Selected cannabis cultivars modulate glial activation: in vitro and in vivo studies.

IF 4.1 Q1 PHARMACOLOGY & PHARMACY
Sigal Fleisher-Berkovich, Nitzan Sharon, Yvonne Ventura, Valeria Feinshtein, Jonathan Gorelick, Nirit Bernstein, Shimon Ben-Shabat
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引用次数: 0

Abstract

Introduction: Multiple sclerosis (MS) is a chronic autoimmune disease of the central nervous system characterized by neuroinflammation, demyelination and axonal loss. Cannabis, an immunomodulating agent, is known for its ability to treat MS effectively. However, due to variations in the profile of secondary metabolites, especially cannabinoids, among cannabis cultivars, the effectiveness of cannabis treatment can vary, with significant variability in the effects on different biological parameters. For screening available cultivars, cellular in vitro as well as pre-clinical in vivo assays, are required to evaluate the effectiveness of the wide range of chemical variability that exists in cannabis cultivars. This study evaluated comparatively three chemically diverse cannabis cultivars, CN2, CN4 and CN6, containing different ratios of phytocannabinoids, for their neuroinflammatory activity in MS model.

Materials and methods: In vitro experiments were performed with lipopolysaccharide (LPS)-activated BV-2 microglia and primary glial cells to evaluate the effect of different cannabis cultivars on nitric oxide (NO) and inflammatory cytokines, as well as inducible nitric oxide synthase (iNOS) protein expression. An in vivo experiment using the experimental autoimmune encephalomyelitis (EAE) MS model was conducted using Myelin oligodendrocyte glycoprotein (MOG) as the activating peptide. The cannabis extracts of the cultivars CN2, CN4, CN6 or vehicle, were intraperitoneally injected with clinical scores given based on observed symptoms over the course of study. At the end of the experiment, the mice were sacrificed, and splenocyte cytokine secretion was measured using ELISA. Lumbar sections from the spinal cord of treated MS mice were evaluated for microglia, astrocytes and CD4+ cells.

Results: Extracts of the CN2 cultivar contained tetrahydrocannabinolic acid (THCA) and tetrahydrocannabinol (THC) without cannabidiol (CBD), and a number of monoterpenes. CN4 contained cannabidiolic acid (CBDA) and tetrahydrocannabidiolic acid (THCA), with significant amounts of THC: CBD in a 1:1 ratio, as well as sesquiterpenes and some monoterpenes; and CN6 contained primarily CBDA and THCA, as well as THC and CBD in a 2:1 ratio, with some sesquiterpenes and no monoterpenes. All extracts were not cytotoxic in glial cells up to 50 µg/ml. Dose dependent inhibition of LPS-induced BV2 as well as primary microglial NO secretion confirmed the anti-inflammatory and anti-oxidative activity of the three cannabis cultivars. CN2 but not CN4 reduced both astrocytosis and microglial activation in lumbar sections of EAE mice. In contrast, CN4 but not CN2 significantly decreased the secretion of TNFα and Interferon γ (IFNγ) in primary splenocytes extracted from EAE mice.

Conclusions: While both cannabis cultivars, CN2 and CN4, significantly reduced the severity of the clinical signs throughout the course of the study, they modulated different inflammatory mediators and pathways, probably due to differences in their phytocannabinoid composition. This demonstrates the differential potential of cannabis cultivars differing in chemotype to regulate neuroinflammation and their potential to treat MS.

精选大麻品种调节神经胶质激活:体外和体内研究。
导言:多发性硬化症(MS)是中枢神经系统的一种慢性自身免疫性疾病,以神经炎症、脱髓鞘和轴突丢失为特征。大麻是一种免疫调节剂,因其能够有效治疗多发性硬化症而闻名。然而,由于大麻栽培品种之间的次级代谢物(尤其是大麻素)特征存在差异,大麻的治疗效果也会不同,对不同生物参数的影响也存在显著差异。为了筛选现有的栽培品种,需要进行细胞体外和临床前体内试验,以评估大麻栽培品种中存在的各种化学变异的有效性。本研究比较评估了 CN2、CN4 和 CN6 这三种化学性质不同的大麻品种(含有不同比例的植物大麻素)在多发性硬化症模型中的神经炎症活性:用脂多糖(LPS)激活的 BV-2 小胶质细胞和原代胶质细胞进行体外实验,以评估不同大麻品种对一氧化氮(NO)和炎症细胞因子以及诱导型一氧化氮合酶(iNOS)蛋白表达的影响。使用髓鞘少突胶质细胞糖蛋白(MOG)作为激活肽,利用实验性自身免疫性脑脊髓炎(EAE)多发性硬化症模型进行了体内实验。腹腔注射种植品种 CN2、CN4、CN6 或载体的大麻提取物,并根据研究过程中观察到的症状进行临床评分。实验结束后,小鼠被处死,用酶联免疫吸附法测定脾细胞细胞因子的分泌量。对接受治疗的多发性硬化症小鼠脊髓腰部切片进行小胶质细胞、星形胶质细胞和 CD4+ 细胞评估:CN2 栽培品种的提取物含有四氢大麻酚酸(THCA)和四氢大麻酚(THC),不含大麻二酚(CBD)和一些单萜。CN4 含有大麻二酚(CBDA)和四氢大麻二酚(THCA),以及大量 THC:CBD(比例为 1:1)、倍半萜和一些单萜;CN6 主要含有 CBDA 和 THCA,以及 THC 和 CBD(比例为 2:1),还有一些倍半萜,没有单萜。所有提取物在 50 µg/ml 以下对神经胶质细胞均无细胞毒性。对 LPS 诱导的 BV2 和原发性小胶质细胞 NO 分泌的剂量依赖性抑制证实了这三种大麻品种的抗炎和抗氧化活性。在 EAE 小鼠的腰部切片中,CN2(而非 CN4)减少了星形胶质细胞增多和小胶质细胞活化。相比之下,CN4 而非 CN2 能显著减少 EAE 小鼠原代脾细胞中 TNFα 和干扰素 γ (IFNγ)的分泌:虽然 CN2 和 CN4 两种大麻在整个研究过程中都能明显减轻临床症状的严重程度,但它们调节的炎症介质和途径却有所不同,这可能是由于它们的植物大麻素成分不同所致。这证明了化学型不同的大麻品种在调节神经炎症方面的不同潜力及其治疗多发性硬化症的潜力。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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