Apoptosis of olfactory sensory neurons (OSNs) induced by olfactory bulbectomy (OBX) leads to the activation of resident macrophages within the olfactory epithelium (OE). These macrophages phagocytose degenerating OSNs and secrete chemokines, which recruit additional macrophages into the OE, and cytokines/growth factors, which regulate basal cell proliferation and differentiation and maturation of OSNs. In this study we apply for the first time the use of liposome-encapsulated clodronate to selectively deplete macrophages during the OSN degeneration/regeneration cycle in order to elucidate the role(s) of macrophages in regulating cellular mechanisms that lead to apoptosis and neurogenesis. Mice were injected intranasally and intravenously with either liposome-encapsulated clodronate or empty liposomes prior to and after OBX or sham OBX. At 48 hours after surgery the numbers of macrophages in the OE of both sham and OBX clodronate-treated mice were significantly reduced compared to liposome-treated controls (38% and 35%, respectively, P < 0.05). The reduction in macrophage numbers was accompanied by significant decreases in OE thickness (22% and 21%, P < 0.05), the number of mOSNs (1.2- and 1.9-fold, P < 0.05), and basal cell proliferation (7.6- and 3.8-fold, P < 0.005) in sham and OBX mice, respectively, compared to liposome-treated controls. In OBX mice there was also increased immunoreactivity for active caspase-3 in the OE and olfactory nerves of clodronate-treated OBX mice compared to liposome-treated controls. These results indicate that macrophages modulate the OSN population in the normal and target-ablated murine OE by influencing neuronal survival and basal cell proliferation, resulting in neurogenesis and replacement of mature OSNs.

Protein kinases are critical modulators of a variety of cellular signal transduction pathways, and abnormal phosphorylation events can be a cause or contributor to disease progression in a variety of disorders. This has led to the emergence of protein kinases as an important new class of drug targets for small molecule therapeutics. A serine/threonine protein kinase, p38α mitogen-activated protein kinase (MAPK), is an established therapeutic target for peripheral inflammatory disorders because of its critical role in regulation of proinflammatory cytokine production. There is increasing evidence that p38α MAPK is also an important regulator of proinflammatory cytokine levels in the central nervous system, raising the possibility that the kinase may be a drug discovery target for central nervous system disorders where cytokine overproduction contributes to disease progression. Development of bioavailable, central nervous system-penetrant p38α MAPK inhibitors provides the required foundation for drug discovery campaigns targeting p38α MAPK in neurodegenerative disorders.

Target ablation [removal of the olfactory bulb (OBX)] induces apoptotic death of olfactory sensory neurons (OSNs) and an immune response in which activation and recruitment of macrophages (ms) into the olfactory epithelium (OE) occupy a central role. Ms phagocytose apoptotic neurons and secrete cytokines/growth factors that regulate subsequent progenitor cell proliferation and neurogenesis. Scavenger receptor A (SR-A) is a pattern recognition receptor that mediates binding of ms to apoptotic cells and other relevant immune response functions. The aim of this study was to determine the impact of the absence of SR-A on the immune response to OBX. The immune response to OBX was evaluated in mice in which functional expression of the m scavenger receptor (MSR) was eliminated by gene disruption (MSR-/-) and wild-type (wt) mice of the same genetic background. OBX induced significant apoptotic death of mature OSNs in the two strains. However, subsequent m infiltration and activation and progenitor cell proliferation were significantly reduced in MSR-/- vs. wt mice. Gene expression profiling at short intervals after OBX demonstrated significant differences in temporal patterns of expression of several gene categories, including immune response genes. Many immune response genes that showed different temporal patterns of expression are related to m function, including cytokine and chemokine secretion, phagocytosis, and m maturation and activation. These studies suggest that impairment of the immune response to OBX in the OE of MSR-/- mice most likely resulted from decreased m adhesion and subsequent reduced infiltration and activation, with a resultant decrease in neurogenesis.

Background:Cluster analyses are used to analyze microarray time-course data for gene discovery and pattern recognition. However, in general, these methods do not take advantage of the fact that time is a continuous variable, and existing clustering methods often group biologically unrelated genes together.

Resident and recruited olfactory epithelial macrophages participate in the regulation of the survival, degeneration, and replacement of olfactory sensory neurons (OSNs). We have reported that liposome-encapsulated clodronate (Lip-C) induced selective and statistically significant depletion of macrophages in the OE of sham and 48 h OBX mice (38 and 35%, respectively) that resulted in increased OSN apoptosis and decreased numbers of mature OSNs and proliferating basal cells compared to controls (Lip-O). The aim of this study was to identify molecular mechanisms by which the selective depletion of macrophages in the OE resulted in these cellular changes by using a microarray expression pattern analysis. A 2ϫ2 ANOVA identified 4,085 overall significantly (P Ͻ 0.01) regulated genes in the OE of Lip-O and Lip-C sham and 48 h OBX mice, and further statistical analysis using pairwise comparisons identified 4,024 genes that had either a significant (P Ͻ 0.01) treatment main effect (n ϭ 2,680), group main effect (n ϭ 778), or interaction effect (n ϭ 980). The mean hybridization signals of immune response genes, e.g., Cxcr4, and genes encoding growth factors and neurogenesis regulators, e.g., Hdgf and Neurod1, respectively, were primarily lower in Lip-C mice compared with Lip-O mice. Apoptosis genes, e.g., Bak1, were also differentially regulated in Lip-C and/or OBX mice. Expression patterns of selected genes were validated with real-time RT-PCR; immunohistochemistry was used to localize selected gene products. These results identified the differential regulation of several novel genes through which alternatively activated macrophages regulate OSN progenitor cell proliferation, differentiation, and maturation, and the survival of OSNs.clodronate; liposomes; microarray; immune response THE DYNAMIC ENVIRONMENT of the olfactory epithelium (OE), where olfactory sensory neurons (OSNs) undergo apoptosis and are replaced from a pool of basal progenitor cells throughout life (9,41,49), continues to provide insight into mechanisms of neuroprotection, neuronal apoptosis, and neurogenesis. More specifically, recent studies (6,23,33) on the interaction between the immune system and neurogenesis in the OE have strengthened the case for the participation of macrophages in the regulation of OSN survival and the proliferation, differentiation, and maturation of OSN basal progenitor cells in the unperturbed OE and during OE remodeling as a result of olfactory bulbectomy (OBX)-induced OSN apoptosis.