title: Effect of full flavor and denicotinized cigarettes exposure on the brain microvascular endothelium: a microarray-based gene expression study using a human immortalized BBB endothelial cell line.
abstract: BACKGROUND
  Tobacco smoke (TS) toxicity to the brain microvasculature is still an understudied area till date. NF-E2 related factor (Nrf2) is a key transcription factor responsible for activating the antioxidant response element (ARE) genes following an oxidative insult. Till date, several studies targeting the blood brain barrier (BBB) have shown some protective role of Nrf2 in ischemia-reperfusion (IR) injury, however, its functional role in chronic smokers subjected to a life-long oxidative stress has never been addressed. This is of crucial importance since smokers have a much higher risk for cerebrovascular stroke and tobacco smoke exposure has been clearly shown to enhance BBB damage following an ischemia/reperfusion injury. Thus, the goal of our study was to investigate the defense pathways activated at the BBB endothelial level by TS exposure. Specifically we focused on Nrf2 and nuclear factor kappa-light-chain-enhancer of activated B signaling response (NF-κβ) as the central protective mechanisms related to oxidative insult.
  
  
  RESULTS
  With the exception of Nicotine, both full flavor (3R4F) and decotinized (ULN) cigarettes activated Nrf2 and NFκβ pathways in hCMEC/D3 endothelial cells. Several detoxification and anti-oxidant genes including downstream products were also activated including NAD(P)H dehydrogenase quinone 1 (NQO-1), heme oxygenase-1 (HMOX-1), catalytic and modifier subunits of glutamate-cysteine ligase (GCL), solute carrier-SLC7A11). Gene expression levels of cytochrome P450s (CYP2S1 and CYP51A1) and efflux transporters P-glycoprotein (P-gp) and multi-drug resistance protein-4 (MRP4) were also enhanced. Increase of P-gp functional activity and depletion of GSH were also observed. Strikingly, toxicity of denicotinized ("reduced exposure") cigarettes was equivalent to 3R4F (or worse).
  
  
  CONCLUSIONS
  This study provides a detailed analysis of Nrf2-related cytoprotective mechanisms activated in response to 3R4F and ULN-derived TS exposure correlating the results with their oxidative and inflammatory potential. Toxicants present in soluble cigarette smoke extracts (CSE) and not nicotine seem to be the primary determinant of vascular toxicity. In this respect our results from this and previous studies suggest that chronic TS exposure can overcome Nrf2 and NFκB-p65 dependent cytoprotective mechanisms of the brain microvascular endothelium possibly leading to BBB impairment and loss of BBB integrity.
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, 1300 S. Coulter Street, Amarillo, TX, 79106, USA. pooja.naik@ttuhsc.edu.
authors.forename: Pooja
authors.initials: P
authors.lastname: Naik
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, 1300 S. Coulter Street, Amarillo, TX, 79106, USA. ravi.sajja@ttuhsc.edu.
authors.forename: Ravi K
authors.initials: RK
authors.lastname: Sajja
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, 1300 S. Coulter Street, Amarillo, TX, 79106, USA. prasad.shikha@ttuhsc.edu.
authors.forename: Shikha
authors.initials: S
authors.lastname: Prasad
authors.affiliation: Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, 1300 S. Coulter Street, Amarillo, TX, 79106, USA. luca.cucullo@ttuhsc.edu.|Center for Blood Brain Barrier Research, Texas Tech University Health Sciences Center, Amarillo, TX, 79106, USA. luca.cucullo@ttuhsc.edu.
authors.forename: Luca
authors.initials: L
authors.lastname: Cucullo
mesh_terms: D001812:Blood-Brain Barrier / Q000187:drug effects* / Q000378:metabolism; D015153:Blotting, Western; D001921:Brain / Q000098:blood supply / Q000187:drug effects* / Q000378:metabolism; D002460:Cell Line; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D046228:Microarray Analysis; D055806:Microvessels / Q000187:drug effects* / Q000378:metabolism; D051267:NF-E2-Related Factor 2 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D060888:Real-Time Polymerase Chain Reaction; D012906:Smoke / Q000009:adverse effects*; D062789:Tobacco Products / Q000009:adverse effects*
chemical_list: D051267:NF-E2-Related Factor 2; D016328:NF-kappa B; C495635:NFE2L2 protein, human; D012906:Smoke; D003577:Cytochrome P-450 Enzyme System