Immune Response In Aged Mice Research Articles

A Low Blood Ethanol Level Is Associated with Improved Cytokine Production in Aged Mice after Traumatic Injury

Many trauma patients have been consuming alcohol at the time of injury. Although high concentrations of alcohol are correlated with poor outcome, few studies have examined the effects of low levels of alcohol. We examined the effects of low alcohol exposure after burn injury using a murine model. Three- and 18-month-old mice were given ethanol or saline 30 minutes before a 15% total body surface area burn injury. Twenty-four hours after injury, cellular immune responses, including delayed-type hypersensitivity response and splenocyte proliferation were examined, along with production of interferon-gamma, interleukin (IL)-2, and IL-4. Alcohol administration resulted in a significant increase in interferon-gamma in the aged, but not young, burn-injured mice. Likewise, slight increases in IL-2, IL-4, and the delayed-type hypersensitivity response were observed. Low levels of ethanol at the time of injury are associated with partial restoration of immune responses in aged mice.

Caloric Restriction Decreases Survival of Aged Mice in Response to Primary Influenza Infection

Caloric restriction (CR) extends life span of healthy rodents compared to those fed ad libitum. Previous studies have shown positive effects of CR on the immune response of aged mice after influenza immunization. To extend these studies, a mouse model of CR was used to determine if CR could modulate primary responses of aged mice to influenza. Although CR delayed the age-related decrease in mitogen-induced lymphoproliferation of aged mice, in stark contrast, CR decreased survival, increased virus titers, and reduced natural killer cell activity in lungs of aged mice after primary influenza infection. Thus, CR has differential effects on immunity of aged mice, as general indices of immune response are maintained, but primary responses to influenza infection are impaired. This suggests that, although CR may positively affect many long-term parameters of aging, increased susceptibility after primary exposure of aged mice to virus, such as influenza, may not be correctable by CR.

Immunostimulant patch enhances immune responses to influenza virus vaccine in aged mice.

Improvement in the immune response to influenza virus vaccination in the elderly represents the primary unmet need in influenza virus vaccination. We have shown that topical application of immunostimulating (IS) patches containing heat-labile enterotoxin of Escherichia coli (LT) enhances immune responses to injected vaccines. We extend these findings and show that LT-IS patch application enhances the antibody responses to influenza virus vaccination in both young and aged mice. LT-IS patches markedly increased influenza virus-specific immunoglobulin G (IgG), hemagglutination inhibition antibody, mucosal antibody, and T-cell responses. The magnitude of the immune responses in aged mice receiving an LT-IS patch was equivalent to or greater than that of the immune responses in young mice given vaccine alone. These results suggest that addition of an LT-IS patch may compensate for the deficient immune function seen in the aged in response to influenza virus vaccination. Therefore, use of an LT-IS patch could be a new, safe, and simple immunization strategy that may significantly improve the outcome of influenza virus vaccination in the elderly.

CpG-DNA stimulates cellular and humoral immunity and promotes Th1 differentiation in aged BALB/c mice

We examined whether CpG-DNA could be used as adjuvant to induce a T helper cell type-1 (Th1) immunity in aged BALB/c mice that showed a Th2 polarization. Bordetella pertussis and complete Freund's adjuvant (CFA) were used as well. Immunization with ovalbumin (OVA)/CpG-DNA showed that the immunoglobulin G (IgG)2a/IgG1 ratio and OVA-specific T cell response were similar in young and aged mice. OVA/CpG-DNA induced the secretion of interferon-gamma (IFN-gamma) and absence of interleukin (IL)-5. Similar results were found in mice immunized with OVA/CFA. When mice were immunized with OVA/B. pertussis, we found that the IgG2a/IgG1 ratio and OVA-specific T cell response were lower in aged mice and elicited IFN-gamma and IL-5. In vitro CpG-DNA stimulated antigen-presenting cells to display IL-12 and up-regulate the expression of major histocompatibility complex class II and B7-2 on B cells as efficiently in aged as in young mice, but the up-regulation of B7-1 was stronger in aged mice. The findings demonstrate that CpG-DNA is able to induce a young-like Th1 specific immune response in aged mice.

CpG DNA functions as an effective adjuvant for the induction of immune responses in aged mice

The present studies demonstrate that the immunization of aged mice with Diphtheria toxoid in formulations containing unmethylated immunostimulatory CpG motifs, promotes the successful development of immune responses that are qualitatively and quantitatively comparable to those induced in young animals vaccinated in a similar manner. Aged mice given vaccines containing CpG oligodeoxynucleotides (ODNs) expressed primary and secondary systemic humoral immune responses having isotype profiles consistent with an enhancement in Th-1 type immunity. The ability to generate common mucosal immunity was also restored in aged animals given CpG ODN-containing vaccines. Dendritic cells (DCs) were determined to represent one of the cellular targets of CpG ODN activities in aged mice since restoration of immune function was observed when DCs from aged donors were pulsed with antigen and CpG ODNs, prior to injection into syngeneic young adult or aged recipients. Interestingly, antigen-pulsed DCs from young donors were fully capable of stimulating immune responses following their injection into syngeneic young adult or aged hosts, without a need for exposure to CpG ODNs. Although the mechanism(s) by which CpG DNA exerts its beneficial adjuvant effects on the aged immune system remains unclear, our findings suggest that the incorporation of CpG ODNs into vaccine formulations provided to the aged could prove useful in the development of more effective vaccines for the elderly.

Immunogenicity and Efficacy of a DNA Vaccine in Aged Mice

The immunogenicity and protective efficacy of a DNA vaccine encoding the circumsporozoite protein of the Plasmodium yoelii malaria parasite was evaluated in young (2 months) versus aged (>26 months) BALB/c mice. The primary and secondary humoral immune response of aged mice was 19- and 7-fold lower, respectively, than that of similarly treated young animals (p < .01). Cytotoxic T lymphocyte activity in aged mice was also lower than in younger animals. The vaccine response of aged animals was characterized by a 6-fold increase in interleukin-4 and a 3-fold increase in interferon-gamma (IFN-gamma) secreting cells, whereas in young animals immunization only stimulated the production of the type 1 cytokine IFN-gamma. Overall, 80% of young vaccinated mice were protected from subsequent challenge with live malaria sporozoites whereas only 40% of aged mice were protected. These results are the first to demonstrate that DNA vaccination induces less effective immunity in aged than young animals.

Melatonin: immune modulation of spleen cells in young, middle-aged and senescent mice

We examined the effects of melatonin on the immune response in aged mice in an in vitro model system. Spleen cells were removed from 2 month old, 11 month old, and 19 month old C57/BL6 male mice and incubated with melatonin concentrations ranging from 50 to 5000 ng melatonin/ml. All age groups demonstrated increased ability to proliferation in response to mitogenic stimulation when melatonin and mitogen were added at the same time, and we demonstrated that 11 month old and 19 month old mice exhibited a greater increase in proliferation response than the young 2 month old mice. We further examined whether this increased responsiveness was due to increased levels of IL-2 and found that while there was a significant age- dependent difference in the levels of IL-2 in the different age groups, melatonin had no effect on IL-2 levels in any age groups despite the fact that proliferation increased in all groups. The underlying mechanism of melatonin action is being investigated further.

Effect of aging on murine macrophages. Diminished response to IFN-gamma for enhanced oxidative metabolism.

The ability of macrophages to secrete reactive oxygen intermediates, as well as reactive nitrogen intermediates, correlates closely with their capacity to perform two critical effector functions: intracellular killing of microorganisms and lysis of tumor cells. In this study, age-associated changes in the ability of caseinate-elicited peritoneal macrophages to release hydrogen peroxide were determined. Macrophages from aged BALB/c mice produced 50% less hydrogen peroxide than those from young mice in response to PMA or opsonized zymosan. In contrast, the production of macrophage-activating cytokines including IFN-gamma was not diminished in splenocyte supernatants from the aged group. Furthermore, no difference was detected in surface expression of IFN-gamma receptor in old and young mice. Macrophage responses to IFN-gamma, however, declined with aging. In vitro, IFN-gamma-induced release of hydrogen peroxide and nitric oxide was 50% lower in old mice than in young mice. IFN-gamma-induced tyrosine phosphorylation of MAPK, an early activation event, was undetectable in macrophages from the aged mice. These data demonstrate that diminished responses of macrophages to activating signals are one aspect of the impaired immune response in aged mice.

Influences of Dietary Restriction on Immunity to Influenza in Aged Mice

Our previous studies of the immune response of aged mice inoculated with influenza A virus revealed age-related decreases in antigen-specific cytotoxic T-lymphocyte function (CTL), T-cell proliferation, IL-2 production, antigen presentation, and antibody production. Because dietary restriction (DR) of rodents has been shown to extend maximum life span, delay the onset of tumors, and improve many immunologic parameters in aged animals, we tested the effect of such a regimen on the immune response to the influenza virus. We report that DR significantly inhibited the age-related decline in antigen presentation and T-cell proliferation. It also reduced the decline in antibody production to the virus. This is the first demonstration of improved immunity to an actual infectious agent resulting from DR. The improvement appears to be on a number of levels and to reflect more than one operative mechanism.

Vitamin E supplementation suppresses prostaglandin E 21 synthesis and enhances the immune response of aged mice

The potential for vitamin E to modulate prostaglandin metabolism and alter immune response in aged mice was studied. Semi-purified diets containing 30 ppm or 500 ppm dl-α-tocopheryl acetate (VitE) were fed for 6 weeks to young (3 months) and old (24 months) C57BL/6J mice. Delayed hypersensitivity skin test to DNFB and the proliferative response of splenocytes to T- and B-cell mitogens were assessed. Ex-vivo synthesis of Prostaglandin E 2 (PGE 2) was measured in spleen homogenates and serum vitamin E was measured by HPLC. Vitamin E supplementation of aged mice enhanced percent ear swelling to DNFB as well as the mitogenic response of splenocytes to Con A and LPS ( P < 0.05). Furthermore, spleen homogenates from old mice fed 30 ppm VitE had a significantly higher PGE 2 level than young mice fed 30 ppm VitE and old mice fed 500 ppm VitE (3.20 ± 0.07 μg/g vs. 2.60 ± 0.08 and 2.3 ± 0.10, respectively). Thus, the vitamin E enhanced immune response of aged mice appears to be mediated by decreased prostaglandin synthesis.

Proliferative and cytotoxic immune functions in aging mice. III. Exogenous interleukin-2 rich supernatant only partially restores alloreactivity in vitro

The ability of exogenous interleukin-2 (IL-2) rich supernatant to restore the defective T cell mediated immune functions of spleen cells from aged C57BL/6 mice was analyzed. Addition of IL-2 rich supernatant to allogeneic mixed lymphocyte cultures (MLC) resulted in an increase in the proliferative response of spleen cells from both young and old mice. The MLC response of cells from old mice was, however, not restored to the level of proliferation seen with splenocytes from young animals. In studying the generation of specific T cell suppressor function, it was found that IL-2 rich supernatant enhanced this function only for spleen cells from those aged animals which demonstrated a defective response in its absence. The response of these mice was thereby restored to the normal level. The response of cells from young control animals and aged mice with normal suppressor activity was not affected by the addition of IL-2 rich supernatant. We conclude that decreased IL-2 production constitutes a functionally important aspect, but is by no means the only defect in the immune response of aged mice. The results also suggest that responsiveness to IL-2 is less affected by age than lymphokine production.

Rescue of the tumor-specific immune response of aged mice in vitro.

In contrast to young mice, old mice fail to reject a transplanted challenge of the highly immunogenic, ultraviolet light-induced tumor 1591-RE. Old mice also fail to mount a cytolytic tumor-specific immune response in vivo, and spleen cells of old mice are defective in their ability to generate tumor-specific T cells in vitro. In the present study we report the results of cell culture mixing experiments that show that this deficiency is due to a decreased responsiveness of the Lyt-2+ tumor-specific cytolytic T cell precursors of the old animals. We also demonstrate with limiting dilution analysis that the defective responsiveness is not due to a clonal exhaustion of the precursors. In fact, the responsiveness could be restored in vitro by culturing the spleen cells of old animals at high density or by the addition of excess Lyt-1-/Lyt-2-/2000-rad-resistant spleen cells from young or old mice. Our results suggest that the rescue of tumor immunity in old individuals may be possible, perhaps by educating effector cells in vitro for adoptive immunotherapy.

Hematological and Immunological Changes with Age

The development of both myeloid cells and immunocytes from a common stem cell precursor, their differentiated pathways, functions and the factors which modulate them are briefly reviewed here. The age‐related decline (or lack of it) for the several cell lines is outlined, as well as the effect of this upon life processes and homeostasis. The results from two approaches to extending high levels of immune responses in aged mice are reviewed.

The immune response of aged mice to influenza: Diminished T-cell proliferation, interleukin 2 production and cytotoxicity

In view of the fact that T lymphocytes play a key role in the recovery from viral infections, and that many immune functions involving T cells show a decline in aged individuals, we examined the ability of old mice to mount a T-cell response to influenza virus. Both primary and secondary cytotoxic T-lymphocyte (CTL) responses following in vivo immunization with influenza virus were minimal in old mice at the time point of peak response in young mice. Analysis of CTL activity generated in an in vitro microculture system revealed both a diminished proliferative response and a decrease in Interleukin 2 (IL-2) production in “old” compared to “young” cultures following stimulation with virus. Furthermore, addition of exogenous IL-2 at the initiation of the culture period augmented both subtype-specific and A-strain cross-reactive CTL activity of old spleen cells. Finally, exogenous IL-2 increased the lower proliferative response of the old cultures. These findings, if also true for elderly humans, might have broad clinical relevance to the problem of suitable prophylactic approaches to influenza infection in the elderly.

Effects of chronic tobacco smoke exposure on immune responses in aged mice.

Balb/c mice were exposed to fresh smoke, equivalent to 30 high-tar filtered cigarettes/day, for 83 wk or less. After groups of mice were subjected to 56, 64, 72, and 80 wk of tobacco smoke (TS) exposure, serum antibody responses to both T-dependent and T-independent antigens, numbers of plaque-forming cells, spleen cell and lymph node PHA responses, and systemic clearance of normal and opsonised antigen were investigated. Aged TS-exposed mice showed some deterioration of immune responsiveness with age, but results varied; in extreme old age the immunological responsiveness of TS-exposed mice was similar to that of age-matched controls. In addition, groups of mice inoculated with fibrosarcoma cells after 78 wk TS-exposure did not show significantly greater susceptibility to the early development of pulmonary metastases.

Partial reactivation of impaired immune competence in aged mice by synthetic thymus factors

Our results clearly show the profound impairment of the humoral, hemolytic, primary, immune response in aged mice (22 months) as compared with this response in young (10 weeks) mice. A partial but significant reactivation of the age-determined impairment of the immunological responsiveness results from subcutaneous administration of two newly synthesized thymus factors and one analog. These synthetic thymus peptide factors correspond to factors previously isolated and identified in thymus glands or blood.

Immune response in aged mice exposed to lead.

Mice were exposed to 0, 13, or 1,300 ppm lead in drinking water for 18 months. The immunological assays examined were mitogen (lipopolysaccharide E. coli, concanavalin A, and phytohemagglutinin-P) stimulation of lymphocytes; erythrocyte-antibody (EA), erythrocyte-antibody-complement (EAC), and phagocytosis of macrophages; and EAC of splenic lymphocytes. As measured by the majority of these assays, the low dosage (13 ppm) of lead tended to stimulate certain immune responses (lymphocyte mitosis, EA, and EAC) while the high dosage (1,300 pm) did not provoke an appreciable alteration. The results were interpreted by comparing data on aged mice with data on young adult mice. It was apparent from this comparison that the aged mice were naturally immunosuppressed. Therefore, the results obtained from lead-exposed mice were unpredictable.