LETTER TO THE EDITOR

We read with great interest the paper by Zhang et al, “Cannabinoid receptor and N‐acyl phosphatidylethanolamine phospholipase D—Evidence for altered expression in multiple sclerosis”(2). The authors found an up‐regulation of CB₁R and CB₂R cannabinoid receptors not only at the site of neuronal or axonal damage, but also in cerebral blood vessels at the site of blood‐brain barrier disruption (in particular for CB₂R). At the latter site also, the immunoreactivity of N‐acyl phosphatidylethanolamine phospholipase D (NAPE‐PLD), a key enzyme of the endocannabinoid system (ECS), was detected. These data are highly suggestive of a neuroprotective role of ECS, in addition to a possible new endothelial reparative function. In keeping with this hypothesis is the finding of a reduced vascular density, where higher CB₂R expression was detected, further supporting a role for ECS in angiogenesis inhibition. Although the study by Zhang et al was not aimed at measuring a clinical correlation with histopathological findings, any investigation of a possible relationship between a higher ECS expression and a better course of the disease in the 16 multiple sclerosis (MS) patients under study could be of great interest. However, in order to perform a correlation analysis between ECS expression and disease stage, a more accessible tissue than brain is needed.

We investigated in 22 MS patients the blood counterpart of the ECS elements studied by Zhang et al, namely, the anandamide [N‐arachidonoylethanolamine (AEA)]‐binding CB₁R and CB₂R receptors, the AEA‐synthesizing enzyme NAPE‐PLD and the AEA‐degrading enzyme fatty acid amide hydrolase (FAAH). The messenger ribonucleic acid (mRNA) expression of these elements was measured in peripheral blood mononuclear cells of 22 patients affected by primary progressive (PP) MS and in those of 25 healthy controls.

Few studies have reported findings about ECS dysregulation in PP MS, where the immuno‐inflammation is associated to a far‐reaching neurodegenerative process. We thought, therefore, that this kind of MS disease is worth investigating in order to add any biological evidence in support of a possible neuroprotective role of ECS besides its well‐known anti‐inflammatory activity.

Quantitative reverse transcription polymerase chain reaction (RT‐PCR) analysis showed an overall up‐regulation of gene expression of all AEA‐related elements of the ECS in PP MS patients, compared with healthy controls (Table 1). The level of CB₁R mRNA was ∼12‐fold and that of CB₂R was ∼20‐fold higher in PP MS patients than in normal subjects (Table 1). In addition, higher mRNA levels of the AEA‐metabolizing enzymes were detected in PP MS patients compared with controls, FAAH increasing by ∼twofold and NAPE‐PLD by ∼24‐fold (Table 1).

Table 1.

mRNA expression (fold over controls). Abbreviations: FAAH = fatty acid amide hydrolase; mRNA = messenger ribonucleic acid; NAPE‐PLD = N‐acyl phosphatidylethanolamine phospholipase D; PP MS = primary progressive multiple sclerosis.

	Healthy controls (n = 25)	PP MS patients (n = 22)
CB1R	0.74 ± 0.55	12.44 ± 3.36*
CB2R	1.08 ± 0.47	20.77 ± 0.95**
FAAH	1.16 ± 0.73	2.35 ± 0.32
NAPE‐PLD	1.21 ± 0.59	24.91 ± 3.37**

CB₁R, CB₂R, FAAH and NAPE‐PLD mRNA expressions in lymphocytes from healthy controls and PP MS patients. Gene levels were measured by quantitative RT‐PCR, using β‐actin as housekeeping gene to normalize the data, and were expressed as fold over healthy controls. Data are reported as mean ± standard deviation (SD).

*P < 0.01 vs. healthy controls; **P < 0.001 vs. healthy controls (unpaired Student's t‐test).

Our new findings confirm the up‐regulation of ECS elements also in PP MS. In 2009, Gilles et al (1) reported that endocannabinoid levels are elevated in MS plasma, but they found that these changes may depend on disease subtype, being an up‐regulation of cannabinoid receptors dramatically higher in progressive forms of MS compared with relapsing‐remitting MS and healthy controls.

The data reported by Zhang et al (2) are noteworthy. In fact, if the neuroprotective role of ECS at the site of vascular and neuronal damages will be corroborated by further studies, any drug enhancing this system will be highly wanted. Our findings appear to mirror at the peripheral level the histological data reported in (2). Considering the ease of access to blood peripheral lymphocytes, it can be anticipated that markers of ECS activation in these cells might be useful to monitor the effects of any drug enhancing ECS within the brain.