Chronic CBD 10 mg/kg/day administration was effective in the modification of cardiac and plasma endocannabinoid levels in two models of systemic hypertension [16
] and exerted beneficial cardiovascular effects in diabetic and septic rats, in mice with diabetic- and doxorubicin-induced cardiomyopathy and experimental autoimmune myocarditis (see [16
]). We decided to examine the protective but not therapeutic effect of CBD, because three weeks after MCT administration rats exhibited a higher mortality rate [25
3.1. Changes Related to PH
Three weeks after MCT injection we observed increases in the following, parameters of the pulmonary (but not systemic heart rate (HR) and systolic blood pressure (SBP)) circulation, which are characteristic for PH—(1) RVSP by about 115%, (2) RV hypertrophy expressed as Fulton index (RV/LV and septum ratio) by about 55% and (3) the adjusted weight of the lungs (by about 55%) suggesting lung edema [28
]. The above hemodynamic alternations in MCT-treated rats could result from morphometric and vascular functional changes, such as endothelial dysfunction; vascular remodeling, as evidenced by media hypertrophy (similarly to [24
]); and/or changes in vascular responsiveness, i.e., excessive PA constriction to thromboxane A2
analogue (U46619) and phenylephrine and attenuation of vasodilatory effects of Ach and SNP, as has been also demonstrated by Christou et al. [29
]. The latter could result (among other potential causes) from the altered expression and function of receptors for the respective agonists in the pulmonary circulation [30
]. The consequence of MCT-induced PH was relatively lower arterial oxygen saturation. PAI-1 inhibits t-PA by rapid formation of an enzyme inhibitor complex, leading to a prothrombotic phenotype of the endothelium [31
]. Moreover, PAI-1 has been demonstrated to induce pulmonary vascular remodeling [32
]. Similar elevations of PAI-1 and t-PA levels were observed previously in MCT-induced PH rats [32
]. In contrast to increases in plasma t-PA and PAI-1 concentrations MCT did not modify the plasma level of TFs, platelet adhesion to collagen or bleeding time. Importantly, routine blood testing revealed more than a twofold increase in leukocyte count. This may indicate an inflammatory process, which has been shown to play an important role in PH pathogenesis [24
]. However, the exact significance of these latter changes requires detailed study.
We are the first researchers who simultaneously determined 13 endocannabinoids and endocannabinoid-related lipids in the lungs (levels of OEA, SEA, HEA, DEA, 2-LG, DHEA, POEA and EPEA have never been determined in the lung). We confirmed that similarly to the lungs of rabbits [34
], rats [35
], mice [36
] and rat hearts [16
], the lungs’ 2-AG concentration was higher (by about 100 times) than the concentration of the well-known endocannabinoid AEA. Moreover, we noticed a similar ordering of endocannabinoid concentrations in rat lungs (the current study) and heart [16
]: 2-AG > PEA ≈ OEA > AEA > LEA ≈ SEA ≈ POEA ≈ NAGly > DHEA ≈ DEA ≈ HEA ≈ 2-LG. Changes in endocannabinoid levels appear to be dependent on the tissue and the model of hypertension. Thus, cardiac concentrations of AEA, 2-AG and some other endocannabinoids decreased in primary arterial hypertension (spontaneously hypertensive, SHR), but increased in secondary arterial hypertension (deoxycorticosterone (DOCA-salt)) [16
]. In contrast, only POEA and NAGly concentrations decreased in the lungs of PH rats.
To date, the role of these endocannabinoids and endocannabinoid-related lipids in the physiology and pathophysiology of the cardiopulmonary system, including PH, is still unknown. As mentioned in the introduction, a few of them regulate pulmonary vascular tone [9
]. However, in the isolated ventilated and buffer-perfused lungs of mice [37
] and rabbits [34
], AEA (but not 2-AG) has been shown to increase the perfusion pressure (reflecting a vasopressor response) by means of its vasoactive products.
3.2. Influence of CBD on the MCT-Induced PH
Chronic administration of CBD (10 mg/kg for three weeks) ameliorated MCT-induced PH in rats. Thus, it reduced by almost by 80% and completely returned to the control value two important changes typical for PH that were stimulated by MCT, namely, (1) elevation in RVSP and (2) decreased blood oxygen saturation, respectively. Interestingly, chronic administration of the same dose of CBD failed to modify blood pressure (BP) and HR in rats with primary (SHR) and secondary (DOCA-salt) hypertension as well as in their normotensive controls [16
]. We confirmed the lack of a CBD influence on the latter cardiovascular parameters of systemic circulation. Thus, it seems that CBD is sufficient to reduce pulmonary but not systemic BP.
How can we explain the beneficial CBD effects in PH? We suggest that the favorable effects of CBD in MCT-induced PH appear to be related mainly to the reduction of pulmonary vascular resistance by this compound, which could result from the reasons listed below.
Firstly, CBD diminished PA hypertrophy by approximately 30%. The potential anti-hypertrophic impact of CBD was demonstrated in previous experiments in which CBD decreased remodeling processes in the model of allergic asthma [18
] and inhibited the proliferation and migration of human umbilical artery smooth muscle cells (HUASMCs) [38
Secondly, CBD attenuated the vasoconstriction of PAs induced by thromboxane A2
analogue (significantly) and phenylephrine (tendency) and improved endothelial-dependent (Ach) and endothelial-independent (SNP) relaxation. Similar chronic administration of CBD-induced beneficial functional changes regarding systemic vascular responsiveness have been described in Zucker diabetic fatty rats [39
], in hypertensive DOCA-salt and SHR rats [40
] and in human brachial artery (improvement of endothelial function and arterial stiffness) [41
]. The direct vasorelaxant properties of acute CBD administration in isolated arteries [4
], including human pulmonary arteries [7
], was described in the introduction. In experiments conducted in vivo, an acute CBD injection reduced diastolic but not systolic BP in pithed rats (dependent on peripheral resistance and cardiac work, respectively) [42
], as well as the stress-induced increase in SBP in patients [41
Thirdly, CBD increased lung concentrations of endocannabinoids (AEA, 2-LG, LEA, POEA, EPEA and NAGly). AEA and NAGly have been demonstrated to possess potential vasodilatory properties, as has AEA in hPAs and rPAs [10
] and NAGly in the systemic arterial bed [43
]. So far, only AEA has been shown to reduce hypoxia-induced vasoconstriction (an important feature of PH), although it failed to modify the vascular caliber under normoxia in murine intra-acinar and pre-acinar arteries [44
]. Metabolites of AEA have been demonstrated to mediate hypoxia-induced PH in mice [37
]. However, we can exclude this possibility, since 24 h after the final dose of CBD, an increase in lung AEA level and a decrease in RSVP were noticed. The detailed role of other endocannabinoids still remains to be investigated. However, one should keep in mind that the upregulation of the endocannabinoid system under inflammatory conditions is recognized as an autoprotective mechanism in inhibiting disease progression [45
As we mentioned in the introduction, CBD can increase endocannabinoid levels via FAAH inhibition. Such a mechanism might take place not only in the case of AEA, but also in the case of other FAAH-sensitive N
-acylethanolamines, i.e., LEA, POEA and EPEA [16
]. NAGly is an endogenous FAAH inhibitor [46
]. We demonstrated previously that cardiac FAAH activity was inhibited 24 h after the final dose (10 mg/kg) of chronic administration in DOCA-salt and SHR. Interestingly, it was connected with a decrease in cardiac concentrations of 2-AG, DEA and OEA in DOCA-salt and no changes in SHR [16
]. Thus, it seems that CBD modifies endocannabinoid levels, dependent on tissue and the model of hypertension.
Fourthly, CBD normalized plasma PAI-1 and t-PA concentrations, which have been found to be strongly enhanced in PH. Similar decreases in plasma t-PA and PAI-1 in rat MCT-induced PH were induced by chronic administration of a phosphodiesterase-5 inhibitor, sildenafil, and a cholesterol lowering drug, simvastatin, which have therapeutic effects in PAH [32
Fifthly, chronic CBD administration reduced by 60% the leukocyte count that was doubled by MCT, which may result from the well-known anti-inflammatory effects of CBD [47
]. They have been described before, e.g., in a murine model of lipopolysaccharide-induced acute lung injury where CBD decreased total lung neutrophil, macrophage and lymphocyte migration into the lungs [17
] and potently reduced the inflammatory lung response [48
]. However, the potential anti-inflammatory effects of CBD in PH require detailed examination.
The beneficial effects of CBD in PH are not related to its influence on MCT-induced RV hypertrophy and lung edema since CBD did not inhibit the development of the above changes at all. CBD has been suggested to exert beneficial effects in cardiac injury [49
] but nobody has so far demonstrated its direct anti-proliferative influence in the heart.
CBD is recognized as a safe drug [50
], e.g., in healthy volunteers, oral and/or pulmonary administration of CBD was reported to be safe and well tolerated (600 mg ≈ 8 mg/kg [41
]). We have confirmed this finding. Thus, in our hands, chronic CBD did not modify parameters (1) that were not changed by MCT in rats with PH, such as bleeding time, platelet adhesion to collagen and blood parameters other than leukocytes, and (2) of control rats. Only the concentration of NAGly was diminished in the CBD-treated control animals.