Differential effects of vasoactive intestinal peptide, substance P, and somatostatin on immunoglobulin synthesis and proliferations by lymphocytes from Peyer's patches, mesenteric lymph nodes, and spleen

We examined the effect of vasoactive intestinal peptide, substance P, and somatostatin on concanavalin A (1 microgram/ml)-induced lymphocyte proliferation and immunoglobulin (IgA, IgM, and IgG) synthesis by cells from spleens, Peyer's patches, and mesenteric lymph nodes. These neuropeptides (10(-7) to 10(-12) M) modulated immune responses in a dose-dependent manner. For a comparative study, neuropeptides were used at 10(-8) M concentration. Both vasoactive intestinal peptide and somatostatin significantly decreased DNA synthesis (30 to 50%), whereas substance P increased synthesis (40%) in lymphocytes from all organs tested. IgA synthesis was significantly altered by all of the neuropeptides tested, whereas IgM synthesis was less affected and IgG synthesis was virtually unchanged. Somatostatin inhibited IgA (20 to 50%) and IgM (10 to 30%) synthesis in lymphocytes from all three organs. Substance P increased IgA synthesis in mesenteric lymph nodes (50%), spleens (70%), and Peyer's patches (300%). It also increased IgM synthesis in Peyer's patches (20%) and spleens (30%), but was without effect on IgM synthesis in mesenteric lymph nodes. Vasoactive intestinal peptide increased the IgA response in mesenteric lymph nodes (20%) and spleens (30%), but inhibited IgA synthesis in lymphocytes from Peyer's patches (60%). Interestingly, in Peyer's patches, IgM synthesis was increased by vasoactive intestinal peptide (80%), whereas it was unchanged in mesenteric lymph nodes and spleen. Thus, not only did these neuropeptides have different effects on the production of different immunoglobulin isotypes, but their effect was also organ-specific. Because neuropeptides which are abundant in the intestine can modulate IgA and other immunoglobulin synthesis in vitro, they may play a significant regulatory role in mucosal immune responses in vivo.     

Analysis of T cell and B cell function in Peyer's patch and lamina propria of New Zealand Black and DBA/2 mice

We have analyzed gastrointestinal immune function in both DBA/2 and spontaneously autoimmune New Zealand Black (NZB) mice. We have studied both in vitro proliferation and differentiation of Peyer's patch cells and have measured immunoglobulin (Ig) secretion by cultured jejunal segments. Peyer's patch B cells and T cells from both DBA/2 and NZB mice showed similar proliferative responses to Con A and lipopolysaccharide (LPS), respectively. Unlike NZB splenic B cells, isolated Peyer's patch B cells from NZB mice did not spontaneously secrete Ig of any isotype. Seven-day cultures of equal numbers of Peyer's patch T cells and B cells resulted in similar patterns of secretion of IgA, IgG, and IgM in both strains. The addition of Con A to cultures of DBA/2 Peyer's patch cells consistently resulted in a onefold to threefold increase in IgA secretion after 7 days. Con A stimulation of NZB Peyer's patch cells did not produce any increment in IgA secretion. LPS stimulation of Peyer's patch cells from either strain resulted in a similar increase in IgG secretion with little effect on IgA secretion. The in vivo correlate of this finding was seen in the IgA to IgG ratio of Ig secreted by cultured jejunal fragments. In DBA/2 mice the rates of IgA/IgG varied from 2.36 to 4.85, whereas in NZB mice the ratio never exceeded 0.5. These experiments show that defects on the T cell compartment of NZB mice encompass gut-associated lymphoid tissue. The possible relationship of these findings and previously observed defects in oral tolerance is discussed.     

Encapsulated Bifidobacterium bifidum potentiates intestinal IgA production

We asked whether Bifidobacterium bifidum regulates the synthesis of IgA by mucosal lymphoid cells. B. bifidum alone, but not Clostridium perfringens, significantly induced total IgA and IgM synthesis by both mesenteric lymph nodes (MLN) and Peyer's patch (PP) cells. We, further, investigated the mucosal antibody production following peroral administration of B. bifidum to mice. Ingested B. bifidum significantly increased the number of Ig (IgM, IgG, and IgA) secreting cells in the culture of both MLN and spleen cells. Nonetheless, B. bifidum itself does not induce the own specific antibody responses, implying that B. bifidum does not provoke unnecessary immune reaction. Subsequently, it was found that encapsulation of B. bifidum further augments the total IgA production in the culture of both MLN and spleen cells. Finally, we found that the immuno-stimulating activity of B. bifidum is due to its cellular components but not due to any actively secreting component(s) from bacteria.     

Immunomodulatory activity of Bengkoang (Pachyrhizus erosus) fiber extract in vitro and in vivo

Bengkoang (Pachyrhizus erosus (L.) Urban) is one of the most popular edible root vegetables in Indonesia. Bengkoang contains fairly large amounts of carbohydrates and crude fiber. The purpose of this research is to evaluate the immunomodulatory effect of the bengkoang fiber extract (BFE) in vitro and in vivo. BFE was prepared by heating the powder of bengkoang fiber suspended in distilled water at 121 °C for 20 min. BFE facilitated IgM production by the human hybridoma cell line HB4C5 cells. In addition, production of IgM, IgG, and IgA by mouse primary splenocytes was facilitated by BFE in a dose-dependent manner. BFE also significantly facilitated production of both interleukin-5 and interleukin-10 by splenocytes. Immunoglobulin production by lymphocytes from the spleen, Peyer’s patch, and mesenteric lymph node were significantly activated by oral administration of BFE to mice for 14 days. The serum immunoglobulin levels of IgG, IgM, and IgA were also significantly enhanced. Furthermore, cytokine production by lymphocytes from the spleen, Peyer’s patch, and mesenteric lymph node were also facilitated by oral administration of BFE. These results suggest that BFE has positive effects on the immune system in vitro and in vivo.     

Selective adherence of IgA to murine Peyer's patch M cells: evidence for a novel IgA receptor

M cells represent the primary route by which mucosal Ags are transported across the intestinal epithelium and delivered to underlying gut-associated lymphoid tissues. In rodents and rabbits, Peyer's patch M cells selectively bind and endocytose secretory IgA (SIgA) Abs. Neither the nature of the M cell IgR nor the domains of SIgA involved in this interaction are known. Using a mouse ligated ileal loop assay, we found that monoclonal IgA Abs with or without secretory component, but not IgG or IgM Abs, bound to the apical surfaces of Peyer's patch M cells, indicating that the receptor is specific for the IgA isotype. Human serum IgA and colostral SIgA also bound to mouse M cells. The asialoglycoprotein receptor or other lectin-like receptors were not detected on the apical surfaces of M cells. We used recombinant human IgA1 and human IgA2 Abs and domain swapped IgA/IgG chimeras to determine that both domains Calpha1 and Calpha2 are required for IgA adherence to mouse Peyer's patch M cells. This distinguishes the M cell IgA receptor from CD89 (FcalphaI), which binds domains Calpha2-Calpha3. Finally, we observed by immunofluorescence microscopy that some M cells in the human ileum are coated with IgA. Together these data suggest that mouse, and possibly human, M cells express an IgA-specific receptor on their apical surfaces that mediates the transepithelial transport of SIgA from the intestinal lumen to underlying gut-associated organized lymphoid tissues.     

Alternate mucosal immune system: organized Peyer's patches are not required for IgA responses in the gastrointestinal tract

The progeny of mice treated with lymphotoxin (LT)-beta receptor (LTbetaR) and Ig (LTbetaR-Ig) lack Peyer's patches but not mesenteric lymph nodes (MLN). In this study, we used this approach to determine the importance of Peyer's patches for induction of mucosal IgA Ab responses in the murine gastrointestinal tract. Immunohistochemical analysis revealed that LTbetaR-Ig-treated, Peyer's patch null (PP null) mice possessed significant numbers of IgA-positive (IgA+) plasma cells in the intestinal lamina propria. Further, oral immunization of PP null mice with OVA plus cholera toxin as mucosal adjuvant resulted in Ag-specific mucosal IgA and serum IgG Ab responses. OVA-specific CD4+ T cells of the Th2 type were induced in MLN and spleen of PP null mice. In contrast, when TNF and LT-alpha double knockout (TNF/LT-alpha-/-) mice, which lack both Peyer's patches and MLN, were orally immunized with OVA plus cholera toxin, neither mucosal IgA nor serum IgG anti-OVA Abs were induced. On the other hand, LTbetaR-Ig- and TNF receptor 55-Ig-treated normal adult mice elicited OVA- and cholera toxin B subunit-specific mucosal IgA responses, indicating that both LT-alphabeta and TNF/LT-alpha pathways do not contribute for class switching for IgA Ab responses. These results show that the MLN plays a more important role than had been appreciated until now for the induction of both mucosal and systemic Ab responses after oral immunization. Further, organized Peyer's patches are not a strict requirement for induction of mucosal IgA Ab responses in the gastrointestinal tract.     

Generalized systemic and mucosal immunity in mice after mucosal stimulation with cholera toxin

Cholera toxin (CT) has been found to be an extremely potent immunogen for mucosal IgA responses when administered via the intestine. This study has examined both mucosal and systemic immune responses after feeding CT and compared these responses with those obtained after feeding keyhole limpet hemocyanin (KLH), another protein that is strongly immunogenic in mice. Feeding CT to mice resulted not only in IgA antibody in intestinal secretions but also resulted in substantial plasma IgG and IgA antibody levels. Feeding KLH in much larger quantity resulted in little or no antibody response in intestinal secretions or plasma. Lymphoid cells from various tissues of mice fed CT were cultured in vitro for 10 days and the supernatant was tested for antibody to CT. Spontaneous antibody synthesis (no antigen added to cultures) was present in cultures of each cell type, but IgG anti-CT was found mainly in cultures of spleen and mesenteric lymph node cells and IgA anti-CT mainly in cultures of Peyer's patch and lamina propria cells. Peyer's patch cells cultured with CT as antigen synthesized both IgG and IgA anti-CT, suggesting that the antibody response to both isotypes originated in this site. Helper T cell activity for both IgA and IgG anti-CT was detected in spleens, mesenteric lymph nodes, and Peyer's patches. Lastly, when KLH and CT were fed to mice at the same time, an intestinal IgA anti-KLH and plasma IgG anti-KLH response was stimulated, a response pattern similar to that occurring to CT after CT was fed alone. We conclude that mucosal stimulation by CT generates both a systemic IgG and mucosal IgA response to this antigen, and that CT can cause a similar pattern of response to an unrelated protein antigen when both are administered into the intestine at the same time. The data favor the idea that both the IgG and IgA responses originate in GALT and then disseminate to other tissues. We propose that CT accomplishes these effects by altering the regulatory environment within GALT.     

Secretory immunoglobulin A antibodies against the sigma1 outer capsid protein of reovirus type 1 Lang prevent infection of mouse Peyer's patches

Reovirus type 1 Lang (T1L) adheres to M cells in the follicle-associated epithelium of mouse intestine and exploits the transport activity of M cells to enter and infect the Peyer's patch mucosa. Adult mice that have previously cleared a reovirus T1L infection have virus-specific immunoglobulin G (IgG) in serum and IgA in secretions and are protected against reinfection. Our aim in this study was to determine whether secretory IgA is sufficient for protection of Peyer's patches against oral reovirus challenge and, if so, against which reovirus antigen(s) the IgA may be directed. Monoclonal antibodies (MAbs) of the IgA isotype, directed against the sigma1 protein of reovirus T1L, the viral adhesin, were produced and tested along with other, existing IgA and IgG MAbs against reovirus T1L outer capsid proteins. Anti-sigma1 IgA and IgG MAbs neutralized reovirus T1L in L cell plaque reduction assays and inhibited T1L adherence to L cells and Caco-2(BBe) intestinal epithelial cells in vitro, but MAbs against other proteins did not. Passive oral administration of anti-sigma1 IgA and IgG MAbs prevented Peyer's patch infection in adult mice, but other MAbs did not. When anti-sigma1 IgA and IgG MAbs were produced in mice from hybridoma backpack tumors, however, the IgA prevented Peyer's patch infection, but the IgG did not. The results provide evidence that neutralizing IgA antibodies specific for the sigma1 protein are protective in vitro and in vivo and that the presence of these antibodies in intestinal secretions is sufficient for protection against entry of reovirus T1L into Peyer's patches.     

Lacrimal gland-directed B cell responses

Although it is accepted that IgA plasma cells predominate in the lacrimal gland, the factors leading to this prevalence are not known. A series of 4-day LPS-driven co-culture experiments performed with dissociated lacrimal gland and lymphoid cell populations was employed to study the direct effect of lacrimal gland cells on B cell differentiation. Lacrimal gland cells, when co-cultured with spleen or mesenteric lymph node cells, were found to suppress differentiation of cells to IgA, IgG, and IgM production. Furthermore, suppression of IgG and IgM responses occurred after co-culture of lacrimal gland cells with Peyer's patch cells. However, these Peyer's patch co-cultures led to a stimulation of the IgA response, a condition that was abrogated by removal of Peyer's patch T cells before co-culturing. Pretreatment of lacrimal gland cells with mitomycin C eliminated the suppression and stimulation previously observed. These results demonstrate the effects of lacrimal gland, both directly and indirectly through T cells, on B cell differentiation. These findings explain in part the preferential accumulation of IgA-plasma cells within the gland.     

Activated T lymphocytes resist the toxic effects of the adenosine deaminase inhibitor, 2-deoxycoformycin

Tonsil lymphocytes from three adults and three children were examined for immunoglobulin (Ig) production before and after Epstein-Barr virus (EBV) transformation. T-cell depletion was required to obtain cell lines from EBV-seropositive individuals. Cytoplasmic Ig was mainly IgG in adult lymphocytes before and after transformation; IgA and IgM were more prominent after than before. IgM and IgG predominated in lymphocytes of children before and after transformation; IgA was more prominent after than before. Cytoplasmic Ig of peripheral blood lymphocytes from these individuals was mainly IgM. Secreted Ig from tonsil lymphocytes was mainly IgA or IgG; after transformation IgM predominated with adult cell lines, and IgG or IgM with cell lines from children. IgE was consistently sparse in spite of ragweed and/or grass allergies of the adults.     

Dietary fructooligosaccharides induce immunoregulation of intestinal IgA secretion by murine Peyer's patch cells

Probiotic supplements induce immunological responses in the host, and dietary fructooligosaccharides (FOS) stimulate the growth of selected intestinal microflora. In this study we investigated the immunological influences of orally administrated FOS. BALB/c mice were orally administered 0-7.5% FOS for 6 weeks, and the intestinal mucosal immune responses were measured. In the 2.5%-FOS group, fecal IgA was significantly increased. IgA secretion by Peyer's patch (PP) cells was upregulated in a dose-dependent way in response to FOS and CD4+ T cells from PP showed a dose-dependent increase in production of interferon-gamma and interleukin (IL) 10, and a high response in production of IL-5 and IL-6. In contrast, FOS suppressed serum IgG1. Our findings suggest that FOS supplementation changes the intestinal environment of microflora, and leads to upregulation of IgA secretion in CD4+ PP cells in intestinal mucosa, and to suppression of the systemic immune response to type 2 helper T (Th2) dominant.     

Evidence for the emergence of a type-1-like immune response in intestinal mucosa of calves recovering from cryptosporidiosis

This study was undertaken to characterize the mucosal response to Cryptosporidium parvum in infected calves that had recovered from diarrhea. Flow cytometric surface phenotypes of lamina propria lymphocyte (LPL) suspensions from infected calves and age-matched controls revealed the presence of a significantly larger proportion of CD25+ LPL in infected calves than in controls. Freshly isolated LPL from infected calves expressed more iNOS and interferon (IFN)-gamma than did controls. Infected calves excreted IgG1 and IgG2 isotype antibodies to C. parvum p23 by the end of the experiment. Moreover, immunohistochemistry of ileal sections revealed the presence of IgG1+ and IgG2+ B lymphocytes in the villi and IgG1+ but not IgG2+ B lymphocytes in continuous Peyer's patch nodules. These data are consistent with the emergence of a type-1-like mucosal immune response in terminal ileal mucosa as calves recover from cryptosporidiosis.     

Role of immunoglobulin A in protection against reovirus entry into Murine Peyer's patches

Reovirus type 1 Lang (T1L) infects the mouse intestinal mucosa by adhering specifically to epithelial M cells and exploiting M-cell transport to enter the Peyer's patches. Oral inoculation of adult mice has been shown to elicit cellular and humoral immune responses that clear the infection within 10 days. This study was designed to determine whether adult mice that have cleared a primary infection are protected against viral entry upon oral rechallenge and, if so, whether antireovirus secretory immunoglobulin A (S-IgA) is a necessary component of protection. Adult BALB/c mice that were orally inoculated on day 0 with reovirus T1L produced antiviral S-IgA in feces and IgG in serum directed primarily against the reovirus sigma1 attachment protein. Eight hours after oral reovirus challenge on day 21, the Peyer's patches of previously exposed mice contained no detectable virus whereas Peyer's patches of naive controls contained up to 2,300 PFU of reovirus/mg of tissue. Orally inoculated IgA knockout (IgA(-/-)) mice cleared the initial infection as effectively as wild-type mice and produced higher levels of reovirus-specific serum IgG and secretory IgM than C57BL/6 wild-type mice. When IgA(-/-) mice were rechallenged on day 21, however, their Peyer's patches became infected. These results indicate that intestinal S-IgA is an essential component of immune protection against reovirus entry into Peyer's patch mucosa.     

Immunomodulatory effect of (--)-matairesinol in vivo and ex vivo

Matairesinol is one of the lignan compounds found in a variety of plant foodstuffs. We investigated the immunomodulatory effects of (-)-matairesinol in vivo and ex vivo by using mice. Although we found no significant differences in the IgG, IgA and IgM levels in the serum, the IgE level was strongly suppressed by the uptake of (-)-matairesinol in both intact and ovalbumin-immunized mice. The immunoglobulin produced by lymphocytes from the spleen was not activated by the intake of (-)-matairesinol. However, lymphocytes in such gut-associated lymphatic tissues as Peyer's patches and mesenteric lymph nodes were activated by the administration of (-)-matairesinol.     

IL-6 is a potent cofactor of IL-1 in IgM synthesis and of IL-5 in IgA synthesis

In these studies we determined the capacity of IL-6 to act as a differentiation cofactor for murine Peyer's patch B cells producing different Ig classes and subclasses. In preliminary studies we determined that sufficient endogenous IL-6 was produced in LPS-induced cell systems to obscure responses to exogenous IL-6. We therefore studied IL-6 effects on Peyer's patch B cells (T cell-depleted cell populations) in the absence of LPS, relying on responses of in vivo-activated cells. rIL-1 alpha or purified IL-6 only slightly enhanced synthesis of IgM over minimal baseline levels in Peyer's patch T cell-depleted cell cultures; however, when IL-6 was added to cultures also containing rIL-1, IgM synthesis was very substantially increased. In addition, rIL-5 alone gave rise to a modest increase in IgM synthesis and its effect was not enhanced by either rIL-1 or IL-6. IgG production (mainly IgG3) followed a similar pattern. In contrast, IgA production was only modestly increased above baseline by rIL-1, rIL-5, or IL-6 alone or by rIL-1 and IL-6 in combination, but was greatly increased by rIL-5 and IL-6 in combination. The effect of IL-6 on Ig synthesis in the above studies was not due to an effect on cell proliferation. In summary, these data indicate that B cells differ in respect to the cytokines supporting maximal terminal differentiation and thus the class of Ig produced may depend on the presence of a particular combination of cytokines and lymphokines.     

Lack of J chain inhibits the transport of gut IgA and abrogates the development of intestinal antitoxic protection.

Recent publications have provided confusing information on the importance of the J chain for secretion of dimeric IgA at mucosal surfaces. Using J chain-deficient (J chain-/-) mice, we addressed whether a lack of J chain had any functional consequence for the ability to resist challenge with cholera toxin (CT) in intestinal loops. J chain-/- mice had normal levels of IgA plasma cells in the gut mucosa, and the Peyer's patches exhibited normal IgA B cell differentiation and germinal center reactions. The total IgA levels in gut lavage were reduced by roughly 90% as compared with that in wild-type controls, while concomitantly serum IgA levels were significantly increased. Total serum IgM levels were depressed, whereas IgG concentrations were normal. Following oral immunizations with CT, J chain-/- mice developed 10-fold increased serum antitoxin IgA titers, but gut lavage anti-CT IgA levels were substantially reduced. However, anti-CT IgA spot-forming cell frequencies in the gut lamina propria were normal. Anti-CT IgM concentrations were low in serum and gut lavage, whereas anti-CT IgG titers were unaltered. Challenge of small intestinal ligated loops with CT caused dramatic fluid accumulation in immunized J chain-/- mice, and only 20% protection was detected compared with unimmunized controls. In contrast, wild-type mice demonstrated 80% protection against CT challenge. Mice heterozygous for the J chain deletion exhibited intermediate gut lavage anti-CT IgA and intestinal protection levels, arguing for a J chain gene-dosage effect on the transport of secretory IgA. This study unequivocally demonstrates a direct relationship between mucosal transport of secretory SIgA and intestinal immune protection.     

Effects of bovine beta-casein (1-28) and its chemically synthesized partial fragments on proliferative responses and immunoglobulin production in mouse spleen cell cultures.

Effects of bovine beta-casein (1-28) having a phosphoserine-rich region (Glu14-SerP-Leu-SerP-SerP-SerP-Glu-Glu21) and its chemically synthesized partial fragments on proliferation of lymphocytes and immunoglobulin production were investigated in mouse spleen cell cultures. The parent fragment 1-28 and all fragments containing SerP-Leu-SerP and/or SerP-SerP-SerP had a significant mitogenic effect, stimulated proliferation of lymphocytes induced by lipopolysaccharide, phytohemagglutinin, or concanavalin A, and increased immunoglobulin (IgG + IgM + IgA) or IgA levels in the cell cultures. In contrast, dephosphorylated beta-casein (14-21) and SerP-SerP amide had hardly any immunoregulatory activity. On the other hand, SerP-Leu-SerP amide reacted little with antibodies specific to bovine beta-casein (1-28), but beta-casein (14-21), and SerP-SerP-SerP amide obviously reacted with the antibody. These results confirm that the immunoregulatory activity of casein phosphopeptides is attributable to SerP-X-SerP, which may well be available as a non-allergic food ingredient having an adjuvant activity for mucosal IgA responses.     

Migration of antigen-presenting B cells from peripheral to mucosal lymphoid tissues may induce intestinal antigen-specific IgA following parenteral immunization.

Parenterally administered immunizations have long been used to induce protection from mucosal pathogens such as Bordetella pertussis and influenza virus. We previously found that i.m. inoculation of mice with the intestinal pathogen, rotavirus, induced virus-specific Ab production by intestinal lymphocytes. We have now used adoptive transfer studies to identify the cell types responsible for the generation of virus-specific Ab production by gut-associated lymphoid tissue (GALT) after i.m. immunization. Three days after i.m. immunization with rotavirus, cells obtained from the draining peripheral lymph nodes of donor mice were transferred into naive recipient mice. We found that intestinal lymphocytes produced rotavirus-specific Igs (IgM, IgA, and IgG) 2 wk after transfer of either unfractionated cells, or unfractionated cells rendered incapable of cellular division by mitomycin C treatment. Additional studies demonstrated that rotavirus-specific IgA, but not IgG, was produced by intestinal lymphocytes after transfer of purified B cells. Ig allotype analysis revealed that rotavirus-specific IgA was produced by intestinal B cells of recipient origin, suggesting that migration of Ag-presenting B cells from peripheral lymphoid tissues to GALT may contribute to the generation of mucosal IgA responses after parenteral immunization. Strategies that promote Ag uptake and presentation by B cells may enhance mucosal IgA production following parenteral immunization.     

One year in Antarctica: mucosal immunity at three Australian stations

The effect of a year's isolation in Antarctica on the human mucosal immune system was assessed during the winter of 1992 at three Australian Antarctic stations: Casey, Davis and Mawson. Saliva samples were collected from each expeditioner prior to their departure from Australia and during each month in Antarctica. The concentrations of salivary immunoglobulins IgA and IgG were significantly different between the three stations, but there were no differences for salivary IgM and albumin. The mean concentrations of IgA were higher at Mawson (P < 0.008), and the mean concentrations of IgG were lower at Davis (P < 0.001) compared with the other stations. Ranges of values observed at the stations over the 12-13 months were similar. The variability of values within individuals showed station differences for salivary IgM and IgG only. The study revealed significant changes in salivary immunoglobulin values over the period in Antarctica, with similar patterns at the three Australian stations. The salivary IgA and IgM levels were lower in the first 4 months in Antarctica (January-April) and increased to maximum values in July-August, before returning to mean levels when isolation was broken in October-November. The patterns of salivary IgA and IgM suggest that stressors due to isolation may play a role in alterations of mucosal immunity in expeditioners in Antarctica.