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IMMUNOBIOLOGY
From the Servicio de Inmunología and Unidad de
Investigación, Hospital Universitario Puerta del Mar,
Cádiz, Spain.
Plasma cells (PCs) are the final B-cell differentiation stage.
Recent evidence reveals relevant functional differences within the PC
compartment. In rodents, early PCs formed in secondary lymphoid tissues
show enhanced apoptosis and short life span, whereas PCs present in a
final destination organ, such as the bone marrow (BM), have reached a
stable prolonged survival state. BM PCs arrive at this organ as
a circulating precursor whose cellular nature remains uncertain. An
initial aim of this study was to characterize this circulating cell. We
hypothesized that antibody-secreting cells detectable in the human
blood after immunization might be a candidate precursor. These
cells were obtained from the blood of volunteers immunized 6 days
earlier with tetanus toxoid (tet), and they were unambiguously
identified as PCs, as demonstrated by their expression of the
CD38h phenotype, by morphology, by immunoglobulin (Ig)
intracytoplasmic staining, and by IgG-tet-secreting capacity in vitro.
In addition, by using the common CD38h feature, human PCs
from tonsil (as a possible source of early PCs), from blood from
tet-immunized donors (as the putative precursors of BM PCs), and from
BM (as a deposit organ) have been purified and their phenotypes
compared. The results show that a variety of differentiation
molecules, proteins involved in the control of apoptosis, the B-cell
transcription factors, positive regulatory domain I-binding
factor 1/B lymphocyte-induced maturation protein 1 and B
cell-specific activating protein and, at least partially, the
chemokine receptor CXCR4 were expressed by human PCs following a
gradient of increasing maturity in the direction: tonsil Plasma cells (PCs) are the morphologically
well-defined cellular end point of the B-lymphocyte differentiation
sequence, and, as such, they exhibit biochemical and structural
features indicative of their full commitment to the synthesis and
secretion of antibody (Ab). Therefore, PCs are ultimately responsible
for the humoral immune response. Experiments in rodents have revealed
that, soon after antigen (Ag) entry, PCs are formed in inductive
territories of the secondary lymphoid tissue, initially within
Ag-activated foci occurring in the T-cell areas of draining lymphoid
tissue and, if the Ag persists, they are also generated in germinal
centers (GCs).1-3 After this early phase, the number of
PCs falls drastically in these areas and they start accumulating in
final deposit locations, namely the bone marrow (BM) and the lamina
propria (LP), for systemic and mucosal humoral responses,
respectively.4-6 PCs present in the deposit organs are not
formed in situ, but they derive from close precursors generated in
distant lymphoid organs as a result of Ag stimulation, which migrate
into these areas through the circulation.7,8 The nature of
this PC precursor has not been fully clarified. It is well established
that PCs present in the deposit organs are the cells responsible for
the majority of Ig and Ab formation in vivo.4,6 Despite
their morphologic and functional similarities, PCs contained in
different organs show marked differences in several respects. Thus,
experiments of 3H-thymidine incorporation in rats indicate
that Ig-secreting cells from inductive areas (lymph nodes, spleen) show
a life span of less than 3 days, whereas the life span of BM PCs is
more than 3 weeks.9 More recently, experiments in mice
using bromodeoxyuridine staining demonstrated the existence of
long-lived (several months after immunization) Ab-secreting PCs
found almost exclusively in the BM, whereas the majority of
early formed Ab-secreting PCs present in peripheral lymphoid
tissues exhibited a very short life span,10,11 and died
soon after formation, apparently by apoptosis.5 In
addition, differences in the number and quality of the IGV
gene somatic mutations within the PC pool have been documented,
indicative of a progressive enrichment in PCs that synthesize Ab of
higher affinity in the deposit organ.12,13 Collectively,
these data indicate that PC functional and maturational levels are not
homogeneous, and that their capabilities increase from the early PC
stage occurring in lymphoid secondary tissues, to the BM PCs. The
mechanisms that determine these differences remain largely unclear.
Less is known about the formation and maturation of PCs in humans. Low
numbers of PCs are present in similar inductive areas of the secondary
lymphoid organs, and it is thought that they are the result of ongoing
Ag activation.14 In addition, human BM becomes the major
reservoir of Ig-secreting PCs in the systemic humoral immune response,
and, as in animal models, human BM PCs are not generated in situ, but
seem to be formed in inductive areas, and then reach the BM as
circulating PC precursors.15 In this regard, following
immunization to a large variety of Ags, specific Ab-forming cells
generated in the local lymphoid tissues can be transiently detected in
the human circulation.16-19 It is conceivable that these
circulating Ab-secreting cells are the precursors of BM PCs.
Furthermore, human Ig-forming cells from secondary lymphoid organs
such as tonsil and lymph nodes exhibit short Ig secretion kinetics (3 days) in culture, whereas BM and LP PCs secrete Ig in vitro for more
prolonged periods.16-23 In addition, human tonsillar
PCs, as well as Ag-induced circulating Ab-forming cells or blood
pre-PCs observed in reactive plasmacytosis, can undergo apoptosis
either spontaneously or induced by CD95 cross-linking.24-26 In contrast, human BM PCs are not
susceptible to this regulatory mechanism.25 Taken
together, these data indicate the existence of relevant differences
within the human PC compartment. Although their significance remains
uncertain, these observations suggest the possibility that human PCs
undergo their maturational progression in a way similar to that
demonstrated for PCs in rodent models.
Terminal differentiation of B lymphocytes into the PC stage is
under the control of several transcription factors, some of which have
recently been identified. First, B cell-specific activating protein
(BSAP), the product of the pax 5 gene, is a transcription factor expressed only by B-lymphoid cells, in which it plays an essential role in the B-cell lineage commitment,27 as well
as in B-cell growth and survival during the ontogenic development and
in the initial activation and differentiation of mature B lymphocytes.28 In fact, the capacity to express BSAP is
lost only in the latter stages of B-cell
differentiation.29 In contrast, the transcription factor
positive regulatory domain I-binding factor 1 (PRDI-BF1), the murine
homolog of which is known as Blimp-1 (B lymphocyte-induced maturation
protein 1), is expressed only in the later B-cell differentiation
stages as documented in both mouse and human systems,30-32
where this factor appears to control a variety of aspects of the
maturation program of PCs.30
A common feature of human PCs is the expression of high levels of CD38
molecules on their surface (CD38h cells),14
and this marker has allowed their identification and, to a certain
extent, their phenotypic characterization and isolation.21,24,33-35 In the present study, circulating
Ag-induced Ab-secreting cells have been unambiguously identified as
CD38h cells as well as PCs, and the common
CD38h expression on human PCs has been used to perform a
comparative and extensive phenotype analysis of: (1) PCs obtained from
an inductive secondary lymphoid organ such as the tonsil, as a possible source of early PCs; (2) PCs released to the circulation after Ag
immunization, as a possible transitional phase; and (3) normal BM PCs,
as an example of PCs from a final destination organ. In addition, the
expression of BSAP and PRDI-BF1/Blimp-1 messenger RNA (mRNA) by
purified PCs from these 3 locations has been determined by
reverse transcription-polymerase chain reaction (RT-PCR). The results
of the phenotyping reveal that, within the PC compartment, a gradient
of increasing maturity was observed in the direction: tonsil Materials
Purification of CD38h and B cells from different
tissues
Cell staining and flow cytometry analysis Cells (200 µL cells at 0.5-5 × 106 cell/mL) were incubated with optimal concentrations of mAb for 20 minutes in the dark at 4°C. For Bcl-2 and VS38c, an intrastaining kit was used following the manufacturer's instructions. After 2 washes, FACS analysis was performed on a FACScalibur cytometer (Becton Dickinson) equipped with an air-cooled argon ion laser emitting 15 mW at 488 nm. The instrument was equipped with 3 fluorescence detector photomultiplier tubes, with green fluorescence (FITC) being collected through a 530/30-nm bandpass, red/orange (PE) through a 585/42-nm bandpass, and red (CyC) through a 650-nm longpass filter. Cell analysis was performed with CELLQUEST software (Becton Dickinson). Light scatter signals were recorded in linear mode and fluorescence signals in logarithmic mode. CD38h cells were gated on a CD20/CD38 dot-plot of tonsillar CD31+ cell fraction and on a CD19/CD38 dot-plot of blood CD19+ cell fraction and BMMCs. The third fluorescence was used to explore the phenotype of these PCs for differentiation markers (CD19, CD20, CD22, CD40, CD45, CD138, VS38c, and HLA-DR), survival factors (CD95 and Bcl-2), chemokine receptors CXCR4 and CXCR5, and adhesion molecules (CD9, CD11a, CD21, CD31, CD44, CD49d, CD50, CD54, and CD62L). Data from 2000 to 5000 CD38h cells/sample were collected, and the percentage as well as the mean fluorescence intensity (MFI) of the CD38h cells positive for each analyzed molecule were monitored. The results are expressed as the mean ± SEM of a variety of experiments. Statistical analysis was carried out by using the Student t test. Differences were considered significant when P < .05.FACS separation and PC identification Sorting of CD38h cells was performed on a FACStar-Plus cytometer (Becton Dickinson) equipped with an air-cooled argon ion laser emitting 100 mW at 488 nm. Fluorescence detector photomultiplier tubes and filters were similar to those described above. Tonsil CD31+ and blood CD19+ cells were labeled for CD20 (FITC) and CD38 (PE). CD38h cells were gated with CELLQUEST software and sorted in C-normal mode. To determine the nature of purified cells, sorted CD38h cells (0.1 × 106 cells in 100 µL phosphate-buffered saline) were cytocentrifuged on slides and stained by the Giemsa technique. The percentage of cells with PC morphology was determined by optical microscopy. Cells containing intracytoplasmic Igs were detected on cytospin cell preparations by a direct immunofluorescence technique. Positive cells were identified by fluorescence microscopy. Blood CD38h sorted cells and the remaining non-CD38h cells were also cultured at 5 × 103 cells in 0.2 mL, and secreted IgG-tet was determined in the culture supernatants by an enzyme-linked immunosorbent assay technique, as previously described.37 Tonsil GC and follicular mantle (FM) B-cell fractions were also purified by sorting of tonsil B cells contained in the CD38+ CD20high gate (GC B cells) and CD38 CD20+ gate (FM B cells), as
reported elsewhere.23
Detection of the B-cell transcription factors PRDI-BF1 and BSAP by RT-PCR The presence of transcripts for PRDI-BF1/Blimp-1 and BSAP was investigated in highly purified human B-cell fractions, including blood CD19 cells from nonimmunized healthy individuals, tonsil GC and FM B-cell fractions, and isolated PCs from tonsil, blood, and BM. To this end, total RNA from each cellular fraction was purified using the acid-guanidine-thiocyanate-phenol-chloroform method. After a DNAse I treatment, first-strand complementary DNA (cDNA) copies were synthesized by using Moloney leukemia virus reverse transcriptase (Roche Diagnostics, Barcelona, Spain) with oligo-dT as a primer in a total volume of 100 µL and then PCR was performed. The following oligonucleotide primers were used for PCR: for PRDI-BF1/Blimp-1 sense primer 5'-ATGCGGATATGACTCTGTGGA-3' and antisense primer 5'-CTCGGTTGCTTTAGACTGCTC-3'; for BSAP (sense) 5'-CAGCATAGTGTCCACTGGCT-3' and (antisense) 5'-CCTGTCAGCGTCGGTGCTGA-3'. cDNA (3 µL) was amplified in a PCR reaction (PTC-100 MJ Thermocycler, MJ-Research, Waltham, MA) using each primer and Taq DNA polymerase (Bioline, London, United Kingdom). The cycler conditions for PRDI-BF1/Blimp-1 and BSAP were a denaturing step at 94°C for 1 minute, an annealing step at 65°C for 1 minute, and an amplification step at 72°C for 1 minute, for 35 cycles, followed by a final additional amplification step at 72°C for 7 minutes. The amplified products were analyzed on a 1.2% agarose gel containing ethidium bromide and visualized by UV light illumination. The amounts of -actin cDNA were evaluated, with sense primer
5'-TACCACTGGCATCGTGATGGACT-3' and antisense primer
5'-CGTCACACTTCATGATGGAG-3', as cDNA internal control. The cycler
conditions for -actin were as above, except that the annealing temperature was 62°C and only 30 cycles were performed.
Identification of in vivo-induced IgG-tet-secreting cells present in the human blood as CD38h cells as well as PCs It is well established that, a few days after in vivo immunization to a variety of Ag, specific Ab-forming cells are transiently detected in the human circulation.16-19 This Ab-secreting cell has not yet been clearly identified. The CD38h phenotype is a feature exhibited by human Ab-secreting cells14 and, consequently, the proportion of CD38h cells was monitored in the blood of healthy volunteers before and 6 days after a conventional tet booster, when IgG-tet-secreting cells are maximally found in the circulation.16 Blood samples obtained before the booster showed either undetectable or very low numbers of CD38h cells (< 0.03% of the blood mononuclear cells), whereas in the samples taken after the booster (postboost), a clear increase of this population was detected (0.60% ± 0.1%; mean ± SEM; n = 11). Initial phenotypic analysis of these postboost CD38h cells showed that they were also CD19+. As a consequence, a protocol was designed to isolate blood postboost CD38h cells that included a first pre-enrichment step using immunomagnetic selection of CD19+ cells, followed by a FACS for CD38h cells. Postboost CD19+-selected cells exhibited a marked increase in the proportion of CD38h cells (22.4% ± 9.1%; mean ± SEM; n = 6). FACS selection of CD38h cells in the latter population yielded an extremely enriched population of CD38h cells. Figure 1 (middle panel) shows an example of this isolation method. As can be seen, CD38h cells isolated from the blood of immunized individuals exhibited intense staining for intracytoplasmic Ig and most of them (96.5% ± 1%; mean ± SEM; n = 4) showed a clear PC morphology, as determined by Wright-Giemsa staining of cytocentrifugal preparations. To test the relationship between the CD38h cells isolated from the blood and the tet immunization, purified PCs and the rest of the blood cells (devoid of CD38h cells) were cultured and their capacity to produce IgG-tet was evaluated in the supernatants. IgG-tet secretion was detected only in the supernatant of cultured PCs (12 and 66 ng/mL versus < 0.05 and < 0.05 ng/mL, for cultured purified CD38h cells versus non-CD38h cells, respectively, in 2 different experiments).
Figure 1 also shows the procedure used for the purification of tonsil and BM PCs. The upper panel shows that tonsil PCs could be isolated by using a previously reported technique that included a first pre-enrichment step using immunomagnetic selection of CD31+ cells (from 1.67 ± 0.3 to 14.2 ± 1.4; mean ± SEM; n = 14), followed by FACS of CD38h cells.35 With this method, a high purification of tonsil PCs, as determined by morphologic criteria, was obtained (96.7% ± 0.9%; mean ± SEM; n = 4). Figure 1, lower panel, shows the technique used to isolate BM PCs. The initial proportion of CD38h cells in the BMMC fraction was 0.44% ± 0.1% (mean ± SEM; n = 12). BM CD38h cells also expressed high levels of CD138. After 2 rounds of MACS selection of CD138+ cells, the proportion of PCs as defined by morphologic criteria reached 97.7% ± 0.9% (mean ± SEM; n = 4). Morphologically, the lymphoplasmocytic type of PC showing a denser nucleus and less developed cytoplasm was more frequent in purified PCs from tonsil and blood. In contrast, typical PCs, with chromatin peripherally condensed and prominent arcoplasm, were the major population in purified BM PCs. In the 3 territories, the proportion of PCs contained in the non-CD38h cell populations remained below 0.1%. Comparative analysis of differentiation and survival marker expression by human PCs from different organs Once the correspondence between CD38h phenotype and PC morphology had been demonstrated for the 3 organs under study, the former feature was used for a comparative analysis of the expression of a wide variety of differentiation and survival markers on PCs from tonsil, blood, and BM. To obtain a clear resolution of CD38h/PCs, these experiments were carried out using tonsil CD31+ cells, blood CD19+ cells, and BMMC fractions. A representative example of this comparison is shown in Figure 2 (upper panel). The results representing the percentage of positive PCs and MFI of expression for a series of experiments are summarized in the lower panel of Figure 2 (left and right graphics, respectively). As can be seen, the expression of the B-cell markers CD20 and CD22 was positive in tonsil PCs, but disappeared in blood and BM PCs. CD45 and DR also showed a decreasing pattern of expression, although a clear reduction was observed only in BM PCs. CD19 was expressed by all tonsil and blood PCs, but was negative in one half of the BM PCs. CD40 was exhibited by PCs from the 3 organs, although a slightly reduced level of expression was observed in the blood PCs. VS38c mAb recognizes p63 protein of the endoplasmic reticulum and has been previously described as a good marker for PCs.38 This molecule was similarly expressed by tonsil and blood PCs, but its expression was enhanced by an average of 6 fold in BM PCs. The expression of CD138 (syndecan 1), a molecule clearly associated with the acquisition of the PC stage,26,29 was low in tonsil PCs and intermediate in blood PCs, and showed an increase of up to 8 times in BM PCs. Finally, 2 molecules that regulate cell survival, the antiapoptotic protein bcl-239 and the death receptor CD95,40 were also examined. Tonsil and blood PCs exhibited low but detectable levels of CD95, whereas most BM PCs failed to express CD95, as previously reported.25 In contrast, the expression of bcl-2 was lower for tonsil PCs, intermediate for blood PCs, and higher for BM PCs.
Expression of the transcription factors BSAP and PRDI-BF1/Blimp-1 by PCs isolated from tonsil, blood, and BM The presence of transcripts for BSAP and PRDI-BF1/Blimp-1 transcription factors was investigated by RT-PCR. To this end, tonsil, blood, and BM PCs were highly purified by the method described in Figure 1. As can be seen in Figure 3, isolated peripheral blood B lymphocytes, FM B cells, and GC B cells clearly expressed BSAP but did not contain PRDI-BF1/Blimp-1. In contrast, the 3 PC populations clearly expressed PRDI-BF1/Blimp-1, but only tonsil PCs additionally expressed BSAP. Identical results were obtained in 3 separate experiments.
Comparative analysis of the pattern of expression of adhesion molecules and chemokine receptors by PCs from tonsil, blood, and BM In the last series of experiments, the pattern of adhesion molecule expression exhibited by PCs from the 3 different locations was determined as in Figure 2. A representative example of this comparative study is depicted in the upper panel of Figure 4. Results representing the percentage and MFI of positive PCs in several experiments are summarized in the lower panel of Figure 4 (left and right graphic, respectively). As can be seen, CD11a was expressed, although at low level, by tonsil and blood PCs, and was virtually absent from BM PCs. CD21 was positive in the PCs from the 3 tissues, but its level of expression was higher for tonsil PCs. Very high expression of CD31 was observed on PCs from the 3 territories, although BM PCs showed a 4-fold increase in the expression level of this molecule. Likewise, CD49d, although positive in all PCs, was maximally expressed by the blood and BM populations. A different pattern of expression was found for CD9, CD50, and CD54, which were equally detected in tonsil and BM PCs, but their expression decreased in blood PCs. In contrast, CD62L was absent in tonsil and BM PCs, but it was present on most blood PCs. Finally, the expression of the chemokine receptors CXCR4 and CXCR5 was also explored in human PCs. Table 1 shows that CXCR4 expression was up-regulated in increasing proportions of PCs, according to the axis tonsil bloodBM. Nevertheless, this phenomenon appeared to be partial because only a fraction of the BM PCs
were positive for this receptor. In contrast, only a marginal expression of CXCR5 was observed in PCs from the 3 tissues.
Plasma cells have been generally considered the final and homogenous stage of B-lymphocyte differentiation. However, experimental data accumulated in recent years from both rodents and humans demonstrate that PCs are a complex cell compartment comprising cell subsets differing in several significant respects. In rodent systems, increasing evidence indicates that early PCs formed in inductive areas of secondary lymphoid organs show an enhanced tendency to undergo apoptosis and, as a consequence, are short-lived, whereas BM PCs, which originate from undetermined circulating precursors, exhibit a markedly prolonged life span and improved Ab affinity.5,9-11 Therefore, it is reasonable to think that human PCs might follow a similar maturational sequence. In fact, the differences observed in the capacity to undergo apoptosis and in the kinetics of Ig secretion by human PCs from different tissues16-22,25 also support this view. In this context, the circulating precursor of BM PCs might represent a transitional stage between early PCs and BM PCs. Because the nature of these precursors has not been fully clarified, we hypothesized that human Ab-secreting cells that are transiently present in the blood after immunization16-19 might have reached the PC stage and might be the precursors of BM PCs. Accordingly, the initial aim of the present study was to characterize this cell population. Thus, prior to a conventional tet immunization, the frequency of blood cells exhibiting the CD38h phenotype, a feature ascribed to human PCs in other organs,14 was extremely low, whereas their number increased by at least 20 times in the sample obtained 6 days after the tet booster, when the presence of IgG-tet-secreting cells is maximally detected.16 These tet-induced CD38h cells were isolated using a 2-step protocol that combined immunomagnetic selection of CD19+ cells followed by FACS of CD38h cells. These highly purified cells could be clearly identified as PCs by both morphologic criteria and analysis of intracytoplasmic Ig content. Furthermore, IgG-tet secretion was confined to this PC population, indicating their relationship with the ongoing antitet response in vivo. This is the first time that Ab-secreting cells that transiently circulate after in vivo Ag stimulation have been purified and clearly identified as PCs. The tet-induced increase of blood CD38h cells fell back to prebooster values by day 21 after the immunization (data not shown), a time frame that is concordant with the kinetics of PC appearance in the BM in animal models of humoral response10,12,13 and, accordingly, supports the view that the Ag-induced circulating PCs might be the BM PC precursor. In parallel experiments, PCs were also isolated from a secondary
lymphoid organ such as the tonsil, as well as from a deposit organ such
as the BM, by previously reported procedures.26,36 The
common CD38h feature demonstrated for the PC obtained from
tonsil, blood, and BM allowed an assessment of a variety of molecules
expressed by these cell subsets. Although studies of this kind have
been previously reported for individual organs,24,33-35 a
broad comparative analysis of PCs from these 3 locations has
not been consistently performed before. In addition, the inclusion of
Ag-induced circulating PCs in this analysis made it possible to
identify a putative transitional stage between the early PCs formed in
inductive organs (tonsil) and those PCs that have reached the BM. The
comparison of B-cell differentiation molecules revealed the existence
of important differences among PCs from the 3 tissues under
study. Figure 5 gives a schematic summary
of all these changes. Thus, tonsil PCs were CD19+
CD20low CD22+ CD45+ DR+
VS38c+ CD138
The comparison of the PC expression of molecules involved in the regulation of cell survival also revealed a maturational transition within the PC compartment (Figure 5). Thus, early PCs obtained from tonsil, as well as tet-induced circulating PCs, expressed low but detectable levels of the death receptor CD95, a molecule almost totally absent from the BM PCs. In addition, the expression of the antiapoptotic protein bcl-2 was lower in tonsil PCs, intermediate in blood PCs, and maximal in normal BM PCs. Thus, early PCs obtained from inductive organs appear to express molecules that confer propensity to undergo apoptosis, whereas PCs that home to the BM seem to express a nonapoptotic phenotype. These data are in good agreement with previous reports that demonstrate an increased tendency to develop cell death by early and reactive PCs occurring in secondary lymphoid tissues and in the circulation, but not by BM PCs.24-26 Furthermore, experimental models of humoral responses induced in mice revealed the occurrence of massive cell death by apoptosis in early PCs.5 Therefore, the acquisition of a nonapoptotic phenotype appears to be an important hallmark in PC maturation, and such a state seems to be largely confined to those PCs that reach deposit organs such as the BM. It is of interest that, in Ag-stimulated mice, only a residual number of long-lived PCs remain in the spleen,10,41 and this state has been connected with the existence in the PC vicinity of auxiliary dendritic cells apparently required for PC survival, and whose number is limited in this nondeposit organ.41,42 In this regard, the apoptosis of human tonsil and blood PCs can be delayed by coculturing these cells with BM stromal cells.24,35 Further work will be needed to clarify the significance of extensive cell death by early PCs as well as the selective mechanisms involved in the transition into the nonapoptotic and long-living PC compartment. The transcription factor PRDI-BF1/Blimp-1 is expressed in B lymphocytes at their latest stages of maturation in humans and mice,30-32 and its presence in this phase determines many features typical of the PC differentiation program.30 In contrast, the ectopic expression of this factor before they reach this maturational stage drives apoptotic signals to B lymphocytes.43 The present results demonstrate the presence of apparently similar quantities of mRNA for this factor in human PCs purified from the 3 territories under study, which is in agreement with previous immunohistochemical data.32 Differentiation to PCs has also been connected with the disappearance of BSAP. The relevant role of the down-regulation of BSAP during PC maturation is emphasized by the observation that its enforced expression blocked the transition to PCs.44 Thus, the progression into the PC stage seems to be regulated by the opposing balance of these 2 factors. The observation that purified tonsil PCs still retained detectable quantities of mRNA for BSAP reinforces the notion that early PCs formed in inductive areas of secondary lymphoid tissues are immature and suggests that the complete loss of BSAP might be a requisite for the triggering of the terminal PC maturation program. Collectively, the results strongly indicate the existence of
distinguishable PC stages that predominate in each of the 3 different organs under study, suggesting that the human PC differentiation program requires the transition through successive maturational and
migratory steps. Despite this finding, certain levels of heterogeneity do exist in the 3 PC populations analyzed. For example, BM PCs consist
of CD19+ and CD19 The present study was also focused on the pattern of adhesion molecule
expression exhibited by human PCs. Although this issue has been
explored previously (for reviews, see Helfrich et al45 and
Thomas and Witzing46), a broad comparative analysis of
normal PCs from different territories has not been as yet carried out. Therefore, in an attempt to delineate their distinctive migratory and
homing capacities, the presence of these molecules on human tonsil,
blood, and BM PCs was compared. An initial conclusion is that these
cells largely differed in the type and quantity of adhesion molecules
expressed on their surface (changes summarized in the scheme of Figure
5). Thus, at one extreme, tonsil PCs exhibited the profile
CD11a+ CD21+ CD31+ and
CD49dlow, whereas, at the other extreme, the BM PC profile
was CD11a
The authors thank J. Martorell for his help in morphologic plasma cell identification and E Roldán for fruitful discussion.
Submitted May 24, 2001; accepted November 22, 2001.
Supported by grants 94/0498, 96/2116, and 01/1590 from Fondo de Investigaciones Sanitarias of Spain.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: José A. Brieva, Servicio de Inmunología, Hospital Universitario Puerta del Mar, Avenida Ana de Viya 21, 11009 Cádiz, Spain; e-mail: jabrieva{at}hpm.sas.junta-andalucia.es.
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M. C. Jaimes, O. L. Rojas, E. J. Kunkel, N. H. Lazarus, D. Soler, E. C. Butcher, D. Bass, J. Angel, M. A. Franco, and H. B. Greenberg Maturation and Trafficking Markers on Rotavirus-Specific B Cells during Acute Infection and Convalescence in Children J. Virol., October 15, 2004; 78(20): 10967 - 10976. [Abstract] [Full Text] [PDF]
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G. H. Underhill, K. P. Kolli, and G. S. Kansas Complexity within the plasma cell compartment of mice deficient in both E- and P-selectin: implications for plasma cell differentiation Blood, December 1, 2003; 102(12): 4076 - 4083. [Abstract] [Full Text] [PDF]
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G. Cassese, S. Arce, A. E. Hauser, K. Lehnert, B. Moewes, M. Mostarac, G. Muehlinghaus, M. Szyska, A. Radbruch, and R. A. Manz Plasma Cell Survival Is Mediated by Synergistic Effects of Cytokines and Adhesion-Dependent Signals J. Immunol., August 15, 2003; 171(4): 1684 - 1690. [Abstract] [Full Text] [PDF]
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H. A. M. Wols, G. H. Underhill, G. S. Kansas, and P. L. Witte The Role of Bone Marrow-Derived Stromal Cells in the Maintenance of Plasma Cell Longevity J. Immunol., October 15, 2002; 169(8): 4213 - 4221. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
5) for each marker expressed as the percentage is shown in part B
and the MFI of positive PCs is shown in part C.
