Impact of cigarette smoke extract and hyperglycemic conditions on blood–brain barrier endothelial cells

Abstract

BackgroundDiabetes and tobacco smoking are significant public health concerns which have been shown to independently impact the blood–brain barrier (BBB). Since smoking is a risk factor for diabetes and shares some of the common pathological pathways leading to metabolic abnormalities, it is hypothesized that their combination would produce additive or synergistic BBB dysfunction. Therefore, the objective of this study was to assess this hypothesis and evaluate the magnitude of these effects in vitro using hCMEC/D3 cells; a well-established human BBB endothelial cell line.MethodsMonolayers of hCMEC/D3 cells were exposed to hyperglycemic conditions (HG; 35 mM) or 5% soluble cigarette smoke extracts (CSE, model of mainstream smoke exposure) for 12–24 h. Cells were then harvested for subsequent biochemical analyses. Transendothelial electrical resistance (TEER) and paracellular permeability to florescent dextrans were used to assess monolayer integrity. Analysis of released factors and cytokines was carried out by ELISA. Western blot (WB) analysis/immunofluorescence of relevant molecular targets was carried out. P-gp efflux activity was measured using rhodamine 123.ResultsImmunofluorescence and WB data showed a significant ZO-1 down-regulation by HG and/or CSE over 24 h exposure. CSE in presence of HG produced a synergistic increase in release of vascular endothelial growth factor that was accompanied by decreased TEER and augmented permeability to labeled dextrans in a size-dependent manner. Moreover, CSE increased the expression of GLUT-1 and SGLT-1 in isolated membrane fractions of hCMEC/D3 cells. The effect was amplified by HG. Both, HG and CSE elicited the membrane upregulation of P-glycoprotein (P-gp) expression which however, was not paralleled by a comparable efflux activity. Interestingly, concomitant exposure to HG and CSE evoked a marked upregulation of PECAM-1 and other pro-inflammatory markers including IL-6 and -8, when compared to each condition alone. Moreover, exposure to all tested conditions amplified (to a different degree) cellular oxidative stress response denoted by increased Nrf2 nuclear translocation.ConclusionOverall, our results have clearly shown an additive pattern in the release of angiogenic and inflammatory factors following concomitant exposure to HG and CSE. This suggests the involvement of common key modulators in BBB impairment by both CS and HG possibly through the activation of oxidative stress responses.

Drugs of abuse and blood-brain barrier endothelial dysfunction: A focus on the role of oxidative stress

“…Activation of oxidative/ inflammatory stress signaling cascades by drugs of abuse is the primary mechanism for increased endothelial activation and leukocyte migration across the BBB into the brain. Recent studies from our group (Sajja et al, 46 Naik et al, 115 and Prasad et al 146 ) and other groups (Alfieri et al 147 and Sandberg et al…”

Cigarette smoke and related risk factors in neurological disorders: An update

“…Similarly, Hawkins and coworkers confirmed that nicotine down regulated the expression of the ZO-1 tight junction protein in the BBB, the follow-on of increased permeability [53]. Furthermore, many other scientific communities revealed that nicotine alters the function of the BBB Na +À K +À 2Cl À co transporter through mediation of a7nAChR [54][55][56].…”

Offsetting the impact of smoking and e-cigarette vaping on the cerebrovascular system and stroke injury: Is Metformin a viable countermeasure?

“…Further, we and others have found that TS promotes glucose intolerance and increases the risk of developing type-2 diabetes mellitus ( 2DM ) [27], [28] with which it shares other pathogenic traits including the high risk of cerebrovascular and neurological disorders like stroke [29] via ROS generation, inflammation, and blood-brain barrier ( BBB ) impairment [30], [31], [32]. Recent findings [33], [34] from our group, supports an additive release pattern of angiogenic, oxidative and inflammatory factors by BBB endothelial cells in response to hyperglycemia ( HG ) and/or stroke conditions with comcomitant exposure to cigarette smoke extracts ( CSE ), thus suggesting the involvement of common pathogenic modulators of BBB impairment. To this end, Metformin ( MF ; a widely prescribed, firstline anti-diabetic drug) before and after stroke injury has been shown to reduce stress and inhibit the inflammatory responses [34], [35].…”