Brenda Brankin

RNAi-mediated reversible opening of the blood-brain barrier

The blood‐brain barrier (BBB) contains tight junctions (TJs) which reduce the space between adjacent endothelial cells lining the fine capillaries of the microvasculature of the brain to form a selective and regulatable barrier.

Neuropathology associated with feline immunodeficiency virus infection highlights prominent lymphocyte trafficking through both the blood-brain and blood-choroid plexus barriers

Feline immunodeficiency virus (FIV) infection in the cat is a well-evaluated model of human immunodeficiency virus (HIV)-1 infection in man with both viruses associated with significant neuropathology. Although studies in both HIV and FIV infections have shown that virus enters the brain in the acute stages of disease, little is known of the mechanisms of viral entry. The dissection of this stage is fundamental to the development of therapies that may prevent or modulate central nervous system (CNS) infection. The present study was designed to characterize the early sequential neuropathological changes following infection with FIVGL8, a strain known to enter the CNS in acute infection. Cats were infected either by the intraperitoneal (n = 13) or intravenous (n = 12) route with 2000 cat infectious units of virus. Histopathological assessments following intraperitoneal infections were at 4 (n = 2), 5 (n = 1), 8 (n = 3), 10 (n = 1), 16 (n = 1), 32 (n = 2), 52 (n = 2), and 104 (n = 1) weeks post infection whereas animals infected intravenously were examined (n = 3) at 1, 4, 10, and 23 weeks post infection. The most significant lesions following both routes of infection were lymphocyte-rich perivascular infiltrates within cerebral and cerebellar meninges, in choroid plexus and spinal cord dura mater and within epineurium of the sciatic nerve. In addition, following intravenous infection perivascular infiltrations were noted in parenchymal blood vessels primarily of cerebral white matter. Infiltrates were composed of CD79+ B cells and CD3+ T cells. The latter population contained a mixture of CD4+ and CD8+ cells. The severity of lesions increased in intensity in the 8- to 16-week period following infection and then began to wane. The evaluation of this large group of cats at multiple time points revealed pathology comparable with that of early stage HIV-1-associated encephalitis. Moreover, in contrast to previous FIV neuropathology studies, transient meningeal, choroid plexus, and parenchymal vascular pathology were consistent significant findings suggesting that, as in HIV-1 infection, blood-brain barrier and choroid plexus brain barrier integrity are both compromised in early infection.

Involvement of MAPKs in endostatin-mediated regulation of blood-retinal barrier function

Purpose: This study aimed to evaluate the effects of endostatin on tight junction (TJ) integrity in retinal microvascular endothelial cells (RMECs) in vitro and in vivo. Moreover, it was hypothesized that endostatin-induced occludin upregulation regulated VEGF165-mediated increases in endothelial cell permeability and involved activation of the MAPK signaling cascade. Endostatin is a 20-kDa fragment of collagen XVIII that has been shown to be efficacious in the eye by preventing retinal neovascularization. Endostatin is a specific inhibitor of endothelial cell proliferation, migration, and angiogenesis and has been reported to reverse VEGF-mediated increases in vasopermeability and to promote integrity of the blood-retinal barrier (BRB). In order to determine the mechanism of endostatin action on BRB integrity, we have examined the effects of endostatin on a number of intracellular pathways implicated in endothelial cell physiology. Methods: C57/Bl6 mice were injected with VEGF165 and/or endostatin, and the distribution of occludin staining was determined using retinal flatmounts. Western blot analysis of RMECs treated with VEGF165 and/or endostatin was used to determine changes in occludin expression and p38 MAPK and extracellular regulated kinase (ERK1/ERK2 MAPK) activation, while FD-4 flux across the RMEC monolayer was used to determine changes in paracellular permeability. Results: Endostatin prevented the discontinuous pattern of occludin staining observed at the retinal blood vessels of mice administered an intraocular injection of VEGF165. It was shown that endostatin activated p38 MAPK 5 min after addition to RMECs and continued to do so for approximately 30 min. Endostatin was also shown to activate ERK1/ERK2 5 min after addition and continued to do so, albeit with less potency, up to and including 15 min after addition. Inhibition of p38 MAPK and ERK1/ERK2 prevented endostatins ability to upregulate levels of occludin expression. Inhibition of these key signaling molecules was shown to prevent endostatins ability to protect against VEGF165-mediated increases in paracellular permeability in vitro. However, it appears that p38 MAPK may play a more important role in VEGF-mediated permeability, as inhibition of ERK1/ERK2 will not prevent VEGF165-mediated permeability compared with control (untreated) cells or cells treated with both a p38 MAPK inhibitor and VEGF165. Conclusions: Occludin is important for the maintenance of tight junction integrity in vivo. In a p38 MAPK and ERK1/ERK2 dependent manner, endostatin was shown to upregulate the levels of expression of the tight junction protein occludin. Inhibition of these key MAPK components may prevent endostatins ability to decrease VEGF165-induced paracellular permeability.

Wild-type measles virus infection upregulates poliovirus receptor-related 4 and causes apoptosis in brain endothelial cells by induction of tumor necrosis factor-related apoptosis-inducing ligand.

Small numbers of brain endothelial cells (BECs) are infected in children with neurologic complications of measles virus (MV) infection. This may provide a mechanism for virus entry into the central nervous system, but the mechanisms are unclear. Both in vitro culture systems and animal models are required to elucidate events in the endothelium. We compared the ability of wild-type (WT), vaccine, and rodent-adapted MV strains to infect, replicate, and induce apoptosis in human and murine brain endothelial cells (HBECs and MBECs, respectively). Mice also were infected intracerebrally. All MV stains productively infected HBECs and induced the MV receptor PVRL4. Efficient WT MV production also occurred in MBECs. Extensive monolayer destruction associated with activated caspase 3 staining was observed in HBECs and MBECs, most markedly with WT MV. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), but not Fas ligand, was induced by MV infection. Treatment of MBECs with supernatants from MV-infected MBEC cultures with an anti-TRAIL antibody blocked caspase 3 expression and monolayer destruction. TRAIL was also expressed in the endothelium and other cell types in infected murine brains. This is the first demonstration that infection of low numbers of BECs with WT MV allows efficient virus production, induction of TRAIL, and subsequent widespread apoptosis.

Feline immunodeficiency virus infection : A valuable model to study HIV-1 associated encephalitis

Feline immunodeficiency virus (FIV), like human immunodeficiency virus (HIV)-1, is a neurotropic lentivirus and is associated with neuropathology in natural and experimental infections. FIV enters the brain early following experimental infection, and virus has been proposed to enter the brain via the blood-brain barrier and blood-CSF barrier, within infected lymphocytes and monocytes/macrophages. However the entry of cell-free virus or the direct infection of brain endothelial cells and astrocytes of the blood-brain barrier may also contribute to CNS infection. This review explores the role played by the FIV model in the elucidation of mechanism of lentiviral entry to the brain and viral interactions with the CNS, particularly in relation to lymphotropic lentiviruses.