Sleep loss disrupts pericyte-brain endothelial cell interactions impairing blood-brain barrier function
Introduction
Pericytes are perivascular cells that surround the endothelium and contribute to blood-brain barrier stabilization through the induction of endothelial tight junctions and establishment of gap junctions with brain endothelial cells (Fujimoto, 1995; Armulik et al., 2010). Pericytes regulate capillary diameter, cerebral blood-flow and control the entry of immune cells to the central nervous system during angiogenesis and blood-brain barrier maturation (Armulik et al., 2005, Shimizu et al., 2008, Stark et al., 2013, Hall et al., 2014 Neuhaus et al., 2017). Reduced pericyte coverage induces microvascular defects (Benjamin et al., 1998; Tarallo et al., 2012) and blood-brain barrier dysfunction (Vates et al., 2010; Bell et al., 2010, Rustenhoven et al., 2017).
The platelet-derived growth factor receptor-β (PDGFR-β) is expressed predominantly in pericytes (Armulik et al., 2005) and its endothelium-secreted ligand, PDGF-B, has been proposed as the signaling molecule that modulates pericyte-endothelial cell crosstalk (Lebrin et al., 2010, Armulik et al., 2011). Animal models with impaired PDGFB-PDGFR-β signaling pathway present brain microvascular dysfunction (Villaseñor et al., 2017, Arango-Lievano et al., 2018) and perinatal death (Leveen et al., 1994). In addition, monocultures of endothelial cells present lower transendothelial electrical resistance (TEER) and higher permeability to low- and large-molecular weight tracers than co-cultures of brain endothelial cells and pericytes (Hayashi et al., 2004, Nakagawa et al., 2009, Wisniewska-Kruk et al., 2012). These results underline the importance of the cell-cell interactions between brain endothelial cells and pericytes for the maintenance of the blood-brain barrier physiology.
Previous research has shown that sleep loss increases blood-brain barrier permeability to 10 kDa and 70 kDa FITC-dextrans, sodium fluorescein, and Evans blue (Gomez-Gonzalez et al., 2013, He et al., 2014, Hurtado-Alvarado et al., 2016a, Hurtado-Alvarado et al., 2017). Concomitant to the entry of exogenous molecules, there is a change in tight junction morphology in brain microvessels (MVs), characterized by a decrease in tight junction protein expression and the presence of wide gaps at the interendothelial junctions. In addition, our group presented ultrastructural evidence of potential pericyte detachment from the capillary wall in the hippocampus of 10-day sleep-restricted rats (Hurtado-Alvarado et al., 2018). The maintenance of the barrier properties of the blood-brain barrier seem to depend on rapid-eye movement (REM) sleep. Using the multiple platform technique for 10 days, which fully suppresses REM sleep phase and a slightly reduces non-REM sleep (around 30%), Gomez-Gonzalez et al. (2013) found large deranges in blood-brain barrier function. He et al. (2014) found that a protocol reducing 20% non-REM sleep time and greatly reducing REM sleep time (up to 80%) during 6 days of sleep restriction, induces modest changes in the barrier phenotype. In both cases chronic sleep loss induces neuroinflammation, as depicted by the increase in the expression of glial markers (eg. Iba1 and GFAP) in several brain regions (Hurtado-Alvarado et al., 2016a, Hurtado-Alvarado et al., 2018, Manchanda et al., 2018) and by the increase in the cyclooxygenase 2 (COX-2) mRNA in brain homogenates (He et al., 2014). Other reports have shown that sleep restriction for 21 days also increases mRNA expression of tumor necrosis factor (TNF)-α in the hippocampus (Manchanda et al., 2018).
Both, the peripheral and the central low-grade inflammatory status during chronic sleep loss (Hurtado-Alvarado et al., 2016b), may affect the intercellular interactions between pericytes and brain endothelial cells, but the mechanisms involved in the regulation of blood-brain barrier function during sleep loss are only recently being studied. Thus, we aimed to determine the effect of sleep restriction on pericyte-brain endothelial cell interactions and their consequences on blood-brain barrier function, characterizing the inflammatory mediators that might be participating in this event.
Section snippets
Animals
Three-month old male Wistar rats were used and randomly divided in two groups: sleep restriction group (SR, n = 33) and intact control group (CON, n = 33). Animals were housed in standard conditions in our laboratory vivarium under a 12 h light-dark cycle (lights on at 11:00 am) at room temperature of 20–25 °C. Commercial rat chow and tap water were provided ad libitum. All experimental animal procedures were performed following the Guidelines for the Care and Use of Mammals in Neuroscience and
Sleep restriction reduces pericyte-brain endothelial cell interactions
To test changes in pericyte-brain endothelial cell interactions, the expression of the proteins PDGFR-β and connexin 43 was evaluated after sleep loss. Western blot analysis showed that sleep restriction significantly decreased PDGFR-β expression in isolated brain MVs from cerebral cortex (t = 2.623, p = 0.0173) (Fig. 1A and B) and hippocampus (t = 5.219, p = 0.0001) (Fig. 2A and B). The observed changes were corroborated by confocal microscopy (Fig. 1C and C). As shown in Fig. 1C PDGFR-β
Discussion
Given the important role of pericytes in the maintenance of the barrier properties of the blood-brain barrier (Duz et al., 2007; Nakagawa et al., 2009, Bell et al., 2010, Rustenhoven et al., 2017), this study aimed to evaluate the changes in pericyte-brain endothelial cell interactions during sleep loss, and tried to characterize the effects of pericyte detachment in blood-brain barrier physiology in sleep-restricted rats. Here, we found that sleep loss promotes pericyte detachment from the
Conclusion
Chronic sleep restriction decreases the interactions between brain endothelial cells and pericytes by decreasing the expression of the proteins that maintain the junctions between them, such as PDGFR-β and connexin 43. The loss of pericyte-endothelial cell interactions concurs with a disruption of the barrier properties by reducing tight junction proteins between endothelial cells and increasing blood-brain barrier permeability to exogenous tracers in sleep-restricted rats. The impairment in
Funding
This work was partially supported by the Universidad Autónoma Metropolitana to BGG. The funding sources had no role in study design; in the collection, analysis and interpretation of data; in the writing of the report; and in the decision to submit the article for publication.
Declaration of Competing Interest
None.
Acknowledgment
to Dr. Óscar Flores Herrera from Dept. Biochemistry, Medicine School, Universidad Nacional Autónoma de México, for the technical support in the oximetry experiments. To MS. María Cristina Acosta García from the Electron Microscopy Lab, Dept. Biology of Reproduction, Biological Science Division of the Universidad Autónoma Metropolitana, Unidad Iztapalapa for their technical support with the scanning electron microscopy experiments.
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2021, Cerebral Circulation - Cognition and BehaviorCitation Excerpt :Supportive of this hypothesis is the evidence that pericytes (cells found surrounding the capillary/basement membrane and in direct contact with astrocyte endfeet) have contractile capabilities, constrict and dilate in response to local neurotransmitters, and may also contribute to controlling blood and CSF flow [105]. Recent evidence also suggests that pericyte function is compromised by sleep restriction [89] and impaired in Alzheimer's disease [144]. Vasomotion appears to be linked to neural activity, particularly power in the gamma spectral band of EEG, which predominates during waking [33,87], and may drive fluctuations in blood oxygenation, potentially contributing to resting state BOLD measures [87].