CD27 Human

What is the expression pattern of CD27 across different human immune cell types?

CD27 shows distinct expression patterns across various immune cell populations. It is expressed by resting cells including CD4+ T cells, CD8+ T cells, and NK cells. In the B cell compartment, CD27 is predominantly found on memory B cells, germinal center (GC) B cells, and plasma cells, with plasma cells exhibiting the highest expression levels of all B-cell subsets . Importantly, CD27 expression patterns differ between organs, with switched memory B cells in the spleen expressing lower levels than those in peripheral blood . This differential expression may reflect distinct functional requirements in various immunological contexts.

How does CD27 expression change during B cell development and activation?

CD27 expression is dynamically regulated during B cell maturation and activation. CD27 is gradually upregulated during the germinal center reaction . At the single-cell level in human tonsils, GC B cells with increased numbers of somatic mutations express higher levels of CD27, consistent with observations of increased CD27+ B cells in the GC light zones of human lymph nodes . This suggests CD27 upregulation correlates with B cell maturation state. CD27dull B cells appear early in life and remain relatively abundant in adults, while this subset decreases in the elderly, coinciding with weakened vaccine responses . Furthermore, longitudinal data from pregnant women indicates that CD27dull memory B cells may be ancestral to CD27bright memory B cells, suggesting a developmental relationship .

What is known about CD27's binding partner CD70 and their interaction?

CD70 is the only known ligand for CD27 and shows a much more restricted cellular expression pattern than CD27. CD70 is expressed during the activation of dendritic cells and by T cells . Its expression on B cells is very limited and is almost exclusively found on cells that also express CD27 . This restricted expression pattern of CD70 suggests a highly regulated interaction between CD27 and its ligand, likely controlling the timing and context of CD27 signaling. The interaction between CD27 and CD70 is crucial for proper immune function, as evidenced by the immunological defects observed in CD27-deficient patients .

What are the optimal methods for identifying CD27+ cell subsets in multiparameter flow cytometry?

For effective identification of CD27+ cell subsets, multiparameter flow cytometry should incorporate CD27 alongside other lineage and activation markers. For B cells, CD27 should be combined with IgD and IgM to differentiate between switched (CD27+IgD-), marginal zone-like (CD27+IgM+IgD+), and IgM-only (CD27+IgM+IgD-) memory B cell subsets . For NK cells, pairing CD27 with CD56 allows discrimination between CD27lo/CD56dim and CD27hi/CD56bright subsets . To distinguish between CD27dull and CD27bright populations, proper compensation and standardized gating strategies are essential. Additionally, incorporating markers such as CD21, CD45RB, CD11c, CD39, CD73, and CD95 can further refine the identification of specific memory B cell subpopulations . Calibration beads should be used to standardize fluorescence intensity measurements across experiments.

How can immunoglobulin sequencing be integrated with CD27 expression analysis to study B cell development?

Integrating immunoglobulin (Ig) sequencing with CD27 expression analysis provides powerful insights into B cell evolutionary pathways. Researchers should first isolate CD27+ and CD27- B cell populations using fluorescence-activated cell sorting (FACS), followed by RNA extraction, cDNA synthesis, and Ig gene amplification with primers specific for V-region genes. Next-generation sequencing of these amplicons allows for analysis of somatic hypermutation (SHM) patterns and clonal relationships . This approach has revealed that CD27+ B cells typically contain more somatic mutations than CD27- counterparts, and that CD27- switched memory B cells can be found in the same lineage trees as CD27+ switched memory B cells, albeit with fewer mutations . For comprehensive analysis, paired heavy and light chain sequencing should be performed to reconstruct the complete B cell receptor. When studying CD27-deficient patients, parallel sampling of peripheral blood and lymph nodes would be particularly informative to understand germinal center-dependent versus independent memory B cell formation .

What controls should be included when studying CD27 signaling pathways?

When investigating CD27 signaling pathways, several critical controls should be included. First, both positive and negative biological controls are essential: CD27-deficient cells serve as negative controls, while cells known to respond strongly to CD27 stimulation (such as activated T cells) serve as positive controls. Second, stimulation controls should include anti-CD27 antibodies alone, CD70 (the natural ligand) alone, and combinations with other stimulatory signals (like TCR stimulation for T cells), as CD27 activation often requires co-stimulation to exert functional effects . For signaling studies, both time-course experiments (from seconds to hours) and dose-response analyses are necessary to capture the full signaling dynamics. Phosphorylation-specific antibodies targeting components of the NF-κB, MAP kinase, and PI3K pathways should be employed to track signal transduction. Additionally, inhibitors of specific signaling molecules can help delineate the contribution of individual pathways to CD27-mediated responses.

What are the implications of CD27 deficiency for human immune function?

CD27 deficiency represents a rare primary immunodeficiency with significant clinical consequences. Patients with mutations in the CD27 gene often suffer from EBV-driven lymphoproliferation, which frequently progresses to lymphoma . The B cell compartment is substantially affected, with significantly reduced numbers of CD27+ memory B cells . These patients typically exhibit smaller germinal centers and impaired responses to T cell-dependent vaccination . The plasma cell compartment can also be affected, resulting in drastically reduced serum levels of all immunoglobulin isotypes (IgG, IgA, and IgM) in some patients . CD8+ T cell function is severely impaired in CD27-deficient individuals, although hematopoietic stem cell transplantation can restore this defect and reduce lymphoma risk . Interestingly, initial reports indicate normal frequency of somatic hypermutation in these patients despite their immunological defects . The complex immunological phenotype of CD27 deficiency underscores the multifaceted role of CD27 in human immunity.

How is CD27 expression altered in B cell malignancies, and what are the research implications?

CD27 expression is frequently altered in B cell malignancies, making it both a diagnostic marker and potential therapeutic target. Multiple B cell malignancies express CD27 on their surface, with expression patterns often reflecting the cell of origin . For researchers studying B cell malignancies, analyzing CD27 expression patterns may provide insights into tumor origin and differentiation state. Experimentally, CD27 can be targeted with monoclonal antibodies like 1F5 (CDX-1127), which has shown potential therapeutic effects against CD27-expressing lymphoma or leukemia through mechanisms such as antibody-dependent cellular cytotoxicity . When designing studies targeting CD27 in malignancies, researchers should consider that anti-CD27 antibodies may have dual effects: direct antitumor activity against CD27-expressing malignant cells and enhancement of anti-tumor immunity through CD27's co-stimulatory properties . This dual mechanism necessitates careful experimental design to distinguish between direct cytotoxic effects and immune-mediated tumor control.

What methodological considerations are important when studying CD27 in autoimmune diseases?

When investigating CD27 in autoimmune contexts, several methodological considerations are crucial. First, researchers should analyze both the frequency and absolute numbers of CD27+ and CD27- memory B cell subsets, as proportional changes may not reflect absolute alterations in cell populations. CD21−/low memory B cells, which are enriched in subjects with autoimmune diseases, contain both CD27+ and CD27- cells and should be specifically examined . Longitudinal sampling is preferable to cross-sectional analysis due to fluctuations in disease activity and treatment effects. Flow cytometric analysis should include markers of B cell activation (CD80/CD86), exhaustion (PD-1, FCRL5), and tissue homing receptors to comprehensively characterize CD27+ B cells in disease states. When possible, parallel analysis of affected tissues and peripheral blood should be performed, as the CD27- memory B cell subsets vary significantly between different lymphoid tissues . Additionally, functional assays examining cytokine production, antibody secretion, and autoreactivity of sorted CD27+ and CD27- B cell subsets can provide mechanistic insights into disease pathogenesis.

How do CD27+ memory B cell subsets differ functionally and phenotypically?

CD27+ memory B cells comprise distinct subsets with unique functional and phenotypic characteristics. These include switched (IgG+/IgA+), IgM+IgD+ (marginal zone-like), IgM-only, and a minor population of IgD-only cells . Switched CD27+ memory B cells are traditionally regarded as the main post-germinal center memory B cells, although IgM+ memory B cell subsets also participate in germinal center reactions . Functionally, switched CD27+ memory B cells readily differentiate into plasmablasts upon Toll-like receptor 9 stimulation, as evidenced by the upregulation of Prdm1 . They represent a highly selected memory population that responds quickly to re-infection by differentiating into antibody-secreting cells . In contrast, CD27+IgM+IgD+ B cells, which resemble splenic marginal zone B cells, show high proliferative capacity . CD27dull memory B cells, which appear early in life and remain abundant in adults, may represent precursors to CD27bright memory B cells . These functional distinctions between CD27+ memory B cell subsets reflect their specialized roles in humoral immunity and highlight the heterogeneity within the CD27+ memory B cell compartment.

What distinct roles do CD27hi versus CD27lo NK cell subsets play in human immunity?

Human NK cells can be divided into distinct subsets based on CD27 and CD56 expression. Most peripheral blood NK cells are CD27lo/CD56dim, while a minor CD27hi NK cell population displays a CD56bright phenotype . These subsets exhibit fundamental differences in receptor expression patterns and typical NK cell functions . The CD27lo/CD56dim subset is generally considered more cytotoxic, while CD27hi/CD56bright NK cells tend to be more proficient at cytokine production. This functional dichotomy parallels the distinction observed in murine NK cell subsets based on CD27 expression . The identification of corresponding CD27-defined subsets in both mice and humans facilitates more accurate translational research, allowing better projections of NK cell subset roles from murine models to human pathologies . When designing experiments to compare these subsets, researchers should isolate pure populations using flow cytometry sorting and assess multiple functional parameters including cytotoxicity, cytokine production, proliferation capacity, and response to various activating stimuli to comprehensively characterize their distinct immunological roles.

How does CD27 expression on T cells correlate with their differentiation state and functional capacity?

CD27 expression on T cells closely correlates with their differentiation state and functional capabilities. CD27 is expressed by resting T cells, including both CD4+ and CD8+ populations . During T cell differentiation, CD27 expression is downregulated on effector T cells but maintained on memory CD4+ T cells . This differential expression pattern makes CD27 a useful marker for distinguishing naive, effector, and memory T cell subsets. Functionally, CD27 provides co-stimulatory signals that enhance T cell activation, proliferation, and survival, but only when combined with T cell receptor stimulation . In research settings, analyzing CD27 in conjunction with other differentiation markers such as CD45RA, CCR7, and CD28 provides a more comprehensive assessment of T cell differentiation states. When investigating CD27's role in T cell responses, it's important to note that CD27 signaling requires interaction with its ligand CD70, which has a restricted expression pattern primarily on activated dendritic cells and T cells . The timely and contextual regulation of this receptor-ligand pair is crucial for proper T cell immunity, as evidenced by the impaired CD8+ T cell function observed in CD27-deficient patients .

What are the best approaches for studying CD27 in tissue samples versus blood?

When studying CD27 in different tissues, researchers must adapt their methodologies to account for tissue-specific considerations. For fresh tissue samples, mechanical dissociation followed by enzymatic digestion (using collagenase with DNase I) is recommended to obtain single-cell suspensions while preserving surface CD27 expression. Cryopreserved samples require careful thawing protocols with DNase I to prevent cell clumping. For immunohistochemistry or immunofluorescence on fixed tissues, antigen retrieval methods must be optimized, as some CD27 epitopes are sensitive to fixation. Multiple anti-CD27 antibody clones should be tested, as staining patterns may vary between tissues. Importantly, CD27 expression differs significantly between organs; for example, switched memory B cells in the spleen express lower CD27 levels than those in peripheral blood . Additionally, CD27- memory B cell subsets vary substantially between different lymphoid tissues (peripheral blood, spleen, tonsils, and gut) . Therefore, direct comparisons between tissues should be interpreted cautiously. For comprehensive analysis, multiparameter approaches combining CD27 with tissue-specific markers and other immune cell identifiers provide the most informative results.

How can researchers effectively distinguish between CD27dull and CD27bright populations in flow cytometric analysis?

Distinguishing between CD27dull and CD27bright populations requires careful flow cytometry optimization and standardized analysis strategies. First, proper antibody titration is essential to ensure optimal signal-to-noise ratio without saturation. Fluorophore selection is critical—bright fluorophores (PE, APC) are recommended for resolving subtle differences in CD27 expression levels. To establish consistent gating, fluorescence minus one (FMO) controls should be included to accurately set the CD27-negative boundary. For longitudinal studies, fluorescent calibration beads should be used to standardize measurements across experiments and instruments. Biexponential or logicle display transformations often provide better resolution of CD27dull from CD27-negative cells compared to standard logarithmic scaling. When analyzing data, both median fluorescence intensity and percent positive cells should be reported. Machine learning-based clustering algorithms (e.g., FlowSOM, SPADE) can help objectively identify CD27 expression continua. Recent research has shown that CD27dull memory B cells appear early in life and remain abundant in adults while decreasing in the elderly . These cells may represent precursors to CD27bright memory B cells, as suggested by longitudinal studies of pregnant women , highlighting the biological significance of accurately distinguishing these populations.

What experimental approaches can determine whether CD27 expression differences represent distinct lineages or developmental stages?

Determining whether CD27 expression differences represent distinct lineages or developmental stages requires multiple complementary experimental approaches. Lineage tracing experiments, though challenging in humans, can be performed by isolating CD27-negative, CD27dull, and CD27bright populations and tracking their phenotypic evolution in vitro under various stimulation conditions. Immunoglobulin gene sequencing of sorted CD27 subsets can reveal clonal relationships—cells within the same clone but displaying different CD27 expression levels would suggest a developmental continuum rather than separate lineages . Single-cell RNA sequencing provides high-resolution insights into the transcriptional continuum between CD27 subsets, potentially revealing transitional states. Epigenetic profiling (ATAC-seq, methylation analysis) can identify stable lineage-specific versus plastic developmental differences in chromatin accessibility. In vivo approaches include analyzing samples from rare patients with CD27 deficiency, which could reveal whether CD27-negative memory B cells in these patients resemble CD27-negative memory B cells in healthy donors . Longitudinal sampling during immune responses or pregnancy has proven informative, with evidence suggesting CD27dull memory B cells are ancestral to CD27bright memory B cells . Together, these approaches can distinguish between stable lineage differences and transient developmental stages defined by CD27 expression.

What considerations are important when developing monoclonal antibodies targeting CD27 for cancer immunotherapy?

Development of CD27-targeting monoclonal antibodies for cancer immunotherapy requires careful consideration of multiple factors. Antibody characteristics must be optimized—epitope selection influences whether the antibody will block or mimic CD70 binding, while isotype selection determines effector functions (ADCC, CDC, ADCP). The human monoclonal antibody 1F5 (CDX-1127) binds with high affinity and specificity to human CD27 and competes with ligand binding . Importantly, such antibodies may operate through dual mechanisms: direct effects on CD27+ tumors and immunomodulatory effects on T cells. 1F5 activates T cells only in combination with T-cell receptor stimulation and does not induce proliferation of primary CD27-expressing tumor cells . Preclinical evaluation should include assessment of both direct tumor cell killing and enhancement of anti-tumor immunity. Safety considerations are paramount—1F5 administration to cynomolgus monkeys at doses up to 10 mg/kg was well-tolerated without significant toxicity or depletion of circulating lymphocytes . Combination strategies with other immunotherapies should be explored, as CD27 stimulation may complement checkpoint inhibition. Patient selection strategies should consider tumor CD27 expression levels, immune infiltration patterns, and biomarkers of potential response to maximize therapeutic efficacy while minimizing off-target effects.

How can CD27 expression analysis be integrated into clinical assessment of lymphoid malignancies?

Integration of CD27 expression analysis into clinical assessment of lymphoid malignancies offers valuable diagnostic, prognostic, and therapeutic insights. Diagnostically, CD27 serves as a surface marker on various B- and T-cell malignancies , helping distinguish between different lymphoma subtypes based on their cell of origin. Flow cytometric panels for hematological malignancies should routinely include CD27 alongside lineage-specific and differentiation markers. Quantitative assessment of CD27 expression levels (rather than simple positive/negative categorization) provides more granular information. For minimal residual disease monitoring, CD27 can be incorporated into multiparameter flow cytometry panels to track malignant populations with aberrant CD27 expression patterns. Prognostically, correlation of CD27 expression levels with clinical outcomes can identify patient subgroups with distinct risk profiles. Therapeutically, CD27 expression analysis helps identify patients who might benefit from CD27-targeted therapies like the monoclonal antibody 1F5/CDX-1127, which has shown potential efficacy against CD27-expressing lymphomas in preclinical models . Integration of CD27 assessment with other molecular and cytogenetic markers provides a more comprehensive risk stratification and treatment selection strategy for patients with lymphoid malignancies.

What methodological approaches can assess the relationship between CD27+ memory B cell subsets and vaccine responses?

Assessing relationships between CD27+ memory B cell subsets and vaccine responses requires comprehensive methodological approaches spanning pre-vaccination baseline assessment through long-term memory evaluation. Before vaccination, researchers should establish baseline frequencies and absolute counts of CD27+ memory B cell subsets (switched, IgM+IgD+, IgM-only) using multiparameter flow cytometry. Plasmablast responses should be measured 7-10 days post-vaccination, with antigen-specific cells identified using fluorescently-labeled vaccine antigens. Memory B cell ELISpot assays with polyclonal stimulation can enumerate antigen-specific memory B cells at various timepoints post-vaccination. Single-cell sorting of antigen-specific CD27+ B cells followed by immunoglobulin sequencing reveals clonal expansion and somatic hypermutation patterns in response to vaccination. Longitudinal sampling is crucial, as the quality and longevity of vaccine responses may correlate with specific CD27+ memory B cell subsets. Age-related considerations are important—CD27dull memory B cells are abundant in adults but decrease in the elderly, coinciding with weakened vaccine responses . Systems biology approaches integrating transcriptional, epigenetic, and functional profiling of CD27+ memory B cell subsets with serological responses can identify correlates of protection. These methodological approaches collectively provide a comprehensive assessment of how different CD27+ memory B cell subsets contribute to effective vaccine responses.