Deficiency of Tfh Cells and Germinal Center in Deceased COVID-19 Patients

Summary:The COVID-19 pandemic caused by SARS-CoV2 is characterized by a remarkable variation in clinical severity ranging from a mild illness to a fatal multi-organ disease.Understanding the dysregulated human immune responses in the fatal subjects is critical for management of COVID-19 patients and the pandemic.In this study,we examined the immune cell compositions in the lung tissues and hilar lymph nodes using immunohistochemistry on 6 deceased COVID-19 patients and 4 focal organizing pneumonia(FOP)patients who underwent lung surgery and served as controls.We found a dominant presence of macrophages and a general deficiency of T cells and B cells in the lung tissues from deceased COVID-19 patients.In contrast to the FOP patients,Tfh cells and germinal center formation were largely absent in the draining hilar lymph nodes in the deceased COVID-19 patients.This was correlated with reduced IgM and IgG levels compared to convalescent COVID-19 patients.In summary,our data highlight a defect of germinal center structure in deceased COVID-19 patients leading to an impaired humoral immunity.Understanding the mechanisms of this deficiency will be one of the key points for the management of this epidemic.

Requirement for cyclin D3 in germinal center formation and function

Germinal centers （GC） of secondary lymphoid tissues are critical to mounting a high-affinity humoral immune response. B cells within the GC undergo rapid clonal expansion and selection while diversifying their antibody genes. Although it is generally believed that GC B cells employ a unique proliferative program to accommodate these processes, little is known about how the GC-associated cell cycle is orchestrated. The D-type cyclins constitute an important component of the cell cycle engine that enables the cells to respond to physiological changes. Cell type- and developmental stage-specific roles of D-type cyclins have been described but the cyclin D requirement during GC reaction has not been addressed. In this study, we report that cyclin D3 is largely dispensable for proliferation and Ig class switching of in vitro activated B cells. In contrast, GC development in Ccnd3^-/- mice is markedly impaired, as is the T cell-dependent antibody response. Within the GC, although both switched and unswitched B cells are affected by cyclin D3 inactivation, the IgM^- pool is more severely reduced. Interestingly, despite a compensatory increase in cyclln D2 expression, a significant number of Ccnd3^-/- GC B cells accumulate in quiescent GO state. Lastly, although cyclin D3 inactivation did not disrupt BCL6 expression in GC B cells, it completely blocked the GC promoting effect of BCL6 overexpression, suggesting that cyclin D3 acts downstream of BCL6 to regulate GC formation. This is the first demonstration that cyclin D3 plays an important and unique role at the GC stage of B cell development.

IMMUNOPHENOTYPING OF CUTANEOUS GERMINAL CENTER CELL-DERIVED LYMPHOMAS

Monoclonal antibodies were used to label cutaneous germinal center cell-derived lymphomas <CGCCL) obtained from 10 patients. According to the Kiel classification, they were classified into 2 types. Eight patients had centroblastic/centrocytic <CB/CC) lymphomas while 2 patients and centrocytic (CC) lymphomas. After monoclonal antibody labelling, the results were consistent with those of the clinical and morphologic analyses. Of the 10 cases, 9 were B1 positive, 6 were K positive, and 4 were λ positive. In 8 cases labeled with immunoglobulin, 6 were IgGFab positive, 2 were IgM positive and 8 were IgA negative. Five cases (CB/CC 3, CC 2) were both Bl, K and IgG positive (γ K). Four cases CB/CC were both Bl and A positive. Only one case (CB/CC) was both K and IgM positive (μ K). Two cases (CB CC) were both A and IgG positive (γ λ). The results indicate that Bl, K and A are the most important markers to phenotype cutaneous B-cell lymphomas. Our findings also show a higher percentage of y K types in CGCCL as compared with Western countries.

The RNA binding proteins TIA1 and TIAL1 promote Mcl1 mRNA translation to protect germinal center responses from apoptosis

Germinal centers(GCs)are essential for the establishment of long-lasting antibody responses.GC B cells rely on post-transcriptional RNA mechanisms to translate activation-associated transcriptional programs into functional changes in the cell proteome.However,the critical proteins driving these key mechanisms are still unknown.Here,we show that the RNA binding proteins TIA1 and TIAL1 are required for the generation of long-lasting GC responses.TIA1-and TIAL1-deficient GC B cells fail to undergo antigen-mediated positive selection,expansion and differentiation into B-cell clones producing high-affinity antibodies.Mechanistically,TIA1 and TIAL1 control the transcriptional identity of dark-and light-zone GC B cells and enable timely expression of the prosurvival molecule MCL1.Thus,we demonstrate here that TIA1 and TIAL1 are key players in the post-transcriptional program that selects high-affinity antigen-specific GC B cells.

Germinal center-dependent and-independent immune responses of tumor-infiltrating B cells in human cancers

B cells play essential roles in immunity,mainly through the production of high affinity plasma cells(PCs)and memory B(Bmem)cells.The affinity maturation and differentiation of B cells rely on the integration of B-cell receptor(BCR)intrinsic and extrinsic signals provided by antigen binding and the microenvironment,respectively.In recent years,tumor infiltrating B(TIL-B)cells and PCs(TIL-PCs)have been revealed as important players in antitumor responses in human cancers,but their interplay and dynamics remain largely unknown.In lymphoid organs,B-cell responses involve both germinal center(GC)-dependent and GC-independent pathways for Bmem cell and PC production.Affinity maturation of BCR repertoires occurs in GC reactions with specific spatiotemporal dynamics of signal integration by B cells.In general,the reactivation of high-affinity Bmem cells by antigens triggers GC-independent production of large numbers of PC without BCR rediversification.Understanding B-cell dynamics in immune responses requires the integration of multiple tools and readouts such as single-cell phenotyping and RNA-seq,in situ analyses,BCR repertoire analysis,BCR specificity and affinity assays,and functional tests.Here,we review how those tools have recently been applied to study TIL-B cells and TIL-PC in different types of solid tumors.We assessed the published evidence for different models of TIL-B-cell dynamics involving GC-dependent or GC-independent local responses and the resulting production of antigen-specific PCs.Altogether,we highlight the need for more integrative B-cell immunology studies to rationally investigate TIL-B cells as a leverage for antitumor therapies.

Mechanisms of action of BCL6 during germinal center B cell development

The transcriptional repressor B cell lymphoma 6(BCL6) controls a large transcriptional network that is required for the formation and maintenance of germinal centers(GC). GC B cells represent the normal counterpart of most human B-cell lymphomas, which are often characterized by deregulated BCL6 expression or BCL6-mediated pathways. BCL6 suppresses gene transcription largely through recruitment of its co-repressors through its distinct repression domain. Understanding the precise biological roles of each repression domain in normal and malignant B cells is helpful for development of targeted inhibition of BCL6 functions that is emerging as the basis for design of anti-lymphoma therapies. This review focuses on recent progress in the molecular mechanisms of action of BCL6 in B cells and discusses remaining unresolved questions related to how these mechanisms are linked to normal and malignant B cell development.

Germinal center kinases in immune regulation

Germinal center kinases （GCKs） participate in a variety of signaling pathways needed to regulate cellular functions including apoptosis, cell proliferation, polarity and migration. Recent studies have shown that GCKs are participants in both adaptive and innate immune regulation. However, the differential activation and regulatory mechanisms of GCKs, as well as upstream and downstream signaling molecules, remain to be fully defined. It remains unresolved whether and how GCKs may cross-talk with existing signaling pathways. This review stresses the progresses in research of GCKs relevant to the immune system.

Ephrin-B1-mediated regulation of germinal center T-cell territoriality and function

Subject Code:C08With the support by the National Natural Science Foundation of China,a study by the research group led by Prof.Qi Hai(祁海)from the School of Medicine,Tsinghua University revealed a novel mechanism that regulates the germinal center reaction,aphysiological process that underlies high-quality

ARTIFICIAL IMMUNE ALGORITHM OF MULTICELLULAR GROUP AND ITS CONVERGENCE

Objective To find out more extrema simultaneously including global optimum and multiple local optima existed in multi-modal functions. Methods Germinal center is the generator and selector of high-affinity B cells, a multicellular group's artificial immune algorithm was proposed based on the germinal center reaction mechanism of natural immune systems. Main steps of the algorithm were given, including hyper-mutation, selection, memory, similarity suppression and recruitment of B cells and the convergence of it was proved. Results The algorithm has been tested to optimize various multi-modal functions, and the simulation results show that the artificial immune algorithm proposed here can find multiple extremum of these functions with lower computational cost. Conclusion The algorithm is valid and can converge on the satisfactory solution set D with probability 1 and approach to global solution and many local optimal solutions existed.

B cell development and antibody responses in human immune system mice:current status and future perspective

Humanized immune system(HIS)mice have been developed and used as a small surrogate model to study human immune function under normal or disease conditions.Although variations are found between models,most HIS mice show robust human T cell responses.However,there has been unsuccessful in constructing HIS mice that produce high-affinity human antibodies,primarily due to defects in terminal B cell differentiation,antibody affinity maturation,and development of primary follicles and germinal centers.In this review,we elaborate on the current knowledge about and previous attempts to improve human B cell development in HIS mice,and propose a potential strategy for constructing HIS mice with improved humoral immunity by transplantation of human follicular dendritic cells(FDCs)to facilitate the development of secondary follicles.

Dhx33 promotes B-cell growth and proliferation by controlling activation-induced rRNA upregulation

Upon recognition of foreign antigens,naïve B cells undergo rapid activation,growth,and proliferation.How B-cell growth and proliferation are coupled with activation remains poorly understood.Combining CRISPR/Cas9-mediated functional analysis and mouse genetics approaches,we found that Dhx33,an activation-induced RNA helicase,plays a critical role in coupling B-cell activation with growth and proliferation.Mutant mice with B-cell-specific deletion of Dhx33 exhibited impaired B-cell development,germinal center reactions,plasma cell differentiation,and antibody production.Dhx33-deficient B cells appeared normal in the steady state and early stage of activation but were retarded in growth and proliferation.Mechanistically,Dhx33 played an indispensable role in activation-induced upregulation of ribosomal DNA(rDNA)transcription.In the absence of Dhx33,activated B cells were compromised in their ability to ramp up 47S ribosomal RNA(rRNA)production and ribosome biogenesis,resulting in nucleolar stress,p53 accumulation,and cellular death.Our findings demonstrate an essential role for Dhx33 in coupling B-cell activation with growth and proliferation and suggest that Dhx33 inhibition is a potential therapy for lymphoma and antibody-mediated autoimmune diseases.

TLR signaling in B-cell development and activation

Expression of Toll-like receptors （TLRs） in B cells provides a cell-intrinsic mechanism for innate signals regulating adaptive immune responses. In combination with other signaling pathways in B cells, including through the B-cell receptor （BCR）, TLR signaling plays multiple roles in B-cell differentiation and activation. The outcome of TLR signaling in B cells is largely context-dependent, which partly explains discrepancies among in vitroand in vivostudies, or studies using different immunogens. We focus on recent findings on how B-cell-intrinsic TLR signaling regulates antibody responses, including germinal center formation and autoantibody production in autoimmune disease models. In addition, TLR signaling also acts on the precursors of B cells, which could influence the immune response of animals by shaping the composition of the immune system. With TLR signaling modulating immune responses at these different levels, much more needs to be understood before we can depict the complete functions of innate signaling in host defense.

A Recombinant Rabies Virus Expressing Fms-like Tyrosine Kinase 3 Ligand(Flt3L) Induces Enhanced Immunogenicity in Mice

Rabies is a zoonotic disease that still causes 59,000 human deaths each year,and rabies vaccine is the most effective way to control the disease.Our previous studies suggested that the maturation of DC plays an important role in enhancing the immunogenicity of rabies vaccine.Flt3L has been reported to own the ability to accelerate the DC maturation,therefore,in this study,a recombinant rabies virus expressing mouse Flt3L,designated as LBNSE-Flt3L,was constructed,and its immunogenicity was characterized.It was found that LBNSE-FU3L could enhance the maturation of DC both in vitro and in vivo,and significantly more TFH cells and Germinal Center B(GC B)cells were generated in mice immunized with LBNSE-FU3L than those immunized with the parent virus LBNSE.Consequently,expressing of Flt3L could elevate the level of virus-neutralizing antibodies(VNA)in immunized mice which provides a better protection from a lethal rabies virus challenge.Taken together,our study extends the potential of Flt3L as a good adjuvant to develop novel rabies vaccine by enhancing the VNA production through activating the DC—Tfh^GC B axis in immunized mice.

Homeostasis and regulation of autoreactive B cells

In contrast to the previous belief that autoreactive B cells are eliminated from the normal repertoire of B cells,many autoreactive B cells actually escape clonal deletion and develop into mature B cells.These autoreactive B cells in healthy individuals perform some beneficial functions in the host and are homeostatically regulated by regulatory T and B cells or other mechanisms to prevent autoimmune diseases.Autoreactive B-1 cells constitutively produce polyreactive natural antibodies for tissue homeostasis.Recently,autoreactive follicular B cells were reported to participate actively in the germinal center reaction.Furthermore,the selection and usefulness of autoreactive marginal zone(MZ)B cells found in autoimmune diseases are not well understood,although the repertoire of MZ B-cell receptors(BCRs)is presumed to be biased to detect bacterial antigens.In this review,we discuss the autoreactive B-cell populations among all three major B-cell subsets and their regulation in immune responses and diseases.

Diffuse large B-Cell lymphoma:from novel molecular classifications to tailored targeted therapies

Diffuse large B-cell lymphoma(DLBCL)is a heterogeneous disease comprising multiple genetic subtypes that translates and impacts clinical outcomes after standard chemoimmunotherapy.Our initial understanding of the complex biological subtypes of DLBCL began with the identification of cell of origin(COO),and now has evolved to include even more specific subtypes defined by genetic signatures and mutations.These newer classifications lend themselves to the application of precision-based medicine,allowing us to tailor new treatment platforms that target specific oncogenic drivers in order to improve DLBCL outcomes.Essential to this is the development of genetic assays and tools that are reliable and readily available to assist in the application of these molecular classifications to real-world use.In this review,we discuss the history of DLBCL classification systems and their implication on clinical investigation as well as novel therapeutic options in DLBCL.