A novel method to isolate retinal and brain microvessels from individual rats: Microscopic and molecular biological characterization and application in hyperglycemic animals

Li, Youhai; Lapina, Natalia; Weinzierl, Nina; Bonde, Lisbeth; Boedtkjer, Ebbe; Schubert, Rudolf; Moshage, Han; Wohlfart, Paulus; Schilling, Lothar

doi:10.1016/j.vph.2018.07.001

Cited by 7 publications

(11 citation statements)

References 24 publications

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“…4 B). Moreover, enrichment of 5 endothelial cell marker genes (endothelial nitric oxide synthetase [NOS3], TEK tyrosin-kinase [Tie2], von Willebrand factor [vWF], claudin 5 [Cld5], solute carrier (SLC) 2,1a [Slc2a1, previously termed Glut-1]) and 3 pericyte markers (platelet-derived growth factor receptor, beta polypeptide [PDGFRb], chondroitin sulfate proteoglycan 4 [CSPG4], and regulator of G-protein signaling 5 [RGS5]) taken from our previous study 13 also displayed a significant correlation with the respective microarray data (Spearman rank test: r = 0.597, n = 16, p < 0.02). These observations strongly suggest good suitability and reliability of our microarray data for comparative gene expression analysis in the microvascular compartments.…”

Section: Resultsmentioning

confidence: 99%

“…Isolation of RMVs and BMVs was performed in exactly the same manner as described previously 13 . In this paper, we provided an in-depth characterization of the microvascular compartments showing that both, RMVs and BMVs were histologically devoid of any contamination by tissue fragments.…”

Section: Discussionmentioning

confidence: 99%

“…This is exemplified by significantly different activity levels of the p-glycoprotein transporter (gene symbol, ABCB1), a member of the ATP-binding cassette (ABC) efflux transporter family 6 and of high and low affinity neurotransmitter / organic acid transporters 7 . Furthermore, indirect evidence comes from studies addressing the level of expression/activity in isolated MVs or purified endothelial cells compared with the respective full tissue 8 – 13 .…”

Section: Introductionmentioning

confidence: 99%

“…Pericytes surround the abluminal side of the capillary wall and contribute to microvascular development, network stabilization and remodeling, and blood flow regulation 15 – 17 . In fact, the ratio of endothelial cells to pericytes is suspiciously high in the retina and brain 13 , pointing to their prominent role in RMVs and BMVs physiology within the neurovascular unit 14 . Therefore, to fully characterize the blood-neuronal barrier properties, isolated RMVs and BMVs consisting of endothelial cells and pericytes are in many respects superior to the use of individual cell types such as isolated endothelial cells or pericytes.…”

Section: Introductionmentioning

confidence: 99%

“…Therefore, we have previously developed a method, which allows high yield high purity isolation of the microvascular compartment from rat brain and retina. We characterized these isolates histologically and by semi-quantitative real time polymerase chain reaction (qRT-PCR) methodology which showed significant enrichment of markers of microvascular-specific cells and a distinct depletion of parenchymal cell markers (covering neuronal, astrocytic, and photoreceptor cells) compared to full tissues 13 . Based on these results we have now performed a transcriptome-wide analysis to systematically compare RMVs and BMVs obtained from the same group of rats.…”

Section: Introductionmentioning

confidence: 99%

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Comparative transcriptome analysis of inner blood-retinal barrier and blood–brain barrier in rats

Faiz

Moshage

et al. 2021

Sci Rep

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Although retinal microvessels (RMVs) and brain microvessels (BMVs) are closely related in their developmental and share similar blood-neural barriers, studies have reported markedly different responses to stressors such as diabetes. Therefore, we hypothesized that RMVs and BMVs will display substantial differences in gene expression levels even though they are of the same embryological origin. In this study, both RMVs and BMVs were mechanically isolated from rats. Full retinal and brain tissue samples (RT, BT) were collected for comparisons. Total RNA extracted from these four groups were processed on Affymetrix rat 2.0 microarray Chips. The transcriptional profiles of these tissues were then analyzed. In the present paper we looked at differentially expressed genes (DEGs) in RMVs (against RT) and BMVs (against BT) using a rather conservative threshold value of ≥ ± twofold change and a false discovery rate corrected for multiple comparisons (p < 0.05). In RMVs a total of 1559 DEGs were found, of which 1004 genes were higher expressed in RMVs than in RT. Moreover, 4244 DEGs between BMVs and BT were identified, of which 1956 genes were ≥ twofold enriched in BMVs. Using these DEGs, we comprehensively analyzed the actual expression levels and highlighted their involvement in critical functional structures in RMVs and BMVs, such as junctional complex, transporters and signaling pathways. Our work provides for the first time the transcriptional profiles of rat RMVs and BMVs. These results may help to understand why retina and brain microvasculature show different susceptibilities to stressors, and they might even provide new insight for pharmacological interventions.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%