abnormal immunophenotypic pattern in their bone marrow specimen. The objective of this study was to evaluate this abnormality in association with the patients' clinical course and development of a B-lineage malignancy. The study population consisted of 11 children aged 4 mo to 13 yrs (mean 4,8 yrs) who had undergone bone marrow diagnostic procedures for haematological disorders: neutropenia (2), thrombocytopenia (8) and Fanconi anemia (1). Bone marrow morphological studies in smears and biopsy samples revealed increased numbers of immature cells(28-60%) in 2/11 patients. Flow cytometry analysis of bone marrow biopsy samples (XL-Coulter) showed in all patients an increased percentage of immature B - cell precursors which expressed the lymphocyte surface markers CD19, CD10 (CALLA) and HLA-DR. Results from the immunophenotyping analysis were expressed as percentage of positive cells. The mean values of CD19+, CD10 and HLA-DR+ cells were 46% (±18,5), 44,5%(±23) and 56,3% (±22) respectively. Patients were treated with the standard protocole treatment according to their diagnosis and were followed up for 6 mo - 3 yrs (mean 2 yrs). Clinical and laboratory evaluation during the follow up time show that 9/11 children have recovered completely and 2/11 are still on treatment for their cytopenia. Nevertheless, none of these children has developed malignant lymphoproliferative disease. These findings indicate that an abnormal immunophenotypic pattern is not always consistent with heamatologic malignancy. However, patients with all these findings must have a long-term follow up in order to reach a definite diagnosis.

The evolution of the main serum opsonins in neonates and infants of varying gestational age was investigated to provide reference values for these opsonins in early infancy. Serum concentrations of immunoglobulins, IgG subclasses, C3, C4 and fibronectin were serially measured from birth until the age of 6 months in term and preterm infants. Measurements were performed by rate nephelometry. Five hundred and sixty six neonates (gestational age 26-41 weeks) were examined at birth, 233 at 1 month, 218 at 3 months, and 147 at 6 months, respectively. The same measurements were performed in 54 pairs of neonatal/maternal samples and in 230 apparently healthy adults. Gestational age had a significant impact on serum IgG, IgG subclasses, C3 and C4 up till the third month, and on fibronectin until the first month. No such impact was observed for IgA and IgM. Sixteen per cent of the neonates had IgM concentrations higher than 0.2 g/l at birth, suggesting that the critical concentration of serum IgM at birth for suspected intrauterine infection should be reconsidered. Concentrations of all opsonins at birth were significantly lower than adult reference values. They only approached or even reached adult values by the third or the sixth month. Data from analysis of the neonatal and the corresponding maternal sera indicate that there is a preferential active transplacental transport of IgG subclasses in the order of IgG1, IgG3, IgG2 and IgG4. These results show that concentrations of immunoglobulins, C3, C4 and fibronectin undergo changes during the first months of life, depending not only on the infants9 postnatal age but also on gestational age.

Aiming to define the evolution pattern of 10 acute-phase proteins in early infancy, we measured nephelometrically the serum concentrations of albumin, prealbumin, retinol-binding protein, transferrin, ceruloplasmin, hemopexin, haptoglobin, alpha 1-acid glycoprotein, alpha 2-macroglobulin, and alpha 1-antitrypsin in 395 term and preterm infants (gestational ages 26-41 weeks). Measurements were performed within 24 h after birth and then at the end of 1 (n = 171), 3 (n = 155), and 6 (n = 90) months afterwards. Data obtained from 250 healthy adults were used as adult reference values. All proteins increased progressively with postnatal age, except for alpha 1-antitrypsin, which remained stable from birth to the 6th month. Concentrations of almost all measured proteins were significantly lower in preterm than in term infants in the first 3 months. Compared with adult values, alpha 2-macroglobulin and alpha 1-antitrypsin were higher in infants throughout the 6 months. The other proteins were significantly lower at birth than adult values but after 6 months, only albumin, prealbumin, retinol-binding protein, and alpha 1-acid glycoprotein still remained lower in infants. Thus both gestational and postnatal age should be considered when interpreting concentrations of these proteins in early infancy.

Objective Recent data in adult Rheumatoid Arthritis support that the Th17 cell-derived cytokine interleukin 17 (IL-17) in the presence of IL-6 and IL-23 plays a critical role in the pathogenesis of chronic destructive arthritis. Data on synovial fluid (SF) concentrations of IL-17 in JIA pts are sparse. We measured concentrations of the above 3 cytokines and assessed the CD4+CD25highFoxP3+ (Treg) and CD4+CD25lowFoxP3T cell subpopulations in the SF of children with JIA. Findings were correlated with SF sRANKL which expresses the osteoclastic activity in active disease.

Familial Mediterranean Fever (FMF), is the second commonest autoinflammatory disease in pediatric Greek patients (pts) after PFAPA. So far, long-term follow-up case series in Greek FMF pts have not been emerged.