Abstract
Intermittent hypoxia (IH) during sleep is a hallmark of sleep apnea, causing significant neuronal apoptosis, and cognitive and behavioral deficits in CNS regions underlying memory processing and executive functions. IH-induced neuroinflammation is thought to contribute to cognitive deficits after IH. In the present studies, we tested the hypothesis that IH would differentially induce inflammatory factor gene expression in microglia in a CNS region-dependent manner, and that the effects of IH would differ temporally. To test this hypothesis, adult rats were exposed to intermittent hypoxia (2 min intervals of 10.5% O2) for 8 hours/day during their respective sleep cycles for 1, 3 or 14 days. Cortex, medulla and spinal cord tissues were dissected, microglia were immunomagnetically isolated and mRNA levels of the inflammatory genes iNOS, COX-2, TNFα, IL-1β and IL-6 and the innate immune receptor TLR4 were compared to levels in normoxia. Inflammatory gene expression was also assessed in tissue homogenates (containing all CNS cells). We found that microglia from different CNS regions responded to IH differently. Cortical microglia had longer lasting inflammatory gene expression whereas spinal microglial gene expression was rapid and transient. We also observed that inflammatory gene expression in microglia frequently differed from that in tissue homogenates from the same region, indicating that cells other than microglia also contribute to IH-induced neuroinflammation. Lastly, microglial TLR4 mRNA levels were strongly upregulated by IH in a region- and time-dependent manner, and the increase in TLR4 expression appeared to coincide with timing of peak inflammatory gene expression, suggesting that TLR4 may play a role in IH-induced neuroinflammation. Together, these data indicate that microglial-specific neuroinflammation may play distinct roles in the effects of intermittent hypoxia in different CNS regions.
Highlights
Intermittent hypoxia (IH) during sleep is a hallmark feature of sleep apnea
We found that the inflammatory genes induced by IH differed in microglia from different CNS regions, and that the general temporal profiles of inflammatory gene expression varied in IH-sensitive and -resistant CNS regions
In cortical tissue homogenates (Fig. 1B), the mRNA levels of all genes showed an increase of approximately 2- fold after 3 days of IH and this increase was maintained at 14 days, they did not reach statistical significance as determined by ANOVA Tumor necrosis factor a (TNFa) came close (p = 0.055; inducible nitric oxide synthase (iNOS) p = 0.09; COX-2 p = 0.299; IL-1b p = 0.25; IL-6 p = 0.11)
Summary
Intermittent hypoxia (IH) during sleep is a hallmark feature of sleep apnea. IH induces significant cognitive and behavioral deficits that involve disruptions of connections among CNS regions underlying memory processing and executive function. Neuronal apoptosis in the CA1 region and cortex (as assessed by single stranded DNA immunostaining) indicates that apoptosis peaks between 1 and 2 days of IH, is significantly decreased by 7 days, and has returned to baseline levels by 14 days [1] This profile coincides with inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) gene expression in cortical homogenates where mRNA and protein levels peak at 1 day of IH and decline thereafter [10,11]. A proteomic study comparing the profile of IH-induced proteins in sensitive (CA1) and resistant (CA3) hippocampal regions showed that several heat shock proteins were upregulated by IH, and some differentially in CA1 versus CA3 [34], suggesting that IH may region- increase endogenous ligands for TLR4 that could contribute to microglial activation and IH-induced neuroinflammation in this region. Microglial TLR4 mRNA levels were strongly upregulated in a region- and time-dependent manner, and its expression often coincided with increased inflammatory gene expression, suggesting that TLR4 may play a role in IH-induced neuroinflammation
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