B cell-derived circulating granzyme B is a feature of acute infectious mononucleosis.

Granzyme B (GzmB) is a serine protease best known for inducing target cell apoptosis when released by cytotoxic T lymphocytes (CTLs) or natural killer cells with pore-forming perforin. As a result, GzmB detected in the serum of virus-infected individuals has typically been attributed to these sources. Here, we show that patients with recently diagnosed infectious mononucleosis caused by Epstein-Barr virus (EBV) have high circulating levels of GzmB that may be derived from infected B cells early in course of disease.

A colorimetric assay that specifically measures Granzyme B proteolytic activity: hydrolysis of Boc-Ala-Ala-Asp-S-Bzl.

The serine protease Granzyme B (GzmB) mediates target cell apoptosis when released by cytotoxic T lymphocytes (CTL) or natural killer (NK) cells. GzmB is the most studied granzyme in humans and mice and therefore, researchers need specific and reliable tools to study its function and role in pathophysiology. This especially necessitates assays that do not recognize proteases such as caspases or other granzymes that are structurally or functionally related.

A method for detecting intracellular perforin in mouse lymphocytes.

Cytotoxic lymphocytes destroy pathogen-infected and transformed cells through the cytotoxic granule exocytosis death pathway, which is dependent on the delivery of proapoptotic granzymes into the target cell cytosol by the pore-forming protein, perforin. Despite the importance of mouse models in understanding the role of cytotoxic lymphocytes in immune-mediated disease and their role in cancer immune surveillance, no reliable intracellular detection method exists for mouse perforin. Consequently, rapid, flow-based assessment of cytotoxic potential has been problematic, and complex assays of function are generally required.

B-CLL cells acquire APC- and CTL-like phenotypic characteristics after stimulation with CpG ODN and IL-21.

CpG oligodeoxynucleotides (CpG) and IL-21 are two promising agents for the treatment of B-cell chronic lymphocytic leukemia (B-CLL). Recently, we reported that the combination of CpG and IL-21 (CpG/IL-21) can induce granzyme B (GrB)-dependent apoptosis in B-CLL cells. Here, we demonstrate that treatment of B-CLL cells with CpG and IL-21 results in the development of antigen-presenting cell (APC)-like cells with cytotoxic features.

Interleukin 21-induced granzyme B-expressing B cells infiltrate tumors and regulate T cells.

The pathogenic impact of tumor-infiltrating B cells is unresolved at present, however, some studies suggest that they may have immune regulatory potential. Here, we report that the microenvironment of various solid tumors includes B cells that express granzyme B (GrB, GZMB), where these B cells can be found adjacent to interleukin (IL)-21-secreting regulatory T cells (Treg) that contribute to immune tolerance of tumor antigens. Because Tregs and plasmacytoid dendritic cells are known to modulate T-effector cells by a GrB-dependent mechanism, we hypothesized that a similar process may operate to modulate regulatory B cells (Breg).

Why do human B cells secrete granzyme B? Insights into a novel B-cell differentiation pathway.

B cells are generally believed to operate as producers of high affinity antibodies to defend the body against microorganisms, whereas cellular cytotoxicity is considered as an exclusive prerogative of natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). In conflict with this dogma, recent studies have demonstrated that the combination of interleukin-21 (IL-21) and B-cell receptor (BCR) stimulation enables B cells to produce and secrete the active form of the cytotoxic serine protease granzyme B (GrB). Although the production of GrB by B cells is not accompanied by that of perforin as in the case of many other GrB-secreting cells, recent findings suggest GrB secretion by B cells may play a significant role in early antiviral immune responses, in the regulation of autoimmune responses, and in cancer immunosurveillance.

Activated mouse B cells lack expression of granzyme B.

Recently, it has been reported that human B cells express and secrete the cytotoxic protease granzyme B (GrB) after stimulation with IL-21 and BCR cross-linking. To date, there are few clues on the function of GrB in B cell biology. As experimental transgenic murine systems should provide insights into these issues, we assayed for GrB in C57BL/6 B cells using an extensive array of physiologically relevant stimuli but were unable to detect either GrB expression or its proteolytic activity, even when Ag-specific transgenic BCRs were engaged.

Human B cells differentiate into granzyme B-secreting cytotoxic B lymphocytes upon incomplete T-cell help.

Recently, CD4(+) T helper cells were shown to induce differentiation of human B cells into plasma cells by expressing interleukin (IL-)21 and CD40 ligand (CD40L). In the present study we show, that in the absence of CD40L, CD4(+) T cell-derived IL-21 induces differentiation of B cells into granzyme B (GzmB)-secreting cytotoxic cells. Using fluorescence-activated cell sorting (FACS) analysis, ELISpot and confocal microscopy, we demonstrate that CD4(+) T cells, activated via their T-cell receptor without co-stimulation, can produce IL-21, but do not express CD40L and rapidly induce GzmB in co-cultured B cells in an IL-21 receptor-dependent manner.

CD5+ B cells from individuals with systemic lupus erythematosus express granzyme B.

Recently, we reported that IL-21 induces granzyme B (GzmB) and GzmB-dependent apoptosis in malignant CD5(+) B cells from patients with chronic lymphocytic leukemia. Several autoimmune diseases (AD) are associated with enhanced frequencies of CD5(+) B cells. Since AD are also associated with elevated IL-21 and GzmB levels, we postulated a link between CD5(+) B cells, IL-21 and GzmB.

Human B cells secrete granzyme B when recognizing viral antigens in the context of the acute phase cytokine IL-21.

Human B cells are currently not known to produce the proapoptotic protease granzyme B (GrB) in physiological settings. We have discovered that BCR stimulation with either viral Ags or activating Abs in the context of the acute phase cytokine IL-21 can induce the secretion of substantial amounts of GrB by human B cells. Importantly, GrB response to viral Ags was significantly stronger in B cells from subjects recently vaccinated against the corresponding viruses as compared with unvaccinated subjects.