SERS-Based Evaluation of the DNA Methylation Pattern Associated With Progression in Clonal Leukemogenesis of Down Syndrome.

Here we show that surface-enhanced Raman scattering (SERS) analysis captures the relative hypomethylation of DNA from patients with acute leukemia associated with Down syndrome (AL-DS) compared with patients diagnosed with transient leukemia associated with Down syndrome (TL-DS), an information inferred from the area under the SERS band at 1005 cm-1 attributed to 5-methycytosine. The receiver operating characteristic (ROC) analysis of the area under the SERS band at 1005 cm-1 yielded an area under the curve (AUC) of 0.77 in differentiating between the AL-DS and TL-DS groups. In addition, we showed that DNA from patients with non-DS myeloproliferative neoplasm (non-DS-MPN) is hypomethylated compared to non-DS-AL, the area under the SERS band at 1005 cm-1 yielding an AUC of 0.78 in separating between non-DS-MPN and non-DS-AL. Overall, in this study, the area of the 1005 cm-1 DNA SERS marker band shows a stepwise decrease in DNA global methylation as cells progress from a pre-leukemia to a full-blown acute leukemia, highlighting thus the potential of SERS as an emerging method of analyzing the methylation landscape of DNA in the context of leukemia genesis and progression.

The value of abdominal ultrasonography compared to colonoscopy and faecal calprotectin in following up paediatric patients with ulcerative colitis.

AIM: To evaluate the value of abdominal ultrasonography (US) in the follow-up of paediatric patients with ulcerative colitis (UC) compared to faecal calprotectin (FC) and colonoscopy. MATERIAL AND METHOD: In this retrospective study we enrolled 30 paediatric patients previously diagnosed with UC, examined by abdominal US and colonoscopy within the same week. FC was also determined during the same week. Disease activity was established using the paediatric ulcerative colitis activity index (PUCAI). The global endoscopic activity was evaluated using the Mayo endoscopic subscore. RESULTS: Endos-copy revealed pathological findings of active disease in 27 out of 30 patients; 3 patients were in endoscopic remission. Only 18 of them had clinical active disease (PUCAI >10), [sensitivity (Se) 66.7% and specificity (Sp) 33% of PUCAI in detecting endoscopic active disease). Twenty-three (76.7%) patients had FC >250 mcg/g, but in 2 of these cases the colonoscopy was normal (Se 77.8% and Sp 33.3% in detecting active disease). At US examination, pathological findings (increased bowel wall thickness, hypervascularity, lymphadenopathies, and/or mesenteric inflammatory fat) were found in 27 patients (90%), all with endoscopic active disease (agreement US - colonoscopy, at patient level, k=1.0, p<0.001, Se 100% and Sp 100%). At seg-ment level (totally 180 bowel segments examined by US), the overall agreement between US and colonoscopy was k=0.767, p<0.001, Se 86.5%, Sp 90.1%. Of the 27 patients with US pathological findings in any of colonic segments, 23 had FC >250 mcg/g (85.1%). The inter-observer agreement for the US measurements had an overall ICC of 0.926 with p<0.001. CONCLUSION: Abdominal US findings demonstrate a good to excellent concordance with endoscopic examination and are correlated with elevated FC levels. Therefore, US appears as an accurate technique in assessing activity in patients with UC and might replace colonoscopic evaluation for the follow-up.

B cell-activating factor (BAFF) in children with inflammatory bowel disease.

BACKGROUND: Assessing the inflammation is important in the follow-up of paediatric patients with inflammatory bowel disease (IBD). We aim to evaluate the value of B cell-activating factor (BAFF) in paediatric IBD as a potential biomarker for follow-up. METHOD: We determined BAFF in serum and faeces and faecal calprotectin (CP) in 32 IBD children-16 Crohn's disease (CD) and 16 ulcerative colitis (UC). Twenty-six healthy children and 10 children with irritable bowel syndrome (IBS) were included as controls. RESULTS: No differences were found in serum BAFF between IBD, IBS, and healthy group: 1037.35, 990.9 and 979.8 pg/ml, respectively, all p > 0.05, but faecal BAFF was higher in the IBD group: 15.1, 8.5 and 8.2 pg/ml, respectively, p < 0.05, and higher in the UC group (55.975 pg/ml) compared to the CD group (10.95 pg/ml), p = 0.015. Splitting the IBD group in relation to the CP level, the serum BAFF had no significantly different values between the subgroups, but the faecal BAFF was significantly higher in the >250 µg/g subgroup. Cut-off values of BAFF were calculated. CONCLUSION: Faecal BAFF is a promising marker for monitoring the children with IBD, higher levels of BAFF being correlated with high CP. IMPACT: Faecal BAFF is a promising marker in monitoring the children with IBD, higher levels of BAFF being correlated with high faecal calprotectin. To our knowledge, this is the first paediatric study concerning BAFF evaluation in IBD. Faecal BAFF levels could be considered a potential non-invasive marker in monitoring IBD activity in paediatric population with clinically mild or inactive disease.

MicroRNA-155-5p Plays a Critical Role in Transient Leukemia of Down Syndrome by Targeting Tumor Necrosis Factor Receptor Superfamily Members.

BACKGROUND/AIMS: Down syndrome associated disorders are caused by a complex genetic context where trisomy 21 is a central component in relation to other changes involving epigenetic regulators and signaling molecules. This unique genetic context is responsible for the predisposition of people with Down syndrome to acute leukemia. Although, the research in this field has discovered some important pathogenic keys, the exact mechanism of this predisposition is not known. METHODS: In this study we applied functional enrichment analysis to evaluate the interactions between genes localized on chromosome 21, genes already identify as having a key role in acute leukemia of Down syndrome, miRNAs and signaling pathways implicated in cancer and cell development and found that miR-155 has a high impact in genes present on chromosome 21. Forward, we performed next generation sequencing on DNA samples from a cohort of patients diagnosed with acute leukemia of Down syndrome and in vitro functional assay using a CMK-86 cell line, transfected with either mimic or inhibitor of the microRNA-155-5p. RESULTS: Our results show that the epigenetic alteration of the TNF superfamily receptors in Down syndrome, which can be correlated to microRNA-155-5p aberrant activity, may play an important role in cell signaling and thus be linked to acute myeloid leukemia. CONCLUSION: Some genes, already shown to be mutated in AML-DS, are potential targets for miR-155. Our results show that the epigenetic alteration of the TNF superfamily receptors in Down syndrome may play an important role in cell signaling and thus be linked to acute myeloid leukemia.

Baseline characteristics of a nationwide cohort of pediatric patients with acute leukemias of Down syndrome.

PURPOSE: Down syndrome (DS) or trisomy 21, brings together some unique aspects from clinical pediatrics. Among the hematological disorders present in DS, by far the most important is the predisposition for developing acute leukemia. Acute myeloid leukemia (AML) of DS has a preleukemic state with the onset in the neonatal period, rarely symptomatic but with the presence of blasts in peripheral blood smear and apparently a spontaneous remission. The unique tumor profile of DS underlines the importance of chromosome 21 in hematopoiesis and it can help understanding leukemogenesis in general. The purpose of this study was to present the very rare cases with DS and transient leukemia and/or acute leukemia that were found in a nationwide survey of Romania, in three centers of pediatric hematology and oncology. METHODS: A nationwide analysis of the very rare cases of transient leukemia of DS are described, involving the three major pediatric hematology centers of Romania: Cluj Napoca, Bucharest and Timisoara. Data analysis was performed using R 3.5.3. Categorical variables were presented as absolute value (percent). Contingency tables were analyzed using the Fisher test. Normality of the distribution was assessed using the Shapiro test and histogram visualization, but also took into consideration the sample size. Non-normally distributed variables were presented as median (quartile 1, quartile 3). Wilcoxon test was used to determine the differences between two non-normally distributed groups. A p value under 0.05 was considered statistically significant. RESULTS: It appears that the more aggressive entity at presentation is represented by CD45 positive leukemia, which is the more frequent of the myeloid lineage and has lower counts at diagnosis. CONCLUSION: We address this manuscript to pediatricians and neonatologists in order to emphasize the importance of diagnosing hematological disorders in children with DS, especially neonates, even if they are asymptomatic.

Approach to the Adult Acute Lymphoblastic Leukemia Patient.

During recent decades, understanding of the molecular mechanisms of acute lymphoblastic leukemia (ALL) has improved considerably, resulting in better risk stratification of patients and increased survival rates. Age, white blood cell count (WBC), and specific genetic abnormalities are the most important factors that define risk groups for ALL. State-of-the-art diagnosis of ALL requires cytological and cytogenetical analyses, as well as flow cytometry and high-throughput sequencing assays. An important aspect in the diagnostic characterization of patients with ALL is the identification of the Philadelphia (Ph) chromosome, which warrants the addition of tyrosine kinase inhibitors (TKI) to the chemotherapy backbone. Data that support the benefit of hematopoietic stem cell transplantation (HSCT) in high risk patient subsets or in late relapse patients are still questioned and have yet to be determined conclusive. This article presents the newly published data in ALL workup and treatment, putting it into perspective for the attending physician in hematology and oncology.

Transient leukemia of Down syndrome.

Childhood leukemia is mostly a "developmental accident" during fetal hematopoiesis and may require multiple prenatal and postnatal "hits". The World Health Organization defines transient leukemia of Down syndrome (DS) as increased peripheral blood blasts in neonates with DS and classifies this type of leukemia as a separate entity. Although it was shown that DS predisposes children to myeloid leukemia, neither the nature of the predisposition nor the associated genetic lesions have been defined. Acute myeloid leukemia of DS is a unique disease characterized by a long pre-leukemic, myelodysplastic phase, unusual chromosomal findings and a high cure rate. In the present manuscript, we present a comprehensive review of the literature about clinical and biological findings of transient leukemia of DS (TL-DS) and link them with the genetic discoveries in the field. We address the manuscript to the pediatric generalist and especially to the next generation of pediatric hematologists.

Joint hyperlaxity prevents relapses in clubfeet treated by Ponseti method-preliminary results.

PURPOSE: The aim of the study was to evaluate the role of joint hyperlaxity (by Beighton score) as a protective factor for clubfoot relapse. METHODS: Patients with idiopathic clubfoot treated with the Ponseti method between January 2004 and December 2012, without other congenital foot deformity, and not previously treated by open surgery were included in either the Relapse group (n = 23) if it was a clubfoot relapse or the Control group (n = 19) if no relapse was noted. Joint laxity was evaluated using the Beighton score at the latest follow-up against the Normal group (n = 22, children matched by sex and age without clubfoot deformity). RESULTS: We found a significantly higher joint laxity in the Control group (4.58, 95% confidence interval [CI]: 2.1-7.06) as compared to the Relapse (3.17, 95% CI: 1.53-4.81, p = 0.032) and Normal (3.14, 95% CI: 1.78-4.5, p = 0.03) groups. The univariate logistic regression showed a 5.28-times increase in the risk of relapse for a Beighton score lower than 4/9 points (odds ratio = 5.28; 95% CI = 1.29-21.5; p = 0.018). CONCLUSIONS: Joint hyperlaxity could be a protective factor for clubfoot relapse.

Echocardiography as the first diagnostic clue to rapidly progressive systemic AL amyloidosis associated with multiple site thrombosis. A case report.

AL-amyloidosis is a rare, but complex disease, with a severe prognosis, cardiac involvement being found in half of the patients. The rapid increase of the LV wall thickness predicts an unfavorable evolution. We report the case of a 63-year-old man diagnosed with AL-amyloidosis, with cardiac involvement, associated with multiple site thrombosis.  Specific echocardiographic methods like tissue Doppler imaging and speckle tracking provided crucial diagnostic and prognostic information.

Fatal hypersensitivity pneumonitis after chemical occupational exposure.

Hypersensitivity pneumonitis (HP; extrinsic allergic alveolitis) is a rare non-immunoglobulin E (IgE)-mediated inflammatory lung disease caused by inhalation exposure (occupational, recreational or ordinary home exposure). A 36-year-old female patient, without significant medical history, is referred to an outpatient pulmonology clinic for dry cough, shortness of breath, fever, fatigue and weight loss. Chest high-resolution computed tomography (HRCT) was performed, and significant lung fibrosis (especially centrilobular and interlobular in bilateral "thick lines"), traction bronchiectasis and alveolitis in both superior lobes are described. Lung function tests showed severe restrictive dysfunction. Transfer factor of the lung for carbon monoxide (TLCO) being very low, the flexible bronchoscopy was contraindicated. Surgical lung biopsy was performed. Histopathological examination showed characteristic lesions of chronic bilateral hypersensitivity pneumonitis. The patient died four days after the surgical intervention due to post-operative complications. Exposure to various chemical substances can form bonds with human proteins molecules and induce an exaggerated immune response in susceptible individuals. A high index of suspicion of occupational exposure can determine an early diagnosis with a better outcome.

Autoimmune hepatitis with sclerosing cholangitis in a patient with thiopurine methyltransferase deficiency: case presentation.

The association between two autoimmune diseases is known in the literature as overlap syndrome. We present the case of an 18-year-old boy, diagnosed at the age of 13 with an overlap syndrome between type I autoimmune hepatitis and sclerosing cholangitis. The response to immunosuppressant therapy was hampered by azathioprine-induced toxicity causing severe pancytopenia, as a result of thiopurine methyltransferase enzyme genetic deficiency. Treatment was replaced by mycophenolate mofetil. Although the relapse rate was reduced, the disease progressed to cirrhosis. Specific features of this case were the overlap syndrome, young age of onset, especially for sclerosing cholangitis, azathioprine toxicity that influenced the prognosis and the treatment problems regarding the use and efficiency of alternative immunosuppressant agents in pediatric patients.

Myositis ossificans traumatica of the neck - a pediatric case.

Myositis ossificans circumscripta (MOC) is an extra-osseous non- neoplastic growth of a new bone. It occurs most commonly in the second and the third decade of life, while it is rare in children. The etiology of MOC is unknown and the quadriceps and brachials are the most affected. The occurrence of traumatic MOC in tissues of the neck is uncommon. We are presenting below a rare case of traumatic myositis ossificans occurring in sternocleidomastoid and trapezius muscles in a 17-year-old girl.

Immunogenicity and safety of human papillomavirus-16/18 AS04-adjuvanted vaccine administered according to an alternative dosing schedule compared with the standard dosing schedule in healthy women aged 15 to 25 years: results from a randomized study.

BACKGROUND: The human papillomavirus (HPV)-16/18 AS04-adjuvanted vaccine is immunogenic, has a clinically acceptable safety profile, and prevents incident and persistent HPV-16/18 infection and cervical precancerous lesions. This study (NCT00552279) evaluated the vaccine when administered according to an alternative dosing schedule (0-1-12 months) compared with the standard dosing schedule (0-1-6 months). METHODS: The study was of randomized open design and was conducted at multiple centers in Europe. Healthy women aged 15 to 25 years were randomized (1:1) to receive HPV-16/18 vaccine according to the standard schedule at months 0, 1, and 6 (n = 401) or an alternative schedule at months 0, 1, and 12 (n = 403). HPV-16 and -18 antibodies were measured by enzyme-linked immunosorbent assay at months 0, 2, and 7 or 13 (depending on group); noninferiority evaluation was performed sequentially for seroconversion rates and geometric mean antibody titers. Primary analysis of immunogenicity was based on the according-to-protocol cohort. Vaccine safety and reactogenicity were assessed on the total vaccinated cohort. RESULTS: Predefined noninferiority criteria were met 1 month after the third vaccine dose when the HPV-16/18 vaccine was administered according to the 0-1-12 month schedule compared with the 0-1-6 month schedule in terms of seroconversion rates for HPV-16 (100% and 100%) and HPV-18 (99.7% and 100%) and geometric mean antibody titers for HPV-16 (11884.7 and 10311.9 ELISA units/mL) and HPV-18 (4501.3 and 3963.6 ELISA units/mL), respectively. The HPV-16/18 vaccine had a clinically acceptable safety profile when administered according to either schedule. CONCLUSIONS: The third dose of the HPV-16/18 vaccine can be administered any time between 6 and 12 months after the first dose, with adequate immunogenicity and a clinically acceptable safety profile.