Role Of BAFF Research Articles

CDNA cloning, expression and bioactivity of porcine BAFF

B cell activating factor belonging to the tumor necrosis factor (TNF) family (BAFF) is critical for B cell survival, maturation and T cell activation by acting through its three receptors, BAFF-R, BCMA and TACI. In the present study, a porcine BAFF cDNA, designated pBAFF, was cloned by RT–PCR and rapid amplification of cDNA ends (RACE) strategies. The full-length cDNA of pBAFF consists of 805 bp with a 702 bp open reading frame, encoding 233 amino acids. The deduced amino acid sequence contains a predicted transmembrane domain and a putative furin protease cleavage site corresponding to other identified BAFF homologues. The amino acid similarity between the functional soluble parts of pBAFF and human BAFF (hBAFF) or chicken BAFF (cBAFF) is 93% and 85%, respectively, with identity at the amino acid level was 88% and 76%, respectively. The characteristic of the three-cysteine residues of BAFF is conserved in pBAFF. RT-PCR showed that BAFF is expressed in many tissues in the pig, including spleen, liver, lung, heart, intestine, kidney, thymus and PBLs. Recombinant soluble pBAFF (psBAFF) fused with His 6 tag was efficiently expressed in Escherichia coli BL21 (DE3) and its expression was confirmed by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and Western blotting. In vitro, purified psBAFF co-stimulates the proliferation of not only porcine B cells but also human B cells. In addition, hsBAFF binds to porcine B cells and has a positive effect on their proliferation. These findings indicate pBAFF plays an important role in proliferation of porcine B cells and functional cross-reactivity occurs between porcine and human BAFF. In vitro expression of bioactive psBAFF provides the basis for further investigation of its potential to be used as an immunoadjuvant for enhancing vaccine efficacy and an immunotherapeutic in pig. It also provides the basis for investigations on the role of BAFF in this important domestic species and an animal model for human diseases.

Role of BAFF in Adhesion and Growth of Human Multiple Myeloma Cells in the Bone Marrow Microenvironment.

Recent studies have underscored the role of B-cell activating factor (BAFF), a member of the tumor necrosis factor superfamily, in promoting the survival of malignant B cells, including human MM. Since the tumor bone marrow microenvironment plays a crucial role in MM cell growth and survival, we here characterize the functional significance of BAFF in the interaction between MM and bone marrow stromal cells (BMSCs), and further defined molecular mechanisms regulating these processes. We first confirmed the expression of BAFF and its receptors (BCMA, TACI, BAFF-R) on majority of MM cell lines and CD138-purified patient MM cells. Significantly, gene expression profiling revealed increased BCMA expression on newly diagnosed and relapsed MM versus normal plasma cells (p<0.0001, student T test, Abstract # 552872). Although BAFF and its 3 receptors are expressed on CD138+patient MM cells (n=10) by RT-PCR analysis, the pattern of expression of TACI and BAFF-R receptors was heterogeneous, assessed by flow cytometric analysis. We next examined BAFF expression in BMSC lines (n=4) and BMSCs derived from patient BM (n=5). Baff expression is easily detected in BMSCs, and Baff levels are approximately 3–10-fold higher in supernatants of BMSCs than equivalent numbers of MM cells. Thus BMSCs are the main source of BAFF in MM patients. We then asked whether MM adhesion to BMSCs further upregulates BAFF secretion from BMSCs. Adhesion of 5 MM cell lines to BMSCs augments BAFF secretion by 2–5 fold, using BAFF ELISA. Immunoblotting using anti-BAFF Ab confirmed markedly higher expression of BAFF in BMSCs than MCCAR MM cells, as well as greater BAFF upregulation induced by MM adhesion. Since NF-kappaB (NF-κB) is crucial for MM adhesion-induced cytokine secretion from BMSCs and the BAFF gene promoter contains at least six NF-κB-binding sites, we next transfected KM104 BMSC line with a luciferase reporter vector carrying the BAFF gene promoter (BAFF-LUC) and then allowed MM cells to adhere to BMSCs for 24 hr, followed by measurement of luciferase activity. NF-κB-LUC reporter was used as positive and pGL3 plasmid as a negative control. Adhesion of MCCAR and MM1S MM lines to KM103 BMSC line with BAFF-LUC and NF-κB-LUC, but not control reporters, significantly enhanced luciferase activity, suggesting that NF-κB activation induced by MM adhesion to BMSCs mediates BAFF upregulation. In parallel, we also asked whether BAFF induces MM adhesion to BMSCs. BAFF (0–100 ng/ml) increases adhesion of 5 MM lines to BMSCs in a dose-dependent manner; conversely, TACI-Ig or blocking anti-BCMA Ab inhibited BAFF stimulation, indicating increased adhesion specific triggered by BAFF. Using adenoviruses expressing dominant-negative and constitutively expressed AKT as well as IkappaB kinase (IKK) inhibitor (PS-1145), we further showed that BAFF-induced MM cell adhesion is primarily mediated via activation of AKT and NF-κB signaling. Finally, BAFF significantly increased adhesion of CD138-expressing patient MM cells to BMSCs. These studies suggest a role for BAFF in localization and growth of MM cells in the BM microenvironment and strongly support novel important therapeutics targeting the interaction between BAFF and its receptors in human MM.

The role of BAFF in immune function and implications for autoimmunity

BAFF [B-cell activating factor belonging to the tumor necrosis factor (TNF) family] is a ligand that is required for peripheral B-cell survival and homeostasis. In addition to mediating B-cell survival, BAFF also regulates expression of certain B-cell-surface proteins, such as CD21/35. BAFF deficiency results in a reduced number of peripheral B cells and a diminished ability to mount robust humoral immune responses. Overexpression of BAFF has been linked to murine and human autoimmunity, and recent data provide clues as to how excess BAFF may allow the emergence of autoreactive B cells. In vivo animal testing with BAFF inhibitors has generated exciting data that support the pathway as a target for modulating B cells. The role of BAFF in B-cell biology, T-cell biology, and autoimmunity is discussed, as well as current efforts to develop BAFF inhibitors for clinical testing in autoimmune disorders.

The role of BAFF in autoimmune diseases

B cell activating factor belonging to the tumor necrosis factor family (BAFF) is a tumor necrosis factor (TNF) superfamily member best known for its role in the survival and maturation of B cells. BAFF is a ligand for three TNF receptor superfamily members: B-cell maturation antigen (BCMA), transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), and BAFF receptor (BAFF-R). Among them, BAFF-R plays the central role in the BAFF system, whereas TACI plays the inhibitory role. BAFF/BAFF receptors appear to span nearly all stages of B-lineage differentiation, ranging from the development, selection, and homeostasis of naive primary B cells to the maintenance of long-lived bone marrow plasma cells. Furthermore, excessive BAFF rescues self-reactive B cells from anergy, which may play a crucial role in the induction and development of autoimmunity. Mice overexpressing BAFF exhibit increased B cell numbers in spleen and lymph node and autoimmune phenotype similar to patients with systemic lupus erythematosus (SLE) and Sjögren's syndrome. Furthermore, inhibition of BAFF by TACI-Ig and BAFFR-Ig has been successful in treating murine models of SLE and rheumatoid arthritis. In humans, previous reports have shown elevated serum BAFF levels in SLE, rheumatoid arthritis, Sjögren's syndrome, and systemic sclerosis patients. Thus, the dysregulation of BAFF/BAFF receptor system may contribute to induction and development of autoimmune diseases and become one of important therapeutic targets.

Excess BAFF Rescues Self-Reactive B Cells from Peripheral Deletion and Allows Them to Enter Forbidden Follicular and Marginal Zone Niches

The role of BAFF in B cell self tolerance was examined by tracking the fate of anti-HEL self-reactive B cells in BAFF transgenic mice using four different models of self-reactive B cell deletion. BAFF overexpression did not affect the development of self-reactive B cells normally deleted in the bone marrow or during the early stages of peripheral development. By contrast, self-reactive B cells normally deleted around the late T2 stage of peripheral development were rescued from deletion, matured, and colonized the splenic follicle. Furthermore, self-reactive B cells normally selectively deleted from the marginal zone repopulated this compartment when excess BAFF was present. Self-reactive B cells rescued by excess BAFF were not anergic. BAFF overexpression therefore rescued only self-reactive B cells normally deleted with relatively low stringency and facilitated their migration into otherwise forbidden microenvironments. This partial subversion of B cell self tolerance is likely to underlie the autoimmunity associated with BAFF overexpression.

BAFFling No More

A number of tumor necrosis factor (TNF)-related ligands and receptors have recently been identified that regulate B cell development and function. One of these is the B cell-activating factor, BAFF, which interacts with two receptors [transmembrane activator and calcium-modulating cyclophilin ligand interacting protein (TACI) and B cell maturation antigen (BCMA)] to promote survival and proliferation of splenic B cells (see the Perspective by Waldschmidt and Noelle ). Scheimann et al. observed a profound defect in B cell development in mutant mice that lack expression of BAFF that was only partially reiterated in TACI- and BCMA-deficient animals. These results implicate the existence of another receptor for BAFF. Thompson et al. describe a candidate receptor that binds BAFF but not any other of the identified TNF-like ligands. The A/WySnJ strain of mice harbors a truncation in the BAFF-R gene that leads to a B cell deficit similar to that observed in BAFF-deficient mice. Together, these studies suggest that the interaction of BAFF with BAFF-R mediates major effects on B cell development. T. J. Waldschmidt, R. J. Noelle, Long live the mature B cell--a BAFFling mystery resolved. Science 293 , 2012-2013 (2001). [Full Text] B. Schiemann, J. L. Gommerman, K. Vora, T. G. Cachero, S. Shulga-Morskaya, M. Dobles, E. Frew, M. L. Scott, An essential role for BAFF in the normal development of B cells through a BCMA-independent pathway. Science 293 , 2111-2114 (2001). [Abstract] [Full Text] J. S. Thompson, S. A. Bixler, F. Qian, K. Vora, M. L. Scott, T. G. Cachero, C. Hession, P. Schneider, I. D. Sizing, C. Mullen, K. Strauch, M. Zafari, C. D. Benjamin, J. Tschopp, J. L. Browning, C. Ambrose, BAFF-R, a newly identified TNF receptor that specifically interacts with BAFF. Science 293 , 2108-2111 (2001). [Abstract] [Full Text]

BAFF mediates survival of peripheral immature B lymphocytes.

B cell maturation is a very selective process that requires finely tuned differentiation and survival signals. B cell activation factor from the TNF family (BAFF) is a TNF family member that binds to B cells and potentiates B cell receptor (BCR)-mediated proliferation. A role for BAFF in B cell survival was suggested by the observation of reduced peripheral B cell numbers in mice treated with reagents blocking BAFF, and high Bcl-2 levels detected in B cells from BAFF transgenic (Tg) mice. We tested in vitro the survival effect of BAFF on lymphocytes derived from primary and secondary lymphoid organs. BAFF induced survival of a subset of splenic immature B cells, referred to as transitional type 2 (T2) B cells. BAFF treatment allowed T2 B cells to survive and differentiate into mature B cells in response to signals through the BCR. The T2 and the marginal zone (MZ) B cell compartments were particularly enlarged in BAFF Tg mice. Immature transitional B cells are targets for negative selection, a feature thought to promote self-tolerance. These findings support a model in which excessive BAFF-mediated survival of peripheral immature B cells contributes to the emergence and maturation of autoreactive B cells, skewed towards the MZ compartment. This work provides new clues on mechanisms regulating B cell maturation and tolerance.

BAFF binds to the tumor necrosis factor receptor-like molecule B cell maturation antigen and is important for maintaining the peripheral B cell population.

The tumor necrosis factor (TNF) family member B cell activating factor (BAFF) binds B cells and enhances B cell receptor-triggered proliferation. We find that B cell maturation antigen (BCMA), a predicted member of the TNF receptor family expressed primarily in mature B cells, is a receptor for BAFF. Although BCMA was previously localized to the Golgi apparatus, BCMA was found to be expressed on the surface of transfected cells and tonsillar B cells. A soluble form of BCMA, which inhibited the binding of BAFF to a B cell line, induced a dramatic decrease in the number of peripheral B cells when administered in vivo. Moreover, culturing splenic cells in the presence of BAFF increased survival of a percentage of the B cells. These results are consistent with a role for BAFF in maintaining homeostasis of the B cell population.