ABSTRACT
Human B-cell differentiation has been extensively investigated on genomic and transcriptomic grounds; however, no studies have accomplished so far detailed analysis of antigen-dependent maturation-associated human B-cell populations from a proteomic perspective. Here, we investigate for the first time the quantitative proteomic profiles of B-cells undergoing antigen-dependent maturation using a label-free LC-MS/MS approach applied on 5 purified B-cell subpopulations (naive, centroblasts, centrocytes, memory and plasma B-cells) from human tonsils (data are available via ProteomeXchange with identifier PXD006191). Our results revealed that the actual differences among these B-cell subpopulations are a combination of expression of a few maturation stage-specific proteins within each B-cell subset and maturation-associated changes in relative protein expression levels, which are related with metabolic regulation. The considerable overlap of the proteome of the 5 studied B-cell subsets strengthens the key role of the regulation of the stoichiometry of molecules associated with metabolic regulation and programming, among other signaling cascades (such as antigen recognition and presentation and cell survival) crucial for the transition between each B-cell maturation stage.
Subject(s)
Antigens/immunology , B-Lymphocyte Subsets/cytology , Cell Differentiation/immunology , Gene Expression Regulation/immunology , Signal Transduction/immunology , Adolescent , Adult , Cells, Cultured , Child , Child, Preschool , Female , Gene Expression Profiling , Gene Expression Regulation/genetics , Germinal Center/cytology , Germinal Center/immunology , Humans , Lymphocyte Activation/immunology , Male , Palatine Tonsil/cytology , Palatine Tonsil/immunology , Proteome/genetics , Transcriptome/genetics , Young AdultABSTRACT
The clinical value of assessing immunoglobulin (Ig)G and IgA subclasses in addition to the isotypes of soluble Igs in serum has been well established. >20years ago, the International Union of Immunological Societies and the World Health Organization performed collaborative studies in order to validate antibody (Ab) clones for the detection of IgG and IgA subclasses for a broad range of laboratory assays, except for flow cytometry. Here we analyzed the performance of commercially available Ab clones to detect IgG and IgA subclasses in memory B-cells and plasma cells (PCs) by flow cytometry. In a first step, 28 Ab clones were evaluated in peripheral blood from healthy donors. Only 17/28 clones showed reactivity against IgG and IgA subclasses expressed on the B-cell and PC surface membrane, including Ab clones for IgG1 (SAG1, HP6188, HP6001 and HP6186), IgG2 (SAG2, HP6014 and HP6002), IgG3 (SAG3, HP6095 and HP6050), IgG4 (SAG4), IgA1 (SAA1, H69-11.4 and B3506B4) and IgA2 (SAA2, 2E2, and A9604D2). In a second step, for each Ig subclass a single clone was selected according to its specificity and fluorescence intensity (resolution power), for further more detailed validation (SAG1, SAG2, SAG3, SAG4, SAA1 and SAA2). This validation process was carried out in 4 different laboratories by testing the selected Ab clones in human peripheral blood, bone marrow and tonsil samples, using different staining protocols (e.g. surface membrane and/or cytoplasmic staining). All selected Ab clones displayed strong positivity, high specificity and optimal resolution between negative and positive cells. Alternative Ab clones were also validated. Thus, our results show the feasibility of using the validated Ig subclass Ab clones in combination with other B cell-associated markers for detailed dissection of the memory B-cell and PC compartments that express distinct Ig subclasses in different human tissues.
Subject(s)
Antibodies, Monoclonal , Antibody Specificity , Flow Cytometry/methods , Immunoglobulin A/analysis , Immunoglobulin G/analysis , Immunophenotyping/methods , Adult , Aged , B-Lymphocytes/immunology , Child, Preschool , Female , Flow Cytometry/standards , Humans , Male , Middle Aged , Plasma Cells/immunologyABSTRACT
Today's imaging studies accurately delineate the extent of glomus tumors of the temporal bone. Microsurgical techniques allow total removal of even the largest tumors with acceptable morbidity. While surgical management is the principal treatment of glomus tumors, stereotactic fractionated radiation therapy may be used as an alternative treatment when there is a risk of cranial neuropathy with surgical management. We should be aware of the full range of management options and base treatment choice on the age and medical condition of the patient and location and size of the tumor.
Subject(s)
Glomus Jugulare , Glomus Tympanicum , Head and Neck Neoplasms/surgery , Paraganglioma/surgery , Skull Neoplasms/surgery , Temporal Bone , Head and Neck Neoplasms/diagnosis , Humans , Paraganglioma/diagnosis , Preoperative Care , Skull Neoplasms/diagnosis , Surgical Procedures, Operative/adverse effects , Surgical Procedures, Operative/methodsABSTRACT
Actualmente, los estudios por imagen permiten conocer con precisión la extensión de los tumores glómicos del hueso temporal. Las técnicas de microcirugía permiten una extirpación completa de grandes tumores, con una morbilidad aceptable. Si bien estamos a favor del abordaje quirúrgico como tratamiento principal en los tumores glómicos, el tratamiento con radiación estereotáxica focalizada puede emplearse como tratamiento alternativo, cuando el riesgo de neuropatía craneal con cirugía sea alto. Debemos conocer todas las opciones terapéuticas y realizar nuestra elección sobre la base de la edad y el estado de salud del paciente, así como de la localización y el tamaño del tumor (AU)
Todays imaging studies accurately delineate the extent of glomus tumors of the temporal bone. Microsurgical techniques allow total removal of even the largest tumors with acceptable morbidity. While surgical management is the principal treatment of glomus tumors, stereotactic fraction atedradiation therapy may be used as an alternative treatment when there is a risk of cranial neuropathy with surgical management. We should be aware of the full range of management options and base treatment choice on the age and medical condition of the patient and location and size of the tumor (AU)
