Ulf Dahlgren

Loss of ileal IgA+ plasma cells and of CD4+ lymphocytes in ileal Peyer's patches of vitamin A deficient rats

Child mortality in diarrhoeal disease is increased significantly by vitamin A deficiency in poor countries. The pathological mechanisms are not known in detail. However, in this paper we report that vitamin A‐deficient Wistar rats had much reduced IgA+ plasma cells in the ileal lamina propria (eightfold reduction from 470 cells/mm2, P = 0·009), as well as a prominent reduction of CD4+ cells in the parafollicular regions of ileal Peyers patches (reduction from 7200 to 105 cells/mm2, P = 0·009). IL‐2Ralpha‐chain (CD25) positive lymphocytes in the ileal Peyers patches were also reduced significantly in vitamin A deficiency (from 1400 to 300 cells/mm2, P = 0·009). The density of CD8 cells tended to be increased relative to the control animals (from 5100 to 6000 cells/mm2, not statistically significant). In conclusion, the marked decrease of lamina propria IgA+ plasma cells may be one cause of the high diarrhoeal mortality in vitamin A deficiency. This, in turn, appears to be related to reduced numbers of activated or regulatory CD4+ T cells in Peyers patches.

Oral Tolerization Leads to Active Suppression and Bystander Tolerance in Adult Rats while Anergy Dominates in Young Rats

Oral tolerance was induced in 4‐week‐old (young) and 12‐week‐old (adult) rats by feeding ovalbumin (OvA)‐containing pellets during 4 weeks. Seven weeks after removal of the OvA‐pellets the rats were immunized with a mixture of OvA and human serumalbumin (HSA) in Freund’s complete adjuvant (FCA), and the following immune response was monitored. Both the young and adult groups of OvA‐fed rats had significantly suppressed OvA‐specific delayed‐type hypersensitivity (DTH) responses and T‐cellproliferation, reflecting a long‐lasting T‐cell tolerance to OvA both in vivo and in vitro. Furthermore, spleen cells from rats tolerized as adults were able to suppress the proliferation of primed T‐cells from normal immunized rats, demonstrating thepresence of antigen‐specific suppressive cells. Accordingly, the adult rats showed bystander suppression of the response to HSA with respect to DTH‐reaction, specific proliferation, and reduced enlargement of the draining lymph nodes after immunization.There was no evidence of active suppression in vitro or bystander tolerance in the orally tolerized young group, indicating that anergy rather than active suppression was prevalent in these rats. Furthermore, in the young group there was no suppression of the antibody response since the IgG and IgE anti‐OvA antibody levels were indistinguishable from those of the controls. Contrary to the young rats, the adult fed group showed transiently elevated levels of IgG anti‐OvA antibodies at 1 weekpost‐immunization, followed by a subsequent significantly suppressed IgG antibody response. In conclusion, the results demonstrate that the induction of anergy or active suppression after antigen feeding can be determined by the age at which the antigenis introduced to the mucosal immune system.

Neonatal colonization of rats induces immunological tolerance to bacterial antigens

We wanted to investigate the immunological events occurring in rats intestinally colonized from birth (neonatally) or at adult age with an ovalbumin (OVA)‐producing Escherichia coli O6K13 strain, carrying type 1 pili. The neonatally colonized animals responded with lower delayed type hypersensitivity (DTH) against OVA and lower levels of IgG antibodies against OVA, O6 lipopolysaccharide (LPS) and type 1 pili compared to age‐matched controls. The IgG antibody response against the bystander antigen, human serum albumin (HSA), was lower in the neonatally colonized animals than in the controls co‐immunized with HSA and E. coli, indicating a release of suppressive factors induced by the bacterial antigens. The adult colonized animals showed an increased DTH and antibody response against OVA after immunization. They also had high pre‐immunization levels of IgG anti‐O6 LPS antibodies compared to controls. However, the relative increase in IgG anti‐O6 LPS antibody levels after the immunization with dead E. coli was much lower in the adult colonized animals. The present results suggest that neonatal animals develop tolerance against antigen on bacterial colonizers of the intestine. In addition, this tolerance contains components of suppression.

Dimeric IgA in the Rat is Transferred from Serum into Bile but not into Milk

Intravenous administration of ducts thoracicus lymph with dimeric IgA antibodies against Escherichia coli 06 to lactating rat dams did not result in transfer of IgA antibodies into the milk, although the antibodies were detectable in serum 1 min after the administration and in bile 60 min later. After intravenous injection of serum from bile‐duct‐occluded (BDO) rats Immunized in the Peyers patchs into lactating rat dams, IgA antibodies appeared in the serum and remained there up to 230 min. At this time no IgA antibodies were seen in the milk while they were present in bile. IgG and IgM 06 antibodies did not appear in bile or after intravenous administration of lymph or serum from BDO rats.

Active suppression in orally tolerized rats coincides with in situ transforming growth factor-beta (TGF-β) expression in the draining lymph nodes

Adult rats were fed pellets containing ovalbumin (OvA) during 4 weeks, and were 2 weeks thereafter immunized subcutaneously with a mixture of OvA and human serum albumin (HSA) in Freunds complete adjuvant (day 0). As a result of the immunization, the draining lymph nodes of the non‐tolerized (control) rats were heavily enlarged from day 10 to day 18; however, this size increase was absent in the OvA‐fed rats. This manifestation of active suppression in the tolerized rats was preceded by the appearance of scattered CD4+ TGF‐β‐expressing T cells in the T cell area of their lymph nodes (days 5–8); correspondingly, the levels of TGF‐β mRNA in the nodes were elevated in the tolerant rats compared with the control rats. The anti‐OvA antibody levels in sera from the rats revealed that there was an initial B cell priming in the OvA‐fed group, with levels higher than in the control group during the first week. Thereafter, suppression governed the response, and from day 10 onwards the anti‐OvA levels were considerably lower than in the controls. When other groups of animals were pretreated with neutralizing anti‐TGF‐β antibodies 1 day before the immunization, the anti‐OvA response of the OvA‐fed rats was restored to the levels of the control group, demonstrating the importance of TGF‐β in the maintenance of suppression. In conclusion, we demonstrate that TGF‐β‐producing cells appear in the draining lymph nodes shortly after immunization in rats made orally tolerant using a relatively high‐dose feeding regime; these cells are probably responsible for the down‐regulation of the immune response observed in the OvA‐fed rats.

Vitamin A deficiency leads to severe functional disturbance of the intestinal epithelium enzymes associated with diarrhoea and increased bacterial translocation in gnotobiotic rats.

A disturbance of the integrity of the intestinal epithelium with an increased risk for bacterial translocation is one of the suggested factors underlying the increased incidence of infections and septicaemia during vitamin A deficiency. In the present study the effects of vitamin A deficiency on the enzymic activity of enterocytes in response to bacterial colonization with a non-pathogenic Escherichia coli strain were studied in monocolonized and conventional Wistar rats. The monocolonized, but not the conventional, vitamin A-deficient rats had markedly reduced weight compared to their pair-fed controls and presented neurological symptoms, such as hind leg weakness, tremor and slow gait. Moreover, only in the monocolonized vitamin A-deficient rats were severe diarrhoea and bacterial translocation to extraintestinal sites-mainly kidneys-detected. Measurements of enterocyte brush-border enzyme activities revealed that lactase, sucrase, gamma-glutamyltranspeptidase (GGT) and dipeptidyl peptidase IV (DPP IV) were significantly reduced in the monocolonized vitamin A-deficient rats compared to the pair-fed controls, indicating a severe functional disturbance of the enterocytes. In conventional vitamin A-deficient rats only sucrase activity was markedly lower than in the respective controls. Our observation, that the deficient vitamin A status led to a strong reduction of enterocyte enzymic activities, associated with diarrhoea and increased bacterial translocation, mainly in the gnotobiotic rats, suggests that the composition of the bacterial flora, i.e. the colonization state, has a strong influence on triggering the severity of the functional disturbances of the intestinal epithelium, and adds to the clinical manifestations of vitamin A deficiency.

Food Allergens Transformed into Tolerogens

Antigen presentation determines immunologic outcome, and by modifying the presentation of allergen to the host one can prevent an allergic response. Under certain conditions, covalent linkage, of ovalbumin to rat IgG, a molecule already tolerated by the host, can make a protein-IgG conjugate which down-regulates the immune response to this food allergen. The suppression is allergen specific. It affects both T and B cell immune responses. Administration of allergens linked to isologous IgG may provide a novel strategy for allergy prevention.

Langerhans cells from human oral epithelium are more effective at stimulating allogeneic T cells in vitro than Langerhans cells from skin

This  report  is  focused  on  the  functional  capacity  of  Langerhans  cells  (LC)  in  the epithelium  of skin and oral mucosa, which both meet different antigenic challenges. The capacity of LC from human oral and skin epithelium to provide co‐stimulatory signals to T cells in vitro was compared. LC in a crude suspension of oral epithelial cells had a significantly enhanced T cell co‐stimulatory capacity compared to skin epithelial cells. This applied both to cultures with concanavalin A (con‐A)‐stimulated syngeneic T cells and to a mixed epithelial cell lymphocyte reaction involving allogeneic T cells. The co‐stimulatory capacity of oral and skin epithelial cells was reduced by >70% if monoclonal antibodies against HLA‐DR, ‐DP and ‐DQ were added to the cultures with allogeneic T cells, indicating the involvement of HLA class II expressing LC. Immunohistochemistry revealed that 6% of the epithelial cells were CD1a + LC in sections from both oral and skin epithelium. Interleukin (IL)‐8 production was higher in cultures of oral epithelial cells and con‐A stimulated T cells than in corresponding cultures with skin epithelial cells as accessory cells. The results suggest that LC in human oral epithelium are more efficient at stimulating T cells than those of skin.

Defence of mucous membranes by antibodies, receptor analogues and non-specific host factors

SummaryMost infections reach man via the mucosal membranes, and more than half of the lymphoid system is found in connection with mucosae. The major antibodies found on mucous membranes are secretory IgA, which function primarily by binding microorganisms and thereby preventing their contact with the host tissues. The optimal mode of immunization to obtain a secretory IgA response is not well defined. Repeated mucosal exposure with antigen may result in oral tolerance, with decreasing circulating antibodies but a remaining secretory IgA response. The secretory IgA response is usually short-lived and can be difficult to boost. IgM as well as IgG antibodies may add to host defence at the mucosal level, but when engaged, they usually induce inflammation in host tissues. Analogues to bacterial receptors on mucosal epithelium may be present in exocrine secretions such as human milk. During an attack on the host, it is possible that such receptor analogues may aid in the prevention of attachment of bacteria to mucous membranes used as an initial site. A number of non-specific host factors support mucosal defence. One of them is lactoferrin. Lactoferrin deficiency seems to result in recurrent bacterial infections, suggesting its importance in normal host defence.ZusammenfassungBeim Menschen gelangen Infektionen zum größten Teil über die Schleimhäute in den Organismus; mehr als die Hälfte des lymphatischen Systems findet sich in Verbindung mit den Schleimhäuten. Sekretorisches IgA stellt die wichtigste Antikörperklasse der Schleimhäute dar; seine Funktion besteht primär darin, daß es Mikroorganismen bindet und damit ihren Kontakt mit den Geweben des Wirtes verhindert. Wie durch Immunisierung am besten eine Antwort von sekretorischem IgA erzielt werden kann, ist bislang nicht hinreichend geklärt. Wiederholte Antigenexposition von Schleimhäuten kann eine orale Toleranz hervorrufen, bei der es zu einer Verminderung der zirkulierenden Antikörper kommt, während die Antwort von sekretorischem IgA erhalten bleibt. Für gewöhnlich hält die Antwort von sekretorischem IgA nur kurze Zeit an, eine Boosterung ist schwierig. IgM — und IgG-Antikörper unterstützen wahrscheinlich die Abwehr im Bereich der Schleimhäute, doch kommt es in der Regel zu einer Entzündung der Gewebe, wenn diese Antikörper beteiligt sind. Exokrine Sekrete wie die Muttermilch enthalten wahrscheinlich Analoga zu Rezeptoren für Bakterien auf dem Schleimhautepithel. Es ist möglich, daß solche Rezeptorenanaloga das Anheften von Bakterien an den Schleimhäuten verhindern helfen, die als primärer Angriffsort beim Eindringen der Erreger dienen. Eine Reihe unspezifischer Wirtsfaktoren unterstützen die Abwehr im Schleimhautbereich. Dazu gehört Lactoferrin. Lactoferrin-Mangel führt offensichtlich zu rezidivierenden bakteriellen Infektionen, was auf seine Bedeutung für die normale Abwehr schließen läßt.

An established immune response against ovalbumin is suppressed by a transferable serum factor produced after ovalbumin feeding: a role of CD25+ regulatory cells.

We have previously demonstrated that rats fed ovalbumin (OVA) develop a tolerogenic activity in serum, which upon transfer induces tolerance to OVA and suppression of the immune response to a bystander antigen. Here, we have extended these studies and analysed if the tolerogenic activity in serum could suppress an established immune response in the recipients. Rats were immunized with OVA, 4 and 1 week prior to the transfer of serum from either OVA‐fed or control animals.