IntroductionDendritic cells (DCs) are the most efficient antigen presenting cells, which are considered a central component of the immune system for their extraordinary capacity to initiate and modulate the immune responses elicited upon recognition of infectious agents. This has made them a major focus of interest in the conception of immunotherapeutic vaccine strategies.Aim of the studyTo standardise a protocol for in vitro differentiation of human peripheral blood monocytes into immature DCs (iDCs) upon treatment with specific growth factors and to compare two monocyte isolation methods including magnetic activated cell sorted (MACS) monocytes by CD14+ immuno-magnetic beads and monocytes separated by adherence.Material and methodsImmature DCs were generated from monocytes of human peripheral blood in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-4 after in vitro culture for seven days. Cultured cells were stained with surface markers of iDCs: FITC-anti-CD14, PE-anti-CD11c, PE-anti-CD1a, PE-Cy5-anti-HLA-DR, and PE-anti-CD83 for flow cytometry analysis.ResultsWe found that the viability of MACS-DCs was higher than DCs derived from monocytes separated by adherence (median 50 and interquartile range 45-50 vs. 25 and 10-30, respectively; p < 0.001). Flow cytometry analysis revealed that the median interquartile percentages of MACS-DCs expressing CD14– was significantly higher compared to the DCs derived from monocytes separated by adherence (median 80.2 and interquartile range 77.7-80.7 vs. 40.2 and 30.4-40.6, respectively; p < 0.001). However, MACS-DCs expressed the same levels of CD11c, CD1a, and HLA-DR as well as CD83 compared to the DCs derived from monocytes separated by adherence with p value > 0.05.ConclusionsBoth positively selected monocytes and monocytes separated by adherence procedure gave the same results as regards cell surface marker expression, although the DCs purity and viability using MACS separated monocytes were better.
Background: The clinical experience gathered throughout the years endorses primary immunodeficiency diseases (PIDs) awareness and guides research into newborn screening and future therapeutic strategies.Combined T-cell receptor excision circle levels (TRECs) and kappa-deleting recombination excision circles (KRECs) assay paves the way to new potential applications in this field. Objectives: We aimed to establish a technique for quantification of TRECs and KRECs in Egyptian individuals in our laboratory and to set a lower threshold of normal for TRECs and KRECs in pediatric population for different age groups as a start for its implementation in newborn screening protocols for PIDs. Methods: 50 apparently healthy children (25 males and 25 females) with age ranging from 1 day to 16 years were analyzed..Combined quantification of TRECs and KRECs in the genomic DNA of peripheral blood mononuclear cells was performed using real-time quantitative PCR. Results: Individuals in the study were divided in to 5 different age groups Data regarding lower threshold of normal for TRECs and KRECs copies per ml of blood among the study group was obtained. A highly significant negative correlation between TRECs and KRECs, both calculated per 10 6 PBMCs and per ml of blood and age was observed. On the contrary, there was no statistically significant differences in the studied parameters between males and females when evaluated regardless of age (p value=0.697). Conclusion: It appeared that it is technically feasible to introduce the TRECs/KRECs quantitation by real time PCR into routine laboratory practice to be used in the near future both for new born screening for PIDs
Article informationBackground: These days, inflammatory bowel disease [IBD] is growing more common.Its diagnosis relies on invasive techniques like colonoscopy and biopsy, and its activity is monitored by fecal calprotectin levels that have a low compliance rate, so there is a pressing need for a serum biomarker that is non-invasive, accepted, and accurate for diagnosing and monitoring IBD activity. The aim of the work: The goal of our research is to study serum calprotectin as a candidate biomarker in IBD. Methods: The study included 50 patients with IBD who were recruited from Ain Shams University Hospitals' Gastroenterology clinic. Sixty percent were diagnosed with ulcerative colitis, with half in activity and the other half in remission, and 40% were diagnosed with Chron's disease, with half in activity and the other half in remission.A control group of 20 apparently healthy individuals comparable in age and sex were also included in the study. All subjects had their serum calprotectin tested by ELISA in addition to their ESR and CRP measurements. Results: Serum calprotectin levels were significantly higher in patients with IBD than in controls and in clinically active patients than those in remission in both UC and CD groups. Although there was a positive association between serum calprotectin levels and CRP and ESR, serum calprotectin had a higher diagnostic value than CRP and ESR due to its higher sensitivity and specificity. Our findings demonstrated that serum calprotectin and platelet count had a direct relationship, while serum calprotectin and serum albumin and hemoglobin levels had an inverse relationship. Conclusion: Serum calprotectin levels are raised and linked to clinical activity in IBD patients, implying that it could be utilised as a clinically useful indicator of disease activity.
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