N-3 fatty acids modulate Th1 and Th2 dichotomy in diabetic pregnancy and macrosomia

We assessed the implication of Th (helper)-cells and the modulation of the Th1/Th2 dichotomy by n-3 polyunsaturated fatty acids (PUFA) in type I diabetic pregnancy (DP) and macrosomia. Female gestant rats fed a standard diet or n-3 PUFA regimen were rendered diabetic by administration of five low doses of streptozotocin. The macrosomic (MAC) offspring were sacrificed at the age of 90 days. The mRNAs of IL-2 and IFN-gamma (Th1 cytokines) and IL-4 (Th2 cytokine) were downregulated in the pancreas and spleen of diabetic pregnant rats. The levels of IL-10 mRNA, another Th2 cytokine, were unchanged in the spleen or upregulated in the pancreas of these animals. Feeding an n-3 PUFA diet to rats with DP upregulated IL-10 mRNA in the pancreas and IL-4 and IL-10 mRNA in the spleen. In MAC offspring, high expression of IL-2 and IFN-gamma mRNA, but not of Th2 cytokines, was observed. The n-3 PUFA diet diminished Th1 mRNA quantities and increased the levels of IL-4, but not of IL-10, mRNA in MAC offspring. Our study shows that DP is associated with a decreased Th1 phenotype and IL-4 mRNA expression in the pancreas and spleen, and an n-3 PUFA diet upregulates Th2 profile. In MAC offspring, the Th1 phenotype is upregulated and an n-3 PUFA diet downregulates this phenomenon.     

Assessment of interleukin-12, gamma interferon, and tumor necrosis factor alpha secretion in sera from mice fed with dietary lipids during different stages of Listeria monocytogenes infection

Recent experimental observations have determined that long-chain n-3 polyunsaturated fatty acids suppress immune functions and are involved in the reduction of infectious disease resistance. BALB/c mice were fed for 4 weeks with one of four diets containing either olive oil (OO), fish oil (FO), hydrogenated coconut oil, or a low fat level. Interleukin-12p70 (IL-12p70), gamma interferon (IFN-gamma), and tumor necrosis factor alpha (TNF-alpha) production in the sera of mice fed these diets and challenged with Listeria monocytogenes were determined by enzyme-linked immunosorbent assay. In addition, bacterial counts from spleens of mice were carried out at 24, 72, or 96 h of infection. Here, we quantified an initial diminution of production of both IL-12p70 and IFN-gamma, which appear to play an important role in the reduction of host resistance to L. monocytogenes infection. In addition, an efficient elimination of L. monocytogenes was observed in spleens of mice fed a diet containing OO at 96 h of infection, despite reductions in IL-12p70 and TNF-alpha production, suggesting an improvement of immune resistance. Overall, our results indicate that the initial reduction of both IL-12 and IFN-gamma production before L. monocytogenes infection represents the most relevant event that corroborates the impairment of immune resistance by n-3 polyunsaturated fatty acids during the different stages of infection. However, we speculate that the modulation of other cytokines must be also involved in this response, because the alteration of cytokine production in mice fed an FO diet in a late phase of L. monocytogenes infection was similar to that in mice fed OO, whereas the ability to eliminate this bacterium from the spleen was improved in the latter group.     

The effect of dietary lipid manipulation on rat lymphocyte subsets and proliferation.

Polyunsaturated fatty acids (PUFA) have been shown to suppress immune cell functions in vitro. Dietary studies investigating the effects of PUFA-containing oils on lymphocyte functions have yielded contradictory findings: such studies are difficult to compare as there are many variations in protocols. The present study investigated the effects of diets containing oils rich in saturated fatty acids, monounsaturated fatty acids, n-6 PUFA or n-3 PUFA on rat lymphocyte proliferation and on receptor and surface marker expression. Rats were fed for 10 weeks on a low-fat (LF) diet (approximately 2% fat by weight) or on one of five high-fat diets, which contained 20% (by weight) hydrogenated coconut oil (HCO), olive oil (OO), safflower oil (SO), evening primrose oil (EPO) or menhaden (fish) oil (MO). Compared with feeding the LF diet, all of the high-fat diets suppressed the proliferation of lymphocytes from the spleen: although there was no significant effect of diet on the proliferation of lymphocytes from the thymus, there was a trend towards decreased proliferation with high-fat feeding. Feeding the OO, EPO or MO diets significantly suppressed proliferation of mesenteric lymph node lymphocytes compared with feeding the LF, HCO or SO diets. Dietary lipid manipulation had no effect on the proportion of T cells, B cells or monocytes/macrophages in the spleen, thymus or lymph nodes. Dietary lipid manipulation also had no significant effect on the proportions of CD4+ or CD8+ lymphocytes in spleen, thymus or lymph nodes, either in freshly prepared cells or in cells cultured in the presence of mitogen. There were no significant effects of dietary lipid manipulation on the expression of IL-2 receptors or transferrin receptors by concanavalin A (Con A)-stimulated lymphocytes. However, there was a trend towards a decrease in transferrin receptor expression by Con A-stimulated lymphocytes from the thymus and lymph nodes of the MO-fed rats and towards a decrease in the expression of IL-2 receptors by lymphocytes from the spleens and thymi of the MO-fed rats. These observations provide evidence that some dietary oils, particularly OO, EPO and MO, possess immunosuppressive properties and so may be useful in the therapy of diseases involving inappropriate lymphocyte activation.     

The influence of dietary protein antigen on Th1/Th2 balance and cellular immunity.

Dietary administration of ovalbumin (OVA) antigen (Ag) into OVA-specific T cell receptor alphabeta-transgenic (TCR-Tg) mice resulted in the induction of activated CD4+ Th cells expressing CD69 early activation Ag. However, the number of CD4+ Th cells rather decreased by dietary administration of OVA antigen. The production of Th1-cytokines such as IFN-gamma and IL-2 markedly reduced in spleen of OVA-fed mice compared to mice fed with normal diet. In sharp contrast, the production of Th2-cytokine, IL-4 greatly increased in spleen of OVA-fed mice though the number of CD4+ T cells decreased to less than 10% of control mouse spleen. The decrease of IFN-gamma production and the increase of IL-4 production by CD4+ T cells was demonstrated at a single cell level by intracellular cytokine staining analysis. Moreover, such a polarized cytokine production pattern was also demonstrated using highly purified CD4+ T cells obtained from mice fed with OVA. In addition to the decrease of Th1-cytokine production, TCR-Tg mice fed with OVA-containing diet showed greatly reduced in vivo generation of NK cells, LAK cells and CTL. These results suggested that dietary protein antigen caused the polarization of Th1/Th2 balance into Th2-dominant immunity and inhibited cellular immunity.     

Dietary lipids modify the cytokine response to bacterial lipopolysaccharide in mice

To investigate the effect of dietary lipids with different fatty acid compositions upon the in vivo cytokine response to bacterial lipopolysaccharide (LPS), mice were fed for 5 weeks on a low-fat diet or on one of four high-fat diets that contained 20%, by weight, of coconut oil (CO), olive oil (OO), safflower oil (SO) or fish oil (FO). The mice were injected intraperitoneally with a non-lethal dose of Escherichia coli LPS (100 micrograms/20 g body weight) and killed 90 or 180 min later. Plasma tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-1alpha, IL-6 and IL-10 concentrations were measured by enzyme-linked immunosorbent assay (ELISA). Plasma TNF-alpha and IL-10 concentrations were higher 90 min postinjection than after 180 min, whereas plasma IL-1beta and IL-6 concentrations were higher 180 min postinjection than after 90 min. Peak plasma TNF-alpha, IL-1beta and IL-6 concentrations were lower in the CO- and FO-fed mice than in those fed the SO diet. Peak plasma IL-10 concentrations were higher in CO-fed mice than in those fed some of the other diets. These observations suggest that, relative to the n-6 polyunsaturated fatty acid-rich SO diet, CO and FO diminish production of proinflammatory cytokines in vivo. This indicates that these fatty acids might be useful therapies in acute and chronic inflammatory diseases. The enhanced production of IL-10 following CO feeding appears to be an additional antiinflammatory effect of this oil, which could give added benefit in various clinical conditions.     

Dietary omega-3 polyunsaturated fatty acids from fish oil reduce interleukin-12 and interferon-gamma production in mice

The objective of this study was to investigate the impact of feeding mice a diet rich in n-3 polyunsaturated fatty acids (PUFA) from fish oil on the interleukin-12 (IL-12) and interferon-gamma (IFNgamma) production during the early stage of an infectious challenge with Listeria monocytogenes. Weanling female C3H/HeN mice were fed AIN-93G experimental diets containing 20%, by weight one of three fat sources: lard (low PUFA), soybean oil (n-6 PUFA) or a mixture (9:1) of menhaden fish oil and corn oil (n-3 PUFA). After 4 weeks, mice were injected intraperitoneally with 10(5) Listeria monocytogenes and the concentration of IL-12(p70) and IFNgamma in serum was determined 24 h post-infection by ELISA. IL-12p35, IL-12p40 mRNA, and IFNgamma mRNA in the spleen were quantified by RNase protection assay. The number of IFNgamma-producing cells in the spleen was determined by flow cytometry using an intracellular staining procedure. We found that n-3 PUFA-fed mice had lower levels of circulating IL-12 at 24 h post-infection than n-6 PUFA- or low PUFA-fed mice (9.7+/-3.4 pg/ml vs. 61.6+/-10.6, and 44.4+/-12.5 pg/ml, respectively; P=0.002, n = 10/trt). The level of IL-12 p35 mRNA did not significantly differ among dietary treatment groups. However, IL-12p40 mRNA was significantly lower in n-3 PUFA- and n-6 PUFA-fed mice compared to low-PUFA-fed mice. Further, the n-3 PUFA group also had the lowest circulating IFNgamma (4.4+/-1.8 ng/ml vs. 9.1+/-1.0, and 9.7+/-2.1 ng/ml, respectively; P = 0.007. n = 8-10/trt). The n-3 PUFA-fed mice had significantly lower IFNgamma mRNA in their spleens compared to the mice fed the other fat sources. In agreement with having lower circulating IFNgamma and lower splenic IFNgamma mRNA, n-3 PUFA-fed mice had a significantly lower percentage of IFNgamma-producing cells in their spleens compared with the n-6 PUFA-fed group (2.1+/-0.6 vs. 4.2+/-0.7%; P = 0.037, n = 10/trt). In summary, feeding mice a diet rich in n-3 PUFA from fish oil significantly lowered the production of both IL-12 and IFNgamma during the early phase of a Listeria infection.     

Dietary n-3 polyunsaturated fatty acids modulate purified murine T-cell subset activation

Studies in humans and murine disease models have clearly shown dietary fish oil to possess anti-inflammatory properties, apparently mediated by the n-3 polyunsaturated fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). To determine the mechanisms by which dietary EPA and DHA modulate mouse T-cell activation, female C57BL/6 mice were fed diets containing either 2% safflower oil (SAF), 2% fish oil (FO), or a 2% purified EPA/DHA ethyl ester mixture for 14 days. Splenic CD4 T cells ( approximately 90% purity) or CD8 T cells ( approximately 85% purity) were incubated with agonists which act at the plasma membrane receptor level [anti(alpha)-CD3/anti(alpha)-CD28], the intracellular level (PMA/Ionomycin), or at both the receptor and intracellular levels (alphaCD3/PMA). CD4 T cells stimulated with alphaCD3/alphaCD28 or PMA/Ionomycin proliferated and produced principally IL-2 (i.e. a Th1 phenotype), whereas the proliferation of CD4 T cells stimulated with alphaCD3/PMA was apparently driven principally by IL-4 (i.e. a Th2 phenotype). The IL-4 driven proliferation of putative Th2 CD4 cells was enhanced by dietary n-3 fatty acids (P = 0.02). Conversely, IL-2 production by alphaCD3/alpha CD28-stimulated CD4 T cells was reduced in FO-fed animals (P < 0.0001). The alphaCD3/alphaCD28-stimulated CD8 cells cultured from FO-fed animals exhibited a significant decrease (P < 0.05) in proliferation. There were no dietary effects seen in alphaCD3/PMA-stimulated CD8 cells, which produced both IL-2 and IL-4, or in PMA/Ionomycin-stimulated CD8 cells, which produced principally IL-2. These data suggest that dietary n-3 fatty acids down-regulated IL-2 driven CD4 and CD8 activation, while up-regulating the activation of the Th2 CD4 T-cell subset. Thus, the anti-inflammatory effects of n-3 fatty acids may result in both the direct suppression of IL-2-induced Th1 cell activation and the indirect suppression of Th1 cells by the enhanced cross-regulatory function of Th2 cells.     

Profiles of Th1 and Th2 Cytokines after Primary and Secondary Infection by Schistosoma mansoni in the Semipermissive Rat Host

In contrast to most mouse strains, rats eliminate the primary schistosome burden around 4 weeks postinfection and subsequently develop protective immunity to reinfection. In rat schistosomiasis, we have shown predominant expression of a Th2-type cytokine response at the mRNA level after primary infection. In the present study, we showed a significant increase in interleukin-4 (IL-4) mRNA expression in inguinal lymph nodes early after a secondary infection. IL-5 mRNA expression showed a significant increase at days 2 and 4 postreinfection in the spleen and lymph nodes, respectively. We did not detect any gamma interferon (IFN-gamma) mRNA after a challenge infection. Analysis of cytokine secretion by stimulated spleen cells after a primary infection showed predominant expression of IL-4 with maximum production on day 21, accompanied by production of IL-5 from day 11 to day 67. A significant increase in IFN-gamma secretion was detected at day 21. Analysis of immunoglobulin G2b (IgG2b) and IgG2c (Th1-related isotypes) showed undetectable levels of IgG2b, but detectable levels of specific IgG2c antibodies were observed from day 42. The analysis of Th2-related isotypes showed high specific IgG1 and IgG2a antibody titers from day 29. After a secondary infection, only IL-4 and IL-5 secretion was sustained. This is supported by the increased production of Th2-related isotypes. These findings showed that S. mansoni infection can drive Th2 responses in rats in the absence of egg production which is required to induce a Th2 response in mice and are in favor of the role of Th2-type cytokines in protective immunity against reinfection.     

Long-chain polyunsaturated fatty acids are consumed during allergic inflammation and affect T helper type 1 (Th1)- and Th2-mediated hypersensitivity differently

Studies have shown that atopic individuals have decreased serum levels of n‐3 fatty acids. Indicating these compounds may have a protective effect against allergic reaction and/or are consumed during inflammation. This study investigated whether fish (n‐3) or sunflower (n‐6) oil supplementation affected T helper type 1 (Th1)‐ and Th2‐mediated hypersensitivity in the skin and airways, respectively, and whether the fatty acid serum profile changed during the inflammatory response. Mice were fed regular chow, chow + 10% fish oil or chow + 10% sunflower oil. Mice were immunized with ovalbumin (OVA) resolved in Th1 or Th2 adjuvant. For Th1 hypersensitivity, mice were challenged with OVA in the footpad. Footpad swelling, OVA‐induced lymphocyte proliferation and cytokine production in the draining lymph node were evaluated. In the airway hypersensitivity model (Th2), mice were challenged intranasally with OVA and the resulting serum immunoglobulin (Ig)E and eosinophilic lung infiltration were measured. In the Th1 model, OVA‐specific T cells proliferated less and produced less interferon (IFN)‐γ, tumour necrosis factor (TNF) and interleukin (IL)‐6 in fish oil‐fed mice versus controls. Footpad swelling was reduced marginally. In contrast, mice fed fish oil in the Th2 model produced more OVA‐specific IgE and had slightly higher proportions of eosinophils in lung infiltrate. A significant fall in serum levels of long‐chain n‐3 fatty acids accompanied challenge and Th2‐mediated inflammation in Th2 model. Fish oil supplementation affects Th1 and Th2 immune responses conversely; significant consumption of n‐3 fatty acids occurs during Th2‐driven inflammation. The latter observation may explain the association between Th2‐mediated inflammation and low serum levels of n‐3 fatty acids.     

Selective induction of Th2 cells in murine Peyer's patches by oral immunization.

We have used three different methods to determine the T helper (Th) cell response, including Th1 and Th2 types, in murine Peyer's patches (PP) following oral immunization with sheep red blood cells (SRBC). These include: (i) use of cytokine-specific (IFN-gamma and IL-5), single cell assays to estimate the frequencies of Th1 and Th2 cells respectively, (ii) cytokine-specific mRNA--cDNA dot blot and Northern gel hybridizations to detect levels of specific mRNA, and (iii) T cell cloning techniques to determine the frequency of Th1 and Th2 clones. Mice were immunized with SRBC by either the oral or i.p. route. The PP and splenic (SP) CD3+ and CD3+ CD4+ T cell subsets were isolated and cultured with antigen, feeder cells, and IL-2, and were assessed at various intervals (days 0, 1, 3, and 6) for numbers of T cells producing either IFN-gamma or IL-5 by use of an enzyme-linked immunospot (ELISPOT) procedure. Cultures of T cells from PP or SP of mice given SRBC by the oral route had a high frequency of IL-5 spot forming cells (SFC), with lower numbers of IFN-gamma SFC. However, cultures of CD3+ T cells and CD3+ CD4+ Th cells from spleens of i.p. immunized mice exhibited predominantly IFN-gamma SFC, with smaller but significant numbers of IL-5 SFC. This distinct pattern of cytokine production was supported by mRNA analysis where high IL-5 specific mRNA levels were noted in PP T cell cultures of orally primed mice, while IFN-gamma mRNA was predominant in the SP CD3+ T cell and CD3+ CD4+ Th cell cultures from i.p. immunized mice. When the frequencies of IFN-gamma or IL-5 SFC were assessed among cloned Th cells from orally- or systemically-immunized mice, 74% of Th cell clones from PP of mice orally immunized with SRBC were IL-5 producers (Th2 type), while 67% of Th cell clones from SP of mice immunized by the i.p. route were IFN-gamma producers (Th1 type). Our studies show that higher frequencies of IFN-gamma producing Th1-type cells occur in SP of mice given antigen by the systemic route, while oral immunization results in predominantly IL-5 producing, Th2-type cells in PP.     

Dietary n-3 polyunsaturated fatty acids suppress splenic CD4(+) T cell function in interleukin (IL)-10(-/-) mice

Our laboratory has demonstrated that down-regulation of proliferation and cytokine synthesis by CD4(+) T cells in mice fed diets rich in n-3 polyunsaturated fatty acids (PUFA) is highly dependent on the involvement of the co-stimulatory molecule, CD28. It has been reported that the inhibitory cytokine interleukin (IL)-10 acts directly on T cells which up-regulate IL-10 receptor (IL-10R) expression following stimulation via CD28 by efficiently blocking proliferation and cytokine production. Thus, it was hypothesized that dietary n-3 PUFA would suppress T cell function through the effects of IL-10. The proliferation of purified splenic CD4(+) T cells activated in vitro with anti-CD3 and anti-CD28 (alphaCD3/CD28) from conventional mice (C57BL/6) fed either a control corn oil (CO)-enriched diet devoid of n-3 PUFA, docosahexaenoic acid (DHA; 22 : 6) or eicosapentaenoic acid (EPA; 20 : 5) for 14 days was suppressed by dietary DHA and EPA. Surprisingly, a similar trend was seen in IL-10 gene knock-out (IL-10(-/-)) mice fed dietary n-3 PUFA. IL-10R cell surface expression was also significantly down-regulated on CD4(+) T cells from both the C57BL/6 and IL-10(-/-) mice fed dietary n-3 PUFA after 72 h of in vitro stimulation with alphaCD3/CD28. Enzyme-linked immunosorbent assay (ELISA) measurements revealed that C57BL/6 mice fed DHA had significantly reduced interferon (IFN)-gamma and IL-10 levels 48 h post-activation. However, CD4(+) T cells from IL-10(-/-) mice fed dietary n-3 PUFA produced significantly greater levels of IFN-gamma than the CO-fed group. Our data suggest that in the absence of IL-10, CD4(+) T cells from n-3 PUFA-fed mice may up-regulate IFN-gamma. Suppressed CD4(+) T cells from n-3 PUFA-fed C57BL/6 mice may use mechanisms other than IL-10 to down-regulate T cell function.     

Differential regulation of Th1-type and Th2-type cytokine profiles in pancreatic islets of C57BL/6 and BALB/c mice by multiple low doses of streptozotocin

In the nonobese diabetic (NOD) mouse, the T helper (Th)1-type inflammatory cytokines interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha play a critical role in the development of type 1 diabetes, whereas the Th2-type anti-inflammatory cytokines interleukin (IL)-4 and IL-10 operate counterregulatory. There are no comprehensive analyses on cytokine profiles in the mouse model of diabetes induced with multiple low doses of streptozotocin (MLD-STZ). Therefore, we used islets to study ex vivo effects of MLD-STZ and in vitro effects of STZ on IFN-gamma, TNF-alpha, IL-4, and IL-10 on both levels of protein-producing cells and the mRNA expression, as well as the mRNA expression of the Th3-type cytokine transforming growth factor TGF-beta1. C57BL/6 and BALB/c mice of both genders were injected intraperitoneally with 40 mg/kg body wt STZ on five consecutive days and islets were isolated on day I and 3 after the fifth STZ-injection. Control mice received the solvent of STZ. In islets of C57BL/6 mice of both genders MLD-STZ similarly stimulated production of IFN-gamma and TNF-alpha, but significantly reduced IL-4 and IL-10 levels in male mice only. Opposite results were obtained in islets of BALB/c mice of both genders. Here, MLD-STZ markedly decreased the levels of IFN-gamma and TNF-alpha, but significantly increased the levels of IL-4 and IL-10. The functional results were in line with MLD-STZ effects on the mRNA expression of the cytokines. Moreover, MLD-STZ effects on the TGF-beta1 mRNA expression were reversed to the effects on IFN-gamma and TNF-alpha. The in vitro effects of STZ in islets, in general, were similar to those exerted by MLD-STZ. Apparently, reduction and upregulation of Th2-type cytokines was more associated with susceptibility and resistance, respectively, to MLD-STZ-induced diabetes than upregulation of Th1-type cytokine levels.     

The initial immune response during experimental cysticercosis is of the mixed Th1/Th2 type

The immunological events that occur during the initial stages of experimental cysticercosis are not known. The studies presented here examined the cytokines produced by peritoneal exudate cells (PECs), splenocytes and mesenteric lymph node (MLN) cells during the first week of infection with larval Taenia crassiceps in BALB/cJ mice. Proliferation assays determined that the earliest time when antigen-specific responses could be measured was 5 days post-infection. Concanavalin A (ConA) stimulation of host cells elicited an initial burst of IL-4 production at 24 h of infection and ConA-stimulated Th2-type cytokine production is predominant by 7 days post-infection. Thus, there are responses at day 1 of infection that seem to promote a Th2-type response. Stimulation of MLN cells, splenocytes and PECs with larval antigens supported previous reports of mixed Th1/Th2-type cytokine production with increases in interleukin (IL)-4, IL-10 and interferon (IFN)-gamma. Ex vivo IFN-gamma production by PECs from infected mice was increased at 3, 5 and 7 days post-infection, whereas at these times reduced ex vivo IL-10 production was observed. This ex vivo IFN-gamma response preceded an increasing IL-10 production by PECs between 3 and 7 days post-infection in parasite-specific and ConA-induced proliferation assays. Thus, infection with larval T. crassiceps results in an initial response mediated by IFN-gamma that is quickly followed by an increase in IL-10 production and subsequent reduction in the amount of IFN-gamma being produced.     

Dietary nucleotides can up-regulate antigen-specific Th1 immune responses and suppress antigen-specific IgE responses in mice

BACKGROUND: It has been reported that dietary nucleotides enhance T helper cell activities. In this study, we have determined the effects of dietary nucleotides on antigen-specific Th1 and Th2 responses and IgE responses. METHODS: Ovalbumin (OVA)-specific T cell receptor (TCR) transgenic (OVA-TCR Tg) mice, 3 weeks old, were fed a nucleotide-free diet (NT(-) diet) or the NT(-) diet supplemented with dietary nucleotides (NT(+) diet) for 4 weeks. Cytokine production by spleen cells and macrophages obtained from these mice was measured in vitro. BALB/c mice, 3 weeks old, immunized intraperitoneally with OVA adsorbed onto alum, were fed the NT(-) diet or the NT(+) diet for 4 weeks. Serum levels of antigen-specific antibodies in the BALB/c mice were determined by ELISA. RESULTS: The level of production of antigen-specific interferon-gamma by spleen cells was significantly higher in the OVA-TCR Tg mice fed the NT(+) diet than in the control mice. The levels of secretion of bioactive IL-12 by spleen cells and peritoneal macrophages were also significantly increased in the NT(+) diet group. The serum OVA-specific IgE level was significantly decreased in BALB/c mice fed the NT(+) diet compared with those fed the NT(-) diet. CONCLUSION: These results show that dietary nucleotides up-regulate the antigen-specific Th1 immune response through the enhancement of IL-12 production and suppress the antigen-specific IgE response.     

Inhibition of intracellular peroxides and apoptosis of lymphocytes in lupus-prone B/W mice by dietary n-6 and n-3 lipids with calorie restriction

Our earlier studies have shown that calorie restriction and n-3 fatty acids inhibit autoimmune disease and prolong life span. Experiments were designed to study the alteration of apoptosis and its mediators in B/W mice fed either n-6 fatty acids [5% corn oil (CO)] or n-3 fatty acids [5% fish oil (FO)] and either allowed access to the diet ad libitum (AL) or restricted in caloric intake by 40% (CR), from 4 weeks of age. At 4 months (young) and 9 months (old) mice were killed, splenic lymphocytes were isolated, and apoptosis was measured with Annexin V and PI staining. Apoptosis was decreased in splenic lymphocytes from both young and old CR mice compared to their respective AL-fed control groups regardless of fat source. Increasing apoptosis with age was observed in CO/AL, CO/CR, and FO/AL mice which correlated closely with significantly higher cellular peroxides measured by flow cytometry using dichlorofluourescein diacetate (DCFH-DA), whereas in both CO/CR and FO/CR peroxide levels remained low in old mice. Furthermore, CR increased the proliferative response of splenic lymphocytes and decreased the Fas (CD95) and Fas-L protein expression in CD4+ lymphocytes from old mice. Higher levels of Fas and Fas-L expression were observed in old mice compared to young mice. Bcl-2 levels were elevated in both young and old CR groups compared to the respective AL groups. Calorie restriction prevented the loss of CD8 cells in old mice fed both the CO and the FO diet. In summary, CR resulted in decreased apoptosis accompanied by alterations in Fas, Fas-L, and Bcl-2 expression in old mice, increased life span, and delayed onset of kidney disease.     

Oral antigen induces antigen-specific activation of intraepithelial CD4+ lymphocytes but suppresses their activation in spleen

Intraepithelial lymphocytes (IELs) are considered to drive immune surveillance of the epithelial layer to the mucosa, which is initially exposed to exogenous antigens. However, how IELs are activated by orally administered antigens remains unclear. To clarify this mechanism, we fed ovalbumin (OVA) to T cell receptor transgenic (TCR-Tg) mice with OVA-specific MHC class II-restricted TCR and found that the cytotoxic activity of IELs was increased against both NK and LAK target cells, but notably reduced after depleting CD8 + IELs. Cytoplasmic staining showed that the production of IFN-gamma and IL-2 was increased in mice fed with OVA both in the supernatant of cultured IELs with immobilized anti-CD3 mAb and in fresh CD4+ IELs. In contrast, the cytotoxic activity against NK and LAK target cells and the production of IL-2 and IFN-gamma was decreased in splenic T cells from mice fed with OVA. However, when the splenic T cells from these mice were cultured with OVA and IL-2, IFN-gamma production recovered. The decreased response demonstrated the clonal anergy of T cells. Furthermore, tumor growth was enhanced in TCR-Tg mice carrying an OVA-transfected counterpart A20 B cell lymphoma (OVA-A20) and fed with OVA. These results indicate that the oral administration of soluble antigens can activate CD4+ IELs in an antigen-specific manner but induces hyporesponsiveness in the spleen. In addition, Th1-type cytokines produced by activated CD4+ IEL might provide a bystander effect on the cytotoxic activity of IELs.     

Fish oil supplementation in pregnancy modifies neonatal allergen-specific immune responses and clinical outcomes in infants at high risk of atopy: a randomized, controlled trial

BACKGROUND: There is growing interest in the potential role of anti-inflammatory n-3 polyunsaturated fatty acids (n-3 PUFAs) in the prevention of allergic disease. OBJECTIVE: We sought to determine whether maternal dietary supplementation with n-3 PUFAs during pregnancy could modify immune responses in infants. METHODS: In a randomized, controlled trial 98 atopic, pregnant women received fish oil (3.7 g n-3 PUFAs per day) or placebo from 20 weeks' gestation until delivery. Neonatal PUFA levels and immunologic response to allergens were measured at birth. RESULTS: Eighty-three women completed the study. Fish oil supplementation (n = 40) achieved significantly higher proportions of n-3 PUFAs in neonatal erythrocyte membranes (mean +/- SD, 17.75% +/- 1.85% as a percentage of total fatty acids) compared with the control group (n = 43, 13.69% +/- 1.22%, P <.001). All neonatal cytokine (IL-5, IL-13, IL-10, and IFN-gamma) responses (to all allergens) tended to be lower in the fish oil group (statistically significant only for IL-10 in response to cat). Although this study was not designed to examine clinical effects, we noted that infants in the fish oil group were 3 times less likely to have a positive skin prick test to egg at 1 year of age (odds ratio, 0.34; 95% confidence interval, 0.11 to 1.02; P =.055). Although there was no difference in the frequency of atopic dermatitis at 1 year of age, infants in the fish oil group also had significantly less severe disease (odds ratio, 0.09; 95% confidence interval, 0.01 to 0.94; P =.045). CONCLUSIONS: These data suggest a potential reduction in subsequent infant allergy after maternal PUFA supplementation. More detailed follow-up studies are required in larger cohorts to establish the robustness of these findings and to ascertain their significance in relation to longer-term modification of allergic disease in children.     

Th1-derived cytokine IFN-gamma is a potent inhibitor of eotaxin synthesis in vitro.

Eotaxin potentially plays an integral role in tissue eosinophilia. Inasmuch as Th2-derived cytokine IL-4 has been shown to stimulate eotaxin generation, we investigated here the effect of Th1-derived cytokine IFN-gamma on human eotaxin production. IFN-gamma but not -alpha or -beta potently inhibited tumor necrosis factor (TNF)-alpha-induced eotaxin generation by dermal fibroblasts. The inhibitory effect was unique to eotaxin, because production of IL-8 or monocyte chemoattractant protein (MCP)-1 protein was not affected by the treatment with IFN-gamma. Furthermore, the suppressive effect of IFN-gamma was not cell-type or stimulus specific. The level of eotaxin mRNA increased within 2 h after activation with TNF-alpha and continued to increase up to 72 h. IFN-gamma did not inhibit, but rather augmented the TNF-alpha-induced accumulation of mRNA in the early phase ( approximately 6 h). However, in the later phase, IFN-gamma completely prevented the subsequent elevation of eotaxin mRNA and sustained it at low levels. Although the protective effect of IFN-gamma against allergic inflammation has been assumed to result from its sole regulation of the proliferation of Th2-type T lymphocytes, these results imply that IFN-gamma can also directly act on stromal cells to inhibit eotaxin production and consequently intervene in eosinophil recruitment.     

Dietary n-3 PUFA affect TcR-mediated activation of purified murine T cells and accessory cell function in co-cultures

Diets enriched in n-3 polyunsaturated fatty acids (PUFA) suppress several functions of murine splenic T cells by acting directly on the T cells and/or indirectly on accessory cells. In this study, the relative contribution of highly purified populations of the two cell types to the dietary suppression of T cell function was examined. Mice were fed diets containing different levels of n-3 PUFA; safflower oil (SAF; control containing no n-3 PUFA), fish oil (FO) at 2% and 4%, or 1% purified docosahexaenoic acid (DHA) for 2 weeks. Purified (>90%) T cells were obtained from the spleen, and accessory cells (>95% adherent, esterase-positive) were obtained by peritoneal lavage. Purified T cells or accessory cells from each diet group were co-cultured with the alternative cell type from every other diet group, yielding a total of 16 different co-culture combinations. The T cells were stimulated with either concanavalin A (ConA) or antibodies to the T cell receptor (TcR)/CD3 complex and the costimulatory molecule CD28 (alphaCD3/alphaCD28), and proliferation was measured after four days. Suppression of T cell proliferation in the co-cultures was dependent upon the dose of dietary n-3 PUFA fed to mice from which the T cells were derived, irrespective of the dietary treatment of accessory cell donors. The greatest dietary effect was seen in mice consuming the DHA diet (P = 0.034 in the anova; P=0.0053 in the Trend Test), and was observed with direct stimulation of the T cell receptor and CD28 costimulatory ligand, but not with ConA. A significant dietary effect was also contributed accessory cells (P = 0.033 in the Trend Test). We conclude that dietary n-3 PUFA affect TcR-mediated by T cell activation by both direct and indirect (accessory cell) mechanisms.