| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
|
FJ
EXPRESS SUMMARY ARTICLE The Full-length version of this article is also available, published online May 26, 2005 as doi:10.1096/fj.05-3776fje. |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
,1
* Division of Nutritional Sciences and
Department of Animal Sciences, University of Illinois, Urbana, Illinois, USA
1Correspondence: E-mail: rwjohn{at}uiuc.edu
SPECIFIC AIMS
Acute cognitive impairment (i.e., delirium) is common in elderly emergency department patients and frequently results from infections unrelated to the central nervous system. Since activation of the peripheral innate immune system induces brain microglia to produce inflammatory cytokines that are responsible for behavioral deficits, the aim of this study was to determine whether aging exacerbated neuroinflammation and sickness behavior after peripheral injection of lipopolysaccharide (LPS).
PRINCIPAL FINDINGS
1. Transcriptional profile in brain of aged mice after activation of the peripheral innate immune system
The transcriptional profile in brain of adult and aged mice was compared after i.p. injection of either saline or lipopolysaccharide (LPS). Total RNA was isolated from brain obtained 4 h after LPS administration and differential gene expression was determined by microarray analysis. A pairwise comparison of adult/saline subjects vs. aged/saline subjects showed a total of 458 genes were differentially expressed in brain of aged mice compared with adults. A notable change in the transcriptional profile was evident in the inflammation category. There were 38 genes associated with inflammation that were differentially expressed between adult and aged brain: 34 were up-regulated and only 4 were down-regulated in the aged brain. Genes that were up-regulated in the aged brain were associated with the complement cascade (complement components C1q, C3, C4, and serum amyloid), defense response (CD68, CD44, CD83, MHC class II antigens A and E, MHC class I, GFAP, lysozyme, and lymphotoxin B), and oxioreductase activity (cytochrome P450, thioredoxin interacting protein, and glutathione peroxidase). These results demonstrate an age-associated increase in markers indicative of primed or activated microglia and suggest a transition to a neuroinflammatory state, even in aged, healthy animals.
The primary objective of the microarray study was to determine whether activation of the peripheral innate immune system with LPS induced a different inflammatory gene expression profile in brain of aged mice compared with adults. There were a number of up- and down-regulated genes in brain 4 h after peripheral LPS for both adult and aged mice. Analysis of the fold change for each gene transcript revealed a subset of 415 genes whose activity was influenced by a LPS x age interaction: a notable change in inflammatory genes was again evident. There were genes that were up- or down-regulated by LPS, but that were affected more so in aged mice than in adults. For example, genes such as C1q-like, IL-1ß, IL-6 signal transducer, oncostatin M receptor, and Janus kinase 1 were up-regulated to a greater extent; and growth hormone, chemokine ligand 9, CREB binding protein, and metallothionein 1 and 2 were down-regulated more so in the brain of aged mice after LPS. There were genes that were affected by LPS in aged mice but not adults. For example, transcripts for amyloid ß precursor protein and binding proteins, cathepsin D, and interferon gamma receptor 2 were only increased after LPS in brain of aged mice, whereas superoxide dismutase and NADPH dehydrogenase were decreased in the brain of aged mice but not adults after LPS.
2. Neuroinflammation is exacerbated in aged mice after activation of the peripheral innate immune system
From the same RNA pool used in the microarray study, MHC class II antigen E, IL-1ß, and IL-6 mRNA were quantified by real-time PCR. The steady-state level of MHC class II antigen E was increased due to age (P<0.001) but was not increased by LPS (P=0.6). The steady-state level of both cytokine mRNAs was increased due to age (IL-1ß, P<0.03 and IL-6, P<0.01) and LPS (P<0.001, for both IL-1ß and IL-6). However, the LPS-induced increase in IL-1ß and IL-6 mRNA was greater in brains of aged mice than adults (IL-1ß, P<0.04 and IL-6, P<0.01). Whereas LPS treatment increased IL-1ß and IL-6 mRNA in brain of adults more than 22-fold, both cytokines increased more than 43-fold in brain of aged mice. To determine whether the increases in cytokine mRNA after LPS were paralleled by increases in cytokine protein, total protein was extracted from brains of adult and aged mice and assayed for IL-1ß and IL-6 (Fig. 1
A, B). Both IL-1ß and IL-6 were increased due to age (IL-1ß, P<0.008 and IL-6, P<0.002) and LPS (P<0.001 for both IL-1ß and IL-6). The LPS-induced increase in IL-6 was greater in brains of aged mice than adults (P<0.002). The increase in IL-1ß caused by LPS was also greater in brains of aged mice (P<0.01). Circulating levels of IL-1ß and IL-6 were determined but the exaggerated neuroinflammatory response was not reliably paralleled by changes in peripheral blood cytokines. Since peripheral administration of LPS induces oxidative stress in brain of young adult rats and reactive oxygen species may induce inflammatory cytokine production in microglia, we measured lipid peroxidation in brain. We found that lipid peroxidation was higher in brain of aged mice than adults (P<0.001). Lipid peroxidation was increased by LPS (P<0.01) but the magnitude of the response tended to be age dependent (P<0.09). Lipid peroxidation in brain after peripheral injection of LPS was markedly higher in aged mice than adults. Taken together, these data suggest signs of neuroinflammation emerge in old age, and that healthy aged individuals suffer an exaggerated neuroinflammatory reaction when the peripheral innate immune system is stimulated.
|
3. Behavioral deficits and neuroinflammation are prolonged in aged mice after activation of the peripheral innate immune system
To determine whether a more severe sickness behavior syndrome is a functional consequence of the exaggerated neuroinflammatory response in aged mice, locomotor activity and motivation to engage in social behavior were assessed in adult and aged mice at several time points after i.p. injection of LPS. Food intake and change in body weight were determined for the 24 h period after LPS. The effects of age and LPS on social behavior are shown in Fig. 2
. In this test, a juvenile was introduced into a subject’s home cage for 10 min. Because a novel juvenile was used for each test, mice generally maintain motivation for social investigation. This point was evident since social behavior was not depressed compared with baseline at anytime in either adult or aged mice given saline. However, social behavior was reduced due to LPS (P<0.001) and age (P<0.03); at 8 (P<0.02) and 24 h (P<0.001) postinjection, the depression in social behavior was greater in aged mice than adults. Whereas adult mice began to recover at 8 h and their social behavior was restored to normal by 24 h, aged mice still suffered a 57% deficit in motivation at 24 h (P<0.001). The effects of age and LPS on locomotor behavior were also assessed. Because this test measured general motor activity (i.e., no motivational stimulus was provided), it was not surprising that locomotor activity was decreased due to age (P<0.01) prior to LPS administration. At 2, 4, 8, and 24 h after injection, locomotor activity was decreased due to age (P<0.001) and LPS (P<0.001). However, at 8 (P<0.05) and 24 h (P<0.001), the depression in locomotor activity was greater in aged mice than in adults, which is similar to what was observed for social behavior. The delayed recovery in social and locomotor behaviors was further corroborated by a greater reduction in food consumption (P<0.04) and body weight (P<0.02) for aged mice given LPS compared with adults.
|
Immediately after the behavioral tests, mice were killed and brains were collected to assess markers of inflammation. On the one hand, the steady-state levels of IL-1ß mRNA remained elevated in both adult and aged mice 24 h after LPS injection, which is of note since only aged mice suffered behavioral deficits at the time. On the other hand, the steady-state level of IL-6 mRNA was elevated in brain of aged mice only (P<0.03), 24 h after LPS injection. Cytokine protein was assayed in extracts from whole brain. Whereas IL-1ß was not detectable, IL-6 was still elevated in brain of aged mice 24 h after LPS (P<0.05). Furthermore, 24 h after peripheral administration of LPS, lipid peroxidation was higher in brain of aged mice compared with adults (P<0.05). Collectively, these data suggest that the prolonged deficits in behavior of aged mice after stimulation of the peripheral innate immune system are related to an exaggerated and protracted neuroinflammatory response.
CONCLUSIONS AND SIGNIFICANCE
Brain microglia are activated in certain stages of neurodegenerative diseases and in young adult subjects when the peripheral innate immune system is stimulated with LPS. Activated microglia produce inflammatory cytokines that are partially responsible for the progression of neurodegenerative disease and for LPS-induced sickness behavior. Primed microglia are morphologically similar to activated microglia, but do not appear to produce appreciable levels of inflammatory cytokines in this state. They are, however, hyperresponsive to secondary stimuli and so can produce an exaggerated cytokine response when further provoked. Because aging may prime microglia to set the stage for an exaggerated response to a secondary stimulus, in this study we investigated whether aging exacerbated neuroinflammation and behavioral deficits that are associated with stimulation of the peripheral innate immune system. The transcriptional profile indicated the presence of primed or activated microglia and increased inflammation in aged brain. The most important finding was that the inflammatory response induced by peripheral LPS was amplified and prolonged in the aged brain. Behavioral deficits induced by LPS were protracted in the aged. Collectively, these results suggest that activation of the peripheral innate immune system leads to exacerbated neuroinflammation in the aged. This dysregulated link between the peripheral and central innate immune system is likely to be involved in the severe behavioral deficits that frequently occur in older adults with systemic infections.
Understanding the mechanisms involved in an exaggerated neuroinflammatory response and increased sickness behavior in the aged is important for several reasons (Fig. 3
). First, aging is associated with immunosenescence and the elderly are more susceptible to infection. Second, many of the behavioral consequences of infection, if prolonged, can have devastating effects. For example, reduction of appetite, which contributes to loss of lean body mass, is positively correlated to increased morbidity and mortality and delayed recovery from injury or surgery. There is also an increased prevalence of acute cognitive impairment in elderly emergency department patients as a result of infections unrelated to the CNS. Moreover, acute cognitive impairment results in a failure of self-care and is associated with increased hospitalization and delayed recovery from illness. Inflammatory cytokines have been linked to depressive withdrawal from normal activities. Finally, because inflammatory cytokines are involved in neurodegenerative disease, it is possible that infection accompanied by increased neuroinflammation is a predisposing factor for neurodegenerative diseases in elderly patients.
|
FOOTNOTES
To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.05-3776fje;
This article has been cited by other articles:
|
|
 |
|
  S. Jang, K. W. Kelley, and R. W. Johnson Luteolin reduces IL-6 production in microglia by inhibiting JNK phosphorylation and activation of AP-1 PNAS, May 27, 2008; 105(21): 7534 - 7539. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
significantly different from adult LPS.