Hypoxia/reoxygenation (H/R) is a component of several brain diseases such as traumatic brain injury, acute respiratory syndrome, obstructive sleep apnea, high altitude cerebral edema, acute mountain sickness, cardiac arrest and ischemic stroke. The objective of current stroke therapy is to restore perfusion to ischemic brain; however, considerable brain cellular damage and BBB dysfunction occurs when blood flow/oxygen supply is re-established. Therefore, there is a critical need for development of novel treatment strategies that can rescue salvageable brain tissue from damage and/or protect BBB integrity during H/R. In this grant, we will test the hypothesis that organic anion transporting polypeptides (Oatps) and multidrug resistance proteins (Mrps), two families of endogenous BBB transporters, can be targeted for treatment of H/R.
Two specific aims will test this hypothesis.
Aim 1 : To investigate CNS drug delivery mediated by Oatps during H/R. In this aim, we will focus on Oatp1a4, the primary drug transporting Oatp at the rodent BBB. We will investigate, in vivo, Oatp1a4 mediated transport of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase inhibitors (i.e., statins) (Aim 1A). We will then correlate changes in Oatp1a4-mediated statin transport with indices of neuroprotective and antioxidant efficacy (Aim 1B). Transforming growth factor (TGF-?) signaling regulates Oatp1a4 expression/activity. Therefore, we will evaluate effects of dorsomorphin and SB431542, two TGF-? receptor inhibitors, on BBB functional expression of Oatp1a4 to evaluate targeting of this pathway for control of CNS drug delivery (Aim 1C).
Aim 2 : To evaluate if pharmacological targeting of Mrps at the BBB protects BBB integrity following H/R. Glutathione (GSH), a critical CNS antioxidant, is a substrate for Mrp1, Mrp2, and Mrp4. Therefore, we will study in vivo H/R-induced changes in expression/activity of these Mrps at the BBB (Aim 2A). We will then examine changes in GSH and its oxidized form GSH disulfide (GSSG) at the BBB resulting from alterations in GSH transport systems and enzymes (Aim 2B). We will evaluate regulation of Mrps and GSH synthetic/metabolic enzymes at the BBB by nuclear factor erythroid 2-related factor 2 (Nrf2) signaling (Aim 2C). Since Nrf2 signaling is activated by oxidative stress, our studies will be conducted in the presence and absence of the reactive oxygen species (ROS) scavenger TEMPOL (i.e., 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) to determine if therapeutic targeting of oxidative stress can control BBB Mrp-mediated transport. Our goal in this grant is to facilitate discovery of novel approaches for treating diseases with an H/R component by therapeutic targeting of endogenous transporters at the BBB.
Treatment of brain diseases with a hypoxia/reoxygenation component requires delivery of drugs to attain effective levels in the brain and/or prevention of loss of the brain's natural antioxidants. Transporters are critical to the function of the blood-brin barrier (BBB) by precisely determining the levels of drugs and antioxidants in the brain. This grant proposal will directly impact public health in the United States by targeting BBB transporters for control of brain drug delivery and antioxidant transport, thereby facilitating discovery of new approaches to treat diseases with a hypoxia/reoxygenation component.
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