Plasmodium chabaudi chabaudi Infection in Mice Induces Strong B Cell Responses and Striking But Temporary Changes in Splenic Cell Distribution This information is current as of September 25, 2021. Ariel H. Achtman, Mahmood Khan, Ian C. M. MacLennan and Jean Langhorne J Immunol 2003; 171:317-324; ; doi: 10.4049/jimmunol.171.1.317 http://www.jimmunol.org/content/171/1/317 Downloaded from References This article cites 52 articles, 20 of which you can access for free at: http://www.jimmunol.org/content/171/1/317.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication by guest on September 25, 2021 *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2003 by The American Association of Immunologists All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Plasmodium chabaudi chabaudi Infection in Mice Induces Strong B Cell Responses and Striking But Temporary Changes in Splenic Cell Distribution1 Ariel H. Achtman,* Mahmood Khan,† Ian C. M. MacLennan,† and Jean Langhorne2* B cells and Abs play a key role in controlling the erythrocytic stage of malaria. However, little is known about the way the humoral response develops during infection. We show that Plasmodium chabaudi chabaudi causes major, but temporary changes in the distribution of leukocytes in the spleen. Despite these changes, an ordered response to infection develops, which includes vigorous extrafollicular growth of plasmablasts and germinal center formation. Early in the response, the lymphocytes in the T zone and follicles become widely spaced, and the edges of these compartments blur. This effect is maximal around the peak of parasitemia. Germinal centers are apparent by day 8, peak at day 20, and persist through day 60. Extrafollicular foci of plasmablasts are visible Downloaded from from day 4 and initiate a very strong plasma cell response. Initially, the plasma cells have a conventional red pulp distribution, but by day 10 they are unconventionally sited in the periarteriolar region of the white pulp. In this region they form clusters occupying part of the area normally filled by T cells. B cells are absent from the marginal zone for at least 30 days after the peak of infection, although flow cytometry shows their continued presence in the spleen throughout infection. Relatively normal splenic architecture is regained by day 60 of infection. These results show that the changes in splenic cell distribution are linked to the presence of parasites and do not seem to interfere with the development of the humoral response. The Journal of Immunology, http://www.jimmunol.org/ 2003, 171: 317–324. cells and Abs are necessary for eliminating the erythro- still be found up to 1 year after infection with P. chabaudi cytic stages of malaria in most experimental models. chabaudi, but their number is significantly reduced compared with B Mice lacking B cells are unable to clear infections with the response 3 mo after infection (14). Together, these data indi- Plasmodium yoelii and Plasmodium chabaudi chabaudi, although cate that the generation or maintenance of memory B cells and in the latter, the early acute phase is controlled to some extent in long-lived plasma cells could be impaired in malaria infection. the absence of Ab (1–3). A role for the humoral immune response One way of examining B cell memory generation is to assess the in protective immunity in human malaria is inferred from the ex- balance between the follicular and extrafollicular B cell response. by guest on September 25, 2021 perimental models and from immuno-epidemiological studies in T-independent Ab responses, which typically induce extrafollicu- which high Ab titers and in some cases restricted Ig isotypes to lar Ab responses, are not associated with functional germinal cen- particular Ags or regions of Ags correlate with immunity. Transfer ters or memory formation (15). Under exceptional conditions, T of immune serum into naive mice or humans can diminish or pre- cell-independent germinal centers can form but do not complete vent infection in the recipient (4, 5). their development (16). By contrast, T-dependent Ags, which pro- Although Ab appears to be important in the blood stages of duce both extrafollicular responses and germinal centers, are as- human malaria, immunity only develops after repeated infections sociated with affinity maturation and sustained Ab responses (17, and is lost without continued exposure to infection. Longitudinal 18). The differentiation between the follicular and extrafollicular studies in malaria-exposed human populations show that Ab levels pathways is performed by studying the four major B cell popula- to several important malarial proteins drop rapidly at the end of the tions of the spleen: naive recirculating B cells, which are mainly malaria transmission season (6–10). This drop in Ab levels is also located in the follicular mantle of the white pulp; germinal center seen after people have left the transmission area (11). These data B cells as the source of memory B cells and long-lived plasma contrast with Ab responses of noninfected mice to nonrenewable cells; plasma cells and their plasmablast precursors; and marginal protein Ags that can be sustained for years (12, 13). In mice, zone B cells, which include both naive and memory cells and give splenic B cells able to respond to infected erythrocytes in vitro can rise to rapid extrafollicular thymus-dependent and thymus-inde- pendent Ab responses (19–21). In the present study, C57BL/6 mice were infected with P. *Division of Parasitology, National Institute for Medical Research, London, United chabaudi chabaudi (AS), and the resulting changes in the splenic Kingdom; and †Medical Research Council Center for Immune Regulation, University architecture as well as the four major B cell populations of the of Birmingham, Edgbaston, Birmingham, United Kingdom spleen were studied by immunohistology and flow cytometry. We Received for publication December 24, 2002. Accepted for publication April 29, 2003. conclude that there are major structural changes in the spleen, but these reflect an ordered response to the parasite rather than a sal- The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance vage response to indiscriminate damage. with 18 U.S.C. Section 1734 solely to indicate this fact. 1 This work was supported by the Medical Research Council, U.K., and in part by the Materials and Methods European Union, International Cooperation with Developing Countries CT980363. Animals 2 Address correspondence and reprint requests to Dr. Jean Langhorne, Division of Parasitology, National Institute for Medical Research, The Ridgeway, Mill Hill, NW7 Female C57BL/6 mice bred in the specific pathogen-free unit at the Na- 1AA London, U.K. E-mail address: [email protected] tional Institute for Medical Research (London, U.K.) were used at 6–12 wk Copyright © 2003 by The American Association of Immunologists, Inc. 0022-1767/03/$02.00 318 SPLENIC HUMORAL RESPONSES IN MOUSE MALARIA of age. They were conventionally housed on sterile bedding, food, and hydrated in graded alcohols, cleared with Histoclear (National Diagnostics, water. Hull, U.K.), and mounted in DPX (BDH). Parasites and infection Flow cytometry P. chabaudi chabaudi clone AS was routinely injected from frozen stocks. Single-cell suspensions of spleens were prepared, the erythrocytes lysed 5 Further infections were initiated by i.p. injection of 10 parasitized RBCs with 0.16 M NH4Cl and the cells resuspended in FACS buffer (1% weight obtained from infected mice before the peak of parasitemia, and the to volume ratio BSA, 5 mM EDTA, 0.01% sodium azide in PBS, or FACS infection monitored by Giemsa-stained thin blood films as previously buffer without sodium azide for syndecan-1 staining). Cells (5 ϫ 105) were described (22). incubated with appropriately diluted Abs in the presence of anti-Fc recep- tor Ab to prevent Ab binding via the Fc receptor. Samples were acquired ␥ 3 Immunization with chicken -globulin (CGG) on a FACSCalibur and analyzed with CellQuest software (BD Biosciences, CGG (1 mg/ml; The Jackson Laboratory, Bar Harbor, ME) was mixed 1:1 San Jose´, CA). with 9% potassium-aluminum-sulfate and precipitated with 1 M sodium hydroxide. Mice were immunized i.p. with 25 ␮g alum-precipitated Ag in Cell death ␮ 100- l saline solution. 7-Aminoactinomycin D (7-AAD, Sigma-Aldrich) was used to detect apo- ptotic cells, with the necrotic cell marker propidium iodide used in parallel Abs and other reagents for immunohistology and flow cytometry ϩ to ascertain that the majority of the 7-AAD cells were actually apoptotic. The following Abs and reagents were used in immunohistology: rat anti- Splenocytes were stained as described in the previous section. In the final mouse CD3 (KT3) and rat anti-mouse IgM H chain (LO-MM-9) (Serotec, step, either 7-AAD (1 ␮g/2.5 ϫ 105 cells) or propidium iodide (1 ␮g/2.5 ϫ Oxford, U.K.); rat anti-mouse syndecan-1 (CD138, 09341D; BD Phar- 105 cells; Sigma-Aldrich) was added for 20 or 5 min, respectively. Cells Mingen, Oxford, U.K.); biotinylated peanut agglutinin (PNA; Vector Lab- were then analyzed on the FACSCalibur without further washes.