Hematopoietic stem cell transplantation (HSCT) is a treatment modality in which hematopoietic stem cells are used as curative therapy for congenital and acquired disorders of the hematopoietic system and metabolic diseases (1). During HSCT, the hematopoietic system is replaced using donor-derived hematopoietic stem cells as allograft. Stem cells can give progeny to functioning erythrocytes, thrombocytes, myeloid lineages and/or lymphocytes, achieving recovery of normal hematopoiesis and immunity.

Restoration of the individual components of the immune system occurs with different dynamics in which innate immunity (neutrophils, monocytes and natural killer cells) typically precedes adaptive immunity (T- and B-lymphocytes). Complete immune reconstitution can take several months up to 2 years after HSCT. Immune reconstitution after allogeneic HSCT has been studied extensively with a main focus on T cell reconstitution. Only limited information is available about B cell reconstitution. In this review we summarize the existing knowledge on B cell reconstitution after pediatric allogeneic HSCT and point out the need and challenges for further investigations. We included studies via systematic literature search in Embase, Medline Epub (Ovid), Cochrane Central and Web of Science including the terms “hematopoietic stem cell transplantation,” “B lymphocyte,” “immune reconstitution,” “child” and synonyms between 01-01-1980 and 31-12-2018. Additional relevant studies were included through references within the identified studies. Lastly, top results of Google Scholar were screened.

Compared to other hematopoietic cell types, B cell reconstitution occurs relatively late after HSCT. The first emerging B cells can be detected in peripheral blood after 1.5–2 months. These are mainly CD24high CD38high transitional B cells (2) (See Figure 1A for all peripheral B-cell subsets). A subdivision of these transitional B cells can be made in the early appearing CD21low B cells, called the T1 cells, and the CD21high B (or T2) cells which develop later (3). Additionally, expansion of CD5+ B cells is reported early after HSCT (4, 5). CD5 is a known marker of human immature B cells, but its expression covers a broader range of B cell developmental/differentiation stages. More mature CD5+ B cell subpopulations are characterized by their regulatory properties such as IL-10 production upon activation by bacterial or parasitic antigens, and their constitutive death-inducing ligand expression (6–10). The CD5+ B cell expansion early after HSCT is likely to be a reflection of B cell immaturity, but may also point to a role for regulatory B cells in controlling immune reactions and autoimmunity after HSCT. Several months after HSCT, the transitional B cells decrease in number and gradually naive mature (CD24int CD38int) B cells emerge to become the predominant population. In the course of the first year following HSCT, naive mature B cells represent more than 80% of the peripheral blood B lymphocytes. (2). To obtain more insight in the mechanism and dynamics of B cell regeneration, κ-deleting recombination excision circles (KRECs) might serve as a useful biomarker for replication history and to evaluate the onset of de novo B-cells, as KRECs have been reported to be positively correlated with B cell numbers after transplantation (11–15). At one year after HSCT, the B cell reconstitution stabilizes reaching age-corrected normal total B cell counts in peripheral blood in most patients (Figure 1B) (16–20). Looking further into the B cell populations, non-switched (CD27+IgM+IgD+/-) and switched (CD27+IgD-IgM-) memory B-cells appear slowly, taking up to two years or longer after HSCT to reach normal age matched levels (16, 17, 20, 21). Especially non-switched B cells seem to remain below normal values, suggesting defects in this maturation stage. During the process of B cell maturation in general, mature B lymphocytes further differentiate into memory B cells, and may undergo isotype switching and affinity maturation in a T cell dependent germinal center reaction (Figure 1A). In this process, cognate interaction between T follicular helper (Tfh) cells and specialized follicular dendritic cells is pivotal. As a consequence, the quality and dynamics of CD4 T cell, and thus also Tfh, reconstitution after HSCT will also impact on B cell differentiation and may thus contribute to an impaired or arrested maturation of B cells. (22–24). However, even in the presence of donor CD4+ T cells that are capable of supporting the process of somatic hyper mutation, the incidence of somatic hypermutation is decreased in recipient B cells in cell culture (25). It could be that treatment given prior to transplantation disrupts secondary lymphoid organs, which are necessary for the introduction of somatic hypermutations in the variable domains of the immunoglobulin molecules and affinity maturation in the germinal centers (26). Immune responses against polysaccharides seem frequently impaired in HSCT patients (27, 28). Polysaccharide antibody responses are important for the T cell independent defense to encapsulated bacteria, in which marginal zone B cells play an important role (29, 30). The impaired reconstitution of this subset might indicate why certain patients encounter specific problems with susceptibility to encapsulated bacteria such as pneumococcus. The counterpart of marginal zone B cells, IgM memory B cells, seems also to be reduced in long term transplanted patients (16, 17, 20, 21).

Immunoglobulin (Ig) levels seem to recover in parallel to B cell reconstitution, in which recovery of Ig subclasses usually occurs in a distinctive order (16, 31–33). After HSCT, Ig levels drop, reflecting the absence of Ig producing B cells. Some Ig production may persist, probably due to surviving long-lived plasma cells of host origin (34). As a reflection of normal ontogeny, IgM production will reconstitute relatively early and, on average, reaches normal levels within the first 6 months after HSCT. Similar to IgM, IgG generally reaches normal levels in the second half of the first year, whereas normalization of IgA levels may take up to 5 years after HSCT. IgG subclasses, on average, reach normal serum levels within 5 months (IgG1), 9 months (IgG3), or up to 2 years (IgG2 and IgG4). However, the time frame is highly variable and can be influenced by several factors such as the underlying disease, stem cell source, and type of donor (16, 31–33).

For complete humoral immune reconstitution after HSCT, generation of a diverse BCR repertoire is necessary. Literature on the diversification of the BCR after HSCT is limited. In the last 10 years, investigation of BCR diversification after HSCT has stagnated and studies performed are limited by older techniques or are difficult to generalize because of the small sample size. Analysis of the pattern of VH3- and VH4-gene usage based on 700 rearrangements in four patients suggested that the B-cell receptor repertoire shows the same (limited) repertoire of VH genes at 90 days and 1 year after HSCT (35). Other studies showed evidence that generation of the new repertoire occurs gradually and suggest that the CDR3 regions post HSCT are similar to CDR3 regions in adults and do not follow fetal ontogeny (36–38). The CDR3 length in the memory B cell compartment has a specific restriction compared to healthy controls, resulting in an oligoclonal repertoire early after HSCT (39). These methods only provide rough information about the BCR repertoire post HSCT. In adults, one study investigated the IGH repertoire before and after HSCT in acute myeloid leukemia patients using next generation sequencing. In general, they observed lower repertoire diversity after HSCT than before (40). Furthermore, each individual appeared to have highly unique and characteristic IGH repertoire of switched memory B-cells, which allowed the investigators to separate donor and recipient derived B cell clones. Interestingly, this showed in some cases persistence of recipient B cells which indicates that recipient B cells may still contribute to protective immunity after HSCT. The study analyzed the VH1 Ig repertoire, which represents only about 10% of all Ig sequences in humans. All of these studies exclusively investigated Ig heavy chain diversity. To the best of our knowledge, there are no studies investigating Ig light chain.

Based on available literature, HSCT-treated patients are frequently affected by an unbalanced, incomplete and, therefore, abnormal BCR repertoire, leading to impaired humoral immunity with associated risks of infectious and/or auto-immune complications. So far, studies have been limited by small cohorts and low-throughput or low resolution techniques. More in-depth analyses are needed to understand the integrated evolution of cellular and repertoire reconstitution and the influence of different HSCT-related factors on immune repertoire formation and B cell function require after transplantation.

In a significant proportion of HSCT recipients, antibody titers of vaccine-preventable diseases decline over the years, if these recipients are not revaccinated (86–89). Vaccination responses after HSCT are dependent on both T- and B cell reconstitution. An exception is the polysaccharide antibody response, which is completely B cell dependent (90). The polysaccharide antibody response plays a role in the immunization against encapsulated bacteria, such as pneumococcus. HSCT recipients are more susceptible to infections during the post transplantation period (91–93). The risk of pneumococcal invasive disease is increased both early and late after HSCT, reaching 30-fold higher risks compared to the general population after 10 years, suggesting long lasting defects in B cell reconstitution even after revaccination (94–97). The risk of pneumococcal disease correlates with presence/occurrence of GVHD, suggesting a link between the functional dysregulation of B cells and GVHD (21, 94, 96). Whereas, an association with hypogammaglobulinemia has been reported, increased susceptibility to pneumococcal disease is most often due to a more selectively impaired immune response against polysaccharides (27, 28, 95). A poor response to polysaccharide vaccines, is hypothesized to be caused by the lack of unswitched memory B cells, which are reduced after transplantation (98). A poor response to polysaccharide vaccines is hypothesized to be caused by reduced MZ B cells or IgM memory B cells, which are reduced after transplantation (16, 17, 20, 21, 98). Reduced numbers of IgM memory B cells and increased risk of encapsulated bacterial infection has also been observed in young children, patients with common variable immune deficiency (CVID) and splenectomized patients (29, 99–101). Revaccination usually occurs with inactivated vaccines, as live attenuated vaccines have the potential to induce active disease in immunosuppressed patients. However below normal values, class switched memory B cells are observed as early as 3 months after HSCT (17). It is largely unknown if these cells are already capable of an immune response, taking into account the slow reconstitution of CD4+ T cells. In current guidelines, revaccination starts 3-6 months after HSCT, but looking at thresholds of the CD4+ T cells and ability for class switch recombination might be a useful biomarker to guide the timing of vaccination compared to fixed time point after HSCT. Live attenuated vaccines could be considered two years after HSCT, in patients without cGVHD or immunosuppression (102).

B cell reconstitution after HSCT including BCR repertoire formation and diversification, is awaiting new insights in order to develop better treatment strategies for prevention of clinical complications due to defects in B cell mediated immunity. For example, clinical outcome is still suboptimal in a proportion of HSCT-treated severe combined immunodeficiency (SCID) patients, due to humoral immune dysfunction. Some of these patients still suffer from persistent (humoral) immunodeficiency, auto-immunity and/or immune dysregulation leading to impaired quality of life (103). Although SCID represents the prototype of inherited immune disorders, an increasing spectrum of patients with inherited immune disorders is being genetically identified (>250 monogenetic diseases) (104). This steadily growing group of transplanted patients with non-SCID inherited immune disorders faces similar challenges regarding the long term quality of their (sometimes partially) corrected immune system. To evaluate the long term quality of immune reconstitution, B cell immunity recovery can serve as an indicator of immune fitness after HSCT.

We have limited information on the diversity of the BCR repertoire in the various B cell subsets in both peripheral blood and BM of HSCT-treated patients. Important insights have been obtained through flow cytometric analysis of peripheral B-cells after HSCT. However, to gain better understanding of numerical and functional B-cell reconstitution more in depth-analysis of cellular dynamics (using flow cytometry and KREC analysis), molecular aspects (BCR repertoire analysis) and antigen specificity is needed. Extensive analysis of B cell reconstitution can be done through modern flow cytometry and sequence techniques or even simultaneously with mass cytometry (105, 106). With modern sequencing techniques, it is possible to look at both the BCR heavy and light chain. Combined with single cell RNA sequencing, the integrated gene expression per cell within individual patients can be investigated (107). These modern techniques/methodology will make it possible to investigate the influence of aforementioned parameters on B cell repertoire development after HSCT in an innovative and more accurate way. This could revolutionize the knowledge about the B cell reconstitution and pinpoint the individual B cell maturation problems of transplanted patients.

NvdM and MvdB contributed conception of the paper; NvdM did literature search and wrote the first draft of the manuscript; DB, MvdB, and AL wrote sections of the manuscript. All authors contributed to manuscript revision, read and approved the submitted version.