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CURATOR
A pinboard by
Fane Mensah

PhD Student, University College London

PINBOARD SUMMARY

I have developed different assays to measure the metabolic activity of B cells throughout maturation

B cells or B lymphocytes are a type of white blood cell (immune cell) that plays a very important role in our humoral immunity. They are most famously known for the production of different types of antibodies that combat pathogens and or infected cells, but they are also vital in assisting other immune cells. During my PhD project, I have developed different assays to measure the metabolic activity of different B cell subsets. This is very important to understand how different energy pathways link up with different B cell functions. Interestingly, B cell dysfunction has been suggested in a condition known as Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (ME/CFS). ME/CFS is a heterogeneous disease of unknown aetiology without an established diagnostic test. The condition is characterised by fatigue, cognitive impairment and post-exertional malaise after mental or physical exertion lasting for at least 6 months. Due to successive trials of B-cell depletion (removing B cells in the periphery, and resetting B cell formation) from ME/CFS patients with rituximab, it is proposed that B-cell function and possibly repertoire is altered in these patients. In my first peer-reviewed article (published in clinical and experimental Immunology 2016), I compared extended B-cell phenotypes between healthy controls and ME/CFS patients. This was mainly based on flow cytometry work, a technique often used in immunology using fluorescent-labeled antibodies to detect proteins or ligands that bind to specific molecules (e.g. surface CD markers or intracellular proteins), such as CD19, CD27 and IgD (markers used to delineate B-cell subsets). In these studies, I have identified an increase in frequency and expression of a B cell marker that plays an important role in maturation. This project was the base for functional studies that I have and will perform throughout my PhD to understand B-cell metabolism through maturation and the possible role these immune cells might play in ME/CFS. I have managed to set up national and international collaboration with leading scientists in the field. I have also demonstrated my ambition, scientific capacity and resourcefulness by successfully writing and obtaining research grants. So far, my work has led to 15 invitations at conferences and/or meetings (of which 10 oral presentations).

4 ITEMS PINNED

Metabolic reprogramming is required for antibody production that is suppressed in anergic but exaggerated in chronically BAFF-exposed B cells.

Abstract: B cell activation leads to proliferation and Ab production that can protect from pathogens or promote autoimmunity. Regulation of cell metabolism is essential to support the demands of lymphocyte growth and effector function and may regulate tolerance. In this study, we tested the regulation and role of glucose uptake and metabolism in the proliferation and Ab production of control, anergic, and autoimmune-prone B cells. Control B cells had a balanced increase in lactate production and oxygen consumption following activation, with proportionally increased glucose transporter Glut1 expression and mitochondrial mass upon either LPS or BCR stimulation. This contrasted with metabolic reprogramming of T cells, which had lower glycolytic flux when resting but disproportionately increased this pathway upon activation. Importantly, tolerance greatly affected B cell metabolic reprogramming. Anergic B cells remained metabolically quiescent, with only a modest increase in glycolysis and oxygen consumption with LPS stimulation. B cells chronically stimulated with elevated BAFF, however, rapidly increased glycolysis and Ab production upon stimulation. Induction of glycolysis was critical for Ab production, as glycolytic inhibition with the pyruvate dehydrogenase kinase inhibitor dichloroacetate sharply suppressed B cell proliferation and Ab secretion in vitro and in vivo. Furthermore, B cell-specific deletion of Glut1 led to reduced B cell numbers and impaired Ab production in vivo. Together, these data show that activated B cells require Glut1-dependent metabolic reprogramming to support proliferation and Ab production that is distinct from T cells and that this glycolytic reprogramming is regulated in tolerance.

Pub.: 13 Mar '14, Pinned: 25 Oct '17

Extended B cell phenotype in patients with myalgic encephalomyelitis/chronic fatigue syndrome: a cross‐sectional study

Abstract: Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a heterogeneous condition of unknown aetiology characterized by multiple symptoms including fatigue, post‐exertional malaise and cognitive impairment, lasting for at least 6 months. Recently, two clinical trials of B cell depletion therapy with rituximab (anti‐CD20) reported convincing improvement in symptoms. A possible but undefined role for B cells has therefore been proposed. Studies of the relative percentages of B cell subsets in patients with ME/CFS have not revealed any reproducible differences from healthy controls (HC). In order to explore whether more subtle alterations in B cell subsets related to B cell differentiation exist in ME/CFS patients we used flow cytometry to immunophenotype CD19+ B cells. The panel utilized immunoglobulin (Ig)D, CD27 and CD38 (classical B cell subsets) together with additional markers. A total of 38 patients fulfilling Canadian, Centre for Disease Control and Fukuda ME/CFS criteria and 32 age‐ and sex‐matched HC were included. We found no difference in percentages of classical subsets between ME/CFS patients and HC. However, we observed an increase in frequency (P < 0·01) and expression (MFI; P = 0·03) of CD24 on total B cells, confined to IgD+ subsets. Within memory subsets, a higher frequency of CD21+CD38– B cells (>20%) was associated with the presence of ME/CFS [odds ratio: 3·47 (1·15–10·46); P = 0·03] compared with HC, and there was a negative correlation with disease duration. In conclusion, we identified possible changes in B cell phenotype in patients with ME/CFS. These may reflect altered B cell function and, if confirmed in other patient cohorts, could provide a platform for studies based on clinical course or responsiveness to rituximab therapy.

Pub.: 22 Feb '16, Pinned: 25 Oct '17

Germinal Center B-Cell-Associated Nuclear Protein (GANP) Involved in RNA Metabolism for B Cell Maturation.

Abstract: Germinal center B-cell-associated nuclear protein (GANP) is upregulated in germinal center B cells against T-cell-dependent antigens in mice and humans. In mice, GANP depletion in B cells impairs antibody affinity maturation. Conversely, its transgenic overexpression augments the generation of high-affinity antigen-specific B cells. GANP associates with AID in the cytoplasm, shepherds AID into the nucleus, and augments its access to the rearranged immunoglobulin (Ig) variable (V) region of the genome in B cells, thereby precipitating the somatic hypermutation of V region genes. GANP is also upregulated in human CD4(+) T cells and is associated with APOBEC3G (A3G). GANP interacts with A3G and escorts it to the virion cores to potentiate its antiretroviral activity by inactivating HIV-1 genomic cDNA. Thus, GANP is characterized as a cofactor associated with AID/APOBEC cytidine deaminase family molecules in generating diversity of the IgV region of the genome and genetic alterations of exogenously introduced viral targets. GANP, encoded by human chromosome 21, as well as its mouse equivalent on chromosome 10, contains a region homologous to Saccharomyces Sac3 that was characterized as a component of the transcription/export 2 (TREX-2) complex and was predicted to be involved in RNA export and metabolism in mammalian cells. The metabolism of RNA during its maturation, from the transcription site at the chromosome within the nucleus to the cytoplasmic translation apparatus, needs to be elaborated with regard to acquired and innate immunity. In this review, we summarize the current knowledge on GANP as a component of TREX-2 in mammalian cells.

Pub.: 29 May '16, Pinned: 25 Oct '17