Age-Associated B Cell Features of the Murine High-Grade B Cell Lymphoma Bc.DLFL1 and Its Extranodal Expansion in Abdominal Adipose Tissues.

Diffuse large B cell lymphoma comprises a heterogeneous group of B cell-derived tumors, with different degrees of aggressiveness, as defined by their cellular origin and tissue microenvironment. Using the spontaneous Bc.DLFL1 lymphoma originating from a BALB/c mouse as a diffuse large B cell lymphoma model, in this study we demonstrate that the lymphoma cells display surface phenotype, IgH V-region somatic mutations, transcription factor characteristics and in vivo location to splenic extrafollicular regions of age-associated B cells (ABCs), corresponding to T-bet+ and Blimp-1+/CD138- plasmablasts derivation. The expansion of lymphoma cells within lymphoid tissues took place in a close arrangement with CD11c+ dendritic cells, whereas the extranodal infiltration occurred selectively in the mesentery and omentum containing resident gp38/podoplanin+ fibroblastic reticular cells. Antagonizing BAFF-R activity by mBR3-Fc soluble receptor fusion protein led to a significant delay of disease progression. The extranodal expansion of Bc.DLFL1 lymphoma within the omental and mesenteric adipose tissues was coupled with a significant change of the tissue cytokine landscape, including both shared alterations and tissue-specific variations. Our findings indicate that while Bc.DLFL1 cells of ABC origin retain the positioning pattern within lymphoid tissues of their physiological counterpart, they also expand in non-lymphoid tissues in a BAFF-dependent manner, where they may alter the adipose tissue microenvironment to support their extranodal growth.

Splenectomy modulates the immune response but does not prevent joint inflammation in a mouse model of RA.

The spleen is the largest secondary lymphoid organ which is involved in the development of B cells and also in systemic (auto)immune responses. Using the recombinant human G1 domain-induced arthritis (GIA) model in splenectomized and control BALB/c mice, we investigated the role of the spleen in the induction and pathogenesis of autoimmune arthritis. Splenectomized mice developed GIA with a similar clinical picture to the control group. However, we observed significant alterations in the humoral and cellular immune responses in splenectomized mice. In the sera of the splenectomized mice, we found lower pro-inflammatory cytokine and anti-rhG1 IgM levels, but higher IL-4, anti-rhG1 IgG1 and anti-CCP and RF antibodies. The arthritis induction in the splenectomized group was associated with a significant expansion of activated helper T cells and an increase in the proportion of the circulating B1 and marginal zone B cell subsets. Importantly, immunization of the splenectomized mice with rhG1 induced the formation of germinal centers in the inguinal- and mesenteric lymph nodes (i/mLNs) which showed an active immune response to rhG1. Finally, both B and T cells from the mLNs of the splenectomized mice showed decreased intracellular Ca2+ signaling than those of the control group. Collectively, these findings indicate that the presence of the spleen is not critical for the induction of GIA, and in its absence the autoimmune arthritis is most likely promoted through the compensatory activity of the i/mLNs. However, our data implies the immunological role of the spleen in arthritis which could be further assessed in human RA.

Foliate Lymphoid Aggregates as Novel Forms of Serous Lymphocyte Entry Sites of Peritoneal B Cells and High-Grade B Cell Lymphomas.

The cellular homeostasis of lymphoid tissues is determined by the continuous interactions of mobile hematopoietic cells within specialized microenvironments created by sessile stromal cells. In contrast to the lymph nodes and mucosal lymphoid tissues with well-defined entry and exit routes, the movement of leukocytes in the peritoneal cavity is largely unknown. In this study, we report that, in addition to the omental milky spots and fat-associated lymphoid clusters, in mice, the serous surface of the mesenteric adipose streaks contains lymphocyte-rich organoids comprised of a highly compacted leaf-like part connected to the adipose tissue that can also efficiently bind B cells and high-grade B cell lymphoma (diffuse large B cell lymphoma) cells. Denoted as foliate lymphoid aggregates (FLAgs), these structures show incomplete T/B segregation and a partially differentiated stromal architecture. LYVE-1-positive macrophages covering FLAgs efficiently bind i.p. injected normal B cells as well as different types of diffuse large B cell lymphoma cells. Within FLAgs, the lymphocytes compartmentalize according to their chemokine receptor pattern and subsequently migrate toward the mesenteric lymph nodes via the mesenteric lymphatic capillaries. The blood supply of FLAgs includes short vascular segments displaying peripheral lymph node addressin, and the extravasation of lymphocytes to the omental and mesenteric adipose tissues is partly mediated by L-selectin. The appearance of i.p. injected cells in mesenteric lymph nodes suggests that the mesentery-associated lymphatics may also collect leukocytes from the fat-associated lymphoid clusters and FLAgs, thus combining the mucosal and serous exit of mobile leukocytes and increasing the range of drainage sites for the peritoneal expansion of lymphoid malignancies.

Intraperitoneal Glucose Transport to Micrometastasis: A Multimodal In Vivo Imaging Investigation in a Mouse Lymphoma Model.

Bc-DLFL.1 is a novel spontaneous, high-grade transplantable mouse B-cell lymphoma model for selective serosal propagation. These cells attach to the omentum and mesentery and show dissemination in mesenteric lymph nodes. We aimed to investigate its early stage spread at one day post-intraperitoneal inoculation of lymphoma cells (n = 18 mice), and its advanced stage at seven days post-inoculation with in vivo [18F]FDG-PET and [18F]PET/MRI, and ex vivo by autoradiography and Cherenkov luminescence imaging (CLI). Of the early stage group, nine animals received intraperitoneal injections, and nine received intravenous [18F]FDG injections. The advanced stage group (n = 3) received intravenous FDG injections. In the early stage, using autoradiography we observed a marked accumulation in the mesentery after intraperitoneal FDG injection. Using other imaging methods and autoradiography, following the intravenous injection of FDG no accumulations were detected. At the advanced stage, tracer accumulation was clearly detected in mesenteric lymph nodes and in the peritoneum after intravenous administration using PET. We confirmed the results with immunohistochemistry. Our results in this model highlight the importance of local FDG administration during diagnostic imaging to precisely assess early peritoneal manifestations of other malignancies (colon, stomach, ovary). These findings also support the importance of applying topical therapies, in addition to systemic treatments in peritoneal cancer spread.

In situ lymphoma imaging in a spontaneous mouse model using the Cerenkov Luminescence of F-18 and Ga-67 isotopes.

Cerenkov luminescence imaging (CLI) is a promising approach to image-guided surgery and pathological sampling. It could offer additional advantages when combined to whole-body isotope tomographies. We aimed to obtain evidence of its applicability in lymphoma patho-diagnostics, thus we decided to investigate the radiodiagnostic potential of combined PET or SPECT/CLI in an experimental, novel spontaneous high-grade B-cell lymphoma mouse model (Bc.DLFL1). We monitored the lymphoma dissemination at early stage, and at clinically relevant stages such as advanced stage and terminal stage with in vivo 2-deoxy-2-[18F]fluoro-D-glucose (FDG) positron emission tomography (PET)/magnetic resonance imaging (MRI) and 67Ga-citrate single photon emission computed tomography (SPECT)/MRI. In vivo imaging was combined with ex vivo high resolution CLI. The use of CLI with 18F-Fluorine (F-18) and 67Ga-Gallium isotopes in the selection of infiltrated lymph nodes for tumor staging and pathology was thus tested. At advanced stage, FDG PET/MRI plus ex vivo CLI allowed accurate detection of FDG accumulation in lymphoma-infiltrated tissues. At terminal stage we detected tumorous lymph nodes with SPECT/MRI and we could report in vivo detection of the Cerenkov light emission of 67Ga. CLI with 67Ga-citrate revealed lymphoma accumulation in distant lymph node locations, unnoticeable with only MRI. Flow cytometry and immunohistochemistry confirmed these imaging results. Our study promotes the combined use of PET and CLI in preclinical studies and clinical practice. Heterogeneous FDG distribution in lymph nodes, detected at sampling surgery, has implications for tissue pathology processing and it could direct therapy. The results with 67Ga also point to the opportunities to further apply suitable SPECT radiopharmaceuticals for CLI.

Role of adipose-associated lymphoid tissues in the immunological homeostasis of the serosal surface.

Although not typical lymphoid organs, analysis of the visceral adipose-associated lymphoid tissues has recently substantially expanded our knowledge about the immunological features of these elusive compartments. Recent data have highlighted their considerable complexity in cellular organization and interactions in several biological processes, including adaptive immune responses, tissue plasticity to accommodate mesenchymal stem cells and progenitors, and providing a suitable microenvironment for serosal tumor propagation. This review aims to present a comprehensive view of the adipose-associated lymphoid tissues in local and systemic immune responsiveness, with particular emphasis on the omental and mesenteric lymphoid tissues in the serosal defense of abdominal organs.