Person:

Lo, Eng

Trophic coupling between cerebral endothelium and their neighboring cells is required for the development and maintenance of blood-brain barrier (BBB) function. Here we report that oligodendrocyte precursor cells (OPCs) secrete soluble factor TGF-β1 to support BBB integrity. Firstly, we prepared conditioned media from OPC cultures and added them to cerebral endothelial cultures. Our pharmacological experiments showed that OPC-conditioned media increased expressions of tight-junction proteins and decreased in vitro BBB permeability by activating TGB-β-receptor-MEK/ERK signaling pathway. Secondly, our immuno-electron microscopic observation revealed that in neonatal mouse brains, OPCs attach to cerebral endothelial cells via basal lamina. And finally, we developed a novel transgenic mouse line that TGF-β1 is knocked down specifically in OPCs. Neonates of these OPC-specific TGF-β1 deficient mice (OPC-specific TGF-β1 partial KO mice: PdgfraCre/Tgfb1flox/wt mice or OPC-specific TGF-β1 total KO mice: PdgfraCre/Tgfb1flox/flox mice) exhibited cerebral hemorrhage and loss of BBB function. Taken together, our current study demonstrates that OPCs increase BBB tightness by upregulating tight junction proteins via TGF-β signaling. Although astrocytes and pericytes are well-known regulators of BBB maturation and maintenance, these findings indicate that OPCs also play a pivotal role in promoting BBB integrity.

Progress in experimental stroke and translational medicine could be accelerated by high-resolution in vivo imaging of disease progression in the mouse cortex. Here, we introduce optical microscopic methods that monitor brain injury progression using intrinsic optical scattering properties of cortical tissue. A multi-parametric Optical Coherence Tomography (OCT) platform for longitudinal imaging of ischemic stroke in mice, through thinned-skull, reinforced cranial window surgical preparations, is described. In the acute stages, the spatiotemporal interplay between hemodynamics and cell viability, a key determinant of pathogenesis, was imaged. In acute stroke, microscopic biomarkers for eventual infarction, including capillary non-perfusion, cerebral blood flow deficiency, altered cellular scattering, and impaired autoregulation of cerebral blood flow, were quantified and correlated with histology. Additionally, longitudinal microscopy revealed remodeling and flow recovery after one week of chronic stroke. Intrinsic scattering properties serve as reporters of acute cellular and vascular injury and recovery in experimental stroke. Multi-parametric OCT represents a robust in vivo imaging platform to comprehensively investigate these properties.

Objective: Rho-associated kinase (ROCK) is a key regulator of numerous processes in multiple cell types relevant in stroke pathophysiology. ROCK inhibitors have improved outcome in experimental models of acute ischemic or hemorrhagic stroke. However, the relevant ROCK isoform (ROCK1 or ROCK2) in acute stroke is not known. Methods: We characterized the pharmacodynamic and pharmacokinetic profile, and tested the efficacy and safety of a novel selective ROCK2 inhibitor KD025 (formerly SLx-2119) in focal cerebral ischemia models in mice. Results: KD025 dose-dependently reduced infarct volume after transient middle cerebral artery occlusion. The therapeutic window was at least 3 h from stroke onset, and the efficacy was sustained for at least 4 weeks. KD025 was at least as efficacious in aged, diabetic or female mice, as in normal adult males. Concurrent treatment with atorvastatin was safe, but not additive or synergistic. KD025 was also safe in a permanent ischemia model, albeit with diminished efficacy. As one mechanism of protection, KD025 improved cortical perfusion in a distal middle cerebral artery occlusion model, implicating enhanced collateral flow. Unlike isoform-nonselective ROCK inhibitors, KD025 did not cause significant hypotension, a dose-limiting side effect in acute ischemic stroke. Interpretation Altogether, these data show that KD025 is efficacious and safe in acute focal cerebral ischemia in mice, implicating ROCK2 as the relevant isoform in acute ischemic stroke. Data suggest that selective ROCK2 inhibition has a favorable safety profile to facilitate clinical translation.

Objective: Brain amyloidosis is a key feature of Alzheimer's disease (AD). It also incorporates cerebrovascular amyloid β (Aβ) in the form of cerebral amyloid angiopathy (CAA) involving neurovascular dysfunction. We have recently shown by retrospective analysis that patients with mild cognitive impairment receiving a vasoactive drug cilostazol, a selective inhibitor of phosphodiesterase (PDE) III, exhibit significantly reduced cognitive decline. Here, we tested whether cilostazol protects against the disruption of the neurovascular unit and facilitates the arterial pulsation-driven perivascular drainage of Aβ in AD/CAA. Methods: We explored the expression of PDE III in postmortem human brain tissue followed by a series of experiments examining the effects of cilostazol on Aβ metabolism in transgenic mice (Tg-SwDI mice) as a model of cerebrovascular β-amyloidosis, as well as cultured neurons. Results: We established that PDE III is abnormally upregulated in cerebral blood vessels of AD and CAA subjects and closely correlates with vascular amyloid burden. Furthermore, we demonstrated that cilostazol treatment maintained cerebral hyperemic and vasodilative responses to hypercapnia and acetylcholine, suppressed degeneration of pericytes and vascular smooth muscle cells, promoted perivascular drainage of soluble fluorescent Aβ1-40, and rescued cognitive deficits in Tg-SwDI mice. Although cilostazol decreased endogenous Aβ production in cultured neurons, C-terminal fragment of amyloid precursor protein expression was not altered in cilostazol-treated Tg-SwDI mice. Interpretation The predominant action of cilostazol on Aβ metabolism is likely to facilitate Aβ clearance due to the sustained cerebrovascular function in vivo. Our findings mechanistically demonstrate that cilostazol is a promising therapeutic approach for AD and CAA.

There is growing interest in the potential neuroprotective properties of gelsolin. In particular, plasma-type gelsolin (pGSN) can ameliorate deleterious inflammatory response by scavenging pro-inflammatory signals such as actin and lipopolysaccharide. In a recent issue of Critical Care, Pan and colleagues report an important association between pGSN and subarachnoid hemorrhage (SAH) disease severity, and found pGSN to be a novel and promising biomarker for SAH clinical outcome. Previous research shows pGSN may be actively degraded by neurovascular proteases such as matrix metalloproteinases in the cerebral spinal fluid of SAH patients. Taken together, these results suggest that pGSN is not only a novel marker of SAH clinical outcome, but may also play an active mechanistic role in SAH, and potentially serve as a future therapeutic target.

White matter dysfunction is an important part of many CNS disorders including multiple sclerosis (MS) and vascular dementia. Within injured areas, myelin loss and oligodendrocyte death may trigger endogenous attempts at regeneration. However, during disease progression, remyelination failure may eventually occur due to impaired survival/proliferation, migration/recruitment, and differentiation of oligodendrocyte precursor cells (OPCs). The ventricular-subventricular zone (V-SVZ) and the subgranular zone (SGZ) are the main sources of neural stem/progenitor cells (NSPCs), which can give rise to neurons as well as OPCs. Under normal conditions in the adult brain, the V-SVZ progenitors generate a large number of neurons with a small number of oligodendrocyte lineage cells. However, after demyelination, the fate of V-SVZ-derived progenitor cells shifts from neurons to OPCs, and these newly generated OPCs migrate to the demyelinating lesions to ease white matter damage. In this mini-review, we will summarize the recent studies on extrinsic (e.g., vasculature, extracellular matrix (ECM), cerebrospinal fluid (CSF)) and intrinsic (e.g., transcription factors, epigenetic modifiers) factors, which mediate oligodendrocyte generation from the V-SVZ progenitor cells. A deeper understanding of the mechanisms that regulate the fate of V-SVZ progenitor cells may lead to new therapeutic approaches for ameliorating white matter dysfunction and damage in CNS disorders.

Background: Intracerebral hemorrhage is a subtype of stroke that has a poor prognosis without an adequate therapy. Recently, the use of anesthetics such as isoflurane has been shown to be protective after cerebral ischemia. However, the potential therapeutic effect of isoflurane after intracerebral hemorrhage (ICH) has not been fully explored. Results: In this study, male Sprague–Dawley rats (SD) were subjected to ICH and randomized into controls and 1.2% or 1.5% isoflurane posttreatment groups. Brain water content, neurological outcomes and matrix metalloproteinase-2 and -9 (MMP2-MMP9) plasma levels were quantified at 24 hours. Isoflurane treatment did not reduce brain edema compared with controls in any of the applied isoflurane concentrations. Moreover, consistent with this lack of effect on brain edema, isoflurane posttreatment did not affect neurological outcomes in any of the tests used. Plasma MMP levels did not change. Conclusion: Our data suggested that there is no neuroprotection after isoflurane posttreatment in a rat model of ICH.

Tissue plasminogen activator (tPA) is the only FDA-approved treatment for reperfusing ischemic strokes. But widespread use of tPA is still limited by fears of inadvertently administering tPA in patients with intracerebral hemorrhage (ICH). Surprisingly, however, the assumption that tPA will worsen ICH has never been biologically tested. Here, we assessed the effects of tPA in two models of ICH. In a mouse model of collagenase-induced ICH, hemorrhage volumes and neurological deficits after 24 hrs were similar in saline controls and tPA-treated mice, whereas heparin-treated mice had 3-fold larger hematomas. In a model of laser-induced vessel rupture, tPA also did not worsen hemorrhage volumes, while heparin did. tPA is known to worsen neurovascular injury by amplifying matrix metalloproteinases during cerebral ischemia. In contrast, tPA did not upregulate matrix metalloproteinases in our mouse ICH models. In summary, our experimental data do not support the assumption that intravenous tPA has a deleterious effect in acute ICH. However, due to potential species differences and the inability of models to fully capture the dynamics of human ICH, caution is warranted when considering the implications of these findings for human therapy.

The repair process after CNS injury shows a well-organized cascade of three distinct stages: inflammation, new tissue formation, and remodeling. In the new tissue formation stage, various cells migrate and form the fibrotic scar surrounding the lesion site. The fibrotic scar is known as an obstacle for axonal regeneration in the remodeling stage. However, the role of the fibrotic scar in the new tissue formation stage remains largely unknown. We found that the number of A-kinase anchoring protein 12 (AKAP12)-positive cells in the fibrotic scar was increased over time, and the cells formed a structure which traps various immune cells. Furthermore, the AKAP12-positive cells strongly express junction proteins which enable the structure to function as a physical barrier. In in vivo validation, AKAP12 knock-out (KO) mice showed leakage from a lesion, resulting from an impaired structure with the loss of the junction complex. Consistently, focal brain injury in the AKAP12 KO mice led to extended inflammation and more severe tissue damage compared to the wild type (WT) mice. Accordingly, our results suggest that AKAP12-positive cells in the fibrotic scar may restrict excessive inflammation, demonstrating certain mechanisms that could underlie the beneficial actions of the fibrotic scar in the new tissue formation stage during the CNS repair process.

What is the organization of cerebral microvascular oxygenation and morphology that allows adequate tissue oxygenation at different activity levels? We address this question in the mouse cerebral cortex using microscopic imaging of intravascular O2 partial pressure and blood flow combined with numerical modeling. Here we show that parenchymal arterioles are responsible for 50% of the extracted O2 at baseline activity and the majority of the remaining O2 exchange takes place within the first few capillary branches. Most capillaries release little O2 at baseline acting as an O2 reserve that is recruited during increased neuronal activity or decreased blood flow. Our results challenge the common perception that capillaries are the major site of O2 delivery to cerebral tissue. The understanding of oxygenation distribution along arterio-capillary paths may have profound implications for the interpretation of BOLD fMRI signal and for evaluating microvascular O2 delivery capacity to support cerebral tissue in disease.