UC Riverside

Microglial activation and the recruitment of peripheral macrophages are the hallmarks of CNS inflammation (reviewed in Carson et al. 2007). These two populations of effector macrophages cannot be reliably distinguished by histology, as CNS-resident microglia express most common macrophage markers. Although CNS-resident microglia and peripheral macrophages are characterized by similar expression of surface markers, a large body of research using irradiation bone marrow chimeras and flow cytometry assays have shown that these cells types are functionally distinct (Hickey and Kimura 1999, Schmid et al. 2009, Byram et al. 2004). This dissertation primarily focuses on the identification of distinct molecular pathways that regulate the process of microglial activation and macrophage recruitment in murine models of CNS inflammation.

Dysregulation of the polyamine biosynthetic pathway has been implicated in a variety of CNS neurodegenerative disorders like Alzheimer's disease, ischemia and amyotrophic lateral sclerosis (Clarkson et al. 2004, Kim et al. 2009, Morrison et al. 1995, Morrison et al. 1998, Virgili et al. 2006). However, the CNS intrinsic functions of polyamine biosynthesis have been not completely defined due to the existence of conflicting data generated from in vitro versus in vivo models of neuroinflammation. (Tjandrawinata et al. 1994, Szabo et al. 1994, Gilad et al. 2004, Soulet and Rivest. 2003). Using a murine model of acute, self resolving CNS inflammation, we demonstrate polyamine biosynthesis plays a dual role in the maintaining tissue homeostasis and sustaining CNS inflammatory responses. In-situ hybridization analysis of brain tissue from mice sacrificed post intracerebral LPS injection revealed that both, neuronal as well as non-neuronal cells rapidly upregulated the expression of ornithine decarboxylase (ODC) mRNA expression, the rate limiting enzyme in polyamine biosynthesis. Interestingly, inhibition of polyamine biosynthesis by intracerebral injections of DFMO (a suicide inhibitor of ODC), resulted in decreased LPS-induced expression of the macrophage chemoattaractant CCL2 as well as macrophage influx into the inflamed CNS. However, polyamine inhibition had no effects of microglial activation in response to CNS inflammation.

In addition to dysregulated polyamine metabolism, a factor in the development of neurodegenerative diseases like Alzheimer's disease, is the propensity of the aged CNS to mount chronic inflammatory responses. Healthy aging has been long regarded as a key factor contributing to microglial immune responses (Bilbo S.D. 2009, Frank M.J et al. 2008). To understand the differential effects of aging on the immune responses of the resident microglia and the infiltrating macrophages, we used healthy 3 months (young adult), 8 months (mid-aged adult) and 15 months (aged adult) mice to induce CNS inflammation. Using flow cytometry we showed that CNS resident microglia in the healthy, non-inflamed aged brain assumed a pro-inflammatory phenotype characterized by upregulated expression of MHC class II, CD40 and B7.2. In addition, the aged CNS displayed a modest increase in the infiltration by peripheral macrophages in the absence of any inflammation. Interestingly, the recruitment of peripheral macrophages to the CNS in response to CNS inflammation was not altered as a function of age. However, aging differentially affected the microglial immune responses upon LPS-induced neuroinflammation. CNS resident microglia from the aged CNS responded with a more robust upregulation of the classical pro-inflammatory activation markers, namely, MHC class II, CD40 and B7.2. In addition, we also observed an age associated increase in the upregulation of markers of alternative activation TREM2, but not mannose receptor (MMR). No such age-associated changes were seen in the immune response of the peripheral macrophages.

Lastly, we have also examined the regulation of a family of receptors, the TREM-like transcripts (TLTs) in response to inflammatory signals (LPS+IFNγ). The TLTs, which are members of the TREM family of receptors have been identified as putative receptors expressed by various cells of the myeloid lineage. Using in vitro and in vivo models of microglial and macrophage activation, we showed that TLT1, TLT2, TLT4 and TLT6 are differentially regulated by LPS+IFNγ in vitro and in vivo.

In summary, our data suggests that

1. The processes of microglial activation and recruitment of macrophages during CNS inflammation are regulated by distinct molecular mechanisms.

2. The CNS may be in active control of the immune responses mounted. Continuous, bi-directional communication between the neurons and the glial population may be critical in the shaping of glial immune responses.