TGF-β1 Regulation of Multidrug Resistance P-glycoprotein in the Developing Male Blood-Brain Barrier

P-glycoprotein (P-gp), an efflux transporter encoded by the abcb1 gene, protects the developing fetal brain. Levels of P-gp in endothelial cells of the blood-brain barrier (BBB) increase dramatically during the period of peak brain growth. This is coincident with increased release of TGF-β1 by astrocytes and neurons. Although TGF-β1 has been shown to modulate P-gp activity in a number of cell types, little is known about how TGF-β1 regulates brain protection. In the present study, we hypothesized that TGF-β1 increases abcb1 expression and P-gp activity in fetal and postnatal BBB in an age-dependent manner. We found TGF-β1 to potently regulate abcb1 mRNA and P-gp function. TGF-β1 increased P-gp function in brain endothelial cells (BECs) derived from fetal and postnatal male guinea pigs. These effects were more pronounced earlier in gestation when compared with BECs derived postnatally. To investigate the signaling pathways involved, BECs derived at gestational day 50 and postnatal day 14 were exposed to ALK1 and ALK5 inhibitors and agonists. Through inhibition of ALK5, we demonstrated that ALK5 is required for the TGF-β1 effects on P-gp function. Activation of ALK1, by the agonist BMP-9, produced similar results to TGF-β1 on P-gp function. However, TGF-β1 signaling through the ALK1 pathway is age-dependent as dorsomorphin, an ALK1 inhibitor, attenuated TGF-β1-mediated effects in BECs derived at postnatal day 14 but not in those derived at gestational day 50. In conclusion, TGF-β1 regulates P-gp at the fetal and neonatal BBB and both ALK5 and ALK1 pathways are implicated in the regulation of P-gp function. Aberrations in TGF-β1 levels at the developing BBB may lead to substantial changes in fetal brain exposure to P-gp substrates, triggering consequences for brain development.

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Abstract 218: Pericytic Laminin Regulates Blood-Brain Barrier Integrity in an Age-Dependent Manner

Stroke ◽

10.1161/str.48.suppl_1.218 ◽

2017 ◽

Vol 48 (suppl_1) ◽

Author(s):

Yao Yao ◽

Jyoti Gautam ◽

Xuanming Zhang

Keyword(s):

Endothelial Cells ◽

Basement Membrane ◽

Blood Brain Barrier ◽

Vessel Density ◽

Brain Barrier ◽

Dependent Manner ◽

Vascular Integrity ◽

Blood Brain ◽

Age Dependent

Introduction: Laminin, a major component of the basement membrane, plays an important role in blood brain barrier (BBB) regulation. At the neurovascular unit, astrocytes, brain endothelial cells, and pericytes synthesize and deposit different laminin isoforms into the basement membrane. Previous studies from our laboratory showed that loss of astrocytic laminin induces age-dependent and region-specific BBB breakdown and intracerebral hemorrhage, suggesting a critical role of astrocytic laminin in vascular integrity maintenance. Laminin α4 (predominantly generated by endothelial cells) has been shown to regulate vascular integrity at embryonic/neonatal stage. The role of pericytic laminin in vascular integrity, however, remains elusive. Methods: We investigated the function of pericyte-derived laminin in vascular integrity using laminin conditional knockout mice. Specifically, laminin floxed mice were crossed with PDGFRβ-Cre line to generate mutants (PKO) with laminin deficiency in PDGFRβ + cells, which include both pericytes and vascular smooth muscle cells (vSMCs). To distinguish the contribution of pericyte- and vSMC-derived laminin, we also generated a vSMC-specific condition knockout line (TKO) by crossing the laminin floxed mice with Transgelin-Cre mice. In this study, mice of both genders on a C57Bl6 background were used. At least 5-6 animals were used in biochemical and histological analyses in this study. Results: Pericyte-derived laminin was abrogated in all PKO mice. However, only old but not young PKO mice showed signs of BBB breakdown and reduced vessel density, suggesting age-dependent changes. Consistent with these data, further mechanistic studies revealed reduced tight junction proteins, diminished AQP4 expression, and deceased pericyte coverage in old but not young PKO mice. In addition, neither BBB disruption nor decreased vessel density was observed in TKO mice, suggesting that these vascular defects are due to loss of pericyte- rather than vSMC-derived laminin. Conclusions: These results strongly suggest that pericyte-derived laminin active regulates BBB integrity and vessel density in an age-dependent manner. I would like this abstract to be considered for the Stroke Basic Science Award.

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Structural, Molecular, and Functional Alterations of the Blood-Brain Barrier during Epileptogenesis and Epilepsy: A Cause, Consequence, or Both?

International Journal of Molecular Sciences ◽

10.3390/ijms21020591 ◽

2020 ◽

Vol 21 (2) ◽

pp. 591 ◽

Cited By ~ 17

Author(s):

Wolfgang Löscher ◽

Alon Friedman

Keyword(s):

Endothelial Cells ◽

Blood Brain Barrier ◽

Defense Mechanism ◽

Extracellular Fluid ◽

Cell Types ◽

Therapeutic Interventions ◽

Brain Barrier ◽

Transport Systems ◽

Blood Brain ◽

The Brain

The blood-brain barrier (BBB) is a dynamic, highly selective barrier primarily formed by endothelial cells connected by tight junctions that separate the circulating blood from the brain extracellular fluid. The endothelial cells lining the brain microvessels are under the inductive influence of neighboring cell types, including astrocytes and pericytes. In addition to the anatomical characteristics of the BBB, various specific transport systems, enzymes and receptors regulate molecular and cellular traffic across the BBB. While the intact BBB prevents many macromolecules and immune cells from entering the brain, following epileptogenic brain insults the BBB changes its properties. Among BBB alterations, albumin extravasation and diapedesis of leucocytes from blood into brain parenchyma occur, inducing or contributing to epileptogenesis. Furthermore, seizures themselves may modulate BBB functions, permitting albumin extravasation, leading to activation of astrocytes and the innate immune system, and eventually modifications of neuronal networks. BBB alterations following seizures are not necessarily associated with enhanced drug penetration into the brain. Increased expression of multidrug efflux transporters such as P-glycoprotein likely act as a ‘second line defense’ mechanism to protect the brain from toxins. A better understanding of the complex alterations in BBB structure and function following seizures and in epilepsy may lead to novel therapeutic interventions allowing the prevention and treatment of epilepsy as well as other detrimental neuro-psychiatric sequelae of brain injury.

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Human pluripotent stem cell–derived brain pericyte–like cells induce blood-brain barrier properties

Science Advances ◽

10.1126/sciadv.aau7375 ◽

2019 ◽

Vol 5 (3) ◽

pp. eaau7375 ◽

Cited By ~ 40

Author(s):

Matthew J. Stebbins ◽

Benjamin D. Gastfriend ◽

Scott G. Canfield ◽

Ming-Song Lee ◽

Drew Richards ◽

...

Keyword(s):

Stem Cells ◽

Endothelial Cells ◽

Blood Brain Barrier ◽

Neurovascular Unit ◽

Barrier Properties ◽

Cell Types ◽

Brain Barrier ◽

Human Model ◽

Blood Brain ◽

Brain Pericytes

Brain pericytes play important roles in the formation and maintenance of the neurovascular unit (NVU), and their dysfunction has been implicated in central nervous system disorders. While human pluripotent stem cells (hPSCs) have been used to model other NVU cell types, including brain microvascular endothelial cells (BMECs), astrocytes, and neurons, hPSC-derived brain pericyte–like cells have not been integrated into these models. In this study, we generated neural crest stem cells (NCSCs), the embryonic precursor to forebrain pericytes, from hPSCs and subsequently differentiated NCSCs to brain pericyte–like cells. These cells closely resembled primary human brain pericytes and self-assembled with endothelial cells. The brain pericyte–like cells induced blood-brain barrier properties in BMECs, including barrier enhancement and reduced transcytosis. Last, brain pericyte–like cells were incorporated with iPSC-derived BMECs, astrocytes, and neurons to form an isogenic human model that should prove useful for the study of the NVU.

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Age-dependent decline of blood–brain barrier P-glycoprotein expression in the canine brain

Neurobiology of Aging ◽

10.1016/j.neurobiolaging.2009.08.014 ◽

2011 ◽

Vol 32 (8) ◽

pp. 1477-1485 ◽

Cited By ~ 9

Author(s):

A. Pekcec ◽

E.L. Schneider ◽

W. Baumgärtner ◽

V.M. Stein ◽

A. Tipold ◽

...

Keyword(s):

Blood Brain Barrier ◽

Brain Barrier ◽

P Glycoprotein ◽

Blood Brain ◽

Canine Brain ◽

Age Dependent

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Blood-Brain Barrier Dysfunction in the Detrimental Brain Function

10.5772/intechopen.94572 ◽

2021 ◽

Author(s):

Alejandro Gonzalez-Candia ◽

Nicole K. Rogers ◽

Rodrigo L. Castillo

Keyword(s):

Endothelial Cells ◽

Blood Brain Barrier ◽

Brain Function ◽

Neurovascular Unit ◽

Cell Types ◽

Brain Barrier ◽

Barrier Dysfunction ◽

Microvascular Endothelium ◽

Blood Brain ◽

And Function

The blood circulation interface and the neural tissue feature unique characteristics encompassed by the term blood -brain barrier (BBB). The barrier’s primary functions are maintenance of brain homeostasis, selective transport, and protection, all of them determined by its specialized multicellular structure. The BBB primarily exists at the level of the brain microvascular endothelium; however, endothelial cells are not intrinsically capable of forming a barrier. Indeed, the development of barrier characteristics in cerebral endothelial cells requires coordinated cell–cell interactions and signaling from glial cells (i.e., astrocytes, microglia), pericytes, neurons, and extracellular matrix. Such an intricate relationship implies the existence of a neurovascular unit (NVU). The NVU concept emphasizes that the dynamic BBB response to stressors requires coordinated interactions between various central nervous system (CNS) cell types and structures. Every cell type makes an indispensable contribution to the BBBs integrity, and any cell’s failure or dysfunction might result in the barrier breakdown, with dramatic consequences, such as neuroinflammation and neurodegeneration. This chapter will focus on the structure and function of the BBB and discuss how BBB breakdown causes detrimental brain function.

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Plasmodium falciparum erythrocyte membrane protein 1 variants induce cell swelling and disrupt the blood–brain barrier in cerebral malaria

Journal of Experimental Medicine ◽

10.1084/jem.20201266 ◽

2021 ◽

Vol 218 (3) ◽

Author(s):

Yvonne Adams ◽

Rebecca W. Olsen ◽

Anja Bengtsson ◽

Nanna Dalgaard ◽

Mykola Zdioruk ◽

...

Keyword(s):

Endothelial Cells ◽

Plasmodium Falciparum ◽

Cerebral Malaria ◽

Blood Brain Barrier ◽

Brain Barrier ◽

Cell Swelling ◽

Dependent Manner ◽

Brain Endothelial Cells ◽

Blood Brain ◽

The Brain

Cerebral malaria (CM) is caused by the binding of Plasmodium falciparum–infected erythrocytes (IEs) to the brain microvasculature, leading to inflammation, vessel occlusion, and cerebral swelling. We have previously linked dual intercellular adhesion molecule-1 (ICAM-1)– and endothelial protein C receptor (EPCR)–binding P. falciparum parasites to these symptoms, but the mechanism driving the pathogenesis has not been identified. Here, we used a 3D spheroid model of the blood–brain barrier (BBB) to determine unexpected new features of IEs expressing the dual-receptor binding PfEMP1 parasite proteins. Analysis of multiple parasite lines shows that IEs are taken up by brain endothelial cells in an ICAM-1–dependent manner, resulting in breakdown of the BBB and swelling of the endothelial cells. Via ex vivo analysis of postmortem tissue samples from CM patients, we confirmed the presence of parasites within brain endothelial cells. Importantly, this discovery points to parasite ingress into the brain endothelium as a contributing factor to the pathology of human CM.

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SCIDOT-41. NANOTHERAPEUTIC TARGETING OF TUMOR ENDOTHELIUM FOR ENHANCING DRUG DELIVERY PAST THE BLOOD-BRAIN BARRIER

Neuro-Oncology ◽

10.1093/neuonc/noz175.1177 ◽

2019 ◽

Vol 21 (Supplement_6) ◽

pp. vi280-vi280

Author(s):

Hiroto Kiguchi ◽

Mandana Manzari ◽

Jake Vaynshteyn ◽

Jeffrey Gerwin ◽

Daniel Tylawsky ◽

...

Keyword(s):

Drug Delivery ◽

Endothelial Cells ◽

Blood Brain Barrier ◽

Low Dose ◽

Tumor Vasculature ◽

Adult Brain ◽

Brain Barrier ◽

Dependent Manner ◽

Shh Pathway ◽

Blood Brain

Abstract OBJECTIVE The Sonic Hedgehog (SHH) medulloblastoma subgroup accounts for ~25% of all cases and has an intermediate prognosis. Current conventional therapies result in devastating morbidities including intellectual disability and secondary malignancies. Although molecularly targeted agents that inhibit the SHH pathway have demonstrated clinical efficacy, recent studies have shown on-target secondary toxicities on bone development suggesting new therapeutic approaches are needed. METHODS We investigated the efficacy of the SHH pathway inhibitor, Vismodegib packaged in a fucoidan-based nanoparticle (Fi-Vis) that targets P-selectin, a protein overexpressed on vascular endothelial cells and induced by low-dose ionizing radiation (XRT) in a time- and dose-dependent manner. This P-selectin targeting nanoparticle drug delivery system shows selectivity toward tumor vasculature and not normal brain vasculature in a genetic SHH medulloblastoma mouse model as assessed by ex vivo infrared imaging and two-photon intravital imaging. RESULTS Quantitative RT-PCR analysis of SHH medulloblastoma tissue following single dose XRT and Fi-Vis treatment (10mg/kg) showed a synergistic inhibition of Gli1 expression (>90% target inhibition). Furthermore, we demonstrate that very low dose XRT (0.25Gy) can induce P-selectin expression specifically within MB tumor vasculature and synergize with low dose Fi-Vis (10mg/kg) to significantly enhance mouse survival (p<0.01) when compared to radiation or Fi-Vis alone. Furthermore, assessment of bone toxicity using micro-CT and histological analysis following Fi-Vis administration in postnatal (P10) mice shows no bone toxicity when compared to free Vismodegib. Finally, in vitro studies using mouse brain endothelial cells suggest at least in part a caveolin-1 mediated transcytosis mechanism of crossing the endothelial blood-brain barrier. CONCLUSIONS These data suggest applicability of combined XRT and tumor vasculature-targeted nanotherapeutic dose de-escalation strategies for SHH medulloblastoma with implications for other pediatric and adult brain tumors.

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P-glycoprotein in blood-brain barrier endothelial cells: interaction and oligomerization with caveolins

Journal of Neurochemistry ◽

10.1046/j.1471-4159.2003.02081.x ◽

2004 ◽

Vol 87 (4) ◽

pp. 1010-1023 ◽

Cited By ~ 74

Author(s):

Julie Jodoin ◽

Michel Demeule ◽

Laurence Fenart ◽

Roméo Cecchelli ◽

Sarah Farmer ◽

...

Keyword(s):

Endothelial Cells ◽

Blood Brain Barrier ◽

Brain Barrier ◽

P Glycoprotein ◽

Blood Brain

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Corticosteroid Regulation of P-Glycoprotein in the Developing Blood-Brain Barrier

Endocrinology ◽

10.1210/en.2010-1227 ◽

2011 ◽

Vol 152 (3) ◽

pp. 1067-1079 ◽

Cited By ~ 25

Author(s):

Majid Iqbal ◽

William Gibb ◽

Stephen G. Matthews

Keyword(s):

Blood Brain Barrier ◽

Fetal Brain ◽

Late Gestation ◽

Brain Barrier ◽

Protective Capacity ◽

Maternal Circulation ◽

Acetoxymethyl Ester ◽

P Glycoprotein ◽

Blood Brain ◽

Brain Microvessel

The early fetal brain is susceptible to teratogens in the maternal circulation, because brain microvessel expression of drug efflux transporter, P-glycoprotein (P-gp), is very low. However, there is a dramatic up-regulation of brain microvessel P-gp in late gestation. This study investigated the role of cortisol and dexamethasone in this up-regulation of fetal brain microvessel P-gp expression. Primary brain endothelial cell (BEC) cultures derived from gestational d (GD)40, GD50, GD65 (term, ∼68 d) and postnatal d 14 male guinea pigs were treated with varying doses (10−8 to 10−5m) of cortisol, dexamethasone, and aldosterone. After treatment, P-gp function was assessed using calcein-acetoxymethyl ester (P-gp substrate; 1 μm for 1 h) and measuring BEC accumulation of calcein. Corticosteroid treatment of BECs derived from postnatal d 14 resulted in increased P-gp activity. BECs derived from GD65 (near term) responded similarly, but these cells were extremely sensitive to the effects of mineralocorticoid receptor agonists (cortisol and aldosterone). BECs derived from GD50 displayed dose-dependent increases in P-gp function with dexamethasone (P < 0.05) and a trend towards increased function with cortisol. Cells derived from GD40 were unresponsive to all treatments. In conclusion, P-gp function in BECs is more responsive to glucocorticoids (GCs) in late gestation. Therefore, the late gestational surge in fetal plasma GCs, which parallels the increase in brain microvessel P-gp expression, may contribute to this P-gp up-regulation. Further, synthetic GCs (administered to pregnant women at risk of preterm delivery) may increase the protective capacity of the developing fetal blood-brain barrier, depending on the timing of GC exposure.

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