Low-dose immune tolerance induction therapy in children of Arab descent with severe haemophilia A, high inhibitor titres and poor prognostic factors for immune tolerance induction treatment success.

INTRODUCTION: Immune Tolerance Induction (ITI) is the first-choice therapy to eradicate Factor VIII (FVIII) neutralizing antibodies in patients with haemophilia A (HA). There is limited published data on ITI from East Mediterranean countries. AIM: To assess the effectiveness of a low-dose ITI regimen to eradicate FVIII neutralizing antibodies in children with severe HA and high-titre inhibitors. METHODS: A prospective, single-arm study was conducted in children with HA (FVIII < 1 IU/dl), high-titre inhibitors and poor prognostic factors for successful ITI. Patients were treated with ∼50 IU/kg plasma-derived FVIII containing von Willebrand factor (pdFVIII/VWF) concentrate (Koate-DVI, Grifols) three times a week. Time to achieve tolerance, total and partial success were analysed after ITI. Annual bleeding rate (ABR), number of target joints, FVIII recovery and school absence were compared before and after ITI. RESULTS: Twenty patients with median (range) age of 6.2 (3-12) years and pre-ITI inhibitor titre of 36.5 (12-169) BU were enrolled. ITI lasted ≤12 months (early tolerization) in 45% of patients. Median follow-up was 12 months (3-22) and total response rate was 80% (60% total success; 20% partial success). Patients with two and three poor prognosis factors achieved overall success rate of 60% and 50%, respectively. ABR, target joints and school absence were reduced after ITI by 60%, 50% and 44.1%, respectively. In successful ITI tolerized patients, FVIII recovery was 90 (60-100)%. CONCLUSION: A low-dose ITI therapy using a pdFVIII/VWF concentrate achieved at least partial tolerance in 80% of patients, and reduced annual bleeds in children with high inhibitor titres and at least one poor prognosis factor for ITI treatment success.

Tissue-inhibitors of metalloproteinase-1 and vascular-endothelial growth-factor in severe haemophilia A children on low dose prophylactic recombinant factor VIII: Relation to subclinical arthropathy.

BACKGROUND: Subclinical synovitis occur long before clinical haemophilic arthropathy (HA). New biomarkers are needed for early detection of HA. AIM: To compare the levels of tissue inhibitors of metalloproteinase-1 (TIMP-1) and vascular endothelial growth factor (VEGF)in severe haemophilia A boys on prophylaxis and on-demand therapy to healthy boys and correlate them with the haemophilia joint health score (HJHS) & the Denver magnetic resonance imaging (MRI) scale; hence, determine their values in early detection of HA. METHODS: Haemophilia joint health score, serum TIMP-1, VEGF and Denver MRI score were assessed in 50 boys with severe haemophilia A (31 on prophylactic factor VIII therapy (62%) with a dose of 15 IU/kg/twice weekly) and 50 age-matched healthy boys. RESULTS: Boys with severe haemophilia A had significantly higher TIMP-1 240 ng/mL, SD200-350 (P < .001) and VEGF 600 pg/mL, SD400-1100 (P < .001). Their mean HJHS was 4.5 ± 3.0 (0-11) and their mean Denver MRI score was 5.55 ± 1.6 (2.00-8.00). A significant positive correlation was found between TIMP-1 and VEGF (P < .001), BMI Z-score (P = .029), HJHS (P = .041)and total MRI score (<.001). Significant correlations were found between VEGF and age (P < .001), HJHS (P = .003) and total MRI score (P < .001). Boys with severe haemophilia A on prophylaxis therapy had significantly lower HJHS (P = .021), VEGF (P < .001), TIMP-1 (P = .002) and total MRI score (P = .021) than those on on-demand therapy. Receiver operating characteristic curve, defined a cut-off value of 160 ng/mL for TIMP-1 with a sensitivity of 90% and specificity of 60% and that of 350 pg/mL for VEGF with a sensitivity of 78% and specificity of 88% for discrimination between severe haemophilia A and healthy boys. CONCLUSION: Vascular endothelial growth factor and TIMP-1 can be used for early detection of HA. Further prospective studies should include larger study populations. In addition, studies should address the role of various anti-VEGFs as potential therapy for HA and their impact on prevention and treatment of HA.

Effect of low-dose factor VIII prophylaxis therapy on bone mineral density and 25(OH) vitamin D level in children with severe haemophilia A.

BACKGROUND: Decreased bone mineral density (BMD) is a significant morbidity in haemophilia. Vitamin D is important for the bone health of people with haemophilia. Regular factor VIII prophylaxis can prevent bleeding and arthropathy. AIM: To determine the 25(OH) vitamin D level in severe haemophilia A patients and correlate it to their Hemophilia Joint Health Score (HJHS) and dual-energy X-ray absorptiometry (DEXA). We also compared the 25(OH) vitamin D and DEXA in haemophilia A and healthy children and in haemophilia A children on prophylaxis versus on-demand therapy. METHODS: Fifty severe haemophilia A patients were compared to 50 age-matched healthy boys. Patients were recruited from the Pediatric Hematology Clinic, Ain Shams University from May 2017 to April 2018. Full medical history was taken with emphasis on frequency of bleeding episodes, duration and amplitude of pain assessed by the pain score. Weight, height, body mass index and HJHS were assessed. 25(OH) vit-D3, calcium, phosphorus and alkaline phosphatase were measured. BMD was assessed using Lunar DEXA, paediatric software. RESULTS: People with haemophilia had significantly lower 25(OH) vit-D3 (P < .001) and DEXA z-score (P < .001) than controls. Seventy per cent of patients were on factor VIII prophylaxis twice weekly (15U/kg/dose). Significant difference was found regarding DEXA z-score (P = .012), 25(OH) vit-D3 (P = .033) and HJHS (P = .022) among patients on prophylaxis and on-demand therapy. CONCLUSION: Severe haemophilia A patients showed significantly lower 25(OH) vit-D3 and DEXA than controls. Hence, vitamin D deficiency should be tested in all people with haemophilia for early diagnosis and treatment. Low-dose prophylaxis in severe haemophilia preserves BMD and increases vitamin D. Further studies are required to evaluate the effect of different prophylaxis protocols on BMD and haemophilic arthropathy.

Angiopoietin-2 as a Marker of Retinopathy in Children and Adolescents With Sickle Cell Disease: Relation to Subclinical Atherosclerosis.

OBJECTIVES: Angiopoietin-2 (Ang-2) is a multifaceted cytokine that functions in both angiogenesis and inflammation. A proangiogenic state has been found in adults with sickle cell disease (SCD), mainly because of elevated Ang-2 levels. We determined Ang-2 level in 40 children and adolescents with SCD compared with 40 healthy controls and assessed its relation to retinopathy as well as carotid intimamedia thickness (CIMT). METHODS: Hematologic profile, serum ferritin, and serum Ang-2 were measured. CIMT was assessed using high-resolution ultrasound. Fundus examination was performed followed by fundus fluorescein angiography. Optical coherence tomography angiography (OCTA) was used to find small vascular changes not clinically manifested. RESULTS: Ang-2 levels and CIMT were significantly higher in SCD patients compared with controls. The incidence of nonproliferative retinopathy was 45%. SCD patients with retinopathy were older in age with a history of sickling crisis of >3 attacks per year and had a higher incidence of sickle cell anemia than sickle ß-thalassemia. Ang-2 cutoff value 9000 pg/mL could significantly detect the presence of retinopathy among SCD patients with 100% sensitivity and specificity. Serum Ang-2 levels were positively correlated with HbS and CIMT. Logistic regression analysis revealed that Ang-2 and HbS significantly contribute to retinopathy among patients with SCD. CONCLUSIONS: Elevated Ang-2 highlights the role of angiogenesis in the pathophysiology of SCD and may be considered a promising marker for screening of patients at risk of sickle retinopathy and vascular dysfunction.

Soluble Fas/FasL ratio as a marker of vasculopathy in children and adolescents with sickle cell disease.

OBJECTIVES: Sickle cell disease (SCD) is characterized by chronic inflammation due to ischemic tissue damage, accentuated during acute complications. Fas and its ligand (FasL) are members of tumor necrosis factor receptor superfamily and a major pathway for induction of apoptosis. Fas/FasL interactions may be related to augmentation of inflammatory response. We assessed the levels of sFas and sFasL in 35 children and adolescents with SCD compared with 35 healthy controls in relation to hemolysis, iron overload, sickle vasculopathy including kidney disease. METHODS: SCD patients, in steady state and asymptomatic for pulmonary hypertension, were studied stressing on hydroxyurea therapy, serum ferritin, urinary albumin creatinine ratio (UACR), high-sensitivity C-reactive protein (hs-CRP) and sFas/sFasL levels. RESULTS: sFas/sFasL ratio was significantly higher in patients compared with controls. sFas/sFasL ratio was elevated in patients with pulmonary hypertension, nephropathy and those who had history of frequent sickling crisis or serum ferritin ⩾2500. SCD patients treated with hydroxyurea had lower sFas/sFasL ratio than untreated patients. sFas/sFasL ratio was positively correlated to transfusion index, white blood cells, hs-CRP, serum ferritin and UACR. The cutoff value of sFas/sFasL at 8.75pg/mL could differentiate SCD patients with and without nephropathy while the cutoff value at 22pg/mL could differentiate SCD patients with and without pulmonary hypertension risk with high sensitivity and specificity. CONCLUSION: sFas/sFasL ratio may be considered as a marker for vascular dysfunction in SCD patients and is related to inflammation, iron overload and albuminuria level. Thus, it may be a reliable method to assess renal impairment in SCD.

The Association Between Circulating Levels of miRNA-181a and Pancreatic Beta Cells Dysfunction via SMAD7 in Type 1 Diabetic Children and Adolescents.

BACKGROUND: miRNA-181a has been implicated in autoimmunity and apoptosis. Therefore, this study was conducted to explore its possible role in pancreatic beta-cells dysfunction. METHODS: miRNA-181a expression was evaluated by real-time PCR in serum of 40 type 1 diabetic children and adolescents and 40 age- and gender-matched healthy controls. RESULTS: miRNA-181a expression was significantly higher in diabetic children and adolescents and it was negatively correlated to fasting C-peptide and SMAD7 levels. CONCLUSION: miRNA-181a appears to play a potential role in pancreatic beta-cells dysfunction via SMAD7.

Kallistatin as a marker of microvascular complications in children and adolescents with type 1 diabetes mellitus: Relation to carotid intima media thickness.

In diabetes, angiogenesis is disturbed, contributing to proliferative retinopathy, nephropathy and neuropathy. Kallistatin, a serine proteinase inhibitor, has anti-angiogenic effects. We assessed serum kallistatin in children and adolescents with type 1 diabetes as a potential marker for microvascular complications and its relation to carotid intima media thickness (CIMT). Sixty patients with type 1 diabetes were divided into two groups according to the presence of microvascular complications and compared with 30 healthy controls. High-sensitivity C-reactive protein (hs-CRP), HbA1c, urinary albumin creatinine ratio (UACR), kallistatin levels and CIMT were assessed. Kallistatin levels were significantly higher in patients with microvascular complications (9.9 ± 2.38 ng/mL) and those without complications (5.0 ± 1.5 ng/mL) than in healthy controls (1.39 ± 0.55 ng/mL; p<0.001). Kallistatin was increased in patients with microalbuminuria compared with the normoalbuminuric group (p<0.001). Positive correlations were found between kallistatin and disease duration, fasting blood glucose, HbA1c, triglycerides, total cholesterol, hs-CRP, UACR and CIMT (p<0.05). A kallistatin cut-off value at 6.1 ng/mL could differentiate patients with and without microvascular complications, with a sensitivity of 96.87% and specificity of 93.75%. Increased kallistatin levels in type 1 diabetes and its relation with CIMT may reflect vascular dysfunction and suggest a link between micro- and macro-angiopathy.

Circulating soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) as diagnostic and prognostic marker in neonatal sepsis.

OBJECTIVE: Triggering receptor expressed on myeloid cells-1 (TREM-1) is an important receptor involved in the innate inflammatory response and sepsis. We assessed soluble TREM-1 (sTREM-1) in 112 septic neonates (63 culture-positive and 49 culture-negative) and 40 healthy controls as a potential early diagnostic and prognostic marker for neonatal sepsis (NS). METHODS: Studied neonates were evaluated for early- or late-onset sepsis using clinical and laboratory indicators upon admission. sTREM-1 was measured on initial sepsis evaluation and at 48h after antibiotic therapy. For ethical reasons, cord blood samples were collected from control neonates and only samples from neonates that proved to be healthy by clinical examination and laboratory analysis were further analyzed for sTREM-1. RESULTS: Baseline sTREM-1 levels were significantly elevated in culture-proven (1461.1±523pg/mL) and culture-negative sepsis (1194±485pg/mL) compared to controls (162.2±61pg/mL) with no significant difference between both septic groups. Culture-positive or negative septic preterm neonates had significantly higher sTREM-1 compared to full term neonates. sTREM-1 was significantly higher in neonates with early sepsis than late sepsis and was associated with high mortality. sTREM-1 was significantly decreased 48h after antibiotic therapy compared to baseline or levels in neonates with persistently positive cultures. sTREM-1 was positively correlated to white blood cells (WBCs), absolute neutrophil count, immature/total neutrophil (I/T) ratio, C-reactive protein (hs-CRP) and sepsis score while negatively correlated to gestational age and weight. hs-CRP and sepsis score were independently related to sTREM-1 in multiregression analysis. sTREM-1 cutoff value of 310pg/mL could be diagnostic for NS with 100% sensitivity and specificity (AUC, 1.0 and 95% confidence interval [CI], 0.696-1.015) while the cutoff value 1100pg/mL was predictive of survival with 100% sensitivity and 97% specificity (AUC, 0.978 and 95% CI, 0.853-1.13). However, hs-CRP cutoff 13.5mg/L could be diagnostic for NS with a sensitivity of 76% and specificity of 72% (AUC, 0.762 and 95% CI, 0.612-0.925) and levels were not related to survival as no significant difference was found between dead and alive septic neonates. CONCLUSIONS: Elevated sTREM-1 could be considered an early marker for NS that reflects sepsis severity and poor prognosis.

D-dimer and portal vein status in splenectomized Egyptian ß-thalassemia major patients: a prospective single-thalassemia center experience.

Splenectomy is a recognized cause of portal vein thrombosis. Thirty-six ß-thalassemia major (ß-TM) patients were followed up for 36 months to evaluate changes in D-dimer levels (as a possible marker for thrombosis development) and portal vein status (by portal duplex ultrasound) at both early and late postlaparoscopic splenectomy periods. They were classified into group I if they were splenectomized in the study period (n = 12), or group II if they were splenectomized during the 5 years preceding the period (n = 24). In group I, D-dimer was measured 5 times: 1 day presplenectomy, the 1st week, 6th week, and 6th month postsplenectomy, and at the study end, whereas in group II, D-dimer was measured twice: at the study entry and end. Portal duplex was done 1 week postsplenectomy (group I) and at study end in both groups. Presplenectomy D-dimer levels in group I were significantly higher compared with the 6th month (P = .042) and study end (P = .03), whereas 1st week (postsplenectomy) D-dimer levels had a high mean of 3497.3 ng/mL, lowered at the 6th week (P = .017), at the 6th month (P = .008), and at study end (P = .005). D-dimer levels in group II showed no difference between study entry and end (P = .104). Portal vein "diameter and flow" were within normal findings in both groups. In this 3-year prospective study, a subclinical hypercoagulable state was detected 1 day prior to splenectomy and in the early postsplenectomy period, as evidenced by high D-dimer levels. Laparoscopic splenectomy was not associated with portal venous thrombosis either clinically or by duplex sonography.

Cognitive functions in children and adolescents with early-onset diabetes mellitus in Egypt.

The objective of this study was to investigate performance on memory, intelligence, and executive functions in children and adolescents with type 1 diabetes mellitus (T1DM) and to investigate the role of glycemic control, hypoglycemic attacks and diabetic ketoacidosis. We compared 50 subjects with T1DM with 30 healthy controls (ages between 7 and 16 years) using Benton Visual Retention Test (BVRT), the Arabic version of the Wechsler Intelligence Scale for Children (WISC), and Wisconsin Card Sorting Test (WCST). We also compared good versus poor glycemic control in T1DM subjects. RESULTS: T1DM subjects had significantly poorer performance than controls on all subtests of the BVRT, on all subscales of the WISC (verbal, performance and total IQ) and on most subtests of WCST (p < 0.05). T1DM subjects with good glycemic control performed significantly better than subjects with poor glycemic control on all subtests of the BVRT and on all subscales of the WISC (p < 0.05), but there was no difference on the WCST. T1DM subjects differed from controls on memory, intelligence, and executive functions. They also differed according to good or poor glycemic control (except on the WCST). Cognitive performance significantly correlated with a number of demographic and clinical variables.

Serum and Urinary Orosomucoid in Young Patients With Type 1 Diabetes: A Link Between Inflammation, Microvascular Complications, and Subclinical Atherosclerosis.

Orosomucoid is an acute-phase serum protein that is upregulated in urine samples of patients with diabetic nephropathy. We assessed serum and urinary orosomucoid levels in children and adolescents with type 1 diabetes and their relation to microvascular complications and carotid intima-media thickness (CIMT). Sixty patients with type 1 diabetes were divided into 2 groups according to the presence of microvascular complications and compared with 60 healthy controls. High-sensitivity C-reactive protein (hs-CRP), hemoglobin A1c (HbA1c), urinary albumin-creatinine ratio (UACR), serum and urinary orosomucoid, and CIMT were assessed. Both serum and urinary orosomucoid levels were significantly increased in patients with and without microvascular complications compared with controls, and the highest levels were in patients with complications (P < .001). Serum and urinary orosomucoid were higher in patients with microalbuminuria than normoalbuminuric group (P < .001). The cutoff value of urinary orosomucoid at 2825 ng/mL could differentiate patients with and without microvascular complications. Serum and urinary orosomucoid were positively correlated. Multiple regression analysis showed that HbA1c, UACR, hs-CRP, and CIMT were independently related to orosomucoid. We suggest that orosomucoid is a significant independent factor for diabetic microvascular complications and can be considered as an early marker of renal injury. High orosomucoid levels in type 1 diabetes reflect endothelial dysfunction and subclinical atherosclerosis.

Retinal changes in children and adolescents with sickle cell disease attending a paediatric hospital in Cairo, Egypt: risk factors and relation to ophthalmic and cerebral blood flow.

BACKGROUND: Sickle cell disease (SCD) is characterised by occlusion of small blood vessels. This study aimed to assess retinal changes in patients with SCD and its correlation with time-averaged mean flow velocity (TAMV) in middle cerebral arteries (MCA) and ophthalmic arteries (OA). METHODS: Sixty SCD patients (aged 3-18 years) attending a paediatric hospital in Cairo, Egypt, during March 2010 to November 2011, were compared with 30 healthy controls. All underwent clinical and fundus examination by indirect ophthalmoscopy, and assessment of TAMV in MCAs and OAs by transcranial Doppler, repeated 1 year later for those with conditional velocities. RESULTS: HbS/ß was diagnosed in 32 patients and HbSS in 28; 50 patients had normal fundus and 10 had bilateral non-proliferative retinopathy. Risk factors for retinopathy included HbSS, age, previous stroke, non-compliant hydroxyurea (HU) therapy, frequency of sickling crises and HbS level. TAMVs were increased in MCAs, but not in OAs, in sicklers. TAMVs in MCAs and OAs increased with non-compliant HU therapy, previous stroke, age, frequency of sickling crises and level of HbS. No significant interhemispheric difference was found. CONCLUSION: Sickle retinopathy was correlated with TAMV in MCAs but not in OAs. A significant difference was found between initial and follow-up TAMVs in the MCAs, after 1 year of regular HU and transfusion therapy, in those with conditional velocities.

Prevalence of Xmnl Gγ polymorphism in Egyptian patients with ß-thalassemia major.

BACKGROUND AND OBJECTIVES: ß-thalassemia results from a deficiency of ß-globin chains leading to an excess in a globin chains resulting in hypochromic microcytic red cells, ineffective erythropoiesis and hemolytic anemia. It is a result of a decline of HbF synthesis during the first year of life. F-cell levels are influenced by a sequence variant (C->T) at position -158 upstream of the -globin gene, so the frequency of the Xmnl Gγ polymorphism in Egyptian patients with b-thalassemia major needed evaluation to decide on the value of HbF augmentation drugs in treating Egyptian b-thalessemia. DESIGN AND SETTING: A cross-sectional study including 30 ß-thalassemia major patients diagnosed and attending the Pediatric Hematology Unit, Children's University Hospital, Ain Shams University, Cairo, Egypt, in the period from October 2008 to October 2009. PATIENTS AND METHODS: The 17 males and 13 females underwent a medical history and physical examination. Tests included a complete blood count, hemoglobin electrophoresis, serum ferritin, and detection of Xmnl Gγ polymorphism by PCR. RESULTS: The mean (SD) age was [2]10.2 (6.9) years. The most frequent genotype observed was homozygosity for the absence of the site Xmnl (-/-) in 96% of cases. Heterozygosity (+/-) genotype was detected in 4% of cases, while homozygosity for the site XmnI (+/+) genotype was absent. Genotype was not related to age at first transfusion, fetal hemoglobin level or transfusion frequency. CONCLUSION: Despite the small sample size, the study demonstrated that Egyptian ß-thalessemia patients have low frequency of positivity for the Xmnl polymorphism whether in heterozygous (+/-) or homozygous (+/+) state.