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Rho-Kinase Inhibition Improves the Outcome of Focal Subcortical White Matter Lesions - PubMed

. 2022 Jul;53(7):2369-2376.

doi: 10.1161/STROKEAHA.121.037358. Epub 2022 Jun 3.

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Rho-Kinase Inhibition Improves the Outcome of Focal Subcortical White Matter Lesions

Sanem A Aykan et al. Stroke. 2022 Jul.

Abstract

Background: Subcortical white matter lesions are exceedingly common in cerebral small vessel disease and lead to significant cumulative disability without an available treatment. Here, we tested a rho-kinase inhibitor on functional recovery after focal white matter injury.

Methods: A focal corpus callosum lesion was induced by stereotactic injection of N5-(1-iminoethyl)-L-ornithine in mice. Fasudil (10 mg/kg) or vehicle was administered daily for 2 weeks, starting one day after lesion induction. Resting-state functional connectivity and grid walk performance were studied longitudinally, and lesion volumes were determined at one month.

Results: Resting-state interhemispheric functional connectivity significantly recovered between days 1 and 14 in the fasudil group (P<0.001), despite worse initial connectivity loss than vehicle before treatment onset. Grid walk test revealed an increased number of foot faults in the vehicle group compared with baseline, which persisted for at least 4 weeks. In contrast, the fasudil arm did not show an increase in foot faults and had smaller lesions at 4 weeks. Immunohistochemical examination of reactive astrocytosis, synaptic density, and mature oligodendrocytes did not reveal a significant difference between treatment arms.

Conclusions: These data show that delayed fasudil posttreatment improves functional outcomes after a focal subcortical white matter lesion in mice. Future work will aim to elucidate the mechanisms.

Keywords: cell differentiation; fasudil; leukoaraiosis; oligodendrocyte; optical imaging.

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Conflict of interest statement

CONFLICT(S)-OF-INTEREST/DISCLOSURE(S)

Dr Ayata is on the scientific advisory board of Neurelis, Inc. and serves as a consultant.

Figures

Figure 1:
Figure 1:. Experimental timeline and protocol.

(A) Experimental timeline showing the timing of assessments and interventions. (B) A representative hematoxylin/eosin-stained coronal section showing the angle and location of L-NIO injection (arrow) and resulting corpus callosum lesion at day 35. (C) Dorsal field of view of the cranial window as well as the RSFC images. Intact skull transparency was achieved by a chronic cover glass preparation. The approximate location of the L-NIO lesion is also shown.

Figure 2:
Figure 2:. Interhemispheric homotopic resting state functional connectivity.

(A) Diagram of interhemispheric homotopic connectivity analysis using mirror pixels (left), the average interhemispheric homotopic connectivity map (right upper), and average interhemispheric homotopic correlation coefficients between mirror pixels averaged across the mediolateral axis and plotted along the anteroposterior axis, in mice with skull preparation only (i.e., no lesion). (B) Average interhemispheric homotopic connectivity maps (upper), and average interhemispheric homotopic correlation coefficients between mirror pixels plotted along the anteroposterior axis (lower) 1 or 14 days after L-NIO injection in vehicle- or fasudil-treated mice. Day 1 and 14 imaging was performed in the same cohort. Two-way ANOVA for repeated measures on Fisher z-transformed data.

Figure 3:
Figure 3:. Neurological deficits.

Grid walk test to examine motor coordination in vehicle and fasudil treated groups longitudinally at baseline and 1,2 and 4 weeks after L-NIO injection. Data (mean ± standard error) are shown as change from baseline for right and left forelimb separately. Two-way ANOVA for repeated measures.

Figure 4:
Figure 4:. Lesion morphometry.

Lesion areas on coronal plane (left) and distance from the midline of the medial and lateral lesion boundaries at 1 month are plotted along the anteroposterior axis. Two-way ANOVA for repeated measures.

Figure 5:
Figure 5:. Immunohistochemical examination.

(A) Left: Representative sections show perilesional reactive astrocytosis (GFAP), nuclear stain DAPI and the merged image obtained at the coronal section in the center of the lesion along the anteroposterior axis. Right: GFAP intensity was quantified in the entire field of view positioned as shown, at 5 coronal section levels at 200 μm intervals centered along the anteroposterior axis of the lesion bilaterally (two-way ANOVA for repeated measures). (B) Left: Representative sections show synaptophysin, nuclear stain DAPI and the merged image obtained at the coronal section in the center of the lesion along the anteroposterior axis. The bright synaptophysin signal in the lesion represented autofluorescence and was excluded from the cortical region of interest. Right: Synaptophysin intensity was quantified in a region of interest outlining the entire cortex within the field of view positioned as shown, at 5 coronal section levels at 200 μm intervals centered along the anteroposterior axis of the lesion bilaterally (two-way ANOVA for repeated measures). (C) Left: Representative sections show merged images of mature oligodendrocytes (GST-pi) and nuclear stain DAPI from the median corpus callosum as shown in the inset. Higher magnification images show examples of GST-pi positive cells. Right: GST-pi positive cells were manually counted in the entire field positioned in the center of the median corpus callosum as shown in the inset (0.0625 mm2) and centered along the anteroposterior axis of the lesion. All images were acquired using identical optical settings for a given marker and analyzed in a blinded fashion. P-values indicate vehicle vs. fasudil.

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References

    1. Bailey EL, Smith C, Sudlow CL, Wardlaw JM. Pathology of lacunar ischemic stroke in humans--a systematic review. Brain Pathol. 2012;22:583–591. doi: 10.1111/j.1750-3639.2012.00575.x - DOI - PMC - PubMed
    1. DeCarli C, Fletcher E, Ramey V, Harvey D, Jagust WJ. Anatomical mapping of white matter hyperintensities (WMH): exploring the relationships between periventricular WMH, deep WMH, and total WMH burden. Stroke. 2005;36:50–55. doi: 10.1161/01.STR.0000150668.58689.f2 - DOI - PMC - PubMed
    1. Su E, Bell M. Diffuse Axonal Injury. Laskowitz D, Grant G, eds. In: Translational Research in Traumatic Brain Injury. 2016.
    1. van der Knaap LJ, van der Ham IJ. How does the corpus callosum mediate interhemispheric transfer? A review. Behav Brain Res. 2011;223:211–221. doi: 10.1016/j.bbr.2011.04.018 - DOI - PubMed
    1. Wardlaw JM, Valdes Hernandez MC, Munoz-Maniega S. What are white matter hyperintensities made of? Relevance to vascular cognitive impairment. Journal of the American Heart Association. 2015;4:001140. doi: 10.1161/JAHA.114.001140 - DOI - PMC - PubMed

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