Heat-Killed Saccharomyces cerevisiae, A Dectin-1 Agonist, Selectively Induces IgG4 Production by Human B Cells

Dectin-1 is a major receptor that recognizes fungal cell wall β-glucan. We previously reported that heat-killed Saccharomyces cerevisiae (HKSC), a Dectin-1 agonist, selectively induces IgG1 class switching in mouse B cells. Dectin-1 is also expressed on human B cells; however, Dectin-1 function in human B cells remains unknown. This study aimed to investigate the direct effect of in vitro stimulation using HKSC on Ig class switching in human B cells. HKSC selectively induced the expression of germline γ4 transcripts (GLTγ4) by human B cell line 2E2, and HKSC significantly augmented GLTγ4 promoter activity. Moreover, HKSC selectively enhanced GLTγ4 expression and IgG4 production by anti-CD40-activated human tonsillar resting B cells. Thus, these results suggest that Dectin-1 maybe involved in selective IgG4 class switching by human B cells.

Toll-like Receptor 1/2 Agonist Pam3CSK4 Suppresses Lipopolysaccharide-driven IgG1 Production while Enhancing IgG2a Production by B Cells

Interaction between pathogen-associated molecular patterns and pattern recognition receptors triggers innate and adaptive immune responses. Several studies have reported that toll-like receptors (TLRs) are involved in B cell proliferation, differentiation, and Ig class switch recombination (CSR). However, roles of TLRs in B cell activation and differentiation are not completely understood. In this study, we investigated the direct effect of stimulation of TLR1/2 agonist Pam3CSK4 on mouse B cell viability, proliferation, activation, Ig production, and Ig CSR in vitro. Treatment with 0.5 µg/ml of Pam3CSK4 only barely induced IgG1 production although it enhanced B cell viability. In addition, high-dosage Pam3CSK4 diminished IgG1 production in a dose-dependent manner, whereas the production of other Igs, cell viability, and proliferation increased. Pam3CSK4 additively increased TLR4 agonist lipopolysaccharide (LPS)-induced mouse B cell growth and activation. However, interestingly, Pam3CSK4 abrogated LPS-induced IgG1 production but enhanced LPS-induced IgG2a production. Further, Pam3CSK4 decreased LPS-induced germline γ1 transcripts (GLTγ1)/GLTε expression but increased GLTγ2a expression. On the other hand, Pam3CSK4 had no effect on LPS-induced plasma cell differentiation. Taken together, these results suggest that TLR1/2 agonist Pam3CSK4 acts as a potent mouse B cell mitogen in combination with TLR4 agonist LPS, but these 2 different TLR agonists play diverse roles in regulating the Ig CSR of each isotype, particularly IgG1/IgE and IgG2a.

Murine γδ T Cells Render B Cells Refractory to Commitment of IgA Isotype Switching

γδ T cells are abundant in the gut mucosa and play an important role in adaptive immunity as well as innate immunity. Although γδ T cells are supposed to be associated with the enhancement of Ab production, the status of γδ T cells, particularly in the synthesis of IgA isotype, remains unclear. We compared Ig expression in T cell receptor delta chain deficient (TCRδ⁻/⁻) mice with wild-type mice. The amount of IgA in fecal pellets was substantially elevated in TCRδ⁻/⁻ mice. This was paralleled by an increase in surface IgA expression and total IgA production by Peyer's patches (PPs) and mesenteric lymph node (MLN) cells. Likewise, the TCRδ⁻/⁻ mice produced much higher levels of serum IgA isotype. Here, surface IgA expression and number of IgA secreting cells were also elevated in the culture of spleen and bone marrow (BM) B cells. Germ-line α transcript, an indicator of IgA class switch recombination, higher in PP and MLN B cells from TCRδ⁻/⁻ mice, while it was not seen in inactivated B cells. Nevertheless, the frequency of IgA+ B cells was much higher in the spleen from TCRδ⁻/⁻ mice. These results suggest that γδ T cells control the early phase of B cells, in order to prevent unnecessary IgA isotype switching. Furthermore, this regulatory role of γδ T cells had lasting effects on the long-lived IgA-producing plasma cells in the BM.

Cyberbullying and adolescent mental health: systematic review / Cyberbullying e saúde mental dos adolescentes: revisão sistemática / Acoso cibernético y la salud mental de los adolescentes: revisión sistemática

Cyberbullying is a new form of violence that is expressed through electronic media and has given rise to concern for parents, educators and researchers. In this paper, an association between cyberbullying and adolescent mental health will be assessed through a systematic review of two databases: PubMed and Virtual Health Library (BVS). The prevalence of cyberbullying ranged from 6.5% to 35.4%. Previous or current experiences of traditional bullying were associated with victims and perpetrators of cyberbullying. Daily use of three or more hours of Internet, web camera, text messages, posting personal information and harassing others online were associated with cyberbullying. Cybervictims and cyberbullies had more emotional and psychosomatic problems, social difficulties and did not feel safe and cared for in school. Cyberbullying was associated with moderate to severe depressive symptoms, substance use, ideation and suicide attempts. Health professionals should be aware of the violent nature of interactions occurring in the virtual environment and its harm to the mental health of adolescents.

Cyberbullying, uma nova forma de violência expressa por meio da mídia eletrônica, tem preocupado pais, educadores e pesquisadores. A associação entre cyberbullying e a saúde mental dos adolescentes será revisada. Revisão sistemática em duas bases de dados: PubMed e a Biblioteca Virtual em Saúde (BVS). A prevalência do cyberbullying variou entre 6,5% a 35,4%. Bullying tradicional prévio ou atual estava associado às vítimas e agressores do cyberbullying. Uso diário de três ou mais horas de Internet, web câmera, mensagens de texto, postar informações pessoais e assediar outros online estavam associados ao cyberbullying. "Cybervítimas" e cyberbullies tinham mais problemas emocionais, psicossomáticos, dificuldades sociais, e não se sentiam seguros e cuidados na escola. O cyberbullying estava associado à sintomatologia depressiva moderada e grave, uso de substâncias, ideação e tentativas de suicídio. Profissionais de saúde devem conhecer as interações de natureza violenta que ocorrem no ambiente virtual e de seus agravos para a saúde mental dos adolescentes.

Se revisa la asociación entre el acoso cibernético y la salud mental de los adolescentes. Se realiza una revisión sistemática de dos bases de datos: PubMed y la Biblioteca Virtual en Salud (BVS). La prevalencia de ciberacoso varió de un 6,5% a un 35,4%. Los acosos cibernéticos tradicionales -pasados o actuales- se asociaron con las víctimas y los acosadores cibernéticos. El uso diario de tres o más horas de Internet, cámara web, mensajes de texto, la publicación de información personal y acosar a los demás se asociaron con el acoso cibernético. Cibervíctimas y acosadores cibernéticos tenían más problemas emocionales, psicosomáticos, dificultades sociales y no se sentían seguros y cuidados en la escuela. El ciberacoso se asoció con síntomas de moderados a graves de depresión, abuso de sustancias, ideación suicida e intentos de suicidio. Los profesionales de salud deben conocer la naturaleza violenta de las interacciones que se producen en el entorno virtual y sus peligros para la salud mental de los adolescentes.

Lactoferrin Combined with Retinoic Acid Stimulates B1 Cells to Express IgA Isotype and Gut-homing Molecules

It is well established that TGF-beta1 and retinoic acid (RA) cause IgA isotype switching in mice. We recently found that lactoferrin (LF) also has an activity of IgA isotype switching in spleen B cells. The present study explored the effect of LF on the Ig production by mouse peritoneal B cells. LF, like TGF-beta1, substantially increased IgA production in peritoneal B1 cells but little in peritoneal B2 cells. In contrast, LF increased IgG2b production in peritoneal B2 cells much more strongly than in peritoneal B1 cells. LF in combination with RA further enhanced the IgA production and, interestingly, this enhancement was restricted to IgA isotype and B1 cells. Similarly, the combination of the two molecules also led to expression of gut homing molecules alpha4beta7 and CCR9 on peritoneal B1 cells, but not on peritoneal B2 cells. Thus, these results indicate that LF and RA can contribute to gut IgA response through stimulating IgA isotype switching and expression of gut-homing molecules in peritoneal B1 cells.

SUMO Proteins are not Involved in TGF-beta1-induced, Smad3/4-mediated Germline alpha Transcription, but PIASy Suppresses it in CH12F3-2A B Cells

TGF-beta induces IgA class switching by B cells. We previously reported that Smad3 and Smad4, pivotal TGF-beta signal-transducing transcription factors, mediate germline (GL) alpha transcription induced by TGF-beta1, resulting in IgA switching by mouse B cells. Post-translational sumoylation of Smad3 and Smad4 regulates TGF-beta-induced transcriptional activation in certain cell types. In the present study, we investigated the effect of sumoylation on TGF-beta1-induced, Smad3/4-mediated GLalpha transcription and IgA switching by mouse B cell line, CH12F3-2A. Overexpression of small ubiquitin-like modifier (SUMO)-1, SUMO-2 or SUMO-3 did not affect TGF-beta1-induced, Smad3/4-mediated GLalpha promoter activity, expression of endogenous GLalpha transcripts, surface IgA expression, and IgA production. Next, we tested the effect of the E3 ligase PIASy on TGF-beta1-induced, Smad3/4-mediated GLalpha promoter activity. We found that PIASy overexpression suppresses the GLalpha promoter activity in cooperation with histone deacetylase 1. Taken together, these results suggest that SUMO itself does not affect regulation of GLalpha transcription and IgA switching induced by TGF-beta1/Smad3/4, while PIASy acts as a repressor.

The Niche of Follicular Helper T Cells in Systemic Autoimmune Diseases / 한양의대학술지

Production of thymus-dependent antibodies by autoreactive B cells requires help from T cells. Follicular helper T (Tfh) cells are a unique lineage of CD4+ T subsets present in the follicles of peripheral lymphoid tissues which functions primarily to provide help to cognate B cells. Within germinal centers Tfh cells stimulate germinal center B cells to undergo affinity maturation, Ig class switching, and differentiation to memory B cells and plasma cells. Proposals that activity of Tfh cells is crucial for long-lived humoral autoimmunity are supported by the correlation of numbers and/or functions of Tfh cells with disease activity in many autoimmune disorders. In this review, we discuss recent findings regarding Tfh cell development and function. In addition, we discuss putative roles of Tfh cells in the pathogenesis and highlight the potential of Tfh cells as therapeutic targets in autoimmune diseases.

Germinal Center Formation Controlled by Balancing Between Follicular Helper T Cells and Follicular Regulatory T Cells / 한양의대학술지

Follicular helper T cells (Tfh) play a significant role in providing T cell help to B cells during the germinal center reaction, where somatic hypermutation, affinity maturation, isotype class switching, and the differentiation of memory B cells and long-lived plasma cells occur. Antigen-specific T cells with IL-6 and IL-21 upregulate CXCR5, which is required for the migration of T cells into B cell follicles, where these T cells mature into Tfh. The surface markers including PD-1, ICOS, and CD40L play a significant role in providing T cell help to B cells. The upregulation of transcription factor Bcl-6 induces the expression of CXCR5, which is an important factor for Tfh differentiation, by inhibiting the expression of other lineage-specific transcription factors such as T-bet, GATA3, and RORgammat. Surprisingly, recent evidence suggests that CD4 T cells already committed to Th1, Th2, and Th17 cells obtain flexibility in their differentiation programs by downregulating T-bet, GATA3, and RORgammat, upregulating Bcl-6 and thus convert into Tfh. Limiting the numbers of Tfh within germinal centers is important in the regulation of the autoantibody production that is central to autoimmune diseases. Recently, it was revealed that the germinal center reaction and the size of the Tfh population are also regulated by thymus-derived follicular regulatory T cells (Tfr) expressing CXCR5 and Foxp3. Dysregulation of Tfh appears to be a pathogenic cause of autoimmune disease suggesting that tight regulation of Tfh and germinal center reaction by Tfr is essential for maintaining immune tolerance. Therefore, the balance between Tfh and Tfr appears to be a critical peripheral tolerance mechanism that can inhibit autoimmune disorders.

Alum Directly Modulates Murine B Lymphocytes to Produce IgG1 Isotype

Aluminum hydroxide (alum) is the most widely used adjuvant in human vaccines. Nevertheless, it is virtually unknown whether alum acts on B cells. In the present study, we explored the direct effect of alum on Ig expression by murine B cells in vitro. LPS-activated mouse spleen B cells were cultured with alum, and the level of isotype-specific Ig secretion, IgG1 secreting cell numbers, and Ig germ-line transcripts (GLT) were measured using ELISA, ELISPOT, and RT-PCR, respectively. Alum consistently enhanced total IgG1 production, numbers of IgG1 secreting cells, and GLTgamma1 expression. These results demonstrate that alum can directly cause IgG1 isotype switching leading to IgG1 production.

Newly Identified TLR9 Stimulant, M6-395 Is a Potent Polyclonal Activator for Murine B Cells

BACKGROUND: Toll-like receptors (TLRs) have been extensively studied in recent years. However, functions of these molecules in murine B cell biology are largely unknown. A TLR4 stimulant, LPS is well known as a powerful polyclonal activator for murine B cells. METHODS: In this study, we explored the effect of a murine TLR9 stimulant, M6-395 (a synthetic CpG ODNs) on B cell proliferation and Ig production. RESULTS: First, M6-395 was much more potent than LPS in augmenting B cell proliferation. As for Ig expression, M6-395 facilitated the expression of both TGF-beta1-induced germ line transcript alpha (GLTalpha) and IL-4-induced GLTgamma1 as levels as those by LPS and Pam3CSK4 (TLR1/2 agonist) : a certain Ig GLT expression is regarded as an indicative of the corresponding isotype switching recombination. However, IgA and IgG1 secretion patterns were quite different--these Ig isotype secretions by M6-395 were much less than those by LPS and Pam3CSK4. Moreover, the increase of IgA and IgG1 production by LPS and Pam3CSK4 was virtually abrogated by M6-395. The same was true for the secretion of IgG3. We found that this unexpected phenomena provoked by M6-395 is attributed, at least in part, to its excessive mitogenic nature. CONCLUSION: Taken together, these results suggest that M6-395 can act as a murine polyclonal activator but its strong mitogenic activity is unfavorable to Ig isotype switching.

Immunoglobulin Isotype Switching in a Plasma Cell Myeloma Patient Treated with High-dose Chemotherapy and Hematopoietic Stem Cell Transplantation

A malignant plasma cell clone usually produces a single abnormal monoclonal protein with a constant isotype. However, switching of paraprotein isotype has been reported to be a transient phenomenon associated with the recovery of B-cell function, and, in some cases, the switching might be misinterpreted as relapse. In August 2008, we encountered a case of a 59-year-old man with proteinuria and high IgG level (5.6 g/dL), kappa free light chain level of 2,660 mg/L, reversed A/G ratio (0.51), and multiple osteolytic lesions. Plasma cells, which accounted for 57% of all the nucleated cells, in bone marrow aspirates were positive for kappa immunostaining. Serum protein electrophoresis showed one M-spike, concentration of 4.87 g/dL in the beta region. Immunofixation electrophoresis revealed the peak as an IgG-kappa monoclonal protein; therefore, a diagnosis of plasma cell myeloma was made. Complete remission was achieved after chemotherapy, and autologous peripheral stem cell collection was performed. In March 2009, the patient underwent high-dose chemotherapy and autologous peripheral stem cell transplantation support. After 2 months, serum protein electrophoresis showed 2 M-spikes in the gamma region with positive IgM-lambda, IgG-lambda, and IgG-kappa, and these bands persisted. The electrophoretic mobility of the IgG-kappa protein was different from that of the original disease protein, and bone marrow results were the same as the previous ones. Although immunoglobulin isotype switch is known to have a benign course, it always requires careful monitoring because, in rare cases, true clonal switching may occur.

Vitamin C acts indirectly to modulate isotype switching in mouse B cells / 대한해부학회지

Vitamin C, one of essential micronutrients, has been reported to modulate the humoral immune responses in some mammals. We investigated whether vitamin C might modulate this response in mice by directly affecting B cells. Splenic B cells were isolated and activated by CD40- and B cell receptor-ligation in vitro. The cells were cultured with a pretreatment of vitamin C from 0 to 1 mM of concentrations. Vitamin C slightly increased apoptosis of B cells dose-dependently and behaved as an antioxidant. We found that in vivo administration of vitamin C by intraperitoneal injection affected isotype switching as previously reported: the titer of antigen-specific IgG1 antibody was decreased, while that of IgG2a was unaffected. Somewhat different from those observed in vivo, in vitro exposure to vitamin C slightly decreased isotype switching to IgG1 and increased isotype switching to IgG2a. Pretreatment with vitamin C in the safe range did not affect either proliferation of cultured B cells or the expression of CD80 and CD86 in those cells. Taken together, in vivo results suggest that vitamin C acts to modulate isotype switching in the mouse. However, because of our in vitro results, we suggest that the modulation exerted by vitamin C in vivo is by indirectly affecting B cells, perhaps by directly influencing other immune cells such as dendritic cells.

Expression of IL-21 in the peripheral blood of myasthenia gravis patients and its correlation with anti-AChR-Ab class switch / 中南大学学报(医学版)

OBJECTIVE@#To explore the role of IL-21 in the pathogenesis of myasthenia gravis (MG) and its influence on the the class switch of anti-AChR antibodies.@*METHODS@#Blood was taken from 26 patients and 18 healthy controls, and the expression of IL-21R mRNA in peripheral blood mononuclear cells (PBMCs) was detected by RT-PCR. The expression of IL-21R on B lymphocytes was measured by flow cytometry, while the concentrations of serum IL-21 and the levels of anti-AChR-IgG and its isotype IgG(1), IgG(2), and IgG(3) were tested by ELISA.@*RESULTS@#The serum concentration of IL-21 in the MG group was higher than that in the control group (31.686±8.499 pg/mL, 15.147±6.366 pg/mL) and the difference was significant (P0.05). Compared with the control group, the expression of IL-21R on B lymphocytes also increased in the MG group (P0.05). Expression of IL-21R mRNA in the PBMCs showed no correlation with the level of serum anti-AChR-IgG and its isotype IgG(1), IgG(2), and IgG(3), respectively(P>0.05); however the expression of IL-21R in B lymphocytes showed positive correlation with anti-AChR-IgG and it's isotype IgG(1) and IgG(3) (P0.05).@*CONCLUSION@#IL-21 might induce the class switch of anti-AChR antibodies to IgG(1) and IgG(3) isotype through IL-21R on B lymphocytes which promotes the pathogenesis of the MG.

Immunoglobulin variable heavy chain region genetic constitution and mutation status in Chinese patients with chronic lymphocytic leukemia / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To evaluate the genetic constitution and mutation status of the immunoglobulin variable heavy chain region (IGVH) gene expression in Chinese patients with chronic lymphocytic leukemia (CLL).</p><p><b>METHODS</b>The IGVH mutation was detected by multiplex PCR and direct sequencing of the purified PCR products from 64 CLL patients. The segments of VH, DH and JH family and mutations were analyzed by IMGT/V-QUEST and IGBlast.</p><p><b>RESULTS</b>In the 64 patients, the most common usage was VH3 (31/64, 48%), followed by VH4 (26/64, 41%), VH1 (4/64, 6%), VH2 (2/64, 3%) and VH7 (1/64, 2%). The results also showed that 44 patients (69%) had mutated VH, 6 cases (9%) had identical germline sequences. Among the 64 sequences of DH segments, DH3 gene family was used most frequently (25/64, 39%), among which 11 cases had unmutated VH. The most frequent usage of the JH segments was JH6.</p><p><b>CONCLUSION</b>There is significant difference in the frequency of the IGVH gene family in Chinese CLL patients compared to Western patients, suggesting the involvement of antigen selection in different ethnic and/or environmental factors in CLL disease initiation, and its prognostic significance needs further investigation.</p>

Tiul1 and TGIF are Involved in Downregulation of TGFbeta1-induced IgA Isotype Expression

TGF-beta1 is well known to induce Ig germ-line alpha (GLalpha) transcription and subsequent IgA isotype class switching recombination (CSR). Homeodomain protein TG-interacting factor (TGIF) and E3-ubiquitin ligases TGIF interacting ubiquitin ligase 1 (Tiul1) are implicated in the negative regulation of TGF-beta signaling. In the present study, we investigated the roles of Tiul1 and TGIF in TGFbeta1-induced IgA CSR. We found that over-expression of Tiul1 decreased TGFbeta1-induced GLalpha promoter activity and strengthened the inhibitory effect of Smad7 on the promoter activity. Likewise, overexpression of TGIF also diminished GLalpha promoter activity and further strengthened the inhibitory effect of Tiul1, suggesting that Tiul1 and TGIF can down-regulate TGFbeta1-induced GLalpha expression. In parallel, overexpression of Tiul1 decreased the expression of endogenous IgA CSR-predicitive transcripts (GLT(alpha), PST(alpha), and CT(alpha)) and TGFbeta1-induced IgA secretion, but not GLT(gamma3) and IgG3 secretion. Here, over-expressed TGIF further strengthened the inhibitory effect of Tiul1. These results suggest that Tiul1 and TGIF act as negatively regulators in TGFbeta1-induced IgA isotype expression.

Onychomadesis in a patient with immunoglobulin class switch recombination deficiency

Immunoglobulin class switch recombination deficiencies [Ig CSR deficiencies] or Hyper IgM syndromes [HIGM] are a group of primary immunodeficiency diseases, characterized by defective CD40 signaling of B cells resulting into a CSR and a somatic hypermutation. The affected patients are characterized with reduced serum levels of IgG and IgA, and normal or elevated level of IgM, which lead to increased susceptibility to infections. We describe a 3 year-old boy with frequent bacterial infections of the skin and respiratory tract, mucosal ulcers, and diarrhea. He experienced onychomadesis in both fingernails and toenails during recent bacterial infection. Quantitative immunoglobulin levels revealed high levels of serum IgM and very low levels of IgG, IgA, and IgE. Clinical and immunologic studies supported the diagnosis of HIGM. Onychomadesis as a finding in HIGM could be considered. Considering exclusion of CD40L, CD40, AID and UNG genes by molecular analysis, new CSR selective deficiencies could be suspected in this case

Lack of correlation between rabies virus replication in the brain and antibody isotype profile in genetically modified mice

The relationship among the phenotypes resistance to infection, virus replication in the brain and isotype production was investigated in genetically modified High (H) or Low (L) antibody responder mouse lines. Although they express the same innate susceptibility to rabies infection, these lines differ as to different viral replication rates in the central nervous system and L mice showed a higher permissible state. After intramuscular infection with the Pasteur rabies strain (PV), the H-L interline differences on the earlier stage of virus replication were 1000 and 80 folds on days 5 and 6, respectively. The isotype profile in sera of the experimentally infected mice reflected an interline difference of 25 folds for IgG2a throughout the infection period, and for the IgE production the H-L difference was highly significant only at the beginning of the process. These results confirm the multi-specific effect of antibody immune responsiveness and the general isotype distribution of antibodies in these genetically selected mice. Contrary to the clear correlation between antibody responsiveness and the acquired resistance to rabies infection, the present study demonstrates that the constitutive genetic character of High and Low responder individuals does not intervene in the degree of resistance following infection. Altogether, this study contributes to the knowledge of the protective role of the general innate responsiveness on the pathological pattern to rabies virus infection

A Study of Susceptibility between Allergic Rhinitis and IL-13 Exon 4 G2044A Gene Polymorphism Study in Korean / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: High total serum IgE level is one of the characteristics seen in allergic rhinitis. IL-13 provides impetus to immunoglobulin class switching to IgE. The IL-13 promoter single nucleotide polymorphism has been shown to be associated with allergic diseases and abnormal IL-13 production. We tested whether a polymorphism in the coding region of IL-13 gene is associated with allergic rhinitis, blood eosinophil counts and total serum IgE levels in Korean. SUBJECTS AND METHOD: Blood samples for genetic analysis were obtained from 307 individuals with allergic rhinitis and from 268 healthy subjects without atopic diseases. Polymerase chain reaction-based assay for IL-13 exon 4 G2044A was used for genotyping. Serum total IgE levels were determined by using the immunoassay. Eosinophil values were determined by eosinophil numbers per total cell numbers per microl. RESULTS: There were no differences in the frequencies of the genotypes of IL-13 in the controls and patients (p>0.05). The frequencies of the IL-13 exon 4 2044A allele were statistically different between controls and patients (p0.05). CONCLUSION: Our result suggests that the IL-13 exon 4 G2044A polymorphism might give susceptibility to the development of allergic rhinitis in Koreans.

Cambio del isotipo en la respuesta a antígenos T independientes por co-inoculación con antígenos T dependientes / Isotype switch in the response against T independent antigens by co-inoculation with T dependent antigens

Un paradigma clásico de la inmulogía plantea que para que ocurra cambio de isotipo en los anticuerpos es condición sine qua non la presentación del antígeno a un linfócito T colaborador por parte de una célula presentadora de antígenos. En el presente trabajo se diseñó un modelo animal, ratones BALB/c, de respuesta inmune frente a dos antígenos típicos. Se utilizo dextrán como antígeno T independiente (AgTI) y seroalbúmina bovina (SAB) como antígeno T dependiente (AgTD), y se estúdio la respuesta, analizando los isotipos de los anticuerpos específicos producidos. Los resultados obtenidos muestran que la respuesta a dextrán en presencia de SAB ocurre con cambio de isotipo (swith), essencialmente de IgM a IgG. Estos experimentos sugieren que la SAB genera un entorno bioquímico inductor de cambio de isotipo tanto en supropia via de procesamiento como en del dextrán. Los resultados señalan que la asociación exclusiva de los AgTDs con las respuestas em las que ocurre cambio de isotipo es incorrecta. Considerando el modelo propuesto resulta poco probable encontrar in vivo y en forma espontânea casos en los que los AgTIs ingreses al organismo aislados; en cambio, es mucho más probable que el ingreso ocurra conjuntamente con AgTDs, y en consecuencia ocurra cambio de isotipo.

Characterization of Mouse B Lymphoma Cells (CH12F3-2A) for the Study of IgA Isotype Switching

BACKGROUND: It is well known that IgA isotype switching is induced by TGF-beta1. LPS-activated mouse normal B cells well differentiate into IgA secreting plasma cells under the influence of TGF-beta1. Nevertheless, there are lots of difficulties in studying normal B cells in detail because it is not simple to obtain highly purified B cells, showing low reproducibility and transfection efficacy, moreover impossible to keep continuous culture. To overcome these obstacles, it is desperately needed to develop B cell line which acts like normal B cells. In the present study, we investigated whether CH12F3-2A lymphoma cells are appropriate for studying IgA isotype switching event. METHODS: CH12F3-2A B cell line was treated with LPS and TGF-beta1, then levels of germ-line (GL) transcripts were measured by RT-PCR, and GLalpha promoter activity was measured by luciferase assay. In addition, membrane IgA (mIgA) expression and IgA secretion were determined by FACS and ELISA, respectively. RESULTS: TGF-beta1, regardless of the presence of LPS, increased level of GLalpha transcripts but not GLgamma2b transcripts. However, IgA secretion was increased dramatically by co-stimulation of LPS and TGF-beta1. Both mIgA and IgA secretion in the presence of TGF-beta1 were further increased by over-expression of Smad3/4. Finally, GLalpha promoter activity was increased by TGF-beta1. CONCLUSION: CH12F3-2A cell line acts quite similarly to the normal B cells which have been previously reported regarding IgA expression. Thus, CH12F3-2A lymphoma cell line appears to be adequate for the investigation of the mechanism(s) of IgA isotype switching at the cellular and molecular levels.