Expression of the recombination-activating genes in extrafollicular lymphocytes but no apparent reinduction in germinal center reactions in human tonsils

Abstract

V(D)J recombination in lymphocytes is (Ig) and T-cell receptor (TCR) genes is mediated by the products of 2 recombinationactivating genes (RAG1 and RAG2) which recognize specific recombination signal sequences (RSS) flanking the V, D, and J gene segments. 1,2 The RAGs are transiently and coordinately expressed during B-cell and T-cell ontogeny in bone marrow and thymus, respectively. [3][4][5] Upon completion of V(D)J rearrangement, RAG expression is shut down. 3 IgM-expressing immature bone marrow B cells recognizing self antigens can re-express the RAGs, leading to further rearrangement of the light chain locus. 6 This process, termed receptor editing, is thought to play a role in the elimination of autoreactive B cells. 6 Mature lymphocytes were generally believed to be incapable of expressing the RAG, thus maintaining allelic exclusion. A number of recent studies, however, have raised the possibility that secondary V(D)J recombination may occur in peripheral lymphoid tissue B cells in the context of germinal center reactions. Thus, RAG transcripts and proteins have been detected in germinal center B cells from lymph nodes, spleens, and Peyer patches in mice. 7,8 Using polymerase chain reaction (PCR), RSS DNA strand breaks characteristic of V(D)J recombination were detected in interleukin 4 (IL-4)-stimulated mature mouse B cells and in germinal center B cells from immunized mice. 9,10 Han et al have suggested that reinduction of RAG expression may occur in germinal center B cells as a result of diminished or abolished antigen binding, thus rescuing cells which would otherwise have been lost to apoptotic cell death. 10 Although initial studies have used mouse models, subsequent reports have provided evidence that similar mechanisms may operate in humans. RAG1 and RAG2 transcripts have been detected in human germinal center B lymphocytes by reverse transcriptase (RT)-PCR. 11 In human tonsils, RSS DNA breaks were detectable in germinal center centrocytes but not in follicle mantle cells. 12 Moreover, RAG, terminal deoxynucleotidyl transferase (TdT), VpreB, and -like transcripts were detected in centrocytes by PCR analysis of fluorescence activated cell sorting (FACS) sorted cell populations. 12,13 Nevertheless, the exact nature and localization of RAG-expressing cells in peripheral lymphoid tissues is still controversial and immunohistochemical studies have not clarified this issue. Expression of RAG genes has been variably reported in the light zones of germinal centers, 8 in germinal center apoptotic bodies, 14 and in scattered cells of germinal center, mantle zone, and extrafollicular area. 13 To resolve this issue, we have examined the expression of RAGs and TdT in human lymphoreticular tissues using in situ hybridization and immunohistochemistry, respectively. Materials and methods TissuesFormalin-fixed and paraffin-embedded tonsillectomy specimens from 25 patients aged 3 to 49 years were studied. All cases showed prominent follicular hyperplasia but no evidence of neoplastic disease. In addition, paraffin ...

Lambda Light Chain Revision in the Human Intestinal IgA Response

“…The expression of RAG in secondary and tertiary lymphoid organs has been debated partly due to divergent findings (15)(16)(17)(18)(19)38). Even among studies that have identified expression of RAGs in secondary lymphoid tissues, expression has been reported in Ignegative cells, consistent with centroblasts as observed here (17), and also in centrocytes (38).…”

“…Evidence of revision in man has been observed in autoimmunity and malignant human tissues (15,16), though it has not been universally accepted to be part of normal B cell physiology. Expression of RAGs, which would be essential for revision, has been observed in germinal centers by some groups but not others (15,17,18). Data from a transgenic mouse model suggested that RAGs are not re-expressed in the periphery but that expression may be retained by recent emigration from the bone marrow (19).…”

B cell receptor revision diminishes the autoreactive B cell response after antigen activation in mice

“…These observations underscore the continuing uncertainty regarding the expression and function of RAG in peripheral B cells. Furthermore, B cells expressing RAG have also been observed outside of GCs in immunized mice (52) and in extrafollicular areas of human tonsil (53). Detection of light chain rearrangement products in RAG-expressing B cells in the periphery clearly suggested that receptor editing or revision may occur in B cells beyond the immature stage (9,(54)(55)(56).…”

Evidence for a stepwise program of extrathymic T cell development within the human tonsil

“…TdT + cells have been identified in the human tonsil near the fibrous scaffold (8)(9)(10)(11). However, they have yet to be characterized with regard to origin, hematopoietic lineage, or function.…”

“…Similarly, CD5 + CD7 + CD3 -cells have been identified in neonatal intestine, and PTCRA, TdT (also known as DNTT), and RAG2 mRNA can be detected in whole lysates of these tissues (7); yet, whether or not CD34 + T cell progenitors reside there is unclear. Finally, several reports have identified TdT + cells in extrafollicular zones of the human tonsil near fibrous scaffolding (8)(9)(10)(11). However, little is known about the function or phenotype of these cells.…”