Protective Effects of Cannabidivarin and Cannabigerol on Cells of the Blood–Brain Barrier Under Ischemic Conditions
Nicole L. Stone, Timothy J. England, and Saoirse E. O’Sullivan
Cannabis and Cannabinoid Research, 2021,
Doi : 10.1089/can.2020.0159
Background and Objectives : Preclinical studies have shown cannabidiol is protective in models of ischemic stroke. Based on results from our recent systematic review, we investigated the effects of two promising neuroprotective phytocannabinoids, cannabigerol (CBG) and cannabidivarin (CBDV), on cells of the blood–brain barrier (BBB), namely human brain microvascular endothelial cells (HBMECs), pericytes, and astrocytes.
Experimental Approach : Cultures were subjected to oxygen-glucose deprivation (OGD) protocol to model ischemic stroke and cell culture medium was assessed for cytokines and adhesion molecules post-OGD. Astrocyte cell lysates were also analyzed for DNA damage markers. Antagonist studies were conducted where appropriate to study receptor mechanisms.
Results : In astrocytes CBG and CBDV attenuated levels of interleukin-6 (IL-6) and lactate dehydrogenase (LDH), whereas CBDV (10 nM–10 lM) also decreased vascular endothelial growth factor (VEGF) secretion. CBDV (300 nM–10 lM) attenuated levels of monocyte chemoattractant protein (MCP)-1 in HBMECs. In astrocytes, CBG decreased levels of DNA damage proteins, including p53, whereas CBDV increased levels of DNA damage markers. Antagonists for CB1, CB2, PPAR-c, PPAR-a, 5-HT1A, and TRPV1 had no effect on CBG (3 lM) or CBDV (1 lM)-mediated decreases in LDH in astrocytes. GPR55 and GPR18 were partially implicated in the effects of CBDV, but no molecular target was identified for CBG.
Conclusions : We show that CBG and CBDV were protective against OG mediated injury in three different cells that constitute the BBB, modulating different hallmarks of ischemic stroke pathophysiology. These data enhance our understanding of the protective effects of CBG and CBDV and warrant further investigation into these compounds in ischemic stroke. Future studies should identify other possible neuroprotective effects of CBG and CBDV and their corresponding mechanisms of action.
Keywords : blood–brain barrier; cannabidivarin; cannabigerol; cannabinoids; ischemia; neuroprotection
The blood–brain barrier (BBB) is a unique interface that separates the central nervous system (CNS) and the periphery, protecting the brain from damaging components found in general circulation, namely peripheral leukocytes, macromolecules, and xenobiotics. 1,2 The barrier itself is formed by microvascular endothelial cells, which are en-compassed by pericytes, and altogether surrounded by astrocyte end feet, which cover 99% of BBB endothelia.3 Cerebral ischemia– reperfusion (IR) initiates a plethora of inflammatory signaling pathways, cytotoxic glutamate release, and oxidative stress, all of which contribute to increases in BBB permeability.4 This loss of BBB integrity ultimately causes uncontrolled immune infiltration into the CNS that perpetuates neuronal injury and hinders poststroke recovery. Although administration of tissue plasminogen activator (tPA) and mechanical thrombectomy are effective licensed therapies to dissolve or remove the culpable clot, at present, there are no available approved therapies that mitigate poststroke injury.5
Cannabidiol (CBD), one of the chemicals found in Cannabis sativa, has displayed a range of neuroprotective qualities, preventing neuronal loss,6,7 attenuating astrocyte reactivity,8 and dampening the neuroinflammatory response.9 Unlike delta9 tetrahydrocannabinol (D9-THC), CBD does not activate the central cannabinoid receptors, CB1 or CB2, but activates a plethora of other targets including PPAR-c, TRPV1, and 5-HT1A receptors.10–13 CBD has formulations (alone and with D9-THC) licensed by GW pharmaceuticals to treat rare childhood epilepsies and spasticity associated with multiple sclerosis. The protective effects of CBD in stroke models has been well documented,14 specifically CBD has been shown to reduce infarct volume,15,16 reduce glutamate toxicity,9,17 attenuate mitochondrial dysfunction18 and glial activation.6,19 In a co-culture BBB model CBD preserved barrier integrity after oxygenglucose deprivation (OGD), which was mediated at least in part by PPAR-c and 5-HT1A receptors.12
Cannabigerol (CBG) and cannabidivarin (CBDV) are neutral cannabinoids present in cannabis and studies have found these compounds share similar pharmacological characteristics to CBD. Like CBD, they do not produce feelings of euphoria and display antioxidant and anti-inflammatory properties, as well as interacting with a range of target proteins including TRPV1,13 PPAR-c,20 5-HT1A, and CB2.21 Recently our group conducted a systematic review focusing on the neuroprotective properties of minor phytocannabinoids (other than D9-THC or CBD) and found that CBG and CBDV show efficacy in models of Huntington’s disease, Alzheimer’s, and epilepsy, with CBG mediating its protective effects through PPAR-c activation,22 the same mechanism by which we have shown that CBD protects BBB integrity.12 However, despite these compounds having neuroprotective effects in other models, no studies have been conducted to test whether CBG or CBDV are protective in IR injury.
In light of the above, we hypothesized these compounds may exhibit protective properties at the BBB in a stroke model. To test this, we treated cells of the BBB with CBG or CBDV in vitro before an OGD protocol and measured various proinflammatory cytokines, adhesion molecules, and cell damage markers.